Evaluation of BluePoint MycoID and MALDI-TOF MS for identification of Nontuberculous Mycobacteria from the Flagged Mycobacterium Growth Indicator Tube system

Background/Aims: Species-level identification of nontuberculous mycobacteria (NTM) is important in making decisions about the necessity and choice of antimicrobial treatment. The reason is predictable clinical significance and the susceptibility profile of specific NTM species. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is recognized as a diagnostic tool for routine identification of bacteria and yeasts in the clinical laboratory based on protein fingerprint analysis. The aim of the study was to evaluate MALDI-TOF MS in the identification of NTM. Methods: A total of 25 NTM isolates from liquid cultures were identified with both polymerase chain reaction (PCR)-based hybridization assay and MALDI-TOF MS at the University Hospital Center Zagreb. Results: PCR-based hybridization assay identified 96% (24/25) and MALDI-TOF MS 80% (20/25) of tested NTM isolates. Five isolates with no reliable MALDI-TOF MS identification belonged to the Mycobacterium avium-intracellulare complex. Seventy percent (14/20) of NTM isolates successfully identified with MALDI-TOF MS had a score higher than 2.0, indicating reliable species identification. Conclusion: MALDI-TOF MS is a promising tool for the identification of NTM. With a further improvement of the protein extraction protocol, especially regarding the M. avium-intracellulare complex, MALDI-TOF MS could be an additional standard method for identification of NTM.

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Impact of updating the MALDI-TOF MS database on the identification of nontuberculous mycobacteria

Journal of Mass Spectrometry ◽

10.1002/jms.3944 ◽

2017 ◽

Vol 52 (9) ◽

pp. 597-602 ◽

Cited By ~ 4

Author(s):

D. Rodriguez-Temporal ◽

D. Perez-Risco ◽

E. A. Struzka ◽

M. Mas ◽

F. Alcaide

Keyword(s):

Nontuberculous Mycobacteria ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Tof Ms

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Evaluation of Two Protein Extraction Protocols Based on Freezing and Mechanical Disruption for Identifying Nontuberculous Mycobacteria by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry from Liquid and Solid Cultures

Journal of Clinical Microbiology ◽

10.1128/jcm.01548-17 ◽

2018 ◽

Vol 56 (4) ◽

Cited By ~ 9

Author(s):

David Rodriguez-Temporal ◽

Daniel Perez-Risco ◽

Eduardo A. Struzka ◽

Mireia Mas ◽

Fernando Alcaide

Keyword(s):

Protein Extraction ◽

Nontuberculous Mycobacteria ◽

Laser Desorption Ionization ◽

Liquid Media ◽

Ionization Time ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Mechanical Disruption ◽

Tof Ms

ABSTRACTMatrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) has proved to be a useful diagnostic method for identifying conventional bacteria. In the case of mycobacteria, a good protein extraction protocol is essential in order to obtain reliable identification results. To date, no such protocol has been definitively established. The aim of this study was to compare the manufacturer's recommended protein extraction protocol (protocol A) with two novel protocols (protocols B and C), which apply different freezing temperatures and mechanical disruption times using an automatic tissue homogenizer. A total of 302 clinical isolates, comprising 41 nontuberculous mycobacteria (NTM) species, were grown in parallel on solid and liquid media and analyzed: 174 isolates were slow-growing mycobacteria (SGM) and 128 isolates were rapid-growing mycobacteria (RGM). Overall, MALDI-TOF MS identified a higher number of NTM isolates from solid than from liquid media, especially with protocol C (83.4 and 68.2%, respectively;P< 0.05). From solid media, this protein extraction method identified 57.9 and 3.9% more isolates than protocols A (P< 0.001) and B (P< 0.05), respectively. In the case of liquid media, protocol C identified 49.7 and 6.3% more isolates than protocols A and B, respectively (P< 0.001). With regard to the growth rate, MALDI-TOF MS identified more RGM isolates than SGM isolates in all of the protocols studied. In conclusion, the application of freezing and automatic tissue homogenizer improved protein extraction of NTM and boosted identification rates. Consequently, MALDI-TOF MS, which is a cheap and simple method, could be a helpful tool for identifying NTM species in clinical laboratories.

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Utility of the MALDI-TOF MS method to identify nontuberculous mycobacteria

Journal of Infection and Chemotherapy ◽

10.1016/j.jiac.2015.09.006 ◽

2016 ◽

Vol 22 (1) ◽

pp. 32-35 ◽

Cited By ~ 16

Author(s):

Masahiro Kodana ◽

Norihito Tarumoto ◽

Tohru Kawamura ◽

Taeko Saito ◽

Hideaki Ohno ◽

...

Keyword(s):

Nontuberculous Mycobacteria ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Tof Ms

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Epidemiology of Nontuberculous Mycobacteria in French Polynesia

Journal of Clinical Microbiology ◽

10.1128/jcm.01560-15 ◽

2015 ◽

Vol 53 (12) ◽

pp. 3798-3804 ◽

Cited By ~ 5

Author(s):

Michael Phelippeau ◽

Djaltou Aboubaker Osman ◽

Didier Musso ◽

Michel Drancourt

Keyword(s):

Nontuberculous Mycobacteria ◽

Sequence Similarity ◽

Gene Sequencing ◽

French Polynesia ◽

Rrna Gene ◽

Mycobacterium Fortuitum ◽

Content Type ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Tof Ms

As few data are available in the Pacific countries and territories of the Oceania region regarding nontuberculous mycobacteria, we retrospectively identified 87 such isolates from French Polynesia from 2008 to 2013 by hybridization using DNA-strip, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and partialrpoBgene sequencing. PartialrpoBgene sequencing classified 42/87 (48.3%) isolates in theMycobacterium fortuitumcomplex, 28 (32.2%) in theMycobacterium abscessuscomplex, 8 (9.2%) in theMycobacterium mucogenicumcomplex, and 5 (5.7%) in theMycobacterium aviumcomplex. Two isolates were identified asMycobacterium acapulcensisandMycobacteriumcosmeticumby partial 16S rRNA gene sequencing. One isolate, unidentified by MALDI-TOF MS and yielding less than 92% and 96% sequence similarity withrpoBandhsp65reference sequences, respectively, was regarded as a potentially new species. Samples from three patients exhibiting ≥2Mycobacterium porcinumisolates and from one patient with emphysema and a lung abscess exhibiting 2Mycobacterium senegalenseisolates fulfilled the American Thoracic Society microbiological criteria for nontuberculous mycobacterial lung infection. Remote geographic areas, such as French Polynesia, are potential sources for the discovery of new mycobacterial species.

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Evaluation of RGM Medium for Isolation of Nontuberculous Mycobacteria from Respiratory Samples from Patients with Cystic Fibrosis in the United States

Journal of Clinical Microbiology ◽

10.1128/jcm.02423-16 ◽

2017 ◽

Vol 55 (5) ◽

pp. 1469-1477 ◽

Cited By ~ 11

Author(s):

Rongpong Plongla ◽

Clair L. Preece ◽

John D. Perry ◽

Peter H. Gilligan

Keyword(s):

Cystic Fibrosis ◽

Burkholderia Cepacia ◽

Nontuberculous Mycobacteria ◽

Culture Method ◽

The United States ◽

Content Type ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Selective Agar ◽

Tof Ms

ABSTRACTA novel selective agar (RGM medium) has been advocated for the isolation of rapidly growing mycobacteria from the sputa of cystic fibrosis (CF) patients. The aim of this study was to compare RGM medium toBurkholderia cepaciaselective agar (BCSA) and a standard acid-fast bacillus (AFB) culture method for the isolation of nontuberculous mycobacteria (NTM) from patients with CF. The applicability of matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) for the identification of NTM isolated on RGM medium was also assessed. Respiratory samples (n= 869) were collected from 487 CF patients and inoculated directly onto RGM medium and BCSA. Cultures were incubated at 30°C and examined for up to 28 days. A subset of 212 samples (from 172 patients) was also cultured by using a mycobacterial growth indicator tube (MGIT) and on Lowenstein-Jensen medium following dual decontamination. By using a combination of all methods, 98 mycobacteria were isolated from 869 samples (11.3%). The sensitivity of RGM medium (96.9%) was significantly higher than that of BCSA (35.7%) for the isolation of mycobacteria (P< 0.0001). The sensitivity of RGM medium was also superior to that of standard AFB culture for the isolation of mycobacteria (92.2% versus 47.1%;P< 0.0001). MALDI-TOF MS was effective for the identification of mycobacteria in RGM medium. RGM medium offers a simple and highly effective tool for the isolation of NTM from patients with CF. Extended incubation of RGM medium for 28 days facilitates the isolation of slow-growing species, including members of theMycobacterium aviumcomplex (MAVC).

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Evaluation of MALDI Biotyper Interpretation Criteria for Accurate Identification of Nontuberculous Mycobacteria

Journal of Clinical Microbiology ◽

10.1128/jcm.01103-20 ◽

2020 ◽

Vol 58 (10) ◽

Author(s):

David Rodriguez-Temporal ◽

Belén Rodríguez-Sánchez ◽

Fernando Alcaide

Keyword(s):

Liquid Medium ◽

Solid Medium ◽

Protein Extraction ◽

Nontuberculous Mycobacteria ◽

Cutoff Score ◽

Content Type ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Interpretation Criteria ◽

Tof Ms

ABSTRACT Identification of mycobacteria by matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) requires not only a good protein extraction protocol but also an adequate cutoff score in order to provide reliable results. The aim of this study was to assess the cutoff scores proposed by the MALDI-TOF MS system for mycobacterial identification. A total of 693 clinical isolates from a liquid medium and 760 from a solid medium were analyzed, encompassing 67 different species of nontuberculous mycobacteria (NTM). MALDI-TOF MS identified 558 (80.5%) isolates from the liquid medium and 712 (93.7%) isolates from the solid medium with scores of ≥1.60. Among these, four (0.7%) misidentifications were obtained from the liquid medium and four (0.5%) from the solid medium. With regard to species diversity, MALDI-TOF MS successfully identified 64 (95.5%) different species, while PCR-reverse hybridization (GenoType Mycobacterium CM and AS assays) identified 24 (35.8%) different species. With MALDI-TOF MS scores of ≥2, all isolates were correctly identified, and with scores in the range from 1.60 to 1.99, most isolates were correctly identified, except for Mycobacterium angelicum, M. parascrofulaceum, M. peregrinum, M. porcinum, and M. gastri. In conclusion, MALDI-TOF MS is a useful method for identifying a large diversity of NTM species. A score threshold of 1.60 proved useful for identifying almost all the isolates tested; only a few species required a higher score (≥2.00) to obtain a valid definitive identification.

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