APHL 2021 Poster Abstracts
Infectious Disease concordant with ID by HPLC . From LJ slants and 7H10 plates , 92 % of mycobacterial isolates had species level ID that matched between MALDI-TOF and HPLC . Of the 8 % remaining , 6 % were discordant and 2 % failed to produce an ID score > 1.8 . All 63 MTBC evaluated were 100 % concordant , with an ID score > 2.0 ( average ID score of 2.256 ). Isolates extracted directly from liquid MGIT culture performed similarly to solid media , with 92 % ( 55 / 60 ) of isolates concordant with HPLC ; all with ID scores > 2.0 . Further testing of discordant and failed isolates is in progress . Both intra- and inter-assay reproducibility studies using reference strains resulted in 100 % concordance with 1.8 can reliably be used to report mycobacterial species identifications , while ID scores < 1.8 require confirmation by a secondary ID method . The Bruker MALDI-TOF method provide us the ability to ID more NTM species compared to HPLC . MALDI-TOF also provides a rapid and accurate ID method for routine use at NYC PHL .
Presenter : Randal Fowler , New York City Public Health Laboratory , rfowler @ health . nyc . gov
Reverse Transcriptase Real-time PCR Assay for the Enumeration of Live Candida auris from the Healthcare Environment
B . L . Freitas , S . Chaturvedi , L . Leach and V . Chaturvedi , New York State Department of Health / Wadsworth Center , Albany , NY
The ongoing healthcare-associated outbreaks of multidrug-resistant Candida auris prompted the development of several rapid DNAbased molecular diagnostic tests . These tests do not distinguish between live and dead C . auris cells , and as a result are not fully useful for environmental surveillance and containment efforts . We addressed this problem by developing a reverse transcriptase ( RT ) real-time ( rt ) PCR assay for rapid detection of live C . auris from the healthcare environment . The assay targets the ITS2 cDNA of C . auris , which was amplified by first obtaining pure RNA followed by the reverse transcription and real-time PCR assay . The assay was highly sensitive , with the detection limit of ten colony-forming units ( CFU ) per RT real-time PCR reaction . Next , the relationship between the detection of ITS2 cDNA and cellular viability in C . auris was assessed by killing C . auris cells with bleach , heat , and ethanol . The ITS2 cDNA was not detected from the bleach-killed cells while It was detected from the heat of the ethanol-killed cells . The RT real time PCR assay was finally validated on the sponge matrix spiked with live , dead ( bleach-killed ) C . auris and other Candida spp . No Ct values were obtained from samples spiked with either dead C . auris ( 1E5 / ml ) or other species of Candida ( 1E5 / ml ). In contrast , the majority of environmental samples spiked with a low to a high number of live C . auris ( 1E2 to 1E5 ) yielded Ct values . Finally , retrospective analysis of 33 environmental samples revealed concordant results with culture . These results proved that the assay Is highly reproducible , robust , and specific . The new RT real-time PCR assay would be an invaluable tool in surveillance efforts to control the spread of live C . auris in the healthcare environment .
Presenter : Bryanna Lexus Freitas , New York State Department of Health / Wadsworth Center , Bryanna . freitas @ health . ny . gov
Development and Retrospective Analysis of a Multiplex Real-time PCR Assay to Detect the Novel Beta-Lactamase Genes , blaSIM , blaIMI / NMC-A and blaSPM
K . M . Hager , L . M . Rose , K . J . Cummings , S . Morris , C . J . Wagner , J . Bodnar , K . Wahl , K . Prussing , J . L . Chan , E . Nazarian and K . A . Musser , New York State Department of Health / Wadsworth Center , Albany , NY
Antibiotic resistance ( AR ) is one of the greatest public health challenges of modern times . The emergence of carbapenemaseproducing bacteria , primarily the gram-negative bacilli , has become a global health threat . CDC lists carbapenem-resistant Enterobacteriaceae , Acinetobacter species , and Pseudomonas aeruginosa as urgent and serious threats . Rapid detection of AR genes by molecular methods can aid in the implementation of infection prevention measures and help to combat the spread of AR organisms . The CDC AR Laboratory Network and the New York State Department of Health employ molecular-based assays that detect the most common carbapenemase genes , blaKPC , blaNDM , blaVIM , blaOXA-48-like , blaIMP , blaOXA-23 , blaOXA-24 / 40 , and blaOXA-58 . Yet , few rapid detection methods exist for emerging β-lactamase ( bla ) resistance genes . Identification requires labor intensive phenotypic methods for carbapenemase and metallo-β-lactamase production in conjunction with whole genome sequencing . First identified in South Korea in 2003 , blaSIM has since been identified in China and Egypt amidst Acinetobacter species and Klebsiella pneumoniae . As a metallo-β-lactamase , blaSIM is a concern because it can hydrolyze carbapenems and other β-lactam antibiotics . The blaSIM gene can be chromosomally encoded or located within mobile genetic elements , which is especially unsettling from a public health perspective as mobile genetic elements can increase the spread of AR genes . For this reason , we sought to develop a multiplex Taqman-based real-time PCR assay to detect the blaSIM gene with the addition of an internal control . Using Geneious and Primer 3 Plus software , primers and probe were designed to conserved regions of the blaSIM gene . In silico results suggest the assay can detect all blaSIM variants . This assay will be validated for both culture isolates and rectal swabs to aid in surveillance efforts of the blaSIM gene . With this newly developed assay , a retrospective analysis of Pseudomonas aeruginosa isolates received through the AR Lab Network since 2019 will be tested to determine the prevalence of this novel AR gene in our isolate submissions . Performance of this assay will allow for the rapid detection of a novel metallo-β-lactamase gene and provide key insights into the emergence and spread of newly evolving genes .
Presenter : Kelli Hager , New York State Department of Health / Wadsworth Center , kelli . hager @ health . ny . gov
Application of Metagenomics Analysis for Pathogen Identification Direct from Cadaveric Blood Cultures
S . LaVoie , E . Omoregie , M . Chowdhury , J . Wu , E . Gonzalez , S . Hughes and J . Rakeman , New York City Public Health Laboratory , New York City , NY
The New York City Public Health Laboratory ( PHL ) conducted a pilot study to compare metagenomics analysis with traditional culture-based methods for identification of pathogenic bacteria from blood cultures . NYC PHL routinely receives inoculated aerobic and anaerobic blood bottles for culture from autopsy cases from the NYC Office of the Chief Medical Examiner ( OCME ). Typically
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LAB MATTERS Summer 2021 |
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