results of the 2018 16s rrnagene sequencing and/or maldi ...nmgroup.ca/document/2018/2018_15.pdf ·...

Results of the 2018 16S rRNA Gene Sequencing and/or MALDI-TOF

National Challenge Test**Sponsored by the NMG and the NML**

Kathryn Bernard and Mark UngerNML- CSCHAH Winnipeg MB

National Molecular Group Annual Meeting

Hospital for Sick Children, Toronto ONDecember 4, 2018

Preparation for the 2018 (7th) Test• Project management done by Mark Unger

(Proteomics Core Facility)

• Test listed on NML Guide to Services/SpecBact

• Previous contacts & ‘new’ respondents were contacted in August 2018 in order to:

–Confirm names/contact info

–Provide provisional dates for shipment

–Arrange for sign off a ‘Transfer Application for Pathogens and Toxins’

–Get courier acc’t no.- to pay for shipment 2

The Panel • Panel of 5 taxa sent using transport media on Oct. 23

or 24, 2018 (each delivery confirmed by Mark).

• Panel consisted of:

– 4 facultative anaerobe or aerobes; BCYE (Legionella agar) also sent (1st attempt to assess member of this genus)

– 1 ‘Bonus Bug’, for testing by MALDI only

• Due date: Friday Nov. 9 2018 (~15 days for work), typo on cover email (Nov. 7, 2018) due date noted/corrected

• No. sites enrolled & provided results = 65• No. different participants from all sites = 69, that is,

multiple participants for same assay from 2 sites• No. sites in 2017= 57; 2016: 47

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Met Due Date; No. by test method• Due date: Nov. 9, 2018 • No. sites which provided results by due date:

= 63/65 (97%) (same as 2017). No. sites which did:

• 16S rRNA gene sequencing only = 5• Both 16S & MALDI-TOF = 17 [TOTAL 16s = 22 SITES]

• 16S and Bruker = 10 • 16S and Vitek = 7• Did MALDI-TOF only = 43 (Bruker= 22, Vitek =21)• Did MALDI (N=60)

• BRUKER= 32/60 (53%); VITEK=28/60(47%)

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Summary of Participant Methods-1 • Panel isolates selected expected to have

(relatively) unambiguous IDs • ‘Best’ results shown for sites with > 1 participant• Beta testing, 16S: done using in-house methods

at 2 separate sites (Sept. 2018)• Beta testing, MALDI-TOF: done separately on

Bruker Microflex and Vitek MS (Sept 2018)• 16S rRNA methods: most sites provided partial seq • Small number = provided nearly full sequence • Only 2 sites cited accession no. for ref. seq (scored

similarity) against which unknown was compared5

Summary of Participant Methods-2, MALDI • Process: most used DIRECT method• Some used ‘on-plate-extraction’, especially for

samples 1 and Bonus Bug• One applied ‘bead beating’ prior to extraction

(adapting a mycobacterial procedure) • ‘Tube extraction’ or ‘extraction’ described esp

for samples 1 and Bonus Bug• Extraction done in second round, if direct failed

to get any peaks or had poor response esp samples 1 and Bonus Bug

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Software / Library vers used; Taxonomy References • BRUKER: 4.1.1, 4.1.7, 4.1.8 (7) or not described• Biotyper: 6903 (2); 7311 (3); 7854 (5);rest not cited• Vitek ver 4.4; 4.4.0.3; 4.5.1-2; 4.5.1-2; 4.6.1• Most did not explicitly state if result was from

IVD or the RUO (Saramis) • Presumed IVD here unless otherwise stated• Ver. 3.0 (6); Ver 3.2 (2) otherwise ver not citedTaxonomy References: from the List of Prokaryotes with Standing from Nomenclature (LPSN) found at http://www.bacterio.net/ or from relevant literature

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N=65 sites. MALDI=Black; 16S only=Blue; 8 new• AB N=5 (0 Bruker, 4 Vitek); N= 1; Total = 5 • BC N= 11 (9 Bruker, 2 Vitek); N= 0; Total = 11

• MB N= 10 (7 Bruker, 1 Vitek); N= 2; Total = 10 (6 NML) • NB N=3 (1 Bruker, 2 Vitek); N= 0; Total = 3

• NL N=1 (1 Bruker, 0 Vitek); N= 0; Total = 1 • NS N=1 (0 Bruker, 1 Vitek); N= 0; Total = 1

• ON N=22 (12 Bruker, 9 Vitek); N= 1; Total = 22 • PE N=2 (2 Bruker, 0 Vitek); N= 0; Total = 2

• QC N = 7 (0 Bruker, 7 Vitek); N= 0; Total = 7 • SK N = 3 (0 Bruker, 2 Vitek); N= 1; Total = 3

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No. Sites by Province [Territories= 0]

Lab primarily provides Diagnostics for:

Human Disease: 62Animal Disease: 3

• 16S rRNA gene sequencing: 2 sites, 2 participants (1 NML) produced/interpreted sequence

• MALDI: 2 sites (one Bruker Microflex [NML, extracted]) one Vitek MS (direct method)

Result Triage/Formats of Results Tables:• Rec’d, (scanned) Excel (M Unger); proof-read (KB)

• Beta test results presented first, followed by participant

• Results subdivided by platform

• Summary of other IDs described in table or comments

• ‘ID would be reported as’: discussed in comments

• Other relevant observations described in comments9

Beta Tester Methods Used

2018-ECT-1 Nocardia cyriacigeorgica• NML identifier 160983, recovered from sputum• Formerly part of ‘Nocardia asteroides complex’• Described in 2001 IJSEM 51:1419-1423 as ‘N

cyriacigeorgici’; Latin corrected to N. cyriacigeorgica• Considered as possibly the most commonly recovered

Nocardia from human disease (2015 MCM 11th pg 516) • 1st Nocardia sent in a panel • Slower growing (48-72h), typical Nocardia colonies • CW of Nocardia:~more difficult to break open(esp for

MALDI): here, 50% = direct, 50% extracted (includes tube extracted, bead, Direct FA)

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ECT-1 Nocardia cyriacigeorgica: NML 160983• 16S: unambiguous ID found here by 16S, as strain had

“>99.6% and ≥ 0.4% separation from TS other species” (ref: Petti et al, MM18-2nd ed re: Nocardia (2018 CLSI MM18-2ed pg 99-103)

• NML often uses secA1 seq as 2° target for Nocardia. • Had 100% to N cyriacigeorgica DSM 444484T (NR_041857)

98.9% N. abscessus IMMID-D-1592T (NR_025059)• However, caveat discovered while reviewing results

(ID= asteroides / cyriacigeorgica):• N. asteroides ATCC 14759 (EF127493) 100% to 160983• ATCC 14759 (N asteroides, original deposit)= NCTC 8595 (N

cyriacigeorgica) = DSM 43208 (N cyriacigeorgica), latter two based on analyses of additional target genes

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ECT-1 ID = Nocardia cyriacigeorgica NML 160983Results by Beta Testers, September 2018

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*averaged among 3 replicates; extraction procedure used # Biotyper ver 7854 used$IVD ver 3.0 used; direct procedure used

Method RESULT Interpretation of ID as…

16s (TESTER 1) 1450/1450 (100%)

DSM 44484T

(NR_041857)

N. cyriacigeorgica (unambiguous)

16S (TESTER 2) 686/686 (100%)

686/686 (100%)

N. cyriacigeorgicaN. asteroides ATCC 14576

ID as Nocardia sppMALDI (1-Bruker*,#) N. cyriacigeorgica

but scores ~1.60

Nocardia spp; noted that

extraction good but no ID due

to low scores obtained

MALDI (2-Vitek MS$) N. cyriacigeorgica 99.% confidence

N. cyriacigeorgica (unambiguous)

ECT-1 ID = Nocardia cyriacigeorica NML 160983

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Response & Interpret ID as[% to number in category]

16S@ Partial / Full seqONLY(N= 5)

MALDI and/or BRUKER16S@ + Bruker = 10Bruker only = 22(N=32)

MALDI and/or VITEK16S@ = 7 (both)Vitek only = 21 (N=28)

N. cyriacigeorgica and REPORT as N. cyriacigeorgica

4/5 (80%) 16S: 6/10 (60%)MALDI*: 9/32 (28%), scores 1.98-2.06

16S: 5/7 (72%) MALDI*: 14/28 (50%); scores all 99.9%

N. cyriacigeorgica but REPORT as ‘N. cyriacigeorgica complex’ or Nocardia spp #

1/5 (20%) 16S: 4/10 (40%)

MALDI: 19/32 (59%)scores 1.6-2.14

16S: 1/7 (14%)

MALDI: 11/28 (39%) scores all 99.9%

N. asteroides / N cyriacigeorica

0/5 (0%) 16S: 0/10 (0%)MALDI: 0/32 (0%)

16S: 1/7 (14%)MALDI: 0/28 (0)%

Clostridium spp C.beijerinckii C. sporogenes

NO ID/No peaks

0/5 (0%) 16S: 0/10 (0%)MALDI: C. beijerinckii 1/32(3%) (score 1.36) 2/32 (6%)

16S: 0/10 (0%)MALDI: C. sporogenes 1/28 (4%) score 99.9% (??)NO ID = 2/28 (7%)

@ some would refer out or sequence a secondary target gene; *Many would refer out to reference centre to confirm; # cited ‘complex’ OR ‘slashline’ between various Nocardia species OR interpreted as (possible) Nocardia sp /genus

Summary, ECT-1, Nocardia cyriacigeorica• 16S: complicated as some N. cyria reference strains still

had older “N asteroides” name in GB. 2° target gene not mandatory here but recommended

• ‘Nocardia cyriacigeorica complex’ described by some labs but does not seem to exist in literature..(older ‘N. asteroides complex’ does exist)

• MALDI MSPs (Bruker) Correct ID came up but many reported ‘Nocardia sp’ type answer

• MSPs (Vitek) Correct ID came up and was reported by Vitek users 50% of the time; others used ‘Nocardia sp’

• Several labs had no ID or provided spurious taxa as ID

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ECT-2 Legionella bozemanae NML 180064 • L. bozemanae (originally L. bozemanii, 1980) here from

bronchial wash; rare respiratory pathogen• NML: 24 referrals, all clinically relevant, since 19781

• NML 180064 was 1st Legionella sent in a CT panel; we provided BCYE plate for those labs which normally do not ever work with legionella

• Assigned to new genus Fluoribacter, as F. bozemanae in 1980 but as discussed in LPSN:Genus name Fluoribacter Garrity et al. 1980 is a correct name (Principle 6). However, the names Fluoribacter Garrity et al. 1980, ¤ Fluoribacter bozemanae Garrity et al. 1980 (the type species of the genus), ¤ Fluoribacter dumoffii (Brenner et al. 1980) Brown et al. 1981, and ¤ Fluoribacter gormanii (Morris et al. 1980) Brown et al. 1981 have never found widespread usage.

(We opt to use Legionella bozemanae)1. Pachco and Bernard 2018 AMMI-CACMID Conference Vancouver BC Poster no. 0049

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ECT-2 Legionella bozemanae • Challenge = species difficult to ID by 16S alone• Some 16S accession nos. for (nearly full seq) L. bozemanae

ATCC 33217T

(=WIGAT=NCTC 11368

T=serogroup 1), are of

very poor quality; (nearly full) sequences are not represented in NCBI-curated database, i.e.,

• M36031 (ATCC 33217T): 87 ambiguous bps (6%), but cited in

LPSN; Z49718 (for ATCC 33217T) had 93% similarity to above

• Z49719 (ATCC 35545= serogroup 2= ‘Toronto 3’)

• 180064 had 99.0% similarity to Z49719 and 98.9% to

Z49718 BUT 100% using mip gene sequencing to these

serogroups; NML ID based on MIP seq• 180064: 99.6% over 518 bps (NR_125584) and to

draft genome NCTC 11368T

(access. No UGGY01)16

ECT-2 ID = Legionella bozemanae NML 180064Results by Beta Testers, September 2018

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*averaged among 3 replicates; extraction procedure used # Biotyper ver 7854 used;@ Bruker Biotyper has not modified MSPs to L. bozemanae from L bozemanii$IVD ver 3.0 used; direct procedure used

Method RESULT Interpretation, ID as…

16s (TESTER 1) 99.0% NCTC 11369 (Z49719); 98.9% NCTC 11368T (Z49718)

L. bozemanae with comment that ID is always confirmedusing mip sequencing

16S (TESTER 2) 99.0% (L bozemanae)97.8%-97.9%) (L steelei, L. santicrucis, L anisa)

Legionella species; would refer to ref centre for species

MALDI (1-Bruker*,#)

Legionella bozemanii score 2.41-2.44

Legionella bozemanii (sic@)(unambiguous)

MALDI(2-Vitek MS$)

Legionella bozemanaeconfidence value 99.9%

Legionella bozemanae(unambiguous)

ECT-2 ID = Legionella bozemanae NML 180064

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16S@

Partial / Full seqONLY N= 5

MALDI and/or BRUKER16S@ + Bruker = 10Bruker only = 22(N=32)

MALDI and/or VITEK16S@ = 7 (both)Vitek only = 21 (N=28)

L bozemanae* and REPORT as L.bozemanae*

2/5 (40%) 16S: 1/10 (10%)MALDI*: 19/32 (59%), scores 1.7-2.51


L. bozemanae orLegionella species AND REPORT as closest to L. bozemanae, L. steelei, L. longbeachae, other

3/5 (60%) 16S: 9/10 (90%)

MALDI#: 12/32 (38%) scores 2.01- 2.57, all >2.0

16S: 1/7 (14%)

MALDI#: 9/28 (32%) scores all 99.9%

NO GROWTH /NOT DONE (can not work on bacteria from respiratory tract)

0/5 (0%) 16S: 0/10 (0%)

MALDI: 1/32 (3%)

16S: 0/7 (0%)

MALDI: 1/28 (4)%

*Legionella bozemanae, L. bozemanii, Fluoribacter bozemanae or F. bozemanae or most similar to/ probable/possible L. bozemanaeall considered as synonyms /’correct’ here. Some mention notifying MOH re: case @ some would send to ref centre or sequence a secondary target gene to confirm# Most obtained L bozemanae with ID high confidence but would report as: one or more Legionella species cited; Gram neg rod; NO COMMENT; NO ID; would refer out as MSP or BCYE not validated in house yet

Summary, ECT-2, Legionella bozemanae• Source/ Growth Medium = respiratory specimen

and/or not having BCYE available for subculture, problematic for some; several=did only as part of test

• 16S: identification problematic due to poor quality (nearly full) sequence from type/ref sequence in GB

• Partial sequence (5’ ~ 1-500 bps) or WGS provided ID

• Unambiguous ID using 2° target (mip) /other

• MALDI MSPs (Bruker or Vitek) both performed very well; high confidence scores to genus and species ID

• Difficult to assess if MALDI would unambiguously ID extremely rare species which are closely related to L. bozemanae (no recent publications, only 1 older one)

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ECT-3 Moraxella catarrhalis NML 151131• 151131 from blood culture (rare cause of

bacteremia) in a 14 month old female• Can cause respiratory infections/otitis media in

young; upper respiratory tract commensal of young• Can spread person-to-person• Occasionally recovered in adults from sterile sites or

respiratory infections• MALDI-TOF ID thought to be unambiguous and

should provide correct ID [2015 Chapter 44, MCM 11th ed pg.817]

• Originally Neisseria catarrhalis then Brahamella(1970) now Moraxella catarrhalis (ref. LPSN)

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ECT-3 ID = Moraxella catarrhalis NML 151131Results by Beta Testers, September 2018

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*averaged among 3 replicates; extraction procedure used # Biotyper ver 7854 used; many reference MSPs (both systems) used Moraxella Branhamella catarrhalis; NML entries we opted to use Moraxella catarrhalis $IVD ver 3.0 used; direct procedure used


16s (TESTER 1) 100% M. catarrhalis Ne11T

(NR_028669); 99.9% ATCC 25238T (NR_118775)

M. catarrhalisunambiguous ID

16S (TESTER 2) 100% M. catarrhalis98.7% M. nonliquefaciens97.8% M. canis

Moraxella species; would refer to ref centre for species level ID

MALDI (1-Bruker*,#)

Moraxella catarrhalis 2.12-2.59 (Biotyper or NML MSPs)

Moraxella catarrhalisUnambiguous ID

MALDI(2-Vitek MS$)

Moraxella (Branhamella) catarrhalis score 99.9%

Moraxella (Branhamella) catarrhalis Unambiguous ID

ECT-3 ID = Moraxella catarrhalis NML 151131

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16S Partial / Full seqN= 5

MALDI and/or BRUKER16S + Bruker = 10Bruker only = 22(N=32)

MALDI and/or VITEK16S +Vitek = 7 Vitek only = 21 (N=28)

M catarrhalis* and REPORT as M. catarrhalis*

4/5 (80%) 16S: 8/10 (80%)MALDI*: 30/32 (94%), scores 2.01-2.6625/30 (83%) scores >2.4


Moraxella catarrhalis/species and REPORT as Moraxella sp#

1/5 (20%) 16S: 2/10 (20%)

MALDI#: 2/32 (6%) scores 2.12-2.48

16S: 1/7 (14%)

MALDI#: 1/28 (4%) score 99.9%

NOT REPORTED (as cannot work on bacteria from blood culture)

0/5 (0%) 16S: 0/10 (0%)

MALDI: 0/32 (0%)

16S: 0/7 (0%)

MALDI: 1/28 (4)%

*Reported as one of: Moraxella catarrhalis, M. catarrhalis (Branhamella) catarrhalis, M catarrhalis sg Branhamella catarrhalis; several labs said would report species depending on specimen type # Result of M. catarrhalis but report as Moraxella species/refer to ref centre; Moraxella genus most probably M. catarrhalis; do not report bacteria recovered from specific sources

Summary, ECT-3, Moraxella catarrhalis • Source = respiratory, ear, other sites

• 16S: unambiguously identified

• MALDI (Bruker or Vitek) unambiguously identified based on a variety of MSPs– Existence of several Latin epithets may be

confusing to new/younger staff

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ECT-4 Trueperella bernardiae NML 150885• Rare human pathogen, from brain / wounds/

abscesses, sterile body fluids/sites, UTIs, septic arthritis, bacteremias, necrotizing fasciitis

• Also detected/cause of disease in animals• 150885 from blood culture of 35y F with sepsis• Has undergone 4 name changes:• Early 1980s: CDC group 2• 1995: Actinomyces bernardiae• 1997: Arcanobacterium bernardiae• 2011: assigned to new genus, TrueperellaRef January 2019. MCM 12th ed Bernard, K. Chapter 28 Coryneforms

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ECT-4 ID= Trueperella bernardiae NML 150885Results by Beta Testers, September 2018

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$ LCDC, Laboratory Centre for Disease Control= NML name when located in Ottawa *averaged among 3 replicates; extraction procedure used # Biotyper ver 7854 used; reference MSPs (Bruker) only used $IVD ver 3.0 used; direct procedure used


16s (TESTER 1) 99.9% T. bernardiae LCDC$

89-0504T (NR_027195) 98.6% T abortisuis MurakamiT

(NR_041607)

T. bernardiaeunambiguous ID

16S (TESTER 2) 100% T. bernardiae98.6% T. abortisuis

Trueperella sp, send to

ref lab for species level ID

if required

MALDI

(1-Bruker*,#)

T. bernardiae 2.20-2.47

(Biotyper or NML MSPs)

Trueperella bernardiaeUnambiguous ID

MALDI

(2-Vitek MS$)

T. bernardiae score 99.9%

Trueperella bernardiae

ECT-4 ID = Trueperella bernardiae NML 150885

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16S Partial / Full seqN= 5

MALDI and/or BRUKER16S + Bruker = 10Bruker only = 22(N=32)

MALDI and/or VITEK16S +Vitek = 7 Vitek only = 21 (N=28)

T. bernardiae*and REPORT as T.bernardiae*

3/5 (60%) 16S: 7/10 (70%)

MALDI*: 24/32 (75%), scores 2.01-2.616/24 (67%) scores ≥2.3

16S: 6/7 (86%)

MALDI*: 26/28 (92%); scores all 99.9%

T. bernardiae or Trueperella spp and REPORT as Trueperella spp#

2/5 (40%) 16S: 3/10 (30%)

MALDI#: 8/32 (25%) scores 2.02-2.617/8 (87%) scores ≥2.3

16S: 1/7 (14%)

MALDI#: 2/28 (8%) score 99.9%

*Reported as one of: T. bernardiae, possible/probable T. bernardiae; T. (formerly Arcanobacterium) bernardiae; several labs said would report species depending on specimen type; report after consult with ID; refer to ref centre# Report as: Trueperella, closest to T. bernardiae; Trueperella species & would refer out; Trueperella spp; Genus Trueperella; low query coverage of top BLAST result, would do MALDI or biochems; would refer out as not usual mandate; have only had 2 strains so is not as yet validated

Summary, ECT-4 Trueperella bernardiae• Source = various sterile or non-sterile sites, in

both humans and animals • By 16S: provides unambiguous ID among

species of this genus• MALDI MSPs (Bruker or Vitek): unambiguous

high confidence identifications by either system • Agent rare, so some labs still validating in-house• Taxonomy complex, with 4 changes in past ~25ys

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ECT-5- Candida auris - CDC AR Bank #0382• Described in instructions as a ‘Bonus Bug’ to be

tested by MALDI-TOF users only

• Panel of 20 Candida strains received at NML as gift from the CDC in the fall 2016

• Able to provide this panel to PHLs in Canada, but was not to be further disseminated (based on CDC requirements in the MTA)

• Provided to assist with recognition, esp. for MDR C. auris

• AR bank 0382 fully susceptible usual antibiotics https://wwwn.cdc.gov/ARIsolateBank/Panel/IsolateDetail?IsolateID=382

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ECT-5- Candida auris• C. auris 1st recovered from ear (Japan) in 20091

• Infections in Asia, Middle East, Africa and UK; some are MDR2,3, resistant to fluconazole, amphotericin B

• Linked to cases in US1,2; 4 unique clones by WGS

• Problem: ID systems could not unambiguously ID C. auris (including MALDI systems 2-4)-making ID difficult

• Canada: few infections by susc. strains but 1st MDR case in 2017 linked to travel to India5

• NML now providing some services for C. auris (since2017)- contact: [email protected]

1. MMWR Nov. 4 2016 vol 65 pg 1-4 2. Lockhart et al 2017 Clin Microbiol Newsl 39:99-103

2. Kathuria et al 2015 JCM 53:1823-1830 4. Hass et al 2016 Diagn. Microbiol. Inf. Dis. 86:385-386

5. Schwartz & Hammond 2017 CCDR 43: 7/8 150-153

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“Reliably identify C. auris 100% of the time”

ECT-4 ID= Candida auris CDC AR 0382Results by Beta Testers, September 2018

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*averaged among 3 replicates; extraction procedure used # Both NMD and Biotyper MSPs analyzed here $IVD ver 3.0 used; direct procedure used


MALDI (1-Bruker*,#)

C. auris 2.23-2.51 (NML MSPs)

C. aurisUnambiguous ID

MALDI (1-Bruker*,#)

C. auris 2.40-2.43 (Biotyper 7854 MSPs)

C. aurisUnambiguous ID

MALDI(2-Vitek MS$)

NO IDscore 99.9%

C. auris not in IVD library ver 3.0

Bonus Bug: Candida auris –BRUKER-2018 vs 2017

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MALDI and/or BRUKER, 2018Strain CDC AR 038216S + Bruker = 10Bruker only = 22 (N=32)

MALDI and/or BRUKER, 2017Strain CDC AR 038116S + Bruker = 7Bruker only = 20 (N=27)

1. C. auris* andREPORT as C.auris*

21/32 (66%) scores 1.73-2.51[14/21& (67%) scores ≥2.07/21& (33%) scores 1.73-1.93]

25/27 (93%) scores 1.74-2.24

2. C. auris = ID other but REPORT as Candida sp/yeast#

10/32 (31%) [scores 1.70.-2.47 (to C auris), but report as Candida sp, yeast, other]

Candida spp 0/27 (0%)

NO ID 1/32 (3%) score 1.62 (would require additional testing)

2/27 (7%)

*Report as: C. auris; report after consult with ID; refer to ref centre; notifiable in province. # Report as Candida sp, yeast, NO ID or No peaks, would refer to ref centre; not in usual mandate; remarked that taxon ‘not as yet validated for MALDI’ Category 1: 4/21 (19%) done by direct, 17/21 (81%) done by some sort of extraction protocolCategory 2 and no ID 3/11 (27%) done by direct, 8/11 (73%) by some sort of extraction protocol

**Categories 1+2= 97% **

Bonus Bug: Candida auris – VITEK-2018 vs 2017

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MALDI and/or VITEK, 2018Strain CDC AR 038216S +Vitek = 7 Vitek only = 21 (N=28)

MALDI and/or VITEK 2017Strain CDC AR 038116S +Vitek = 8Vitek only = 16 (N=24 VITEK)

1. C. auris* & REPORT as C. auris*

6/28 (21%) 1/24 (4%) , score 99.9%, (lab possibly had newer ver IVD)

2. C. auris but REPORT as NO ID

3/28 (11%) includes a 50:50 with C. duobushaemulonii

0/24 (0%)

C. haemulonii REPORT as Candida sp or C auris

2/28 (7%), 1) probable C. haemulonii; 2)C. auris response using RUO

C. haemulonii: 7/24 (29%);C. pulcherrima or C rugosa 2/24 (8%)

Millerozyma farinosa (ID and report as)

1/28 (4%) -

NO ID / REPORT as NO ID&

16/28 (57%) 14/24 (58%)

*Report as: C. auris or presumptive C. auris; consult with ID; refer to ref centre; notifiable in prov; confirm with sequencing; & Ran on Vitek 2 panel scores 96%-98% C auris -anticipate C auris with ver 3.2; MALDI can not ID C auris at present; forward to ref centre if sterile site; ‘yeast’; other method needed; lab does not process specimen type; left blank



Summary, BONUS BUG Candida auris• Second time yeast sent in the NMG panel (to

see if ID improved from 2017)

• MALDI (Bruker): high confidence scores; 97% got ID as C auris, some with low scores; 31% elected to not ID as C auris

• MALDI (Vitek): modest improvement for ID in 2018; roll out of IVD with C. auris slower than anticipated but ID expected to dramatically improve with that

• Suggest: confirm ID & AST by ref centre if C. auris /closely related spp recovered

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28 Taxa to date: (underlined= strict anaerobes)GPR: Coryne. kroppenstedtii (2012), Unidentifiable Propionibacteriaceae (2014), Coryne. propinquum (2014), Eisenbergiella tayi (2016), Actinomyces neuii (2016), Actinotignum urinale (2017), Mycobacterium smegmatis (2017); Nocardia cyriacigeorgica (2018); Trueperella bernardiae (2018)GPC: Streptococcus suis (2013), UNID Aerococcaceae (2013), Streptococcus mitis (2016)GNC: Neisseria sp, closest to N. polysaccharea (2015); Moraxella catarrhalis (2018)GNR: Chryseobacterium hominis (2012), Klebsiella pneumoniae (2013), Burkholderia thailandensis (2014), Bacteriodes fragilis (2014), Yersinia pseudotuberculosis (2015), Burkholderia cenocepacia (2015), Eggerthella lenta (2015), Burkholderia stabilis (2016), Elizabethkingia anophelis (2017), Aggregatibacter aphrophilus (2017); Legionella bozemanae (2013)Yeast: Candida auris (2017), Candida auris (2018)

NMG-NML Summary of 7 Years of sending out Challenge Test

2013

Challenge Test / CMUG Updates• ‘6 years of the Challenge Test’ presented in brief at

ECCMID (Madrid)• Depending on staff availability, expect Challenge Test to

roll out with similar time lines to 2018• Will request that shipping be limited to major couriers CMUG (Canadian MALDI-TOF Users Group) • Annual meeting on Friday April 5, 2019 from 12-2pm

(during the AMMI-CACMID conference, Westin Hotel, Ottawa ON) http://www.cacmid.ca/2018/09/2019_ottawa/

• Includes review of highlights from this test• Free for CACMID attendees

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GRDI Project (to create NMD) Update• Original GRDI project began in mid-2014 • Original project ended Mar. 31, 2017 (2.5y) • Got 2y ‘extension’ (April 2017-March 31, 2019) • Extension= 1 /2 of funding from original project• MALDI manager M Unger works on this ½ time• ~NMD currently has ~750 entries in NMD, now

being exhaustively curated • ~20% of NMD entries-new genera and/or spps• Need to develop protocols to enable downloads

of MSPs which conserve the IP37

NEW Call for GRDI Proposals Underway• GRDI call for new proposals, LOI due Dec. 10/18

• We plan to propose continuation of existing NMD project and add new research initiatives

ROLE OF MALDI-TOF USERS for new GRDI

• We will be contacting MALDI-TOF users to provide concrete support for this GRDI proposal, as part of this post-pilot round

• This will probably involve writing letters of support in January 2019.

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Acknowledgements for the Challenge Test Team

Photo by KB in the Galapagos, April 2016

We gratefully acknowledge that this work was carried out in a facility located on Treaty One Land and the traditional

home of the Métis People

For assistance in characterizing taxa and preparations of shipments:

• Special Bacteriology: Tamara Burdz, Debbie Wiebe and Ana Luisa Pacheco

• COOP, U St-Boniface: Dominic Kielich• NML Shipping: Don Ashley and Mike

Pushka• NML’s DNA Core Facility for assistance

with sequencing• NML’s Proteomic Core Facility , for

maintaining the MALDI-TOF systems, for advice and considerable assistance with this work

results of the 2018 16s rrnagene sequencing and/or maldi ...nmgroup.ca/document/2018/2018_15.pdf ·...

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