Antibiotic Susceptibility Testing Research Articles

Antibacterial activity of fat from black soldier fly (Hermetia illucens) larvae against antibiotic resistant Campylobacter spp. strains

Abstract In this study, the antibacterial effect of fats derived from black soldier fly larvae (BSFL) were evaluated against various antibiotic-resistant strains of Campylobacter jejuni and Campylobacter coli. BSFL fats with different free fatty acids (FFAs) contents were obtained based on the larvae’s processing conditions. Initially, antibiotic susceptibility testing revealed that all Campylobacter strains were resistant to at least one antibiotic, with two strains classified as multidrug-resistant. The antibacterial activity of the BSFL fats was directly correlated with their FFAs content. Fats obtained through freeze-drying and defatting via supercritical fluid extraction (SFE) exhibited the strongest antibacterial effects against Campylobacter spp., reducing bacterial growth by 4.1 to 5.6 log CFU/mL in C. jejuni and demonstrating bactericidal activity (reducing bacterial growth >6 log CFU/mL) against all C. coli strains. An analysis of the individual contributions of FFAs in BSFL fat revealed that lauric and linoleic acids were bactericidal (reducing bacterial growth >8 log CFU/mL), completely inhibiting the growth of C. jejuni and C. coli strains, while myristic, palmitic, and oleic acids showed no effect on Campylobacter growth. These findings suggest that BSFL fat, which has been shown to enhance broiler growth, could also serve as an effective strategy to combat Campylobacter spp. contamination in the food chain.

Assessment of typing methods, virulence genes profile and antimicrobial susceptibility for clinical isolates of Proteus mirabilis

Proteus mirabilis (P. mirabilis) is one of the most important causative pathogens associated with complicated urinary tract infections with a 20% incidence. For epidemiological determinations, several phenotypic and molecular typing methods have been implicated. Sixty P. mirabilis isolated undergo antibiotic susceptibility test by standard Kirby Bauer method. They showed high resistance to nitrofurantoin and trimethoprim/sulfamethoxazole that appear mainly in 3rd age group. The 2nd age group comprised most of the resistant isolates to the tested antibiotics. A total of 73.33% of isolates were classified as multi drug resistance (MDR) and 78.3% of isolates were distributed in several antibiotypes with MAR index over 0.2. Twenty-one isolates were strong biofilm-producers and they were significantly related to MDR. Different virulence factors as protease, urease and hemolysin production are detected. Detection of several virulence genes by PCR; zapA and ureC were harbored by all isolates, followed by rsbA (95%), ureA and flaA (93%), hpmA (91.7%) and mrpA (73.3%). Determination of genetic diversity between isolates was performed by different methods (RAPD, ISSR, ERIC, BOX-AIR and REP-PCR) by using several parameters as typeability and discriminatory power indicating that ERIC-PCR was the best method followed by REP-PCR 1R. Rand’s & Wallace coefficients were used for calculating the congruence among typing methods. Conclusions: The results obtained from both conventional and molecular typing methods indicated that molecular methods are superior to conventional methods in the discrimination of isolates. ERIC-PCR and Rep-PCR provide high discrimination ability among P. mirabilis clinical isolates contributing to epidemiological studies.

Multidrug-Resistant Corynebacterium spp., Escherichia coli, and Klebsiella spp. Isolated from the Lungs of a Six-Year-Old Arewa Stallion

This manuscript presents a case of multidrug-resistant (MDR) bacterial infections in the respiratory system of a6-year-old Arewa stallion, thus highlighting the importance of diagnostic microbiology. History revealed that the stallion exhibited clinical signs, including high fever, hemoglobinuria, and edema of the head and neck, and this was followed by death despite medical intervention. Postmortem was pneumonia; hence lung sample was sent for microbiological analyses. The diagnostic microbiological techniques utilized were culture on blood MacConkey and Eosin Methylene Blue agars, and biochemical tests were used to confirm the bacterial identities, followed by antibiotic susceptibility testing. The outcomes revealed the isolation of Corynebacterium spp., Escherichia coli, and Klebsiella spp. Antibiotic susceptibility testing revealed extensive resistance, with Corynebacterium spp. resistant to Septrin, Chloramphenicol, Sparfloxacin, Ciprofloxacin, and more. E. coli and Klebsiella spp. were resistant to Ciprofloxacin, Gentamicin, Nitrofurantoin, Ofloxacin, Cefuroxime, Ceftazidime, Augmentin, and Cotrimoxazole. These findings emphasize the diagnostic challenges caused by MDR pathogens in equine respiratory diseases.

Antibiotic Susceptibility Patterns in Gram-Positive and Gram-Negative Isolates from Post-Operative Wound Infections

Background: Post-operative wound infections (POWIs) are a leading cause of morbidity and increase in healthcare costs, especially in low resource settings. The rising prevalence of antibiotic resistance among pathogens also increases the complexity of treatment strategies, highlighting the importance of local surveillance of microbiological and resistance patterns. This study aims to find out the antibiotic susceptibility patterns of gram positive and gram negative isolates from the post-operative wound infections from women undergoing obstetric and gynecological surgeries. Methods: A cross-sectional study was conducted at Department of Obstetrics and Gynecology, Dhaka Medical College Hospital, Dhaka from October 2019 to September 2021. A total of 200 women with post-operative wound infections were purposively selected. Among them, 134 had culture-positive findings. Bacteriological analysis and antibiotic susceptibility testing were performed on isolates following standard microbiological methods. Results: Among the 134 culture-positive cases, 33.6% of isolates were gram-positive, with Staphylococcus aureus being predominant. Gram-negative bacteria accounted for 66.4% of isolates, with E. coli and Pseudomonas aeruginosa equally prevalent (20.9%). Gram-positive isolates showed high sensitivity to piperacillin-tazobactam (91.1%) and sulbactam (53.3%) but were resistant to ceftriaxone (48.9%) and cefixime (51.1%). Gram-negative bacteria exhibited sensitivity to amikacin and meropenem but demonstrated widespread resistance to ceftriaxone and doxycycline. Conclusion: This study describes the diverse microbiological profile and high prevalence of antibiotic resistance in POWIs. Novel antibiotic therapy and strict infection control measures are essential to curb the burden of these infections.

Evaluation of the In vitro efficacy of ceftazidime-avibactam against Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa isolates from respiratory tract cultures in intensive care units

Aims: Worldwide, an increase in multidrug resistance is observed in Escherichia coli (E. coli), Klebsiella pneumoniae (K. pneumoniae), and Pseudomonas aeruginosa (P. aeruginosa) isolates, leading to challenges in the treatment of infections caused by these pathogens. This study aims to investigate the in vitro efficacy of ceftazidime-avibactam (CZA) against isolates containing K. pneumoniae, E. coli, and P. aeruginosa strains obtained from respiratory tract samples sent from intensive care units. Methods: A retrospective analysis was conducted on 653 Enterobacterales (E. coli, K. pneumoniae) and P. aeruginosa isolates obtained from respiratory tract cultures, including sputum, tracheal aspirates, and bronchial lavage, from patients over 18 years old admitted to the intensive care units of Ordu University Training and Research Hospital between May 1, 2021, and May 1, 2024. Automated systems were used to identify the pathogens and perform antibiotic susceptibility testing. Discriptive data analysis was conducted using SPSS version 24.0. Results: A total of 653 isolates from respiratory tract samples were included in the study, consisting of 368 Enterobacteriaceae [61 E. coli (9.3%) and 307 K. pneumoniae (47%)] and 285 P. aeruginosa (43.7%). These samples were isolated from endotracheal aspirate (69.5%), sputum (27.9%), and bronchoalveolar lavage (2.6%). Among all isolates, 364 (55.7%) were found to be sensitive to carbapenems, while 289 (44.3%) were carbapenem-resistant. Of the samples, 631 (96.6%) were sensitive to CZA, while 22 (3.4%) were resistant. Although resistance to CZA was detected in 3.6% of K. pneumoniae isolates and 3.9% of P. aeruginosa isolates, no resistance was detected in E. coli. Colistin resistance was observed in 15.3% of K. pneumoniae and 5.6% of P. aeruginosa isolates, but was absent in E. coli isolates. Resistance rates to other antibiotics were as follows for E. coli, K. pneumoniae, and P. aeruginosa isolates, respectively: amikacin (3.3%, 46.6%, 8.1%), ciprofloxacin (73.8%, 73.6%, 85.9%), ceftazidime (67.2%, 77.8%, 35.8%), piperacillin-tazobactam (26.2%, 70%, 37.2%), and trimethoprim-sulfamethoxazole (52.5%, 66.4%, 0%). Conclusion: In our study, CZA was found to be the most effective antibiotic against multidrug-resistant Enterobacterales and P. aeruginosa isolates, followed by colistin.

ANTIBIOTIC SUSCEPTIBILITY PATTERN OF E. COLI ISOLATED FROM THE URINE SAMPLES OF PATIENTS VISITED CIVIL HOSPITAL MADYAN SWAT

Background: Antibiotic resistance is a global challenge, particularly in urinary tract infections (UTIs) caused by Escherichia coli (E. coli), which is a leading pathogen in both nosocomial and community-acquired infections. Regular monitoring of antibiotic susceptibility profiles is essential due to the rising prevalence of drug-resistant bacteria. This study focused on evaluating the antibiotic susceptibility pattern of E. coli isolated from urine samples of patients attending Civil Hospital Madyan, Swat, to guide effective empirical therapy. Objective: To determine the antibiotic susceptibility profile of E. coli isolated from urine samples of UTI patients visiting Civil Hospital Madyan, Swat. Methods: A cross-sectional study was conducted from December 2022 to July 2023 in the Department of Microbiology, Government Degree College Madyan, Swat. A total of 450 mid-stream urine samples were collected from patients with suspected UTIs. The samples were cultured on MacConkey agar and incubated at 37°C for 24–48 hours under aerobic conditions. Isolated bacterial colonies were identified using gram staining, microscopy, and biochemical tests. Antibiotic susceptibility testing was performed on Mueller-Hinton agar using the Kirby-Bauer disc diffusion method with multiple antibiotic discs. The bacterial isolates were categorized into resistant (R), sensitive (S), or intermediate (I) groups by measuring inhibition zone diameters. Data were statistically analyzed and presented in figures and tables. Results: Out of 450 urine samples, 125 (27.0%) were culture-positive, with E. coli being the most frequently isolated bacterium (60%). Culture positivity was higher in females (70%) than males (30%). Among hospital areas, samples from the outpatient department (70%) showed the highest culture positivity, followed by wards (25%) and the emergency department (5%). Antibiotic susceptibility testing revealed that E. coli exhibited the highest resistance to ampicillin (97.1%), ceftriaxone (92%), moxifloxacin (88.2%), cefixime (86%), and ceftazidime (80.1%). Conversely, the highest sensitivity was observed to fosfomycin (95.3%), sulzone (85.2%), imipenem (85.0%), and amikacin (78%). Conclusion: This study confirmed that E. coli is the predominant pathogen causing UTIs and displayed significant resistance to commonly used antibiotics such as ampicillin, ceftriaxone, moxifloxacin, cefixime, and ceftazidime. However, fosfomycin, imipenem, and amikacin were found to be the most effective antibiotics against E. coli. These findings underscore the importance of conducting antibiotic susceptibility testing to guide effective therapy and combat antimicrobial resistance.

Genomic Characterization of Carbapenemase-Producing Enterobacteriaceae from Clinical and Epidemiological Human Samples.

Background/Objectives: Infections caused by multidrug-resistant (MDR)bacteria pose a significant public health threat by worsening patient outcomes, contributing to hospital outbreaks, and increasing health and economic burdens. Advanced genomic tools enhance the detection of resistance genes, virulence factors, and high-risk clones, thus improving the management of MDR infections. In the Autonomous Community of Aragon, the diversity and incidence of carbapenemase-producing Enterobacteriaceae (CPE) have increased during the last years. This study analyses CPE trends at a tertiary hospital in Spain from 2021 to 2023, aiming to optimize personalized medicine. Methods: CPE isolates were the first isolate per patient, year, species, and carbapenemase from January 2021 to December 2023. Additional metadata were collected from the laboratory's information system. Antibiotic susceptibility testing was performed by broth microdilution. Whole-genome sequencing (WGS) was performed using Illumina short reads. De novo assembly was used to generate draft genomes in order to determine their complete taxonomic classification, resistome, plasmidome, sequence type (ST), core-genome multilocus sequence typing (cgMLST), and phylogenetic relationships using a suite of bioinformatics tools and in-house scripts. Results: Between 2021 and 2023, 0.4% out of 38,145 Enterobacteriaceae isolates were CPE. The CPE rate tripled in 2022 and doubled again in 2023. The most common species was Klebsiella pneumoniae (51.8%) and the most common carbapenemase was blaOXA-48. WGS revealed concordant species identification and the carbapenemase distribution in detail. Resistance rates to critical antibiotics, such as carbapenems, were variable, but in most cases were above 70%. Genetic diversity was observed in WGS and phylogenetic analyses, with plasmids often mediating carbapenemase dissemination. Conclusions: The increasing rate of CPE in healthcare settings highlights a critical public health challenge, with limited treatment options. Genomic characterization is essential to understanding resistance mechanisms, aiding therapy, limiting outbreaks, and improving precision medicine.

Isolation, Identification, and Antimicrobial Susceptibilities of Bacteria from the Conjunctival Sacs of Dogs with Bacterial Conjunctivitis in Different Regions of Wuhan, China.

In order to investigate the bacterial species present in the conjunctival sacs of dogs with bacterial conjunctivitis in Wuhan (Hongshan District, Wuchang District, Jiangxia District, and Huangpi District) and their resistance to aminoglycoside antibiotics, samples of conjunctival sac secretions were collected from 56 dogs with bacterial conjunctivitis in various regions of Wuhan. Drug susceptibility testing for aminoglycoside antibiotics was performed on the most commonly isolated gram-positive and gram-negative bacteria. The expression of two aminoglycoside modifying enzyme genes, aacA-aphD and aac (6')-Ib, and three 16S rRNA methyltransferase genes, rmtB, rmtE and npmA, were analyzed by PCR. The results showed that a total of 123 bacterial strains were cultured from 56 conjunctival sac secretion samples, with Staphylococcus being the most commonly isolated species, followed by Escherichia. Among them, 14 strains of Staphylococcus pseudointermedius were not resistant to tobramycin, amikacin, gentamicin or neomycin, but the resistance rates to streptomycin and kanamycin were 35.71% and 42.86%, respectively. Among them, 14 Escherichia coli strains were not resistant to tobramycin and gentamicin, but they showed high resistance rates to neomycin and kanamycin (both at 50%). The detection rate of the aacA-aphD gene in Staphylococcus pseudointermedius strains was 100%. The detection rates of the rmtB gene and rmtE gene in Escherichia coli were 85.71% and 28.57%, respectively, while the aac(6')-Ib gene and npmA gene were not detected.

Assessment of biofilm formation, antibiotic resistance patterns, and the prevalence of adhesion-related genes in clinical Staphylococcus aureus isolates.

This study aimed to evaluate the biofilm formation abilities of clinical Staphylococcus aureus strains, assess their antibiotic susceptibility patterns, and identify the prevalence of adhesion-associated genes. In this study, a total of 60S. aureus strains were collected from urine, pus, wounds, blood, body fluid, and sputum in health centers affiliated with Abadan University of Medical Sciences, Iran. Strains were identified via microbiological methods and polymerase chain reaction (PCR) to target the nuc gene. Antibiotic susceptibility testing (AST) was conducted via the disc diffusion method. Methicillin-resistant S. aureus (MRSA) strains were identified by cefoxitin disc diffusion and PCR targeting the mecA gene. Biofilm formation was assessed via a microtiter plate assay, and the prevalence of adhesion-encoding genes was evaluated via PCR. The data were analyzed in Excel and SPSS via statistical methods, with P-values <0.05 considered significant. Using AST, daptomycin and linezolid were the most effective antibiotics (100% susceptibility rate). According to the results of the cefoxitin disc test, 48.3% (n=29/60) of the strains were MRSA. All the MRSA strains harbored the mecA gene. In total, 32% of the strains were biofilm producers. Moreover, 56.2%, 28.1%, and 15.6% of the strains produced weak, moderate, and strong biofilms, respectively. There were no significant differences between the MRSA and MSSA strains in terms of the association of biofilm formation with antibiotic resistance except for erythromycin (P-value=0.0087), gentamicin (P-value=0.0009), and penicillin (P-value=0.0009). The most prevalent biofilm-encoding genes were icaA (76.7%), followed by icaD (70%), clfA (65.0%), and fnbA (53.3%). This study identified MRSA strains with biofilm-forming abilities that possess adhesion-associated genes. The most prevalent biofilm-encoding gene was icaA. To prevent further spread of these strains, regional preventive measures are needed.

WAR IN THE MIDDLE EAR: MICROBIOLOGY OF CHRONIC SUPPURATIVE OTITIS MEDIA WITH SPECIAL REFERENCE TO ANAEROBES AND ITS ANTIMICROBIAL SUSCEPTIBILITY PATTERN: OPTIMIZING ANTIMICROBIAL THERAPY IN A TERTIARY CARE HOSPITAL OF RURAL INDIA

Objective: To isolate etiological organisms of Chronic Suppurative Otitis Media (CSOM), study the antimicrobial susceptibility pattern, study the risk factors and associated co-morbid conditions with CSOM, and detect biofilm production in isolated bacteria. Methods: An ear discharge specimen was obtained from the diseased ear of the patient, using three separate sterilized swabs. One of the swabs was used for Gram staining and Aerobic culture. Antibiotic susceptibility testing was done by the Kirby-Bauer standard disc diffusion method all the g-negative isolates were screened for Extended-Spectrum Beta-Lactamase (ESBL) production and Amp C ß-lactamase production. Methicillin-Resistant Staphylococcus Aureus (MRSA) screening was done using a Cefoxitin disc. Biofilm production was done using the Tube method and Microtiter plate method. Anaerobic culture was inoculated into Robertson’s Cooked Meat (RCM) broth. Potassium hydroxide (KOH) mount was done using 10% KOH for the presence of budding yeast cells, fungal hyphae, and spores. Tubes showing positive cultures were examined by LactoPhenol Cotton Blue (LPCB) mount. Final Candida speciation and Antifungal susceptibility testing was done using VITEK-2. Results: A total of 500 patients were included in the study, with 62.8% being males. The most common organism isolated was Pseudomonas aeruginosa. Fungi accounted for 30 isolates (5%) and anaerobes were 5 isolates (1%). Gram-negative bacteria were found to be most sensitive to Piperacillin-tazobactam (96%) while the highest resistance was noted to Amoxicillin-clavulanate (65%). 100% of Staphylococcus aureus isolates were sensitive to Vancomycin, Linezolid, and Teichoplanin.13% of S. aureus were found to be Cefoxitin resistant (MRSA). The highest resistance in S. aureus isolates was noted to Ciprofloxacin (65%).13% of isolates of Gram-negative bacteria had ESBL production while Amp C β-Lactamase production was noted in 9% of isolates. Out of 186 isolates tested for biofilm production, 149 isolates (80%) showed biofilm production. A total of 30 fungal isolates were obtained in culture. Aspergillus spp. Accounted for 66.6% of fungal isolates while Candida spp. accounted for 33.3% of fungal isolates. The sensitivity of Candida isolates was tested to Fluconazole, Itraconazole, Voriconazole, and Flucytosine and they were found to be 100% sensitive to all the antifungals tested. Conclusion: CSOM often has polymicrobial pathology and creates a war in the middle ear environment, augmented by mixed aerobic and anaerobic organisms and synergism between aerobic and anaerobic organisms. A high rate of Clindamycin resistance was seen in anaerobe which is noteworthy because this is a frequently utilized antimicrobial for treating anaerobic infections.

Prevalence, aetiology and antimicrobial resistance profile of diabetic individuals suffering from community-acquired urinary tract infection.

Introduction. The rise in antimicrobial resistance poses a significant threat to global health, particularly among diabetic patients who are prone to urinary tract infections (UTIs).Hypothesis. Pathogens that cause UTI among diabetic patients exhibit significant multidrug resistance (MDR) patterns, necessitating more precise empirical treatment strategies.Aim. This study aimed to determine the prevalence of UTI among diabetic patients and study the antimicrobial susceptibility profiles of uropathogens, detected and identified the potential differences in age groups and between genders, focusing on MDR and gender-based variations, causing a global concern in deciding empirical treatment.Methodology. A prospective study was conducted from August 2021 to December 2023 in Gujarat, India. During the period, 1023 diabetic patients with symptoms of UTI were diagnosed by urine culture and 280 individuals tested positive for UTIs. Antibiotic susceptibility testing was carried out on these 280 micro-organism isolates.Results. Among the 280 UTI-positive patients, 166 (59.29%) were females and 114 (40.71%) were males, with the prevalence of UTI in diabetic females being 27.34% (166/607) and males being 27.40% (114/416). Among the isolated uropathogens, Escherichia coli (56.78%) was the predominant organism followed by Pseudomonas aeruginosa (13.57%) and Klebsiella (13.21%). High resistance was noted to various antibiotics in Gram-negative bacteria including both genders. In E. coli, resistance was predominantly high against the penicillin sub-class of the beta-lactam group (70.23%, 69.58%), cephalosporins (66.23%, 76.52%) and least against nitrofurans (30.10%, 40%) in males and females, respectively. Klebsiella has shown higher resistance to cephalosporins (66.23%, 76.52%) and aminoglycosides (60.92%, 62.66%) and least resistance to carbapenem (41.67%) and phosphonic (33.33%) in males and females, respectively. A high proportion of isolates, ~82.5%, exhibited MDR. Among these MDR isolates, those from female patients accounted for a higher percentage (58.44%) compared with males (41.55%). The highest prevalence of MDR was observed in the 41-60-year age group. This pattern highlights notable differences in MDR prevalence across gender and age groups.Conclusion. The high prevalence of UTI caused by MDR organisms based on gender and age group highlights the need for clinicians to choose antibiotics more judiciously for empirical treatment, thereby reducing misuse and overuse in the community. For diabetic UTI patients in this region, nitrofurantoin may be recommended for uncomplicated cases due to low resistance in E. coli, while fosfomycin could be a viable alternative for Klebsiella-related infections. Carbapenems may be reserved for severe cases with MDR pathogens, and combination therapy could be considered for complicated infections, particularly in high-risk age groups.

Clinical Profile and Antibiotic Susceptibility Patterns of Cronobacter sakazakii in the Northern Region of Oman

Abstract Background: Cronobacter sakazakii is an opportunistic pathogen that mostly affects neonates, infants, and elderly people with weakened immune systems. No study has reported the frequency and antibiotic susceptibility patterns of C. sakazakii from Oman, and thus this study was conducted to fill this gap in the literature. Materials and Methods: This single-center retrospective study included C. sakazakii isolates identified from different clinical samples of patients treated at Sohar Hospital, Oman, between January 2017 and December 2023. Bacterial identification and antibiotic susceptibility testing were done using the VITEK II automated microbiological system in accordance with the Clinical Laboratory Standards Institute (CLSI) guidelines. Results: A total of 185 C. sakazakii isolates were included, most commonly from patients aged &gt;60 years (42.7%) and &lt;1 year (11.4%). C. sakazakii strains had high susceptibility (&gt;80%) to most of the tested antibiotics; however, for beta-lactam antibiotics, it ranged from 0% to 50%. Approximately 26.5% of the strains were multidrug resistant. Independent risk factors for increased frequency of multidrug-resistant strains were urinary catheterization (P = 0.002), surgery (P = 0.021), previous antibiotic therapy (P = 0.047), and critical care unit admission (P = 0.048). About one-fifth of the patients experienced life-threatening C. sakazakii infections such as septicemia (15%) and pneumonia (4.7%). All deaths due to septicemia occurred in the &gt;60 years (n = 12) and &lt;1 year (n = 4) age groups. Conclusions: Cronobacter sakazakii isolates from the North Batinah region of Oman were most frequently isolated from elderly and infant patients and had high antibiotic susceptibility; however, the significant resistance against beta-lactams suggests their low effectiveness. The high number of multidrug-resistant strains coupled with the independent risk factors suggests the need for following stricter antibiotic stewardship protocols and infection control practices.

Molecular characterization and genome sequencing of selected highly resistant clinical isolates of Pseudomonas aeruginosa and its association with the clustered regularly interspaced palindromic repeat/Cas system.

The presence of the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas system in the superbug Pseudomonas (P) aeruginosa presents a unique opportunity to precisely target and edit bacterial genomes to modify their drug resistance. The objective was to detect the prevalence of CRISPR in extensively and pan-drug-resistant Pseudomonas aeruginosa and to determine the utility of whole-genome sequencing (WGS) for the analysis of the entire genome for such strains. The antimicrobial susceptibilities of one hundred isolates were assessed using the antibiotic susceptibility test (AST) card of the VITEK system. The presence of the CRISPR/Cas system was determined via specific primers using conventional polymerase chain reaction (PCR). Further, WGS was conducted using a DNA nanoball sequencing platform via BGI-Tech for the isolates of interest. Out of 54 resistant Pseudomonas aeruginosa isolates, 33 (33.0%) were metallo-β-lactamase producers. Cas1, Cas3, CRISPR1, and CRISPR2 were positive in 6.0% of isolates, while incomplete CRISPR1-Cas systems alone were found in 15.0%. Also, CRISPR2-type was found intact in 26% of isolates. The prevalence of resistance to antimicrobials in P. aeruginosa isolates was significantly greater in the CRISPR/Cas-negative group compared to the CRISPR/Cas-positive. Significant relationships for variables were examined using Fisher's exact tests using Chi-squared and a P-value of <0.05 as a statistical threshold. Further, on examination of CRs as a collective entity, encompassing both extensive drug resistance (XDR) and pan-drug resistance (PDR), it becomes evident that the vast majority of these strains (n=29; 87.8%) lacked CRISPR/Cas systems. In phylogenic analysis, PDR-P. aeruginosa revealed a very close evolutionary relationship with those originating from Kazakhstan, while XDR was globally unique. Further, the entire genome showed the presence of unique virulence and resistant pseudomonal genes. The CRISPR/Cas system and drug resistance are antagonistic to one another. XDR and PDR P. aeruginosa represent a potential threat to public health and contribute to the seriousness of associated illnesses by leading to resistant infections. Further, WGS for the two strains revealed resistance to multiple antibiotics. It is important to examine specific antimicrobial resistance (AMR) pathways, which suggests that a significant number of resistant genes in these isolates indicate a loss of CRISPR genes in the two strains. Furthermore, the WGS approach can lead to a better understanding of the genomic mechanism of pseudomonal resistance to antibiotics.

Vancomycin-resistant Enterococcus – a study on its prevalence from different clinical samples in a rural medical college hospital

Background: Vancomycin-resistant Enterococcus (VRE) is an important cause of nosocomial infection with Enterococcus faecium causing most of the VRE infections. Widespread use of glycopeptides in health care facilities has led to the development of VRE and enterococcal infections with high-level resistance to aminoglycosides, beta-lactamase production and glycopeptide (including vancomycin) resistance are difficult to treat and often pose a therapeutic challenge to health care facilities. Aims and Objectives: This study aimed to determine the antibiotic susceptibility pattern of Enterococcus species from various clinical specimens and to find out the occurrence rate of vancomycin-resistant enterococci. Materials and Methods: This study was conducted at the Department of Microbiology of Tamralipto Government Medical College and Hospital, East Midnapore, West Bengal. A total of 688 clinical samples were the total sample size taken. Isolation and identification of Enterococcus spp. were done by standard microbiological procedures such as culture, Gram staining, and suitable biochemical tests were conducted. Antibiotic susceptibility testing was done by the Kirby–Bauers disc diffusion method on Mueller–Hinton agar and results were interpreted according to Clinical and Laboratory Standards Institute (CLSI) guidelines 2023. Teicoplanin sensitivity was performed for those isolates showing resistance to vancomycin. For strains showing resistance to vancomycin by the Kirby–Bauers disc diffusion method, vancomycin minimum inhibitory concentrations (MIC) were performed by E-test (Hi media) with MIC breakpoints between 4 and 32 as per CLSI criteria. This was compared with the control strain of American Type Culture Collection Enterococcus faecalis 29212 as per CLSI 2023 guidelines. Results: A total of 48 Enterococcus isolates were obtained from 688 clinical samples; 31 (8.05%) were detected from 385 urine samples, 9 (6.72%) were detected from 134 blood samples, 5 (5.15%) were detected from 97 pus/wound swab, and 3 (4.17%) were detected from 72 bronchoalveolar lavage fluid samples. Among the 48 Enterococcus isolates, 13 (27.08%) were vancomycin-resistant out of which nine were E. faecalis and four were E. faecium. Enterococcus species showed maximum resistance toward ciprofloxacin followed by ampicillin and maximum sensitivity toward teicoplanin and linezolid. Conclusion: Implementation of strict infection control measures, antimicrobial policies, and proper surveillance are required to identify, contain, and treat VRE infections to reduce mortality and morbidity.

Antibiotic resistance and virulence profile of Klebsiella pneumoniae isolated from wild Sumatran Orangutans (Pongo abelii)

Objective: Orangutans (Pongo abelii), as endemic primates of Indonesia, are characterized by a predominantly arboreal lifestyle. Klebsiella pneumoniae (K. pneumonia) and other Gram-negative bacteria are present in the Indigenous flora of many mammals, including orangutans. This study aimed to investigate the antibiotic resistance and virulence profile of K. pneumonia isolated from wild Sumatran orangutans. Materials and Methods: This study investigated 10 fecal samples from wild Sumatran orangutans from the Gunung Leuser National Park, Aceh, Indonesia. Biochemical and molecular identification of K. pneumoniae using the RNA polymerase subunit b gene and detection of virulence-associated genes. In addition, molecular detection of antibiotic resistance genes was performed to character¬ize the resistance mechanisms in the isolates. Results: K. pneumonia was detected in 6 out of 10 fecal samples from wild Sumatran orangutans. The virulence genes mrkD and entB were detected in all (100%) of the isolates, whereas wabG was identified in 83.33% of the strains. Antibiotic susceptibility testing against K. pneumoniae revealed that three isolates were susceptible to streptomycin (S) and nalidixic acid (NA), while all six isolates were susceptible to chloramphenicol and ciprofloxacin. One isolate demonstrated intermediate resistance to NA, while the remaining two exhibited intermediate resistance to S. Six isolates were resistant to ampicillin, tetracycline, and erythromycin, indicating multidrug resis¬tance. Furthermore, antibiotic resistance genes were detected in the isolates with the following prevalence: blaTEM gene (six isolates; 100%), blaSHV (six isolates; 100%), blaCTX-M gene (four isolates; 66.67%), and tetA gene (four isolates; 66.67%). Conclusion: This study revealed the virulence and resistance profile of K. pneumoniae bacterium isolated from wild Sumatran orangutans, which is essential for formulating effective conservation and healthcare strategies.

Seroprevalence of Typhoid Fever and Its Associated Risk Factors Among Clinically Diagnosed Febrile Patients Visiting the Outpatient Department at Debark Hospital and Drug Susceptibility Patterns of Isolates

Typhoid fever is caused by the bacterium Salmonella Typhi, which poses major health problems in developing countries, including Ethiopia. However, there is limited information regarding typhoid fever, contributing factors with it, and its drug susceptibility pattern in the research area. The aim of this study was to determine the seroprevalence of typhoid fever and its associated risk factors among clinically diagnosed febrile patients at Debark Hospital and evaluate the drug susceptibility patterns of the isolates. A hospital‐based cross‐sectional study was conducted among 158 febrile patients from December 2022 to April 2023. Blood and stool samples were collected from each febrile patient. The Widal test was used to test Salmonella Typhi O and H antigens sera from blood, and bacteria were cultured from the stool. Gram staining and biochemical analyses were carried out for each isolate. Antibiotic susceptibility testing was performed for the isolates using Kirby–Bauer disk diffusion techniques. Descriptive statistics and logistic regression were used for analysis. In this study, the seroprevalence of typhoid fever among febrile patients at Debark Hospital was 22.8%. Gender (adjusted odds ratio (AOR): 95% confidence interval (CI): 0.02, 0.31, p ≤ 0.001), marital status (AOR: 95% CI: 2.63, 4.66, p ≤ 0.001), family size (AOR: 95% CI: 0.01, 0.31, p ≤ 0.001), residence (AOR: 95% CI: 0.09, 0.83, p = 0.021), practice of using toilet (AOR: 95% CI: 0.08, 0.086, p = 0.027), washing fruits and vegetables before eating (AOR: 95% CI: 0.12, 0.87, p = 0.025), and awareness of typhoid fever transmission and prevention (AOR: 95% CI: 0.12, 0.91, p = 0.032) are the associated risk factors for typhoid fever. Fifty percent of the isolates showed multidrug resistance to two or more antibiotics. It was suggested that improving personal hygiene, providing safe drinking water, and careful use of antibiotics could considerably reduce the prevalence of typhoid fever in the study area.

A Study of Bacteriological Profiles, Biofilm Production, and Antibiotic Susceptibility among Isolates of Chronic Suppurative Otitis Media at a Tertiary Care Hospital

ABSTRACT Background: Chronic suppurative otitis media (CSOM) is defined as chronic inflammation of the middle ear and mastoid mucosa. Biofilms act as non-selective physical barriers that obstruct antibiotic diffusion and hinder immune responses facilitating chronic bacterial infections and antibiotic resistance. Aims and Objectives: To study biofilm production among bacteria causing CSOM and their antibiotic resistance. Materials and Methods: This hospital-based descriptive type of observational study was done in the ENT department and Department of Microbiology. Ear discharge samples from 100 clinically diagnosed CSOM patients were collected and processed. Bacterial isolates were identified, and drug susceptibility testing was done using the Kirby–Bauer disc diffusion method. Biofilm production was detected by the microtiter plate method. Results: Among 100 patients included in the study, the most predominant isolate was Staphylococcus aureus (44.0%), followed by Pseudomonas aeruginosa (36.0%). Biofilm production was detected in 59.0% of isolates. Biofilm production was observed in 97.7% S. aureus isolates. Among Gram-negative isolates, biofilm production was observed in 40.0% of Klebsiella spp., 36.0% P. aeruginosa, and 22.2% E. coli. All isolates of S. aureus were susceptible to vancomycin, tigecycline, and linezolid (100%). Methicillin resistance was found in 25.6% S. aureus isolates. A susceptibility of 100% was observed with colistin and piperacillin-tazobactam among P. aeruginosa isolates. Conclusion: Ear discharge should be sent for culture and antibiotic susceptibility testing before initiation of empirical antibiotic treatment. Biofilm production is frequently associated with multi-drug resistance, so it should be tested routinely in laboratories. This will help in effective management of CSOM, prevent development of complications, and thereby prevent deafness.

Probiotic potential of riboflavin-overproducing Bacillus subtilis ACU-I163MR and ACU-I11MR, isolated from fermented African locust beans

Riboflavin (vitamin B2) is a water-soluble compound that plays an important role in multiple cellular functions. This study evaluates the probiotic potential of riboflavin-overproducing Bacillus subtilis strains isolated from fermented African locust beans. After strain improvement, B. subtilis ACU-I11MR and ACU-I163MR were selected due to their higher riboflavin production (0.01905±0.0005 mg l−1 to 0.0259±0.0077 mg l−1 and 0.0195±0.0054 mg l−1 to 0.0267±0.0013 mg l−1, respectively). Their safety was confirmed through haemolytic assay, antibiotic susceptibility tests and the absence of gelatinase and biogenic amine activity. Probiotic potential was assessed via in vitro assays including resistance to low pH, bile salts, phenol, temperature and NaCl; auto-aggregation; cell hydrophobicity; biofilm formation; antibacterial activity; and enzyme and exopolysaccharide production. Both strains were non-haemolytic and negative for gelatinase and biogenic amine activity. They showed significant viability at pH 2 (survival 85.05; 87.09%), 1% bile salts (survival 88.82; 87.64%) and 0.5% phenol (survival 48.80; 59.52%), respectively. ACU-I11MR was susceptible to 9 out of 12 antibiotics, while ACU-I163MR was 100% susceptible. The strains demonstrated strong cell surface adhesion and auto-aggregation and inhibited several pathogenic bacteria. They produce amylase, protease and exopolysaccharide and thrive under various temperature and NaCl conditions. B. subtilis ACU-I163MR, showing superior probiotic potential, could be a promising candidate for developing riboflavin-enriched Bacillus-fermented functional foods.

Prevalence of ESBL resistance genes and fecal carriage of multidrug-resistant Pseudomonas aeruginosa isolates from patients with chronic kidney disease at the Laquintinie Hospital in Douala, Littoral Region, Cameroon.

Pseudomonas aeruginosa is an opportunistic Gram-negative bacterium capable of causing severe infections in immunocompromised patients such as those suffering from chronic kidney disease (CKD). This study aimed to determine the resistance profile of Pseudomonas aeruginosa, and the prevalence of extended-spectrum β-lactamase (ESBL) resistance genes in patients with chronic kidney disease. The prevalence of Pseudomonas aeruginosa was investigated in 458 patients, including 197 CKD patients and 261 patients suffering from gastrointestinal infections. The study was conducted at Laquintinie Hospital in Douala from January 2022 to December 2023. Stool samples were used to isolate Pseudomonas aeruginosa on Cetrimide agar. Antibiotic susceptibility testing was carried out using the Kirby-Bauer diffusion method. extended-spectrum β-lactamase (ESBL) resistance genes were detected by the polymerase chain reaction (PCR) method. The prevalence of fecal carriage of Pseudomonas aeruginosa was 9.17 % (n = 42/458), including twenty-nine (69.05 %) in patients with chronic kidney disease and thirteen (30.95 %) in patients without chronic kidney disease. The Pseudomonas aeruginosa isolates had a high rate of resistance to ceftazidime (72.41 %) in patients with CKD compared to patients without CKD (69.23 %). All isolates had a high resistance to ticarcillin (93.10 % and 92.31 %). The prevalence of multidrug-resistant (MDR) Pseudomonas aeruginosa isolates was 73.81 %. The MDR Pseudomonas aeruginosa were higher (70.97 % vs. 29.03 %) in patients suffering from CKD compared to patients without CKD. Up to 85.71 % of the Pseudomonas aeruginosa isolates harbored at least one ESBL gene. The blaTEM type gene (66.67 %) was the most frequently detected gene, followed by blaCTX-M (61.90 %) and blaSHV (47.62 %). ESBL resistance genes were more common in CKD patients (72.22 %) compared to patients without CKD (27.78 %). These results demonstrate that antibiotics belonging to the carbapenem and aminoglycoside classes could be used for Pseudomonas aeruginosa infection. This highlights the importance of regular surveillance of multidrug resistance and extended-spectrum β-lactamase production for Pseudomonas aeruginosa infections in patients with chronic kidney disease.

A self-priming digital microfluidic chip for single-cell antibiotic susceptibility testing

A self-priming digital microfluidic chip for single-cell antibiotic susceptibility testing