Hawaii State Laboratories Required Submission Information for APHL RFP: Validation of the MALDI-TOF for the identification of Neisseria gonorrhoeae
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2 Hawaii State Laboratories Required Submission Information for APHL RFP: Validation of the MALDI-TOF for the identification of Neisseria gonorrhoeae 1. Please describe the laboratory s current MALDI-TOF testing practices including pathogens detected. a. Describe how many years the MALDI-TOF has been in use, which MALDI-TOF is in use, how often testing is performed, pathogens tested, and amount of experience laboratory staff have in using the methodology (years, training, and consistency performing method Hawaii State Laboratories Division has been performing bacterial identification using a Bruker Biotyper MALDI-TOF since MALDI-TOF testing is performed two times per week by medical microbiology (MMB) staff and as needed by Laboratory Preparedness and Response Program (LPRP). For this project, samples could either be run during the routine Tuesday / Thursday runs by the primary analyst in MMB or on any other day as needed by a secondary. The individuals trained to perform MALDI-TOF are Precy Calimlim (LPRP), Cheryl Daquip (LPRP), Lori Suyama (MMB), Lance Chinna (MMB) and Amy Woron (MMB, in-progress). 2. Please describe the laboratory s experience in testing for N. gonorrhoeae including methodologies. a. Describe how many years each method has been in use, how often testing is performed (times per week), annual volume, and amount of experience laboratory staff have in using the methodology (years, training, and consistency performing method). Hawaii State Laboratories Division Medical Microbiology Branch has been performing culture and identification of N. gonorrhoeae for more than 4 decades, and nucleic acid amplification testing (NAAT) for more than 3 decades. Culture is performed daily whenever specimens or isolates are received (up to 5 days a week). NAAT is performed daily (4-5 times per week). Our annual culture volume of N. gonorrhoeae testing was 2,443 in 2016, and NAAT volume was 20,406 (also in 2016). Currently, 3 lab
3 staff are trained to perform culture for N. gonorrhoeae and 5 staff are trained to perform NAAT. All staff have been trained and are competency assessed annually. In 2016, SLD tested 22 N. gonorrhoeae isolates, 5 N. meningitidis, isolates, and 6 commensals by both MALDI-TOF/MS and API-NH, and got 100% agreement. Also in 2016 we participated in the Quality Control for Molecular Diagnostics (QCMD) 2016 MALDI-TOF EQA Pilot Study, and we plan to participate in this QC program again in All N. gonorrhoeae and Neisseria species from normally sterile site are archived, subject to space limitaions. 3. Please describe the current methodologies for testing non-gonococcal Neisseria species: Process for testing non-gonococcal Neisseria species?? Currently, we use conventional media (sheep blood agar, chocolate agar, Lewis-Martin, and MacConkey) for culture at 35C in CO 2 incubator. We perform oxidase and a gram stain on representative colonies. Single colony subcultures are used for both MALDI-TOF/MS and API-NH. If N. meningitidis, we will try serogrouping with antisera, and also, we will do superoxol testing. If unable to obtain an identification from the MALDI-TOF/MS and API-NH, we can perform16s ribosomal sequencing and additional biochemical tests such as catalase, DNase, nitrate, phenylalanine, urease, pigment production and growth on nutrient agar at 35C. How many years each method has been in use? Institutional knowledge for conventional cultivation and biochemical testing (CDC methods) as mentioned above, goes back to about 1984, but may have been in use earlier. The API-NH has been in use since Sequencing (16s) has been available in-house since MALDI-TOF/MS has been performed since How often is testing performed? Testing is performed whenever unknown Neisseria spp. are submitted to us, or when we isolate Neisseria spp. not N. gonorrhoeae on cultures from our STD clinic. What pathogens are detected?
4 The most common non-gc pathogen is probably Neisseria meningitidis and is usually from a sterile site such as blood or CSF. We get occasional Moraxella catarrhalis and rarely Kingella types, The more commensal Neisseria spp. We see are lactamica, sicca, subflava, weaver, canis, cinerea, elongata, flavescens and mucosa. What is the experience of lab staff in the different methodologies? Two to three staff are trained and competency assessed in each methodology in Medical Microbiology Branch (MMB), with the exception of 16s sequencing, which is performed by the Laboratory Preparedness and Response Program (LPRP). Furthermore, there is redundant expertise in MALDI- TOF/MS in LPRP. Current strategies to mitigate biosafety risk of N. meningitides? All primary isolation plates are initially evaluated in the biological safety cabinet (BSC). Any suspect or confirmed meningiococci are only manipulated in the BSC. Tube extractions are performed on isolates for MADLI-TOF/MS, and we have data to demonstrate this process kills the bacteria. Staff are encouraged to discuss meningiococcus vaccination options with their providers. 4. Please describe the current collection of non-gonococcal Neisseria species. a. Provide an Excel list of commensal Neisseria species as well as M. catarrhalis and K. denitrificans your lab currently have on hand, when possible include the number of isolates for each species, and describe the storage method (frozen, lyophilize). Excel list of isolates attached. Relevant References: 1. Katz, AR, AY Komeya, RD Kirkcaldy, AC Whelen, OO Soge, JR Papp, EN Kersh, GM Wasserman, NP O Connor, PS O Brien, DT Sato, EV Maningas, GY Kunimoto, JE Tomas Cluster of multidrug-resistant Neisseria gonorrhoeae isolates with high-level azithromycin resistance and decreased ceftriaxone susceptibility, Hawaii, Clin Infect Dis.[ePub ahead of Print] with editor comment.
5 2. Papp, JR, JA McLean, E Nash, AR Katz, RD Kirkcaldy, NP O Connor, PS O Brien, DH Harauchi, EV Maningas, EN Kersh, JE Tomas, GM Wasserman, GY Kunimoto, DL Trees, AC Whelen Characterization of a cluster of Neisseria gonorrhoeae isolates from Hawaii with decreased in vitro susceptibility to azithromycin and ceftriaxone. Emerg Infect Dis. 23(5): Trees, D*, G Kubin, B Kirkcaldy, A Whelen, and J McLean Establishment of Whole Genome Sequencing of Neisseria gonorrhoeae isolates in State Public Health Laboratories for use in the Development of a Pipeline Predicting Antimicrobial Resistance. STI & HIV World Congress, Rio de Janeiro, Brazil. 4. Raymond, S, M. Sonson, T. Enomoto, C. Ying, T. Koyamatsu, A.C. Whelen, W. Kim, and M.J. Bankowski* Use of MALDI-TOF MS for the Rapid Identification of Emerging Corynebacterium diphtheriae from Patients with Skin and Soft Tissue Infections in Hawaii. Annual ASM Meeting. 5. Kidd, S., M.V.C. Lee, E. Maningas, A. Komeya, G. Kunimoto, N. O Connor, A. Katz, G. Wasserman, R.D. Kirkcaldy, A.C. Whelen Gonococcal susceptibility to cephalosporins Hawaii, Sexually Transmitted Diseases 40(9): Katz A.R., A.Y. Komeya, O.O. Soge, M.I. Kiaha, M.V.C. Lee, G.M. Wasserman, E. Maningas, A.C. Whelen, R.D. Kirkcaldy, S.J. Shapiro, G.A. Bolan, K.K. Holmes Neisseria gonorrhoeae with high-level resistance to azithromycin: case-report of the first isolate identified in the United States. Clin. Infect. Dis. 54:
6 Species # of isolates Source Storage Neisseria meningitidis 3 throat Yes/frozen Neisseria meningitidis 1 urethral Yes/frozen Neisseria meningitidis 10 CSF Yes/lyophilized Neisseria meningitidis 10 blood Yes/lyophilized Neisseria lactamica 1 throat Yes/frozen Neisseria sicca 1 blood Yes/lyophilized/frozen Neisseria subflava 1 blood Yes/frozen Neisseria subflava 1 blood Yes/lyophilized Neisseria subflava 1 synovial fluid Yes/lyophilized Neisseria mucosa 1 unknown Yes/frozen Neisseria mucosa 4 blood Yes/lyophilized Neisseria canis 1 dog bite Yes/lyophilized Neisseria cinerea 1 urine Yes/lyophilized Moraxella catarrhalis 2 blood Yes/lyophilized Moraxella catarrhalis 1 blood Yes/frozen Moraxella catarrhalis 5 sputum Yes/frozen Moraxella catarrhalis 2 throat Yes/frozen Moraxella catarrhalis 1 nasal Yes/frozen Total 47 Note: Most lyophilized isolates are from late 1980s and early 1990s
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