Vitek MS System Research Articles

Performance of common primary and chromogenic culture media for MALDI-TOF MS identification of clinically relevant yeasts.

Timely and accurate identification of yeasts is essential for adequate treatment, considering the increase in antifungal resistance of some species, particularly for C. auris. Current matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) manufacturer's protocol for identification of yeasts requires 24- to 72-h cultivation on Sabouraud dextrose media (SAB), but not some of the mainstay primary culture media used in mycology such as inhibitory mold agar (IMA), Mycosel, CHROMagar Candida Plus, and CHROMagar Candida. As culture media can influence MALDI-TOF MS identification results, this study evaluated the accuracy and performance of identification of clinically relevant yeasts on these first-line media using the VITEK-MS MALDI-TOF MS system.IMPORTANCEIn this study, a panel of 140 strains (21 species) was used to assess the performance of the selected media. Although not in the manufacturer's list of accepted media, IMA and chromogenic media are suitable for the identification of yeasts on the VITEK-MS systems. CHROMagar Candida Plus allowed the identification of 135/140 isolates tested after 24-h incubation similar to SAB reference media (137/140). Yeast isolates that grew on Mycosel selective media were also reliably identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. VITEK-MS system with IVD database V3.2 correctly identified C. auris strains to the species level on CHROMagar Candida Plus alleviating the need for subcultivation and reduced turnaround time (24-72 h) to identification for patient screening.

Comparative genomics of four lactic acid bacteria identified with Vitek MS (MALDI-TOF) and whole-genome sequencing

Lactic acid bacteria (LAB) can be used as a probiotic or starter culture in dairy, meat, and vegetable fermentation. Therefore, their isolation and identification are essential. Recent advances in omics technologies and high-throughput sequencing have made the identification and characterization of bacteria. This study firstly aimed to demonstrate the sensitivity of the Vitek MS (MALDI-TOF) system in the identification of lactic acid bacteria and, secondly, to characterize bacteria using various bioinformatics approaches. Probiotic potency-related genes and secondary metabolite biosynthesis gene clusters were examined. The Vitek MS (MALDI-TOF) system was able to identify all of the bacteria at the genus level. According to whole genome sequencing, the bacteria were confirmed to be Lentilactobacillus buchneri, Levilactobacillus brevis, Lactiplantibacillus plantarum, Levilactobacillus namurensis. Bacteria had most of the probiotic potency-related genes, and different toxin-antitoxin systems such as PemIK/MazEF, Hig A/B, YdcE/YdcD, YefM/YoeB. Also, some of the secondary metabolite biosynthesis gene clusters, some toxic metabolite-related genes, and antibiotic resistance-related genes were detected. In addition, Lentilactobacillus buchneri Egmn17 had a type II-A CRISPR/Cas system. Lactiplantibacillus plantarum Gmze16 had a bacteriocin, plantaricin E/F.

Heterogeneity of Antibiotic-Resistant Isolates of Listeria Monocytogenes Isolated from Food Products in Moscow

Relevance. Listeria monocytogenes is a ubiquitous bacterium that causes listeriosis, which represents a widespread infectious disease currently inflicting great damage to livestock production and posing a serious threat to human health.Aim. To analyze the population structure and assess the pathogenic potential of Listeria monocytogenes isolates isolated on the territory of the Russian Federation.Materials and methods. A total of 79 listeria isolates were isolated from food products. Species identification and phenotypic analysis for antibiotic resistance were performed using VITEK MS system (bioMerieux, Marcyl’toile, France). Thirty-five antibiotic-resistant isolates were characterized by analysis of whole-genome sequencing data.Results. Whole genome sequences of thirty-five antibiotic-resistant Listeria monocytogenes isolates of food origin were analyzed. We determined clonal structure of this population and revealed a small number of antibiotic resistance determinants (fosX, tetM и сlpL), extensive set of virulence factors, as well as the presence of CRISPR/Cas systems. Most of the isolates belonged to phylogenetic line II and were divided into nine clonal complexes with the prevalence of CC121, which was one of the epidemiologically significant genetic clones. Two CC2 isolates belonging to the most pathogenic phylogenetic lineage I were also found. Thirteen isolates were characterized by the presence of putative CRISPR/Cas systems of IB and IIA types. All ST 121 isolates contained two types of identified adaptive immunity systems simultaneously in their genomes. Correlation analysis confirmed their functionality.Conclusion. We believe that the whole genome data obtained for the foodborne Listeria monocytogenes isolates will facilitate and complement further epidemiological studies of this pathogen, as well as the investigations of its genome variability in terms of the acquisition of various genetic elements associated with adaptation, antimicrobial resistance, and virulence. Moreover, the results of such studies will help to develop preventive measures to effectively solve problems associated with the bacterial contamination of animal products and ensure food safety in production conditions and the «farm-to-table» chain.

In Vitro Antibiofilm Effect of N-Acetyl-L-cysteine/Dry Propolis Extract Combination on Bacterial Pathogens Isolated from Upper Respiratory Tract Infections.

Bacterial biofilms play an important role in the pathogenesis of chronic upper respiratory tract infections. In addition to conventional antimicrobial therapy, N-acetyl-L-cysteine (NAC) and propolis are dietary supplements that are often recommended as supportive therapy for upper respiratory tract infections. However, no data on the beneficial effect of their combination against bacterial biofilms can be found in the scientific literature. Therefore, the aim of our study was to investigate the in vitro effect of N-acetyl-L-cysteine (NAC) and dry propolis extract in fixed combinations (NAC/dry propolis extract fixed combination) on biofilm formation by bacterial species isolated from patients with chronic rhinosinusitis, chronic otitis media, and chronic adenoiditis. The prospective study included 48 adults with chronic rhinosinusitis, 29 adults with chronic otitis media, and 33 children with chronic adenoiditis. Bacteria were isolated from tissue samples obtained intraoperatively and identified using the MALDI-TOF Vitek MS System. The antimicrobial activity, synergism, and antibiofilm effect of NAC/dry propolis extract fixed combination were studied in vitro. A total of 116 different strains were isolated from the tissue samples, with staphylococci being the most frequently isolated in all patients (57.8%). MICs of the NAC/dry propolis extract fixed combination ranged from 1.25/0.125 to 20/2 mg NAC/mg propolis. A synergistic effect (FICI ≤ 0.5) was observed in 51.7% of strains. The majority of isolates from patients with chronic otitis media were moderate biofilm producers and in chronic adenoiditis they were weak biofilm producers, while the same number of isolates in patients with chronic rhinosinusitis were weak and moderate biofilm producers. Subinhibitory concentrations of the NAC/propolis combination ranging from 0.625-0.156 mg/mL to 10-2.5 mg/mL of NAC combined with 0.062-0.016 mg/mL to 1-0.25 mg/mL of propolis inhibited biofilm formation in all bacterial strains. Suprainhibitory concentrations ranging from 2.5-10 mg/mL to 40-160 mg/mL of NAC in combination with 0.25-1 mg/mL to 4-16 mg/mL of propolis completely eradicated the biofilm. In conclusion, the fixed combination of NAC and dry propolis extract has a synergistic effect on all stages of biofilm formation and eradication of the formed biofilm in bacteria isolated from upper respiratory tract infections.

A-290 Evaluation of a Commercial Turbidity Monitoring System Followed by Mass Spectrometry to Rapid Identification of Microorganisms from Urine Samples

Abstract Background Urinary tract infections (UTIs) are one of the most common bacterial infections in worldwide and represent a worried public health problem, especially due to a significant rise in global antibiotic use in recent years. The purpose of the study was to evaluate the performance of Uro-Quick (UQ) system (Alifax) and the Vitek-MS system (bioMérieux) to identify bacteria isolates directly from urine samples. Methods We analyzed 352 clinical samples of urine. From automated screening by UQ system was used 500 µL of homogenized urine sample manually pipetted into a UQ vial containing 4 mL of broth. The inoculated vials were manually introduced into the Uro-Quick reading unit and incubated for 3 h. The signals are processed by software which monitors the growth curves and calculates the microbial count as colony forming units (CFU/mL). Sample preparation to the mass spectrometry (Vitek-MS) was performed using an in-house protocol based on a centrifugation-wash (CW) method. In parallel with the Vitek-MS direct identification, all the urine samples were submitted to standard culture (SC) method on ChromID™CPS® plate by semi-quantitative technique using 1 µL disposable loop, incubated at 35–37°C and the colony counting were read after 24 and 48h of incubation. Bacterial isolates from positive cultures were identified by Vitek®2 System, according to the manufacture’s recommendations. Cultures with the presence of three or more microorganisms were considered negative. Results Out of 334 clinical samples, 261 (78.1%) were negative for the growth by UQ system and for the presence of microorganisms by SC, 78 (21.9%) samples were positive with growth as ≥104 CFU/mL by UQ system, of which 32 samples were also positive with microorganism isolated by SC. Among the 78 microorganisms detected on the samples with growth curve by UQ system, 74 (94.8%) were identified to the species level by the mass spectrometry and 4 (5.2%) were not identified. The microorganisms were Escherichia coli (n = 40), Enterococcus faecalis (n = 12), Klebsiella pneumoniae (n = 6), Proteus mirabilis (n = 4), Citrobacter koseri (n = 4), Streptococcus agalactiae (n = 3), Morganella morganii (n = 2), Enterobacter cloacae (n = 1), Citrobacter freundii (n = 1) and Gardnerella vaginalis (n = 1). Among the 32 microorganisms detected by SC, 28 showed the same identification at species level when was compared with direct identification by Vitek-MS. Conclusion This study showed that both UQ and CW methods by Vitek-MS technology used together were able to provides a rapid result (<4 h) for bacterial identification directly from urine samples, that can be useful in clinical practices, especially in severe infectious cases like pyelonephritis. So, future studies should address with a larger number of samples to improve the sensibility of this test for different groups of microorganisms.

A Single-laboratory Performance Evaluation of MALDI-TOF MS in Rapid Identification of Staphylococcus aureus, Cronobacter sakazakii, Vibrio parahaemolyticus, and Some Closely Related Bacterial Species of Public Health Importance.

Staphylococcus is a genus of Gram-positive bacteria, known to cause food poisoning and gastrointestinal illness in humans. Additionally, the emergence of methicillin-resistant S. aureus (MRSA) strains have caused major healthcare burden worldwide. Cronobacter is a group of Gram-negative bacteria, that can survive in extreme dry conditions. Cronobacter sakazakii is known to contaminate the powdered infant formula and cause life-threatening infections in neonates. Vibrio is a genus of human-pathogenic Gram-negative bacteria that can cause foodborne illness by consuming undercooked or raw seafood. Vibrio parahaemolyticus can cause serious gastrointestinal disease in humans. Thus, rapid identification of Staphylococcus spp., Cronobacter spp. and Vibrio spp. is crucial for the source tracking of contaminated food, and to measure the transmission dynamics of these bacterial pathogens causing foodborne diseases and outbreaks. This single-laboratory performance evaluation study used the VITEK MS system to evaluate the potential of MALDI-TOF MS technology for rapid identification of S. aureus-like, C. sakazakii-like, and V. parahaemolyticus-like, isolates of public health importance. A total of 226 isolates recovered from various food, environmental surveillance samples and other sources were identified by bioMérieux VITEK 2 and VITEK MS systems as Staphylococcus spp., Cronobacter spp., and Vibrio spp. Five American Type Culture Collection (ATCC) reference Gram-positive and Gram-negative bacterial isolates were also tested to complete the study. In addition, for some Staphylococcus spp. isolates whole genome sequencing (WGS), and DNA sequencing of 16S rRNA partial region was also performed for species identification. The VITEK MS system was able to provide species identification to all 96 isolates of Staphylococcus spp. and to all 29 isolates of Vibrio spp. examined with a high confidence value (99.9%). Similarly, species identification was observed for the majority of spots (245 of 303) for the 101 Cronobacter spp. isolates (∼82.0%) with a high confidence value (99.9%), and genus level identification was noticed for the rest of the Cronobacter spp. isolates (18.0%; 58 of the 303 spots) analyzed. Species identification data generated by VITEK 2 system was comparable to data obtained by VITEK MS system. The VITEK MS system is a reliable high-throughput platform that can rapidly identify Staphylococcus, Vibrio, and Cronobacter to the genus level, as well as S. aureus, C. sakazakii, V. parahaemolyticus, and other closely related foodborne isolates, and bacterial isolates from additional sources, in most cases. The VITEK MS system can be used in the rapid genus and species identification of human-pathogenic Staphylococcus spp., Cronobacter spp., and Vibrio spp. isolates.

PERFORMANCE OF THE VITEK MS MATRIX-ASSISTED LASER DESORPTION IONIZATION-TIME OF FLIGHT MASS SPECTROMETRY SYSTEM FOR RAPID IDENTIFICATION OF CLINICAL MICROBIOLOGY ISOLATES

Background: Rapid and precise microbial identification is essential for timely patient management and control of infection. Conventional microbiological procedures are time-consuming, laborious, and require expertise. Recent coalition of MALDI-TOF in the microbiology workflow has yielded excellent results so far. Here we have assessed this quick proteomic based technique for clinical bacterial isolates identification. Material and Methods: All fresh bacterial isolates were selected for VITEK MS (bioMérieux, France) analysis, the bacteria were identified by using MALDI-TOF System (IVD 3.2) software. For same isolates, API (bioMérieux, France) was set as per kit instructions and conventional/biochemical tests were performed according to ASM Guidelines. Results: Among 200 isolates 99% were accurately identified by the VITEK MS system to the species level, one of two isolates was mis-identified (Shigella as E. coli) while the other was identified later by re-spotting. On testing these samples in parallel by APIs, 91.50% were correctly identified, while 8.50% (17 samples) showed discrepant results. These were re-analyzed by VITEK-2 (bioMérieux, France) semi-automated system which showed the same results as those of VITEK MS. Our findings revealed diagnostic accuracy of VITEK MS in comparison with APIs in terms of time, cost and patient management. Conclusion: For bacterial identification MALDI-TOF MS is an expeditious, authentic and comparatively inexpensive system. Our results emphasize that it is speedy technique which can replace the traditional identification methods for most of bacterial strains on their routine isolation. This ingenious approach complies with advanced patient management and therapeutic intervention. Key Words: MALDI-TOF MS, bacterial identification, VITEK-2, Analytical profile index (API), Clinical microbial isolates

Performance of Two Matrix-Assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS) Systems for Identification of the Viridans Group Streptococci.

Due to similar colony morphology among viridans group streptococci (VGS), the differentiation of VGS species remains difficult in routine clinical microbiology. Recently, matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) has been described as a fast method for identifying various bacteria at species level, and also for the VGS strains. A total of 277 VGS isolates were identified with the two MALDI-TOF MS systems (VITEK MS and Bruker Biotyper). The tuf and rpoB gene sequencing was used as the reference identification method for comparison. Based on tuf and rpoB gene sequencing, 84 isolates were S. pneumoniae and 193 strains were other VGS isolates including S. anginosus group (n=91, 47.2%), S. mitis group (n=80, 41.5%), S. bovis group (n=11, 5.7%), S. salivarius group (n=10, 5.2%), and S. mutans group (n=1, 0.5%). VITEK MS and Bruker Biotyper accurately identified 94.6% and 89.9% of all VGS isolates, respectively. VITEK MS showed better identification results than Bruker Biotyper for S. mitis group including S. pneumoniae and S. bovis group, but for other VGS isolates, two MALDI-TOF MS systems showed comparable identification performance. However, VITEK MS was able to identify S. gallolyticus to the subspecies level with high-confidence (S. gallolyticus ssp. pasteurianus), while the Bruker Biotyper system could not. While Bruker Biotyper system could be able to correctly differentiate the subspecies of S. salivarius from S. vestibularis, VITEK MS poorly identify. This study demonstrated that two MALDI-TOF MS systems allowed discrimination for most VGS isolates with different identification performance, but Bruker Biotyper could produce more misidentifications and VITEK MS system. It is crucial to be familiar with the performance of MALDI-TOF MS systems used in clinical microbiology.

Antibiotic susceptibility of Pseudomonas aeruginosa isolated at 108 Military Central Hospital from 01/2020 to 06/2022

Objective: To determine antibiotic susceptibility patterns of Pseudomonas aeruginosa isolated at the 108 Military Central Hospital from January 2020 to June 2022. Subject and method: A total of 784 Pseudomonas aeruginosa strains were isolated from patient specimens from different departments. Bacteria identification and antibiotic susceptibility test were performed using the Vitek MS system and the Vitek-2 Compact system, respectively. Multidrug-resistant and/or carbapenem-resistant Pseudomonas aeruginosa (MDR/CPR P. aeruginosa) strains were determined. Colistin susceptibility test was done using cation-modified Muller-Hinton broth diffusion method (CBDE, colistin broth disk elution). Result: Pseudomonas aeruginosa was highly resisting to fluoroquinolones (62.8%), and aminoglycosides (53.4%). The proportion of MDR/CPR P. aeruginosa was 54.3% (426/784), of which the highest proportion was found in the Department of Infectious Diseases (65.52%), ICU (64.39%) and the Internal Respiratory Department (45.69%). Carbapenem-resistant Pseudomonas aeruginosa was highly prevalent in urine specimens (65.38%) and respiratory specimens (56.76%). Of 258 MDR/CPR P. aeruginosa isolates, 11.2% isolates were resistant to colistin, 26.6% and 33.3% of these remained susceptible to amikacin and piperacillin/tazobactam, respectively. Conclusion: The prevalence of antibiotic resistante Pseudomonas aeruginosa remains high, including resistance to carbapenem and colistin. Antibiotic regimen should include amikacin or piperacillin/tazobactam in combination with other antibiotics. The study suggested the routine application of diagnostic tools for the rapid detection of resistant strains.

Accurate Identification and Virulence Detection of Aeromonas: a Single-Center Retrospective Study on the Clinical Features and Outcomes Associated with Aeromonas Bacteremia in Southwestern China.

In this study, Aeromonas spp. were re-identified, and the clinical aspects associated with Aeromonas bacteremia, as well as drug resistance and virulence genes, were elucidated. A total of 188 isolates were classified into 7 Aeromonas spp. using housekeeping gene sequencing, which was the standard to assess the accuracy of the VITEK MALDI-TOF system and the VITEK2 Compact system. The VITEK MS system and housekeeping gene sequencing had a 39.89% clear coincidence rate, whereas the VITEK2 Compact system and the standard had a 2.13% coincidence rate. Aeromonas bacteremia was associated with septic shock, hematologic malignancy, and post-hepatitic cirrhosis. Hematological malignancy, hypoproteinemia, systemic steroid use, central venous catheterization, and virulence genes act and ast were linked to poor outcomes. Aeromonas bacteremia had a 37.5% mortality rate; however, differences in mortality rates among Aeromonas spp. were observed. According to the broth microdilution method, over 90% of isolates were sensitive to most antimicrobials, except ceftriaxone (83.33%) and imipenem (83.33%). Polymerase chain reaction and DNA sequencing verified the presence of drug resistance genes; blaCphA was detected in 3 isolates, while blaNDM-1 was found in one isolate. In summary, common methods for identifying Aeromonas spp. are ineffective. Immunocompromised patients have a higher risk of infection and mortality. Furthermore, carbapenem resistance is a serious problem.

Option appraisal of matrix-assisted laser desorption ionisation time of flight systems for a diagnostic microbiology laboratory

A major challenge for a clinical diagnostic bacteriology/mycology laboratory is to identify the causal microorganism to the species level within a limited time frame. Conventional microbiological techniques such as culture, Gram stain, biochemical tests and molecular tests like 16S rRNA analysis can take days to yield the desired result. Matrix-assisted laser desorption ionisation-time of flight (MALDI-TOF) mass spectrometry (MS) system serves as a potential solution as it can provide accurate identification results within minutes. While several MALDI-TOF MS instruments are in the market, choosing the right system and comparing all the relative pros and cons can be difficult. Our present study focuses on this aspect as we have performed an option appraisal between two leading MALDI-TOF MS instruments, namely the MBT Sirius IVD system (Bruker) and the VITEK-MS system (Biomerieux). We have discussed all the parts of MALDI-TOF systems in detail followed by a cost analysis categorising the total cost into capital and consumable cost which will provide clarity of the estimated budget. We have also discussed the principle involved in organism identification and have compared the two systems concerning technology, accuracy, database, optimal operating conditions, etc. This article aims to provide readers with a better understanding of MALDI-TOF systems and facilitate informed decision-making regarding the acquisition of such systems.

Comparison of MALDI-TOF mass spectrometry and rpoB gene sequencing for the identification of clinical isolates of Aeromonas spp.

Aeromonas spp., widely present in rivers and soil, cause mild gastroenteritis, severe septicemia, and soft tissue infections in humans. Treatment of these infections require accurate identification of pathogenic Aeromonas spp. However, identification at the species level using conventional methods is highly challenging. In this study, we aimed to compare the accuracy of two different approaches developed for bacterial identification: (i) housekeeping gene sequencing (rpoB) in conjunction with phylogenetic analysis and (ii) matrix-assisted laser desorption ionization mass spectrometry-time of flight (MALDI-TOF MS) (MALDI Biotyper and VITEK MS), for differentiating Aeromonas spp. We analyzed 58 Aeromonas isolates recovered from patients at different medical institutions in Japan using both identification methods. The rpoB sequencing method was the most accurate, identifying all Aeromonas isolates at the species level. Meanwhile, the MALDI Biotyper system correctly identified 53 (91.4%) isolates at the genus level and an additional 30 (51.7%) at the species level. The VITEK MS system correctly identified 58 (100%) isolates at the genus level and an additional 34 (58.6%) at the species level. Thus, MALDI Biotyper and VITEK MS accurately identified isolates at the genus level, but differences were found in the accuracy of identification of species. However, the low cost and ease of analysis make MALDI-TOF MS-based methods strong candidates for use in clinical laboratories that require easy-to-use identification methods.

A side-by-side comparison of the performance and time-and-motion data of VITEK MS.

Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry systems are designed for rapid and reliable microbial identification. VITEK MS PRIME is the bioMérieux's new generation instrument equipped with a continuous load-and-go sample loading system, urgent slide prioritization for critical patient samples and new internal components for faster identification. The aim of this study was to assess the performance of VITEK MS PRIME and to compare it to that of the VITEK MS system. In addition, at two sites, we performed a time-and-motion study to evaluate the efficiency of sample analysis from colony picking to slide removal from the instrument. We analyzed by VITEK MS and VITEK MS PRIME a total of 1413 isolates (1320 bacterial and 76 yeast) deriving from routine diagnostic samples that came into four laboratories in Canada, France, Italy, and Spain. VITEK MS PRIME and VITEK MS were concordant to the species and genus level for 1354/1413 (95.8%) and to the species level for 1341/1413 (94.9%). The identification and concordance rates in individual centers were largely homogenous. Overall, VITEK MS PRIME identified 1370/1413 (97.0%) of isolates compared to 1367/1413 (96.7%) identified by VITEK MS. Identification rates were consistently high for all microorganism categories. A time-and-motion study showed that the use of VITEK MS PRIME was associated with significant time saving. VITEK MS PRIME performs as well as VITEK MS and reduces the time necessary for pathogen identification. To fully optimize the laboratory process and obtain maximum efficiency, VITEK MS PRIME must be integrated into the laboratory workflow.

Performance of BD MAX Group B Streptococcus (GBS) Assay without Enrichment for the Detection of GBS.

Group B streptococcus (GBS) is an important pathogen causing neonatal early-onset disease. We evaluated the diagnostic performance of BD Max GBS assay (Becton Dickinson, Franklin Lakes, NJ, USA) without enrichment (direct BDM) for detecting GBS using vaginal and rectal specimens in comparison with culture. In total, 716 specimens collected from 358 pregnant women between June 2018 and May 2020 were included in this study. Bacterial culture was performed using ChromID Strep B agar (bioMérieux, Marcy-l’Étoile, France), and species identification results were confirmed using the VITEK-MS system (bioMérieux). The sensitivity of direct BDM for vaginal and rectal specimens was 75.0% and 100%, respectively. Thirteen specimens showed discrepant results: 10 false-negative results in the vaginal specimens and three false-positive results in the rectal specimens. The overall agreement between direct BDM and culture was 98.9% (354/358). The final sensitivity and specificity of direct BDM were 98.5% and 99.0%, respectively. Discrepant results—one false-negative and three false-positives—were obtained for four specimens. Direct BDM shows a good diagnostic performance and will be useful for GBS screening within a few hours.

Evaluation of the Vitek Ms system for the identification of filamentous fungi.

The use of MALDI-TOF for fungal identification remains a challenge. In this study, using a commercial protein extraction kit and updated database, VITEK MS system was able to identify up to 89.0% of a diverse collection of 200 filamentous fungi representing 43 fungal species.

The dilemma of identifying Peptoniphilus species by using two MALDI-TOF MS systems

Two commercial MALDI-TOF MS systems were used to identify 18 isolates, belonging to the Peptoniphilus genus; also the 16S rRNA sequencing identity was compared against the MALDI-TOF MS system results. Bruker Biotyper system provided higher accuracy than Vitek MS system, however, adding spectra could allow a more reliable species level identification.

Application of MALDI-TOF mass spectrometry, and DNA sequencing-based SLST and MLST analysis for the identification of Cronobacter spp. isolated from environmental surveillance samples.

Cronobacter spp. are emerging infectious foodborne bacteria that can cause acute meningitis and necrotizing enterocolitis in neonates and immunocompromised individuals. Although, little is known about its reservoirs or transmission routes, it has been linked to powdered infant formula worldwide. Three Cronobacter spp. (C. sakazakii, C. malonaticus, and C. turicensis) have been described as more virulent, and isolated frequently from infant meningitis cases. The estimated mortality rates are as high as 80% in infants. Thus, surveillance and typing of Cronobacter spp. isolated from food and environmental samples is essential to prevent contamination and spread of this pathogen. In this study, we have characterized 83 Cronobacter isolates recovered from various environmental samples by conventional microbiologic protocols. Species identification was accomplished by VITEK 2 system and real-time PCR analysis. Subsequently, these isolates were analyzed using VITEK MS system. Single locus sequence typing (SLST) was achieved by characterizing the regions of 16S rRNA and rpoB genes. Multilocus sequence typing (MLST) was performed by sequence characterization of seven housekeeping genes (atpD, fusA, glnS, gltB, gyrB, infB, and pps) using ABI 3500XL Genetic Analyzer. VITEK MS system identified, the majority of isolates as Cronobacter sakazakii with a high confidence value (99.9%). MLST analysis ascertained 12 distinct clonal complexes (CC1, CC4, CC8, CC13, CC17, CC21, CC31, CC40, CC52, CC64, CC73, and CC83) for the recovered C. sakazakii isolates. The results suggest that the MALDI-TOF MS is a reliable diagnostic tool for rapid species identification whereas 7-loci MLST is a powerful technique to discriminate and differentiate Cronobacter spp. isolates.

MALDI-TOF Mass Spectrometry and 16S rRNA Gene Sequence Analysis for the Identification of Foodborne Clostridium Spp.

Clostridium is a genus of Gram-positive, spore-forming, anaerobic bacteria comprising approximately 100 species. Some Clostridium spp. (C. botulinum, C. perfringens, C. tetani, and C. difficile) have been recognized to cause acute food poisoning, botulism, tetanus, and diarrheal illness in humans. Thus, rapid identification of Clostridium spp. is critical for source-tracking of contaminated food and to understand the transmission dynamics of these foodborne pathogens. This study was carried out to rapidly identify Clostridium-like isolates by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS and rRNA sequencing methods. Thirty-three Clostridium-like isolates were recovered from various baby food and surveillance samples. Species identification of these isolates was accomplished using the VITEK MS system. Sequence characterization of the 16S rRNA region was done on an ABI 3500xL Genetic Analyzer. The VITEK MS system identified 28 of the 33 Clostridium-like isolates with a high confidence value (99.9%); no identification was observed for the remaining five isolates. Nucleotide sequencing of the 16S rRNA region identified all 33 Clostridium-like isolates. Furthermore, while characterizing the 16S rRNA gene, 11 distinct Clostridium spp. (Clostridium aciditolerans, Clostridium aerotolerans, Clostridium argentinense, Clostridium beijerinckii, Clostridium bifermentans, Clostridium butyricum, Clostridium cochlearium, Clostridium difficile, Clostridium perfringens, Clostridium sporogenes, and Clostridium subterminale) were recognized among the 33 Clostridium-like isolates. One of the Clostridium-like isolates was identified as Citrobacter amalonaticus by both diagnostic methods. The generated 16S rRNA sequences matched completely (100%) with sequences available in GenBank for Clostridium and Citrobacter species. Species identification attained using the VITEK MS for the Clostridium-like isolates was comparable to that from the 16S rRNA sequencing-based data. The VITEK MS and 16S rRNA sequence analysis can be implemented in the species identification of Clostridium spp. isolates of public health importance. MALDI-TOF MS and 16S rRNA sequencing can be used in the species identification of Clostridium species.

Comparative evaluation of the Bruker Biotyper and Vitek MS matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) systems for non-albicans Candida and uncommon yeast isolates

IntroductionRapid and accurate diagnosis is critically important in invasive and disseminated fungal infections for appropriate antifungal treatment. HypothesisMALDI-TOF MS systems are effective for fast and accurate identification of Candida species. AimWe aimed to compare two MALDI-TOF MS systems for the rapid identification of non-albicans Candida and rare clinical yeast species. MethodologyThis study included 157 isolates representing 23 yeast species. All isolates were identified using Bruker MALDI Biotyper and VITEK MS systems. If both MALDI-TOF MS systems yielded the same results for a certain isolate, the identification is regarded as correct. We performed internal transcribed spacer (ITS) DNA sequencing on five fungal isolates with discordant species names or that were unidentified by the two MALDI-TOF MS systems. ResultsThe yeast identification sensitivity of MALDI Biotyper was 98.7%, whereas that of VITEK MS was 96.8%. Both MALDI-TOF MS systems correctly identified all strains belonging to four prevalent species, namely, Candida parapsilosis, Candida tropicalis, Candida glabrata, and Candida krusei. For the 19 rare clinical yeast species, identification rates were 96.7% for MALDI Biotyper and 91.7% for VITEK MS. The ITS sequence analysis of five isolates yielded two Meyerozyma caribbica, two Cyberlindnera fabianii, and one Candida dubliniensis. ConclusionsThis study showed the high performance of both MALDI-TOF MS systems, identifying over 90% of yeast isolates in a short time. The disadvantages of these systems are that some species are not present in the databases and it cannot distinguish closely related species. The sensitivity of MALDI-TOF MS systems constantly improves with the expansion of databases in parallel with taxonomic developments for the identification of rare clinical yeast species.

Evaluation of Autof MS 1000 and Vitek MS MALDI-TOF MS System in Identification of Closely-Related Yeasts Causing Invasive Fungal Diseases.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been accepted as a rapid, accurate, and less labor-intensive method in the identification of microorganisms in clinical laboratories. However, there is limited data on systematic evaluation of its effectiveness in the identification of phylogenetically closely-related yeast species. In this study, we evaluated two commercially available MALDI-TOF systems, Autof MS 1000 and Vitek MS, for the identification of yeasts within closely-related species complexes. A total of 1,228 yeast isolates, representing 14 different species of five species complexes, including 479 of Candida parapsilosis complex, 323 of Candida albicans complex, 95 of Candida glabrata complex, 16 of Candida haemulonii complex (including two Candida auris), and 315 of Cryptococcus neoformans complex, collected under the National China Hospital Invasive Fungal Surveillance Net (CHIF-NET) program, were studied. Autof MS 1000 and Vitek MS systems correctly identified 99.2% and 89.2% of the isolates, with major error rate of 0.4% versus 1.6%, and minor error rate of 0.1% versus 3.5%, respectively. The proportion of isolates accurately identified by Autof MS 1000 and Vitek MS per each yeast complex, respectively, was as follows; C. albicans complex, 99.4% vs 96.3%; C. parapsilosis complex, 99.0% vs 79.1%; C glabrata complex, 98.9% vs 94.7%; C. haemulonii complex, 100% vs 93.8%; and C. neoformans, 99.4% vs 95.2%. Overall, Autof MS 1000 exhibited good capacity in yeast identification while Vitek MS had lower identification accuracy, especially in the identification of less common species within phylogenetically closely-related species complexes.