Guidance for Submission of AMR Surveillance Isolate for External

Quality Assessment and Reporting Emerging AMR Alerts

National AMR Surveillance Network

NCDC

National Programme on Containment of Antimicrobial Resistance National Centre for Disease Control, New Delhi

March 2019

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Table of Contents

I. Expectation from AMR surveillance laboratories ................................................................................................. 2

II. Preparation of nutrient agar media stabs ............................................................................................................... 2

III. AST results reporting ............................................................................................................................................ 3

IV. Standard Precautions: ............................................................................................................................................ 3

V. List of strains to shipped to NCDC for External Quality Assessment (EQAS) .................................................... 4

VI. Isolate packaging and transportation according to International Air Transport Association’s Dangerous

Goods Regulations (IATA-DGR) ......................................................................................................................... 5

VII. Checklist for sending EQAS or ALERT strains to NCDC ................................................................................... 7

VIII. List of antibiotics to be tested for priority bacterial pathogens before shipping the isolates to NCDC.............. 8

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Guidance for Submission of AMR Surveillance Isolate for External

Quality Assessment and Reporting Emerging AMR Alerts.

I. Expectation from AMR surveillance laboratories

i. All participating laboratories should enrol with IAMM EQAS and share copy of the EQAS

reports and scores to NCDC every 4 months

ii. All participating laboratories are expected to submit 1% of isolates for each pathogen under

surveillance (as mentioned on page 4) to NCDC every QUARTER for confirmatory testing as

per below mentioned timeline.

a. Quarter 1 isolates (January 1 to March 31) should be submitted by APRIL 15

b. Quarter 2isolates (April 1 to June 30) should be submitted by JULY 15

c. Quarter 3 isolates (July 1 to September 30) should be submitted by OCTOBER 15

d. Quarter 4 isolates (October 1 to December 31) should be submitted by JANUARY 15 of next

year

iii. All AMR alert pathogen isolates has to be shipped to NCDC, New Delhi for confirmation and

further guidance.

a. The strain identity should be confirmed before sending to NCDC

b. The confirmed isolate has to be reported immediately

c. Alert isolate should reach NCDC within one week of confirmation

iv. All the bacterial isolates for NCDC EQAS or Emerging AMR Alert should be sent in nutrient

agar media stabs in 1.5mL/2mL sterile plastic vials and a duplicate stock should be preserved

at the reporting lab.

II. Preparation of nutrient agar media stabs

i. Prepare and autoclave nutrient agar media. Ensure media contains 1.5% agar in it.

ii. Dispense 1ml of Nutrient agar media in airtight, autoclaved 1.5 mL/2.0 mL plastic vials

(approx. 2/3 full). (Autoclavable cryovials are ideal for this, if available)

iii. Stab Inoculate the media with the pure colonies from fresh culture using a sterile straight wire

and incubate overnight at 37°C.

iv. Next day, seal the plastic vials tightly and cover the edges using parafilm.

v. Label the isolates as Lab ID or Sample ID/Lab code/month/Year.

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For e.g. sample ID is18004325; Lab code is GMC; the organism was submitted in the month of

October 2019 (October/2019); then the label on the strain should be

18004325/GMC/OCT/2019.

III. AST results reporting

i. Report AST results only in zone diameters/MIC values and not as interpretation (RIS).

ii. Send the list of isolates for EQAS in prescribed EXCEL format with the packet containing

isolates.

iii. Email the soft copy of isolate list to AMR surveillance secretariat at

amrsurveillance@gmail.com

iv. For Alert strains, send duly filled alert form along with the isolates both in hard and soft copy.

v. Ensure that the AST is performed for ALL the listed antibiotics in the surveillance AST panel

for each isolate sent to NCDC for EQAS or ALERT reporting and confirmation

vi. All labs should report MIC values based on broth microdilution AST for

a. Colistin AST for all gram negative priority pathogens (E. coli, Klebsiella species,

Acinetobacter species and Pseudomonas species), isolated from blood or other sterile

body fluids.

b. Vancomycin AST for Staphylococcus aureus isolated from blood and other sterile body

fluids

c. Linezolid AST for Staphylococcus aureus and Enterococcus species isolated from

blood and other sterile body fluids

IV. Standard Precautions

The persons performing the culture and AST should follow “standard precautions” and wear

PPEs (Mask, glove, gown, close-toed shoes) while handling the isolates for packaging"

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V. List of strains to shipped to NCDC for External Quality Assessment (EQAS)

Pathogen Resistant strain (1%)

+

AMR Alerts (All)

Sensitive strains (1%) Intermediate

Resistant strain

(All)

Enterococcus

species

Vancomycin (All)

Linezolid (All)

Vancomycin (1)

Linezolid (1)

Staphylococcus

aureus

Vancomycin (All)

Linezolid (All)

Vancomycin (1)

Linezolid (1)

Vancomycin

(All)

Escherichia coli Imipenem (1)

Colistin (All)

Imipenem (1)

Colistin (All)

Klebsiella species Imipenem (1)

Colistin (All)

Imipenem (1)

Colistin (1)

Acinetobacter

species

Imipenem (1)

Piperacillin Tazobactum (1)

Colistin (All)

Minocycline (1)

Imipenem (1)

Piperacillin Tazobactum (1)

Colistin (1)

Pseudomonas

species

Imipenem (1)

Piperacillin Tazobactum (1)

Colistin (All)

Imipenem

Piperacillin Tazobactum (1)

Colistin (1)

Salmonella enterica

serotype Typhi

Ceftriaxone (All)

Ciprofloxacin (All)

Azithromycin (All)

Imipenem (All)

Ceftriaxone (1)

Azithromycin (1)

Imipenem (1)

Ceftriaxone (All)

Azithromycin

(All)

Imipenem (All)

Salmonella enterica

serotype Paratyphi

Ceftriaxone (All)

Ciprofloxacin (All)

Azithromycin (All)

Imipenem (All)

Ceftriaxone (1)

Azithromycin (1)

Imipenem (1)

Ceftriaxone (All)

Azithromycin

(All)

Imipenem (All)

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VI. Isolate packaging and transportation according to International Air Transport

Association’s Dangerous Goods Regulations (IATA-DGR)

i. The specimens or isolates of priority AMR pathogens are covered by UN 3373 shipment

procedures and IATA packing instruction 650.

ii. Packaging must be of good quality, strong enough to withstand the shocks and loadings

normally encountered during transport. Packaging must be so constructed and closed so as

to prevent any leakage or loss of contents during transport.

iii. The packaging must consist of three components: (Refer to Image 1, Page 6)

a) A primary container: This can be a screw-capped container or cryovials. If multiple primary

containers are used they should be individually wrapped or separated to prevent contact.

Adequate absorbent material must be packed around the primary receptacle/s to absorb any fluid

leakage from the primary receptacle/s.

b) A leak-proof secondary container/packaging: The primary container/s and adsorbent material

are put into a leak-proof secondary container. Sealed plastic bags (zip lock bag) is a good

alternative for the recommended secondary container. An itemized list of contents and

requisition forms must be enclosed in a separate sealed bags (zip lock bag). Pre-frozen ice-packs

should be packed around the secondary container.

c) A rigid outer packaging/box [Tertiary Container]: The secondary container containing the

primary container must be put into a shipping container with adequate cushioning material. At

least one surface of the outer packaging must have a minimum dimension of 100 mm x 100

mm. The outer packaging must be marked with UN 3373 and ‘Biological substances,

Category B’ label adjacent to the diamond shaped mark.

Image 1:Triple packaging recommended by

IATA

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Image 2: Zip-lock bags: Alternative for the secondary container used commonly in resource poor settings

Image 3: The UN labels for the Tertiary container for specimen/isolate transport/referral

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VII. Checklist for sending EQAS or ALERT strains to NCDC

Sl

No

Things to do before shipping isolates and alert strains to NCDC Please tick

1. Observe the growth of culture after overnight incubation inside the vial ☐Yes ☐No

2. Vials sealed tightly ☐Yes ☐No

3. Vials edges covered with Parafilm ☐Yes ☐No

4. Labelled neatly and correctly as listed ☐Yes ☐No

5. Hardcopy of list of isolates for EQAS with the AST results in zone

diameters/ MIC enclosed as per the format provided by NCDC

☐Yes ☐No

6. Culture vials label & isolate list enclosed with the package cross

checked

☐Yes ☐No

7. Triple layer packaging done as recommended by IATA ☐Yes ☐No

8. AMR alert reporting form (Annexure 3)* is completely filled and duly

signed by the AMR nodal officer (Applicable only when the lab detects emerging antimicrobial resistance of public health

importance)

☐Yes ☐No

9. Package labelled with legibly written “From address” and “addressed

To”

Incharge AMR Programme

Bacteriology and Drug resistance testing lab,

Division of Microbiology,

National Centre for Disease Control,

22 Shamnath Marg,

New Delhi – 110054.

☐Yes ☐No

10. Scanned copy of the list of isolates and their AST details sent to

amrsurveillance@gmail.com

☐Yes ☐No

*As referenced in page 17-18 AMR Surveillance SOP February 2019.

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VIII. List of antibiotics to be tested for priority bacterial pathogens before shipping the isolates to NCDC

Organism

Sp

ecim

en

Antibiotics

Am

ika

cin

30

μg

Am

pic

illi

n 1

0 μ

g

Azi

thro

my

cin

15

μg

Cef

epim

e 3

0 μ

g

Cef

ota

xim

e 30

μg

Cef

ox

itin

30 μ

g

Cef

tazi

dim

e 30

μg

Cef

tria

xo

ne

30

μg

Ch

lora

mp

hen

icol

30

μg

Cip

rofl

ox

aci

n 5

μg

Cli

nd

am

yci

n 2

μg

Do

xy

cycl

ine

30

μg

Ery

thro

my

cin

15μ

g

Gen

tam

icin

10

μg

Gen

tam

icin

Hig

h 1

20

μg

Imip

enem

10

μg

Lin

ezo

lid

30

μg

Min

ocy

clin

e 30

μg

Nit

rofu

ran

toin

300

μg

Pip

era

cill

in/T

azo

ba

ctu

m

10

0/1

0 μ

g

Tet

racy

clin

e 3

0μ

g

TM

P/S

MX

*

Va

nco

my

cin

30μ

g

Va

nco

my

cin

scr

een

Ag

ar

Co

list

in M

IC

Va

nco

my

cin

MIC

Comments

Enterococcus species B P O**

✓

✓ ✓

✓

✓

✓

Enterococcus species U ✓

✓

✓ ✓

✓

✓

Staphylococcus aureus B, P,O

✓

✓ ✓ ✓ ✓ ✓

✓

✓

✓

✓

Escherichia coli B, P, O

✓

✓ ✓

✓

✓

✓

✓

Escherichia coli U

✓

✓ ✓

✓

✓

✓

✓

Klebsiella species B, U, P, O

✓ ✓

✓

✓

✓

✓

Acinetobacter species B, U, P, O

✓ ✓

✓

✓ ✓ ✓

✓

✓

Pseudomonas species B, U, P, O

✓ ✓

✓

✓

✓

✓

✓

Salmonella enterica

serotype Typhi B

✓ ✓

✓ ✓ ✓

✓

✓

Salmonella enterica

serotype Typhi S

✓

✓ ✓ ✓

✓

✓

Salmonella enterica

serotype Paratyphi B

✓

✓ ✓ ✓

✓

✓

*TMP/SMX- Trimethoprim Sulfamethoxazole 1.25/23.75 µg; **B – Blood, U- Urine, O – Other sterile body fluids, S- Stool. All tick marked (✓) antibiotics to

be mandatorily tested before shipping to NCDC

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