Conventional Biochemical Tests Research Articles

Severe co-infection caused by difficult-to-diagnose hypermucoviscous Klebsiella pneumoniae K1-ST82 in a patient with COVID-19: a case report

BackgroundCo-infection with Klebsiella pneumoniae presents a significant concern in hospitalized patients with coronavirus disease (COVID-19), increasing the risk of severe disease progression. Hypervirulent (hv) and hypermucoviscous (hm) K. pneumoniae (Kp) has gained prominence in Asia due to its capacity to cause invasive community-acquired infections. However, recognition of hvKp/hmKp co-infections in the context of COVID-19 remains limited. We report a severe case of rapidly progressing co-infection with hmKp exhibiting “difficult-to-diagnose” phenotypes in a hospitalized patient with COVID-19.Case presentationA 61-year-old woman with COVID-19 initially exhibited mild symptoms resembling the common cold. However, her condition rapidly deteriorated over 7 days, leading to hospital admission with the development of dyspnea. The patient required supplemental oxygen, antibiotic treatment, and mechanical ventilation. Gram-negative bacteria with atypical phenotypes were isolated from alveolar lavage fluid and blood cultures. Both strains formed small, glossy, non-lactose-fermenting colonies on clinically relevant media and were susceptible to ampicillin. Conventional biochemical tests failed to identify the Enterobacteriales strains owing to the urease-negative phenotype. Consequently, the identification of K. pneumoniae was difficult until matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis was performed. A positive string test indicated mucoviscosity, but with variability in the material used for stretching colonies. Whole-genome sequencing performed on the MiSeq and GridION platforms revealed the blood-derived strain JARB-RN-0063 as belonging to serotype K1 and sequence type (ST) 82. The hvKp-associated genes rmpA and iroCD were located on a 5.0-Mb chromosome, and iucABCD-iutA was identified on a 217.9-kb IncFIB(K)/IncR-type plasmid. Therefore, JARB-RN-0063 was genetically classified as hvKp with a Kleborate virulence score of 3. The intrinsic penicillinase gene blaSHV was defective owing to an IS1F element insertion, resulting in the strain being atypically susceptible to ampicillin.ConclusionsThis is the first case of severe COVID-19-associated co-infection with a difficult-to-diagnose K. penummoniae strain. Notably, co-infection by the hmKp K1-ST82 clone exhibited atypical phenotypes, including stunted growth, non-lactose fermentation, urease-negative reaction, ampicillin susceptibility, and abnormal mucoviscosity, posing diagnostic challenges for clinical laboratories and impedes the identification of hvKp/hmKp. Delayed identification may worsen patient outcomes, highlighting the need for increased clinical awareness of such difficult-to-diagnose clones to prevent deterioration.

The Burden of Acinetobacter baumannii Among Pet Dogs and Cats with Respiratory Illness Outside the Healthcare Facilities: A Possible Public Health Concern.

Background: Researchers paid more attention to nosocomial Acinetobacter baumannii in veterinary hospitals worldwide; however, the research scope toward community-acquired A. baumannii infections among animals is largely ignored. Therefore, the current study aimed to investigate the role of diseased dogs and cats suffering from respiratory illness in transmission of community-acquired A. baumannii infection and its public health threat. Materials and methods: Oral swabs were collected from 154 pet animals with respiratory signs, including 80 cats and 74 dogs (outpatient visits). The obtained swabs were cultured on CHROMagar™ MH Orientation media for isolation of A. baumannii, and identification of suspected isolates was conducted via Gram staining, conventional biochemical tests, and molecular detection of the blaOXA-51-like gene. Antimicrobial susceptibility testing of A. baumannii isolates was carried out using the disc diffusion method. Results: Overall, 10 (6.5%) out of 154 diseased pet animals were positive for A. baumannii, where 6 (8.1%) and 4 (5%) dogs and cats were positive, respectively. Multidrug-resistant (MDR) A. baumannii was found in 3.9% of the examined animals. The phylogenetic tree analysis revealed that the obtained sequences from dogs and cats were closely related to human and animal sequences. Conclusion: The occurrence of MDR A. baumannii among dogs and cats suffering from respiratory illness highlights the potential role of pet animals in the dissemination of MDR A. baumannii in the community.

Whole genome analysis of Pantoea species identified from sepsis patients in selected Ethiopian referral hospitals: emerging pathogens

BackgroundThe burden of sepsis worsens due to the continuation of emerging pathogens such as multidrug-resistant Pantoea species.MethodsA multicenter study was conducted between October 2019 and September 2020 at four hospitals located in central, southern, and northern parts of Ethiopia. A total of 1416 sepsis patients were recruited and blood cultures were performed. At each study site, positive cultures were characterized by their colony characteristics, gram stain, and conventional biochemical tests. All Pantoea species were identified using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI TOF) and subjected to whole genome sequencing (WGS) using Illumina HiSeq 2500. The phylogeny structure of Pantoea isolates was calculated using IQ-TREE v1.6.12 from single-nucleotide polymorphisms detected by Snippy v.4.6.0 and filtered by Gubbins v.2.3.4. Average nucleotide identity was estimated by using OrthoANI v.0.93.1 on Shovill v.1.1.0 assemblies. Antimicrobial resistance genes and plasmid replicons were detected using ARIBA v.2.14.6. Phylogenetic trees were visualized using iTOLv.6.5.2.ResultsMultiple Pantoea species include: P. dispersa (n = 19), P. septica (n = 1), and a novel Pantoea spp. (n = 1), were identified among sepsis patients. All P. dispersa isolates and the novel Pantoea species were isolated at Dessie Referral Hospital and displayed phylogenetic clonality, including the ubiquity of an IncM1 plasmid and identical antimicrobial resistance (AMR) gene profiles, encoding blaCTX−M−15, blaTEM−1D, blaSCO−1, and aac(3)-lla. The novel Pantoea spp. isolate harboured blaCTX−M−9 and blaTEM−1D and carried an IncN3 plasmid replicon. The P. septica was isolated at Tikur Anbessa Specialized Hospital in Addis Ababa and carried no detectable acquired AMR genes.ConclusionThe emerging Pantoea spp. carrying multiple AMR genes were identified from sepsis patients. Implementation of strong infection prevention strategies and building surveillance capacity with advanced bacteriology laboratories capable of identifying multidrug-resistant emerging pathogens is strongly recommended.

Listeria grayi Meningitis in a Polypathological Patient: A Case Report

Background: The various species of the Listeria genus are frequently isolated from food and drink products. They are classified into two categories according to their pathogenicity. Pathogenic species are frequently incriminated in infections of the elderly, immunocompromised and pregnant women. The Listeria grayi species is rarely isolated as a pathogen in human pathologies. We report a case of L. grayi meningitis in a polypathological patient. Antimicrobial susceptibility testing revealed resistance to ampicillin and susceptibility to Meropenem. Therapeutic failure was recorded despite in vitro sensitivity Methods: Conventional biochemical test and Api Listeria gallery was used for identification of the isolate. Results: We report a case of L. grayi meningitis in a polypathological patient. Antimicrobial susceptibility testing revealed resistance to ampicillin and susceptibility to Meropenem. Therapeutic failure was recorded despite in vitro sensitivity. Conclusion: Given its infrequent reporting, Listeria grayi meningitis warrants consideration, especially in elderly patients with multiple pathologies.

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.

Emergence of Virulent Extensively Drug-Resistant Vancomycin-Resistant Enterococci Among Diarrheic Pet Animals: A Possible Public Health Threat on the Move.

Background: Vancomycin-resistant enterococci (VRE) have become an increasing public health concern in the past few decades, being associated with serious multidrug-resistant (MDR) infections. This study was conducted to investigate the role of diarrheic pet animals as potential reservoirs for virulent extensively drug-resistant (XDR) VRE and their threat on human health. Materials and Methods: Rectal swabs were collected from 153 diarrheic pet animals (80 dogs and 73 cats). The collected swabs were cultured on CHROMagarTMVRE for the isolation of vancomycin-resistant Enterococcus faecalis and Enterococcus faecium, and then suspected colonies were identified as enterococci after Gram staining, conventional biochemical tests, and molecular techniques. VRE were basically identified using the disk diffusion method; however, molecular identification of vanA and vanB genes was carried out among confirmed VRE isolates. Moreover, three virulence genes (cytolysin A, cylA; enterococcal surface protein, esp; and hyaluronidase, hyl) were investigated in VRE isolates. Thereafter, VRE strains that harbored virulence genes were tested for antimicrobial susceptibility. Results: Eighteen out of 153 animals (11.8%) were positive for VRE, which were obtained from 15% and 8.2% of the examined dogs and cats, respectively. None of the obtained isolates carried the vanA gene, whereas the vanB gene was detected in E. faecalis (4/10) with a prevalence rate (40%). Of the obtained VRE isolates, five possessed esp and/or cylA, while all strains were negative for the hyl gene. Furthermore, four virulent VRE isolates exhibited an XDR pattern, and one isolate was MDR. Conclusion: Diarrheic pet animals could represent a potential zoonotic reservoir for virulent XDR vancomycin-resistant E. faecalis, which may have serious public health implications.

Tigecycline Resistant Pattern among Carbapenem Resistant Gram-negative Bacilli: Prospective Cross-sectional Study

Introduction: Tigecycline is a unique tetracycline class of semi-synthetic, last-line broad spectrum antibiotic against multi-drug-resistant bacteria. However, recently, resistance to this antibiotic is on the rise. Aims: This study was conducted to determine the prevalence of tigecycline resistance amongst carbapenem-resistant Gram-negative bacilli (GNB) isolated from clinical samples (pus and sputum) as well as to evaluate their antimicrobial susceptibility pattern. Study design: Prospective cross sectional Place and Duration of Study: Department of Microbiology at Sanjay Gandhi Post Graduate Institute of Medical Sciences Lucknow, between January 2023 and December 2023. Methodology: Identification of GNB grown on culture was done by conventional biochemical tests and later validated by MALDI-TOF MS. The antimicrobial sensitivity testing of isolates was done using the E-test, and disk diffusion method. Minimum inhibitory concentration determination was done by Broth micro-dilution (BMD) method. Results: Amongst 8326 pus and respiratory samples, GNBs were recovered from 63.15% (5258/8326). Of 5258 GNB isolates, 50.74% (2668) were carbapenem-resistant, while 7.85% (413) demonstrated resistance to both tigecycline and carbapenem. Common isolates in this group were Klebsiella pneumoniae (37.04%), Acinetobacter spp. (25.18%), Enterobacter spp. (14.28%) and Escherichia coli (12.59%). BMD results demonstrated highest activity of tigecycline against carbapenem-resistant E. coli, followed by Citrobacter and Enterobacter. It works against resistant strains of Acinetobacter baumannii and K. pneumoniae as well, but in higher concentrations. Conclusion: High tigecycline resistance (one of the last-resort drugs) among carbapenem resistant GNB isolates is a matter of clinical concern, leaving physicians with limited options for treatment of such infections. Proper adherence to the policies of antimicrobial stewardship programs can reduce the emergence of resistance.

Molecular characterisation of Histoplasma capsulatum sensu lato from Ethiopian horses reveals two distinct phylogenetic clades.

Epizootic lymphangitis (EL) is a highly prevalent and contagious infectious disease affecting horses in many parts of Ethiopia caused by Histoplasma capsulatum sensu lato ('var. farciminosum'). In this study, 12 suspected isolates of H. capsulatum sensu lato or yeasts unidentified by conventional biochemical tests isolated from Ethiopian horses with EL were characterised by internal transcribed spacer sequencing. Six of the 12 isolates were identified to be members of H. capsulatum sensu lato and the other six were Pichia kudriavzevii (synonym: Candida krusei) (n=3), Trichosporon asahii (n=1), Geotrichum silvicola (n=1) and Moesziomyces aphidis (n=1), respectively. The six H. capsulatum sensu lato isolates were further characterised by multilocus sequence analysis. Four distinct gene loci (arf [462 bases], H-anti [410 bases], ole1 [338 bases] and tub1 [272 bases]) of these six isolates as well as those of two H. capsulatum sensu lato ('var. farciminosum') reference strains (ATCC 58332 and ATCC 28798) were polymerase chain reaction (PCR)-amplified and sequenced. Phylogenetic analyses of their concatenated nucleotide sequences showed that three of the isolates and the reference strain ATCC 58332 were identical and belonged to the Eurasia clade within Latin American (LAm) A (H. suramericanum), and those of the other three isolates and the reference strain ATCC 28798 were identical and belonged to the Africa clade. At least two distinct phylogenetic clades of H. capsulatum sensu lato were circulating in Ethiopian horses with EL. Advanced molecular technologies and bioinformatics tools are crucial for the accurate identification and typing of pathogens as well as the discovery of novel microorganisms in veterinary microbiology.

Epidemiology and antimicrobial resistance of toxin-producing Klebsiella oxytoca clinical isolates from children admitted to the oncology chemotherapy center in Mofid Children's Hospital in Tehran, Iran: A cross-sectional study.

Klebsiella oxytoca (K. oxytoca) is the second bacterial cause of nosocomial infections in the general population after K. pneumoniae. This study surveyed the frequency of cytotoxin-producing strains of K. oxytoca and their antibiotic susceptibility profile in a cohort of children admitted to a referral hospital with different malignancies. The Stool samples of children admitted to the Cancer Chemotherapy Unit of the Mofid Children's Hospital, Tehran, Iran were analyzed using conventional biochemical tests and polymerase chain reaction targeting the pehX gene to identify K. oxytoca. The antibiotic susceptibility profile of isolated K. oxytoca against commonly prescribed antibiotics used in treating infection at the facility was determined using the Kirby-Bauer disk diffusion technique. Also, the prevalence of genes encoding toxins among K. oxytoca was identified by PCR assay. The Stool samples of 280 participants were taken for the study of which 38 samples [(55.3% (21/38) 42 males and 44.7% (17/38) females)] tested positive for various Klebsiella spp. Out of this, K. oxytoca was identified in 2.5% (7/280) stools using cultures and conventional biochemical tests. Also, the stools of 2.9% (8/280) of the participants tested positive for K. oxytoca using PCR assay. Using PCR, (2/7) of the K. oxytoca isolates tested positive for the npsA and npsB genes and were identified as toxigenic K. oxytoca strains. The prevalence of toxin-producing K. oxytoca strains in stool samples of children diagnosed with cancer in Iran is relatively low. Most of the K. oxytoca isolates were susceptible to tested antibiotics. Globally, active surveillance of toxigenic K. oxytoca strains in patients with different malignancies or immunocompromised patients is recommended in healthcare settings.

Extended-spectrum beta-lactamase and carbapenemase producing Klebsiella Pneumoniae isolated from patient with suspected urinary tract infection

Urinary tract infection (UTI) is the most common bacterial infections in humans and serious health problem in many parts of the world. Carbapenem resistant Enterobacteriaceae (CRE) are increasingly reported from healthcare facilities in Nepal and around the world. This study is carried out with an objective to assess the Prevalence of ESBL and carbapenemase producing Klebsiella pneumoniae isolate from patient with suspected of UTI. Total 880 urine sample was collected during 6-month period and processed and identified by using conventional microbiological procedure and biochemical test. ESBL screening isolates were done by using Ceftazidime and Ceftazidime with clavulanic acid. Production of ESBL was determine by combination Disc Test. For the confirmation of carbapenemase producing K. pneumoniae Modified Hodge Method, Carbapenem Inactivation Method and EDTA Impregnated Disc test (MBL) was performed. Out of total sample, 9.30% (82) showed significant growth. Prevalence of UTI seen more in male as compare to female patients. Out of positive isolates, 37.8% were K. pneumoniae. K. pneumoniae were resistance with Ceftriaxone (54.8%) followed by 51.61% Ceftazidime, Cotrimoxazole. Multi drug resistance (MDR) was found to be 54.83%. Out of 31 isolates, 10(32%) were confirmed as ESBL positive by Combination Disc Assay. 14 isolates were screened as carbapenemase producers but 2(14.28%) showed positive through Modified Hodge Method, 4(28.57%) Modified Carbapenem Inactivation Method and 7(50%) EDTA impregnated Disc Test (MBL). Analytically, the sensitivity of MHT, CIM, and MBL tests was 42.81%, 58.10%, and 76.96%, respectively, all with 100% specificity. Colistin was the drug of choice, with 93.54% of isolates showing sensitivity to it. Early detection of ESBL and MBL-producing isolates is crucial for reducing mortality rates and preventing the intra-hospital spread of these strains.

Improved Blue Carba test and Carba NP test for detection and classification of major Class A and B carbapenemases, including dual producers, among Gram-negative bacilli.

Rapid and accurate identification of carbapenemase-producing organisms is of vital importance in guiding appropriate clinical antibiotic treatments and curbing their transmission. The emergence of negative bacilli carrying multiple carbapenemase combinations during and after the severe acute respiratory syndrome coronavirus 2 pandemic has posed a challenge to the conventional biochemical tests typically used to determine the specific carbapenemase type in the isolated strains. Several initiatives have aimed to enhance colorimetric methods, enabling them to independently identify the presence of Class A or Class B carbapenemases. Notably, no previous efforts have been made to distinguish both classes simultaneously. Additionally, these modifications have struggled to differentiate between carriers of multiple carbapenemases, a common occurrence in many Latin American countries. In this study, we introduced specific Class A and Class B carbapenemase inhibitors into the Blue Carba test (BCT) and Carba NP-direct (CNP) colorimetric assays to identify the type of carbapenemase, even in cases of multiple carbapenemase producers within these classes. These updated assays demonstrated exceptional sensitivity and specificity (≥ 90%) all within a rapid turnaround time of under 2 hours, typically completed in just 45 minutes. These in-house enhancements to the BCT and CNP assays present a rapid, straightforward, and cost-effective approach to determining the primary carbapenemase classes. They could serve as a viable alternative to molecular biology or immuno-chromatography techniques, acting as an initial diagnostic step in the process.

Association of Nosocomial Burns and Bacterial Colonization in Iraqi Patients

Burn injury is a prevalent and severe form of trauma, posing a significant global public health concern. This study included isolating causative agents involved in nosocomial burns in Iraqi patients. 65 swabs from burn wounds were collected aseptically from a specialty hospital in Baghdad city. The swabs were cultured on routinely used culture media prepared in the laboratory and diagnosed using conventional biochemical tests, and the VITEK-2 compact system validated their findings. The findings revealed that 47 (72.3%) isolates had growth, and the remaining 27.7% of the swabs did not grow. Pseudomonas aeruginosa was the predominant species bacteria isolated in percent (43%), followed by Staphylococcus aureus in percent (26%), Corynebacterium spp. (11%), Proteus vulgaris (6%), whereas "Escherichia coli, Enterococcus faecalis, and Serratia marcescens" isolated in similar percent (4%), and Klebsiella spp. in the lowest percent (2%). Moreover, antibiotic susceptibility testing was also done using VITEK-2 compact system against Amoxiclav, Ticarcillin, Ceftriaxone, Imipenem, Gentamicin, Amikacin, Tetracycline, Doxycycline, Ciprofloxacin, and Erythromycin.

A STUDY ON IN VITRO ACTIVITY OF FOSFOMYCIN AGAINST ENTEROBACTERIACAE UROPATHOGEN ALONG WITH MICROBIOLOGICAL PROFILE AND ANTIBIOGRAM OF UROPATHOGENS

Background:Urinary tract infection (UTI) is one of the most common infection throughout the world that may affect both the lower and the upper urinary tracts. Aims & Objective:The present study was done to determine various organisms causing Urinary tract infections and to analyse antimicrobial sensitivity patterns of those isolates. Materials and Methods:This prospective study was conducted from April 2020 to March 2021at SIMS& RH, Tumkur. All urine samples were received in Microbiology laboratory for routine microbiology investigations were inoculated on Blood agar and MacConkey agar culture media using calibrated wire loop . Urinary isolates with significant bacteriuria were further processed by conventional biochemical tests and antimicrobial susceptibility testing was done as per standard methods. Results: A total of 697 urine samples were processed in that 324 (46.48%) samples were shown as significant growth of bacteria. Most of the cases seen in (67.28%) female and under age group of 21-30(22.22%).The predominantly Gram-negative bacteria (85.18%) were isolated followed by Gram positive bacteria(10.49%) and Candida spp(4.32%). The Gram negative bacteria showed high sensitivity to Ampicillin sulbactum (88.27%) followed by Amikacin (87.24%). All Gram-positive bacteria isolates is showed sensitivity to Vancomycin (100%).We found that 235( 92.88%) Enterobacteriaceace isolates were is susceptible for Fosfomycin. Conclusion:. Antibiotic resistance has become a major clinical problem worldwide and has increased over the years.Hence antibiotic susceptibility testing should be done to diagnose and treat the drug resistant UTI cases. Fosfomycin may be given empirically in patients with uncomplicated urinary tract infection due to Enterobacteriaceace.

Prevalence of multidrug resistance Salmonella species isolated from clinical specimens at University of Gondar comprehensive specialized hospital Northwest Ethiopia: A retrospective study.

Multidrug resistance Salmonellosis remains an important public health problem globally. The disease is among the leading causes of morbidity and mortality in developing countries, but there have been limited recent studies about the prevalence, antimicrobial resistance, and multidrug resistance patterns of Salmonella isolates from various clinical specimens. Aimed to assess the prevalence, antimicrobial resistance, and multidrug resistance patterns of Salmonella isolates from clinical specimens at the University of Gondar Comprehensive Specialised Hospital, northwestern Ethiopia. A retrospective hospital-based cross-sectional study was conducted to determine the prevalence, antimicrobial resistance, and multidrug resistance patterns of isolated from all clinical specimens at the University of Gondar Salmonella Comprehensive Specialised Hospital from June 1st, 2017 to June 3rd, 2022. A total of 26,154 data points were collected using a checklist of records of laboratory registration. Clinical specimens were collected, inoculated, and incubated for about a week with visual inspection for growth and gram staining. The isolates were grown on MacConkey agar and Xylose Lysine Deoxycholate agar. Pure colonies were identified with a conventional biochemical test, and those unidentified at the species level were further identified by the analytical profile index-20E. Then, antimicrobial susceptibility was determined by the Kirby-Bauer disc diffusion technique. The multidrug resistance Salmonella isolates was identified using the criteria set by Magiorakos. Finally, the data was cleaned and checked for completeness and then entered into SPSS version 26 for analysis. Then the results were displayed using tables and figures. Of the total 26,154 Salmonella suspected clinical samples, 41 (0.16%) Salmonella species were isolated. Most of the Salmonella isolates, 19 (46.3%), were in the age group of less than 18 years, followed by the age group of 19-44 years, 11 (26.8%). In this study, S. enterica subsp. arizonae accounts for the highest 21 (51%), followed by S. paratyphi A 9 (22%). Of the Salmonella isolates, S. typhi were highly resistant to ampicillin (100%), followed by tetracycline and trimethoprim-sulfamethoxazole, each accounting for 83.3%. Furthermore, S. paratyphi A was resistant to ampicillin (100%), tetracycline (88.9%), and chloramphenicol (88.9%). The overall multi-drug resistance prevalence was 22 (53.7%; 95% CI: 39.7-61). Accordingly, S. paratyphi A was 100% multidrug-resistant, followed by S. typhi (66.6%). A low prevalence of Salmonella species was observed in the past six years. Moreover, most S. typhi and S. paratyphi strains in the study area were found to be resistant to routinely recommended antibiotics like ciprofloxacin and ceftriaxone, compared to what was reported earlier. In addition, all isolates of S. paratyphi A and the majority of S. typhi were multidrug resistant. Therefore, health professionals should consider antimicrobial susceptibility tests and use antibiotics with caution for Salmonellosis management.

Prevalence of Salmonella spp., Shigella spp., and intestinal parasites among food handlers working in University of Gondar student's cafeteria, Northwest Ethiopia.

Food-borne infections continue to be a major public health problem at the international level. The issue becomes more serious in developing countries like Ethiopia. This study aimed to examine the prevalence of Salmonella and Shigella species and intestinal parasites, as well as antimicrobial resistance patterns and associated factors among food handlers at the University of Gondar cafeteria in northwest Ethiopia. An institutional-based cross-sectional study was conducted from February to June 2021 in the University of Gondar cafeterias. Data related to the socio-demographic characteristics and hygienic practices of study participants were collected using structured questionnaires. A total of 290 stool samples were collected from food handlers. Culture and conventional biochemical tests were used to isolate the Salmonella and the Shigella species. Wet mount, Formol-ether concentration, and Kato Katz techniques were applied to identify intestinal parasites. Additionally, drug susceptibility tests were performed using the disk diffusion method. Statistical analysis was done using SPSS version 26. Of 290 food handlers' stool samples analyzed, Twenty-seven 27 (9.3%) were positive for both Salmonella and Shigella species. The prevalence of Salmonella and Shigella species was 16 (5.5%) and 11 (3.8%), respectively. Most of the isolated pathogens were resistant to tetracycline 19 (70.4%), and trimethoprim/sulphamethoxazole 19 (70.4%). The overall rate of multi-drug resistant Shigella and Salmonella isolate was 59.3%. Besides, Fifty-seven 57 (19.7%) of the participants were positive for one or more intestinal parasites. The most prevalent intestinal Parasitosis was E. histolytica/dispar 22 (7.6%), followed by G. lamblia 13 (4.5%), and Ascaris lumbricoides 11 (3.8) not washing hands after using the toilet (AOR: 4.42, 95% CI: 1.57, 10.56), and consuming unpasteurized milk (AOR: 3.14, 95% CI: 1.65, 3.96), were factors significantly associated with the prevalence of Salmonella, and Shigella infection. Similarly, not washing hands after using the toilet (AOR: 2.19, 95% CI: 1.0, 1.4), and consuming unpasteurized milk (AOR: 10.4, 95% CI: 3.8, 28.8), were factors significantly associated with the prevalence of intestinal parasites infection. The prevalence of intestinal parasites, Salmonella, and Shigella species was high. Therefore, it is imperative to implement a public health policy that includes ongoing microbiological surveillance.

Bacteriological evaluation, antibiogram, and phenotypic detection of MBL-producing gram-negative bacteria isolated from outdoor patients with Otorrhea attending Ayub Medical Complex

Otitis media is frequently prevalent in developing countries and is primarily caused by bacteria. The current study aimed to study the patients to assess the occurrence rate, susceptibility pattern, and prevalence rate of metallo beta-lactamase (MBL) producing gram-negative bacteria to prevent the prevalent strains and direct the physicians to appropriate antibiotic therapy. The research study was conducted over nine-months, from November 2020 to July 2021, in which a total of 110 samples were collected from the outdoor-patients’ ear, nose, and throat (ENT) wards using sterile swabs and inoculated immediately on suitable culture media. The isolates were identified via conventional biochemical tests, and the susceptibility pattern of each isolate was tested using the Kirby-Bauer disc diffusion method. Out of the 110 total samples, merely 88.18% (97/110) ear samples were detected positive for the various strains of bacteria both in male and female patients with a mean age of 18.21 ± 15.86, contributing 54.64% and 45.36%, respectively. The most prevalent bacterial strain was Staphylococcus aureus (49.48%), followed by Pseudomonas aeruginosa (16.50%), Proteus spp. (13.40%), S. epidermidis (7.22%), E. coli (6.19%), S. saprophyticus (4.12%), and Klebsiella spp. (3.09%). Of the gram-negative isolated strains, only 18.75% (3/16) of the P. aeruginosa strains exhibited MBL production. A high prevalence rate was observed in the age-group 0–4 years (19.58%), followed by the age-groups, 5–9 and 15–19 (16.49% each), and the age-group >39 (10.31%). Levofloxacin, ciprofloxacin, amikacin, and fusidic acid were highly effective against the gram-positive, whereas ciprofloxacin, amikacin, and levofloxacin were potent against the gram-negative (except P. aeruginosa), while carbenicillin, meropenem and imipenem against the P. aeruginosa. The MBL-producers were highly (100%) resistant to IMP and MEM while highly (100%) sensitive to CAZ. In conclusion, the current study showed a high prevalence of S. aureus, P. aeruginosa, and MBL-producing P. aeruginosa (18.75%). Nevertheless, a high degree of resistance to antibiotics was noted, which could not be ignored.

Aerobic Microbiological Spectrum and Antibiotic Resistance in Children Operated for Anorectal Abscesses.

(1) Background: Anorectal abscesses are a relatively rare pathology in childhood. Most often, male children under 1 year of age are affected. The importance of microbiological examination for the diagnosis and treatment of such patients remains debatable among surgeons, resulting in scarce data being available in the literature. We aimed to identify the aerobic microbiological spectrum and antibiotic resistance of isolates in children undergoing operation to treat anorectal abscesses. (2) Methods: We performed a case series of 102 children diagnosed and operated for anorectal abscesses over a period of 10 years (2010-2019). Purulent wound exudate was used for microbiological evaluation, which was subsequently cultured on 5% sheep-blood agar and eosin-methylene blue agar. For microbiological identification, conventional biochemical tests and semi-automated (API 20, bioMerieux, Marcy-l'Étoile, France) tests were used, as well as automated systems (Vitek-2 Compact, bioMerieux, France). Antimicrobial susceptibility testing was performed by the disk diffusion method of Bauer-Kirby and by determining the minimal inhibitory concentrations for glycopeptides. The results were interpreted according to the EUCAST standard for the corresponding year. (3) Results: Microbiological testing in children operated for anorectal abscesses mainly identified the gut commensals that normally reside in the rectal mucosa. Monocultures were found in just over half of the cases. Escherichia coli, Klebsiella pneumoniae complex, and Proteus mirabilis were the most frequently isolated. In addition, Staphylococcus aureus was found in 7% of patients. In Gram-negative bacteria, antibiotic resistance was most often observed in penicillins, cephalosporins, sulfonamides, and fluoroquinolones. (4) Conclusions: The increasing rates of antimicrobial resistance impose the need for the local monitoring of circulating commensal bacteria associated with anorectal abscesses in children to guide antibiotic therapy when indicated.

Molecular Detection of Some Most Important Pathogenic Bacteria by Multiplex PCR for Diagnosis of Subclinical Mastitis in Bovine

Background: The dairy sector faces formidable obstacles in the early detection of the bacteria causing subclinical mastitis and in the proactive treatment of those cases. The objective of the study was to access multiplex PCR assays (MPCR) and conventional methods for the concurrent identification of significant bacteria that cause sub-clinical mastitis and to compare these methods. 200 samples of pooled milk from cows and buffaloes in Rajasthan were gathered between 2020 and 2021. Methods: The primary bacterial pathogens in milk samples were identified using conventional methods such multiplex PCR, biochemical testing and culture. Result: From 200 combined samples of cow and buffalo milk, the traditional approach was able to extract 97 different strains. Staphylococcus aureus 54 (27%), Streptococcus spp. 30 (15%) and E. coli 13 (6.5%) were shown to be prevalent as single or mixed infections, respectively. Staphylococcus was the main pathogen that was discovered. concurrently, S. aureus, Streptococcus and E. coli. Direct detection of Staphylococcus 65 (32.5%), Streptococcus 37 (18.5%), E. coli and 16 (8%) by multiplex PCR was found in milk samples. The analysis revealed that because multiplex PCR assays have higher specificity and sensitivity than conventional procedures, they are more reliable. The multiplex PCR method employed in the current study was a simple and quick technique to identify the major pathogens and it has the potential to be a very helpful tool for determining the pathogens that cause environmental mastitis and evaluating the health of the herd.

Antimicrobial resistance trends in clinical Escherichia coli and Klebsiella pneumoniae in Ethiopia.

Clinicians rely on local antimicrobial resistance pattern data to guide empiric treatment for seriously ill patients when culture and antimicrobial susceptibility testing results are not immediately available. This study aimed to analyse 5-year trends in antimicrobial resistance profiles of Escherichia coli and Klebsiella pneumoniae isolates. Bacteriology reports from 2017 to 2021 at the Ethiopian Public Health Institute were analysed retrospectively. Isolates were identified using either the VITEK 2 Compact system, the BD Phoenix M50 instrument, or conventional biochemical tests. Antimicrobial susceptibility testing was conducted using either the Kirby-Bauer disk diffusion method or the VITEK 2 Compact system and BD Phoenix M50 systems available at the time of testing. The Cochran Armitage trend test was employed to test the significance of antimicrobial resistance trends over time. P-values less than 0.05 were considered statistically significant. Of the 5382 bacteriology reports examined, 458 (9%) were on E. coli and 266 (5%) were on K. pneumoniae. Both K. pneumoniae (88%) and E. coli (65%) demonstrated high resistance to extended-spectrum cephalosporins. However, both K. pneumoniae (14%) and E. coli (5%) showed lower rates of resistance to carbapenems compared to other antimicrobials. In K. pneumoniae, resistance to carbapenems (from 0% to 38%; p < 0.001) and ciprofloxacin (from 41% to 90%; p < 0.001) increased significantly between 2017 and 2021. Both organisms showed very high resistance to broad-spectrum antibiotics. Additionally, K. pneumoniae demonstrated a statistically significant rise in ciprofloxacin and carbapenem resistance. This study emphasises the significance of regular reporting of local antimicrobial resistance patterns as this information can guide appropriate empiric therapy and efforts to address antimicrobial resistance issues.

Random Amplified Polymorphic DNA (RAPD) Markers Protocol of Bacterial Isolates from Two selected General Hospitals Wastewater (HWW)

In this research work, Random amplified polymorphic DNA (RAPD) markers Protocol was used to characterize bacterial isolates from two selected General Hospitals Waste Water (HWW). The hospital environment and its wastes accumulate diseases from both inward and outward patients. It is pertinent to investigate the wastewater and study its microbial community. Wastewater samples were collected from two major General Hospitals in Akoko area, namely, Ikare and Akungba-Akoko General Hospitals. Samples were microbiologically examined for the presence of bacterial colonies. Isolated bacterial species were preliminarily identified through conventional biochemical tests and were further characterized using 16s rRNA sequencing protocol. The bacterial gene was amplified using selected primers. DNA was isolated, purified, and amplified using 16s rRNA gene sequence protocol, and the results were analyzed and assessed with the standard bacteria sequence in the NCBI database. Molecular evolutionary analyses and Phylogenetic trees were mapped out from the results obtained. it was observed that the Bacterial counts in Cfu/ml range between 10.0 x 103Cfu/ml to 24.0 x 103Cfu/ml. The highest bacterial count was observed from the test sample from Ikare followed by Akungba Akoko General Hospitals with the Cfu/ml value of 24.0 x 103 and 18.0 x103 Cfu/ml respectively. Molecular data sequence, when compared with NCBI database using BLAST showed 99.60 –99.87% bactaria phylogenic identity and E-value equal to 0 for all closely related taxa. The following bacteria were characterized using Random amplified polymorphic DNA (RAPD) markers protocol namely: Streptococcus pneumoniae, Staphylococcus aureus, Bacillus cereus, and Bacillus subtilis. this study has revealed the presence of different bacterial species in hospital waste water which pose threat to public health. Hence, proper monitoring and sewage treatment should be encouraged in different hospital settings before the disposal of these wastewaters to the public water runways