Background and Objectives:In most laboratories, diagnosis of invasive aspergillosis (IA) depends on culture and histopathological examination of the involved tissues. Galactomannan (GM) is a polysaccharide antigen detectable in body fluids and represents an ideal assay for diagnosing invasive pulmonary aspergillosis. The main objectives of this study were to estimate the proportion of patients with proven, probable, and possible aspergillosis and to compare different diagnostic methods. Materials and Methods:Samples for culture, microscopy, and GM assay were taken from a total of 60 patients. The enrolled patients were divided into proven, probable, and possible aspergillosis groups. Results:Among the 60 patients in the study, there were 9 (15%) cases of probable aspergillosis. galactomannan assay was positive in 19 patients. Galactomannan assay had a higher sensitivity (66.7%) when compared to other fungal assays for diagnosing probable aspergillosis. Conclusion:Diagnosing invasive aspergillosis is extremely challenging, and biomarkers such as galactomannan assay are important for early diagnosis. Raising the cutoff to >1 can eliminate false-positive results associated with the GM assay.
Background and Objectives:Pseudomonas aeruginosa is a major nosocomial pathogen implicated in a wide range of hospital-acquired infections. Prompt detection and effective control of outbreaks caused by this organism are essential to minimize associated morbidity and mortality. Multiple strains may circulate within healthcare settings due to cross-transmission and persistent environmental reservoirs. Various molecular typing techniques have proven valuable in tracing the sources and transmission dynamics of outbreaks. The present study aimed to investigate the antibiogram profiles and enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) patterns of P. aeruginosa isolates recovered from non-respiratory clinical samples at a tertiary care hospital. Materials and Methods:A total of 157 clinical isolates phenotypically identified as P. aeruginosa were subjected to 16S rDNA-PCR for species confirmation. Among these, 150 isolates were confirmed as P. aeruginosa and subsequently analysed for antimicrobial susceptibility and molecular typing. Antimicrobial susceptibility testing was performed according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Molecular typing was carried out using ERIC-PCR. The resulting ERIC-PCR profiles were analysed using PAST version 4.03 software, and a dendrogram was constructed based on Dice similarity coefficients to assess genetic relatedness among isolates. Fisher's exact test was employed to determine associations between specific ERIC types and antibiotic resistance profiles. Results:Overall, 84% of the P. aeruginosa isolates were identified as multidrug-resistant (MDR), with widespread distribution among both inpatients and outpatients. The highest resistance rate was observed against ticarcillin/clavulanic acid, followed by ceftazidime, whereas no resistance was detected against colistin sulphate. A total of 54 distinct antibiotypes were identified, indicating substantial variability in antimicrobial resistance patterns. All 150 isolates were successfully typed using ERIC-PCR, which revealed 41 distinct clusters at an approximate similarity coefficient of 70%, demonstrating considerable genetic diversity among the isolates. Conclusion:This study enhances the understanding of the epidemiology of P. aeruginosa infections in healthcare settings by characterizing both the molecular diversity and antimicrobial resistance patterns of clinical isolates. The high prevalence of MDR strains and the circulation of multiple genetically diverse strains among hospital patients highlight the ongoing risk of transmission within the healthcare environment. Notably, the findings suggest a comparatively higher rate of cross-infection among burn patients. These observations underscore the urgent need for stringent infection prevention and control measures, particularly in burn units, to limit the spread of MDR P. aeruginosa.
Background and Objectives:Apart from spontaneous deamidation and isomerization of Asp/Asn residues, factors like ageing, oxidative stress, salinity and heat stress may also form abnormal isoaspartate (iso-Asp) residues compromising protein structure and function. PIMT helps in restoration of such iso-Asp residues. Pseudomonas aeruginosa is a gram-negative bacteria responsible for diseases with varied degree of pathogenesis in humans and animals. They can survive a wide range of temperature between 25°C to 42°C. The objective of the current study was to monitor the expression dynamics of PIMT in Pseudomonas aeruginosa upon exposure to different thermal stresses. Materials and Methods:The mid log phase culture of Pseudomonas aeruginosa was exposed to different temperature (37°C, 42°C, 56°C, 80°C) for an hour. The expression levels of PIMT were quantified by Western blot and ELISA. Results:A relatively higher level of expression of PIMT was observed in the group exposed to 42°C, 56°C, 80°C in comparison to 37°C. The expression of PIMT increased around 1.5 times in the group exposed to 42°C. Conclusion:A temperature dependent overexpression of PIMT was observed in Pseudomonas aeruginosa.
Background and Objectives: Malassezia spp. are lipid-dependent yeasts that colonize the skin of humans and animals and may act as opportunistic pathogens. This study aimed to characterize virulence-associated enzymes and their encoding genes in Malassezia spp. isolated from companion animals in northern Iran. Materials and Methods: A total of 300 clinical samples were collected from dogs and cats with dermatological or otic lesions. Species identification was performed using ITS-based PCR. Multiplex PCR was used to detect LIP, PLB, and KER genes. Enzymatic activity was evaluated using the Pz index. Statistical analyses included Chi-square and Spearman’s cor- relation tests (p < 0.05). Results: Eighty isolates were identified as Malassezia pachydermatis (87.5%) and Malassezia nana (12.5%). LIP, PLB, and KER genes were detected in 90%, 62.5%, and 50% of isolates, respectively. Strong enzymatic activity (Pz ≤ 0.69) was observed in 41.2% of isolates. A significant association was found between the presence of the PLB gene and phospholipase activity. Conclusion: Malassezia pachydermatis was the predominant species. Virulence-associated genes were associated with high- er enzymatic activity, supporting their role in pathogenic potential.
Background and Objectives:Flavoprotein monooxygenases (FPMOs) participate in various biological processes, including lignin degradation, natural product biosynthesis, and xenobiotic detoxification. This study aimed to investigate the heterologous expression of FPMO and assess its functional activity by examining its capacity to inactivate ampicillin. Materials and Methods:In silico analyses were performed to predict the enzyme's secondary and tertiary structures. The target gene was isolated from Pseudomonas aeruginosa, cloned into the pET-22b vector, and expressed in Escherichia coli BL21 (DE3). Heterologous protein expression was examined using SDS-PAGE, and the protein was purified through nickel-affinity chromatography. Enzymatic activity was measured by spectrophotometric monitoring of NADPH oxidation at 340 nm. Antibacterial activity was tested using an agar well-diffusion assay, measuring the inhibition zone diameter in E. coli treated with ampicillin. Results:Homology modeling indicated that the 3D structure of FPMO is very similar to cyclohexanone monooxygenase, supporting its classification as a Baeyer-Villiger monooxygenase (BVMO). Molecular docking proposed that ampicillin might be a substrate for FPMO, with predicted interactions at Ile141 and Val159. Enzymatic assays confirmed that FPMO catalyzes the oxidation of ampicillin, using FAD and NADPH as cofactors. Additionally, well-diffusion tests showed decreased ampicillin antibacterial activity after treatment. The enzyme's activity was further confirmed using an ampicillin inactivation assay. Conclusion:The heterologously expressed protein was functionally active. These findings suggest that the studied FPMO may play a role in antibiotic resistance in P. aeruginosa by oxidatively inactivating ampicillin.
