Gram-negative Bacterial Isolates Research Articles

Antibacterial Effect of Nigerian Honey on Staphylococcus aureus and Escherichia coli

Background: Honey exhibits antimicrobial activities against a wide range of bacteria in different milieu. Objective: To determine the antibacterial effect of Nigerian honey types on selected bacterial isolates; Staphylococcus aureus and Escherichia coli. For this reason raw and commercial honey samples were investigated.Methods: Two honey samples were used in this study sourced from two different institutions: raw (sunshine) and commercial (shoprite; Labsan) honey. They were tested on clinically selected Grampositive and Gram-negative bacterial isolates: Staphylococcus aureus and Escherichia coli respectively.Results: The honey samples examined were effective in inhibiting the growth of the test bacteria and the inhibitory effects were found to be superior from sunshine honey on Staphylococcus aureus. Among the selected bacteria used in this assay, Staphylococcus aureus was the most susceptible to the honey samples used and sunshine (raw) honey was found more effective.Conclusion: The Nigerian honey types used in this study were found to be alternative natural antibiotic to conventional synthetic antibiotics.International Journal of Human and Health Sciences Vol. 08 No. 01 Jan’24 Page: 65-68

Selective inhibitory activity of multidrug-resistant bacteria by zinc oxide nanoparticles

The emergence of multidrug-resistant bacteria has attracted much attention from the global community due to their potential to cause harm to health. In this context, ZnO nanoparticles (NPs) produced under different synthesis conditions were successfully applied to inhibit several species of multiresistant bacteria from clinical isolates. Five gram-positive (Staphylococcus epidermidis ATCC 35984, Staphylococcus aureus ATCC 25923, S. aureus ATCC 8095, Enterococcus faecalis ATCC 29212, and Enterococcus faecium ATCC 700221) and four gram-negative bacterial isolates (Klebsiella pneumoniae ATCC 700603, Escherichia coli ATCC 25922, Acinetobacter baumannii ATCC 19606, and Pseudomonas aeruginosa ATCC 27853), known to be involved in healthcare-associated infections and multidrug resistance, were selected for this study. Due to their physicochemical properties, we found that ZnO NPs can exhibit selective inhibition. The minimum inhibitory concentration (MIC) for multidrug-resistant bacteria sensitive to ZnO NPS was determined from varying contact time and catalyst concentration from exhaustive tests. Different characterization techniques were studied to determine the crystalline, optical, morphological, and photocatalytic characteristics of ZnO NPs. The bacterial inhibition mechanisms were discussed in detail with support of photocatalytic assays and cytotoxicity of plant species. In addition, ZnO NPs have also shown great potential for application in the environmental remediation of ecosystems contaminated by organic pollutants due to their efficiency in degrading the dye methylene blue (MB) under UV light with a photocatalytic efficiency of 75% in 60 min. The ZnO NPs showed great potential for multidrug-resistant bacteria, with MICs ranging from 256 to 512 mg/L. This demonstrates that the ZnO NPs produced here have the potential to be used as a technology with great application potential in materials used in hospital environments to mitigate the growth and development of these classes of multi-resistant bacteria.

Antibiotic susceptibility and virulence factors of bacterial species among cancer patients

Antibiotic resistance is one of the most significant challenges of the 20-s century, and the misuse of antibiotics is a driver of antimicrobial resistance. This study aimed to assess the prevalence of multidrug resistance, and detection of its produce virulence factors, including extended-spectrum β-lactamases (ESβLs), biofilm, and siderophores produced by bacterial species isolated from cancer patients. One hundred and seventy-five Gram-negative bacterial isolates were isolated from different samples collected from cancer patients admitted to the National Cancer Institute (NCI), Cairo, Egypt, and processed by standard microbiological methods. One hundred and forty-three bacterial isolates were recovered from adult patients, and 32 were recovered from children. Escherichia coli showed the highest frequency (36%), followed by Klebsiella pneumonia (30.85%), Acinetobacter baummannii (14.28%), and Pseudomonas sp. (9.14%). Antibiotic profiles revealed that bacterial isolates are highly resistant to the most commonly available antibiotics. Amikacin and gentamicin were the most effective antibiotics against isolated Gram-negative bacteria. Moreover, the vast majority of bacterial stains produce virulence factors, including EsβLs, biofilm, and siderophores. E. coli isolates produced ESβLs with rates of 25.28%, Klebsiella pneumonia (11.0%), and Pseudomonas sp. (25.0%). Among these collected bacterial isolates, 132 (75.4%) have the ability to form a biofilm to different degrees. Also, the majority of the bacteria isolates generated siderophores, with 133 (75.94%). This study revealed that a significant distribution of multidrug-resistant pathogenic bacteria may increase the burden on healthcare to prevent infections in cancer patients.

Türkiye'de Yetiştiriciliği Yapılan Levrek Balıklarında (Dicentrarchus labrax) Aeromonas veronii biovar veronii Enfeksiyonu

Fish in aquaculture systems are susceptible to infectious agents as they are kept in a densely populated and often physically restricted environment. Genus Aeromonas include well-known pathogens, and the member Aeromonas veronii has been reported to cause diseases in both humans and animals, either as primary infection or as mixed infection with other pathogens. This study describes a low mortality epizootic caused by Aeromonas veronii biovar veronii in European seabass (Dicentrarchus labrax) farmed in the Aegean Sea. The gills, kidneys and livers of moribund fish were pale. Erythema, haemorrhage and superficial ulcerative lesions were detected on the skin. In addition, petechial haemorrhage was observed on the tongue, maxilla, and operculum. The spleen was enlarged and multiple granulomas were detected in both the kidney and the spleen. Some fish had skin depigmentation, ecchymosis in the liver, and a bloody exudate in the abdominal cavity. The intestinal walls were lined with a clear yellowish fluid. Twenty-nine motile, Gram-negative bacterial isolates were obtained from the internal organs of diseased fish. According to morphology, biochemical properties and 16S rRNA gene sequencing results, all isolates were identified as Aeromonas veronii bv. veronii. All isolates were resistant to amoxicillin and ampicillin, and sensitive to oxytetracycline, enrofloxacin, ciprofloxacin, florfenicol, and flumequine.

Comparative Evaluation of Broth Microdilution With Disc Diffusion and VITEK 2 for Susceptibility Testing of Colistin on Multidrug-Resistant Gram-Negative Bacteria.

Background The rise of antibiotic resistance, particularly in Gram-negative bacteria, poses a significant global health threat. Colistin, a last-resort antibiotic, has witnessed renewed use. However, accurate susceptibility testing for colistin is challenging, with various methods available, leading to potential discrepancies. Ensuring reliable testing is crucial for effective patient treatment and antimicrobial stewardship. This study addresses the need to compare different colistin susceptibility testing methods, providing insights into their accuracy and relevance in clinical settings. Methods In this one-year prospective observational cross-sectional study conducted atIndira Gandhi Institute of Medical Sciences (IGIMS), Bihar, India, atertiary care hospital from July 2021 to June 2022, we aimed to evaluate the concordance between two widely used methods, VITEK 2 and Disc Diffusion, for antibiotic susceptibility testing in clinical multidrug-resistant Gram-negative bacterial isolates. These isolates, including species like Klebsiella pneumoniae, Acinetobacter baumannii, Klebsiella oxytoca, Pseudomonas aeruginosa, Citrobacter freundii, and Escherichia coli, were isolated from various clinical specimens. After rigorous species-level identification and quality control measures, antibiotic susceptibility testing was performed using both methods, and their agreement was assessed through Percentage Agreement analysis. Results In our study, we isolated and identified bacterial isolates from 105 patients, with a mean age of 47.30 years, demonstrating a wide age range. Pus samples were the most common type (25.7%), and K. pneumoniae was the most prevalent organism (45.7%). Antibiotic resistance patterns revealed significant challenges in treating infections caused by K. pneumoniae and A. baumannii, with resistance rates exceeding 70% for certain antibiotics. Among the 48 isolates of K. pneumoniae, the agreement was 93.8%, with 89.6% being sensitive and 6.3% being resistant by Disc Diffusion, while VITEK 2 indicated 0% resistance. E. coli isolates (n=21) had an agreement of 90.5%, with 90.5% sensitivity and 9.5% resistance by Disc Diffusion, and no resistance by VITEK 2. Conclusion The comparative analysis of antibiotic susceptibility testing methods reveals the superior performance of the VITEK 2 system, particularly in sensitivity and negative predictive value, emphasizing its potential as a reliable tool for guiding antibiotic therapy decisions.

Assessment of in vitro colistin susceptibility of carbapenem-resistant clinical Gram-negative bacterial isolates using four commercially available systems & Whole-genome sequencing: A diagnostic accuracy study

AimTo analyze the diagnostic utility of commercially available platforms and Whole-genome sequencing (WGS) for accurate determination of colistin susceptibility test results. Material & methodsAn exploratory diagnostic accuracy study was conducted in which sixty carbapenem-resistant Gram-negative bacteria were subjected to identification and AST using MALDI-TOF MS & MicroScan walkaway 96 Plus. Additional AST was performed using the BD Phoenix system and Mikrolatest colistin kit. The test isolates were subjected to Vitek-2 and WGS at CRL, Bengaluru. ResultsThere was no statistically significant agreement between the colistin susceptibility results obtained by WGS, with those of commercial phenotypic platforms. The MicroScan 96 Plus had the highest sensitivity (31 %) & NPV (77 %), and the BD Phoenix system had the highest specificity (97 %) and PPV (50 %), respectively, for determining colistin resistance. ConclusionThe utility of WGS as a tool in AMR surveillance and validation of phenotypic AST methods should be explored further.

A comparative evaluation of colistin Minimum Inhibitory Concentration determination by reference broth microdilution with other commonly used phenotypic methods in Multidrug-Resistant Gram-negative bacilli

Background: over the past decade, the dependency on colistin as a last resort antibiotic has driven the global emergence of colistin resistance among many bacterial species. This study comparatively evaluated the colistin Minimum Inhibitory Concentration (MIC) by various phenotypic methods, including the reference method of Broth Microdilution (BMD), other approved methods of Colistin Broth Disk Elution (CBDE), and Colistin Agar Test (CAT) and widely available method of Epsilometer Test (E-test) among Multidrug Resistant (MDR) Gram-negative bacteria. Methods: ninety Gram-negative bacterial isolates that were resistant to three or more classes of drugs (MDR) were included in the study. All the MDR isolates were subjected to colistin susceptibility determination by BMD, CBDE, CAT, and E-test. Results: amongst 1118 samples, 90 (8.05%) samples yielded MDR Gram-negative bacilli. All the MDR Gram-negative isolates were colistin intermediate by all four methods of phenotypic colistin susceptibility. Three Acinetobacter baumannii and two Klebsiella pneumoniae isolates that had MIC of 2 μg/mL by BMD, displayed MIC of <1 μg/mL by CBDE and CAT. Three isolates (2 Citrobacter koseri and 1 Enterobacter spp.) showed higher MIC by the E-strip method in comparison to BMD. Conclusions: our study holds significance, as there is a paucity of data comparing the four phenotypic methods for colistin MIC determination; BMD is the most reliable, gold standard method, but it is labor-intensive and requires technical expertise. In the present study, CBDE and CAT methods showed good concordance with BMD, and are easy to perform with limited logistics. Thus, they can be used as an alternative to BMD. We found that even though the Etest method was less accurate, it can still be used with caution to exclude the possibility of colistin resistance.

1698. Activity of ceftolozane/tazobactam in vitro against Gram-negative isolates from pediatric patients: SMART 2017-2021

Abstract Background The study aims to evaluate the in vitro activity of ceftolozane/tazobactam (C/T) and other common-used antibiotics against clinical Gram-negative bacterial (GNB) isolates obtained from Chinese pediatric patients using data from The Study for Monitoring Antimicrobial Resistance Trends (SMART). Methods From 2017-2021, 554 Gram-negative isolates were collected from 16 pediatric departments across China. GNB isolates from pediatric departments were mainly derived from bloodstream (n = 135), intraperitoneal (n = 129), lower respiratory tract (n = 198) and urinary tract (n = 92) infections (Figure 1). The minimum inhibitory concentrations (MICs) were tested using a Trek Diagnostic System (Thermo Fisher Scientific). Susceptibility was determined by CLSI broth microdilution and interpreted with CLSI M100 (2021) breakpoints. Results E. coli (34.30%) and K. pneumoniae (27.62%) were the most common pathogens, followed by P. aeruginosa (15.34%). Susceptibility of species with more than 20 isolates were showed in Table 1. The susceptibility of P. aeruginosa to C/T was 89.41%, which was the highest among beta-lactams and was second only to amikacin. The susceptibilities of E. coli and K. pneumoniae isolates to C/T were 92.63% and 58.17%, respectively. When exclude carbapenem-resistant E. coli and K. pneumoniae, the susceptibility to C/T increased to 96.17% and 86.14%. C/T showed similar activities to E. coli and K. pneumoniae isolated from pediatric patients < 1 year (Table 2). Thirteen P. aeruginosa isolates were collected from patients < 1 year and 13/13 were susceptible to C/T (Table 3). Conclusion P. aeruginosa and E. coli obtained from pediatric patients in China showed a high susceptibility to C/T. When excluding carbapenem resistant isolates, C/T also showed good activities against K. pneumoniae. Disclosures Pengcheng Li, M.D., MSD China: Employee of MSD China

Integration of phenotypic, qPCR and genome sequencing methodologies for the detection of antimicrobial resistance and virulence in clinical isolates of a tertiary hospital, India.

The emergence of antimicrobial resistance (AMR) and virulence in clinical isolates is a significant public health concern. The rapid and accurate detection of these traits in clinical isolates is essential for effective infection control and treatment. We demonstrated the integration of multiple detection methodologies, including phenotypic testing, quantitative polymerase chain reaction (qPCR), and genome sequencing, to detect AMR and virulence in clinical isolates. One hundred sixty-two gram-negative bacterial clinical isolates were selected for this study from the Shri Vinoba Bhave Civil Hospital, Silvassa, a tertiary government hospital. Antimicrobial susceptibility was detected by determining the Minimum Inhibitory Concentration (MIC) using Vitek-2, whereas the combined disk (CD) method was used for phenotypic detection of carbapenemase activity. The highest sensitivity rates were obtained for antibiotics colistin 87.93%, amikacin 67.52%, tigecycline 63.39%, nitrofurantoin 60.87%, and gentamycin 56.08%. The most resistant antibiotics were ceftazidime (71.93%), ciprofloxacin (67.95%) and trimethoprim/sulfamethoxazole (65.56%). Approximately 46.91% (76) of all the isolates were MBL isolates. The qPCR results confirmed the presence of blaNDM-1 in 29.01% of the isolates. The blaNDM-1 harbouring isolates in descending order, were Acinetobacter, Enterobacter cloacae, and Klebsiella pneumoniae. Klebsiella and Acinetobacter isolates were extensively drug-resistant. Whole genome sequencing performed on one of the Klebsiella pneumoniae isolates revealed the presence of many virulence factors, which increased the pathogenicity of the clinical isolates. The results showed that antimicrobial resistance, including carbapenem resistance, blaNDM-1, and virulence factors, was highly prevalent among isolates from tertiary clinical hospitals. The integration of multiple detection methodologies can potentially improve the detection of AMR and virulence in clinical isolates, leading to better patient outcomes and a reduced spread of these essential traits.

A retrospective analysis of the 5-year trends of antimicrobial resistance in gram-negative bacterial isolates from an intensive care unit at a tertiary care hospital.

Intensive care units (ICUs) in developing countries constitute a high risk for patients acquiring infection by multidrug-resistant organisms (MDROs). The rise in antimicrobial resistance (AMR) threatens the effective prevention and treatment of an increasing range of infections. The present study analyzed the local trends of AMR in Gram-negative isolates of ICU patients from a tertiary care facility in North India. This retrospective study was conducted over 5 years (January 2018-December 2022). All bacterial isolates from patients admitted to ICU during the study period were included in the study, and their AMR pattern was analyzed. In addition, sensitivity trends of different antimicrobials against the common Gram-negative bacteria were analyzed, and AMR trends were analyzed over the study period. Klebsiella spp. was the most common isolate in samples received from ICU. A rise of carbapenem-resistant microorganisms was observed over the study period. Escherichia coli and Klebsiella spp. showed around 10% and a 17% decrease in susceptibility to carbapenems, respectively. In contrast, a marked 29% decrease in sensitivity to carbapenems was observed in Acinetobacter spp. The inception of integrated stewardship measures has shown a rising trend in susceptibility and is the need of the hour to prevent the spread of MDROs. Surveillance studies help us understand the impact of AMR in hospitals and help plan prevention programs.

BACTERIAL PROFILE AND ANTIBIOTIC SUSCEPTIBILITY PATTERN OF UROPATHOGENS CAUSING URINARY TRACT INFECTIONS: OBSERVATIONSINA NEW TERTIARY CARE INSTITUTE OF A RURAL AREA OF NORTHERN INDIA

Introduction: Urinary tract infections (UTIs) are among the most common and burdensome diseases affecting outpatients and hospitalized patients, Escherichia coli being the most commonly implicated bacterium. UTIs have been treated empirically in locations where microbiological facilities are either non-existent or too expensive. This study was conducted to fill in our lack of knowledge regarding the bacterial composition and antimicrobial susceptibility patterns of uropathogens in Raebareli (Uttar Pradesh), India, and to help formulate an antibiotic policy. Materials and Methods: This retrospective study was conducted at a 600-bed tertiary care hospital in rural India, from 11th August 2021 to 10th July 2022. The study excluded patients who were on antibiotic therapy and had been hospitalized a week before their OPD visit. Midstream urine samples collected aseptically were cultured semi-quantitatively. Organisms with significant colony counts were identified using routine biochemical techniques and antimicrobial susceptibility was tested against various antibiotics as per CLSI guidelines. The data was condensed and analyzed using SPSS version 23 softwares descriptive statistics feature. Results: A total of 5,982 urine samples were considered eligible during the study period. Most of the patients were 19-39 years old (40.67%), and the majority were females (55.25%). Most of the samples were received from General Medicine (32.77%). 056 (17.65%) samples showed significant bacterial growth. 258 (4.31%) samples were not considered for further analysis due to contamination during collection. E. coli was the most common organism isolated (53.5%), followed by Enterococcus faecalis (21.4%). E. coli isolates exhibited good sensitivity to Fosfomycin (90.97%), Gentamicin (80.71%), and Imipenem (80%). Vancomycin and Linezolid showed >90% susceptibility to all the gram-positive isolates. 71.79% of the Staphylococcal isolates were MRSA. Discussion: This study marks the primary investigation into the susceptibility patterns displayed by bacterial strains causing urinary tract infections in Raebareli, Uttar Pradesh (UP) and enables a comparison with other areas of the nation. Age, sex and pyuria were all crucial factors in the incidence of UTI, and must be considered when assessing and managing UTI patients. There was no significant difference in the prevalence of UTI among various departments. E. coli is the leading cause of UTI worldwide, as in our study. The bacterial isolates exhibited noteworthy resistance to the first-line therapeutic agents which are commonly employed for treating UTIs. Our results indicate that fosfomycin, gentamicin, and nitrofurantoin are viable options for treating E. coli infections, while carbapenems and piperacillin-tazobactam can potentially manage other Gram-negatives. Vancomycin, linezolid and nitrofurantoin were the available options for Gram-positive isolates, Conclusion: Our study has revealed a significant number of Gram-negative bacterial isolates, with Escherichia coli being the main cause of UTIs. We have also conducted a comprehensive analysis of antibiogram data. Based on our findings, Nitrofurantoin and Fosfomycin are recommended to initially treat UTIs in our region. These results will apply to our inpatient and outpatient facilities and other departments within our organization.

A-263 Prevalence and Antimicrobial Susceptibility Profiles of Escherichia coli Isolated from Retail Meat Products Imported from USA and Domestically Produced on Caribbean Islands of St. Kitts and Grenada

Abstract Background Escherichia coli (E. coli) is a commensal flora of the gastrointestinal tract of humans and livestock animals. However, pathogenic and opportunistic variants are becoming a significant cause for concern in human healthcare (CDC, 2014). The emergence of these resistant strains is attributed to the overuse of antibiotics in agricultural practices (Llor, 2014). These microbes carry antimicrobial resistance (AMR) genetic markers that cause severe complications in treating intestinal and extra-intestinal infections in humans (Crecencio, 2020). The role of global meat industrialization and trade as a potential reservoir and source of transmission for AMR bacteria is considered a priority for OneHealth Initiatives, with collaborations between researchers, physicians, veterinarians, and food industries. Methods This study aims to compare the prevalence and antimicrobial susceptibility profiles of E. coli isolated from pork and poultry products imported from the USA to those domestically produced on the Caribbean Islands of St. Kitts and Grenada. This study was conducted over a 6-year period at the University of Medicine and Health Sciences (UMHS) in St Kitts and St. George’s University (SGU) in Grenada. Meat product samples were collected from retail stores around the Caribbean Islands. These meat samples were incubated at 36°C in Buffered media for 60 min and 24 h, then spread onto MacConkey agar to select for Gram-negative bacterial isolates. Lactose fermenting colonies were isolated and subcultured onto Nutrient agar. Species identification was performed using standard biochemical assays and Biomerieux API-20E test kits. Isolates were introduced to Mueller Hinton agar, and antimicrobial susceptibility was tested using the Kirby-Bauer disc diffusion. Results 100 meat product samples (50 domestic, 50 imported) were processed for investigation on the Caribbean Islands. On the Island of St. Kitts, 64 and 60 bacterial colonies were isolated from the domestic and imported samples, respectively. The prevalence of E. coli was 16% (n = 20) for imported samples and 12% (n = 15) for domestically produced samples. As Multiple drug resistance to >3 classes of antibiotics in E. coli isolates was 50% of imported isolates and 27% domestically produced. Resistance to amoxicillin was observed in 60% and 33% of E. coli isolates from Imported and Domestic origins, respectively. Resistance to trimethoprim-sulfamethoxazole was 25% and 13% in imported and domestic, respectively. Resistance to ciprofloxacin was 35% and 27% for imported and domestic, respectively. Resistance to cephalosporins was 60% in isolates from imported meat products and 33% in isolates from meat products domestically produced on the island of St. Kitts. Investigations are still ongoing for bacteria colony isolates from meat product samples from Grenada. Conclusion Imported meats products exhibit a higher prevalence of resistant E. coli, particularly in the beta-lactam class of antibiotics, including amoxicillin and cephalosporins. Therefore, the risk of importing meat products is a concern for introduction of AMR genetic markers to human populations. Active surveillance must be conducted to better understand the risk of exposure to humans from contaminated retail meat products to inform antibiotic therapy and resistance control continuously.

Colistin Susceptibility Testing by Colistin Broth Disk Elution MIC Method among Carbapenem-resistant Gram-negative Blood Culture Clinical Isolates in a Tertiary Care Setting, East Delhi, India.

Antibiotic resistance is a growing concern for bloodstream infections (BSIs), especially with the emergence of multidrug-resistant (MDR) gram-negative bacteria. In this study, we aimed to assess the pattern of colistin susceptibility using the colistin broth disc elution (CBDE) method among carbapenem-resistant gram-negative clinical isolates from blood cultures in a high burden tertiary healthcare setting in East Delhi. A total of 106 carbapenem-resistant gram-negative clinical isolates were tested. The most common isolates were Klebsiella pneumoniae, Escherichia coli, Enterobacter species, and Klebsiella oxytoca by CBDE method. All the carbapenem resistant gram-negative bacterial blood culture isolates showed intermediate colistin susceptibility. This was statistically significant by chi-square test (p<0.5). This study highlights the need to monitor colistin resistance trends in the face of increasing antimicrobial resistance. Accurate surveillance of emerging colistin resistance is crucial for effective management of BSIs caused by carbapenem-resistant gram-negative bacteria.

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

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

A Prospective, Observational and Comparative Study of In-Vitro Susceptibility of Isepamicin against Clinical Isolates from Various Clinical Sources (Triple I Study - Isepamicin in India)

Objectives: This study aims to investigate the in-vitro susceptibility of clinical isolates against Isepamicin and compared it with Gentamicin and Amikacin.&#x0D; Patients and Methods: In this multicentre prospective study, clinical specimens of patients were collected from three different regions of India. Clinical isolates from urine, intra-abdominal, broncho-alveolar lavage, endotracheal secretion, and sterile blood were included. The E-test was used to quantify the minimal inhibitory concentration (MIC) for Isepamicin, Gentamicin, and Amikacin. The percentages of bacterial isolates were categorized as susceptible, intermediate, and resistant according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines as per Comité de l'Antibiogramme de la Société Française de Microbiologie (CA-SFM) recommendation.&#x0D; Results: A total of 8 different bacterial isolates were collected from 150 clinical samples obtained from 50 patients. Respiratory (63 [42%]) and urine (44 [29.3%]) specimens were the most common sources for bacterial strains. The most identified bacterial isolates were K. pneumoniae (40 [26.6%]) and P. aeruginosa (38 [25%]). Isepamicin was found to be highly effective in urine samples and showed excellent sensitivity against E. coli (93.3%), followed by P. aeruginosa (57.9%) and K. pneumoniae (55.0%). Antimicrobial sensitivity was highest for Isepamicin (60/108 [56%]) at MIC≤1 mg/L and was most effective against Gram-negative bacterial isolates from the intensive care units (ICUs).&#x0D; Conclusions: Isepamicin could treat E. coli infections and could be an effective therapy in the treatment of urinary tract infections (UTIs). Moreover, it could also be used as an alternative to Gentamicin and Amikacin against resistant cases.&#x0D; Keywords: Aminoglycosides, Amikacin, E. coli, Gentamicin, Gram-negative bacteria, Isepamicin.

Endophytic fungal species Nigrospora oryzae and Alternaria alternata exhibit antimicrobial activity against gram-positive and gram-negative multi-drug resistant clinical bacterial isolates

BackgroundThe emergence of multidrug-resistant pathogens and the lack of new antimicrobial drugs is a major public health concern that needs urgent and innovative solutions. Endophytic fungi living in unique niches such as in endosymbiosis with plants are increasingly drawing attention as alternative sources of novel and chemically diverse compounds with unique mechanisms of action.MethodsIn the present study, ten endophytic fungi isolated from the medicinal plant, Sclerocarya birrea were screened for bioactivity against a panel of indicator bacteria. Three bioactive endophytic fungi (strains P02PL2, P02MS1, and P02MS2A) were selected and identified through ITS-rDNA sequencing. The whole broth extracts of the three selected isolates were further screened against contemporary drug-resistant bacterial pathogens. This was followed by partial purification by solid phase extraction and GC–MS analysis of bioactive fractions.ResultsThe bioactive endophytic fungi were identified as Alternaria alternata species (strains P02PL2 and P02MS1) and Nigrospora oryzae (strain P02MS2A). The whole broth extracts from N. oryzae P02MS2A exhibited a MIC of one μg/mL and 16 μg/mL against gram-negative, MDR Pseudomonas 5625574 and gram-positive MRSA 25775 clinical isolates, respectively. After partial purification and GC–MS analysis of whole broth extract from A. alternaria PO2MS1, 2-fluorobenzoic acid heptadecyl was putatively identified as the active compound in fraction C of this extract. This compound was also putatively identified in fraction E of A. alternata P02PL2, fraction B of A. alternata P02MS1 and fraction B of N. oryzae P02MS2A, and interestingly, all these fractions retained activity against the two MDR clinical isolates.ConclusionThe putative identification of 2-fluorobenzoic acid heptadecyl compound showing a broad-spectrum of activity, more especially against gram-negative MDR contemporary pathogens is highly encouraging in the initiative at developing novel drugs to combat multi-drug resistance.

Analyses of Extended-Spectrum-β-Lactamase, Metallo-β-Lactamase, and AmpC-β-Lactamase Producing Enterobacteriaceae from the Dairy Value Chain in India.

The consumption of milk contaminated with antibiotic-resistant bacteria poses a significant health threat to humans. This study aimed to investigate the prevalence of Enterobacteriaceae producing β-lactamases (ESBL, MBL, and AmpC) in cow and buffalo milk samples from two Indian states, Haryana and Assam. A total of 401 milk samples were collected from dairy farmers and vendors in the specified districts. Microbiological assays, antibiotic susceptibility testing, and PCR-based genotyping were employed to analyze 421 Gram-negative bacterial isolates. The overall prevalence of β-lactamase genes was 10% (confidence interval (CI) (7-13)), with higher rates in Haryana (13%, CI (9-19)) compared to Assam (7%, CI (4-11)). The identified β-lactamase genes in isolates were blaCMY, blaMOX, blaFOX, blaEBC, and blaDHA, associated with AmpC production. Additionally, blaCTX-M1, blaSHV, and blaTEM were detected as ESBL producers, while blaVIM, blaIMP, blaSPM, blaSIM, and blaGIM were identified as MBL producers. Notably, Shigella spp. were the dominant β-lactamase producers among identified Enterobacteriaceae. This study highlights the presence of various prevalent β-lactamase genes in milk isolates, indicating the potential risk of antimicrobial-resistant bacteria in dairy products. The presence of β-lactam resistance raises concern as this could restrict antibiotic options for treatment. The discordance between genotypic and phenotypic methods emphasizes the necessity for comprehensive approaches that integrate both techniques to accurately assess antibiotic resistance. Urgent collaborative action incorporating rational and regulated use of antibiotics across the dairy value chain is required to address the global challenge of β-lactam resistance.

Boosting voltage generation in dual-chambers microbial fuel cells through multi-parametric optimization of effects of selected factors using Box Behnken design

Low electricity output of microbial fuel cells (MFCs) has continued to limit their large-scale applications, as bioenergy sources. Thus effects of surface area of anode (0.005 to 0.015 m2), surface area of cathode (0.005 to 0.015 m2) and volume of substrate in anode chamber (750 to 1500 ml), on MFCs voltage output, were optimized. Replicated Box Behnken Design (Minitab) gave 30 runs. After 25 days operation, average voltage generated by MFCs ranged from 7.76±0.28 mV to 34.32±3.2 mV, across 10 kΩ. Response Optimizer (Minitab) indicated 0.011 m2 as optimal surface area of anode, 0.015 m2 for cathode and 1500 mL for volume of substrate in anode chamber, with estimated maximum voltage of 41.83 mV, when used. This gives 1:1.3:136,363 ratio for surface area of anode, surface area of cathode and volume of substrate in anode, which could be useful in scaling up the device. On application of these optima, highest and lowest average voltages of 54.5±3.2 mV and 20.1±2.7 mV were generated. This maximum voltage was 30.3% higher than the estimate by Response Optimizer, and 58.8% higher than the highest average voltage recorded without optimization. Again, the lowest average voltage (20.13±2.7 mV) obtained after optimization was 159.4% higher than the lowest voltage (7.76±0.28 mV) recorded without optimization. BOD of piggery wastewater, used as substrate, reduced by 18.9%, while COD declined by 31.4%. Diverse Gram-positive and Gram-negative bacterial isolates were identified in the wastewater. Therefore, Box Behnken design is useful optimization of factors, to boost the output of MFCs.

Changing Trends of Antimicrobial Resistance in Clinical Isolates Yielded from Lower Respiratory Tract Specimens of ICU Patients-A Two-Year Study

Objective: To determine changing trends in antimicrobial resistance among Gram-negative bacterial isolates yielded in lower respiratory tract specimens in intensive care settings.&#x0D; Study Design: Cross-sectional study&#x0D; Place and Duration of Study: Armed Forces Institute of Pathology, Rawalpindi Pakistan, Jul 2018 to Jun 2020.&#x0D; Methodology: The study was carried out on 819 isolates from lower respiratory tract specimens collected from ICUs of multiple centres all over Pakistan. The antimicrobial susceptibility testing was performed according to performance standards provided by CLSI and EUCAST. Antimicrobial resistance trends were analyzed.&#x0D; Results: In Acinetobacter baumannii, resistance increased against Carbapenems (92% to 97.4%) and Polymyxins (0% to 5.3%).Pseudomonas aeruginosa showed increasing resistance, with Imipenem (33.3% to 46.9%), Meropenem (27.3% to 43.6%) and Polymyxin (0% to 3%). In Klebsiella pneumoniae, the resistance against Carbapenems rose from 60.5% to 79.2%, for Imipenem 68.4% to 83% for Meropenem. Polymyxin resistance was much higher in K. pneumoniae (increasing from 22% to 24.5%). In Escherichia coli, resistance increased from 23.5% to 41.7% for Imipenem 35.5% to 50% for Meropenem, and 0% to 8.3% for Polymyxins, whereas Tigecycline showed decreasing resistance trend. Other antimicrobials tested showed increasing resistance as well.&#x0D; Conclusion: Antimicrobial resistance is emerging so rapidly that the post-antibiotic era is approaching sooner than later. Extensive and up-to-date insight regarding antimicrobial resistance rates and trends against significant pathogens is required to deal with this emerging dilemma.

Characterization of Carbapenemase- and ESBL-Producing Gram-Negative Bacilli Isolated from Patients with Urinary Tract and Bloodstream Infections.

A total of 199 Gram-negative bacterial isolates from urinary tract infections and 162 from bloodstream infections were collected from 12 healthcare systems throughout the United States between May 2021 and August 2022. The isolates, phenotypically non-susceptible to 2nd or 3rd generation cephalosporins or carbapenems, were characterized through antimicrobial susceptibility testing and whole genome sequence analysis to obtain a broad snapshot of beta-lactamase-mediated resistance among these two sample types. Overall, 23 different carbapenemase genes were detected among 13 species (20.5% of isolates). The blaKPC-3 and blaKPC-2 subtypes were the most common carbapenemase genes identified, followed by blaNDM and the co-carriage of two different blaOXA carbapenemases by Acinetobacter baumannii isolates. All carbapenemase-producing A. baumannii isolates were mCIM negative. Extended-spectrum beta-lactamase genes were identified in 66.2% of isolates; blaCTX-M-15 was the most common. AmpC genes, both plasmid and chromosomal, were detected in 33.2% of isolates. Importantly, 2.8%, 8.3%, and 22.2% of blaKPC-positive organisms were susceptible to ertapenem, imipenem, and meropenem, respectively. The correlation between broth microdilution and disk diffusion results was high for most drugs except cefepime, where the detection of resistance was statistically lower by disk diffusion. Thus, there were gaps in the accuracy of susceptibility testing for some mechanisms of resistance.