Characterization of biofilm formation and multi-drug resistance among Pseudomonas aeruginosa isolated from hospital wastewater in Dhaka, Bangladesh.

Hospital wastewater has been identified as a hotspot for the emergence and transmission of multidrug-resistant (MDR) pathogens that present a serious threat to public health. Therefore, we investigated the current status of antibiotic resistance as well as the phenotypic and genotypic basis of biofilm formation in Pseudomonas aeruginosa from hospital wastewater in Dhaka, Bangladesh. The disc diffusion method and the crystal violet assay were performed to characterize antimicrobial resistance and biofilm formation, respectively. Biofilm and integron-associated genes were amplified by the polymerase chain reaction. Isolates exhibited varying degrees of resistance to different antibiotics, in which >80% of isolates showed sensitivity to meropenem, amikacin, and gentamicin. The results indicated that 93.82% of isolates were MDR and 71 out of 76 MDR isolates showed biofilm formation activities. We observed the high prevalence of biofilm-related genes, in which algD+pelF+pslD+ (82.7%) was found to be the prevalent biofilm genotypic pattern. Sixteen isolates (19.75%) possessed class 1 integron (int1) genes. However, statistical analysis revealed no significant association between biofilm formation and multidrug resistance (χ2 = 0.35, P = 0.55). Taken together, hospital wastewater in Dhaka city may act as a reservoir for MDR and biofilm-forming P. aeruginosa, and therefore, the adequate treatment of wastewater is recommended to reduce the occurrence of outbreaks.

Genomic epidemiology of multidrug-resistant clinical Acinetobacter baumannii in Bangladesh.

The rising frequency of multidrug-resistant (MDR) Acinetobacter baumannii infections represents a significant public health challenge in Bangladesh. Genomic analysis of bacterial pathogens enhances surveillance and control efforts by providing insights into genetic diversity, antimicrobial resistance (AMR) profiles, and transmission dynamics. In this study, we conducted a comprehensive bioinformatic analysis of 82 whole-genome sequences (WGS) of A. baumannii from Bangladesh to understand their genomic epidemiological characteristics. WGS of the MDR and biofilm-forming A. baumannii strain S1C revealed the presence of 28 AMR genes, predicting its pathogenicity and classification within sequence type ST2. Multi-locus sequence typing (MLST) genotyping suggested heterogeneity in the distribution of clinical A. baumannii strains in Bangladesh, with a predominance of ST575. The resistome diversity was evident from the detection of 82 different AMR genes, with antibiotic inactivation being the most prevalent resistance mechanism. All strains were predicted to be multidrug-resistant. The observed virulence genes were associated with immune evasion, biofilm formation, adherence, nutrient acquisition, effector delivery, and other mechanisms. Mobile genetic elements carrying AMR genes were predicted in 68.29% (N = 56) of the genomes. The "open" state of the pan-genome and a high proportion of accessory genes highlighted the genome plasticity and diversity of A. baumannii in Bangladesh. Additionally, phylogenomic analysis indicated clustering of A. baumannii strains into three separate clades according to sequence type. In summary, our findings offer detailed insights into the genomic landscape of A. baumannii in Bangladesh, contributing to our understanding of its epidemiology and pathogenicity and informing strategies to combat this pathogen.

Phenotypic characterization and genomic analysis of a Salmonella phage L223 for biocontrol of Salmonella spp. in poultry.

The escalating incidence of foodborne salmonellosis poses a significant global threat to food safety and public health. As antibiotic resistance in Salmonella continues to rise, there is growing interest in bacteriophages as potential alternatives. In this study, we isolated, characterized, and evaluated the biocontrol efficacy of lytic phage L223 in chicken meat. Phage L223 demonstrated robust stability across a broad range of temperatures (20-70 °C) and pH levels (2-11) and exhibited a restricted host range targeting Salmonella spp., notably Salmonella Typhimurium and Salmonella Enteritidis. Characterization of L223 revealed a short latent period of 30 min and a substantial burst size of 515 PFU/cell. Genomic analysis classified L223 within the Caudoviricetes class, Guernseyvirinae subfamily and Jerseyvirus genus, with a dsDNA genome size of 44,321 bp and 47.9% GC content, featuring 72 coding sequences devoid of antimicrobial resistance, virulence factors, toxins, and tRNA genes. Application of L223 significantly (p < 0.005) reduced Salmonella Typhimurium ATCC 14,028 counts by 1.24, 2.17, and 1.55 log CFU/piece after 2, 4, and 6 h of incubation, respectively, in experimentally contaminated chicken breast samples. These findings highlight the potential of Salmonella phage L223 as a promising biocontrol agent for mitigating Salmonella contamination in food products, emphasizing its relevance for enhancing food safety protocols.

A comprehensive immunoinformatic analysis of chitin deacetylase's and MP88 for designing multi-epitope vaccines against Cryptococcus neoformans.

Cryptococcus neoformans causes life-threatening pneumonia and meningitis and is regarded as one of the leading killers of immunocompromised individuals. There is currently no vaccine against this pathogen. Recently, WHO placed it at the top among the critical priority groups in the fungal priority pathogens to accelerate the development of effective treatments. Numerous studies suggested the potential of subunit vaccines to overcome the challenges associated with live and inactivated whole-cell vaccines. Therefore, this study exploited integrated reverse vaccinology and immunoinformatic approach to construct and characterize multi-epitope vaccines targeting chitin deacetylases (Cda1, Cda2, Cda3) and MP88 of C. neoformans. 4 CTL, 8 HTL and 6 B cell epitopes were fused with different adjuvants and appropriate linkers to design two multi-epitope vaccines (VC1 and VC2). Both chimeric constructs were predicted to be highly antigenic, non-allergenic, non-toxic, soluble and had satisfactory physicochemical properties. Molecular docking and binding free energy calculation revealed strong binding interactions between vaccine constructs and human TLRs (TLR-2 and TLR-4). Classical MD Simulation and Normal mode analysis verified the stability of the vaccine-TLR complex in the biological environment. Codon adaptation, cloning and in silico expression suggested the efficient expression of recombinant vaccine proteins in E. coli. Both candidates also generated robust immune profiles comprising innate, adaptive and humoral immune responses. Taken together, experimental validations of our findings through extensive in vitro and in vivo testing might provide an effective vaccine for prophylactic control of C. neoformans.Communicated by Ramaswamy H. Sarma.

Use of Phages to Treat Antimicrobial-Resistant Salmonella Infections in Poultry.

Salmonellosis is one of the most common bacterial infections that impacts both human health and poultry production. Although antibiotics are usually recommended for treating Salmonella infections, their misuse results in the evolution and spread of multidrug-resistant (MDR) bacteria. To minimize the health and economic burdens associated with antimicrobial resistance, a novel antibacterial strategy that can obliterate pathogens without any adverse effects on humans and animals is urgently required. Therefore, therapeutic supplementation of phages has gained renewed attention because of their unique ability to lyse specific hosts, cost-effective production, environmentally-friendly properties, and other potential advantages over antibiotics. In addition, the safety and efficacy of phage therapy for controlling poultry-associated Salmonella have already been proven through experimental studies. Phages can be applied at every stage of poultry production, processing, and distribution through different modes of application. Despite having a few limitations, the optimized and regulated use of phage cocktails may prove to be an effective option to combat infections caused by MDR pathogens in the post-antibiotic era. This article mainly focuses on the occurrence of salmonellosis in poultry and its reduction with the aid of bacteriophages. We particularly discuss the prevalence of Salmonella infections in poultry and poultry products; review the trends in antibiotic resistance; and summarize the application, challenges, and prospects of phage therapy in the poultry industry.

Draft genome analysis of a multidrug-resistant Pseudomonas aeruginosa CMPL223 from hospital wastewater in Dhaka, Bangladesh.

OBJECTIVES: Multidrug-resistant (MDR) clones of Pseudomonas aeruginosa can cause complicated infections in human. The emergence of ST664 of MDR P. aeruginosa has been reported in Nepal, Iran and China. Here, we present the draft genome analysis of a MDR P. aeruginosa CMPL223 isolated from hospital wastewater in Bangladesh to understand antimicrobial resistance trends and pathogenicity. METHODS: Cetrimide agar was used for isolation of P. aeruginosa. Polymerase chain reaction (PCR) was carried out for detection of biofilm and integron related genes. Bacterial susceptibility to antibiotics was determined by disc diffusion method. Sequencing of whole genomic DNA was performed using Illumina iSeq 100 platform. Following quality checking of raw reads, assembly and annotation of sequences, a wide array of in silico tools were used for characterization of draft genome. RESULTS: The isolate was a strong biofilm former, carried integron 1 in chromosomal DNA, and was predicted to be pathogenic. It belongs to sequence type ST664 and O7 serogroup. The assembled genome contained 12 acquired antimicrobial resistant (AMR) genes, 2 prophage regions, 240 virulence genes, 71 drug targets, 142 insertion sequences, and 1 CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) array. The isolate was resistant to 21 out of 23 antibiotics, except colistin and imipenem. Comprehensive Antibiotic Resistance Database and ResFinder revealed that bacteria harboured blaOXA-50, blaOXA-796, blaPDC-374, fosA, tet(G), sul1, catB7, aph(3')-iib and ant(4')-IIb genes, conferring resistance to different classes of antibiotics. The results of in vitro characterization were consistent with the possible expression of detected antibiotic resistant genes through in silico analysis. CONCLUSION: Our data suggested the emergence of MDR P. aeruginosa ST664, which needs control measures for limiting its dissemination.

Assessment of foodborne transmission of Salmonella enteritidis in hens and eggs in Bangladesh.

OBJECTIVES: Salmonella is considered one of the leading causes of foodborne illnesses worldwide. Information about the transmission of pathogens to poultry and poultry products is necessary to implement control measures for reducing both human exposure and economic loss. The aim of this study was to analyze and evaluate the transmission characteristics of Salmonella enteritidis to laying-type hen flocks and their laid eggs. MATERIALS AND METHODS: For this purpose, 15 pairs of laying hens were used in which each pair consisted of one inoculated and one contact exposed hen. The eggs and cloacal swabs from these hens were subsequently analyzed. RESULTS: Of the 15 in-contact hens tested, 60% were found to be positive for S. enteritidis within 61 days postinoculation, of which 26.7% transmission occurred within the first 31 days postinoculation. Among the collected laid eggs tested, S. enteritidis was detected on 58% eggshells and 5.33% eggs internal contents. We also observed a 33.33% reduction in egg production from S. enteritidis-infected hens. In a cross-contamination study, we demonstrated that an experimentally inoculated container can act as a potential source of Salmonella spp. CONCLUSIONS: Our results will help establish effective monitoring programs to reduce the transmission of Salmonella spp. in poultry and poultry products.