Determining the prevalence and genetic diversity of plasmid-mediated sulfonamide resistance in Escherichia coli from commercial broiler samples.

Sulfonamides are commonly used antibacterials in commercial poultry, contributing toward the development of multidrug-resistant (MDR) phenotypes among Escherichia coli and that has emerged as global concern. The current study aimed to assess the sulfonamide resistance among isolated E. coli strains among commercial broilers. The bacterial strains were identified from fecal samples (n = 100) using selective media, followed by initial identification based on biochemical profiles. The susceptibility was determined by measuring the minimum inhibitory concentration (MIC) against sulfamethoxazole. The study also evaluated mobile genetic elements (MGEs), the mediators of antibiotic resistance, by amplification of plasmid DNA using specific primer PCR. Additionally, the isolates were subjected to multilocus sequence typing (MLST) analysis to investigate the genetic diversity among E. coli carrying sulfonamide resistance genes. The results revealed that 58% (58/100) E. coli strains were resistant to sulfonamides, with 36.20% (21/58) of the strains exhibiting an MIC breakpoint ≥512 µg/mL. PCR analysis showed that 42.85% (9/21) of the strains harbored the sul-1 gene, while 38.09% (8/21) carried the sul-2 gene, and 19.04% (4/21) had both genes. No isolate showed the presence of the sul-3 gene. Furthermore, class 1 and class 2 integrons were identified among 80.95% (17/21) and 19.04% (4/21) of the strains, respectively. MLST analysis confirmed that the strains belonged to sequence types (STs) including ST1638, ST155, ST48, ST350, ST23, ST156, and ST746. These findings underscore the diversity among E. coli strains in commercial poultry, which poses a significant risk.

Fisetin Attenuates Arsenic-Induced Hepatic Damage by Improving Biochemical, Inflammatory, Apoptotic, and Histological Profile: In Vivo and In Silico Approach.

Arsenic (As) is a toxic metalloid and human carcinogen that may cause hepatotoxicity. Fisetin (3, 3', 4', 7-tetrahydroxyflavone) is a phytoflavonoid, which shows diverse therapeutic activities. This study aimed to examine the remedial potential of fisetin against As-instigated hepatotoxicity in adult male rats. To accomplish this aim, albino rats (N = 48) were evenly classified into 4 groups: control group, As (10 mg/kg) group, fisetin (2.5 mg/kg) + As (10 mg/kg) group, and fisetin (2.5 mg/kg) group. After one month of treatment, biochemical assay, total protein content (TPC), hepatic serum enzymes, inflammatory as well as pro- or anti-apoptotic markers, and histopathological profile of hepatic tissues were estimated. As administration disordered the biochemical profile by decreasing activities of antioxidant enzymes i.e., catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GSR), and glutathione (GSH) content while escalating the levels of reactive oxygen species (ROS), and thiobarbituric acid reactive substances (TBARS). TPC was also considerably reduced after exposure to As. Furthermore, As markedly raised the levels of liver serum enzymes such as aspartate transaminase (AST), alkaline phosphatase (ALP), and alanine transaminase (ALT) as well as the levels of inflammatory markers, i.e., nuclear factor- κB (NF-κB), tumor necrosis- α (TNF-α), interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and cyclo-oxygenase-2 (COX-2) activity. Besides, it lowered the level of antiapoptotic markers (Bcl-2) and upregulated the levels of proapoptotic markers (Bax, Caspase-3, and Caspase-9). Additionally, As exposure led to histopathological damage in hepatic tissues. However, fisetin administration remarkably alleviated all the depicted hepatic damages. For further verification, the screening of several dock complexes was performed by using the GOLD 5.3.0 version. Based on docking fitness and GOLD score, the ranking order of receptor proteins with fisetin compound is superoxide dismutase, interleukin, aspartate aminotransferase, alkaline phosphatase, TNF-alpha, alanine transaminase, cyclo-oxygenase 2, antiapoptotic, and glutathione reductase. Out of these three receptor proteins superoxide dismutase, interleukin, and aspartate aminotransferase showed the best interaction with the fisetin compound. In vivo and in silico outcomes of the current study demonstrated that fisetin could potentially ameliorate As-instigated hepatotoxicity.

Genetic Diversity of Escherichia coli Coharboring mcr-1 and Extended Spectrum Beta-Lactamases from Poultry.

Background: The emergence of resistance to beta-lactam agents in poultry results in multidrug-resistant (MDR) phenotypes in Escherichia coli isolates from poultry birds. The appearance of mobile colistin resistance (mcr) genes in the poultry sector has further worsened the situation. Therefore, the current study is aimed at investigating the molecular epidemiology of mcr harboring colistin-resistant E. coli among poultry. Methods: The isolation and identification of colistin-resistant E. coli (CR-Ec) were done from the broiler's fecal samples through culturing using selective media supplemented with colistin sulfate (4 µg/ml). The antibiogram studies of the isolates were performed using the disc diffusion method and broth microdilution method as per CLSI guidelines. The screening for the genes conferring resistance to colistin as well as beta-lactam agents was performed by PCR. The genetic diversity of mcr-positive strains was assessed by multilocus sequencing typing (MLST). Results: Out of 500 fecal samples, 7% (35/500) were found positive for the presence of colistin-resistant E. coli (CR-Ec). Among the CR-Ec isolates, 74.28% (26/35) were detected as ESBL producers carrying the blaCTX-M-1 gene in 15/35 (42.85%) isolates and blaCTX-M-15 and blaTEM genes in 21/35 (60%) and 35/35 (100%) isolates, respectively. E. coli isolates were found positive for the presence of mcr-1, although none of the isolates exhibited the mcr-2 or mcr-3 genes. The MLST of CR-Ec has shown the ST1035 as the most prevalent genotype, while 82.85% (29/35) of CR-Ec strains belonged to clonal complex (CC) 131 comprising ST1035, ST131, ST1215, ST1650, and ST2279. Conclusions: The findings suggest a continuous monitoring system in veterinary and clinical settings to avoid unnecessary antibiotics. Further studies are needed at the national level to help control the increasing resistance among Enterobacterales in poultry settings.

The occurrence of multidrug-resistant Mycobacterium tuberculosis from patients of pulmonary tuberculosis.

INTRODUCTION: Multidrug-resistant Mycobacterium tuberculosis (MDR-TB) is one of the leading causes of death in the world. The resource constraints make it difficult to diagnose and monitor the cases of MDR-TB. GeneXpert is a recognized tool used to diagnose the patients of pulmonary tuberculosis in clinical settings across the globe. METHODOLOGY: The present one-year cross-sectional study was conducted to estimate the occurrence of MDR-TB in patients with pulmonary TB. A total of 1000 patients suspected of pulmonary tuberculosis were included in this study. A random convenient sampling technique was done to collect the sputum samples (twice) from the patients. Samples were processed for the detection of Mycobacterium tuberculosis using conventional detection methods like the Ziehl Nelson staining method and fluorescent microscopy. Additionally, Cepheid GeneXpert was used for molecular detection of MDR-TB in smear-positive samples of pulmonary tuberculosis by amplifying the rifampicin resistance determining region (RRDR; rpoB gene). All the tests were performed in the biosafety level III lab of District Headquarters Hospital Nankana Sahib. RESULTS: It was observed that 103 (10.3%) individuals were diagnosed as positive for tuberculosis among 1000 patients. Among these 103 TB positive cases, there were 11 (10.7%) patients diagnosed with rifampicin resistance gene (RR-Gene) of Mycobacterium tuberculosis. CONCLUSIONS: Overall findings of the study showed that MDR-TB is prevalent in pulmonary TB patients and GeneXpert is the most sensitive technique for early diagnosis of the disease, which may be very helpful in the treatment and control of this public health menace in low and middle-income countries.

Identification of Ascorbic Acid and Gallic Acid as Novel Inhibitors of Secreted Frizzled-Related Protein for the Treatment of Obesity-Induced Type 2 Diabetes.

Type 2 diabetes mellitus (T2D) has been reported as major public health issue rising at an alarming rate worldwide, and obesity is the leading risk factor for the development of T2D. Secreted frizzled-related protein 4 (SFRP4) released with inflammatory mediators from adipose tissues constrains the exocytosis of insulin containing granules from the pancreatic islets that leads towards the development to T2D. The significant overexpression of SFRP4 in diabetic patients and its involvement in islet dysfunction suggest its critical role in the development of diabetes. Thus, this study was designed to explore the potential of ascorbic acid (AA) and gallic acid (GA) against SFRP4 for the treatment of diabetes. Molecular docking approach was used for the prediction of binding interactions of AA and GA at the active pocket of SFRP4. Docking analysis indicated strong binding interactions of AA and GA to the amino acid residues at the active site of SFRP4. A significant reduction in the level of SFRP4 was observed in transfected cells treated with AA and GA. For the evaluation of the cytotoxicity of AA and GA against HepG2 cells, MTT assay was performed. The results of MTT assay demonstrated that AA and GA are non-cytotoxic towards HepG2 cells at concentration of 15 µM. The oral administration of AA and GA to diet-induced obese mice caused significant reduction in body weight, blood glucose level, and SFRP4 expression. The results of this study suggest that AA and GA have potential for the treatment of obesity-induced T2D.

Distribution and antimicrobial susceptibility profile of bacterial and fungal pathogens isolated from burn wounds in hospitalized patients.

OBJECTIVE: Current study was designed to isolate the pathogens from burn wounds and determine the antibiogram of these isolates. METHODS: A total of 85 samples were collected from burn patients with the history of different weeks of hospitalization in various public and private hospitals of Faisalabad during September 2017-July 2019 and shifted to Department of Microbiology, Government College University, Faisalabad for further processing. Isolation and identification of the pathogens was done through conventional microbiological procedures. Disc diffusion method was used for the determination of antibacterial and antifungal activity. RESULTS: A total of 40(91%) samples were found positive for the presence of bacterial or fungal pathogens. Commonly isolated pathogens were Staphylococcus aureus 15 (21.4%), Pseudomonas aeruginosa 15 (21.4%), Bacillus subtilis 11(15.7%), Escherichia coli 10(14.2%), Candida albicans 8(11.4%), Aspergillus flavus 6(8.5%) and Salmonella Typhi 5(7.1%). Highest resistance was found against S. aureus and P. aeruginosa. Cefotaxime was the least effective antibiotic, while Gentamicin and Amphotericin-B were the mosteffective antimicrobial drugs against bacterial and fungal pathogens, respectively. CONCLUSIONS: Taking together it was concluded that most isolated pathogen was S. aureus and P. aeruginosa followed by B. subtilis, E. coli, C. albicans, A. flavus and S. typhi from burn wound in hospitalized patients. Anti-biogram studies showed S. aureus and P. aeruginosa were the most resistant pathogens whereas S. typhi, C. albicans and A. flavus were susceptible to various commonly used antibiotics. Cefotaxime was the least effective antibiotic, while Gentamicin and Amphotericin-B were the most effective antimicrobial drugs against bacterial and fungal pathogens, respectively. It is suggested that alternate anti-microbial agents should be investigated to control the infections.

First Case Report on Quantification of Antimicrobial Use in Corporate Dairy Farms in Pakistan.

Intensive livestock farming has become indispensable to meet the rapidly increasing demand for animal-based nutrition in low- and middle-income countries (LMICs) where antimicrobials are frequently used for treatment and prophylactic or metaphylactic purposes. However, very little is known about the trends of antimicrobial use (AMU) in dairy animals in LMICs. The objective of this study was to quantify AMU in two large commercial dairy farms in Pakistan. A retrospective study was conducted at two large corporate commercial dairy farms located in Punjab province for the year 2018. AMU was calculated using three metrics: active ingredient (AI; kg) and milligrams per population unit (mg/PU; mg/kg), which quantifies the amount of AI used, and antimicrobial treatment incidence (ATI; DDDA/1,000 cow-days), which estimates the per-day number of treatments to 1,000 cows. Total on-farm AMU was found to be 138.34 kg, 65.88 mg/kg, and 47.71 DDDA/1,000 cow-days. Measured in ATI, aminoglycosides (11.05 DDDA/1,000 cow-days), penicillins (8.29 DDDA/1,000 cow-days), and tetracyclines (8.1 DDDA/1,000 cow-days) were the most frequently used antimicrobial classes. A total of 42.46% of all the antimicrobials used belonged to the critically important antimicrobials for human medicine as defined by the World Health Organization. Considerably high AMU was found compared to other farm-level studies across the world. This was the first study to quantify AMU in the dairy industry in Pakistan. Our results showed that corporate commercial dairy management practices are associated with increased antimicrobial consumption and highlight the need for antimicrobial stewardship programs to encourage prudent use of antimicrobials in commercial dairy.

Biosynthesis of ZnO Nanoparticles Using Bacillus Subtilis: Characterization and Nutritive Significance for Promoting Plant Growth in Zea mays L.

Nano-fertilizer(s), an emerging field of agriculture, is alternate option for enhancement of plant growth replacing the synthetic fertilizers. Zinc oxide nanoparticles (ZnO NPs) can be used as the zinc source for plants. The present investigation was carried out to assess the role of ZnO NPs in growth promotion of maize plants. Biosynthesized ZnO NPs (using Bacillus sp) were characterized using Scanning Electron Microscope (SEM), Transmission Electron Microscope (TEM), X-ray diffraction (XRD) and Zeta potential. Different concentrations of ZnO NPs (2, 4, 8, 16 mg/L) were explored in pot culture experiment. Size of ZnO NPs ranged between 16 and 20 nm. A significant increase in growth parameters like shoot length (61.7%), root length (56.9%) and significantly higher level of protein was observed in the treated plants. The overall pattern for growth biomarkers including the protein contents was maximum at 8 mg/L of ZnO NPs. It was observed that application of biosynthesized ZnO NPs has improved majority of growth biomarkers including plant growth parameters, protein contents and leaf area. Therefore, biosynthesized ZnO NPs could be considered as an alternate source of nutrient in Zn deficient soils for promoting the modern agriculture.

Insecticidal and Genotoxic effects of some indigenous plant extracts in Culex quinquefasciatus Say Mosquitoes.

Five different weed plants viz. Convulvulus arvensis, Chenopodium murale, Tribulus terrestris, Trianthema portulacastrum, and Achyranthes aspera were investigated for their entomocidal and genotoxic effects against Culex quinquefasciatus mosquitoes. High mortality was observed at 72 hours in a dose dependent manner. Among all the tested plants, A. aspera was found highly significant which showed 100% mortality at 250 ppm after 72 hours with LC50 of 87.46, 39.08 and 9.22 ppm at 24, 48, respectively. In combination with Bacillus thuringiensis israelensis (Bti); A. aspera also caused 100% mortality at 250 ppm concentration after 72 hours (LC50 8.29 ppm). Phytochemical analysis of all the tested weed plants showed the presence of flavonoids, saponins, tannins, steroids, cardiac glycosides, alkaloids, anthrequinones and terpenoids. Random Amplification of Polymorphic DNA-Polymerase chain reaction (RAPD-PCR) and comet assay were performed to assess the genotoxic effect of A. aspera but no change in DNA profile was observed. Furthermore, FTIR showed the presence of phenolic compounds in A. aspera extract. It is suggested that certain phenolic compounds such as flavonoids modulate the enzymatic activity and, hence, cause the death of larvae of Cx. quinquefasciatus. Altogether, current study would serve as an initial step towards replacement of synthetic insecticides to plant-microbe based biopesticide against Culex mosquitoes in future.

Association of biofilm formation with multi drug resistance in clinical isolates of Pseudomonas aeruginosa.

Pseudomonas aeruginosa is considered as foremost cause of hospital acquired infections due to its innate and plasmid mediated resistance to multiple antibiotics making it a multi drug resistant (MDR) pathogen. Biofilm formation is a pathogenic mechanism harbored by this pathogen which further elevates its resistance to antibiotics and host defense system. The aim of the present study was to evaluate the biofilm forming potential and distribution of pslA gene in multi drug resistant Pseudomonas aeruginosa isolates obtained from different clinical samples. A total of 200 different clinical samples were collected after obtaining written consent from the patients. The samples were subjected to isolation and identification of P. aeruginosa by standard microbiological procedures. Confirmation of isolates was done by polymerase chain reaction targeting oprL gene. Kirby Bauer method was performed for detection of MDR isolates. Congo red agar (CRA) test and Microtiter plate assay (MPA) for observing the biofilm forming ability and amplification of pslA gene was also performed on MDR isolates. The results showed that from 200 samples 52 (26 %) were P. aeruginosa and among them 20 (38.46 %) were MDR isolates. The CRA showed 23 (44.23 %) while MPA detected 49 (94.23 %) isolates as biofilm producers while all the MDR isolates showed biofilm formation by MPA method. The pslA gene was detected in all biofilm forming isolates while 90 % in MDR P. aeruginosa. It was concluded that biofilm forming P. aeruginosa are more resistant to tested antibiotics and biofilm formation is strongly associated with presence of pslA gene.

Insecticidal, biological and biochemical response of Musca domestica (Diptera: Muscidae) to some indigenous weed plant extracts.

In the current study; insecticidal, growth regulation, oviposition deterrence and repellency of petroleum ether extracts of Azadirachta indica, Penganum harmala, Datura stramonium, Tribulus terrestris and Chenopodium murale against 2nd instar larvae of housefly was investigated. Five different concentrations (5%, 10%, 15%, 20% and 25%) were used through larval feeding and the mortality data was recorded after 24, 48 and 72 hrs. Highest mortality was induced by P. harmala (63.87%) followed by D. stramonium (62.78%), A. indica (53.84%), T. terrestris (41.86%) and C. murale (4.09%) after 72 h at 25% concentration, respectively. Increased mortality was observed with increased time duration and concentration. Longest larval duration (9.33 ±â€¯0.33 days) and pupal duration (7.33 ±â€¯0.33 days) days) was recorded in larvae treated with 25% concentration of P. harmala which also caused a decrease in the activity of AChE, ACP, AKP, α-Carboxyl, and ß-Carboxyl enzymes. However, at 25% concentration, C. murale showed highest oviposition deterrence activity (81.88%) followed by D. stramonium (79.58%). In comet assay test, at highest concentration (25%) the mean comet tail lengths represented by Penganum harmala, Datura stramonium and Azadirachta indica (Reference plant) were 10.20 ±â€¯0.49, 9.20 ±â€¯0.37 and 7.80 ±â€¯0.49 µm while percent DNA damage was 10.56 ±â€¯0.77, 10.67 ±â€¯1.62 and 8.11 ±â€¯0.85% respectively compared to controls cells. Phytochemical analysis indicated the presence of flavonoids, steroids, saponins, cardiac glycosides, tannins, alkaloids, terpenoids and anthraquinones. Fourier Transform Infrared spectroscopy (FTIR) analysis revealed the presence of phenolic flavonoids, saponins, tannins as major functional groups. Further studies are needed to explore and thus, to incorporate weed plant extracts for the management of house flies.

Isolation and molecular characterization of the indigenous Staphylococcus aureus strain K1 with the ability to reduce hexavalent chromium for its application in bioremediation of metal-contaminated sites.

BACKGROUND: Urbanization and industrialization are the main anthropogenic activities that are adding toxic heavy metals to the environment. Among these, chromium (in hexavalent: Cr+6 and/or trivalent Cr+3) is being released abundantly in wastewater due to its uses in different industrial processes. It becomes highly mutagenic and carcinogenic once it enters the cell through sulfate uptake pathways after interacting with cellular proteins and nucleic acids. However, Cr+6 can be bio-converted into more stable, less toxic and insoluble trivalent chromium using microbes. Hence in this study, we have made efforts to utilize chromium tolerant bacteria for bio-reduction of Cr+6 to Cr+3. METHODS: Bacterial isolate, K1, from metal contaminated industrial effluent from Kala Shah Kaku-Lahore Pakistan, which tolerated up to 22 mM of Cr6+ was evaluated for chromate reduction. It was further characterized biochemically and molecularly by VITEK®2 system and 16S rRNA gene sequencing respectively. Other factors affecting the reduction of chromium such as initial chromate ion concentration, pH, temperature, contact-time were also investigated. The role of cellular surface in sorption of Cr6+ ion was analyzed by FTIR spectroscopy. RESULTS: Both biochemical and phylogenetic analyses confirmed that strain K1 was Staphylococcusaureus that could reduce 99% of Cr6+ in 24 hours at 35 °C (pH = 8.0; initial Cr6+ concentration = 100 mg/L). FTIR results assumed that carboxyl, amino and phosphate groups of cell wall were involved in complexation with chromium. Our results suggested that Staphylococcusaureus K1 could be a promising gram-positive bacterium that might be utilized to remove chromium from metal polluted environments.

Molecular identification of blaCTX-M and blaTEM genes among multi-drug resistant Enteropathogenic Escherichia coli isolated from children.

In this cross sectional study (June 2016 to June 2017), we studied the isolation and molecular characterization of multi-drug resistant Escherichia coli (MDR-E. coli) from children suffering from diarrhea. For this purpose, a total of 100 fecal samples were collected with the consent of the parents/ guardians on a prescribed form. The bacterial isolation was done by employing conventional and standard microbiological procedures. Subsequently, all the isolates were identified on the basis of biochemical tests and were further characterized by amplification of 16S rRNA gene followed by di-deoxy sequencing of the amplified product. Afterwards, the isolates were subjected to antimicrobial susceptibility profiling using Kirby Bauer disc diffusion method. A total of 87 E. coli isolates were identified in the current study and majority of the isolates were found sensitive to all or few antimicrobials. However, 14 E. coli isolates were found resistance to multiple drugs including amoxicillin-clavulanic acid, ciprofloxacin, gentamicin, cefoperazone and ofloxacin, hence termed as MDR-E. coli. All of the 14 isolates were further analyzed for the identification of blaCTX-M and blaTEM genes through PCR using specific primers. This resistant was found to be associated with the presence of plasmid encoded beta lactamases genes including blaCTX-M (13/14 E. coli isolates) and blaTEM (9/14 E. coli isolates). Altogether, it was found that ESBLs harboring E. coli is potential source of diarrhea among pediatric diarrheal patients. Therefore, molecular identification and characterization of bacterial pathogens along with antimicrobial susceptibility are critical to understand MDR- E. coli infections.

Secreted frizzled-related protein 4 and its implication in obesity and type-2 diabetes.

Secreted frizzled-related protein 4 (SFRP4) is a member of secreted protein family with sequence similarity to frizzled receptors of wingless-related integration site (Wnt) signaling pathways. These proteins control diverse functions from embryonic development to adults in many organisms including humans. Initially, SFRPs were recognized as antagonists of Wnt signaling and supposed to interact with Wnts. Further research demonstrated their interactions to frizzled receptors and a functional diversity was related to these proteins, Wnt signaling potentiation in addition to modulation. SFRP4 is the largest member of SFRP family and is implicated in many diseases including obesity, type 2 diabetes (T2D), and cancer. SFRP4 acts as a biomarker for T2D and was expressed several years before clinical diagnosis of disease. This review mainly focusses on the role of SFRP4 in obesity and how it can lead to ß-cell failure and ultimately to T2D. The role of SFRP4 in adipose tissues causing increased production of adipokines lead to the oxidative stress in pancreas that particularly have low amount of antioxidant enzymes in pancreatic ß-cells leading to failure in exocytosis of insulin containing granules causing T2D. Obesity-induced inflammation is a principal factor in pathogenesis of insulin resistance as well as metabolic syndrome. Pro-inflammatory cytokines have potential to cause insulin resistance in skeletal muscles, adipose tissue, and liver via inhibition of insulin signal transduction. Secretion of SFRP4 is mediated by interleukin 1-ß (IL1-ß). This review highlights the molecular mechanisms by which SFRP4 leads to T2D. Understanding of molecular mechanism and targeting SFRP4 could help to eradicate or reduce chances of developing T2D.

Cross-sectional epidemiological investigations of Giardia lamblia in children in Pakistan.

BACKGROUND: The prevalence of Giardia lamblia in Pakistani children is currently unknown. The aim here was to evaluate the prevalence and risk factors of Giardia lamblia in children exhibiting diarrhea. DESIGN AND SETTING: Cross-sectional study at different district healthcare hospitals in Pakistan. METHODS: A total of 800 samples were collected from children aged 0-10 years. Information regarding personal data, demographic data and supposed risk factors was collected through a structured questionnaire. Giardia lamblia was detected through direct microscopy and antigens through the enzyme-linked immunosorbent assay (ELISA). RESULTS: The prevalence of Giardia lamblia was 2.75% through direct microscopy and inflated to 9.5% through ELISA. The demographic factors positively associated with occurrences of giardiasis were age (P = 0.035; odds ratio, OR = 1.96; 95% confidence interval, CI = 1.094-3.533), mother's educational level (P = 0.031; OR = 2.67; 95% CI = 1.186-6.045) and father's educational level (P = 0.004; OR = 3.56; 95% CI = 1.612-7.899). Similarly, among the supposed risk factors, rural residency (P = 0.032; OR = 1.76; 95% CI = 1.098- 2.851), absence of proper sewerage system (P = 0.000; OR = 6.60; 95% CI = 4.029-10.841) and unavailability of safe drinking water (P = 0.000; OR = 4.08; 95% CI = 2.207-7.547) were the factors strongly connected with giardiasis. Abdominal discomfort was a prominent clinical sign with 46% frequency. CONCLUSION: Various risk factors were associated with occurrences of Giardia, thus emphasizing the importance of parents' education, safe drinking water and proper sewerage systems for Pakistani children's health.

Effect of Lactobacillus casei on serum interleukins following enteropathogenic E. coli infection in experimental rabbits.

In the present study we investigated the pro-inflammatory and anti-inflammatory effect of Lactobacillus casei following infection with multi-drug resistant enteropathogenic Escherichia coli infection in experimental rabbits. For this purpose, 40 adult rabbits were divided into different groups and were infected with multi-drug resistant E. coli AZ1 strain except the control groups. The rabbits were orally administered with L. casei SABA6 strain in two different ways i.e. pre-treatment and post-treatment and both were continued for 7 days. The rabbits were sacrificed sequentially at 0, 4, 7 and 10 days post infection (dpi). Serum and intestinal tissue samples were collected from each rabbit. Intestinal tissue samples were subjected to histopathological examination that showed microscopic lesions at 4 and 7 dpi among infected group. The serum samples were processed for determination of Interleukin-6 (IL-6, pro-inflammatory) and Interleukin-10 (IL-10, anti-inflammatory) using ELISA. It was found that oral administration of L. casei SABA6 reduces the eruption of intestinal epithelial cells and reduces the incidence of diarrhea. Further, L. casei SABA6 also resulted in immuno modulation by significant increase in concentration of IL-6 and IL-10 particularly at 4 and 7 dpi and protects against E. coli AZ1 infection. Altogether, it was concluded that increased IL-6 and IL-10 levels were responsible for protection against EPEC infections. The sequential sacrifice of experimental animals could be adopted for future studies to find out pathogenesis and virulence mechanism of EPEC infections along with protective efficacy of different probiotics.

Cross-sectional epidemiological investigations of Giardia lamblia in children in Pakistan

ABSTRACT BACKGROUND: The prevalence of Giardia lamblia in Pakistani children is currently unknown. The aim here was to evaluate the prevalence and risk factors of Giardia lamblia in children exhibiting diarrhea. DESIGN AND SETTING: Cross-sectional study at different district healthcare hospitals in Pakistan. METHODS: A total of 800 samples were collected from children aged 0-10 years. Information regarding personal data, demographic data and supposed risk factors was collected through a structured questionnaire. Giardia lamblia was detected through direct microscopy and antigens through the enzyme-linked immunosorbent assay (ELISA). RESULTS: The prevalence of Giardia lamblia was 2.75% through direct microscopy and inflated to 9.5% through ELISA. The demographic factors positively associated with occurrences of giardiasis were age (P = 0.035; odds ratio, OR = 1.96; 95% confidence interval, CI = 1.094-3.533), mother's educational level (P = 0.031; OR = 2.67; 95% CI = 1.186-6.045) and father's educational level (P = 0.004; OR = 3.56; 95% CI = 1.612-7.899). Similarly, among the supposed risk factors, rural residency (P = 0.032; OR = 1.76; 95% CI = 1.098- 2.851), absence of proper sewerage system (P = 0.000; OR = 6.60; 95% CI = 4.029-10.841) and unavailability of safe drinking water (P = 0.000; OR = 4.08; 95% CI = 2.207-7.547) were the factors strongly connected with giardiasis. Abdominal discomfort was a prominent clinical sign with 46% frequency. CONCLUSION: Various risk factors were associated with occurrences of Giardia, thus emphasizing the importance of parents' education, safe drinking water and proper sewerage systems for Pakistani children's health.

Efficacy of Azadirachta indica organic extracts against clinical methicillin resistant Staphylococcus aureus isolates.

In current study we investigated the efficacy of organic extracts of Azadirachta indica leaves against Methicillin Resistant Staphylococcus aureus (MRSA) clinical isolates. For this purpose fresh leaves were used to prepare ethanol, methanol and chloroform extract. Secondly, a cross sectional study was conducted to isolate MRSA in clinical samples from patients having surgical/ non-surgical wounds from Allied Hospital and District Head Quarter Hospital, Faisalabad. The S. aureus isolates were initially identified by biochemical characterization, followed by identification of MRSA using cefoxitin disc diffusion test that was finally confirmed by genomic amplification of mecA gene, responsible for resistance. All MRSA isolates were tested to find vancomycin resistant S. aureus (VRSA) using E-strips (M.I.C. EvaluatorTM, Oxide, UK). The data showed an overall 37% prevalence of S. aureus including 56.75% clinical MRSA isolates while none of the isolated S. aureus showed resistance to vancomycin. The antimicrobial activity was measured as mean zone of inhibition for each extract against all MRSA isolates and it was found as 15.38±2.26, 16.09±3.09 and 17.42±2.48 for methanol, ethanol and chloroform extracts respectively. Chloroform extract showed significantly high antimicrobial activity against MRSA isolates. Altogether, the current study exposed the high prevalence of MRSA isolates from tertiary care hospitals. However, all MRSA isolates were found susceptible to organic extracts of A. indica leaves.

Toxicity, Phytochemical Composition, and Enzyme Inhibitory Activities of Some Indigenous Weed Plant Extracts in Fruit Fly, Drosophila melanogaster.

Drosophila melanogaster being used as model organism is considered as pest of homes, restaurants, and fruit markets. The damaged fruits are also reported to serve as a carrier for various diseases. The current study was designed to evaluate the toxicity of petroleum extract of some weed plants, namely, Euphorbia prostrata, Parthenium hysterophorus, Fumaria indica, Chenopodium murale, and Azadirachta indica, against D. melanogaster. Mortality at 10, 20, and 30% concentrations after 24 and 48 hours was found comparatively low. E. prostrata caused high mortality (51.64%) at 30% concentration and was found more toxic (LC50 27.76; P value 0.00) after 72 hours. A. indica showed high LC50 value (P value 0.15) compared to other weed plants. The combination of E. prostrata and Bti showed highest mortality (100%; LC50 12.49; P value 0.00) after 72 hours. Similarly, the same combination caused maximum reduction in the activity of AChE, AcP, AkP, α-Carboxyl, and ß-Carboxyl enzymes. Phytochemical analysis showed the presence of flavonoids, saponins, tannins, steroids, cardiac glycosides, alkaloids, anthraquinones, and terpenoids. FTIR analysis of E. prostrata showed the presence of phenolic compounds. It is suggested that further studies are needed in order to incorporate weed plant extracts in combination with Bti for the management of fruit flies.