Clinical Microbiology Laboratory Research Articles

Detection of Chlamydia trachomatis and Neisseria gonorrhoeae (and Its Resistance to Ciprofloxacin): Validation of a Molecular Biology Tool for Rapid Diagnosis and Treatment

Background and Objectives: Neisseria gonorrhoeae and Chlamydia trachomatis can cause similar clinical syndromes and may coexist in infections. In emergency medicine, empirical treatment targeting both pathogens is often employed, potentially contributing to antibiotic resistance. Gonococcal resistance has emerged against first-line antimicrobials, necessitating prior testing for fluoroquinolone susceptibility. Certest Biotec developed two molecular diagnostic products for simultaneous detection: VIASURE C. trachomatis & N. gonorrhoeae Real-Time PCR Detection Kit and VIASURE Neisseria gonorrhoeae Ciprofloxacin-Resistant Real-Time PCR Detection Kit. To evaluate these products, clinical performance assessments were conducted at the Clinical Microbiology Laboratory of Miguel Servet University Hospital in Zaragoza, Spain. Results and Conclusions: Both VIASURE assays under study demonstrated high clinical sensitivity and specificity compared to reference molecular assays and Sanger sequencing. These kits offer an accurate diagnosis, facilitating appropriate treatment choices while addressing concerns about emerging antibiotic resistance. Methods: A total of 540 clinical samples from 540 patients already characterized as positive or negative for CT and NG by a molecular assay and by antibiotic susceptibility testing for ciprofloxacin using a concentration gradient diffusion method were used for the clinical evaluation. In cases where sensitivity results were unavailable, conventional PCR and Sanger sequencing were employed.

Lock, stock, and Leuconostoc: an unusual presentation of a rare pathogen

Leuconostoc mesenteroides is a Gram-positive bacterium in the Lactobacillaceae family. This species is rarely encountered clinically and in the past was regarded as a contaminant when cultured. Recently it is being recognized as a pathogen responsible for opportunistic infections in immunocompromised patients. Compared to other Gram-positive cocci, Leuconostoc spp. carry intrinsic resistance to vancomycin. It is not common for clinical microbiology laboratories to isolate this organism and instead incorrectly report enterococcus and streptococcus organisms. Here, we present a case of bacteremia resulting in febrile illness and pneumonia in an 81-year-old woman with a history of rheumatoid arthritis and a notable lapse between immunomodulator therapy exposure. She was successfully treated when this organism was identified, and her antibiotic therapy was switched from vancomycin to ampicillin-sulbactam. This case highlights the importance of considering L. mesenteroides as a source of infection in patients predisposed to an immunocompromised state despite this organism’s being a rare pathogen. Keywords: Leuconostoc mesenteroides, rheumatoid arthritis, immunomodulators; vancomycin resistance

Identifying Candida auris transmission in a hospital outbreak investigation using whole-genome sequencing and SNP phylogenetic analysis.

Candida auris poses a global public health challenge, causing multiple outbreaks within healthcare facilities. Despite advancements in strain typing for various infectious diseases, a consensus on the genetic relatedness threshold for identifying C. auris transmission in local hospital outbreaks remains elusive. We investigated genetic variations within our local isolate collection using whole-genome-based single nucleotide polymorphism (SNP) phylogenetic analysis. A total of 74 C. auris isolates were subjected to whole-genome sequencing (WGS) and SNP phylogenetic analysis via the QIAGEN CLC Genomics Workbench. Isolates included known related strains from the same patient, strains from different hospitals, strains from our hospital patients with no epidemiological link, and 19 patient isolates from a recent C. auris outbreak. All but three isolates were identified to be Clade IV. By examining the genetic diversities of C. auris within patients and between patients, we identified a SNP variation range of 0-13 for identifying related isolates. During an outbreak investigation, utilizing this range, maximum likelihood phylogenetic analysis revealed two distinct clusters that aligned with the epidemiological links. Determining a SNP variation range to delineate genetic relatedness among isolates is crucial for the application of WGS and SNP phylogenetic analysis in identifying C. auris transmission during hospital outbreak investigations. The use of WGS SNP phylogenetic analysis via the CLC Genomics Workbench has emerged as a valuable method for typing C. auris in clinical microbiology laboratories.

Analyzing Antibiotic Resistance: A Comparative Study of E. coli Isolates from Wastewater and Clinical UTI Samples at a University Community in Grenada, West Indies

Abstract Introduction/Objective The rise of antibiotic resistance poses a significant global health threat, needing a robust monitoring system to track its emergence and spread. However, most developing countries lack the infrastructure to monitor such resistance effectively. Wastewater-based epidemiology (WBE) has been proven effective in surveilling various outbreaks and offers a promising method for monitoring antibiotic resistance at the community level. This study aims to assess and compare the antibiotic resistance of Escherichia coli isolated from raw wastewater with those isolated from clinical urinary tract infection (UTI) samples from the same community. Methods/Case Report Conducted from January 2022 to October 2023, this study analyzed antibiogram data from eighty-six Escherichia coli isolates sourced from wastewater and thirty-four isolates derived from clinical urinary tract infections (UTIs). The clinical samples were obtained and processed at the Clinical Microbiology Laboratory of a university campus. The isolates were evaluated against the eleven most prescribed antibiotics for UTIs in the University Clinic including: Amoxicillin/Clavulanic Acid, Ampicillin, Cefazolin, Cefoxitin, Ceftriaxone, Cefuroxime, Ciprofloxacin, Gentamicin, Nalidixic Acid, Nitrofurantoin, and Sulfamethoxazole-trimethoprim. Results (if a Case Study enter NA) Explaining the observed trends in both wastewater and clinical samples from the perspective of AWaRE-WHO classification (Access, Watch, Reserve), we see similar trends of resistance in both data sets. A) The antibiotics used in the Access category (Empiric first or second choice for treatment of most common infections) demonstrates similar resistance patterns for both wastewater and clinical isolates except for Nitrofurantoin. B) The antibiotics used in the Watch category (Antibiotics that have higher toxicity issues and higher potential to negatively impact AMR) resistance pattern shows negligible risk in the university community evaluated. Conclusion It is recommended that the university clinic prescribe Nitrofurantoin as the last antibiotic from the Access list to the university community to treat UTI patients. Furthermore, WBE is a useful tool to determine patterns of resistance under the studied settings.

Case Series: Emerging Pathogens Aerococcus urinae and Aerococcus sanguinicola at a Canadian tertiary care hospital

Abstract Introduction/Objective Background: Aerococcus urinae and sanguinicola are emerging opportunistic pathogen linked with urinary tract infections. Unmanaged Aerococcus infections have the propensity to precipitated into urosepsis. The ongoing gestalt of managing these infections needs standardization. We reviewed A. urinae and sanguinicola isolates from our clinical microbiology laboratory to better characterize its spectrum of clinical presentation, sources of infection, microbiological characteristics, and antimicrobial management. Methods/Case Report Methods: The electronic microbiology database was utilized to review the retrospective clinical data of patients that grew positive cultures for Aerococcus urinae and sanguinicola on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) in Saskatchewan from January to July 2023. Demographic variables, site of presentation, microbiological parameters surrounding culture and sensitivities, and antimicrobial management were recorded. The results were analyzed on SPSS 28 software. Results (if a Case Study enter NA) Results: This cohort (n=115) had a median age 82 years [Range: 7 to 103] with a female predilection (F:M = 78:37). A. urinae and sanguinicola were primarily involved in UTI (n=96), obstructive uropathy (n=9), and urosepsis (n=6). The hypothyroidism (26.9%, n = 31) rate in this population was 3-fold higher than the age-matched background rate for our province. Aerococcus infections were predominantly monomicrobial (73.9%, n= 85 out of 115) and showed sensitivity for ceftriaxone, penicillin G, and vancomycin. Antimicrobials were seldom used in the UTI cohort (31.2%, n=30 out of 96). Amoxicillin-clavulanate (29.8%, n=25 out 84) was the most popular choice at patient discharge in monomicrobial Aerococcus infections only. Conclusion Conclusion: Aerococcus urinae and sanguinicola are emerging uropathogen. Despite this, the clinical significance of these Aerococcus species remains undervalued. Our case series shows that untreated or recurrent A. urinae and A. sanguinicola infections can precipitate into urosepsis. Clinicians should utilize the antimicrobial susceptibility profiles for these Aerococcus species to provide appropriate antimicrobial coverage and manage these pathogens effectively. Further studies should explore the link between hypothyroidism and these aerococcal infections.

Evaluation of protein extraction protocols for MALDI-TOF Biotyper analysis of mycobacteria

Infections caused by Mycobacterium tuberculosis and nontuberculous mycobacteria represent a significant global threat and medical concern. Therefore, accurate and reliable methods must be employed to identify mycobacteria rapidly. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a technique that compares the cellular protein profiles of unknown isolates with reference mass spectra in a database to identify microorganisms. However, the thick and waxy lipid layer, which is rich in mycolic acids and is present in mycobacterial cells, makes protein extraction challenging. To identify the optimal protocol for correctly identifying bacilli using MALDI-TOF mass spectrometry, this study compared four different cellular protein extraction methods. Four strains of M. bovis BCG were selected as representatives of slow-growing mycobacteria, while three strains of fast-growing mycobacteria were also included: M. peregrinum, M. smegmatis, and M. farcinogenes. The extraction method that proved most effective was the extraction of inactivated cells with chloroform and methanol, which partially delipidates the cells. These cells were then extracted with formic acid, as is standard practice for protein extraction. The advantage of this method is that it allows the parallel analysis of cellular lipids and proteins from a single sample. It is therefore important to optimize mycobacterial protein extraction for MALDI-TOF MS analysis in clinical microbiology laboratories.

Evaluating the Efficacy of a Polymerase Chain Reaction Assay for Pneumocystis carinii Detection in Respiratory Samples

Abstract Introduction: Accurately identifying patients with pneumocystis pneumonia (PCP) presents considerable challenges, primarily relying on clinical suspicion. The manifestations of PCP lack specificity, encompassing symptoms such as cough, dyspnoea, fever, hypoxaemia, malaise and weight loss. Therefore, this study assesses the efficacy of a polymerase chain reaction (PCR) assay intended for use in clinical microbiology laboratories for the identification of Pneumocystis carinii from respiratory specimens. Materials and Methods: In this retrospective study, the medical records of patients aged 18 years and above were examined, focusing on those assessed using pneumocystis PCR-based tests. The study spanned from 2019 to 2022 in our tertiary care centre at Max Hospital, Saket. In this study, sputum and bronchoalveolar lavage (BAL) samples were proceeded for both staining and PCR for multi-copy mtLSU gene and single-copy dihydropteroate synthase polymorphisms (DHPS) fas gene of P. jirovecii. Results: In the analysis of 112 BAL specimens, 17 were identified as positive for P. carinii through both Grocott-Gomori’s Methenamine Silver Staining (GMS) staining and PCR. In addition, two BAL specimens were negative through GMS staining but showed positive results with PCR. On reviewing the clinical details of the patients associated with these two specimens, it was discovered that both individuals were under systemic steroid treatment, yet neither exhibited clinical signs of pneumocystis pneumonia (PCP). Consequently, these instances were classified as false positives by PCR. In comparison to GMS, the PCR assay demonstrated a sensitivity of 100% and specificity of 98% for detecting P. carinii in BAL specimens. Conclusions: The study underscores the necessity for individualised approaches, with each laboratory assessing the positive and negative predictive values based on their specific prevalence rates. In the ongoing pursuit of improving PCP diagnosis, PCR emerges as a promising screening tool that when used judiciously in conjunction with confirmatory measures, can enhance diagnostic precision in immunosuppressed populations.

Implementation of a Clinical Decision Support System for Antimicrobial Prescribing in Sub-Saharan Africa: Multisectoral Qualitative Study.

Suboptimal use of antimicrobials is a driver of antimicrobial resistance in West Africa. Clinical decision support systems (CDSSs) can facilitate access to updated and reliable recommendations. This study aimed to assess contextual factors that could facilitate the implementation of a CDSS for antimicrobial prescribing in West Africa and Central Africa and to identify tailored implementation strategies. This qualitative study was conducted through 21 semistructured individual interviews via videoconference with health care professionals between September and December 2020. Participants were recruited using purposive sampling in a transnational capacity-building network for hospital preparedness in West Africa. The interview guide included multiple constructs derived from the Consolidated Framework for Implementation Research. Interviews were transcribed, and data were analyzed using thematic analysis. The panel of participants included health practitioners (12/21, 57%), health actors trained in engineering (2/21, 10%), project managers (3/21, 14%), antimicrobial resistance research experts (2/21, 10%), a clinical microbiologist (1/21, 5%), and an anthropologist (1/21, 5%). Contextual factors influencing the implementation of eHealth tools existed at the individual, health care system, and national levels. At the individual level, the main challenge was to design a user-centered CDSS adapted to the prescriber's clinical routine and structural constraints. Most of the participants stated that the CDSS should not only target physicians in academic hospitals who can use their network to disseminate the tool but also general practitioners, primary care nurses, midwives, and other health care workers who are the main prescribers of antimicrobials in rural areas of West Africa. The heterogeneity in antimicrobial prescribing training among prescribers was a significant challenge to the use of a common CDSS. At the country level, weak pharmaceutical regulations, the lack of official guidelines for antimicrobial prescribing, limited access to clinical microbiology laboratories, self-medication, and disparity in health care coverage lead to inappropriate antimicrobial use and could limit the implementation and diffusion of CDSS for antimicrobial prescribing. Participants emphasized the importance of building a solid eHealth ecosystem in their countries by establishing academic partnerships, developing physician networks, and involving diverse stakeholders to address challenges. Additional implementation strategies included conducting a local needs assessment, identifying early adopters, promoting network weaving, using implementation advisers, and creating a learning collaborative. Participants noted that a CDSS for antimicrobial prescribing could be a powerful tool for the development and dissemination of official guidelines for infectious diseases in West Africa. These results suggest that a CDSS for antimicrobial prescribing adapted for nonspecialized prescribers could have a role in improving clinical decisions. They also confirm the relevance of adopting a cross-disciplinary approach with participants from different backgrounds to assess contextual factors, including social, political, and economic determinants.

Can flow cytometry be an alternative method for the detection of antimicrobial resistance?

Background: The inappropriate use of antibiotics may cause antibiotic resistance, cause side effects, and eventually cause an increase in healthcare costs. This study aimed to determine a flow cytometry-based test to detect ESBL E. coli and MDR p. aeruginosa in a short time. Methods: The study included 25 E.coli isolates and 25 P.aeruginosa isolates identified by Phoenix TM 100 automated system [Becton Dickinson, USA]. In the flow cytometric method, the percentages of death cells exposed to cephalosporin including ceftazidime [CAZ] or cefotaxime [CTX] and clavulanic acid [CLA] combination, were compared with death cells exposed only to a cephalosporin [CAZ or CTX]. CLA index values [CAZ-CLA and CTX-CLA indices] were obtained for CTX and CAZ. Index values that were higher than 1.5 just for one cephalosporin were accepted as positive. Results: Cell viability was performed after 1-hour exposure to the drugs. When PI staining was applied to ESBL-treated and MDR-treated bacteria, 65% and 85% had nonviable membranes, indicating the method efficiently identified only cells with damaged membranes. Conclusion: Flow cytometry is a rapid and reliable method for the detection of ESBL and MDR in clinical microbiology laboratories Keywords: Antibiotic resistance; ESBL; flow cytometry; MDR.

A-326 Testing for Multidrug-Resistant Acinetobacter baumannii with a Microfluidic Device

Abstract Background Multidrug-resistant (MDR) Acinetobacter baumannii in hospitals contribute to high mortality rates and entail expensive infection control procedures. Conventionally, species and resistance determination of MDR bacteria is performed by mass spectrometry and labor-intensive phenotypical assays in clinical microbiology laboratories. Some institutions also apply time-consuming and expensive whole genome sequencing for in-depth resistance and transmission analysis. In contrast, faster and more cost-efficient MDR typing can be achieved by on-site application of real-time PCR-based point-of-care tests (POCTs) that target species- and resistance-specific genes and enable rapid screening of large sample sizes. Methods To construct a real-time PCR-based POCT system for species and resistance determination of MDR A. baumannii, we selected two species-specific genes (rpoB, OXA-51-like) and 18 resistance genes found in MDR A. baumannii (OXA-23-like, OXA-24/40-like, OXA-51-like, OXA-58-like, OXA-143-like, mcr, ADC, NDM, GIM, VIM, IMP, GES, PER, TEM, SHV, VEB, CTX-M, KPC) as targets. For each gene, one to six sets of primers and fluorophore-labeled probes that amplify conserved genetic regions were designed and their functionality was tested in vitro. Parallel, a fully automated system comprising microfluidic chips and a bench-top analyzer able to perform DNA extraction and real-time PCR starting from swab eluate was developed. Results All oligonucleotides tested so far reliably detected 100 to 100,000 copies of DNA extracted from MDR bacteria in single- and multiplex experiments in vitro. Tests with the microfluidic system showed that lysis of different bacterial species could be accomplished via sonication on a membrane located on the microfluidic chip. In addition, chip implementation of all PCR reagents like lyophilized master mix as well as primers and probes was successful and can be stored on chip at least one year. So far, the system delivered detectable fluorescence signals within one hour when performing test runs with bacteria eluted from swabs. Conclusions A real-time PCR-based Point-of-Care screening platform was developed for the detection of antibiotic-resistant A. baumannii from various patient sample materials starting directly from swabs, which detects colonization/infection and at the same time the relevant resistance genes within about an hour.

B-205 Performance of a Novel Fluorogenic Assay for Detection of Carbapenemase-producing Enterbacteriaceae

Abstract Background In recent years, under the selective challenge of antibiotics, the variety and number of drugresistant pathogenic microorganisms have increased significantly, which brings great challenges to clinical diagnosis and treatment, especially the infection caused by carbapenem resistant Enterobacteriaceae (CRE). Carbapenemases production is the main mechanism of drug resistance of Enterobacteriaceae to carbapenems. Drug resistance of Carbapenems can be caused by three mechanisms, resulting in the production of five major Carbapenemases. These are Klebsiella pneumoniae enzyme (KPC), New Delhi metal β-lactamase (NDM), carbapenem hydrolyzed oxalase (OXA-48 like), integrin-encoded metal β- lactamase (VIM) and IMP (Imipenemase). The real-time fluorescence quantitative PCR (qPCR) method based on molecular beacons was combined with the melting curve analysis to identify five drug resistance genes simultaneously by a single PCR reaction, with rapid detection, high sensitivity and strong specificity. Based on this principle, we developed the novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae (Dynamiker Biotechnology (Tianjin) Co., Ltd.). Methods We evaluated the performance of the novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae, including the limit of detection (LoD) and cross-reactivity, and compared it with the lateral flow immunochromatography assay (LFA). Results The LoD ranged from 75-450 CFU/mL for the five carbapenemase genes. The analytical specificity for target genes was 100%, as assessed with a panel of 15 pathogens, which indicated no cross-reactions. Comparison of qPCR and LFA results from twenty-three CRE clinical isolates with characterized carbapenemase content demonstrated a complete agreement (Table 1). Conclusions The novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae is an accurate and rapid method to identify KPC, NDM, VIM, IMP and OXA-48-like carbapenemases in the clinical microbiology laboratory, which can guide infection control programs to limit the spread of these organisms.

B-225 Bridging the Gap Between Clinical Microbiology and Molecular Diagnostics: Correlation of PCR Cycle Threshold Values for Establishing Semi-Quantitative CFU/mL Ranges of Bacterial Targets

Abstract Background Molecular diagnostics (MolDx) has surpassed culture in the clinical microbiology laboratory. Wong (2023) states PCR is more sensitive than urine culture in detecting polymicrobial infections (para. 8), marking an advantage in MolDX for urinary tract infections. This advancement poses a challenge for clinicians in treatment decision-making. To address requests for numerical MolDx result ranges, a validation accessed semi-quantitative Colony Forming Units per milliliter (CFU/mL) through the assessment of PCR cycle threshold (CT) values. Methods Bacterial targets of UTI and STI PCR panels were inoculated in nutrient broth (Hardy diagnostics cat#K43) and incubated overnight in a shaking incubator at 35±2°C. The density of the cultures was measured using the OD600 function of a NanoDrop One/One after 18-24 hour incubation. The density of stock cultures was adjusted to 106 CFU/mL. Serial dilutions of stock were made in 1:10 steps covering the range of 106 to 100 CFU/mL. The stock and dilutions were extracted, and PCR was performed in a minimum of 2 replicates of each sample for the UTI and STI PCR panels. Mean CT values, standard deviations and coefficients of variation were calculated for each sample. The linearity of CT values within each organism’s stock and dilution samples were assessed by evaluating the CT differences between each dilution step. Corresponding CFU/mL concentrations were tabulated against each mean CT value. Results The average CT values for reportable CFU/mL ranges were established for each stock and dilution. Mean CT values for each CFU/mL breakdown across all organisms were also calculated, along with standard deviations, to investigate universal CT-CFU/mL correlations. A final interpretation table (Table 1) was constructed according to the universal range calculations. Conclusions The study confirmed correlation between CT values and CFU/mL ranges. Further data analysis and comparison with other laboratories’ results will enhance accuracy and precision of reportable CFU/mL ranges.

Emergence and spread of resistant and biofilm-forming Acinetobacter baumannii in critically ill COVID-19 patients

The COVID-19 pandemic has raised concerns beyond the viral infection itself. Bacterial co-infections, particularly those involving Acinetobacter baumannii, have become a significant worry in critically ill COVID-19 patients. A. baumannii is an opportunistic pathogen that can cause nosocomial infections, especially in patients with compromised immune systems. This study investigated 36 A. baumannii isolates obtained from COVID-19 patients during a concurrent outbreak. The isolates were collected over two years through routine medical requests sent to the Clinical Microbiology laboratory. Identification of the strains was confirmed through biochemical tests, the Phoenix BD® Automated Microbiology System, and MALDI-TOF Mass Spectrometry. The study assessed the antimicrobial sensitivity of the isolates, with a specific focus on resistance to the beta-lactam group as well as aminoglycosides. The presence of specific antibiotic resistance genes (blaOXA-23, -24, -51 and -58, blaKPC, blaSHV, blaIMP, blaVIM, aac(6')-Ib, ant(3'')-Ia, and aph(3')-Ia) was investigated using PCR and Sanger DNA sequencing. Biofilm-forming capabilities of the isolates were also evaluated. The findings revealed diverse resistance profiles, with a high prevalence of resistant strains, including resistance to carbapenems. Genetic analysis suggested potential clonal spread of certain strains within the hospital setting. Moreover, a significant proportion of the isolates demonstrated strong biofilm-forming abilities, which can enhance persistence and antibiotic resistance. In conclusion, this study highlights the need for vigilant monitoring and targeted interventions to address bacterial co-infections in COVID-19 patients. The diversity in resistance patterns, potential clonal spread, and robust biofilm-forming abilities among A. baumannii isolates underscore the importance of addressing this issue to better manage and treat critically ill COVID-19 patients.

Identification methods as a factor affecting the performance of clinical microbiology laboratories participating in an external quality assessment program: a cross-sectional, retrospective analysis.

Introduction. Laboratory participation in external quality assessment (EQA) programmes including proficiency testing (PT) is a requirement of clinical laboratory conformance to ISO 15189:2022 Medical laboratories - Requirements for quality and competence. PT is one EQA method whereby laboratories are sent blinded samples for characterization by routine laboratory diagnostic methods. Importantly, PT enables a laboratory's performance to be evaluated in comparison to the standard reference methods and to the performance of other peer laboratories using similar diagnostic methods.Gap statement. The desired outcome of participating in PT is to help laboratories identify possible sources of error in each step of the total testing process and particularly in their test methods during the analytical phase.Aim. This cross-sectional study investigated the impact of using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) compared to conventional phenotypic biochemical testing on laboratory performance in a clinical bacteriology PT scheme.Methodology. During a 6-year period from 2017-2022, the Canadian Microbiology Proficiency Testing implemented 112 PT challenges comprising 22 different sample types and included 61 different bacterial species. This was translated into 5883 graded test events for analysis. Multiple logistic regression techniques were employed to explore the association between the test method employed and laboratory performance. The sample type and aerobic classification of challenge organisms were included as confounding variables.Results. Laboratories using MALDI-TOF MS performed significantly better in characterizing microorganisms than laboratories using phenotypic biochemical testing alone [odds ratio OR = 5.68, confidence interval (CI): 3.92, 8.22] regardless of the sample type and aerobic classification. Notably, our analysis identified a significant association between anaerobic organisms and laboratory performance (OR: 0.24, CI: 0.17-0.35), suggesting that culturing and identifying fastidious organisms remains a significant obstacle for many clinical microbiology laboratories.Conclusions. Although no method is infallible and its performance will depend on the validation and quality assurance procedures, this finding may help the management in the decision for implementing MALDI-TOF MS in the microbiology laboratory. This study highlights the important role PT providers play in the objective assessment of laboratory performance and how it can provide evidence for quality improvement.

Uropathogenic Bacterial Detection and Surveillance of Antimicrobial Resistance Pattern in Urine Specimens Referred to Al anbar Province

For clinicians treating patients, antimicrobial resistance is becoming a significant concern. This brief study was therefore conducted to find out how common it is for bacterial isolates that are resistant to multiple drugs (MDR), XDR, and PDR, as well as extensively drug-resistant (XDR), in Anbar hospital. The microbiology department cultivated the clinical samples and identified the bacterial strains. The detection of MDR, XDR, and PDR microorganisms was accomplished by analyzing the antibiotic susceptibility profile of various bacterial isolates. 257 children and adult patients from both sexes were included in this study. The total number of urine samples that gave positive bacterial culture results were 114 (44.4%) and the number of samples that gave negative results was 143(55.6%) . The antibiotic susceptibility profile of 114 bacterial strains was studied. 90(78.9) of gram-negative bacteria strains were MDR, (8.9%) (72.2%) XDR, and (18.9%) PDR was isolated. 24 (21.1%) Gram positive bacterial egative bacteria strains were MDR, (33.3%) (54.2%) XDR, and (12.5%) PDR was isolated. To effectively combat the threat of antibiotic resistance, all clinical microbiology laboratories need to closely monitor MDR, XDR, and PDR.

Uropathogenic Bacterial Detection and Surveillance of Antimicrobial Resistance Pattern in Urine Specimens Referred to Al anbar Province

For clinicians treating patients, antimicrobial resistance is becoming a significant concern. This brief study was therefore conducted to find out how common it is for bacterial isolates that are resistant to multiple drugs (MDR), XDR, and PDR, as well as extensively drug-resistant (XDR), in Anbar hospital. The microbiology department cultivated the clinical samples and identified the bacterial strains. The detection of MDR, XDR, and PDR microorganisms was accomplished by analyzing the antibiotic susceptibility profile of various bacterial isolates. 257 children and adult patients from both sexes were included in this study. The total number of urine samples that gave positive bacterial culture results were 114 (44.4%) and the number of samples that gave negative results was 143(55.6%) . The antibiotic susceptibility profile of 114 bacterial strains was studied. 90(78.9) of gram negative bacteria strains were MDR, (8.9%) ,(72.2%) XDR, and (18.9%)PDR was isolated. 24 (21.1%)Gram positive bacterial egative bacteria strains were MDR, (33.3%) ,(54.2%) XDR, and (12.5%) PDR was isolated. To effectively combat the threat of antibiotic resistance, all clinical microbiology laboratories need to closely monitor MDR, XDR, and PDR.

An in vitro experimental study on the interference of glyphosate on the urease enzyme

Purpose: Exposure to glyphosate is increasing due to the density of agricultural areas in Türkiye. In this study, the possible interference effect of glyphosate on urease, an enzyme that is frequently used in the diagnosis and follow-up of many diseases and in the measurement of urea in biological samples was examined. Materials and Methods: First, glyphosate was observed to have a negative interference in experiments using solutions of varying concentrations of urea. Second, blood samples were examined using the urease-glutamate dehydrogenase (GLDH) and indirect nesslerization procedures to determine the effects of glyphosate on the results before and after its addition. To determine the morphological and chemical alterations, scanning electron microscope (SEM) and Fourier-transform infrared spectroscopy (FTIR) analyses were conducted, and binding patterns were established through molecular docking. Urea measurements conducted with urease-GLDH and indirect nesslerization demonstrated a negative interference on the results with glyphosate concentrations of 10–3, 10–4, and 10–5 M. Results: Morphological changes observed in the SEM analysis were supported by the 3228.25 (O-H), 1642.08 (C=C), and 1531.20 (N-O) cm–1 bonds formed in the FTIR analysis. Furthermore, the molecular docking analysis showed that glyphosate affected the urease via hydrogen bonding (Gly13, Ser12, Lys14, Thr15, and Asp37) and hydrophobic interactions (Val10, Asp37, and Glu98). It was hypothesized that these interacting amino acids limit the accessibility of the urease’s active catalytic conformation and/or impact the stability of the catalytic transition state. Conclusion: Glyphosate leads to negative interference in human serum urea assays, leading to incorrect test results in clinical biochemistry, microbiology, and agricultural laboratories. This effect should be considered when conducting analysis, and clinicians as well as hospital information management systems should be informed ahead of time, with special emphasis devoted to this interference.

Simulation to optimize the laboratory diagnosis of bacteremia.

Optimization of laboratory pathways and resource availability has a direct impact on the clinical management of patients with bloodstream infection. This research offers an insight into the laboratory processing of blood cultures at a system level and allows clinical microbiology laboratories to explore the impact of changes to processes and resources.

Impact of ESKAPE Pathogens on Bacteremia: A Three-Year Surveillance Study at a Major Hospital in Southern Italy.

ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.) pose a serious public health threat as they are resistant to multiple antimicrobial agents. Bloodstream infections (BSIs) caused by ESKAPE bacteria have high mortality rates due to the limited availability of effective antimicrobials. This study aimed to evaluate the prevalence and susceptibility of ESKAPE pathogens causing BSIs over three years in a large tertiary hospital in Salerno. Conducted at the Clinical Microbiology Laboratory of San Giovanni di Dio e ''Ruggi D'Aragona'' Hospital from January 2020 to December 2022, blood culture samples from different departments were incubated in the BD BACTEC™ system for 5 days. Species identification was performed using MALDI-TOF MS, and antimicrobial resistance patterns were determined by the VITEK2 system. Out of 3197 species isolated from positive blood cultures, 38.7% were ESKAPE bacteria. Of these, 59.9% were found in blood culture samples taken from men, and the most affected age group was those aged >60 years. (70.6%). Staphylococcus aureus was the main BSI pathogen (26.3%), followed by Klebsiella pneumoniae (15.8%). Significant resistance rates were found, including 35% of Staphylococcus aureus being resistant to oxacillin and over 90% of Acinetobacter baumannii being resistant to carbapenems. These results highlight the urgent need for antimicrobial stewardship programs to prevent incurable infections.

Barriers to implementation of rapid identification and antimicrobial susceptibility testing technologies in the clinical microbiology laboratory: an American perspective.

Clinical microbiology laboratories are responsible for confirming the aetiology of infectious diseases and providing antimicrobial susceptibility testing results. Traditional culture-based testing can be augmented by more rapid testing modalities to provide clinically actionable information as quickly as possible. Despite improvements in patient outcomes, many clinical microbiology laboratories are facing challenges to in-sourcing these technologies. Depending on a multitude of factors, including size, location and patient population served, these barriers may affect some laboratories and hospital systems to greater or lesser extents than others. It will be up to each individual facility to ascertain its ability to overcome barriers. To aid in this self-assessment, we present for thoughtful consideration a discussion of the barriers to implementation of rapid identification and antimicrobial susceptibility testing technologies, with specific attention to matters of financial cost, staff expertise, operational issues and stakeholder buy-in.