Bruker MALDI Biotyper Research Articles

Hazard Characterization of Antibiotic-resistant Aeromonas spp. Isolated from Mussel and Oyster Shellstock Available for Retail Purchase in Canada

Surveillance and monitoring of foods for the presence of antimicrobial-resistant (AMR) bacteria are required to assess the risks these bacteria pose to human health. Frequently consumed raw or lightly cooked, live bivalve shellfish such as mussels and oysters can be a source of exposure to AMR bacteria. This study sought to determine the prevalence of third-generation cephalosporin (3GC) and carbapenem-resistant bacteria in live mussel and oyster shellstock available for retail purchase through the course of one calendar year. Just over half of the 180 samples (52%) tested positive for the presence of 3GC-resistant bacteria belonging to thirty distinct bacterial species. Speciation of the isolates was carried out using the Bruker MALDI Biotyper. Serratia spp., Aeromonas spp., and Rahnella spp. were the most frequently isolated groups of bacteria. Antibiotic resistance testing confirmed reduced susceptibility for 3GCs and/or carbapenems in 15 of the 29 Aeromonas isolates. Based on AMR patterns, and species identity, a subset of ten Aeromonas strains was chosen for further characterization by whole genome sequence analysis. Genomic analysis revealed the presence of multiple antibiotic resistance and virulence genes. A number of mobile genetic elements were also identified indicating the potential for horizontal gene transfer. Differences in gene detection by the bioinformatic tools and databases used (ResFinder. CARD RGI, PlasmidFinder, and MobSuite) are discussed. This study highlights the strengths and limitations of using genomics tools to perform hazard characterization of diverse foodborne bacterial species.

Application of MALDI-TOF mass spectrometry for identification of Nocardia species

BackgroundNocardiosis, despite its rarity and underreporting, is significant due to its severe impact, characterized by high morbidity and mortality rates. The development of a precise, reliable, rapid, and straightforward technique for identifying the pathogenic agent in clinical specimens is crucial to reduce fatality rates and facilitate timely antimicrobial treatment. In this study, we aimed to identify Nocardia spp. in clinical isolates, using MALDI-TOF MS as the primary method, with molecular methods as the gold standard. Clinical Nocardia isolates were identified using 16S rRNA/hsp65/gyrB/secA1/rpoB gene sequencing. Identification performance of the Bruker MALDI Biotyper 3.1 (V09.0.0.0_8468) and MBT Compass 4.1 (V11.0.0.0_10833) for Nocardia identification was evaluated.ResultsSeventy-six Nocardia isolates were classified into 12 species through gene sequencing. The MALDI Biotyper 3.1 (V09.0.0.0_8468) achieved 100% genus-level accuracy and 84.2% species accuracy (64/76). The MBT Compass 4.1 with the BDAL Database (V11.0.0.0_10833) improved species identification to 98.7% (75/76). The updated database enhanced species level identification with scores > 1.7, increasing from 77.6% (59/76) to 94.7% (72/76), a significant improvement (P = 0.001). The new and simplified extraction increased the proportion of strains identified to the species level with scores > 1.7 from 62.0% (18/29) to 86.2% (25/29) (P = 0.016). An in-house library construction ensured accurate species identification for all isolates.ConclusionsThe Bruker mass spectrometer can accurately identify Nocardia species, albeit with some variations observed between different database versions. The MALDI Biotyper 3.1 (V09.0.0.0_8468) has limitations in identifying Nocardia brasiliensis, with some strains only identifiable to the genus level. MBT Compass 4.1 (V11.0.0.0_10833) effectively addresses this shortfall, improving species identification accuracy to 98.7%, and offering quick and reliable identification of Nocardia. Both database versions incorrectly identified the clinically less common Nocardia sputorum as Nocardia araoensis. For laboratories that have not upgraded their databases and are unable to achieve satisfactory identification results for Nocardia, employing the new and simplified extraction method can provide a degree of improvement in identification outcomes.

Intact cell lipidomics using the Bruker MBT lipid Xtract assay allows the rapid detection of glycosyl-inositol-phospho-ceramides from Aspergillus fumigatus.

Glycosyl-inositol-phospho-ceramides (GIPCs) or glycosylphosphatidylinositol-anchored fungal polysaccharides are major lipids in plant and fungal plasma membranes and play an important role in stress adaption. However, their analysis remains challenging due to the multiple steps involved in their extraction and purification prior to mass spectrometry analysis. To address this challenge, we report here a novel simplified method to identify GIPCs from Aspergillus fumigatus using the new Bruker MBT lipid Xtract assay. A. fumigatus reference strains and clinical isolates were cultured, harvested, heat-inactivated and suspended in double-distilled water. A fraction of this fungal preparation was then dried in a microtube, mixed with an MBT lipid Xtract matrix (Bruker Daltonik, Germany) and loaded onto a MALDI target plate. Analysis was performed using a Bruker MALDI Biotyper Sirius system in the linear negative ion mode. Mass spectra were scanned from m/z 700 to m/z 2 000. MALDI-TOF MS analysis of cultured fungi showed a clear signature of GIPCs in Aspergillus fumigatus reference strains and clinical isolates. Here, we have demonstrated that routine MALDI-TOF in the linear negative ion mode combined with the MBT lipid Xtract is able to detect Aspergillus fumigatus GIPCs.

2485. Candida auris Hospital Cluster: Were Our Unwelcome Guests Driven Away by the COVID-19 Delta Surge?

Abstract Background Candida auris (CA) is an MDR pathogen that persists in the environment, causing outbreaks. NUH is an academic medical centre in Singapore. We describe a cluster of CA cases from April 2019 – October 2021. Our last case was during our Delta surge, just before we changed our routine inpatient cleaning to bleach based from quarternary-ammonium. Methods Any CA from the micro lab is escalated to the Infection Prevention Team to initiate isolation, cleaning, and contact tracing (CT) up to 30 days prior to positive cultures. In September 2020, we started enhanced surveillance via NAG (Nasal, axilla and groin) swabs in all new CRE (Carbapenem-Resistant Enterobacterales) patients. Surveillance cultures used sabouraud dulcitol agar with 10% saline incubated at 40°C. Identification and susceptibility testing done using Bruker MALDI Biotyper and Etest (Biomerieux). Hospital onset (HO) is defined as positive cultures sent after day 3 of admission. CA isolates were sent for whole genome sequencing (WGS). Results The 10 cases included 7 from clinical isolates, 2 from enhanced surveillance and 1 from CT. 8 were HO. 7 deaths were during the CA admission, but only 1 due to CA, and 1 occurred > 60 days after positive culture [Table 1]. Of 231 contacts swabbed all were negative, except for Case 8, in whom one contact (Case 9) was positive. Positive yield from enhanced surveillance of CRE patients, was 0.84% (2/239). WGS results indicated all except Case 9 belonged to Clade I with Y132F mutation in ERG111 gene [Table 2]. Cases 2, 3, 5, 6 were closely related with 2-7 SNPs difference, but spaced in years. Cases 7 and 8 differed by 3 SNPs hence, closely related. There were 2 related clones being transmitted, and 4 unrelated sub-types of CA. Case 8 and 9 were not related, though 9 was a contact of 8. Possible links were shared equipment or admission to a common room however, admissions were either non-overlapping or non-successive [Table 3]. Case 5 was admitted to the same room after a long stay by Case 3, with one patient in between. No links found between Cases 7 and 8. Conclusion WGS results suggest that various clones of CA remain in the environment as reservoirs, and related cases are not often spatiotemporally related. There have been no CA cases since Oct 2021, possibly due to the change to bleach-based cleaner during our Delta surge. Disclosures All Authors: No reported disclosures

614. Experience with Using MALDI-TOF MS to Diagnose Clinically Significant Filamentous Fungal Isolates

Abstract Background Matrix assisted laser desorption/ionisation time of flight mass spectrometry (MALDI-TOF MS) technology has revolutionised bacterial diagnostics. Filamentous fungi are a varied and important group of pathogens that cause a wide spectrum of clinical disease. Identification of these traditionally relies upon highly experienced practitioners identifying unique fungal morphological characteristics, along with specific molecular techniques, but MALDI-TOF MS may be a useful alternative. The aim of this retrospective review was to determine how useful MALDI-TOF MS was as an adjunct to filamentous fungi identification at our laboratory service. Methods All fungal diagnoses made at St Vincent’s Pathology Melbourne, a large metropolitan laboratory servicing multiple tertiary hospitals and community pathology centres, from 2018 to 2022 were extracted. For clinically significant filamentous fungal isolates, the method of diagnosis was reviewed and categorised into; morphological diagnosis, Bruker MALDI Biotyper® diagnosis, and national reference laboratory diagnosis. All clinically significant isolates where MALDI-TOF MS was attempted were also reviewed. Results A total of 256 clinically significant isolates were extracted and reviewed. MALDI-TOF MS was attempted in 59 of these, with success in 39 (66.1%). Of the 20 cases where MALDI-TOF MS was attempted but did not identify the isolate, 16 were diagnosed morphologically and four required send out to the national reference laboratory. 71.2% of MALDI-TOF MS attempts were in the 2021-22 period when successful identification occurred 80.9% of the time. No species/genus appeared more or less readily identifiable. Of the remaining cases, 208 were identified morphologically and nine using the national reference laboratory. Conclusion MALDI-TOF MS performed modestly well at identifying filamentous fungi during this study period. Increased use and successful isolate identification in the 2021-22 period likely reflects improvement in the Bruker MALDI-TOF MS database for filamentous fungi. The MALDI-TOF MS system is a useful option in the identification of clinically significant filamentous fungi however morphological and molecular techniques remain important. Disclosures All Authors: No reported disclosures

Microbial Load of Hand Sanitizer Dispensers—A University Hospital Study

Hospital-acquired infections are a significant concern in healthcare settings, leading to patient safety risks, increased morbidity and mortality, and financial burdens. Hand hygiene is crucial in preventing the spread of bacteria in hospitals and communities. Manual hand sanitizer dispensers can harbor presumptive pathogenic bacteria and act as fomites for bacterial transmission. This study aimed to assess the microbial contamination of manual hand sanitizer dispensers in a hospital setting and to study their antibiotic resistance profiles. Samples were collected using sterile cotton swabs and then inoculated into brain heart infusion broth. Subsequent subcultures were performed on both blood and MacConkey agar. The isolates were then identified using the Bruker MALDI Biotyper (Bruker Daltonik, Bremen, Germany) to the species level. Sampling was conducted in various wards and in the hospital and the University areas on dispenser levers and nozzle areas. The results showed that all samples yielded one or more bacterial species. Bacterial isolates identified belonged to species commonly found on the skin microflora and some Gram-negative enteric bacilli. Higher colonization was observed on the dispenser lever. Among Gram+ microorganisms, most bacterial species were shown to be sensitive to β-lactams, with the exception of Staphylococcus spp., resistant to AMP (Ampicillin) and Penicillin. However, no Methicillin resistant isolates were detected. Gram− microorganisms such as Pseudomonas luteola were shown to be sensitive to all tested antibiotics, while Pantoea agglomerans was shown to be resistant to AMC (amoxicillin–clavulanic acid). Rifampicin tested only against Bacilli showed resistance. Based on the findings, it is recommended to implement systematic cleaning and proper maintenance of manual dispenser areas or to use automated dispensers to reduce hand contact and minimize microbial contamination. Monitoring the presence of microorganisms in hand sanitizing gels and dispensers is an essential infection control strategy.

An easy adjustment of instrument settings (‘Peak MALDI’) improves identification of organisms by MALDI-ToF mass spectrometry

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF MS) is a mature technology with ‘auto-execute’ instrument settings and peak processing parameters tailored for rapid bacterial identification. Adoption for other organisms has been problematic, with optimisation efforts focusing on sample preparation. Using the Bruker MALDI Biotyper, we demonstrate ‘Peak MALDI’: easily-applied settings that immediately enhance sensitivity, improve spectrum quality, and increase identification confidence for any target, establishing its potential value for all MALDI-ToF MS systems.

Improved identification of Streptococcus bovis-Streptococcus equinus-complex species and subspecies by MALDI-TOF MS using a novel library

ObjectivesTo develop an in-house matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) library for improved identification of species and subspecies of the Streptococcus bovis/Streptococcus equinus-complex (SBSEC). MethodsA total of 236 SBSEC isolates from blood stream infections and culture collections, determined by whole genome sequencing to subspecies level, were grown in brain heart infusion broth. Mass spectra were collected using the Bruker MALDI Biotyper system after ethanol-formic acid extraction. Main spectral profiles from 117 isolates were used to create the “SBSEC-CMRS library.” The remaining 119 spectra were used for evaluation of Bruker MALDI Biotyper (MBT) Compass Library Revision K (2022) and the SBSEC-CMRS library. ResultsThe Bruker library correctly identified species and subspecies in 72 of 119 (61 %) isolates, while the SBSEC-CMRS library identified 116 of 119 (97 %), using a cutoff score of ≥2.0. ConclusionsThe SBSEC-CMRS library showed sufficient diagnostic accuracy, and can be implemented in clinical practice for SBSEC species and subspecies identification.

Identification of Francisella tularensis Subspecies in a Clinical Setting Using MALDI-TOF MS: An In-House Francisella Library and Biomarkers.

Francisella tularensis is a zoonotic bacterium that is endemic in large parts of the world. It is absent in the standard library of the most applied matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) systems: the Vitek MS and the Bruker Biotyper system. The additional Bruker MALDI Biotyper Security library contains F. tularensis without subspecies differentiation. The virulence of F. tularensis differs between the subspecies. The F. tularensis subspecies (ssp.) tularensis is highly pathogenic, whereas the subspecies holarctica displays lower virulence and subspecies novicida and F. tularensis ssp. mediasiatica are hardly virulent. To differentiate the Francisellaceae and the F. tularensis-subspecies, an in-house Francisella library was built with the Bruker Biotyper system and validated together with the existing Bruker databases. In addition, specific biomarkers were defined based on the main spectra of the Francisella strains supplemented with in silico genome data. Our in-house Francisella library accurately differentiates the F. tularensis subspecies and the other Francisellaceae. The biomarkers correctly differentiate the various species within the genus Francisella and the F. tularensis subspecies. These MALDI-TOF MS strategies can successfully be applied in a clinical laboratory setting as a fast and specific method to identify F. tularensis to subspecies level.

Performance of the MSI-2 Database for Fungal Identification by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry from Cleanroom Environments.

Accurate mold identification by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is dependent on robust organism representation in available databases. The Mass Spectrometry Identification (MSI) platform has proven successful for mold identification in clinical and veterinary settings but has yet to be studied with a large set of environmental isolates. Here, we performed a retrospective study using spectra collected by the Bruker MALDI Biotyper (MBT) v4.1 microflex LT instrument to evaluate the MSI-2 database alongside the combined use of the Bruker MBT (including the MBT Filamentous Fungi Library) and the National Institutes of Health (NIH) mold database (MBT/NIH databases). Analysis was performed for 462 environmental fungal isolates (representing 73 different fungi) cultured from the hospital pharmacy and cellular therapy suites as part of the current good manufacturing practices (cGMP) environmental monitoring program at the NIH. When used alone, MSI-2 identified 237 spectra (51.3%) at its higher score threshold (index A), while the MBT/NIH databases identified only 183 spectra (39.6%; P < 0.001) at the equivalent threshold of ≥2.0. The combination of all three databases at the respective high thresholds improved identification sensitivity to 327 spectra (70.8%). The combination of MSI-2 with the MBT/NIH databases at a lowered threshold (index B or ≥1.7, respectively) identified 400/462 environmental spectra (86.6%). Our results show that the MSI-2 database, in combination with existing databases, may be useful for environmental surveillance, particularly by clinical or industry laboratories involved in cGMP or current good tissue practices (cGTP) applications, such as cellular therapy manufacturing facilities and sterile compounding pharmacies.

A Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Direct-from-Urine-Specimen Diagnostic for Gram-Negative Pathogens.

Urinary tract infections (UTIs) pose a major public health burden. The vast majority of UTIs are caused by Gram-negative bacteria. Current culture-based pathogen identification methods may require up to 24 to 48 h of incubation. In this study, we developed and evaluated a method for Gram-negative pathogen identification direct from urine, without culture, via matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) in approximately 1 h. Urine samples were collected (n = 137) from the University of Maryland Medical Center clinical microbiology laboratory. To identify bacteria direct from urine, two methods were evaluated. First, 1 μL of urine was directly spotted onto the MALDI target plate, and second, 1 mL of urine was centrifuged at 8,000 rpm for 5 min before processing using the fast lipid analysis technique (FLAT). Mass spectra were acquired on the Bruker MALDI Biotyper sirius system in the negative-ion mode. Results were compared to those of standard culture methods. When 1 μL of urine was directly spotted, positive agreement was 81.5% (101/124) and, after centrifugation, 94.4% (117/124) relative to that of standard culture methods. Negative agreement for both methods was 100% (13/13). The time to results for both of the specimen preparation methods using the FLAT extraction protocol was approximately 1 h, with minimal hands-on time required (<5 min). The ability to rapidly identify pathogens directly from urine, without the need for culture, allows for faster turnaround times and, potentially, improved patient outcomes. Overall, the FLAT extraction protocol, in combination with lipid A identification, provides a reproducible and accurate method to rapidly identify urinary pathogens. IMPORTANCE This study describes and evaluates a direct-from-urine extraction method that allows identification of Gram-negative bacteria via MALDI-TOF MS within 1 h. Currently, identification of urinary pathogens requires 24 h of culture prior to identification. While this method may not replace culture, we demonstrate its utility in screening for common urinary pathogens. By providing identifications in under 1 h, clinicians can potentially treat patients sooner with more-targeted antimicrobial therapy. In turn, earlier treatment can improve patient outcome and antimicrobial stewardship. Furthermore, MADLI-TOF MS is a readily available, easy-to-use diagnostic tool in clinical laboratories, making implementation of this method possible.

W084 Assessment of a novel matrix-assisted desorption/ionization time-of-flight mass spectrometry platform, asta microidsys, for identification of various acinetobacter species, compared with bruker maldi biotyper

W084 Assessment of a novel matrix-assisted desorption/ionization time-of-flight mass spectrometry platform, asta microidsys, for identification of various acinetobacter species, compared with bruker maldi biotyper

Impact of MALDI-TOF MS identification on anaerobic species and genus diversity in routine diagnostics

ObjectivesIntroduction of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has revolutionized bacterial identification in the last decade. In 2013, MALDI-TOF MS was implemented for the identification of anaerobic bacteria at our laboratory. This study analyzed the impact of MALDI-TOF MS on the number of different anaerobic genera and species identified in diagnostics. Methods155 anaerobic human clinical isolates, representing the most frequently isolated anaerobic species at our laboratory, were identified by conventional biochemical methods and by a Bruker MALDI Biotyper (Bruker Daltonics, Bremen, Germany). Discrepancies were resolved by partial 16S rRNA gene sequence analysis. In addition, we compared the frequencies of anaerobic genera and species prior to the implementation of MALDI-TOF MS from 2008 to 2012 to the frequencies of anaerobes from 2013 to 2020 when MALDI-TOF MS was used for identification. ResultsThe diversity of anaerobic bacteria increased from 12 genera and 20 species in 2012, before the introduction of MALDI-TOF MS, to 16 genera and 31 species in 2013 and to 20 genera and 41 species in 2020 when MALDI-TOF MS was used as primary identification method. MALDI-TOF MS allowed species assignment within closely related species such as the Bacteroides fragilis group in accordance with 16S identification, and correctly identified newly described anaerobic species. ConclusionIntroduction of MALDI-TOF MS identification increased genus and species diversity of the reported anaerobes at our laboratory. Updates to the MALDI-TOF MS database and new species descriptions will further increase the diversity of anaerobic bacteria isolated from infectious processes.

Microbial Identification in the Clinical Microbiology Laboratory Using MALDI-TOF-MS.

MALDI-TOF mass spectrometry has been applied with huge success to the identification of cultured microorganisms in clinical microbiology laboratories where it has become the reference method because it is simple, fast, and highly reproducible. We describe the different procedures used in the routine for pathogen identification using the Bruker MALDI Biotyper® system.

Comparison of novel rapid diagnostic of blood culture identification and antimicrobial susceptibility testing by Accelerate Pheno system and BioFire FilmArray Blood Culture Identification and BioFire FilmArray Blood Culture Identification 2 panels

BackgroundConventional turnaround time (TAT) for positive blood culture (PBC) identification (ID) and antimicrobial susceptibility testing (AST) is 2–3 days. We evaluated the TAT and ID/AST performance using clinical and seeded samples directly from PBC bottles with different commercial approaches: (1) Accelerate Pheno® system (Pheno) for ID/AST; (2) BioFire® FilmArray® Blood Culture Identification (BCID) Panel and/ or BCID2 for ID; (3) direct AST by VITEK® 2 (direct AST); and (4) overnight culture using VITEK® 2 colony AST.ResultsA total of 141 PBC samples were included in this evaluation. Using MALDI-TOF (Bruker MALDI Biotyper) as the reference method for ID, the overall monomicrobial ID sensitivity/specificity are as follows: Pheno 97.9/99.9%; BCID 100/100%; and BCID2 100/100%, respectively. For AST performance, broth microdilution (BMD) was used as the reference method. For gram-negatives, overall categorical and essential agreements (CA/EA) for each method were: Pheno 90.3/93.2%; direct AST 92.6/88.5%; colony AST 94.4/89.5%, respectively. For gram-positives, the overall CA/EAs were as follows: Pheno 97.2/98.89%; direct AST 97.2/100%; colony AST 97.2/100%, respectively. The BCID/BCID2 and direct AST TATs were around 9–20 h (1/9-19 h for ID with resistance markers/AST), with 15 min/sample hands-on time. In comparison, Pheno TATs were around 8–10 h (1.5/7 h for ID/AST) with 2 min/sample hands-on time, maintains a clinically relevant fast report of antibiotic minimal inhibitory concentration (MIC) and allows for less TAT and hands-on time.ConclusionIn conclusion, to the best of our knowledge, this is the first study conducted as such in Asia; all studied approaches achieved satisfactory performance, factors such as TAT, panel of antibiotics choices and hands-on time should be considered for the selection of appropriate rapid ID and AST of PBC methods in different laboratory settings.

Sensitivity of vaginal and mammary bacterial microflora of sows from patients with postpartum dysgalactic syndrome to antibacterial drugs

Purpose: Investigation of the sensitivity of conditionally pathogenic and pathogenic causative agents of postpartum disgalacting syndrome isolated from the pathological discharge of the vagina and the secretion of the mammary glands of patients with sorts to the most common antibacterial drugs.Materials and methods. The fence of the biological material was carried out within 2-3 days after supporting in sows of different ages and parity with the clinical manifestation of postpartum disgalacting syndrome from June to August 2021. In sows, the purse was taken in a mosper, milk, discharge from the vagina for the purpose of bacteriological research.Samples of vaginal wasches were sent in a special transportation environment of Ames. Samples of colostrum and milk (3-5 ml) were gained in sterile test tubes in compliance with the rules of antiseptics (the mammary glands were laid with warm water and 70% ethyl alcohol were treated). For the allocation and study of pure cultures of microorganisms from the above biomaterials produced crops on various nutrient media.The material brought to the laboratory was studied as follows: from the transport medium was carried out primary sowing on triptica-soybean agar, tryptichase-soybean broth, triptichase-soy agar with the addition of 5% of the defibrous blood of the ram; Samples were incubated in aerobic conditions at 37; The growth was taken into account after 24 hours. Then, pure cultures were isolated for the study of the cultural and morphological properties of the microorganisms obtained.The primary identification of the strains of microorganisms was carried out using the Microflex® LRF Bruker Maldi Biotyper system. The accuracy of the results obtained was confirmed by classical microbiological methods based on morphological, cultural and biochemical signs of microorganisms.The resistance of the isolated and identified pure crops to antibiotics was determined by diffusion in agar. In the study, the "advanced set of disks to estimate the antibiotic sensitivity of enterobacteria was used.Results. Inflammatory processes in the reproductive path of sows with postpartum diskalactic syndrome are caused by predominant gram-negative microorganisms, in lactic glands - associations of gram-positive and gram-negative microflora. The main conditionally pathogenic and pathogenic pathogens of endometritis and / or mastitis associated with postpartum diskalactic syndrome are the microorganisms of Escherichia, Enterobacter, Klebsiella, Actinobacillus, Rothia, Weisella, Pseudomonas, Proteus, Enterococcus, Streptococcus and Staphylococcus. It has been established that the overwhelming majority of microorganisms are sensitive to cephalosporine antibiotics: cefepim, zefisim, cefotaxim, ceftazidim, ceftriaxone, ceftinibuthen, cefuroxime.Conclusion. With respect to a large number of the most frequently used antibacterial drugs on this pig-breeding enterprise, high resistance is observed in microorganisms. Treatment of postpartum diskalactic syndrome sowers using antibacterial drugs is recommended to be carried out taking into account sensitivity to them allocated conditional and pathogenic microorganisms.

MALDI-TOF MS as a method for rapid identification of Phytophthora de Bary, 1876.

The number of described species of the oomycete genus Phytophthora is growing rapidly, highlighting the need for low-cost, rapid tools for species identification. Here, a collection of 24 Phytophthora species (42 samples) from natural as well as anthropogenic habitats were genetically identified using the internal transcribed spacer (ITS) and cytochrome c oxidase subunit I (COI) regions. Because genetic identification is time consuming, we have created a complementary method based on by matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS). Both methods were compared and hypothesis that the MALDI-TOF MS method can be a fast and reliable method for the identification of oomycetes was confirmed. Over 3500 mass spectra were acquired, manually reviewed for quality control, and consolidated into a single reference library using the Bruker MALDI Biotyper platform. Finally, a database containing 144 main spectra (MSPs) was created and published in repository. The method presented in this study will facilitate the use of MALDI-TOF MS as a complement to existing approaches for fast, reliable identification of Phytophthora isolates.

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.

Analysis of Cryptosporidium spp. from clinical samples by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.

To purify Cryptosporidium spp. oocysts from clinical stool samples and evaluate using an up-to-date mass spectrometry protocol producing high-quality reference spectra. A refined purification protocol was developed for oocysts from stools, involving salt flotation and potassium bromide density centrifugation. Purified oocysts were prepared for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) by formic acid extraction, and the extracts analysed using the Bruker MALDI Biotyper system. Individual spectral markers were identified by their specific mass peaks. Cryptosporidium parvum oocysts (Iowa strain) propagated in vivo, and C. parvum (n=2) and Cryptosporidium hominis (n=1) oocysts from clinical stool samples produced distinct spectra that were considered specific to Cryptosporidium spp. with no evidence of contamination. The production of distinct spectra demonstrated the utility of the purification method for oocysts from clinical stool samples and provided reference spectra. The use of MALDI-TOF MS and other mass spectrometry techniques has been limited previously to C. parvum oocysts propagated in vivo. Appropriate purification of oocysts can achieve sufficient biomass, enabling analysis by MALDI-TOF MS and potentially other mass spectrometry platforms, facilitating peptide and protein discovery and identification of biomarkers from a much wider range of Cryptosporidium spp. from natural infections.

Detection of Active BoNT/C and D by EndoPep-MS Using MALDI Biotyper Instrument and Comparison with the Mouse Test Bioassay.

Botulinum neurotoxins (BoNTs) are among the most poisonous known biological substances, and therefore the availability of reliable, easy-to use tools for BoNT detection are important goals for food safety and human and animal health. The reference method for toxin detection and identification is the mouse bioassay (MBA). An EndoPep-MS method for BoNT differentiation has been developed based on mass spectrometry. We have validated and implemented the EndoPep-MS method on a Bruker MALDI Biotyper for the detection of BoNT/C and D serotypes. The method was extensively validated using experimentally and naturally contaminated samples comparing the results with those obtained with the MBA. Overall, the limit of detection (LoD) for both C and D toxins were less than or equal to two mouse lethal dose 50 (mLD50) per 500 µL for all tested matrices with the exception of feces spiked with BoNT/C which showed signals not-related to specific peptide fragments. Diagnostic sensitivity, specificity and positive predictive value were 100% (95% CI: 87.66–100%), 96.08% (95% CI: 86.54–99.52%), and 93.33% (95% CI: 78.25–98.20%), respectively, and accuracy was 97.47% (95% CI: 91.15–99.69%). In conclusion, the tests carried out showed that the EndoPep-MS method, initially developed using more powerful mass spectrometers, can be applied to the Bruker MALDI Biotyper instrument with excellent results including for detection of the proteolytic activity of BoNT/C, BoNT/D, BoNT/CD, and BoNT/DC toxins.