Wood ash as a forest soil amendment: effects on seedling

growth and nutrition, and red-backed salamander

abundance

by

Adam Gorgolewski

A thesis submitted in conformity with the requirements

for the degree of Master of Science

Faculty of Forestry

University of Toronto

© Copyright by Adam Gorgolewski, 2015

ii

Wood ash as a forest soil amendment: effects on seedling growth and nutrition, and

red-backed salamander abundance

Adam Gorgolewski

Master of Science

Faculty of Forestry

University of Toronto

2015

Abstract

Wood ash is being considered as a soil amendment in acidified forests of eastern North America.

However, research is required to ensure ash does not negatively affect native species. The short-term

effects of ash on growth and nutrition of tree seedlings native to eastern North America was assessed in

greenhouse experiments, finding relatively neutral effects up to dosages of 10 Mg ha-1, and negative effects

at dosages of 15-20 Mg ha-1. Responses of red-backed salamander (Plethodon cinereus) abundance to ash

additions up to 8 Mg ha-1 in a tolerant hardwood forest were also assessed in a field trial. Neutral and

positive effects of ash on salamander abundance were observed, and positive effects were driven by

increases in soil pH and moisture. These results tentatively support ash additions in acidified forests of

eastern North America, but more research is needed into the longer-term effects of ash and its potential

toxicants.

iii

Acknowledgements

I am sincerely grateful to my supervising professors Dr. Nathan Basiliko and Dr. John Caspersen, for

their continuous support, enduring patience, and many opportunities and resources they have offered me

during my MSc. I am also particularly grateful for the methodological support, funds, and personnel that

committee member Dr. Trevor Jones has provided, and to committee members Dr. Honghi Tran and Dr.

Tat Smith for their encouraging comments and feedback on my thesis. A special thanks also goes out to

Emma Horrigan for her support in the field, for her input and feedback regarding the projects.

Thanks to Dr. George Arhonditsis and to DeChang Gao for providing statistical consultation and

support, and to the plethora of field assistants, lab assistants, technicians, and students who have invested

time into the project, including John Perron, Darren Derbowka, Jeff Kokes, Genevieve Noyce, George

Dong, and Joey Aulakh. This project would also not have been possible without the grants and scholarships

provided to me and my supervisors by NSERC and the University of Toronto. I am truly grateful to Peter

Schleifenbaum and the staff of Haliburton Forest and Wildlife Reserve for providing accommodations,

logistical support during fieldwork, and a beautiful forest to work in. Finally, I would like to thank my

family, girlfriend, personal mentors, and the many animals in my life, who have all encouraged me and

put up with me throughout my MSc degree.

iv

Contents

Introduction: .................................................................................................................................. 1

Ash Properties: .......................................................................................................................................... 2

Effects of Ash on Forest Soils: ................................................................................................................. 3

Effects of Ash on Forest Plants: ................................................................................................................ 4

Ash Regulation: ........................................................................................................................................ 6

Study Summary: ........................................................................................................................................ 7

Thesis Objectives: ..................................................................................................................................... 8

Chapter 1: Wood Ash as a Forest Soil Amendment: Responses of Betula alleghaniensis, Pinus

strobus, Pinus resinosa, and Picea glauca seedlings to fly and bottom ashes, and mixed wood ash, in

a greenhouse experiment

Abstract: ................................................................................................................................................... 9

Introduction:............................................................................................................................................. 9

Methods: ................................................................................................................................................ 12

Experimental Design: ......................................................................................................................... 12

Biochemical Analysis: ....................................................................................................................... 13

Statistical Analyses: ........................................................................................................................... 14

Results and Discussion: ......................................................................................................................... 15

Seedling leaf tissue chemistry: ........................................................................................................... 15

Seedling growth responses: ................................................................................................................ 21

Conclusion: ............................................................................................................................................ 25

Figures and Tables: ................................................................................................................................ 27

Chapter 2: Wood Ash as a Forest Soil Amendment: Responses of Red-Backed Salamander

(Plethodon cinereus) Abundance in a Northern Hardwood Forest

Abstract: ................................................................................................................................................. 44

Introduction: ........................................................................................................................................... 44

Methods: ................................................................................................................................................ 46

Site Description: ................................................................................................................................. 46

Plot Setup and Experimental Design: ................................................................................................ 46

Field and Laboratory Sampling and Anlyses: .................................................................................... 47

Statistical Analyses: ........................................................................................................................... 48

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Results: ................................................................................................................................................... 49

Discussion: ............................................................................................................................................. 51

Conclusion: ............................................................................................................................................ 54

Figures and Tables: ................................................................................................................................ 56

General Conclusions: ............................................................................................................................. 65

Literature Cited: .......................................................................................................................................... 67

1

Introduction

Wood ash from biomass boilers is generated in large quantities by forestry industries in North America,

when waste wood is disposed of in biomass boilers to generate on-site bioenergy. In Canada alone over

0.75 million tonnes of ash are annually produced (Elliott & Mahmood, 2006), and in the USA over 3

million tonnes are produced (Risse & Harris, 2013). Production of ash is likely to increase in the future

due to the growing bioenergy industry, which is projected to triple by 2050 (IEA, 2013). Ash has a high

pH and neutralizing capacity, and except for N, retains nutrients in similar proportions to those required

by growing trees (Vance, 1996). In eastern Europe (and a growing number of other countries), ash from

biomass boilers is applied to acidic and low nutrient forest soils in order to neutralize soil pH and reinstate

depleted nutrients, and to avoid disposal in landfills (Stupak et al., 2008). Conversely, in North America

ash is generally sent to landfills at a cost (Pitman, 2006; Elliot & Mahmood, 2006), but interest is growing

in using wood ash as a forest soil amendment to reinstate acidified and low nutrient forest soils, and to

avoid the cost of landfilling.

There are growing concerns that forest harvesting in North America is inducing soil nutrient

depletions, particularly over long timescales and several harvest rotations (Federer et al., 1989; Johnson et

al., 1991; Phillips & Watmough; 2012; Grand et al., 2014). Given that there is also interest in intensifying

extraction of forest biomass to support the growing bioenergy industry (Puddister et al., 2011) the potential

for forest soil nutrient depletions following harvests may be exacerbated (Shultze et al., 2014), but by

applying ash to soils of harvested forests the nutrients that were removed in the biomass of harvested trees

could be reinstated (Vance, 1996). Many forests soils in eastern North America also suffer from

acidification and associated losses of nutrients due to historic acid rain (Likens & Bormann, 1974; Driscoll

et al., 2001; Watmough & Dillon, 2003). Although regulations have been relatively successful at

alleviating emissions of SO2 (the primary acid rain causing pollutant), soil acidity and nutrient imbalances

persist (Watmough & Dillon, 2003), and acid rain continues to a lesser extent due to emissions of NOx

(Driscoll et al., 2001). Interest is growing in liming these acidified forest soils to reinstate soil pH and

nutrient status (Long et al., 2011; Moore et al., 2012), and wood ash from biomass boilers may be an

effective, cheap, and convenient liming agent.

Although ash is already commonly used as a forest soil amendment in eastern Europe, it is primarily

applied to intensively managed forests and plantations (Stupak et al., 2008), and little research exists on

the impacts of ash in North American forests which are often less intensively managed, and in which ash

additions may pose more significant ecological risks. Most ash-based research has focused on soil and tree

responses to ash additions, with a heavy focus on eastern European tree species (Augusto et al., 2008).

2

Before ash is applied to forest soils in North America, precautionary research into the effects of ash on

native forest species, and on other important aspects of forest ecosystems is necessary.

Ash Properties

From the standpoint of being used as a forest fertilizer, ash is an alkaline substance with an average

pH between 8-13 (Augusto et al., 2008), and is generally a rich source of the plant-essential macronutrients

Ca, Mg, K, and P, and the micronutrients Fe, Mn, Zn, B, Cu, and Mo (Demeyer et al., 2001; Pitman, 2006).

Calcium is present in the largest concentrations, followed by K, Mg, Al, Fe, and P (Reid & Watmough,

2014). K is the most soluble nutrient in ash, and Ca then Mg are the next most soluble (Eriksson, 1998;

Demeyer et al., 2001). The solubility of P is generally lower than that of K, Ca, and Mg (Eriksson, 1998;

Demeyer et al., 2001). N is not present in significant quantities in wood ash, as the organic N compounds

in wood vaporize when combusted (Ingerslev et al., 2011). The solubility of metals in wood ash is

generally low (Perkiomaki et al., 2003), as most metals are less reactive in higher pH environments (Brady

& Weil, 2010).

There are several properties of ash that could be detrimental to forest ecosystems when applied at high

dosages. Firstly, the initial pH of ash can be above 13 (Pitman, 2006), which could be caustic to susceptible

plants and fauna. Ash also retains traces of heavy metals (including As, Cd, Ni, Pb, Mn, Zn, Cu, B, Ni)

which could be toxic to plants when applied at high dosages (Pitman, 2006; Moilanen et al., 2006; Dahl et

al., 2010). Finally, ash can contain high concentrations of Na salts that dissolve rapidly following

application (Huotari et al., 2015), and ashes with particularly high Na concentrations have been shown to

cause salt toxicity to tree seedlings on poorly drained soils (Staples & van Reese, 2000).

Biomass boilers produce fly ash – a volatile lighter ash extracted from flue gasses, and bottom ash –

heavier particles that fall to the bottom of boilers (Ingerslev et al., 2011). Fly ash typically retains more

nutrients than bottom ash, but also retains more metals (Dahl et al., 2010), which has led some authors to

dismiss the use of fly ash as a forest soil amendment (Pitman, 2006). However, as the metals in ash tend

to be unavailable to plants after application due to its neutralizing effect, fly ash may still be an acceptable

soil amendment (Augusto et al., 2008; Perkiömäki et al., 2003). Specific properties of ash can also vary

depending on the feedstock, boiler type, and boiler operating conditions (Pitman, 2006; Bostrom et al.,

2010), which makes it challenging to draw comparisons between studies that used different ashes.

Although in its raw form ash can have some properties that are potentially detrimental when applied

to forest soils, in Europe the ash is often ‘hardened’ prior to application by leaving the ash outdoors during

the summer to be weathered for 1-3 months (Dong et al., 2014). This is done to decrease its pH, as the

high initial pH of un-hardened ash can damage susceptible ground vegetation and fauna (Kellner &

3

Weibull, 1998). Alongside hardening, ash can also be granulated to mitigate the potential negative effects

of its high initial pH, to increase the duration of its positive effects to soil, and to make it easier to apply

(Pitman, 2006). In order to make granulated ash, it is mixed with water and then rolled into balls before

being dried to <5% moisture content (Kellner & Weibull, 1998). This process of granulation has also been

shown to alter the chemical composition of ash, and Pitman (2006) reported that more processed ash

generally has higher P and Ca concentrations.

Effects of Ash on Forest Soils

As a soil amendment ash increases soil pH, delivers nutrients, and alters nutrient availability by

increasing soil pH and by stimulating microbial activity (Augusto et al., 2008).

Increases in soil pH following ash additions are caused by its CaO/Ca(OH)2 and CaCO3 contents (Dong

et al., 2014). Immediately following application, CaO/Ca(OH)2 causes a rapid and short lived spike in soil

pH, and then CaCO3 causes longer term and sustained increases in soil pH (Dong et al., 2014). Hardened

ash has less CaO/Ca(OH)2 and more CaCO3 (Dong et al., 2014), and does not cause the rapid spike in pH

that untreated loose ash can induce (Pitman, 2006).

Ash is a particularly rich source of Ca, K, Mg, Al, Fe, and P (Reid & Watmough, 2014), although the

biological availability of these nutrients after application is strongly dependent on soil pH. Most of the

essential plant macro-nutrients (N, P, K, Ca, Mg) are most available in soils with pH values of 6-7 (Brady

& Weil, 2010), and the neutralizing effect of ash generally increases their availability (Demeyer et al.,

2001). Of the plant essential macronutrients that wood ash delivers to soil, K is the most soluble and readily

available to plants, followed by Ca, Mg, and P (Demeyer et al., 2001). The neutralizing effect of ash also

limits the availability of potentially toxic heavy metals (Perkiomaki et al., 2003), most of which are more

available at pH values below 5 (Brady & Weil, 2010). When Perkiomaki et al., (2003) applied ash to acidic

and Ni and Cu contaminated soils, decreases in availability of Ni and Cu were reported, and it was

concluded that ash may be an effective tool to remediate metal-polluted soils. However, the long-term

mobility of metals following ash additions is still in question, (Huotari et al., 2015), and is an important

research question that requires long term studies to be assessed.

The neutralizing effect of ash has also been shown to stimulate microbial activity in the organic layers

of soil (Augusto et al., 2008; Perkiomaki et al., 2003; Mahmood et al., 2003), which in turn increases

availability of certain plant nutrients. For example, although ash does not directly supply N to the soil, it

has been found to stimulate rates of microbial N mineralization in organic soils, which then increases N

availability to plants (Pitman, 2006; Augusto et al., 2008). Ash has also been shown to increase abundances

of collembolans, mites, and enchytraeid populations in soil (Nieminen, 2012), due to increases in soil

4

moisture. However, ash does not always increase soil moisture, and this effect depends on the type of ash

and its C content (Pugliese et al., 2014). Little research exists regarding the effects of ash on other species

of forest fauna beyond soil invertebrates, and no studies have assessed the effects of ash on vertebrate

population abundances. Before being applied to forest soils in North America, the effects of ash on native

soil fauna and on ecologically significant vertebrate species should be assessed.

Effects of Ash on Forest Plants

Most literature regarding the effects of ash to plants focuses on mature trees native to eastern Europe,

growing in intensively managed forests and in plantations. The effects of ash are highly dependent upon

the soil type that it is applied to, as soil type governs the changes in pH and subsequent availability of

nutrients following ash additions (Pitman, 2006; Reid & Watmough, 2014; Huotari et al., 2015). As N is

typically limiting in forest soils (Aber et al., 1989), most authors conclude that the lack of N in wood ash

limits its potential for increasing the productivity and yields of forest stands growing on mineral soils

(Pitman, 2006; Augusto, 2008; Huotari et al., 2008; Jacobson et al., 2014; Kloseiko et al., 2014; Huotari

et al., 2015). However, the tendency of ash to stimulate N-mineralizing microbes when applied to organic

soils can increase N supply to plants, resulting in increases in tree yields (Pitman, 2006; Augusto et al.,

2008; Huotari et al., 2015). Ash can also increase tree growth and yields when applied to forest soils with

higher N concentrations such as drained peatlands, which also have low concentrations of most other plant

essential nutrients (Pitman et al., 2006; Stupak et al., 2008; Augusto et al., 2008; Huotari et al., 2015).

Most forest soils in North America are N limited, although in certain areas of eastern North America acid

deposition of NOx has caused them to move away from N-limitation and toward either P or Ca limitation

(Watmough & Dillon, 2003; Gradowski & Thomas, 2006). On these soils not only could wood ash be used

to restore soil pH and nutrient status, but could also potentially increase tree growth and yield.

The effects of ash on tree productivity are also dependent on the time since application. In a meta-

analysis of ash addition literature, Reid & Watmough (2014) found that ca. 30% of the reviewed studies

reported positive effects on tree growth from ash additions; however, temporal variation in the dataset may

explain many of the zero-effect results, as 50% of the reviewed studies were conducted over less than 6

years, and longer term studies more often report beneficial effects to tree growth. This is likely because

the effects of ash on forest soil nutrient status and pH are a series of dynamic processes which can take

several years to be realized (Demeyer et al., 2001).

Although ash does not always increase tree productivity in the short term, increases in foliar

concentrations of K and Ca are generally seen within 1-2 years following ash additions, irrespective of soil

type (Augusto et al., 2008; Reid & Watmough, 2014). Ash can also increase foliar B concentrations, but

5

this tends to be dependent on soil type (Augusto et al., 2008). Increases in foliar Mg and P are variable,

and tend to take 3-4 years to be realized (Augusto et al., 2008; Huotari et al., 2015). The foliar

concentrations of N generally remain unchanged following ash additions. However, increases in foliar N

have been observed due to the stimulation of N-mineralizing microbes (Pitman, 2006; Augusto et al., 2008;

Huotarti et al., 2015). Decreases in foliar N concentrations have also been observed, and these were

attributed to a “dilution effect”, whereby ash additions stimulated the growth of trees and their N content

was then distributed over a larger biomass (Huotari et al., 2015). The concentrations of metals in tree

foliage tends to remain low following ash additions, due to their low availability in the more neutral pH

environments that ash creates. Minor increases in foliar Cu and Zn have been reported in foliage of Picea

abies initially following application (Osteras et al., 2005), and of Cd in select grasses and shrubs (Moilanen

et al., 2002; Huotari et al., 2011), but these changes tend to be transitory and return to previous levels or

even lower several years following application (Huotari et al., 2015). However, metals often accumulate

more in the roots rather than the shoots of plants, and no studies exist regarding the effect of wood ash on

heavy metal concentrations in plant roots (Huotari et al., 2015).

A handful of studies have also investigated the effects of ash on tree seedlings, finding mixed effects.

Although direct positive effects to seedlings productivity are generally not expected on mineral soils, at

dosages of up to 10 Mg ha-1 and over time periods of 1-5 growing seasons ash has been found to increase

seedling height, root collar diameter, and shoot length in Pinus sylvestris (Kloseiko et al., 2014), Picea

abies (Kloseiko et al., 2014; Nieminen, 2009), Betula pendula (Mandre et al., 2010), and Alnus glutinosa

(Parn et al., 2009), even in the absence of supplemental N. Ash has also been found to decrease growth of

Picea glauca (Staples & Van Rees, 2001) and Picea abies (Mandre et al., 2004), and to have no effect on

Picea abies (Nieminen, 2009). The variable effects of ash seem to depend on the type of soil that was used,

the length of the study period, the age of the seedlings, the properties of the ashes and the dosages at which

they were applied, and the seedling species.

The influence of soil type on the effects of ash to seedlings appears to be similar to mature trees, and

when applied to peat soils ash consistently increases seedling growth and yields (Mahmood et al., 2003;

Parn et al., 2009; Mandre et al., 2010), but more variable results are reported on mineral soils (Mandre et

al., 2004; Nieminen, 2009). Similar to mature trees, the length of the study period also plays a role in the

effects of ash to seedlings. For example, Solla Gullon et al., (2008) explored the effects of ash (bark

derived, fly and bottom ash mixed) at dosages of 5 and 10 Mg ha-1 on growth of Pinus radiata seedlings,

and only found significant effects on plant growth after 5 years. This is consistent with the results of

Nieminen (2009), who found ash (unspecified type and feedstock) to have no effect on Picea abies biomass

in a greenhouse experiment over 93 days. The seedlings’ age may also play a role in how ash effects their

6

growth and nutrition (Augusto et al., 2008), and most studies use seedlings that were 1 year old or older.

Younger seedlings are more susceptible to changes in their environment than older seedlings and mature

trees, and thus the initial spike in pH that ash often causes and changes in soil nutrient balances could have

more negative effects. The physical and chemical properties of ashes can be highly variable (Demeyer et

al., 2001; Pitman, 2006; Ingerslev et al., 2011), and may also play an important role in its effects to

seedlings. For example, Staples & Van Rees, (2001) applied ash mixed with pulp sludge that had relatively

high concentrations of Na salts to Picea glauca seedlings, and after 2 growing seasons dosages of 5 Mg

ha-1 had increased soil electrical conductivity (EC) to a level that caused decreases in seedling growth.

Most studies do not specify the type of ash that was used or the feedstock that was burned, making accurate

comparisons between studies challenging (August et al., 2008). Due to the variability in ash properties,

the effect of ash dosage is also highly variable. However, there appears to be a lack of studies that have

investigated the effects of ash dosage on plant growth and nutrition, with most studies testing ash at only

2 or 3 dosages that generally do not exceed 10 Mg ha-1. Finally, the effects of ash can vary based on the

species of seedling that was used. For example, Parn et al., (2009) found ash additions to favour the growth

of Alnus glutinosa over Betula pendula seedlings. In an ash addition meta-analyses Reid & Watmough

(2014) also found that tree species was one of the main variables that determined the effects of ash on tree

productivity. Vance (1996) suggested that wood ash may be more beneficial to hardwoods rather than

softwoods due to their higher nutrient demands, although this has never been explicitly proven. It could

also be hypothesized that ash would have a more positive effect on fire-adapted tree species, as these have

evolved in conjunction with natural ash additions to the soil from forest fires.

Effects of ash on forest understorey and ground vegetation have also been reported on. The initial

spike in soil pH caused by untreated wood ash has been shown to reduce ground vegetation cover at

dosages of 7-9 Mg ha-1, particularly of bryophytes and lichens (Kellner & Weibull, 1998; Jacobson &

Gustafsson, 2001). However, such effects tend to only occur at ash dosages above 7 Mg ha-1 and are short

lived, with vegetation generally recovering within 5-10 years (Pitman, 2006). The effects of ash on ground

vegetation also vary with the surface area of ash that comes into direct contact with the plants, and

granulated ash does not cause these detrimental effects (Pitman, 2006).

Ash Regulation

In eastern European countries, regulations and guidelines for use of ash as a forest soil amendment are

already in place, and ash additions of between 1-8 Mg ha-1 are commonly prescribed to replenish depleted

soil nutrients and neutralize acidic soils (Stupak et al., 2008). These guidelines focus on using ash to

increase pH and to re-establish nutrient balances in forest soils, whilst restricting contamination from

7

heavy metals and salts (Demeyer et al., 2001; Pitman et al., 2006; Augusto et al., 2008; Stupak et al.,

2011). In Canada, land application of ash falls under provincial regulation for land application of industrial

waste products, with the Ministry of Environment and Climate Change (MOECC) setting regulations in

Ontario, and in the USA ash additions are federally regulated by the Environmental Protection Agency

(EPA) (EPA, 1993; MOE, 2002). Regulations for ash additions by North American agencies focus on

minimizing heavy metal contamination, and are broadly applied to land application of most industrial

waste products. Specific regulations or guidelines do not exist for applying ash to forest soils. However,

regulations for the application of wood ash to agricultural soils do exist in Alberta (Alberta Environment,

2002), and some forestry companies have had their ash certified by the Canadian Food Inspection Agency

(CFIA) as an agricultural fertilizer (CFIA, 2014).

A challenge of regulating ash as a soil amendment is the variability in its properties (Pitman et al.,

2006; Bostrom et al., 2010; Ingerselv et al., 2011). Current methods of regulating ash additions in European

countries involve testing the ashes before application, to ensure that nutrient and metal concentrations are

within specified limits (Stupak et al., 208). As biomass boiler technologies are constantly evolving and

becoming more and more efficient, precautionary testing of ash chemistry before application is likely to

be a necessity in regulation of ash additions to North American forest soils.

Study Summary

The variability in ash chemistry and variability in its effects to soils and plants justifies extensive

precautionary research before ash is approved for large-scale additions in forests of North America. A

substantial amount of research has been done in eastern European forests, but the effects of ashes on North

American forests must be assessed to ensure it does not detrimentally effect native species. Ash may be a

particularly effective soil amendment on acidified and nutrient depleted forest soils of eastern North

America; however, these soils are typically much less intensively managed than forests where ash is

applied in eastern Europe, and the effects of ash on broader ecosystem health should also be assessed. The

importance of long-term studies to assess the long term effects of ash and the mobility of its potential

toxicants is also evident. Several gaps in the literature exist, such as the effects of ash on forest vertebrate

populations, and systematic experimental evaluations regarding the effects of different ash types, applied

at different dosages, on different tree species, growing on different soil types.

For this thesis, several seedling trials in which the effects of fly and bottom ashes, from different

boilers, at different dosages, was assessed on the growth and nutrition of seedling species native to eastern

North America, and the results are presented in this thesis. Field trials were also established in Haliburton

Forest and Wildlife Reserve, a managed tolerant hardwood forest in central Ontario. The field trial was

8

intended to become a long-term monitoring site to assess the effects of wood ash in a forest that has

suffered from soil acidification and nutrient depletions due to acid rain, and in which ash may provide

additional beneficial effects to tree productivity due to high levels of N in the soil. Several datasets were

established to assess the effects of ash additions on many aspect of the forest ecosystem, including soil

chemistry and biology, responses of plants (understorey and overstorey), heavy metal accumulation in

forest food sources, and abundances of ecologically significant indicator species. Alongside the effects of

ash additions to seedling growth and nutrition, this thesis also presents results regarding the short term (1

year) effects of ash additions on red-backed salamander (Plethodon cinereus) populations, which are an

ecologically significant indicator species in forests of eastern North America.

The chapters of this thesis have been presented as stand-alone manuscripts.

1. Wood ash as a forest soil amendment: Responses of Betula alleghaniensis, Pinus strobus, Pinus

resinosa, and Picea glauca seedlings to fly and bottom ashes, and mixed wood ash, in a greenhouse

experiment

2. Wood ash as a forest soil amendment: Responses of red-backed salamander (Plethodon cinereus)

abundance in a northern hardwood forest. (This manuscript has been submitted to the Canadian

Journal of Forest Research and is in review)

Thesis Objectives

The objectives of this thesis were to begin determining whether wood ash from biomass boilers is

a safe and effective soil amendment when applied to acidified and low nutrient forest soils in eastern North

America, focusing on short-term effects of ash. Specifically, I set out to determine:

1. How do ashes from different boilers, and different ash types (fly and bottom ash), effect growth

and nutrition of native germinant seedling species when applied to a low nutrient and acidic soil,

at dosages from 1-20 Mg ha-1, and what is the optimum ash dosage?

2. How does ash alter the natural habitat, and relative abundance of red-backed salamander

populations when applied to an acidified northern hardwood forest?

9

Wood ash as a forest soil amendment: Responses of Betula alleghaniensis,

Pinus strobus, Pinus resinosa, and Picea glauca seedlings to fly and bottom

ashes, and mixed wood ash, in a greenhouse experiment

Abstract

Wood ash from biomass boilers may be a useful forest soil amendment in North America to correct

soil acidification and nutrient depletions. Short term effects of ash were tested in greenhouse experiments

on the growth and foliar concentrations of P, K, Ca, Mg, and Mn, of Betula alleghaniensis, Pinus strobus,

Pinus resinosa, and Picea glauca seedlings. Two experiments were conducted to test the effects of 1) fly

and bottom ash mixed in a 1:1 ratio (mixed wood ash), from 2 different boilers, at dosages of 0, 1, 2, 5,

10, 15, & 20 Mg ha-1, and 2) fly and bottom ash applied separately, from 2 different boilers, at dosages of

0, 1, 5, & 10 Mg ha-1. Metal concentrations in all ashes were within the limits set by the MOECC and

EPA, although the Na and As concentrations in the ashes were relatively high. Irrespective of ash type or

boiler type, at dosages below 10 Mg ha-1 ash had a neutral effect on seedling biomass and leaf/needle

area, presumably due to the low N concentrations in the ash. The root:shoot ratios and foliar P, K, and

Ca status of red pine suggested that ash dosages of 10 Mg ha-1 were optimum. At dosages above 10 Mg

ha-1 the biomass and leaf/needle areas of white pine decreased, and at 20 Mg ha-1 that of yellow birch and

red pine also decreased, and this is expected to be due to either the antagonistic effects of Na, toxicity by

certain metals, or excessive increases in soil pH. This study tentatively supports applying low dosages of

ash to mineral soils to counteract soil acidification and associated nutrient depletions, and to redirect ash

away from landfills. Future studies should investigate the effects of ash additions on different soil types

and over longer timeframes, and on the toxicity potential of heavy metals in ash.

Introduction

Wood ash is generated in large quantities by forestry industries in North America, who use biomass

boilers to burn waste wood and to generate on-site bioenergy. The residual ash is generally sent to landfill

at a cost, with Canada annually producing over 0.75 million tonnes of ash of which approximately 80% is

sent to landfills (Elliott & Mahmood, 2006), and the USA producing over 3 million tonnes of ash (Risse

& Harris, 2013) of which 90% is sent to landfills (Pitman, 2006). The use of bioenergy as a carbon neutral

10

energy source is also projected to triple by 2050 in response to increasing oil prices, energy security, and

climate change (IEA, 2013), which will create more waste ash.

Wood ash has a high pH and neutralizing capacity, and except for N, retains nutrients in similar

proportions to those needed by growing trees (Demeyer et al., 2001). These characteristics make ash an

effective soil amendment on acidic and low nutrient soils (Pitman, 2006; Augusto et al., 2008; Stupak et

al., 2008; Reid & Watmough, 2014; Huotari et al., 2015). In eastern Europe over 80 years of research

exists regarding the use of ash as a forest soil amendment in intensively managed forests and plantations,

and ash additions of between 1-8 Mg ha-1 are commonly prescribed to neutralize acidic soils and to

replenish depleted soil nutrients (Stupak et al., 2008). N is the primary limiting nutrient in temperate forest

soils (Aber et al., 1989) and as ash does not retain significant quantities of N (Ingerslev et al., 2011), it

rarely increases forest productivity (Jacobson et al., 2014), particularly in the short term (Reid &

Watmough, 2014). However, ash can correct soil nutrient imbalances (Demeyer et al., 2001; Stupak et al.,

2008) and improve the foliar nutrient status of trees (Demeyer et al., 2001; Pitman, 2006; Augusto et al.,

2008; Reid & Watmough, 2014; Huotari et al., 2015), and when added to N rich sites such as drained

peatlands ash can also increase tree productivity by over 9 times (Huotari et al., 2015).

In North America there are concerns regarding forest soil acidification and nutrient depletions, and ash

is being considered for use as a soil amendment to rectify these issues. Historic acid rain has degraded

many forest soils in eastern North America, causing soil acidification, loading of N, and associated losses

of base cations (Likens & Bormann, 1974; Driscoll et al., 2001; Watmough & Dillon, 2003). Although

regulations have been relatively successful at alleviating emissions of SO2, soil acidity and low base cation

levels persist (Watmough & Dillon, 2003), and acid rain continues to a lesser extent due to emissions of

NOx (Driscoll et al., 2001). There are also ongoing concerns that intensive forest harvesting can induce

soil acidification and nutrient depletions, particularly over long timescales and several harvest rotations

(Federer et al., 1989; Johnson et al., 1991; Grand et al., 2014). Although some studies find that even

intensive harvests have a relatively small impact on soil fertility (McLaughlin, 2014), longer term studies

are needed over many cut-cycles. Indeed, Phillips & Watmough (2012) found that even under low intensity

harvest regimes there is still potential for nutrient depletions and acidification in tolerant hardwood forests.

There is also growing interest in intensifying extraction of forest biomass to support the growing bioenergy

industry (Puddister et al., 2011), which may accelerate existing problems of forest soil acidification and

nutrient depletions (Shultze et al., 2014).

Wood ash may be a convenient soil amendment to counteract soil acidification and nutrient depletions

in forests of North America, as it has a strong neutralizing capacity and high concentrations of base cations.

This would redirect ash away from landfills and save the cost of disposal, and as ash is often generated in

11

close proximity to managed forests the potential transportation costs might be relatively small. However,

although much research exists on the use of ash in intensively managed forests and plantations in eastern

Europe, little research exists on the benefits and impacts of ash in North American forests, which are often

less intensively managed and in which ash additions may pose more significant ecological risks.

Ash has several properties that may be detrimental to more susceptible forest flora and fauna. Firstly,

ash can have an initial pH of over 13 (Demeyer et al., 2001), and although this decreases rapidly after

application as the ash equilibrates with the soil (Dong et al., 2014), its initial pH could be caustic to certain

species (Pitman, 2006). Ash also retains traces of heavy metals (including As, Cd, Ni, Pb, Mn, Zn, Cu, B)

and salts (Na), all of which have the potential to be toxic at large enough dosages (Moilanen et al., 2006;

Pitman, 2006; Dahl et al., 2010). Although present in ash, metals rarely exceed regulatory limits for land

application of industrial waste residues in Canadian contexts (Pugliese et al., 2014), and most metals are

also less biologically available in the higher pH environments that ash creates in soil (Perkiomaki et al.,

2003). The salt content of ash can vary widely (Karltun, 2015), and may be an issue when high amounts

are applied to poorly drained soils (Staples & Van Rees, 2001; Pugliese et al., 2014) or to particularly salt-

sensitive species.

The nutrient composition of ash can vary between different boilers, and different ash types. Biomass

boilers generally produce both fly ash - a volatile, lighter, more reactive ash, extracted from flue gasses,

and bottom ash – heavier, less reactive particles that fall to the bottom of boilers (Ingerslev et al., 2011),

and conclusions regarding their suitability as soil amendments vary. Furthermore, the type of ash used in

published literature is often unspecified (Augusto et al., 2008), making it challenging to differentiate

between ash sources or types in evaluating effects as soil amendments.

A handful of studies have investigated the effects of ash on tree seedling growth parameters and foliar

nutrient concentrations, finding a mix of positive (Parn et al., 2009; Kloseiko et al., 2014; Mandre et al.,

2010; Nieminen, 2009), negative (Staples & Van Rees, 2001; Mandre et al., 2004), and neutral (Nieminen,

2009) effects. In general, these studies investigated the effect of ash at 2-3 dosages that did not exceed 10

Mg ha-1, and there appears to be no studies that have explored the effects of ash dosage on seedling growth

and nutrition. Making direct comparisons between studies can be complicated by the variety of soil types

and ash types that were used. If ash is to be used as a soil amendment in North America, its effects on

native tree species and seedlings must be systematically assessed, and the optimal dosages must be

determined. Additionally, the effects of fly ash and bottom ash, and ashes from different boilers must be

differentiated in order for appropriate guidelines and regulations to be imposed.

This study assessed the effects of wood ash fertilization on growth and health of 4 species of germinant

tree seedlings native to eastern North America, in a greenhouse experiment conducted over 24 weeks. The

12

following research questions were addressed: 1) How do native seedling species respond to wood ash

additions? 2) Do fly and bottom ash differ in their effects on seedling growth and leaf tissue chemistry? 3)

Do ashes from different boilers differ in their effects on seedling growth and foliar tissue chemistry? 4)

What is the optimum ash dosage to benefit seedling growth and health? The effects of ash on foliar nutrient

status and growth parameters of seedlings were expected to vary with species, as a result of the differing

nutrient requirements and tolerances of different seedling species. As ash was not applied in combination

with N, positive effects to seedling productivity were not expected. The variability in nutrient

concentrations between fly and bottom ashes and ashes from different boilers were expected to cause

varying effects on the seedlings’ foliar nutrient concentrations. Mid-range dosages of 5-10 Mg ha-1 were

expected to be the most beneficial, with the potential for negative effects at higher dosages.

Methods

Experimental design

Two ash fertilization experiments were conducted using germinant tree seedlings in a temperature and

light controlled greenhouse. The effects of ash on seedling biomass, leaf/needle area, root:shoot ratio, and

foliar nutrient status was assessed.

1. The first experiment tested the effects of fly and bottom ash when mixed in a 1:1 ratio by weight

(from here on referred to as ‘mixed wood ash’). Mixed wood ash from 2 different boilers was

applied at dosages of 0, 1, 2, 5, 10, 15, and 20 Mg ha-1, to yellow birch (Betula alleghaniensis

Britt.), white pine (Pinus strobus L.), red pine (Pinus resinosa Alt.), and white spruce (Picea glauca

(Moench) Voss.) seedlings in a full factorial experiment, conducted over 24 weeks (n=7).

2. The second experiment tested the effects of fly and bottom ash separately, from 2 different boilers,

at dosages of 0, 1, 5, and 10 Mg ha-1, on white spruce, red pine, and yellow birch seedlings, in a

full factorial experiment conducted over 24 weeks (n=8).

Ash was obtained from biomass boilers used in pulp and paper mills in eastern Canada. Boiler 1 is a

vibrating-stoker grate boiler (‘Detroit RotoStoker’) - an older system that is common in Canadian pulp and

paper mills. Boiler 2 is a newly installed Wellons gasification boiler with a separate furnace and

combustion chamber. Both boilers were primarily burning a combination of spruce/pine/fir (SPF) bark.

Seeds from the Algonquin seed zone were provided by Angus Seed Orchards, soaked in aerated water

for 24 hours, and then exposed to cold stratification at 2.5°C. White pine seeds were exposed for a period

of 20 days, yellow birch and white spruce for 28 days, and white pine for 60 days. A sandy and acidic

(pH=4.8) soil with low organic carbon and nutrient content was collected from the arboretum beside the

13

OFRI buildings in Sault Ste Marie, Ontario. Soil was manually homogenized, then autoclaved twice for

20 minutes at 121°C to remove biological pests. Thirty ml of peatmoss-Pearlite mixture was placed in 500

ml nursery plugs, which were then filled with soil to within 2-3cm of the top. One seed was planted in

each plug before being left in the greenhouse to germinate. To ensure the seedlings were able to survive

in such a low nutrient soil, 25 ppm of a general purpose 20-20-20 NPK water soluble fertilizer was added

uniformly to every sample (including the control). Watering occurred as needed by the crop (this was

visually determined based on the moisture of the soil) using overhead sprinklers, and an automatic lighting

system kept daylight at 18 hours per day. Temperatures were regulated to a minimum of 26°C during the

day and 18°C at night; however, the experiment occurred during the summer months and temperatures

regularly exceeded these. After the seeds had sprouted and the first set of needles or leaves had set,

treatments occurred by adding ash directly to the soil surface. The amount of ash added is expressed in

Mg ha-1, and was calculated based on the surface area of the nursery plugs. One extra treatment where the

NPK fertilizer was applied at 100 ppm was also included, from here on referred to as the ‘NPK’ treatment.

This treatment was used when interpreting the results, to compare ash-treated seedlings with those treated

with a conventional fertilizer, and the seedlings in the NPK treatment were considered ‘healthier’ than the

control with respect the their root:shoot ratios, biomasses, and leaf/needle areas.

Seedlings were harvested 24 weeks following ash additions. Harvested seedlings were rinsed to remove

any soil particles and then dried in a drying oven for 48 hours at 50°C. The roots, stems, and foliage were

then each weighed on a balance with 0.0001 g readability. Foliage was also scanned, and leaf/needle area

was determined using WinFolia and WinSeedle software.

Biochemical analyses

As the chemical properties of wood ash can vary between boilers and between ash types (Ingerslev et

al., 2011; Demeyer et al., 2001), 2 samples of each ash were randomly analyzed for nutrient content. Ash

chemistry was determined using a NCS combustion analyzer (Vario EL III, Elementar Americas, Mt.

Laurel, NJ) for total C, N, and S, and with an inductively coupled argon plasma (ICAP) spectrometer

(Varian Analytical Instruments, Walnut Creek, CA) following high temperature microwave acid digestion

for P, K, Ca, Mg, Al, Na, S, Fe, Cu, Mn, As, Zn, V, Cd, Co, Cr, Mo, Ni, Pb, Ba, Be, Si, La, Li, Se, and Sr

(EPA standard method 3052).

Foliage samples from a subset of ash dosages (0, 2, 10 & 20 Mg ha-1 for experiment 1, and 0, 5 & 10

Mg ha-1 for experiment 2) were ground in a Wiley mill to pass through a 2 mm mesh, and then all replicates

from each treatment were pooled for analyses (to obtain a large enough sample - approximately 2 g). The

ground samples were digested in a Se solution, then analyzed for Ca, Mg, Mn, P, and K using ICP analysis.

14

Total N was determined using the dry combustion method. Initially, a more complete foliar elemental

analyses was planned that also included C, Na, and metals, but restriction in the laboratory, and in some

cases insufficient sample sizes, resulted in only Ca, Mg, Mn, P, K, being tested, and N in certain seedlings

only.

The foliar nutrient concentrations of seedlings was expected to be lower than published “normal”

values for healthy seedlings, as the soil used was of particularly low nutrient quality. Therefore, the foliar

nutrient concentrations of treated seedlings were compared to the control groups, and vector analysis was

also employed to diagnose the foliar nutrient status (Timmer & Stone, 1978). Vector analysis is a graphical

method in which foliar nutrient concentration, nutrient content, and foliar biomass are all assessed in

unison and relative to a control, because foliar nutrient concentration can vary due to changes in seedling

biomass and nutrient uptake (Figure 1). This allows for more accurate diagnoses of fertilization effects

(dilution, luxury consumption, deficiency, sufficiency or excess (Figure 2)) than when using nutrient

concentration alone.

Statistical analysis

The extra treatment with 100 ppm of NPK fertilizer was excluded from analyses (but was included in

graphs for visual comparisons). Normality was assessed using normal probability plots of residuals,

frequency histograms, and the Kolmogorov-Smirnov, Lilliefors, and Shapiro-Wilks tests, and

homoscedasticity of variances was assessed using the Levene’s test (p>0.05). Variances differed

substantially between each of the seedling species and transformations could not suitably correct this, so

statistical analysis was conducted on each species separately.

For the first experiment that tested the effects of mixed wood ashes, biomass and leaf/needle area data

for white spruce, and root:shoot data for white pine and yellow birch were log-transformed to meet the

assumption of normality. The independent variables were ‘ash dosage’ and ‘boiler type’, and the dependent

variables were ‘seedling biomass’, ‘leaf/needle area’, and ‘root:shoot ratio’. GLMs were constructed for

each seedling species, for each dependent variable, using ash dosage as a continuous variable and boiler

type as a categorical variable. The dependent variables were expected to have either a linear or quadratic

dosage response. Thus, two GLMs were run for dependent variable using ash dosage as a linear and then

a quadratic variable, and Akaike’s Information Criterion (AIC) was used to select the most parsimonious

model. If the difference between the two AIC scores was less than 1, then the simpler model was used.

For the second experiment that tested the separate effects of fly and bottom ashes, all dependent

variables had normal distributions except for white spruce biomass and needle area, and transformations

could not normalize the data. However, the white spruce dataset had several outliers, where approximately

15

10 seedlings had biomasses an order of magnitude or more greater than the others. These outliers were

distributed relatively evenly across the treatment groups, indicating that this was unlikely to be a treatment

effect. Therefore, all data from white spruce seedlings that had biomass greater than 0.18 g was removed,

and the biomass and needle area data was then log-transformed to meet the assumption of normality. The

independent variables for experiment 2 were ‘ash dosage’, ‘boiler type’, and ‘ash type’, and the dependent

variables were ‘seedling biomass’, ‘leaf/needle area’, and ‘root:shoot ratio’. Similar to experiment 1,

GLMs were constructed for each seedling species, for each dependent variable, using ash dosage as a

continuous variable and boiler type and ash type as categorical variables. Again, the dependent variables

were expected to have either a linear or quadratic dosage response to ash dosage, and two GLMs were run

using ash dosage as both a linear and quadratic variable, and Akaike’s Information Criterion (AIC) was

used to select the most parsimonious model. If the difference between the two AIC scores was less than 1,

then the simpler model was used.

All statistical tests were considered significant at the p<0.05 level.

Results and Discussion

Seedling leaf tissue chemistry

The effects of ash on nutrient uptake by seedlings are a result of its influence on soil pH, nutrient

content, and nutrient availability. Neutralization of soil acidity by ash increases the availability of P, K,

Ca, and Mg, which are most available in soils with pH values of 6-7 (Brady & Weil, 2010), and limits the

availability of potentially toxic heavy metals (Perkiomaki et al., 2003), most of which are more available

at lower pH values (Brady & Weil, 2010). Ash also delivers nutrients to the soil, and is a particularly good

source of K, Ca, and Mg, and to a lesser extent P (Eriksson, 1988; Demeyer et al., 2001; Augusto et al.,

2008). In an 8 week soil incubation study, Pugliese et al., (2014) added ashes from the same boilers that

were used in this experiment at 5 Mg ha-1 to a soil with similarly low organic matter content, and found

increases in soil pH of 0.36-1.91 units, increases in extractable P, K, Ca, and Mg, and decreases in

extractable Cu, Zn, and Pb. Assuming that a similar response happened in the soil of this experiment, its

pH would have been raised from 4.8 to 5.1-6.7 (or greater at higher dosages), causing increases in plant

available macronutrients and decreases in availability of most metals.

Foliar K concentrations generally increased compared to the control groups in all seedlings species

treated with both mixed wood ash and fly and bottom ash (Tables 1, 2), and vector analyses for red pine

and white pine in generally indicated a slight shift towards luxury consumption of K (Figures 4b, 5b, 7b).

K is the most soluble nutrient in ash (Eriksson, 1988; Augusto et al., 2008), and increases in foliar K were

16

expected and are consistent with previous studies using other seedling species (Mahmood et al., 2003;

Mandre et al., 2004; Mandre et al., 2010; Kloseiko et al., 2014). However, vectors of certain treatment

groups for white pine and yellow birch also indicated a minor shift towards antagonism of K (Figures 3b,

5b, 6b, 7b). Antagonism occurs when 2 nutrients are in competition for uptake by plants, and affect each

other’s availability. A common form of antagonism in soils is brought about by excessive levels of Na,

which can limit plant uptake of K, Ca, and Mg (Hall, 2008). Excessive soil salts can also detrimentally

affect plant health by causing water stress due to differences in osmotic pressures between soil water and

plant roots, and by accumulating to toxic levels (Brady & Weil, 2010). The ashes used in this study had

relatively high levels of Na, but Na values were still within the normal range for ashes from boilers burning

similar feedstocks (Table 3). When the same ashes were used in the aforementioned soil incubation study

by Pugliese et al., (2014), fly ash from boiler 1 caused extractable soil Na to increase from 7.23 mg kg-1

to 199.32 mg kg-1, and the other ashes raised soil Na by up to 16.36 mg kg-1. However, the sodium

adsorption ratio (the ratio of cation charges from Na to that of Ca and Mg) was <13, which is within the

safe range for forest soils. In this experiment higher dosages of ash were used, which could have induced

Na:K antagonism. Also, germinant seedlings are more susceptible to changes in their immediate

environment than mature seedlings or adult trees, and may have been more sensitive to the potential

detrimental effects of Na. Staples & Van Rees (2001) applied a wood ash-pulp sludge mix that had a

similar Na concentration as fly ash from boiler 1 in this experiment to Picea glauca seedlings growing on

a clay-loam soil, finding that at dosages of 5 Mg ha-1 salt toxicity was induced. Salts are highly soluble

and are generally more detrimental on soils with poor drainage (Pugliese et al., 2014), and the negative

effects seen at relatively low dosages of ash by Staples & Van Rees (2001) may have been due to the poor

soil drainage. In this study, the sandy soil had good drainage, and this may have mitigated some of the

negative effects from the ashes’ high concentrations of Na salts, particularly at lower dosages.

In general, the ashes in this experiment had lower than average Ca levels when compared to other ashes

from boilers burning similar feedstocks (Table 3). Foliar Ca concentrations generally increased in yellow

birch and red pine, and the increase was most notable for red pine seedlings treated with mixed wood ash

at 10 Mg ha-1 (Tables 1, 2), which exhibited a shift towards luxury consumption of Ca (Figure 4b). Ash is

a particularly rich source of Ca (Augusto et al., 2008; Reid & Watmough, 2014), and as Ca is most

available at neutral soil pH values (Brady & Weil, 2010) it is often added to acidic and Ca depleted forest

soils to increase their pH and Ca levels (Stupak et al., 2008; Reid & Watmough, 2014). Previous studies

examining ash additions to seedlings also typically report increases in foliar Ca concentrations (Staples &

Van Rees, 2001; Mahmood et al., 2003; Mandre et al., 2004; Solla-Gullon et al., 2008). However, in the

present study Ca concentrations in white pine needles decreased by up to 31% with fly and bottom ash

17

treatments (Table 2), and for mixed wood ash decreases of up to 50% were observed, and interestingly the

tissue Ca concentrations decreased as ash addition increased (Table 1). Vector analyses diagnosed either

toxicity or antagonism for high dosages of mixed wood ash on all seedling species (Figures 3c – 5c).

Similar to K, the high levels of Na in the ash may have also induced antagonism of Ca, decreasing its

availability to seedlings at high dosages. Where vector analyses suggested toxicity, it is likely that another

nutrient was having a toxic effect on the seedlings (as opposed to a direct toxic effect of Ca) and causing

biomass to decrease, which was concentrating the foliar Ca. White pine seedlings had much higher foliar

Ca concentrations in the mixed wood ash treatments than the fly and bottom ash treatments (Tables 1, 2),

a pattern that is difficult to explain.

The sensitivity of the seedling species used in this experiment to soil Na varies, with white spruce and

yellow birch generally reported as being moderately tolerant of soil salinity (McKenzie, 1988), whereas

red pine is less tolerant and white pine is the least tolerant (Townsend & Kwolek, 1987). In this experiment,

white pine seedlings exhibited the most substantial negative effects from ash additions to foliar Ca levels

(Figures 5c, 7c; Tables 1, 2), supporting the theory that salts in the ash were having a negative effect on

seedling growth and health. As red pine is also sensitive to soil salinity, and yellow birch is only moderately

sensitive, it would be expected that red pine experienced antagonism more than yellow birch. However,

vector analyses for red pine did not indicate strong antagonism of K, or Ca, whereas for yellow birch

certain treatments did. Another common form of antagonism in plants is between K, Ca, and Mg. Indeed,

Mandre et al., (2010) reported decreases in foliar Ca levels in seedlings treated with ash, and suggested

this was due to competition with the abundant K supply. It is also possible that K was antagonising uptake

of Ca in this study, although the concentrations of K in the ashes did not seem disproportionately high

compared to Ca in any of the ashes (Table 3). Although vector analyses of white pine foliar nutrient status

in the fly and bottom ash treatments generally indicated luxury consumption of K and antagonism of Ca,

which does indicate K:Ca antagonism, this trend was not clearly evident in the mixed wood ash treatments

or for any other seedling species.

Foliar concentrations of Mg did not vary by more than 21% that of the control for any of the seedling

species in any treatment (Tables 1, 2), and vector analyses generally diagnosed either minor antagonism

or toxicity for all seedling species, particularly at the higher dosages of mixed wood ash (Figures 3d - 5d).

Mg is the next most available nutrient in ash after K and Ca (Eriksson, 1998), but increases in foliar Mg

levels of plants can take several years to be recognized (Reid & Watmough, 2014; Augusto et al., 2008).

Previous studies on seedlings report decreases (Mandre et al., 2010), no effect (Staples & Van Rees, 2001;

Kloseiko et al., 2014), and increases (Solla-Gullon et al., 2008; Mahmood et al., 2003) in foliar Mg

concentrations following ash additions. Mg uptake by plants can also be antagonised by Na, and at higher

18

dosages the Na concentrations in the ashes may have induced mild antagonism. Where vector analyses

suggested toxicity, it is likely that another nutrient was having a toxic effect on the seedlings and causing

biomass to decrease, which was concentrating the foliar Mg.

The solubility and subsequent availability of P in ash is lower than K, Ca, and Mg (Eriksson, 1998),

and the benefits to plants of increased soil P from ash additions often takes several years to be recognized

and can be highly variable (Augusto et al., 2008). Accordingly, foliar P concentrations were variable

between treatments and seedling species, differing by up to 42% that of the control. Vector analysis of

foliar P status was also variable between treatments and seedling species. For most seedling species there

were few large effects at lower and mid-range dosages of both mixed wood ash and fly and bottom ash,

except for red pine treated with mixed wood ash which exhibited luxury consumption (Figures 3a - 7a).

Higher dosages of mixed wood ash generally appeared to induce toxicity (Figures 3a - 7a). P availability

does not increase linearly with soil pH in acidic to neutral soils and decreases at pH values between 7.5

and 8.5 (Brady & Weil, 2010). Previous work suggests that the variability in P availability from ash arises

due to differences in soil type and pH buffering capacity (Erich & Ohno, 1992). In this experiment, the

higher dosages of wood ash may have increased soil pH to the range in which P is less available, and could

explain any decreases in foliar P. Where vector analyses suggested toxicity, it is likely that another nutrient

was having a toxic effect on the seedlings and causing biomass to decrease, which was concentrating the

foliar P. Previous studies of seeding responses to ash additions have reported decreases (Mandre et al.,

2004), increases (Mahmood et al., 2003; Mandre et al., 2010), and no effect (Staples & Van Rees, 2001;

Solla-Gullon et al., 2008; Kloseiko et al., 2014) of ash on foliar P levels of seedlings.

Foliar concentrations of Mn generally decreased, except for yellow birch in the mixed wood ash

treatments where increases of up to 71% that of the control (Table 1) were seen. Vector analyses generally

indicated antagonism, except for yellow birch treated with mixed wood ash where vector analyses

diagnosed luxury consumption (Figures 3e – 7e). Mn is most available in soil with lower pH values (Brady

& Weil, 2010), and as ash is known to increase soil pH, decreases in foliar Mn concentrations are expected

and consistent with current literature (Augusto et al., 2008). Iyer et al., 1971) reported average values for

foliar Mn concentrations in red pine seedlings of 182 mg kg-1, and the two lowest Mn concentrations in

red pine foliage in this experiment were 174.1 and 214.2 mg kg-1. Yellow birch seedlings are reported by

Hoyle (1972) to experience Mn deficiency below 60 mg kg-1, and toxicity above 1300 mg kg-1. For the

mixed wood ash treatments foliar Mn of yellow birch experiment were well within this range (Table 1),

but for the fly and bottom ash treatments ranged between 2730.5-4765 mg kg-1 (and was 4732.4 mg kg-1

in the control group), an order of magnitude larger than for the mixed wood ash. This discrepancy is

challenging to explain, although despite having potentially toxic levels of Mn, the seedlings’ growth

19

parameters did not appear to be affected (Figures 11c, 12c). In fact, the yellow birch seedlings in the fly

and bottom ash treatments had consistently higher biomass than those in the mixed wood ash treatments

(Figures 8d, 11c). Therefore, it is unlikely that Mn toxicity or deficiency drove the negative responses in

seedling biomass to ash additions at high dosages. It is also challenging to explain the increase in foliar

Mn concentrations of yellow birch treated with mixed wood ash. Another discrepancy in foliar Mn levels

occurred between mixed wood ash and the fly and bottom ash treatments for white pine, where Mn

concentrations ranged from 148-330 mg kg-1 in the mixed wood ash treatments, and from 848.9-1062 mg

kg-1 in the fly and bottom ash treatments. However, these were still within the reported critical range for

white pine of 100-5000 mg kg-1 according to Timmer (1991).

Unfortunately, the concentrations of other elements in seedling foliage beside N, P, K, Ca, Mg, and

Mn were not tested for, which makes it challenging to truly discern the cause for positive and negative

effects of ash on seedling foliar nutrient status. Had the foliar concentrations of Na been obtained, a more

accurate diagnosis of Na antagonism with K, Ca, and Mg would have been attainable. It is also possible

that at the higher ash dosages the soil pH was raised beyond the optimal range for growth of the seedlings

used in this experiment, which is 4.5-6.5 (Uchytil, 1991; Carey, 1993; Sullivan, 1994; Hauser, 2008). At

pH values greater than this, certain plant essential micronutrients become less available and deficiencies

can be induced (Brady & Weil, 2010). Finally, it is also important to consider the foliar concentrations of

metals, in order to eliminate the possibility of metal toxicity. Indeed, the higher concentration of potentially

toxic metals in fly ash has led some authors to disregard its use as a soil amendment (Pitman, 2006). Metal

concentrations were generally at, near, or below previously reported averages, except for Fe and As in fly

ash from both boilers which were up to ca. 150% and 550% higher respectively, and Cd which was 150%

higher than average in ash from boiler 2 (Table 3). However, none of the metal concentrations in the ashes

used in this experiment exceeded EPA or MOECC limits for land application of industrial wastes (Table

3). Additionally, the availability of most metals decreases in conjunction with increases in pH (Brady &

Weil, 2010), and the neutralizing effects of ash on soil pH have been shown to reduce the toxicity of metal-

contaminated soils (Perkiomaki et al., 2003). Excessive levels of As are known to disrupt plant metabolism

(Brady & Weil, 2010), and in this study it is possible that the drastically higher-than-average

concentrations of As had a detrimental effect on growth and health of seedlings even despite being within

the limits set by the MOECC and EPA, especially when ash was applied at high dosages. Previous ash

fertilization studies typically report that concentrations of metals in tree foliage remain low following ash

additions (Huotari et al., 2015). Minor increases in foliar Cu and Zn have been reported in foliage of Picea

abies (Osteras et al., 2005), and of Cd in select grasses and shrubs (Moilanen et al., 2002; Huotari et al.,

2011), but these changes occur shortly after application of ash, and tend to be transitory, returning to

20

previous levels or even lower several years following application. Metals are known to accumulate

differentially between the roots and shoots of plants (Malik et al., 2010), but no studies exist regarding the

effect of wood ash on heavy metal concentrations in plant roots (Huotari et al., 2015), and future studies

should focus on assessing the concentrations of metals in all compartments of ash-fertilized tree seedlings.

Overall, and as was expected, seedling specific differences in nutrient uptake were evident. There also

appeared to be differences in the uptake of some nutrients between fly and bottom ash and mixed wood

ash, and this is likely due to the differences in nutrient concentrations between the ashes (Table 3). The

higher mixed wood ash dosages of 15-20 Mg ha-1 may have induced negative effects due to excessive Na,

As, or other nutrient imbalances, whereas the mid-range dosages appeared to be most beneficial. Increases

in foliar K, Ca, Mg, and decreases in Mn, are expected to be a result of increased soil nutrient content due

to addition of nutrients via ash, and changes in nutrient availability due to increases in soil pH.

There are several factors that must be considered in this study that could have affected the responses

of seedlings to ash additions. Firstly, the nutrients in ash become available to plants at varying rates after

application, and if this study had been allowed to continue for more than 24 weeks then more of the benefits

that ash can provide may have been documented. For example, the P in ash is in a relatively slowly-soluble

form, and benefits of ash to soil P and plant P status can take several years to be recognized (Jacobson et

al., 2004; Clarholm, 1994 & 1998). In a meta-analyses of ash addition literature Reid & Watmough (2014)

found that the large variation in reported effects of ash to mature trees can be primarily attributed to

differences in the durations of the studies, with longer term studies being more likely to report beneficial

effects to tree growth. Secondly, the soil in this study was autoclaved prior to planting seedlings, which

destroyed the microbial biomass (Nieminen, 2009). Ash has been shown to stimulate microbial activity

due to increases in soil pH and dissolved organic C, which stimulates rates of N mineralization (Augusto

et al., 2008). Tree seedlings are also known to form symbiotic associations with mycorrhizal fungi, which

increase the supply of nutrients from soil to seedlings. Mahmood et al., (2002) isolated six species of

ectomycorrhizal fungi that were colonising roots of ash-treated Picea abies seedlings, which were then

used in a greenhouse fertilization experiment that explored the crossed effects of wood ash (unspecified

fly/bottom) and colonization by ectomycorrhizal fungi on seedling growth and health (Mahmood et al.,

2003). Mycorryhizal associations with seedling roots had a significant and substantial positive effect on

growth of seedlings treated with ash. Beneficial associations between seedlings and mycorrhizal fungi,

and N mineralizing microbes were not at play in this study, and may have prevented the seedlings from

capitalizing on the full suite of benefits that ash can provide. However, it is possible that during the course

of the study a few seedlings accidentally became inoculated with mycorryhizal fungi. Indeed, this could

explain the outliers that were removed from the white spruce dataset, that had biomass an order of

21

magnitude or more greater than most of the other seedlings. Mycorrhizae also tend to protect seedlings

against metal and salt stresses, alongside aiding in nutrient uptake. The potential positive and protective

effects of mycorryhizal fungi to seedlings treated with ash is an important future research question that

should be explored.

Seedling growth responses

There was a significant quadratic dosage effect of mixed wood ash on red pine and yellow birch

biomass and red pine leaf/needle area (Tables 4, 5), and this appeared to be driven by the negative effect

of ash at the highest dosage of 20 Mg ha-1, as there appeared to be little or no effect of ash at dosages of

1-15 Mg ha-1 (Figures 8a,d, 9a,d). There was also a significant linear effect of ash dosage on white pine,

in which higher dosages of mixed wood ash appeared to decrease seedling biomass and needle area

(Figures 8b, 9b; Tables 4, 5). However, although the most parsimonious model for the dosage response

was linear, there did not appear to be substantial decreases in seedling biomass or needle area below

dosages of 10 Mg ha-1 (Figures 8b, 9b). Although the effects of mixed wood ash on white spruce seedlings

were not significant (Table 4), there was a consistent mean increase in biomass and leaf/needle area across

all treatment groups (Figure 8c, 9c) that suggests ash additions had a positive effect. Ash dosage was not

a significant explanatory variable for any of the seedling species in the fly and bottom ash treatments

(Table 6), and this is likely because dosages up to only 10 Mg ha-1 were tested, and the dosage response

in experiment 1 was driven by the negative effects of ash at the highest dosages of 15 and 20 Mg ha-1.

There were also no statistically significant differences between effects of ashes from different boilers, or

the effects of fly and bottom ash on seedling biomass and leaf/needle area (Table 6). Therefore, it can be

concluded that irrespective of ash type or boiler type, and up to dosages of 10 Mg ha-1, ash had a neutral

effect on seedling biomass and leaf/needle area, presumably due to the low N concentrations in the ash

(Table 3). The decreases in biomass and leaf/needle area exhibited by yellow birch, white pine, and red

pine treated with higher dosages of mixed wood ash (Figures 8a-d, 9a-d) are expected to be due to either

the antagonistic effects of Na, or toxicity by certain metals (perhaps As).

Ash dosage also had a significant effect on red pine root:shoot ratios in the mixed wood ash treatments

(Table 4), which were lower (and closer to that of the NPK treated seedlings) at the mid-range dosages of

10-15 Mg ha-1 (Figure 10a). This implies that the nutrient status of red pine seedlings treated with ash at

10-15 Mg ha-1 was most optimal, which agrees with the previously discussed results where red pine foliar

K, Ca, and P status exhibited luxury consumption at mid-range dosages of mixed wood ash.

Seedlings treated with additional NPK fertilizer consistently had substantially larger biomass and

leaf/needle areas than all other treatments (Figures 8a-d, 9a-d), supporting the hypothesis that ash did not

22

have a significant positive effect on seedling biomass due to limited N supply. White spruce seedlings

showed the strongest response to the NPK treatment (Figures 8d, 9d), suggesting that they were the most

nutrient-starved compared to the other seedling species. Different tree species have different nutritional

requirements and were expected respond slightly differently to ash. Vance (1996) suggested that hardwood

trees may benefit more from ash additions due to their higher requirement of base cations compared to

softwoods. However, in this study yellow birch growth parameters or foliar nutrient status did not appear

to benefit substantially more than the conifers. It could also be expected that tree species which experience

forest fires as part of their natural ecology would be more adapted to periodic influxes of ash to the soil,

and would thus respond more positively to application of ash generated in biomass boilers. Forest fires are

an important part of white and red pine ecology, and these species have evolved to rely on forest fires for

successful establishment and competition (Carey, 1993; Hauser, 2008). Although yellow birch is not

specifically adapted to rely on fire, it is opportunistic and establishes readily in the years post-fire (Sullivan,

1994). White spruce is a mid-successional species, and although it does grow on soils that have burned in

the past, it does not generally colonize immediately after a fire (Uchytil, 1991). It does not appear that the

species with natural adaptations to forest fires had a more positive effect than those without in this study.

Although there were no significant differences between ashes from different boilers, or fly and bottom

ash, on seedling biomass and leaf/needle area, boiler type did have a significant effect on white pine

root:shoot ratio in the mixed wood ash treatments (Table 4) where ash from boiler 2 caused higher

root:shoot ratios than boiler 1 (Figure 10b). The root:shoot ratio of seedlings from boiler 2 were more

similar to the NPK treatment, suggesting that these seedlings were healthier than those treated with ash

from boiler 1. However, contrary to this presumption, boiler 2 also consistently decreased white pine

needle area and biomass more than boiler 1 (Figure 8b, 9b). The root:shoot ratios of white pine seedlings

was also influenced by ash type (Table 6), and were generally higher (and closer to that of the NPK

treatment) for fly ash (Figure 13b). The lack of differences between ash types and boiler types on seedling

biomass and leaf/needle area may be because despite having different pH values and concentrations of

mineral nutrients, the ashes all had similarly low quantities of N (Table 3), and were thus unable to

stimulate seedling productivity. However, changes in the nutrient balance of soils and seedlings

irrespective of N can manifest as changes in root:shoot ratio (e.g. Solla-Gullon et al., 2008). If the ashes

were applied in combination with an N fertilizer, differences in seedling biomass and leaf/needle area

between ash types and boiler types may have become apparent as the other nutrients would have become

limiting.

There is a discrepancy in previous literature regarding the safety of using fly ash as a soil amendment,

with Pitman (2006) concluding that fly ash should be avoided due to its higher concentrations of heavy

23

metals, but Perkiomaki et al., (2003) suggested using fly ash as a tool to remediate metal contaminated

sites due to its neutralizing effect on soil pH and subsequent decreases in metal availability. The results

from this study suggest that up to dosages of 10 Mg ha-1 fly ash can be safe to seedlings.

If future studies consistently find a lack of differences between effects of ashes from different boilers

and between fly and bottom ashes on seedling and tree health, regulation of ash additions in Canada will

be much simpler, as boiler-specific regulation would not be necessary. Current methods of regulating ash

additions in European countries often involve testing the ashes before application to ensure that nutrient

concentrations are within specified limits (Stupak et al., 2008). As biomass boiler technologies are

constantly evolving and becoming more and more efficient, precautionary testing of ash composition

before application is likely to be a necessity in regulation of ash additions to forest soils.

Effects of ash on mature trees are relatively well studied in Eastern Europe, but only a handful of

studies have investigated the effects of ash on growth parameters of seedlings, with somewhat mixed

results, and few have focused on Canadian species. Positive, neutral, and negative effects of ash on

seedling growth and health have all been reported.

Consistent with results from this study, most authors conclude that the lack of N in wood ash limits its

potential for increasing biomass of forest stands on mineral soils (Pitman, 2006; Augusto, 2008; Huotari

et al., 2008; Jacobson et al., 2014; Kloseiko et al., 2014; Huotari et al., 2015). For example, ash was found

by Nieminen (2009) to have no effect on Picea abies biomass and root:shoot ratios in greenhouse

experiment over 93 days. However, the effects of ash are generally found to be more beneficial to plant

productivity on other soils such as drained peatlands, which are rich in N but low in most other nutrients

(Ferm et al., 1992; Pitman, 2006; Huotari et al., 2008; Moilanen et al., 2013; Huotari et al., 2015). Certain

forest soils in eastern North America have also been found to be moving away from the state of N limitation

and towards either P or Ca limitation due to inputs of N from acid-rain (Gradowski & Thomas, 2006;

Watmough & Dillon, 2003). On these soils not only could wood ash be used to restore soil pH and nutrient

status, but could also increase tree growth and yield.

Although direct positive effects to seedlings productivity are generally not expected on mineral soils,

at dosages of up to 10 Mg ha-1 and over time periods of 1-5 growing seasons ash has been found to increase

seedling height, root collar diameter, and shoot length in Pinus sylvestris (Kloseiko et al., 2014), Picea

abies (Kloseiko et al., 2014; Nieminen, 2009), Betula pendula (Mandre et al., 2010), and Alnus glutinosa

(Parn et al., 2009), even in the absence of supplemental N. In this study the age of the seedlings, the

duration of the study, the soil type, and the autoclaving of the soil may have prevented more substantial

positive effects of ash on seedling growth parameters. Firstly, ash was applied after only the first set of

leaves/needles on the seedlings had emerged, and most previous studies that report positive effects from

24

ash to seedlings used growing stock that is 1 year old or more. Younger seedlings tend to be more

susceptible to changes in their environment, and the seedlings in this study may have been detrimentally

affected by ash due to potential as increases in soil salinity. Secondly, the positive effects of ash can take

several years to be recognized (as was discussed in the previous section; Reid & Watmough, 2014). Indeed,

Solla Gullon et al., (2008) explored the effects of ash (bark derived, fly and bottom ash mixed) at dosages

of 5 and 10 Mg ha-1 on growth of Pinus radiata seedlings, and only found significant effects on plant

growth after 5 years. If this study had occurred over a longer period of time, more of the longer-term

benefits that ash can provide to plants may have been realized and translated into to positive growth

responses. It is also known that the effects of ash to plants is highly dependent on soil type (Pitman, 2006;

Reid & Watmough, 2014; Huotari et al., 2015), and the low nutrient status, low C content, and sandy

texture of the soil may have prevented positive effects from ash. When applied to peat soils that are rich

in organic matter and N, ash has been shown to increase growth of Betula pendula (Mandre et al., 2010;

Parn et al., 2009), Alnus glutinosa (Parn et al., 2009), and Picea abies (Mahmood et al., 2003) seedlings

in the short term. Future studies should explore the effects of ashes on the growth and nutrition of seedlings

grown on a variety of soil types. Finally, and as was previously mentioned, another important consideration

is the autoclaving of the soil, which likely prevented microbial activity in the soil and symbiotic

associations between the seedlings and mycorrhizal fungi (Nieminen, 2009). These factors may have

prevented the seedlings from capitalizing on the full suite of benefits ash can provide to plants. Indeed, the

white spruce seedlings with drastically higher biomasses and leaf areas that were removed from the dataset

as outliers could have been benefitting from unplanned mycorrhizal inoculation.

Negative responses of tree seedlings to ash have also been reported in the literature. For example, in

an outdoor pot experiment conducted over 2 growing seasons, Madre et al., (2004) found that wood ash

(unspecified fly/bottom) applied at 10 Mg ha-1 caused nutrient imbalances and decreases in N and P

availability in the soil, which decreased the height of Picea abies seedlings, and dry mass of the shoots,

needles, and stems., Staples & Van Rees (2001) also found that a wood ash/sludge mix applied at 5 Mg

ha-1 decreased growth of Picea glauca seedlings due to salt phytotoxicity. However, they used a sludge/ash

mix applied to soils with poor drainage, which may have intensified the detrimental effects of ash and

could explain why negative effects were seen at lower dosages than in this study, where negative effects

from ash were only seen at 15-20 Mg ha-1.

The high initial pH of ash also has potential to negatively affect seedling health by being caustic to the

seedlings, particularly immediately after application (Dong et al., 2014). Average pH values for ash range

between 8-13, with a mean of 12 (Augusto et al., 2008). In this experiment, bottom ash from boilers 1 and

2 and fly ash from boiler 1 had pH values in the lower range of normal (Table 3), and fly ash from boiler

25

2 was in the higher range of normal. However, although the initial pH of fly ash from boiler 2 was 12.7,

this did not appear to have a negative effect on the seedlings 24 weeks after application.

Conclusions

Before being implemented as a soil amendment in Canada, the effects of ash on native tree species

must be assessed. This study provides an important view of the short-term effects of ash on the foliar

nutrient status and growth responses of tree seedlings native to Ontario, and is the first known experimental

attempt to discern the effects of ash dosage on growth and nutrition of seedlings.

Metal concentrations in all ashes were within the limits for land application of industrial wastes set by

the MOECC and EPA, although the Na and As concentrations in the ashes were relatively high.

Irrespective of ash type or boiler type, up to dosages of 10 Mg ha-1 ash had a neutral effect on seedling

biomass and leaf/needle area, presumably due to the low N concentrations in the ash. The root:shoot ratios

and foliar P, K, and Ca status of red pine suggested that ash dosages of 10 Mg ha-1 were optimum. At

dosages above 10 Mg ha-1 the biomass and leaf/needle areas of white pine decreased, and at 20 Mg ha-1

that of yellow birch and red pine also decreased. These negative effects are expected to be due to either

the antagonistic effects of Na, metal toxicity, or excessively high soil pH.

The young age of the seedlings used in this study, the short duration of the study, and the sterilization

of the soil via autoclaving may have prevented the seedlings from capitalizing on the full suite of benefits

that ash can provide, and longer term studies using non-sterilized soils are recommended. Heavy metal

accumulation in all seedling compartments (roots, stems, and shoots) should also be assessed.

This study tentatively supports the use of ash additions when applied to mineral soils at low dosages.

Although immediate benefits to seedlings may not occur on sandy and acidic soils, ash could be safely

used to counteract soil acidification and associated losses of base cations, and to redirect ash away from

to landfills. When applied to other soil types with higher organic matter and N contents, more immediate

positive effects of ash to seedlings may occur, and this is an important research question that should be

addressed in the context of North American soil types. In certain areas of eastern North America where

forest soils have been shown to be moving away from the natural state of N limitation and towards P or

Ca limitation due to acid rain, ash could neutralize soil pH, reinstate depleted nutrients, and may also have

a positive effect on forest productivity.

26

Tables

Table 1: Elemental composition of foliar tissues from a subsample of treatments 24 weeks after addition of mixed ash.

Tree Species Ash Type Ash Dosage Ca

(mg kg-1)

K

(mg kg-1)

Mg

(mg kg-1)

Mn

(mg kg-1)

P

(mg kg-1)

TN

(%)

Yellow Birch Control 0 25353.1 12128.3 7220.8 275.5 734.8

IPP 2 33113.7 11700.7 8782.0 632.4 761.9

10 23917.7 12998.6 5742.9 378.9 528.1

20 28956.2 10573.3 8280.0 682.4 1041.5

LUP 2 24095.3 16402.2 6282.9 395.6 660.8

10 32788.9 15343.5 8487.3 742.1 959.0

20 26181.1 11676.4 7561.2 644.1 739.3

Red Pine Control 0 5425.0 2160.4 1608.3 603.5 282.0

IPP 2 5649.6 3072.0 1536.3 581.9 363.4

10 7698.0 3773.1 1648.0 344.2 305.2

20 7135.2 4208.4 1555.6 174.1 338.3

LUP 2 4912.1 2609.3 1514.2 443.0 314.7

10 7823.3 4111.7 1594.5 283.2 312.2

20 6918.3 5981.6 1525.7 214.2 332.7

White Pine Control 0 30313.6 7896.8 2776.5 330.7 401.9 1.22

IPP 2 32612.0 8604.1 2634.0 148.0 389.9

10 20071.1 7345.9 2386.7 292.5 358.3 1.10

20 19998.7 11084.0 2224.2 236.2 427.8 0.85

LUP 2 34652.7 8063.7 2821.5 298.8 368.5 1.04

10 20460.3 7828.6 2675.6 222.6 366.3 1.02

20 15099.9 12814.0 2758.8 240.4 504.3

White Spruce Insufficient sample size

27

Tree

species

Boiler

type

Ash type Dosage Ca

(mg kg-1)

K

(mg kg-1)

Mg

(mg kg-1)

Mn

(mg kg-1)

P

(mg kg-1)

TN

(%)

Yellow

Birch

None None 0 20924.4 10716.0 5924.1 4732.4 700.0 0.83

IPP Fly 5 21964.7 12127.4 5010.1 3969.5 749.2 0.78

10 24359.6 14521.3 4954.9 2730.5 815.6 0.88

Bottom 5 20074.1 11706.2 5431.4 4765.0 796.7 0.79

10 21528.3 11860.9 6114.6 3631.6 735.9 0.80

LUP Fly 5 22203.5 11059.8 6764.1 3692.6 726.2 0.82

10 20789.9 12359.1 6341.6 4601.3 862.8

Bottom 5 21038.1 12592.8 5895.4 3666.5 723.0 0.78

10 22881.8 11624.5 5966.3 4682.4 852.4 0.68

White

Spruce

None None 0

IPP Fly 5

10 7692.2 7755.1 1902.3 1243.7 621.4

Bottom 5 7209.6 5670.4 1579.5 689.9 692.9

10 7296.3 7451.8 1779.1 1355.7 885.7

LUP Fly 5

10

Bottom 5 7896.3 7290.3 2018.2 1326.4 588.3

10 8271.0 6328.8 1616.0 881.1 612.9

White Pine None None 0 8127.0 6819.9 2306.4 916.4 432.2 1.36

IPP Fly 5 7793.0 9952.3 2332.5 877.9 431.5 1.30

10 9749.1 8668.2 2610.9 1012.7 613.8

Bottom 5 8023.4 6366.1 2549.3 887.1 523.2

10 8811.6 6516.4 2378.8 938.5 560.1 1.44

LUP Fly 5 9178.8 7904.8 2331.0 848.9 415.4 1.45

10 8184.4 8240.7 2659.3 988.2 582.6 1.26

Bottom 5 5603.1 9410.4 2274.9 1041.8 471.7

10 7929.1 10623.4 2580.0 1062.2 450.2

Table 2: Elemental composition of foliar tissues 24 weeks after addition of fly and bottom ash, from 2 separate boilers.

Missing values due to insufficient sample sizes

28

Boiler

1 Fly

Ash

Boiler 1

Bottom

Ash

Boiler 2

Fly Ash

Boiler 2

Bottom

Ash

Average values

for Fly ash1

(+/- 95%)

Average values

for Bottom

ash1 (+/- 95%)

EPA limits

for land

application2

MOECC limits

for land

application3

pH 9.26 8.93 12.67 8.31

Total N (%) 0.06 0.00 0.08 0.30

Total C (%) 5.52 0.18 4.60 33.23

Total S (%) 2.52 0.16 1.26 0.36 5.8 (3.5)

LOI (%) 8.97 0.20 2.70 40.41 20.0 (15.2)

P (%) 0.36 0.15 0.76 0.11 1.1 (0.1) 0.6 (0.2)

K (%) 2.67 2.22 6.43 1.56 5.9 (1.5) 3.9 (0.3)

Ca (%) 6.62 2.24 13.37 2.34 21.3 (3.7) 12.5 (2.3)

Mg (%) 0.71 0.56 1.56 0.35 1.9 (0.2) 1.5 (0.1)

Na (%) 2.41 1.38 1.39 8.61 3.12 (2.10) 1.10 (9.06)

Fe (mg kg-1) 19179.0 17765.7 24680.5 11208.9 9930 (5710) 17270 (9820)

Cu (mg kg-1) 99.7 39.5 104.9 16.1 81.8 (10.8) 62.7 (32.3) 4300 1700

Mn (mg kg-1) 7189.7 2530.6 11579.4 1993.4 11080 (2630) 5070 (520)

Zn (mg kg-1) 503.6 98.1 3330.8 106.7 2702.2 (1078.1) 7500 4200

Al (mg kg-1) 29891.6 38089.6 38043.5 25947.8 21920 (15780) 53570 (16300)

As (mg kg-1) 35.7 11.4 34.2 3.6 5.53 (2.8) 2.3 (2.5) 75 170

Ba (mg kg-1) 1298.2 650.3 1773.4 457.1 2359.0 (657.9) 1210.0 (1905.9)

Be (mg kg-1) 0.6 0.8 0.8 0.7 0.4 (0.3)

Hg (mg kg-1) 0.4 (0.2) 0.0 (0.1) 57 11

Cd (mg kg-1) 4.2 0.7 24.0 0.9 9.4 (3.1) 3.5 (13.8) 85 34

Co (mg kg-1) 10.7 8.1 16.0 6.4 10.5 (1.1) 9.9 (7.0) 340

Cr (mg kg-1) 58.6 29.2 46.5 18.5 66.1 (25.8) 82.7 (57.0) 2800

La (mg kg-1) 12.6 12.2 17.0 11.2 3.7 (4.6)

Li (mg kg-1) 15.8 17.7 25.7 11.8

Mo (mg kg-1) 2.4 0.8 5.9 1.0 7.8 (5.8) 2.8 (0.7) 75 94

Ni (mg kg-1) 18.5 13.0 28.2 10.0 80.4 (22.3) 43.0 (26.3) 420 420

Pb (mg kg-1) 16.9 16.8 68.3 4.4 40.0 (9.9) 53.0 (22.1) 840 1100

Se (mg kg-1) 0.7 0.0 13.1 1.0 100 34

Sr (mg kg-1) 270.5 133.8 575.1 138.5 910.6 (126.0)

V (mg kg-1) 40.1 38.4 58.5 24.6 35.0 (11.0) 31.7 (5.7)

Table 3: Elemental composition of ashes prior to application, regulatory limits for land application of industrial wastes set

by the MOECC and EPA, and average nutrient concentrations in fly and bottom ashes from boilers burning similar

feedstocks 1 - average values obtained from www.woodash.slu.se (an ash composition database) by selecting fly ash or bottom ash, from

all boiler types, and bark or wood-and-bark fuels. n varied from 3-11 for fly ash, and was 3 for bottom ash. 2 – EPA limits obtained from EPA (1993) 3 – MOECC limits obtained from OFIA (1999)

29

Biomass Leaf/Needle Ares Root:shoot

Seedling

species Variable SS DF F p SS DF F p SS DF F p

Red Pine

Ash dosage 0.00 1 2.43 0.1233 11.73 1 4.00 0.0487 0.60 1 6.85 0.0106

Ash dosage^2 0.01 1 4.38 0.0395 17.02 1 5.81 0.0182 0.60 1 6.84 0.0106

Boiler type 0.00 1 0.78 0.3786 0.78 1 0.27 0.6074 0.07 1 0.85 0.3605

White

Pine

Ash dosage 0.05 1 10.11 0.0021 51.08 1 9.31 0.0031 0.03 1 0.70 0.4069

Boiler type 0.00 1 0.00 0.9939 3.18 1 0.58 0.4485 0.29 1 6.06 0.0160

White

Spruce

Ash dosage 0.29 1 0.99 0.3222 0.09 1 1.25 0.2674 0.05 1 0.51 0.4780

Boiler type 0.24 1 0.85 0.3598 0.06 1 0.93 0.3383 0.00 1 0.02 0.8925

Yellow

Birch

Ash dosage 0.01 1 2.62 0.1095 63.42 1 2.29 0.1339 0.03 1 0.41 0.5223

Ash dosage^2 0.02 1 4.13 0.0455 97.88 1 3.54 0.0636 - - - -

Boiler type 0.00 1 0.02 0.8882 7.20 1 0.26 0.6112 0.13 1 1.72 0.1931

Table 4: Mixed wood ash seedling biomass, leaf/needle area, and root:shoot GLM results.

30

Dependent

variable Independent variables DF Red Pine

White

Pine

White

Spruce

Yellow

Birch

Biomass Ash dosage + Boiler type 2 -311.753 -203.222 136.7442 -196.271

Ash dosage + Ash

dosage^2 + Boiler type

3 -314.345 -201.229 138.6752 -198.372

Leaf Area Ash dosage + Boiler type 2 336.6162 384.3406 15.95660 522.9178

Ash dosage + Ash

dosage^2 + Boiler type

3 332.7620 385.0224 17.89280 521.3110

Root:shoot Ash dosage + Boiler type 2 42.04394 -14.7777 44.55788 55.39235

Ash dosage + Ash

dosage^2 + Boiler type

3 37.18987 -13.7892 45.35461 57.29254

Table 5: AIC scores for models using ‘ash dosage’ as a linear and a quadratic variable in the mixed wood ash

treatments. When the difference between two AIC scores was less than 1, the simpler model was chosen.

31

Biomass Leaf/Needle Area Root:Shoot

Seedling

species Variable SS DF F p SS DF F p SS DF F p

White

Pine

Ash

dosage 0.00 1 0.36 0.5521 1.40 1 0.16 0.6865 0.04 1 0.55 0.4614

Boiler type 0.00 1 0.04 0.8426 0.79 1 0.09 0.7615 0.19 1 2.54 0.1145

Ash type 0.01 1 1.04 0.3104 1.70 1 0.20 0.6560 0.43 1 5.63 0.0198

Ash type*

Boiler type 0.00 1 0.47 0.4963 16.18 1 1.89 0.1721 0.07 1 0.87 0.3524

Yellow

Birch

Ash

dosage 0.00 1 0.06 0.8070 42.38 1 0.61 0.4366 0.05 1 1.31 0.2546

Boiler type 0.01 1 0.66 0.4202 265.04 1 3.82 0.0538 0.03 1 0.87 0.3537

Ash type 0.00 1 0.00 0.9577 21.25 1 0.31 0.5814 0.02 1 0.58 0.4501

Ash type*

Boiler type 0.00 1 0.15 0.7009 155.23 1 2.24 0.1383 0.00 1 0.01 0.9131

White

Spruce

Ash

dosage 0.21 1 1.59 0.2117 0.10363 1 0.81935 0.368089 0.01448 1 0.4651 0.4972

Boiler type 0.02 1 0.12 0.7329 0.04517 1 0.35710 0.551811 0.01043 1 0.3349 0.5644

Ash type 0.00 1 0.01 0.9119 0.02246 1 0.17756 0.674611 0.01201 1 0.3858 0.5363

Ash type*

Boiler type 0.04 1 0.30 0.5843 0.11621 1 0.91875 0.340692 0.00895 1 0.2873 0.5935

Table 6: Seedling biomass, leaf/needle area, and root:shoot GLM results, for fly vs bottom ash treatments

32

Dependent

variable Independent variables DF

White

Pine

White

Spruce

Yellow

Birch

Biomass

Ash dosage + Ash type + Boiler type +

Ash type*Boiler type 4 -218.461 440.216 -144.459

Ash dosage + Ash dosage^2 + Ash type +

Boiler Type + Ash type*Boiler type 5 -217.066 440.216 -143.636

Leaf Area

Ash dosage + Ash type + Boiler type +

Ash type*Boiler type 4 483.244 70.235 677.259

Ash dosage + Ash dosage^2 + Ash type +

Boiler Type + Ash type*Boiler type 5 483.048 70.913 677.077

Root:shoot

Ash dosage + Ash type + Boiler type +

Ash type*Boiler type 4 30.116 -57.987 -38.796

Ash dosage + Ash dosage^2 + Ash type +

Boiler Type + Ash type*Boiler type 5 30.844 -56.152 -37.239

Table 7: AIC scores for models using ‘ash dosage’ as a linear and a quadratic variable in the fly and bottom ash treatments.

When the difference between two AIC scores was less than 1, the simpler model was chosen.

33

Figures

Figure 1: Relationships between seedling biomass, nutrient content, and nutrient concentration, with respect to vector

analysis. Reproduced based on Timmer & Stone (1978)

34

Vector

Direction

Change in

Foliar

Change in Foliar

Nutrient

Change in Foliar

Nutrient Interpretation Possible

diagnosis Mass Concentration Content

A 0 + + Luxury consumption Non-toxic

B + + + Deficiency Limiting

C + 0 + Sufficiency Non-limiting

D + - + Dilution Non-limiting

E +/- - +/- Depletion

F - - - Excess Antagonistic

G - + +/- Excess Toxic

Figure 2: Diagnosis of seedling nutrient status based on vector diagrams. Reproduced based on Timmer & Stone (1978)

G

F D

C

B

A

E

Rel

ati

ve

Nu

trie

nt

Co

nce

ntr

ati

on

(%

)

Relative Nutrient Content (%)

Relative Leaf/Needle Mass (%)

35

Yellow Birch Vector Diagrams: Boiler 1 vs boiler 2

Figures 3a-3e: Vector diagrams depicting nutrient status of yellow birch seedlings 24 weeks after addition of mixed ash. Red

and green circles are drawn around points from the highest and lowest addition rates from both boilers respectively. One

vector drawn for the highest addition rates.

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(a) Phosphorus

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(b) Potassium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(c) Calcium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(d) Magnesium

0

50

100

150

200

250

300

0 100 200 300

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(e) Manganese

36

Red Pine Vector Diagrams: Boiler 1 vs boiler 2

Figures 4a-4e: Vector diagrams depicting nutrient status of red pine seedlings 24 weeks after addition of mixed ash. Red and

green circles are drawn around points from the highest and lowest addition rates from both boilers respectively. One vector

drawn for the highest addition rates.

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(a) Phosphorus

0

50

100

150

200

250

300

0 50 100 150 200 250 300

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(b) Potassium

0

20

40

60

80

100

120

140

160

180

200

0 50 100 150 200

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(c) Calcium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(d) Magnesium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(e) Manganese

37

White Pine Vector Diagrams: Boiler 1 vs boiler 2

Figures 5a-5e: Vector diagrams depicting nutrient status of white pine seedlings 24 weeks after addition of mixed ash. Red

and green circles are drawn around points from the highest and lowest addition rates from both boilers respectively. One

vector drawn for the highest addition rates.

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(a) Phosphorus

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(b) Potassium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(c) Calcium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(d) Magnesium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(e) Manganese

38

Yellow Birch Vector Diagrams: Fly ash vs bottom ash

Figures 6a-6e: Vector diagrams depicting nutrient status of white pine seedlings 24 weeks after addition of fly and bottom

ash, from 2 separate boilers. Red and green circles are drawn around points from the highest and lowest addition rates from

all ash types. One vector drawn for the highest addition rates (but no vector drawn for Mg as there was not apparent

trend/direction)

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(a) Phosphorus

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(b) Potassium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(c) Calcium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(d) Magnesium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(e) Manganese

39

White Pine Vector Diagrams: Fly ash vs bottom ash

Figures 7a-7e: Vector diagrams depicting nutrient status of white pine seedlings 24 weeks after addition of fly and bottom

ash, from 2 separate boilers. Red and green circles are drawn around points from the highest and lowest addition rates from

all ash types. One vector drawn for the highest addition rates.

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(a) Phosphorus

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(b) Potassium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(c) Calcium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(d) Magnesium

0

20

40

60

80

100

120

140

160

0 50 100 150

Nutr

ient

Co

nce

ntr

atio

n (

% o

f co

ntr

ol)

Nutrient Content (% of control)

(e) Manganese

40

(a) Red Pine

Ash dosage (Mg ha-1)

Bio

mass

(g)

0 2 4 6 8 10 12 14 16 18 200.00

0.05

0.10

0.15

0.20

0.25

0.30

(a) White Pine

Ash dosage (Mg ha-1)

Bio

ma

ss (

g)

0 2 4 6 8 10 12 14 16 18 200.0

0.1

0.2

0.3

0.4

0.5

(a) White Spruce

Ash dosage (Mg ha-1)

Bio

ma

ss (

g)

0 2 4 6 8 10 12 14 16 18 200.0

0.1

0.2

0.3

0.4

0.5

(a) Yellow Birch

Ash dosage (Mg ha-1)

Bio

ma

ss (

g)

0 2 4 6 8 10 12 14 16 18 200.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

Figure 8a-8d: Biomass of seedling species 24 weeks after being treated with mixed ash from 2 separate boilers. 95%

confidence intervals shown. Horizontal dotted line drawn at the level of the control treatment.

41

(a) Red Pine

Ash dosage (Mg ha-1)

Lea

f A

rea

(cm

2)

0 2 4 6 8 10 12 14 16 18 200

2

4

6

8

10

12

14

16

18

(b) White Pine

Ash dosage (Mg ha-1)

Lea

f A

rea

(cm

2)

0 2 4 6 8 10 12 14 16 18 200

2

4

6

8

10

12

14

16

(c) White Spruce

Ash dosage (Mg ha-1)

Lea

f A

rea

(cm

2)

0 2 4 6 8 10 12 14 16 18 200

5

10

15

20

25

30

(d) Yellow Birch

Ash dosage (Mg ha-1)

Lea

f A

rea

(cm

2)

0 2 4 6 8 10 12 14 16 18 200

10

20

30

40

50

60

70

Figure 9a-9d: Leaf/needle areas of seedling species 24 weeks after being treated with mixed ash from 2 separate boilers.

95% confidence intervals shown. Horizontal dotted line drawn at the level of the control treatment.

42

(a) Red Pine

Ash dosage (Mg ha-1)

Root:

Sh

oo

t

0 2 4 6 8 10 12 14 16 18 200.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

2.0

(b) White Pine

Ash dosage (Mg ha-1)

Ro

ot:

Sh

oo

t

0 2 4 6 8 10 12 14 16 18 201.0

1.2

1.4

1.6

1.8

2.0

2.2

2.4

2.6

(c) White Spruce

Ash dosage (Mg ha-1)

Root:

Sh

oo

t

0 2 4 6 8 10 12 14 16 18 200.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

(d) Yellow Birch

Ash dosage (Mg ha-1)

Ro

ot:

Sh

oo

t

0 2 4 6 8 10 12 14 16 18 200.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

Figure 10a-10d: Root:shoot of seedling species 24 weeks after being treated with mixed ash from 2 separate boilers. 95%

confidence intervals shown. Horizontal dotted line drawn at the level of the control treatment.

43

(a) White Pine

Ash Dosage (Mg ha-1)

Bio

ma

ss (

g)

0 1 2 3 4 5 6 7 8 9 100.0

0.1

0.2

0.3

0.4

0.5

0.6

(b) White Spruce

Ash Dosage (Mg ha-1)

Bio

ma

ss (

g)

0 1 2 3 4 5 6 7 8 9 100.0

0.1

0.2

0.3

0.4

0.5

0.6

(c) Yellow Birch

Ash Dosage (Mg ha-1)

Bio

ma

ss (

g)

0 1 2 3 4 5 6 7 8 9 100.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

Figure 11a-11c: Biomass of seedling species 24 weeks after being treated with fly or bottom ash, from 2 separate boilers.

95% confidence intervals shown. Horizontal dotted line drawn at the level of the control treatment.

44

(a) White Pine

Ash Dosage (Mg ha-1)

Lea

f A

rea

(cm

2)

0 1 2 3 4 5 6 7 8 9 100

2

4

6

8

10

12

14

16

(b) White Spruce

Ash Dosage (Mg ha-1)

Lea

f A

rea

(cm

2)

0 1 2 3 4 5 6 7 8 9 100

5

10

15

20

25

30

(c) Yellow Birch

Ash Dosage (Mg ha-1)

Lea

f A

rea

(cm

2)

0 1 2 3 4 5 6 7 8 9 100

10

20

30

40

50

60

Figure 12a-12c: Leaf/needle area of seedling species 24 weeks after being treated with fly or bottom ash, from 2 separate

boilers. 95% confidence intervals shown. Horizontal dotted line drawn at the level of the control treatment.

45

(a) White Pine

Ash Dosage (Mg ha-1)

Ro

ot:

Sh

oo

t

0 1 2 3 4 5 6 7 8 9 100.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

2.0

2.2

2.4

(b) White Spruce

Ash Dosage (Mg ha-1)

Ro

ot:

Sh

oo

t

0 1 2 3 4 5 6 7 8 9 100.0

0.2

0.4

0.6

0.8

1.0

1.2

(c) Yellow Birch

Ash Dosage (Mg ha-1)

Ro

ot:

Sh

oo

t

0 1 2 3 4 5 6 7 8 9 100.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

Figure 13a-13c: Root:shoot of seedling species 24 weeks after being treated with fly or bottom ash, from 2 separate boilers.

95% confidence intervals shown. Horizontal dotted line drawn at the level of the control treatment.

46

Wood Ash as a Forest Soil Amendment: Responses of Red-Backed

Salamander (Plethodon cinereus) Abundance in a Northern Hardwood

Forest

Abstract

Wood ash may be an effective soil amendment in North America to restore acidified and low nutrient

forest soils, but little research exists beyond its effects on soil and plants. Eastern red-backed salamander

(Plethodon cinereus) abundance was assessed in a northern hardwood forest 1 year following an ash

addition field trial. Plots (400 m2) were established with fly and bottom ash treatments of 0, 1, 4, and 8

Mg ha-1 (n=4), and cover boards were positioned both with and without ash beneath. The following year,

8 salamander surveys were conducted and soil samples were collected and analyzed. Salamander

abundance, soil pH, and electrical conductivity (EC) increased under boards with fly ash beneath

compared to those without. At 8 Mg ha-1, 1 more salamander was counted (per plot, per survey), soil pH

increased by 2.0 units, and soil EC increased by 168.5 ms m-1. The moisture holding capacity of fly ash

was 60% higher than the soil, and for bottom ash was 63% lower. Bottom ash had no significant effect on

salamander abundance, and soil pH and EC. These results suggest that ash altered salamander abundance

via soil pH and moisture, and would not inhibit their movement over the forest floor.

Introduction

Forest soils in eastern North America have experienced acidification and nutrient depletions resulting

from acid rain (Likens & Bormann, 1974; Watmough & Dillon, 2003; McLaughlin, 2014). Low soil pH

and nutrient imbalances from acid rain persist (Driscoll et al., 2001; Gradowski & Thomas, 2006), and

forest harvesting also has the potential to deplete nutrients in certain forest soils (Phillips & Watmough,

2012). Declines in the growth and health of sugar maple (Acer saccharum Marsh.) have already been

recorded in response to soil acidification and Ca declines in northern hardwood forests (Horsley et al.,

2002; Long et al., 2011), and there is growing interest in using soil amendments to restore soil pH and

nutrient status (Long et al., 2011; Moore et al., 2012).

Wood ash is produced in significant quantities in pulp and paper and lumber mills in North America

(Elliott & Mahmood, 2006), and may be a practical forest soil amendment as it has a high pH, and other

than N, contains nutrients in similar proportions to those needed by growing trees (Mandre et al., 2010).

47

In certain European countries ash is already commonly applied to intensively managed forests to raise soil

pH and reinstate depleted nutrients (Emilsson, 2006; Stupak et al., 2008), and is also used as a soil

amendment in certain agricultural settings (Elliott & Mahmood, 2006). In Canada and the USA ash is

typically sent to landfill, but interest has been growing in using ash as a forest soil amendment (Elliott &

Mahmood, 2006; Pugliese et al., 2014). Production of ash is likely to increase in the future due to a growing

bioenergy sector (use of bioenergy has been projected to triple by 2050), with Canada as a major supplier

of biomass fuel (IEA, 2013).

Although ash is already used to increase soil pH and address nutrient deficiencies in agriculture and

intensively managed forests in Europe, its use in North American forests is not well studied. European

forests where ash additions are employed are intensively managed for timber production, and applying ash

to less intensively managed forests in North America may pose more significant risks. There are several

properties of ash that could harm susceptible forest species, especially when applied at high dosages. These

include a high pH, and retention of salts and traces of heavy metals (Pitman, 2006; Augusto et al., 2008).

Current literature focuses on soil and plant responses to ash additions, and little is known regarding its

effects on other components of forest ecosystems.

Biomass boilers typically produce both fly ash - a volatile, lighter, more reactive ash extracted from

flue gasses, and bottom ash – heavier, less reactive particles that fall to/remain on the bottom of combustion

chambers or grates. These ashes vary in their physical and chemical properties (Ingerslev et al., 2011), and

conclusions regarding their suitability as soil amendments vary. Precautionary research is required to

investigate the effects of these ashes on other components of forest ecosystems beyond plants and soils

before they are endorsed as forest soil amendments in North America.

Eastern red-backed salamanders (Plethodon cinereus) are one of the most abundant amphibian species

in northern hardwood forests, and their importance in many forest ecosystem functions such as litter

decomposition, carbon cycling, and biodiversity has been widely explored in the literature (Wyman, 1998;

Homyack et al., 2010; Hocking & Babbitt, 2014). Several factors make them good indicator species and

their responses to soil conditions and disturbances convenient to study. First, they are abundant; some

forests can sustain populations of more than 1 breeding pair in every 1m2, and their total biomass can

exceed that of all other small vertebrates combined (Semlitsch et al., 2014; Burton & Likens, 1975).

Second, they live beneath cover objects such as fallen logs, and their relative abundances can be monitored

with cover object surveys (Welsh & Droege, 2001; Moore, 2005). Third, red-backed salamander

populations are reasonably stable from year to year, and do not drastically fluctuate in the absence of

disturbances (Moore, 2005). Fourth, salamanders are relatively central in the forest food web, and can be

used as indicators of forest ecosystem health and integrity (Welsh & Droege, 2001; Davic & Welsh, 2004).

48

Finally, they respond quickly to environmental disturbances and changes, as they occupy the LFH layer

of soil and are both ectothermic and lungless, breathing directly through their skin (Welsh & Droege,

2001). Key environmental factors that influence the presence and abundance of red-backed salamanders

include soil pH, Ca, and moisture, and moist soils with a circumneutral pH and high Ca levels are preferred

(Sugalski & Claussen, 1997; Beier et al., 2012).

Although amending acidic northern hardwood forest soils with wood ash presents a practical

alternative to landfilling, research is needed regarding the impacts of ash on ecologically important forest

species such as red-backed salamanders. To our knowledge, no studies exist regarding the effects of ash

on salamanders or any other vertebrate communities. The neutralizing properties and Ca content of ash

may increase red-backed salamander abundance, however potential toxicants could have negative effects

at large dosages. This study assessed how fly and bottom ashes influenced soil pH, moisture, salinity, and

eastern red-backed salamander abundance (using cover board surveys), 1 year after application in a

northern hardwood forest. We also measured whether salamanders favored or avoided ash by residing

under cover boards with or without ash beneath.

Methods

Site description

The study site was located in Haliburton Forest and Wildlife Reserve, Central Ontario (45’11N,

78’35W), a mixed-deciduous Great Lakes-St. Lawrence forest (Rowe, 1972) dominated by sugar maple

(Acer saccharum) and American beech (Fagus grandifolia Ehrh.). The mean annual temperature in the

region is 5.0oC, ranging from -9.9oC in January to 18.7oC in July, with an average of 1074 mm of annual

precipitation (Environment Canada, 2014). Soils are shallow, rocky, and medium to coarse-textured and

belong to the Orthic or Eluviated Dystric Brunisol subgroups based on the Canadian System of Soil

Classification (Soil Classification Working Group, 1998). Much of the soil parent material is derived from

granite or granitic-gneiss deposits atop the Precambrian shield, with pH values ranging from 4 to 5.5. The

study site was harvested using the single tree selection silviculture system (OMNR, 2000) in 2003, and

subsequently salvage logged for beech due to the arrival of beech bark disease in the stand. Average basal

area in the study site in 2013 for trees >8 cm DBH was 16.4 m2 ha-1.

Plot setup and experimental design

49

In the summer of 2012 an area of forest was selected and 28 20m×20 m plots (including a 2.5 m

buffer) were established in a grid. Bottom and fly ashes were applied at rates of 1, 4, and 8 Mg ha-1 over 4

days in August, 2013. A control treatment (0 Mg ha-1 ash) and each ash application treatment were

replicated four times across the study area. To spread the ash evenly, each plot was sectioned into a 5×5

m grid, and the amount of ash needed for each square was weighed, carried to the plot using buckets, and

manually spread as uniformly as possible. Ashes for the trials came from a large RotoStoker VGC biomass

boiler system fired with bark after bole de-barking for pulp production (Detroit Stoker Company, Monroe,

Michigan, USA). The furnace had air and feedstock injected from above, and was equipped with a

horizontal vibrating stoker grate to move the fuel bed through the combustion chamber and remove the

bottom ash, which comprised of heavier, coarse, sandy particles. Fly ash represented a mix of material

recovered from both cyclone and electrostatic precipitators emissions control systems on the boiler.

Cover boards were cut to 30×30×2 cm from locally harvested, rough cut, untreated hemlock (Figure

14), and placed in each plot in August of 2013. Four cover boards were placed in each plot 1 week prior

to ash additions, evenly spaced along a diagonal transect running between opposite corners of each plot.

Another 4 were placed immediately after ash additions along the opposite transect, making 8 per plot: 4

with ash beneath, and 4 without. Cover boards were placed flat against the ground and avoiding rocks and

logs, then left to weather for 9 months before being monitored for salamanders.

Field and laboratory sampling and analyses

Three samples each of fly and bottom ashes were placed in sealable plastic bags for pH and elemental

analysis. Ash chemistry was determined using a NCS combustion analyzer (Vario EL III, Elementar

Americas, Mt. Laurel, NJ) for total C, N and S and with an inductively coupled argon plasma (ICAP)

spectrometer (Varian Analytical Instruments, Walnut Creek, CA) following high temperature microwave

acid digestion for P, K, Ca, Mg, Al, Na, S, Fe, Cu, Mn, As, Zn, V, Cd, Co, Cr, Mo, Ni, Pb, Ba, Be, Si, La,

Li, Se, and Sr (EPA standard method 3052).

The plots were assessed for canopy openness in July of 2013 and for soil moisture content in August

of 2014, to be used as covariates in statistical analysis. Soil moisture was determined by pooling 4

randomly located soil cores from the top 5 cm of the LFH layer of soil in each plot. Samples were sieved

through a 2 mm mesh, 10 g was weighed out, then dried in a drying oven at 90oC for 48 hours. The soil

was reweighed, and moisture content was calculated as: ((wet mass - dry mass) / dry mass) × 100. Canopy

openness was determined by taking 5 photographs at evenly spaced locations in each plot with a digital

camera facing vertically upwards and at 1.3 m above the ground, on standard settings. Photographs were

50

then imposed with a 5×5 grid, and assessed to determine the percentage of canopy cover compared to open

sky.

Salamander sampling occurred 8 times between May-October, 2014, always at least 1 week apart. The

order of plots visited was reversed every second sampling to avoid temporal sampling bias. To check for

salamanders each board was lifted, the number of salamanders was recorded, and the board was returned

to its original position. All sampling methods complied with the Guide to the Care and Use of Experimental

Animals (CCAC, 1993), and were approved by the University of Toronto’s Animal Care Committee.

In August of 2014, a 1 cm diameter soil core of the surface 5cm of the organic (LFH) layers of soil

were sampled from beneath each cover board, and from 4 randomly located soil cores in each plot. Samples

were pooled to obtain 3 samples per plot, providing 1 each from under boards with/without ash beneath,

and from uncovered soil. Soil moisture content was determined as described previously. pH and electrical

conductivity (EC) were measured after sieving soil using a 2mm mesh, then making a water:soil slurry

using a 4:1 ratio. The slurry was stirred, allowed to sit for 30 minutes, and stirred again before taking pH

and EC readings using an Accumet pH and EC meter (ThermoFisher, Waltham, MA, USA). The pH and

EC probes were calibrated after every 14 samples.

To assess the moisture holding capacity of ash, fly and bottom ashes were first oven dried at 90oC for

48 hours. Ten fine mesh bags (mesh opening size approximately 0.01 mm) were each then filled with

approximately 50 g of fly ash, and another 10 with 50 g of bottom ash. In October, 2014, 10 cover boards

were placed approximately 5m apart along a transect near the ash addition experiment. One bag each of

fly and bottom ash were placed beneath each cover board, and left for 2 weeks. The bags were then

retrieved, a 1 cm diameter soil core of the surface 5cm of the LFH layers of soil was taken from beneath

each board, and the moisture content of each ash and soil sample was determined using the same methods

described above.

Statistical analysis

The average number of salamanders observed per survey based on the pooled number of salamanders

found in each survey, for each plot, was used to represent salamander abundance. Salamander abundance,

soil pH, soil EC, and soil moisture content were the dependent variables. Normality was assessed using

normal probability plots of residuals, frequency histograms, and the Kolmogorov-Smirnov, Lilliefors, and

Shapiro-Wilks tests, and homoscedasticity of variances was assessed using the Cochran, Hartley, and

Bartlett tests (p>0.05). To achieve normality, salamander abundance was transformed using the square

root function, soil pH was transformed using 1/x2, and soil EC and moisture content were log transformed.

51

For salamander abundance, ‘ash type’ (fly and bottom ash) and ‘ash dosage’ (0 (control), 1, 4, and 8

Mg ha-1) were tested in a full factorial ANOVA, with ‘cover boards with/without ash beneath’ included as

a nested factor within ‘ash dosage’ and ‘ash type’, and interactive terms included between ‘ash type’ and

‘ash dosage’. Canopy openness and soil moisture content were included as covariates. For soil pH, EC,

and moisture content, ‘ash type’ (fly and bottom ash) and ‘ash dosage’ (0 (control), 1, 4, and 8 Mg ha-1)

were again tested in a full factorial ANOVA, with ‘cover boards with/without ash beneath, and uncovered

soil’ was included as a nested factor within ‘ash dosage’ and ‘ash type’, and interactive terms included

between ‘ash type’ and ‘ash dosage’.

Two comparisons are being made to test the null hypotheses that ash has no effect on salamander

abundance: 1) comparing treated plots to controls, and 2) comparing cover boards with ash beneath to

those without ash beneath within each treatment. Dunnett’s post-hoc test was used to determine differences

between treated plots and controls, and Tukey’s HSD test was used to determine all pairwise comparisons.

For soil chemistry data, soil from beneath cover boards in the control plot was used as the reference

treatment in Dunnett’s post-hoc test, as opposed to uncovered soil from the control plots.

To assess differences between moisture holding capacity of ash and soil, a one-way ANOVA was used

with ash/soil moisture as the dependent variable and sample type (fly ash, bottom ash, or soil) as the

independent variable. Tukey’s HSD test was then used to assess all pairwise comparisons. Statistica 7

(Statsoft inc, 2007) was used for all analyses, and all results were considered significant at p<0.05.

Results

Ash type, ash dosage, and presence of ash beneath the cover boards all had significant effects on soil

pH (Table 6). At the highest dosage of 8 Mg ha-1 boards with fly ash beneath increased pH from 5.2 to 7.2

when compared to the control plots, and from 5.0 to 7.2 relative to boards in the same plots without ash

beneath (Figure 15). Dunnett’s test found boards with fly ash underneath at 4 and 8 Mg ha-1 to be

significantly different to control whereas Tukey’s did not, but Tukey’s test did yield significant differences

between boards with/without ash beneath in the same treatments. Although not statistically significant, for

all fly ash treatments the pH of uncovered soil increased compared to soil from under boards without ash

beneath (Figure 15). Increases in soil pH under cover boards with ash beneath were ca. double those of

uncovered soil within the same treatment (Figure 15). Bottom ash did not significantly increase soil pH,

yet at 8 Mg ha-1 an average pH increase of 1.1 units occurred under boards with ash beneath compared to

those without (Figure 15).

52

Ash type and presence of ash beneath the cover boards had significant effects on soil EC (Table 6).

Presence of fly ash beneath the cover boards increased soil EC at all dosages compared to control plots,

and at 8 Mg ha-1 soil EC increased from 64-239 ms m-1 compared to the control plots (Figure 16), and

from 70-239 ms m-1 compared to boards without ash beneath in the same plots. Results from Dunnett’s

test corresponded with those from Tukey’s except for boards with fly ash underneath at 1 Mg ha-1.

Although not statistically significant, the EC of uncovered soil increased compared to soil from under

boards without ash beneath (Figure 16). Increases in soil EC under cover boards with ash beneath were ca.

double those of uncovered soil within the same treatment (Figure 16). Smaller and non-significant

increases in soil EC compared to control plots were seen in the bottom ash treatments (Figure 16).

The only significant effect on soil moisture of the LFH horizon was that soil moisture content was

consistently higher under cover boards when compared to uncovered soil (Figure 17), and although results

from Dunnet’s test did not always correspond with those from Tukey’s, but both support this result. When

comparing boards with/without ash beneath, or between treatments, there were no significant differences

(Figure 17). Although non-significant, bottom ash consistently caused minor decreases in soil moisture

content when comparing cover boards with/without ash beneath, and fly ash consistently increased soil

moisture content (Figure 17).

After 2 weeks of exposure to soil, fly ash had a significantly higher moisture content than the soil by

60%, whereas bottom ash had a significantly lower moisture content by 63% (Figure 18) (p<0.001).

One hundred and eighty two red-backed salamanders were counted in total. Ash type and presence of

ash beneath the cover boards were significant predictor variables for red-backed salamander abundance

(Table 7). Fly ash had a positive effect on salamander abundance, and at the highest dosage of 8 Mg ha-1

ca 1 more salamander was found under boards with fly ash beneath compared to both controls and boards

without ash beneath within the same treatment (Figure 19). Dunnett’s test found boards with fly ash

underneath at 8 Mg ha-1 to be significantly different to control whereas Tukey’s did not, but Tukey’s test

did yield significant differences between boards with/without ash beneath in the same treatment. Bottom

ash had no significant effect on salamander abundance (Figure 19, Table 7). Canopy openness between

the plots ranged from 12-60% (33% average), and soil moisture content ranged from 34-259% (93%

average), and these both had a significant effect on salamander abundance (Table 7). Including canopy

openness as a covariate also indirectly accounted for differences in both soil temperature and availability

of natural cover objects, which also influence red-backed salamander abundance (Grover, 1998) but were

not formally assessed. Varying amounts of dead wood (with both large and small diameters) between the

plots mostly arose from a recent beech salvage harvest, and plots with more open canopies had been more

53

intensively harvested and had more resulting downed woody debris or slash. Soil temperatures also vary

with canopy openness, as more sun exposure increases the temperature of soil.

Discussion

The liming effects of ash observed in this study are consistent with current literature (Demeyer et al.,

2001; Pitman, 2006; Augusto et al., 2008; Ingerslev et al., 2011; Reid & Watmough, 2014). The

neutralizing capacity of ash is based on its CaO/Ca(OH)2 and CaCO3 contents, and after addition the fast

dissolution of CaO/Ca(OH)2 causes a short-lived spike in soil pH (Dong et al., 2014), and the slower

dissolution of CaCO3 then causes longer term and sustained increases in soil pH which can take several

years to be recognized (Reid & Watmough, 2014). In this study soils were sampled 1 year after ash

additions, and for uncovered soils the initial pH spike caused by dissolution of CaO/Ca(OH)2 had likely

ended, and the effects of CaCO3 may have still not fully occurred , explaining the lack of significant effects.

Nevertheless, a consistent increase in soil pH with dosage occurred on uncovered soil for both ash types,

and at 8 Mg ha-1 fly and bottom ash increased soil pH by 0.8 and 0.5 units respectively compared to boards

that did not have any ash beneath (Figure 15). In a meta-analysis of ash addition literature, Augusto et al.,

(2008) report 1-5 year increases in soil pH to average 1.0 at dosages of 4-8 Mg ha-1, and a more recent

meta-analysis by Reid & Watmough (2014) found pH increases in organic soil horizons to average 1.04.

Similar and significant increases in the pH of uncovered soil may be observed in this study site in future

years. The larger and significant increase in soil pH under cover boards with ash beneath is because the

soil had been protected from direct rainfall and subsequent leaching of base cations and alkalinity lower

in the soil profile.

Fly and bottom ashes from the same boiler that was used in the present study have been previously

used in a soil incubation experiment where 5 Mg ha-1 of ash was added to a Brunisol collected from

Haliburton Forest (the same region that this study was conducted in), and incubated for 8 weeks (Pugliese

et al., 2014). Fly ash caused no significant change in soil pH, but contrary to this study bottom ash slightly

decreased soil pH. Other studies have also reported minor short term decreases in soil pH following ash

additions over similar timescales (Liimataineu et al., 2014).

The pH of the ashes used in this experiment were 8.6 and 9.7 for fly and bottom ash respectively (Table

8). These values are near the lower end of the ranges for the average pH of wood ash, reported by Vance

(1996) as 7.8-13.1 and Pitman (2006) as 11.7-13.1. In European forests where ash additions are employed,

the ash is often ‘hardened’ to decrease its pH before being used as a soil amendment, by leaving the ash

outdoors for 1-3 months during the summer (Dong et al., 2014). This is done as the high initial pH of un-

54

hardened ash can damage susceptible ground vegetation and fauna (Kellner & Weibull, 1998). If ash

additions are employed in North American forests, hardening (or reducing the pH by other means) is likely

to be an important component of reducing the potential negative effects of ash to ground flora and fauna.

The moisture holding capacity of bottom ash was 63% lower than the soil whereas for fly ash was 60%

higher (Figure 18), and although effects of ash on soil moisture content were not statistically significant,

fly ash consistently caused minor increases and bottom ash consistently caused minor decreases (Figure

17). The influence of ash on soil moisture content is based on the size of ash particles and their swelling

properties (Pugliese et al., 2014), and in this study the sandy consistency and low C content (Table 8) of

the bottom ash (large particles with limited swelling capacity) and the finer consistency and higher C

content of the fly ash (Table 8) likely drove their effects on soil moisture. Pugliese et al.’s (2014) soil

incubation study using ash from the same boiler as in this experiment also found fly ash to increase soil

moisture, and bottom ash had no strong effect.

Soil EC is often measured as a proxy for cation exchange capacity and salinity in soils (Brady & Weil,

2010; Khan, 2013). Many cationic nutrients are most available at circumneutral soil pH levels (Brady &

Weil, 2010), and as ash both supplies nutrients and neutralizes acidic soils, the increase in soil EC in

conjunction with pH in response to ash additions is expected and consistent with current literature (Staples

& Van Rees, 2001; Chirenje & Ma, 2002). The sodium content of ash has been suggested as a potential

toxicant in certain forest soils (particularly dry soils) by causing excessive salinity (Pugliese et al., 2014).

The typical Na values in ash vary highly, with Augusto et al., (2008) reporting a range of 2,000-5,000 mg

kg-1, but in a Swedish database of ash elemental composition the average Na levels in ash from bark (and

wood and bark mixed) feedstocks is 18,420 +/- 10,680 mg kg-1 (Karltun, 2015). Pugliese et al., (2014)

present the Na concentrations of fly and bottom ashes from 3 Canadian boilers, and found Na

concentrations above 18,000 mg kg-1 for both fly and bottom ash from 2 of the 3 boilers tested. It did not

appear that the salt content of the ashes in this study had a detrimental effect on salamander abundance,

and to our knowledge no other studies exist regarding the effects of soil salinity on red-backed salamander

health or populations.

The EC range indicative of salt toxicity to plants varies on the soil type and plant species, but for forest

soils an EC>400 ms m-1 can be considered detrimental (Khan, 2013), which was not reached in any of the

treatments in this study (Figure 16). However, other studies have suggested soil EC levels as low as 50 ms

m-1 can be detrimental to white spruce seedlings (Maynard et al., 1997). Staples & Van Rees (2001) found

that an ash/sludge mix with Na levels of 28,380 mg kg-1 applied at 5 Mg ha-1 increased soil EC from 20 to

100 ms m-1, decreasing the growth of white spruce seedlings after 2 growing seasons. In this study, soil

EC increased from 70 to 239 ms m-1 for fly ash at 8 Mg ha-1 (Figure 16), and more research is needed to

55

investigate the potential negative effects of elevated salt levels in ash on other aspects of the forest

ecosystem.

Red-backed salamanders prefer moist soil with a circumneutral pH (Sugalski & Claussen, 1997), and

the neutralizing effect of fly ash (Figure 15) and its higher moisture holding capacity compared to soil

(Figure 18) likely explains the observed increases in red-backed salamander abundance under cover boards

with fly ash beneath (Figure 19). Although the effects of ash on soil moisture content were not significant

(Table 6), the moisture holding capacity of the ash itself may be more reflective of the salamander’s

environment than the moisture content of the entire LFH horizon, as the salamanders occupying boards

with ash underneath were found in direct contact with the ash. For bottom ash, non-significant but

consistent increases in soil pH were seen (Figure 15) and its moisture holding capacity was lower than the

soil (Figure 18), and we suggest that these 2 potentially competing factors controlling salamander

abundance explain why there were not significant effects seen with bottom ash additions.

Although salamander abundance was greater beneath boards with fly ash beneath compared to those

without (Figure 19), the boards without ash beneath represent a more likely scenario in terms of how forest

management and operational ash applications would manifest on the ground. If forest application of ash is

to be employed on a large scale it is likely to occur immediately after harvests, as this decreases the cost

of application and would maximize the benefits to regenerating trees. During a harvest, large amounts of

downed woody debris are created, but a comparatively small amount is added in subsequent years. If ash

was applied following a harvest, there would be many logs without ash underneath (but concentrated

around the edges) that fell during the harvest, and very few with ash beneath. However, red-backed

salamanders move through the forest floor to defend and mark their territories, and hunt for food. As they

do not appear to avoid cover boards with ash underneath, we suggest that presence of ash on the forest

floor would not impede their ability to move or hunt.

To our knowledge, no studies exist regarding the effects of ash on salamanders, or any other vertebrate

communities, and responses of red-backed salamanders to other forest soil amendments has also been

minimally studied. Moore (2014) found no effect of finely ground and sandy textured CaCO3 when used

as a forest liming agent on red-backed salamander body weight and mortality in an eastern U.S.A.

hardwood forest, but the effect of lime texture was assessed as opposed to changes in soil pH, and

salamander abundance was not monitored. Forest liming has been shown to indirectly benefit songbird

populations, as the increases in soil pH and Ca levels increase snail populations, which are a staple food

source of many songbirds. A similar effect of ash on red-backed salamanders could also occur, as wood

ash has been shown to increase abundances of collembolans, mites, and enchytraeid populations in soil

(Nieminen et al., 2012), which are food sources of red-backed salamanders. In untreated northern

56

hardwood forests, abundances of red-backed salamanders have also been shown to increase in conjunction

with soil Ca levels (Beier et al., 2012).

The potentially high initial pH of ashes that have not been hardened could be caustic to salamanders

(and other ground fauna and vegetation) immediately after application. Until the initial pH drops,

salamanders may avoid ash by utilizing cover objects without ash beneath, or by burying into the soil.

Although the ash used in this experiment had a lower than average pH, the observed positive effects of ash

on salamander abundance may still have only occurred after its pH had equilibrated with the soil. This is

an important future research question that should be addressed, for example with behavioural “choice

tests” in controlled soil mesocosm environments using salamanders or other soil fauna (e.g. McTavish et

al., 2013).

Heavy metals in ash could also have a detrimental effect on salamander health and abundance.

Concentrations of heavy metals in ashes used in this study (Table 8), and in ashes from a variety of other

biomass boilers in Canadian mills (Pugliese et al.,2014) are generally low, and their mobility in forest

ecosystems is not normally a concern as the higher pH of ash renders them less available to plants

(Perkiomaki et al., 2003). Land application of industrial waste residues is regulated federally in the USA

by the Environmental Protection Agency (EPA), and provincially in Canada by the Ministry of

Environment and Climate Change (MOECC) in Ontario. For both ashes used in this study, metals were

within EPA and MOECC limits (Table 8) and are unlikely to have been affecting salamander abundance.

However, the mobility of metals increases at soil pH values below 5.5, and if soil pH returns to pre ash

addition levels at some point in the future then the metals from ash could pose a toxicity issue, highlighting

the importance of long term studies.

It is important to consider that this study was assessing relative abundance of red-backed salamanders

between treatments and is not a representation of their absolute abundance in northern hardwood forests,

of which there has been much attention in current and past literature (Burton & Likens, 1975; Semlitsch

et al., 2014). As mentioned previously, the observed increases and decreases in salamander abundance

could have actually been a redistribution of their location in the soil horizons. Changes in absolute

abundance may take longer to be recognized, unless ash directly causes mortality of salamanders which

was not observed in this study.

Conclusions

Fly ash beneath the cover boards increased soil pH and had a higher moisture holding capacity than

the soil, causing increases in red-backed salamander abundance. Bottom ash under the cover boards had

57

no significant effect on soil pH, EC, moisture, or salamander abundance, but had a lower moisture holding

capacity than soil. The salamander boards with ash beneath did not represent a common habitat scenario

from a management perspective, but the finding that salamanders do not avoid boards with ash beneath

suggests that ash would not prevent their movement on and through the forest floor. Both ashes had

relatively high Na levels but soil EC did not indicate excessive soil salinity. Heavy metal concentrations

in the ash were minimal and did not appear to negatively affect salamander abundance. These results

tentatively support use of ash as a soil amendment in acidified hardwood forests of eastern North America,

but more research is needed into the longer-term effects of ash and mobility of its potential toxicants. This

study provides just one aspect of the evidence needed to make an informed decision on whether ash

addition is an acceptable practice in North American forests.

58

Tables

Table 6: p values from nested ANOVA for soil pH, EC, and moisture

Variables DF pH EC (ms m-1) LFH moisture content (%)

1 - Ash type 1 0.0187 0.0108 0.8121

2 - Ash dosage 2 0.0099 0.0713 0.5378

3 - Cover boards with/without ash beneath,

or uncovered soil (1*2)

13 0.0003 0.0000 0.0001

4 - Ash type*Ash dosage 2 0.5523 0.2441 0.2542

59

Table 7: p values for nested ANOVA for salamander abundance

Variables DF p

1 - Ash type 1 0.0001

2 - Ash dosage 2 0.1158

3 - Cover boards with/without ash beneath (1*2) 6 0.0059

4 - Ash type*Ash dosage 2 0.6113

5 - Soil Moisture Content 1 0.0473

6 – Canopy Openness 1 0.0001

60

Table 8: Elemental composition of fly and bottom ash (means; n=3), and untreated Haliburton soil* (0-20 cm depth

including LFH), and EPA† and MOECC‡ limits for metals.

Element Fly ash Bottom ash Untreated soil EPA limits for

land application

MOECC limits for

land application

pH 8.63 9.67 4.08

Total C (%) 17.57 0.52 7.8

LOI (%) 20.42 0.43 13.76

TN (%) 0.09 0.00 0.46

P (%) 0.38 0.17 0.07

K (%) 3.07 1.43 1.97

Na (%) 3.64 1.63 2.1

Al (%1) 2.33 4.47 6.58

Fe (%1) 1.54 2.82 0.35

Ca (%) 10.11 4.36 1.77

Mg (%) 0.87 0.84 0.45

S (%) 4.22 0.29 0.00

Mn (%) 0.80 0.32 0.04

Ba (%) 0.17 0.09

Co (mg kg-1) 10 13 15 340

As (mg kg-1) 10 12 13 75 170

Be (mg kg-1) 0 1

La (mg kg-1) 10 18

Li (mg kg-1) 12 22

Cu (mg kg-1) 52 38 34 4300 1700

Pb (mg kg-1) 21 15 55 840 1100

Cd (mg kg-1) 6 1 3 85 34

Hg (mg kg-1) 57 11

Se (mg kg-1) 10 3 5 100 34

Mo (mg kg-1) 3 1 3 75 94

Zn (mg kg-1) 691 140 74 7500 4200

V (mg kg-1) 40 63

Cr (mg kg-1) 28 33 31 2800

Ni (mg kg-1) 16 20 11 420 420

Sr (mg kg-1) 387 218 0

*Pugliese et al., (2014); †EPA (1993); ‡OFIA (1999)

61

Figures

Figure 14: Rough-cut hemlock cover board in bottom ash treated plot. The red four indicates that it was the 4th board with

ash beneath in the plot

62

Figure 15: pH of LFH beneath cover boards with/without ash beneath, and for uncovered soil. 95% confidence intervals

shown. Significant differences to control (Dunnett) indicated with a *, and significant differences from all pairwise

comparisons (Tukey) indicated with differing letters

3

4

5

6

7

8

0 1 4

Bottom

8 1 4

Fly

8

pH

Ash Treatment (Mg ha-1)

Uncovered soilBoards without ash beneathBoards with ash beneath a*

ab

bc bc

bc

c

c c

c c

c

abc

abc

abc

abc abc

abc

abc

abc

abc

63

Figure 16: Electrical conductivity (EC; ms m-1) of LFH beneath cover boards with/without ash beneath, and for uncovered

soil. 95% confidence intervals shown. Significant differences to control (Dunnett) indicated with a *, and significant

differences from all pairwise comparisons (Tukey) indicated with differing letters

0

50

100

150

200

250

300

350

400

450

0 1 4

Bottom

8 1 4

Fly

8

EC

(m

s m

-1)

Ash Treatment (Mg ha-1)

Uncovered soilBoards without ash beneathBoards with ash beneath a*

ab*

abc*

c c c c

c c

c bc

c

bc bc c

c

c

abc abc

abc

64

Figure 17: LFH moisture content (%) beneath cover boards with/without ash beneath, and for uncovered soil. 95%

confidence intervals shown and bound at 0. Significant differences to control (Dunnett) indicated with a *, and significant

differences from all pairwise comparisons (Tukey) indicated with differing letters

0

100

200

300

400

500

600

700

800

0 1 4

Bottom

8 1 4

Fly

8

So

il m

ois

ture

(%

)

Ash Treatment (Mg ha-1)

Uncovered soil

Boards without

ash beneathBoards with ash

beneath

*a

b

ab

ab

ab ab

*ab

*ab

ab

ab

*ab

ab ab

ab

ab

ab ab ab

ab ab

65

Figure 18: Moisture content of fly ash, bottom ash, and soil, after ashes had been dried and left under cover boards for 2

weeks. 95% confidence intervals shown. Significant differences of pairwise comparisons (Tukey) indicated with differing

letters

0

20

40

60

80

100

120

140

160

Soil Bottom ash Fly ash

Mo

istu

re c

on

ten

t (%

)

Treatment

a

c

b

66

Figure 19: Average number of salamanders found per sampling event, per plot. 95% confidence intervals shown and bound

at 0. Significant differences to control (Dunnett) indicated with a *, and significant differences from all pairwise comparisons

(Tukey) indicated with differing letters

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2

0 1 4

Bottom

8 1 4

Fly

8

Av

era

ge

nu

mb

er o

f sa

lam

an

der

s

Ash treatment (Mg ha-1)

Boards without ash beneath

Boards with ash beneatha*

b

b

b

b

b

b b

b

ab

ab

ab

ab

67

General Conclusions

The general objectives of this thesis were to begin determining whether ash is a safe and effective soil

amendment when applied to acidified and low nutrient forest soils in eastern North America. The seedling

fertilization experiments demonstrated that at lower range dosages (up to 10 Mg ha-1) and regardless of

the type of ash (fly or bottom) and the boiler it came from, ash has relatively neutral effects on seedling

growth and can benefit foliar nutrient status. Longer term effects of ash are expected to be more positive,

as the benefits of ash to forest soils often take several years to transpire. The salamander study

demonstrated that wood ash increases soil pH and electrical conductivity following additions, and

depending on the effects of ash on soil moisture, this has either neutral or positive effects on the

abundances of red-backed salamanders at dosages up to 8 Mg ha-1. As red-backed salamanders are an

ecologically significant species in forests of eastern North America and have been suggested as good

bioindicators of forest health, the neutral and positive effects of ash on their abundance supports use of ash

in these forests. Together, the results from the seedling trials and the field trials provide 2 pieces of

evidence supporting the notion that in the short term, ash can be a safe and effective soil amendment on

acidified and low nutrient soils of eastern North America.

Although there were no differences in the effects of fly and bottom ash or ashes from different boilers

on the growth of seedlings, they did have different effects on the root:shoot ratios of certain seedlings,

which presumably occurred due to differences in the nutrient contents of the ashes. Fly and bottom ash

also differentially effected salamander abundance, presumably due to their differences in moisture content.

Further research is necessary to determine whether fly and bottom ash can be used interchangeably, or

whether they should be differentiated, or mixed. Either way, precautionary testing of ashes before they are

applied to forest soils is likely to be a necessity if ash additions are to become a regular occurrence. In

particular, the salt and metal contents of ash, and its pH should be monitored, as these have potential to

induce negative effects.

Although this thesis provides tentative evidence to support ash additions, there are several limitations

that still need to be addressed. The main limitation is that only 1 soil type was experimented with in both

the seedling trials and the salamander study, and therefore the conclusions made in this thesis are specific

to the soil types that were experimented with. Future studies should investigate the effects of ash on

seedlings growing on other soil types, and on ecologically significant vertebrate species that reside on

other soil types. Another limitation is that only the short term effects of ash were tested, and the effects of

ash often take several years to be realized. Nevertheless, many of the potential negative effects that ash

can induce would occur in the short term, and it is the positive effects that usually take longer to be realized.

Therefore, although this thesis does not provide evidence of the potential long-term beneficial effects that

68

ash could have on forest health, it has begun to eliminate the possibility of short term negative effects

when ash is applied to acidic and low nutrient forest soils. When ash is applied to other soil types with

higher organic matter and N contents, more immediate positive effects to plant productivity may occur,

and this is an important research question that should be addressed in the context of North American soil

types. In certain areas of eastern North America where forest soils have been shown to be moving away

from the natural state of N limitation and towards P or Ca limitation due to acid rain, ash could neutralize

soil pH, reinstate depleted nutrients, and may also have a positive effect on forest productivity. In future

years, the field trial that was established in which the salamander study was conducted should provide data

to validate this presupposition.

69

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