Emerging Issues on Antibiotic-Resistant Bacteria Colonizing Plastic Waste in Aquatic Ecosystems.

Antibiotic-resistant bacteria (ARB) adhesion onto plastic substrates is a potential threat to environmental and human health. This current research investigates the prevalence of two relevant human pathogens, Staphylococcus spp. and Klebsiella spp., and their sophisticated equipment of antibiotic-resistant genes (ARGs), retrieved from plastic substrates submerged into an inland water body. The results of microbiological analysis on selective and chromogenic media revealed the presence of colonies with distinctive phenotypes, which were identified using biochemical and molecular methods. 16S rDNA sequencing and BLAST analysis confirmed the presence of Klebsiella spp., while in the case of Staphylococcus spp., 63.6% of strains were found to be members of Lysinibacillus spp., and the remaining 36.3% were identified as Exiguobacterium acetylicum. The Kirby-Bauer disc diffusion assay was performed to test the susceptibility of the isolates to nine commercially available antibiotics, while the genotypic resistant profile was determined for two genes of class 1 integrons and eighteen ARGs belonging to different classes of antibiotics. All isolated bacteria displayed a high prevalence of resistance against all tested antibiotics. These findings provide insights into the emerging risks linked to colonization by potential human opportunistic pathogens on plastic waste commonly found in aquatic ecosystems.

Effect of fenugreek and flaxseed polysaccharide-based edible coatings on the quality attributes and shelf life of apple fruit during storage.

The present study was designed to explore the potential of fenugreek and flaxseed polysaccharide-based edible coatings to enhance the postharvest storage life of apple fruit. The experimental plan involved the preparation of five different coating formulations, which were subsequently applied to the fruit. The coated fruit was then stored at a temperature of 25 ± 2°C for a duration of 35 days. The effects of these coatings on physicochemical and biochemical attributes (weight loss, firmness, acidity, pH, sugar content, antioxidant activity, microbial growth, and sensory properties) of coated and uncoated samples were evaluated at regular intervals: 0, 7, 14, 21, 28, and 35 days of storage. The experimental results revealed a significant difference (p ≤ .05) in the physicochemical parameters of uncoated and coated apple at different storage times. The coated apple fruits showed significantly (p ≤ .05) lower weight loss, pH, total sugars, total soluble solids, and maximum retention of ascorbic acid, firmness, acidity, and antioxidant contents, leading to enhanced organoleptic properties. The application of edible coatings extended the shelf-life of the apples by inhibiting microbiological spoilage without substantial impact on sensory and nutritional properties. Based on these results, it is concluded that the edible coating formulation labeled T 1 (containing 2.5 g fenugreek polysaccharide and 1.5 g flax polysaccharide) effectively preserved the valuable physicochemical and organoleptic characteristics of the apple fruit throughout the storage period.

Uropathogenic bacteria and deductive genomics towards antimicrobial resistance, virulence, and potential drug targets.

Urinary tract infections (UTIs) are among the most prevalent bacterial infections affecting people in inpatient and outpatient settings. The current study aimed to sequence the genome of uropathogenic Escherichia coli strain CUI-B1 resourced from a woman having uncomplicated cystitis and pyelonephritis. Followed by deductive genomics towards potential drug targets using E. coli strain CUI-B1, strain O25b: H4-ST131, Proteus mirabilis strain HI4320, Klebsiella pneumoniae strain 1721, and Staphylococcus saprophyticus strain ATCC 15305 uropathogenic strains. Comparative genome analysis revealed that genes related to the survival of E. coli, P. mirabilis, K. pneumoniae, and S. saprophyticus, such as genes of metal-requiring proteins, defense-associated genes, and genes associated with general physiology, were found to be highly conserved in the genomes including strain CUI-B1. However, the genes responsible for virulence and drug resistance, mainly those that are involved in bacterial secretion, fimbriae, adherence, and colonization, were found in various genomic regions and varied from one species to another or within the same species. Based on the genome sequence, virulence, and antimicrobial-resistant gene dataset, the subtractive proteomics approach revealed 22 proteins mapped to the pathogen's unique pathways and among them, entB, clbH, chuV, and ybtS were supposed to be potential drug targets and the single drug could be utilized for all above-mentioned strains. These results may provide the foundation for the optimal target for future discovery of drugs for E. coli-, P. mirabilis-, K. pneumoniae-, and S. saprophyticus-based infections and could be investigated further to employ in personalized drug development.

Identification, Genome Sequencing, and Characterizations of Helicobacter pylori Sourced from Pakistan.

The stomach's colonization by Helicobacter pylori (H. pylori) results in gastritis, ulcers, and stomach cancer. Frequently, pain is treated with medication, but resistant H. pylori infections are not. Therefore, it is important to find pharmacological targets and improved treatments for resistant H. pylori strains. The aim of the current study was sampling, identification, drug susceptibility testing following genome sequencing and comparative genome-wide analysis of selected H. pylori strains from Pakistan with three representative strains for virulence and drug-resistant characteristics. Based on culture, biochemistry, and molecular biology, 84 strains of H. pylori were identified, which made up 47% of the enrolled cases. Among all H. pylori strains, the highest resistance was reported for metronidazole with 82 H. pylori strains (98%), followed by clarithromycin with 62 resistant strains (74%). Among metronidazole-resistant strains, 38 strains (46%) were also resistant to clarithromycin, contributing 61% of clarithromycin resistant cases. Two strains, HPA1 and HPA2, isolated from 'gastritis' and 'gastric ulcer' patients, respectively, were further processed for WGS. The draft genome sequences of H. pylori strains HPA1 and HPA2 encode 1.66 Mbp and 1.67 Mbp genome size, 24 and 4 contiguous DNA sequences, and 1650 and 1625 coding sequences, respectively. Both the genomes showed greater than 90% similarity with the reference strain H. pylori ATCC 43504/PMSS1. The antibiotic-resistant genes were identified among all the strains with overall similarity above 95%, with minor differences in the sequence similarity. Using the virulent gene data obtained from the Virulence Factor Database, 75 to 85 virulent genes were identified in the five genome assemblies with various key genes such as cytolethal distending toxin (cdt), type IV secretion system, cag PAI, plasticity region, cell-motility- and flagellar-associated genes, neutrophil-activating protein (HP-NAP), T4SS effector cytotoxin-associated gene A (cagA), and urease-associated genes ureA and ureB, etc. Sequence similarity between the virulence factors found in this study and reference genes was at least 90%. In summary, the results of our study showed the relationship between clinical results and specific H. pylori strains' (HPA1 and HPA2) genetics such as antibiotic resistance and specific virulence factors. These findings provide valued understanding of the epidemiology of H. pylori-associated diseases. Moreover, identification and genomics analysis have provided insights into the epidemiology, genetic diversity, pathogenicity, and potential drug resistance genes of H. pylori strains, offering a foundation for developing more targeted and effective medical interventions, including anti-virulent medications.

Diverse Repertoire and Relationship of Exopolysaccharide Genes in Cold-Adapted Acinetobacter sp. CUI-P1 Revealed by Comparative Genome Analysis.

This study focused on the exploration of microbial communities inhabiting extreme cold environments, such as the Passu and Pisan glaciers of Pakistan, and their potential utilization in industrial applications. Among the 25 initially screened strains, five were found to be suitable candidates for exopolysaccharide (EPS) production, with strain CUI-P1 displaying the highest yield of 7230.5 mg/L compared to the other four strains. The purified EPS from CUI-P1 was tested for its ability to protect probiotic bacteria and E. coli expressing green fluorescence protein (HriGFP) against extreme cold temperatures, and it exhibited excellent cryoprotectant and emulsification activity, highlighting its potential use in the biotechnological industry. Furthermore, the genome of Acinetobacter sp., CUI-P1 comprised 199 contigs, with a genome size of 10,493,143bp and a G + C content of 42%, and showed 98.197% nucleotide identity to the type genome of Acinetobacter baumannii ATCC 17978. These findings offer promising avenues for the application of EPS as a cryoprotectant, an essential tool in modern biotechnology.

Detection of Morganella morganii bound to a plastic substrate in surface water.

OBJECTIVES: Around the globe, escalation in rare opportunistic microbial infections is alarming as they are heading steadily towards 'superbug' status. In aquatic ecosystems, plastic fosters multidrug-resistant pathogenic bacteria and plays a significant role in trafficking antibiotic-resistant genes. In this study, we focused on a multidrug-resistant bacterial strain isolated from microbial communities found on plastic substrates of a volcanic lake in central Italy. METHODS: Extended-spectrum beta-lactamase-producing strains were isolated from both raw water and plastic substrates for a comparative investigation using microbiological and molecular methods, and antibiotic susceptibility profiling was performed against a panel of ten antibiotics. RESULTS: Molecular identification and Basic Local Alignment Search Tool analysis confirmed an almost identical sequencing pattern of two isolated strains and their homology with Morganella morganii. Antibiotic susceptibility tests revealed their resistance to almost all tested antibiotics. Class 1 integron-associated gene (intI1) and seven antibiotic resistance genes were detected in both strains, confirming their superbug status. CONCLUSION: To our knowledge, this is the first study on the characterization of extended-spectrum beta-lactamase-producing M. morganii isolated from the biofilm of plastic substrates, depicting the potential toxicity of plastic in harbouring and dispersing virulent, multidrug-resistant, opportunistic human pathogens.

Optimization and cholesterol-lowering activity of exopolysaccharide from Lactiplantibacillus paraplantarum NCCP 962.

Exopolysaccharides (EPSs) are biological polymers with unique structural features have gained particular interest in the fields of food, chemistry and medicine, and food industry. EPS from the food-grade lactic acid bacteria (LAB) can be used as a natural food additives to commercial ones in the processing and development of functional foods and nutraceuticals. The current study was aimed to explore the EPS-producing LAB from the dahi; to optimize the fermentation conditions through Plackett-Burman (PB) and response surface methodology (RSM); and to study its physicochemical, rheological, functional attributes, and cholesterol-lowering activity. Lactiplantibacillus paraplantarum NCCP 962 was isolated among the 08 strains screened at the initial stage. The PB design screened out four independent factors that had a significant positive effect, i.e., lactose, yeast extract, CaCl2, and tryptone, while the remaining seven had a non-significant effect. The RSM exhibited lactose, yeast extract, and CaCl2, significantly contributing to EPS yield. The maximum EPS yield (0.910 g/L) was obtained at 6.57% lactose, 0.047% yeast extract, 0.59% CaCl2, and 1.37% tryptone. The R2 value above 97% explains the higher variability and depicts the model's validity. The resulted EPS was a heteropolysaccharide in nature with mannose, glucose, and galactose monosaccharides. FTIR spectrum reflected the presence of functional groups, i.e., O-H, C-H, C = O, C-O-H, and CH2. SEM revealed a porous and rough morphology of EPS, also found to be thermally stable and negligible weight loss, i.e., 14.0% at 257 °C and 35.4% at 292.9 °C was observed in the 1st and 2nd phases, respectively. Rheological attributes revealed that strain NCCP 962 had high viscosity by increasing the EPS concentration, low pH, and temperature with respectable water holding, oil capacities, foaming abilities, and stability. NCCP 962 EPS possessed up to 46.4% reduction in cholesterol concentration in the supernatant. Conclusively, these results suggested that strain NCCP 962 can be used in food processing applications and other medical fields. KEY POINTS: • The fermentation conditions affect EPS yield from L. paraplantarum and significantly increased yield to 0.910 g/L. • The EPS was heteropolysaccharide in nature and thermally stable with amorphous morphology. • Good cholesterol-lowering potential with the best rheological, emulsifying, and foaming capacities.