Vitek MS Research Articles

Multi-year comparison of VITEK MS performance for identification of rarely encountered pathogenic Gram-negative organisms (GNOs) in a large integrated Canadian healthcare region.

This multi-year study (2014-2019) compared identification of rare and unusual Gram-negative organisms (GNOs) by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) (VITEK MS, bioMérieux, Laval Que.) to 16S rRNA gene sequencing (16S) according to our laboratories routine workflow; 16S is done if initial MALDI-TOF MS gave discordant, wrong, or no results. GNB isolates were first analyzed by standard phenotypic methods and MALDI-TOF MS using direct deposit-full formic acid extraction; proteomics was repeated if no result occurred. Medically approved 16S analyses were done using fast protocols. Isolate sequences were analyzed using the Integrated Database Network System (IDNS3) bacterial database (SmartGene, Lausanne, Switzerland). Three hundred thirty-one GNOs including 251 (76%) aerobic Gram-negative bacilli (GNB), 63 (19%) fastidious Gram-negative coccobacilli (fGNCBs), and 17 (5%) Campylobacterales (CAMPB) isolates were recovered from 304 specimens; >1 isolate was recovered from 19 (6%). GNOs were mainly recovered from blood cultures (31.6%) and lower respiratory specimens (43%) (one-half were isolated from cystic fibrosis patients). Accurate genus vs species identities were obtained for 67.7% and 32.5% aerobic GNBs, 73% and 60% fGNCBs, and 23.5% CAMPB (with no discrepant species), respectively. Wrong or no results were obtained for 81 (32.3%) aerobic GNBs, 17 (27%) fGNCBs, and 13 (72.2%) CAMPB. No results or misidentifications occurred for 33% of aerobic GNBs, 26% of fGNCBs, and 76.5% of CAMPB due to absence of species in the instrument's database. VITEK MS performance remained stable for aerobic GNBs and fGNCBs but improved for CAMPB with addition of Campylobacter rectus and Campylobacter curvus to the database. 16S remains important for identification of GNOs when proteomics fails.IMPORTANCEMatrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has transformed the identification of commonly encountered Gram-negative organisms (GNOs) in the clinical laboratory, but rare and unusual organisms continue to challenge the technology. This study verified performance of VITEK MS for identification of a broad range of rare and unusual clinical GNO isolates by our large reference laboratory workflow over a multi-year period. Although most GNOs were accurately identified by MALDI-TOF MS, a small number of clinical isolates (~1%-6%) required 16S sequencing for identification depending on the GNO category. Approximately one-third of aerobic Gram-negative bacilli (GNBs) and two-thirds of Campylobacterales could not be accurately identified by proteomics due to lack of an organism in the instrument's database. MALDI-TOF MS databases should be continuously updated and validated, and laboratories should have a workflow for identification of unusual or rarely encountered aerobic, fastidious, and Campylobacterales GNOs that includes 16S rRNA gene sequencing whenever proteomics cannot give a definitive identification.

Distribution and antifungal susceptibility profiles of Candida species isolated from candidemia patients admitted to Egyptian tertiary hospitals: a cross-sectional study

BackgroundCandidemia is a widespread threat that can lead to significant complications in healthcare settings.ObjectivesOur study aimed to identify isolates of Candida isolated from blood culture bottles of patients with candidemia and assess their antifungal susceptibility profiles.MethodsWe conducted a cross-sectional study at Cairo University tertiary care hospitals over 16 months including 90 patients. Candida isolates were collected from blood culture bottles, and identified using MALDI-TOF MS technology of VITEK MS PRIME (bioMérieux) with the corresponding database VITEK IVD Database 3.2. followed by antifungal susceptibility testing using VITEK 2 Compact system.ResultsCandida albicans was the most common species isolated from both pediatric and adult patients with percentages of 47.3% and 36.4% respectively, followed by Candida parapsilosis with percentages of 32.6% and 25.0% respectively. Voriconazole showed the highest antifungal activity at 90.9% of isolates in adults and 95.7% in pediatrics, followed by caspofungin and micafungin. The mean hospital stays for adults ranged from 8 to 30 days and from 10 to 42 days in the pediatric group.ConclusionsC. albicans remains the predominant species isolated from both pediatric and adult candidemia patients, despite a notable increase in other species. C. tropicalis and C. parapsilosis are considered the most common non-albicans Candida (NAC) species. The rise in Candida species other than albicans highlights the urgent need for effective antifungal stewardship programs. Voriconazole exhibited the higher antifungal activity followed by caspofungin and micafungin.

A-290 Prevalence and Antimicrobial Resistance Pattern of Microorganisms Recovered From Bloodstream Infections From Public Hospitals of São Paulo City, Brazil

Abstract Background Nosocomial Bloodstream Infections (nBSI) are an important cause of morbidity and mortality. Brazilian studies concerning bloodstream infections are important tools that can bring specific issues related to antimicrobial resistance. The objective of this study was to describe the frequency and antimicrobial susceptibility profile of bacterial isolates from BSI in patients admitted to public hospitals in 2022 and 2023. Methods This study was a retrospective analysis from 12 hospitals (January 2022 to December 2023) based on BSI isolates laboratory data detected by the clinical microbiology department of a medical diagnostic laboratory. The identification and susceptibility tests were performed using MALDI-TOF mass spectrometry (Vitek MS) and Vitek®2 systems, according to the manufacture’s recommendations. Coagulase Negative Staphylococci (CoNS) BSI was only considered in the presence of at least two sets of positive blood cultures with the same organism. Results During the period (January 2022 to December 2023) a total of 19,588 microorganisms isolated from BSI were obtained from 12 hospitals. The frequency of organism group was Gram-positive (10,707; 54.7%), Gram-negative (7,874; 40.2%), yeasts (961; 4.9%) and filamentous (46; 0.2%). The order of prevalence (top ten) organisms was CoNS (5,149; 26.3%); Staphylococcus aureus (2,769; 14.1%); Klebsiella spp. (2,355; 12.0%); Escherichia coli (1,301; 6.6%); Acinetobacter spp. (1,293; 6.6%); Streptococcus spp. (1,125; 5.7%); Enterococcus spp. (1,112; 5.7%); Candida spp. (937; 4.8%); Pseudomonas spp. (749; 3.8%) and Proteus spp. (553; 2.8%). Extended spectrum beta-lactamase rates for E. coli and Klebsiella pneumoniae were 31.6% (268/848) and 74.9% (1027/1371), respectively. The resistance to carbapenems was detected mainly in 74.1% (661/892) of Acinetobacter spp. isolates, 60.7% (832/1371) of Klebsiella pneumoniae; 24.5% (114/465) of P. aeruginosa; 9.7% (34/349) of Proteus spp. and 1.4% (12/828) of Escherichia coli. Antimicrobial resistance was observed mainly in 87.2% of CoNS (2831/3247) and 44.5% of S. aureus (743/1669) to methicillin; 22.9% (146/637) of Enterococccus spp. to vancomycin. Conclusions The frequency of Klebsiella spp. isolates from nBSI has progressively increased and effective infection control measures should be taken to prevent them from spreading. The rates of antimicrobials resistance reported for Klebsiella pneumoniae, Acinetobacter spp., CoNS, S. aureus and Enterococcus spp. reinforce the high resistance rates to antimicrobials in São Paulo city.

A-282 Purpura fulminans caused by Haemophilus influenzae: case report of unusual etiology

Abstract Background Fulminant purpura is a clinical syndrome characterized by a purplish, ischemic-looking skin rash, commonly accompanied by systemic symptoms ranging from acute febrile syndrome to septic shock. Due to high lethality of possible infectious etiologies, immediate recognition and early initiation of empirical antimicrobial therapy are crucial to ensure better outcomes. Interdisciplinary collaboration among healthcare teams, infectious disease specialists and microbiologists is essential to ensure accurate and early microbiological diagnosis. Methods This work aims to report a case of Haemophilus influenzae-induced purpura fulminans that occurred in Fortaleza-CE, a coastal city in northeastern Brazil, highlighting clinical recognition and microbiological identification, based on review of medical records. Results Case report A 59-year-old man, originally from Germany, was on vacation in Brazil. He presented to emergency medical service with an acute onset of fever, abdominal pain and diarrhea, progressing to decreased level of consciousness and purplish ischemic skin lesions with hemorrhagic bullae on trunk and extremities (Figure 1), with no history of trauma. Upon admission, the blood exams showed bicytopenia - 1900 leukocytes/µL (86% segmented) and 51,000 platelets/µL - and very high C-reactive protein (41.58 mg/dL). Although empirical antimicrobial therapy with vancomycin and piperacillin-tazobactam was initiated, patient progressed to septic shock, hepatic dysfunction, requiring mechanical ventilation, vasopressor support, and renal replacement therapy. Blood cultures from hospital admission were positive after about 14 hours and allowed isolation of small and translucent colonies in Chocolate PVX plate Agar (BioMerieux™). Maldi tof analysis by Vitek MS identified H. influenzae (99.9 % accuracy). Despite prolonged hospitalization due to multiple organ dysfunctions, patient had a positive outcome. Conclusions This case illustrates the importance of immediate recognition of the syndrome coupled with assertive empirical treatment and early microbiological diagnosis for a positive clinical outcome, even in a severe presentation. Multiprofessional approach is essential for accurate and early diagnosis.

Investigations of Antibiotic Susceptibilities of S. aureus Strains Isolated from Various Clinical Samples

Objective: Staphylococcus aureus (S. aureus) is a critical microorganism that causes a range of infections with high morbidity and mortality rates, including skin and soft tissue infections, urinary tract infections, endocarditis, pneumonia, septic arthritis, osteomyelitis, and sepsis in both community and healthcare settings. The objective of this study was to ascertain the antimicrobial resistance rates of methicillin-resistant S. aureus (MRSA) and Methicillin-susceptible S. aureus (MSSA) isolates derived from a range of clinical samples submitted to the Medical Microbiology Laboratory of our hospital and to examine the resistance profile specific to our hospital. Methodology: The study included 229 S. aureus isolates collected between 2022 and 2023. The isolates were identified through the application of conventional methods and the MALDI-TOF-MS system (VITEK MS, bioMerieux France). The antimicrobial susceptibility of the isolates was determined by the BD Phoenix automated system (Becton Dickinson, USA) in accordance with the criteria set forth by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Results: The MRSA rate in the one-year period was 25.77%. When the distribution of S. aureus isolates was analyzed, it was determined that blood cultures were the most common clinical specimens from which S. aureus was isolated. Resistance to glycopeptides and linezolid was determined in MRSA isolates, albeit at a low rate. No resistance to glycopeptide and linezolid was detected in MSSA isolates. MSSA isolates were found to have a more sensitive profile to other antibiotics than MRSA isolates. The highest resistance rate was detected against penicillin with 100% and 87.06% in MRSA and MSSA isolates, respectively. In addition, the most sensitive antibiotics were determined to be glycopeptide, linezolid, daptomycin, and aminoglycoside. Conclusion: In conclusion, knowing the resistance profiles of S. aureus isolates in our hospital, will guide empirical treatment. Furthermore, the implementation of effective infection control measures and a cautious approach to antibiotic use will contribute to the management of MRSA infections.

Study of hygienic hand disinfection of staff and dental students at the Medical University – Plovdiv

<b>Introduction:</b> Transient microbial flora on the hands of medical personnel is a prerequisite for healthcare-associated infections. Unlike permanent this micro flora can be removed by washing and disinfection. The hands of the dental staff are factor number one in the transfer of microorganisms: from patient to patient as well as on objects from the dental environment.<br /> <b>The aim</b> of the present study was to microbiologically examine hand washes of 5th year dental students and medical staff before starting work with a patient in order to evaluate the quality of hygienic disinfection performed.<br /> <b>Methodology:</b> The hands of a total of 115 doctors and students from the Faculty of Dental Medicine of the Medical University of Plovdiv were wiped with a sterile swab after treatment with a disinfectant. Isolates were identified to species using Vitek MS and MALDI-TOF technology.<br />  <b>Results: </b>Coagulase-negative staphylococci with a microbial number of 10<sup>3</sup>-10<sup>4</sup> were found to have the highest microbial count (70%). 9% of the samples with coagulase-negative staphylococci had microbial count ≥10<sup>5</sup>. The most common isolates detected were: <i>Staphylococcus hominis</i>, <i>Staphylococcus epidermidis</i> and <i>Micrococcus luteus</i>.<br /> <b>Conclusion:</b> The presence of coagulase-negative staphylococci in a high microbial count, enterococci and representatives of <i>Bacillus simplex</i> is undesirable, especially in immunosuppressed patients. These results highlight the need to increase knowledge about hand disinfection and its actual application before working with a patient.

Evaluation of Antimicrobial Resistance Patterns of Pseudomonas aeruginosa Strains Isolated among COVID-19 Patients in Brazil Typed by Fourier-Transform Infrared Spectroscopy.

This study aimed to characterize Pseudomonas aeruginosa strains isolated from hospitalized patients during the COVID-19 pandemic. This was achieved using phenotypic and molecular techniques, including their antimicrobial resistance profile and biofilm formation. Eighteen strains were isolated from a hospital in Rio de Janeiro, Brazil, and identified by VITEK®2, MALDI-TOF/MS (VITEK MS® and MALDI Biotyper®), and 16S rRNA sequencing. Fourier-transform infrared (FTIR) spectroscopy, antimicrobial susceptibility testing, and biofilm formation and disinfectant tolerance tests were applied to evaluate the virulence characteristics of the strains. VITEK®2 (≥99%), VITEK MS® (≥82.7%), and MALDI Biotyper® (score ≥ 2.01) accurately identified the P. aeruginosa strains, but 16S rRNA sequencing did not differentiate the species P. aeruginosa from P. paraeruginosa. FTIR typing identified three different clusters, but no correlation between the phenotypical or antimicrobial susceptibility testing patterns was found. Most strains exhibited resistance to various antimicrobials. The exceptions were sensitivity to amikacin and norfloxacin, and consequently, these could be considered potential treatment options. Most strains (n = 15, 83.3%) produced biofilms on polystyrene. Sodium hypochlorite treatment (0.5%/15 min) was shown to be the most effective disinfectant for biofilm elimination. P. aeruginosa biofilm formation and tolerance to disinfectants demonstrate the need for effective cleaning protocols to eliminate contamination by this organism in the hospital environment and medical equipment.

Evaluation of VITEK MS Version 3.0 MALDI-TOF for the identification of anaerobes, mycobacteria, Nocardia, and moulds

PurposeThe identification of anaerobes, Mycobacterium and Nocardia species, and moulds by MALDI-TOF-MS remains a challenge. This study aimed to evaluate the performance of MALDI-TOF in the identification of these organisms. MethodsA total of 382 strains, comprising 128 (33.5 %) anaerobes, 126(33.0 %) mycobacterial, 113(29.6 %), mycelial fungi, and 15(3.9 %) Nocardia species were evaluated by VITEK MS Version 3.0. The results were compared with the identification of the isolates by DNA sequence analysis. The DNA sequences used for analysis were the 16S rRNA for anaerobic bacteria, hsp65 gene for mycobacteria, whereas both 16S rRNA and hsp65 gene for Nocardia species, and internal transcribed spacer (ITS) and 28S rRNA gene's D1/D2 regions of fungi. ResultsThe VITEK-MS accurately identified 78.3 % (299/382) of the strains at the species, and 9.4 % (36/382) at the genus level. Misidentifications were observed in 3.9 % (15/382) isolates. Of isolates tested, 8.4 % (32/382) were not identified by the system, and 7.06 % (27/382) were not included in the IVD database. ConclusionAn upgraded VITEK MS V3.0 database provides reasonably accurate and rapid identification of clinically relevant anaerobes, mycobacteria, Nocardia species, and moulds to the species level.

EVALUATION OF CANDIDA SPP. ADHERENCE TO 3D-PRINTED AND HEAT-CURED PMMA RESIN DENTURES

Introduction: Despite the progress of additive manufacturing, there are different opinions regarding microbial adhesion on the surface of 3D printed dentures and the biocompatiability of the materials. The purpose of this in-vivo study was to determine the presence of Candida species in the saliva, oral mucosa and on dentures of patients wearing heat-cured polymethylmethacrylate (PMMA) and 3D-printed prosthetic appliances after a three-month period of observation. Materials and methods: The clinical study included 40 edentulous patients, divided into two groups - first group (A) –wearing conventional dentures and second group (B) – wearing 3D printed dentures. It was used chrom agar Candida (Bio Merieux) for identification and quantity assessment of the presence of Candida spp. and spectrophotometric analysis using VITEK MS (Bio Merieux). Results: After three months of wearing the dentures, Candida spp was not isolated in the saliva in 60% of patients in group A and 70% of group B, with no statistically significant difference between the results before the treatment and after the third month. In the third month, Candida spp was isolated from the oral mucosa of 20% of group A and in group B – 10%, with no significant difference between the groups (p<0.05). No Candida spp was isolated from 90% of denture surfaces in both groups. Conclusion: The adhesion of Candida spp. to 3D printed dentures is not higher than conventional dentures made from heat-curing resin. Also, there is no difference between the amount of Candida spp. in the saliva, oral mucosa and denture surfaces.

Formic acid sandwich method is well-suited for filamentous fungi identification and improves turn around time using Zybio EXS2600 mass spectrometry

ObjectivesMatrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is extensively employed for the identification of filamentous fungi on MALDI Biotyper (Bruker Daltonics) and Vitek MS (biomerieux), but the performance of fungi identification on new EXS2600 (Zybio) is still unknow. Our study aims to evaluate the new EXS2600 system's (Zybio) ability to rapidly identify filamentous fungi and determine its effect on turnaround time (TAT) in our laboratory.MethodsWe tested 117 filamentous fungi using two pretreatment methods: the formic acid sandwich (FA-sandwich) and a commercial mold extraction kit (MEK, Zybio). All isolates were confirmed via sequence analysis. Laboratory data were extracted from our laboratory information system over two 9-month periods: pre-EXS (April to December 2022) and post-EXS (April to December 2023), respectively.ResultsThe total correct identification (at the species, genus, or complex/group level) rate of fungi was high, FA-sandwich (95.73%, 112/117), followed by MEK (94.02%, 110/117). Excluding 6 isolates not in the database, species-level identification accuracy was 92.79% (103/111) for FA-sandwich and 91.89% (102/111) for MEK; genus-level accuracy was 97.29% (108/111) and 96.39% (107/111), respectively. Both methods attained a 100% correct identification rate for Aspergillus, Lichtheimia, Rhizopus Mucor and Talaromyces species, and were able to differentiate between Fusarium verticillioides and Fusarium proliferatum within the Fusarium fujikuroi species complex. Notably, high confidence was observed in the species-level identification of uncommon fungi such as Trichothecium roseum and Geotrichum candidum. The TAT for all positive cultures decreased from pre EXS2600 to post (108.379 VS 102.438, P < 0.05), and the TAT for tissue decreased most (451.538 VS 222.304, P < 0.001).ConclusionsThe FA-sandwich method is more efficient and accurate for identifying filamentous fungi with EXS2600 than the MEK. Our study firstly evaluated the performance of fungi identification on EXS2600 and showed it is suitable for clinical microbiology laboratories use.

Candiduria: Analysis of Changing Trends and Antifungal Dynamics in Acute Healthcare Settings with Special Reference to Rare Candida Species

Background: Candiduria is a common phenomenon among hospitalized patients, particularly those in intensive care units (ICUs), wherein multiple predisposing factors such as comorbidities, indwelling urinary catheters, and antimicrobial exposure contribute to its prevalence. Objective: This study aims to investigate the epidemiological trends and antifungal susceptibility patterns of Candida spp. isolated from urine samples of patients at a tertiary care center in Western Maharashtra, India. Methods: Of the 7,642 urine samples received between 01 June 2022 to 31 May 2023, 142 samples (9%) displayed the growth of Candida species. Upon retesting samples from these patients, only 70 true positive results were identified. All the isolates were detected via VITEK MS (bioMerieux, France), while antifungal susceptibility testing (AFST) was conducted using the VITEK 2 Compact with an AST-YS08 card. Results: The most common isolate was Candida tropicalis (50%), followed by Candida albicans (25.7%). Rare Candida species (e.g., Candida ciferrii, Candida kefyr, Candida metapsilosis, Candida utilis) collectively contributed to 7.1% of all Candida species. The results of the AFST revealed a high sensitivity to amphotericin B and echinocandins. Fluconazole resistance was observed in 6% of the isolates. Candida ciferrii emonstrated a distinct pattern, indicating a statistically significant association (p = 0.015) when the patient had two or more comorbidities and resistance to azoles, which was statistically significant (p-values for fluconazole (p = 0.015), voriconazole (p = 0.004), and flucytosine (p = 0.002)) when compared with other Candida species. Conclusion: Resistance trends in India have been changing with increased fluconazole-resistant Candida strains being frequently isolated, emphasizing the importance of customized therapeutic strategies based on culture and sensitivity reports.

In vitro antimicrobial activity of silver nanoparticles against selected Gram-negative and Gram-positive pathogens.

Infections caused by pathogenic bacteria increase patient morbidity and mortality and significantly raise treatment costs. The use of silver nanoparticles as an alternative treatment for S aureus, E coli, MRSA, E faecalis, K pneumoniae and P aeruginosa indicates their antibacterial effect and prompts medical research to consider the next generation of antibacterial drugs that could change antibiotic therapy. By combining silver nanoparticles with different classes of antibiotics, the antibacterial effect is evidenced by increased values of the inhibition zone compared to the values obtained for some antibiotics commonly used in the treatment of bacterial infections. This study focuses on comparing the antibacterial activity of antibiotics versus antibiotics combined with silver nanoparticles against various bacteria, by comparing inhibition zones obtained for both. We aim to prove that the size of the inhibition zone for antibiotics combined with silver nanoparticles is greater, thus confirming the improved antibacterial effect. In this study we tested the antibacterial activity of solutions of silver nanoparticles alone or in combination with different antibiotics. We used standard bacterial strains, ATCC, both Gram positive bacteria Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212, as well as Gram negative bacteria Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, but also on clinical isolates: a strain MRSA (Methicillin Resistant Staphylococcus aureus) and a PDR strain (pan drug resistant) of Klebsiella pneumoniae. Bacterial identification was performed using Vitek MS analyzer (bioMerieux). Antibiotic susceptibility determination was performed with VITEK2 COMPACT SYSTEM (bio Merieux, Inc Durham NC) with ready to use VITEK AST cards. The interpretation of the results was done in compliance with EUCAST 2023-2024 standards. Testing was performed for several classes of antibiotics, silver nanoparticle solutions in 2 concentrations (10 μg/mL and 100 μg/mL) and for combinations of antibiotics with silver nanoparticle solutions. The diameter of the inhibition zone (ZOI) for silver nanoparticles, antibiotics and silver nanoparticles combined with antibiotic against each bacterium was expressed in millimeters. The Kirby-Bauer disk-diffusion method, in accordance with current EUCAST standards, was used to analyze the antibacterial effect of antibiotics, silver nanoparticles, and antibiotics combined with silver nanoparticles at biocompatible doses of 10 and 100 μg/mL. The experiments were conducted in triplicate, and the results were almost identical. The results of this study show that the silver nanoparticles displayed antibacterial activity, proven by the appearance of the inhibition zone, in various sizes, for all bacteria studied. The antibiotic classes tested were beta-lactamins, first, second, third and fourth generation cephalosporins, macrolides, fluoroquinolones, lincosamides, aminoglycosides, glycopeptides, tetracyclines, oxazolidinones, sulfonamides, rifamycins, amphenicols. Testing S aureus ATCC 29213, the highest zone of inhibition was demonstrated for cephalosporins (32.6667 ± 0.701 mm), macrolides (31.6667 ± 0.701 mm, and lincosamides (29.6667 ± 0.701 mm). Testing MRSA (internal code GR0333), the highest zone of inhibition for combination of silver nanoparticles and antibiotics was demonstrated for fluoroquinolones (36.3333 ± 0.701 mm), lincosamides (32.3333 ± 0.701 mm), Fusid acid (32.3333 ± 0.701 mm) and aminoglicosides (31.3333 ± 0.701 mm). Testing E coli ATCC 25922 the highest zone of inhibition was for Fosfomycine, 39 mm and for E faecalis ATCC 29212 for aminoglicosides was 19 mm. For K pneumoniae (internal code GQ8575) the inhibition zone for silver nanoparticles 100 μg/mL was 12.3333 ± 0.701 mm and for P aeruginosa ATCC 27253 was 16 ± 1.214 mm. The use of metallic nanoparticles, especially silver ones, as antimicrobial agents with definite bactericidal activity has led medical specialists to consider this new treatment which may change antibacterial therapy. Studies of in vitro combinations between silver nanoparticles and different classes of antibiotics represent a highly efficient and effective new antibacterial treatment against multidrug-resistant bacteria. To avoid the problem of antimicrobial resistance associated with conventional antibiotics, it is necessary to understand the adaptive mechanisms of bacteria under the action of metal nanoparticles, which could be exploited in future studies. Further in vitro and in vivo studies that would assess specify the biocompatibility and toxicity of silver nanoparticles will make these super nanomaterials the medicines of the future.

Clinical Evaluation of VITEK MS PRIME with PICKME Pen for Bacteria and Yeasts, and RUO Database for Filamentous Fungi.

The VITEK MS PRIME (bioMérieux, Marcy-l'Étoile, France), a newly developed matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) system, alongside the VITEK PICKME pen (PICKME), offers easy sample preparation for bacteria and yeasts. The VITEK MS PRIME also offers two software platforms for filamentous fungi: the IVD database and the RUO database. Our study evaluated its identification agreement on 320 clinical isolates of bacteria and yeasts, comparing PICKME and traditional wooden toothpick sampling techniques against MicroIDSys Elite (ASTA) results. Additionally, we assessed the IVD (v3.2) and SARAMIS (v4.16) RUO databases on 289 filamentous fungi against molecular sequencing. The concordance rates for species-level identification of bacteria and yeasts were about 89.4% (286/320) between the PICKME and wooden toothpick, and about 83.4-85.3% between the VITEK MS PRIME and ASTA MicroIDSys Elite. Retesting with PICKME improved concordance to 91.9%. For filamentous fungi, species-level identification reached 71.3% with the IVD database and 85.8% with RUO, which significantly enhanced basidiomycetes' identification from 35.3% to 100%. Some strains in the IVD database, like Aspergillus versicolor, Exophiala xenobiotica, and Nannizzia gypsea, failed to be identified. The VITEK MS PRIME with PICKME offers reliable and efficient microorganism identification. For filamentous fungi, combined use of the RUO database can be beneficial, especially for basidiomycetes.

Identifying organisms directly from BacT/Alert FAN plus blood culture bottles using Vitek MS in a state-wide laboratory network

Identifying organisms directly from positive blood culture bottles using matrix-assisted laser desorption-ionisation time-of-flight mass spectrometry (MALDI-TOF MS) has many advantages to patients, clinical services, and laboratories. However, few published methods have demonstrated good performance using the current BioMérieux culture bottles and MALDI-TOF system: BacT/Alert FAN plus and Vitek MS. The effect of transporting bottles on test performance has not been assessed for any direct-from-bottle MS method. In this study, 802 positive blood culture bottles were analysed including 234 requiring inter-laboratory transport, using a method involving protein extraction with formic acid and acetonitrile. Correct identification rates were high for Staphylococcus aureus (58/58 of new diagnostic samples), Enterococcus faecalis (27/27), Gram-negative bacilli (160/176, 90.1%), and coagulase-negative Staphylococcus species (108/132, 81.8%). Three false identifications were made, none with clinical significance. For Gram-positive cocci in pairs or chains, more correct identifications were made from bottles analysed immediately compared to transported bottles (67% vs 44%, p=0.016), and longer transport time was associated with slightly lower probability of correct identification (OR 0.984 per additional hour, p=0.040). Transportation was not associated with a difference for other organism types. This technique is a vastly more cost-effective alternative to molecular techniques for rapid identification of bacteraemia isolates, and performance is minimally affected by inter-laboratory transport of bottles at ambient temperature.

Pathogenic characteristics of an aggregated diarrhea event caused by Plesiomonas shigelloides from stream.

This study aimed to investigate the cause of a foodborne disease outbreak in Huzhou on August 14, 2023. Multiple enteropathogens were detected using FilmArray, and the pathogen was subsequently isolated and cultured from anal swabs of the cases and stream water. The isolated strains were identified using VITEK MS, and antimicrobial susceptibility test, pulsed field gel electrophoresis (PFGE) molecular typing, and whole genome sequencing (WGS) were performed on the isolates of Plesiomonas shigelloides. Gene annotation and sequence alignment were used to analyze the virulence genes and drug resistance genes of the strains. A phylogenetic tree was constructed based on single nucleotide polymorphism (SNP), and homology analysis was conducted to trace the origin of P. shigelloides. A total of 7 strains of P.shigelloides were isolated, with 3 from stream water and 4 from anal swabs. All 7 strains exhibited the same PFGE pattern and showed resistance to amikacin, trimethoprim-sulfamethoxazole, chloramphenicol, tetracycline, cefazolin, streptomycin, and florfenicol. The isolated strains carried the same resistance genes and virulence factors. In the sequences of the isolated strains from this outbreak, 11 mutation sites were detected. The phylogenetic tree based on SNP sites showed that these strains were homologous. This foodborne disease outbreak caused by P.shigelloides was the first reported in Huzhou. WGS can be used as a complementary method to PFGE for epidemiological investigations of disease outbreaks.

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.

Bacteria-antagonists of the agents of soryz bacterial diseases

Aim. To isolate and identify bacteria with antagonist properties for biocontrol of the agents of bacterial diseases of soryz (Sorghum oryzoidum) and sorghum crops. Methods. The studies were conducted in 2021-2023. Spore-forming bacteria were isolated from the soryz samples, collected in the fields of the experimental farm of the Uman National Horticulture University (Cherkasy region, Uman). Lactic acid bacteria were isolated from soryz plants, collected in the private land plot, located between the villages of Teolyn, Vladyslavchyk, Kniazhyky in Monastyryshche com- munity, Uman district, where Pershotravneve hamlet used to be situated. A total of 1,250 samples were analyzed. The experiment had three repeats. Spore-forming and lactic acid bacteria were isolated from the surface of soryz plants on the firm ripe stage in summer while isolating phytopathogenic bacteria. The isolates of lactic acid bacteria- antagonists were also isolated from the inner part of winter stubble stalk of soryz, collected from the tilled field. The antagonistic activity of the strains of lactic acid bacteria and spore-forming bacteria, isolated from different ecological niches, to phytopathogens of soryz and sorghum crops was determined in vitro. The strains of Pseudomonas syringae, the agents of soryz bacterial spots, were used as test-cultures: 211141a, 211141, 210341, 21034, and 210521, along with the collection strains of phytopathogens: Pseudomonas syringae 8299, Pseudomonas syringae subsp. syringae UKM B-1021, X. oryzae 8375, Dickeya chrysanthemi 8683, Diskeya chryzanthemi 8683. The antagonistic activity of the extracted isolates of spore-forming and lactic acid bacteria was studied using the method of radial strokes (joint cultivation of the antagonist and the strains under investigation). The bacterial isolates were deemed inactive if the growth delay zone was 0–5 mm (–), from 5 to 10 mm (+) – low activity, 11–20 mm (++) – moderate activity, over 20 mm (+++) – high activity regarding the test-cultures. To check the effect of the isolate-antagonist of phytopathogenic bacteria, artificial infecting was conducted in the field conditions. For this purpose, a diurnal culture of the antagonist was introduced into the stalk of plants in the concentration of 1×108 colony-forming units, and 24 h later, a culture of test-strain of the phytopathogen was administered above the previous puncture. The results were evaluated 7–14 days after the artificial infection. The experiment had three repeats. The isolates of bacteria which demonstrated their an- tagonistic properties regarding the phytopathogenic bacteria were identified by their morphological properties, Gram staining, catalase test, profile of carbohydrate fermentation and mass-spectrometry (MALDI-TOF – Matrix Assisted Laser Desorption/Ionization) using VITEK MS mass-spectrometer. Results. Thirty-eight spore-forming bacterial iso- lates were extracted from soryz; among these, 21030, 21095, 21040, ASV1, ASV3, B4 demonstrated their antagonistic activity towards the investigated phytopathogenic bacteria. Isolate 21040 showed high antagonistic activity to most test-strains of P. syringae from soryz (the zone of negative culture – 23–30 mm) and lower activity regarding the collection cultures. Isolates B4 and AVS3 demonstrated their selective activity regarding the investigated phytopatho- gens. Twenty isolates of lactic acid bacteria were extracted. Higher antagonistic activity was noted for the isolates of lactic acid bacteria 8/1 and F1 to the strains of P. syringae, isolated from soryz and collection cultures. The highest antagonistic activity of isolate 8/1 was noted regarding test-strains of P. syringae 210521 and X. oryzae 8375 (the zone of negative culture – 40–35 mm). In the field conditions, the treatment of sorghum plants with F1 affected the pathological process that developed due to the impact of the phytopathogenic bacteria P. syringae, which led to the reduction in disease symptoms. The taxonomic position of the isolates of bacteria, which seem to be promising for the control of disease agents, was determined. In terms of morphology of cells and colonies, the biochemical profile, and mass-spectrometry MALDI-TOF, the spore-forming isolates 21040 and B4 were identified as Bacillus subtilis, and ASV3 – as Bacillus vallismortis. The identified isolates of lactic acid bacteria were Lactobacillus pentosus F1 and Lactobacillus sakei 8/1. Conclusions. In addition to phytopathogenic bacteria, from soryz plants we isolated the strains of spore-forming bacteria Bacillus subtilis 21040, B4, Bacillus vallismortis AVS3 and such lactic acid bacteria as Lactiplantibacillus pentosus and Lactobacillus sakei 8/1 (Latilactobacillus sakei 8/1), promising for the elaboration of methods for the biocontrol of the agents of bacterial diseases.

The Usage of an AMCA-Modified Aptamer Facilitates the Rapid Detection of Vibrio vulnificus

Background: Vibrio vulnificus can cause serious infections in human beings associated with the consumption of raw oysters or cuts exposed to seawater. The traditional method for culturing V. vulnificus is time-consuming and has a high failure rate. Objectives: This study aims to detect V. vulnificus using an AMCA-modified specific DNA aptamer. Methods: Common pathogenic microorganisms present in the seawater of the Fujian Sea area were collected, cultured, and identified. The samples were found to contain mainly V. vulnificus, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, V. alginolyticus, V. parahaemolyticus, and V. cholerae. AMCA was conjugated with 5’ ends of the aptamer using N-hydroxysuccinimide ester (NHS ester) to target V. vulnificus and produce a fluorescent signal upon binding. The aptamer was screened and optimized for rapid detection of V. vulnificus. We collected the 50 bacterial strains isolated from clinical secretion samples and used a fluorescence microscope to determine whether the sample contained V. vulnificus or not. We compared these results with those obtained from VITEK MS (considered the gold standard) to test the sensitivity and specificity of the AMCA-modified aptamer using IBM SPSS Statistics 22. Results: In this experiment, the sensitivity and specificity of the modified aptamer for detecting V. vulnificus were determined to be 100% [95% CI (0.39, 1)] and 93.4% [95% CI (0.81, 0.98)], respectively. The positive predictive value was 57% [95% CI (0.20, 0.88)], and the negative predictive value was 100% [95% CI (0.89, 1)]. These findings indicate that V. vulnificus specimens can be rapidly detected via fluorescence reaction within 30 minutes. Conclusions: Our results suggest that this modified DNA aptamer has the potential to be used for diagnosing V. vulnificus. Further research is needed to explore the application of aptamers in pathogen infections.

Cholangitis with bacteremia due to Pseudomonas nitroreducens in a patient with pancreatic neuroendocrine tumors: a case report

BackgroundPseudomonas nitroreducens is a non-fermenting, gram-negative, rod-shaped bacterium commonly inhabiting soil, particularly soil contaminated with oil brine. To our knowledge, no cases of human infection with P. nitroreducens have been previously reported. Here, we present the first documented case of cholangitis caused by P. nitroreducens in a patient with bacteremia.Case presentationA 46-year-old Japanese man with an advanced pancreatic neuroendocrine tumor was hospitalized with fever and chills. Four days before admission, the patient developed right upper abdominal pain. Two days later, he also experienced fever and chills. Endoscopic retrograde cholangiopancreatography was performed on the day of admission, and the patient was diagnosed as having cholangitis associated with stent dysfunction. Gram-negative rods were isolated from blood cultures, but attempts to identify the bacteria using VITEK2 and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) with VITEK MS ver. 4.7.1 (bioMérieux Japan Co. Ltd., Tokyo, Japan) were unsuccessful. Finally, the organism was identified as P. nitroreducens using MALDI-TOF MS with a MALDI Biotyper (Bruker Daltonics Co., Ltd., Billerica, MA, USA) and 16 S ribosomal RNA sequencing. Despite thorough interviews with the patient, he denied any exposure to contaminated soil. The patient was treated with intravenous cefepime and oral ciprofloxacin for 16 days based on susceptibility results, achieving a good therapeutic outcome. At the outpatient follow-up on day 28, the patient was in good general condition.ConclusionsThis is the first reported human case of cholangitis with bloodstream infection caused by P. nitroreducens. This report provides clinicians with novel insights into the clinical manifestations and diagnostic methods necessary for the accurate diagnosis of P. nitroreducens, along with guidance on treatment.

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.