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Abstract

Microbiological Quality of Packaged Ice from Various Sources in Southern California

Introduction

Microbial contaminations in food and water may post a threat to public health. Ice is defined as a food by the U.S. Food and Drug Administration (FDA). According to the International Packaged Ice Association (IPIA), approximately 2 billion bags of ice are sold from retail, wholesale, and vending producers each year in the U.S. Out of 700 commercial ice-making companies, 200 of the aforementioned are not represented by the IPIA and do not comply to specific packaged ice processing standards. Potential sources of microbial contamination in ice may come from water, equipment, and handlers. Non-IPIA complied samples were collected from gas stations, liquor stores, or convenient stores in Los Angeles and Orange counties. The microbiological quality of non-IPIA complied ice samples were compared the results with the IPIA-complied packaged ice samples using microbiological, molecular, and sequencing analyses. Among the 132 non-IPIA complied packaged ice samples analyzed, 15 samples contained unsatisfactory level of heterotrophs (≥ 500 Most Probable Number [MPN]/100 ml), 12 samples contained unsatisfactory level of coliforms (≥ 1 MPN/100 ml), 19 samples had Staphylococci, and 70 samples had yeast/molds. None of the 24 IPIA-complied samples had unacceptable microbial levels. None of the samples analyzed showed the presence of the pathogen, Salmonella. Our results revealed the microbiological quality of non-IPIA and IPIA complied ice samples in Southern California. These findings may lead to a better enforcement of processing standards on packaged ice.

Phillip M. Lwin, Stefan F. Riedel, Kun Justin Lee, Liana S. Ab Samad, Ashley Magin, and Wei-Jen Lin Biological Sciences Department, California State Polytechnic University, Pomona

Materials and Methods

Each sample was transferred to a sterile WhirlPak

Each sample was transferred to another sterile WhirlPak and melted overnight

Heterotrophic count(TPC)

Filter 100 mL of each sample

Food handler’s hygiene

Staphylococcus aureus

Baird-Parker agar

Fecal contamination

Total ColiformsEscherichia coli

MI agar

Intestinal pathogens

Salmonella sp.

Real time PCR

Bacterial diversity and prevalence

Next generation sequencing(NGS)

ResultsIce is a staple presence in the American household and is now produced on an industrial

scale, for many uses including food processing and storage, chemical manufacturing, concrete mixing and curing. To make easy consumption for consumers, packaged ice became popular.

According to the International Packaged Ice Association (IPIA), approximately 2 billion bags of ice are sold from retail, wholesale, and vending producers each year in the U.S. The Association of Food and Drug Officials (FDA) have published good manufacturing practices (GMPs) to regulate the sanitary manufacturing of packaged ice. All ice manufacturing companies are required to follow these basic GMPs but are enforced differently from state to state. The IPIA has published the Packaged Ice Quality Control Standards (PIQCS) manual, which is based on GMPs but is tailored specifically to packaged ice. According to IPIA, aproximately 200 out of the 700 commercial ice-making companies do not comply with IPIA specific packaged ice processing standards, which can lead to potential contamination in ice from water, equipment and handlers. The high consumption of ice by consumers makes microbial contamination in ice packages a threat to public health.

A recent study performed at the University of Georgia showed that manufactured ice had much better microbiological and chemical quality as compared to the packaged ice from retails and vending machines in Georgia (Mako et al, 2014). The IPIA requested a similar, smaller scaled study to be done for Southern California.To scale down the study, we would like to focus the study on the 6 counties in Southern California‐ Los Angeles, San Bernardino, Riverside, Orange, San Diego, and Imperial. The findings of microbiological quality of IPIA-standardized versus non-IPIA packaged ice will potentially lead to a better enforcement of processing standards on packaged ice.

(A) (B) (C)

(D) (E) (A) & (B): Non-IPIA retailer-packaged machine and ice(C): In store bagger (ISB) machine(D) & (E): IPIA-manufactured ice

(A)

Figure 1. (A) Percentage of samples within acceptable limits sorted by county. (B) Enumeration of Staphylococcus sp. from manufactured packaged ice and various counties. (C) Enumeration of indicator organisms using the most probable number conversion. (*) indicates acceptable limit of heterotrophic growth

Conclusions

Future Studies•  Report findings to IPIA and expand studies to other institution in the United States•  Increase samples size to represent each county populations

Acknowledgements•  This project was fully funded by IPIA•  Metropolitan Water District La Verne: vacuum apparatus

•  All manufactured ice passed the acceptable limits for microbiological quality set by IPIA.•  San Diego/Imperial county had the lowest acceptable percentage of coliform.Orange county had the

most number of plates that contained Staphylococcus species and yeasts/molds.•  Proteobacteria phylum was the most dominant microbial community in our packaged ice samples. •  Four samples from LA, 2 samples from OC, 3 samples from SB/R, and 4 samples from SD/Imperial

had unacceptable number of heterotrophic counts (>500 MPN/mL). Samples from SD/I counties have the highest average heterotrophic counts, followed by Orange County and LA County.

•  Salmonella was not detected in any of the samples.•  There was no significant microbiological quality difference between ice produced by IPIA-ISB and

non-IPIA ISB machines.•  Overall, non-IPIA retailer-packaged ice showed poor microbiological quality compared to IPIA-

manufactured and in-store-bagged ice.

4. Manufactured and ISB ice has higher overall quality than retailer-packaged ice based on IPIA PIQCS standard for ice

100 100

78.8

Figure 4. (A) Percentage of acceptable ice samples that meet the IPIA PIQCS standards. IPIA standards: <500 MPN/100 mL and no coliform growth. (B) Percentage of acceptable ice samples from retailer packaged ice sorted by different counties

2. Examples of test results

Figure 2. (A) Total plate count method. Pink wells indicate positive. (B) Staphylococci colonies on Baird-Parker agar. (C) Growth of coliforms on MI agar showed fluorescence when applied long UV wavelength.

UV

(A) (B) (C) (F) (G)

(F) Sample size represented by the counties (G) Sample size based on various sources.

1. Microbial quality of on-site packaged ice from different counties

(C)(B)

3. Microbial community analysis by next generation sequencing (NGS)

(A)

(C) Figure 3. (A) Alpha rarefaction curves showing the richness of bacterial species. The three samples, On-site A, On-site B, and ISB, are shown in red, blue and orange lines respectively. (B) Microbial communities analysis based on a Bray-Curtis distance matrix, observed-OTUs, and Chao1. (C) Bacterial community compositions at the phylum level

Phylum On-siteA On-siteB ISBAcidobacteria 0.19% 0.14% 0.00%Actinobacteria 6.44% 0.41% 0.01%Bacteroidetes 2.37% 1.37% 0.00%Cyanobacteria 0.69% 0.28% 0.00%Firmicutes 0.01% 1.51% 0.01%Nitrospirae 0.00% 0.55% 0.00%Planctomycetes 0.01% 0.00% 0.00%Proteobacteria 89.96% 91.87% 99.98%Verrucomicrobia 0.26% 3.88% 0.00%WPS-2 0.08% 0.00% 0.00%

(B)

24

25 27

21

LA OC SB/R SD/I

(A) (B)