Multidrug resistance among uropathogenic clonal group A E. Coli isolates from Pakistani women with uncomplicated urinary tract infections.

OBJECTIVE: Multi-drug resistance (MDR) has notably increased in community acquired uropathogens causing urinary tract infections (UTIs), predominantly Escherichia coli. Uropathogenic E. coli causes 80% of uncomplicated community acquired UTIs, particularly in pre-menopausal women. Considering this high prevalence and the potential to spread antimicrobial resistant genes, the current study was conducted to investigate the presence of clinically important strains of E. coli in Pakistani women having uncomplicated cystitis and pyelonephritis. Women belonging to low-income groups were exclusively included in the study. Seventy-four isolates from urine samples were processed, phylotyped, and screened for the presence of two Single Nucleotide Polymorphisms (SNPs) particularly associated with a clinically important clonal group A of E. coli (CgA) followed by antibiotic susceptibility testing and genome sequence analysis. RESULTS: Phylogroup B2 was most prevalent in patients and 44% of isolates were positive for the presence of CgA specific SNPs in Fumarate hydratase and DNA gyrase subunit B genes. Antibiotic susceptibility testing showed widespread resistance to trimethoprim-sulfamethoxazole and extended-spectrum beta-lactamase production. The infection analysis revealed the phylogroup B2 to be more pathogenic as compared to the other groups. The genome sequence of E. coli strain U17 revealed genes encoding virulence, multidrug resistance, and host colonization mechanisms. CONCLUSIONS: Our research findings not only validate the significant occurrence of multidrug-resistant clonal group A E. coli (CgA) in premenopausal Pakistani women suffering from cystitis and pyelonephritis but also reveal the presence of genes associated withvirulence, and drug efflux pumps. The detection of highly pathogenic, antimicrobial-resistant phylogroup B2 and CgA E. coli strains is likely to help in understanding the epidemiology of the pathogen and may ultimately help to reduce the impact of these strains on human health. Furthermore, the findings of this study will particularly help to reduce the prevalence of uncomplicated UTIs and the cost associated with their treatment in women belonging to low-income groups.

Preparation and in Vitro Evaluation of Tamoxifen-Conjugated, Eco-Friendly, Agar-Based Hybrid Magnetic Nanoparticles for Their Potential Use in Breast Cancer Treatment.

Tamoxifen is the drug of choice as hormonal therapy for hormone receptor-positive breast cancers and can reduce the risk of breast cancer recurrence. However, oral tamoxifen has a low bioavailability due to liver and intestinal metabolic passes. To overcome this problem and utilize the potential of this drug to its maximum, inorganic nanoparticle carriers have been exploited and tested to increase its bioavailability. Biocompatibility and unique magnetic properties make iron oxide nanoparticles an excellent choice as a drug delivery system. In this study, we developed and tested a "green synthesis" approach to synthesize iron nanoparticles from green tea extract and coated them with agar for longer stability (AG-INPs). Later, these hybrid nanoparticles were conjugated with tamoxifen (TMX). By using this approach, we synthesized stable agar-coated tamoxifen-conjugated iron nanoparticles (TMX-AG-INPs) and characterized them with Fourier-transform infrared (FTIR) spectroscopy. The average particle size of AG-INPs was 26.8 nm, while the average particle size of tamoxifen-loaded iron nanoparticles, TMX-AG-INPs, was 32.1 nm, as measured by transmission and scanning electron microscopy. The entrapment efficiency of TMX-AG-INPs obtained by the drug release profile was 88%, with a drug loading capacity of 43.5%. TMX-AG-INPs were significantly (p < 0.001) efficient in killing breast cancer cells when tested in vitro on the established breast cancer cell line MCF-7 by cell viability assay, indicating their potential to control cell proliferation.

A Cross-Sectional Study of Potential Antimicrobial Resistance and Ecology in Gastrointestinal and Oral Microbial Communities of Young Normoweight Pakistani Individuals.

Antimicrobial resistance (AMR) is a major global public health concern mainly affecting low- and middle-income countries (LMICs) due to lack of awareness, inadequate healthcare and sanitation infrastructure, and other environmental factors. In this study, we aimed to link microbial assembly and covariates (body mass index, smoking, and use of antibiotics) to gut microbiome structure and correlate the predictive antimicrobial gene prevalence (piARG) using PICRUSt2. We examined the gastrointestinal and oral microbial profiles of healthy adults in Pakistan through 16S rRNA gene sequencing with a focus on different ethnicities, antibiotic usage, drinking water type, smoking, and other demographic measures. We then utilised a suite of innovative statistical tools, driven by numerical ecology and machine learning, to address the above aims. We observed that drinking tap water was the main contributor to increased potential AMR signatures in the Pakistani cohort compared to other factors considered. Microbial niche breadth analysis highlighted an aberrant gut microbial signature of smokers with increased age. Moreover, covariates such as smoking and age impact the human microbial community structure in this Pakistani cohort.

Molecular investigation of possible relationships concerning bovine leukemia virus and breast cancer.

Worldwide, breast cancer has an eminent morbidity and mortality rate, as it is a neoplastic disease among females. The query of the prospective danger of bovine leukemia virus (BLV) to humans is an old but exceedingly topical focus of scientific debate. The objective of the current study was to determine the possible relationship between BLV and breast cancer. A total of 2710 formalin-fixed paraffin-embedded (FFPE) breast cancer samples were selected regardless of the age, ethnicity, or municipality origin of the subjects. The presence of BLV in human breast cancer was determined through nested PCR by amplifying tax and gag genes followed by partial sequencing. Homology was confirmed by using the online BLAST Tool. BLV genes were found to be positive in 26.8% (728/2710) of the samples from breast cancer patients and 10% (10/80) of the samples without cancer (negative control). The results indicated a correlation between the presence of the BLV gene and breast cancer (odds ratio = 0.3889; confidence interval = 1,18; p = 0.0029). The current findings suggest a possible link between BLV and human breast carcinoma. Therefore, screening cattle herds and milk products is suggested to reduce the viral transmission risk to humans.

Identification of novel bacterial urease inhibitors through molecular shape and structure based virtual screening approaches.

The enzyme urease is an essential colonizing factor of the notorious carcinogenic pathogen Helicobacter pylori (H. pylori), conferring acid resistance to the bacterium. Recently, antibiotic resistant strains have emerged globally with little to no alternative treatment available. In this study we propose novel urease inhibitors capable of controlling infection by H. pylori and other pathogenic bacteria. We employed hierarchal computational approaches to screen new urease inhibitors from commercial chemical databases followed by in vitro anti-urease assays. Initially ROCS shape-based screening was performed using o-chloro-hippurohydroxamic acid followed by molecular docking studies. Out of 1.83 million compounds, 1700 compounds were retrieved based on having a ROCS Tanimoto combo score in the range of values from 1.216 to 1.679. These compounds were further screened using molecular docking simulations and the 100 top ranked compounds were selected based on their Glide score. After structural classification of the top ranked compounds, eight compounds were selected and purchased for biological assays. The plausible binding modes of the most active compounds were also confirmed using molecular dynamics (MD) simulations. Compounds 1, 2 and 3 demonstrated good urease inhibitory properties (IC50 = 0.32, 0.68 and 0.42 µM) compared to the other compounds. Enzyme kinetic studies revealed that compounds 1 and 3 are competitive inhibitors while 2 is a mixed type inhibitor of the urease enzyme. Cell based urease inhibition and MTT assay showed that these compounds blocked H. pylori urease activity, affecting bacterial growth and acid tolerance.

Impact of Helicobacter pylori Virulence Factors on the Host Immune Response and Gastric Pathology.

Helicobacter pylori chronically infects nearly half the world's population, yet most of those infected remain asymptomatic throughout their lifetime. The outcome of infection-peptic ulcer disease or gastric cancer versus asymptomatic colonization-is a product of host genetics, environmental influences, and differences in bacterial virulence factors. Here, we review the current understanding of the cag pathogenicity island (cagPAI), the vacuolating cytotoxin (VacA), and a large family of outer membrane proteins (OMPs), which are among the best understood H. pylori virulence determinants that contribute to disease. Each of these virulence factors is characterized by allelic and phenotypic diversity that is apparent within and across individuals, as well as over time, and modulates inflammation. From the bacterial perspective, inflammation is probably a necessary evil because it promotes nutrient acquisition, but at the cost of reduction in bacterial load and therefore decreases the chance of transmission to a new host. The general picture that emerges is one of a chronic bacterial infection that is dependent on both inducing and carefully regulating the host inflammatory response. A better understanding of these regulatory mechanisms may have implications for the control of chronic inflammatory diseases that are increasingly common causes of human morbidity and mortality.

Encapsulation of Cardamom Essential Oil in Chitosan Nano-composites: In-vitro Efficacy on Antibiotic-Resistant Bacterial Pathogens and Cytotoxicity Studies.

Natural antimicrobial agents, particularly essential oils present an excellent alternative to current antibiotics due to their potent and broad-spectrum antimicrobial potential, unique mechanisms of action and low tendency to induce resistance. However their potential as a viable therapeutic alternative is greatly compromised due to their hydrophobic and volatile nature. The objective of the current research was to explore the anti-pathogenic potential of essential oils in a bio-based nano-carrier system. Six different essential oils were tested on multidrug-resistant bacterial pathogens. However, cardamom oil was selected for nano-encapsulation because of most potent anti-microbial activity. Cardamom oil loaded chitosan nano-particles were prepared by ionic gelation method with an encapsulation efficiency of more than 90% and size was estimated to be 50-100 nm. The Zeta potential was more than +50 mV that indicate a stable nano-dispersion. Cytotoxicity analysis indicated non haemolytic and non-cytotoxic behaviour on human corneal epithelial cells and HepG2 cell lines. Cardamom oil loaded chitosan nano-particles were found to exhibit excellent anti-microbial potential against extended spectrum ß lactamase producing Escherichia coli and methicillin resistant Staphylococcus aureus. Our results suggested safety and efficacy of cardamom oil loaded chitosan nano-particles for treating multidrug-resistant pathogens hence offer an effective alternative to current antibiotic therapy.

Helicobacter bilis gamma-glutamyltranspeptidase enhances inflammatory stress response via oxidative stress in colon epithelial cells.

Helicobacter bilis (H. bilis) infection is associated with cases of inflammatory bowel Disease, thyphlocolitis, hepatitis and cholecystitis. However, little is known about the bacterial virulence determinants or the molecular mechanisms involved. Recently, H. bilis γ-glutamyltranspeptidase (HBgGT) was shown to be a virulence factor decreasing host cell viability. Bacterial gGTs play a key role in synthesis and degradation of glutathione and enables the bacteria to utilize extracellular glutamine and glutathione as sources of glutamate. gGT-mediated loss of cell viability has so far been linked to DNA damage via oxidative stress, but the signaling cascades involved herein have not been described. In this study, we identified enhanced ROS production induced by HBgGT as a central factor involved in the activation of the oxidative stress response cascades, which finally activate CREB, AP-1 and NF-κB in H. bilis infected colon cancer cells. IL-8, an important pro-inflammatory chemokine that is a common downstream target of these transcription factors, was up-regulated upon H. bilis infection in an HBgGT dependent manner. Moreover, the induction of these signaling responses and inflammatory cytokine production in host cells could be linked to HBgGT-mediated glutamine deprivation. This study implicates for the first time HBgGT as an important regulator of signaling cascades regulating inflammation in H. bilis infected host epithelial cells that could be responsible for induction of inflammatory disorders by the bacterium.