Expression profile of KIR3DS1/KIR3DL1 receptors in association with immunological responses in TB, HIV and HIV/TB infected patients.

Several studies investigated KIR3DS1 and KIR3DL1 in the context of various infections. However, none of the studies were performed on KIR3DS1/L1 in association with IFN-É£/IL-10 in TB, HIV-1, and their confections. We aimed to evaluate KIR3DS1/KIR3DL1 expression in association with IFNÉ£/IL-10 in HIV-1 and TB mono-infections and HIV-1/TB confection and compared with uninfected controls using RTq PCR. We also performed correlation analysis between KIR3DS1, KIR3DL1, IFN-É£ and IL-10 in the respective cohorts. The overall expression of KIR3DS1 was found to be downregulated in all groups, whereas in HIV-1 and HIV-1/TB, the frequency of KIR3DS1(+) expression was significantly (p < 0.05) associated with undetected HIV-1 viral load. However, expression of KIR3DL1 was found to be significantly (p < 0.05) upregulated in HIV-1 only. In addition, IFNÉ£ expression was significantly (p < 0.05) decreased in TB, whereas in HIV-1/TB, IFNÉ£ expression was significantly (p < 0.05) increased. In contrast, IL-10 expression was significantly (p < 0.05) increased in HIV-1 and HIV-1/TB but not in TB. Also, we found significant positive correlation (p < 0.05, r = 0.61) between KIR3DL1 and IFNÉ£ expression in TB and negative correlation (p < 0.05, r = - 0.62) between KIR3DS1 and IL-10 in HIV-1/TB. In conclusion, we suggest that expression of KIR3DS1/L1 is associated with IFNÉ£/IL-10 responses and it is involved in modulating disease severity in HIV-1 and TB infections.

Association of trinucleotide repeat polymorphisms CAG and GGC in exon 1 of the androgen receptor gene with male infertility: a cross-sectional study.

BACKGROUND: : Infertility is a global problem that brings about serious sexual and social consequences that strain the health sector and society. The expansion of CAG and GGC repeats in androgen receptor (AR) gene (Ensembl number ENSG00000169083) may lead to reduced fertility. Our objective was to determine the association of CAG and GGC repeats with altered sperm parameters in male infertile subjects. METHODS: This was a cross-sectional study conducted at Aga Khan University, Karachi, Pakistan. A total of 376 males were recruited, out of which group A (N = 208) and group B (N = 168) were comprised of subjects with normal and altered sperm parameters, respectively, from 18 to 60 years. The numbers of CAG and GGC repeats were determined by using PCR amplification and sequence analysis using the Molecular Evolutionary Genetic Analysis (MEGA) software version 6.0. Statistical analysis was performed using the SPSS version 20 and the P-value of <0.05 was considered significant. RESULTS: The mean androgen receptor gene CAG repeats were significantly longer in males with altered sperm parameters as compared to male subjects with normal sperm parameters (P < 0.001). There was no significant difference found for GGC repeats for subjects with altered sperm parameters. DISCUSSION: Longer CAG length corresponded to greater severity of spermatogenic defect and may lead to subfertility recommendations.

A comparative study on production of extracellular hydrolytic enzymes of Candida species isolated from patients with surgical site infection and from healthy individuals and their co-relation with antifungal drug resistance.

BACKGROUND: Surgical site infection (SSI) is a crucial dilemma of surgery. Patients with SSIs not only face difficulty in treatment but also bear extra cost with high mortality rate. Resistant strains of Candida have emerged as an important nosocomial pathogen. Proteinase and phospholipase are exo- enzymes of Candida species, have importance with respect to their contribution in diseases. This study focused on prevalence of Candida species in surgical wound, their resistance to antifungal drugs, co-relation of these resistance with virulence potential of Candida species and comparison of production level of exo-enzymes of Candida species isolated from patients with SSIs and healthy individuals to highlights their role in SSIs. RESULTS: A total of (n = 555) swab samples were investigated. (n = 450) samples were collected from patients with SSIs and (n = 105) were collected from healthy individuals. Samples were subjected for the identification of Candida species which were subsequently investigated for antifungal susceptibility, MICs and enzymatic activity of Candida species. Out of 128 strains of Candida spp. isolated from SSIs, 54(42.18%) were identified as C. albicans followed by C. glabrata 32(25%), C. parapsilosis 17(13.28%), C. krusei 13(10.16%) and C. tropicalis 12(9.38%). C. albicans isolates showed 100% susceptibility to voriconazole and amphotericin B followed by itraconazole 98% and fluconazole 89%. Out of 6 fluconazole resistant C. albicans 5(83.33%) were able to produce phospholipase while out of 48 fluconazole-susceptible strains 17(35.42%) were found to be phospholipase producer. Out of 54 C. albicans isolated from surgical wound 46(85.18%) and 49(90.74%) were found to be phospholipase and proteinase producer respectively, whereas out of 20 C. albicans isolates from healthy subjects 14(70%) produce proteinase and 12(60%) produce phospholipase. There were significant statistical differences found between the level of enzyme production by C. albicans, in relation to both sites (P = 0.014). CONCLUSION: Study revealed that prevalence of Candida species is high in SSIs. Phospholipase and proteinase activity were more pronounced in Candida Species from surgical wound in contrast to species from healthy individuals suggests these enzymes may have been responsible for the severity of infection in surgical wound patients.

The antibiofilm activity of Acetylsalicylic acid, Mefenamic acid, Acetaminophen against biofilms formed by P. aeruginosa and S. epidermidis.

OBJECTIVE: To evaluate the antibacterial activity of Aspirin, Mefenamic acid and Acetaminophen against Pseudomonas aeruginosa and Staphylococcus epidermidis biofilms. METHODS: The study was conducted AKU Karachi in collaboration with DIHE Karachi from March 2018 to December 2018.Quantitative spectrophotometric method was used to study the reduction and removal of the Pseudomonas aeruginosa and Staphylococcus epidermidis formed biofilms. Statistical tests were performed using Graph Pad Prism software. . RESULTS: Acetaminophen showed maximum biofilm reduction activity against the biofilms formed by Pseudomonas aeruginosa, and Staphylococcus epidermidis. Mefanamic acid showed maximum biofilm removal potential against Pseudomonas aeruginosa, while Aspirin and Mefanamic acid were equally effective in removing biofilms formed by Staphylococcus epidermidis as well. CONCLUSIONS: There is a continuous need to look for non-antibiotic agents for their potential antimicrobial and antibiofilm potential.

Camellia sinensis Mediated Enhancement of Humoral Immunity to Particulate and Non-particulate Antigens.

The most common drinking beverage in large portion of the world is Camellia sinensis (green tea). In the present study, we evaluated the adjuvant effect of green tea and tea polyphenols to particulate and non-particulate antigens. BALB/c mice were immunized with particulate and non-particulate antigens. Modulation of immunoglobulin-secreting splenocytes, IgG-mediated and IgM-mediated immunity, was evaluated by hemolytic plaque assay and enzyme-linked immunosorbent assay, respectively. Dose-dependent response of tea polyphenols was also assayed. Phenolic content was measured in crude preparations of green tea. We observed a stimulatory effect of green tea preparations on humoral immune response mediated by the increased number of antibody-secreted cells in spleen. A significant increase in IgM-mediated and IgG-mediated immune response to non-particulate antigen was also observed in green tea-treated animals. A dose-dependent adjuvant effect was seen in the case of tea polyphenols for a longer period of time compared with crude tea preparations. This study indicates polyphenols as major constituents responsible for the enhanced and sustained adjuvant activity of green tea. We suggest that tea polyphenols might be considered for real-life evaluation during adjuvant-mediated vaccination trial programs.

Physical activity, smoking, and genetic predisposition to obesity in people from Pakistan: the PROMIS study.

BACKGROUND: Multiple genetic variants have been reliably associated with obesity-related traits in Europeans, but little is known about their associations and interactions with lifestyle factors in South Asians. METHODS: In 16,157 Pakistani adults (8232 controls; 7925 diagnosed with myocardial infarction [MI]) enrolled in the PROMIS Study, we tested whether: a) BMI-associated loci, individually or in aggregate (as a genetic risk score--GRS), are associated with BMI; b) physical activity and smoking modify the association of these loci with BMI. Analyses were adjusted for age, age(2), sex, MI (yes/no), and population substructure. RESULTS: Of 95 SNPs studied here, 73 showed directionally consistent effects on BMI as reported in Europeans. Each additional BMI-raising allele of the GRS was associated with 0.04 (SE = 0.01) kg/m(2) higher BMI (P = 4.5 × 10(-14)). We observed nominal evidence of interactions of CLIP1 rs11583200 (P(interaction) = 0.014), CADM2 rs13078960 (P(interaction) = 0.037) and GALNT10 rs7715256 (P(interaction) = 0.048) with physical activity, and PTBP2 rs11165643 (P(interaction) = 0.045), HIP1 rs1167827 (P(interaction) = 0.015), C6orf106 rs205262 (P(interaction) = 0.032) and GRID1 rs7899106 (P(interaction) = 0.043) with smoking on BMI. CONCLUSIONS: Most BMI-associated loci have directionally consistent effects on BMI in Pakistanis and Europeans. There were suggestive interactions of established BMI-related SNPs with smoking or physical activity.

Correlation of ABH blood group antigens secretion with Helicobacter pylori infection in Pakistani patients.

OBJECTIVES: A and B blood group antigens are fucosylated carbohydrate present on human erythrocytes and body secretions. Their presence in body secretions depends on the expression of a dominant allele of secretor gene FUT2 and is correlated with susceptibility to various infectious and non-infectious diseases. We investigated the correlation of blood group and ABH antigen secretion with Helicobacter pylori infection and gastroduodenal symptoms and analysed the distribution of babA gene among ABH secretors and non-secretors. METHODS: Two hundred and ninety patients who underwent gastroduodenal endoscopy during 2011 to 2012 participated. Gastric biopsy, saliva and blood samples were obtained from every patient. Gastric biopsies were subjected to rapid urease test and PCR for the detection of H. pylori and babA gene. Blood grouping and ABH antigens secretions were determined by Lewis blood group phenotyping and haemagglutination inhibition test. RESULTS: 50.34% of patients were ABH antigen secretors and 45.51% non-secretors. Distribution analysis of blood group revealed that 40 blood group B, 67 blood group A 20 blood group O and 19 blood group AB patients secreted ABH antigens in saliva. Fifty-six blood group O, 19 blood group B, 32 blood group A and 17 blood group AB patients were non-secretors. Gastroduodenal complaints were common among non-secretors. Sixty-two percent of patients with a combination of duodenal ulcer and gastro-oesophageal reflux and 54% of patients with gastritis were non-secretors. Of 290 samples, 31.02% were positive for H. pylori. Thirty percent of these tested positive for babA gene; the majority belonged to non-secretor blood group O. CONCLUSIONS: Our results suggest that the infection of H. pylori is correlated with ABO blood groups and blood group antigens secretion in body fluids.

Cervical cancer screening and HPV genotype distribution among asymptomatic patients of Karachi Pakistan.

BACKGROUND: Cervical cancer is the second most prevalent cancer in females worldwide. Human papillomavirus (HPV) infection is a sexually transmitted infection. However, in addition to HPV infection, other factors exist that influence the risk of developing cervical cancer. In Pakistan most women who developed cervical cancer have been infrequently or never screened. OBJECTIVE: To determine the prevalence of HPV infection and its subtype profile among asymptomatic patients with pre cancerous cervical intraepithelial lesion. METHODS: In this hospital-based descriptive study, 160 asymptomatic females attending gynecology clinics were subjected to HPV screening after obtaining informed consent. Cervical Scrapings were examined by cytopathology and colposcopic directed biopsies taken. High-grade intraepithelial lesion (HSIL) CIN-2, and Low-grade intraepithelial lesion (LSIL) CIN-1 were selected. Samples were analyzed for the presence of HPV-DNA general and type specific genotype 16 and 18. HPV- DNA was extracted by QIA amp DNA kit protocol and amplification was done by polymerase chain reaction (PCR) and genotyped by type specific primers. RESULTS: Out of 160, 17 Pap smear tests were positive, 6 (35.3%) with abnormal results (HSIL) CIN-2 were HPV-DNA positive. Among them, 5 (83.3%) had subtype 16 and in 1 (16.7%) case the genotype was undetectable. The remaining 11(6.9%) with pre cancer minimal abnormal (LSIL) CIN-1 presented. Out of them 3 (27.3%) were HPV-DNA positive with subtype 16. Five (45.4%) were followed by repeated pap smear every six months for two years, and the rest of 3 (27.3%) patients refused for the test. CONCLUSION: A high incidence of Human papillomavirus (HPV) infection is found in women with pre cancerous lesion of cervix in Pakistani women.

Development of an in-house enzyme-linked immunosorbent assay based on surface whole cell antigen for diagnosis of Helicobacter pylori infection in patients with gastroduodenal ulcer disease.

Helicobacter pylori (H. pylori) is a causative agent of gastritis, gastroduodenal ulcers and gastric adenocarcinoma. More than 50% world population is colonized by H. pylori, which is closely related to the chronic gastritis and gastric ulcer infection. In this study, a total of 214 gastritis patient's serum samples were screened for anti-H. pylori IgG antibody. A 96-well plate coated with 20 µg/ml antigen and hundred-fold diluted patient's serum was allowed to react. After extensive washing with buffer, 1:2,500 diluted conjugated secondary antibody was added. Later substrate was added to observe positivity by measuring the intensity of color. Statistical analyses were performed, and p value of <0.01 was taken as significant; 84% male patients and 89% female patients, respectively, tested positive for H. pylori, while agewise distribution was 35-45 years males (40%) and 35-55 years females (52%) were found highest number of H. pylori infected patients. In-house ELISA based on surface whole cell antigen (wELISA) showed a sensitivity of 93%, specificity of 100%, accuracy 94% and κ value 0.86 with significant correlation R-0.77020; p < 0.0001. We conclude that H. pylori local isolates surface antigen was satisfactory for diagnosis as different parameters were adjusted according to the local H. pylori isolates. Fluctuations in serum antibody titer predict the variation in an individual's response of the host against H. pylori. In-house wELISA could provide a reliable and a clinically useful method for the diagnosis of H. pylori infection in patients of Karachi, Pakistan.

Prevalence and Susceptibility Pattern of Multi Drug Resistant Clinical Isolates of Pseudomonas aeruginosa in Karachi.

OBJECTIVE: To determine the frequency and susceptibility pattern of multi-drug resistant (MDR) Pseudomonas aeruginosa isolated from clinical specimens in Karachi. METHODS: This cross sectional study was conducted in Microbiology Department, University of Karachi, from January 2012 to January 2013. Clinical specimens were collected from different hospitals of Karachi. Clinical isolates were identified by standard and specific microbiological methods. The antibiotic susceptibility pattern was determined by Kirby Bauer Disc diffusion method. Clinical and Laboratory Standards Institute (CLSI) guidelines were used to determine the results. RESULTS: The frequency of MDR P. aeruginosa isolated from different clinical specimens was found to be 30%. Amikacin was found to be the most effective antibiotic, followed by Co-trimaxazole and Quinolones. CONCLUSION: Antibiotic resistant P. aeruginosa are emerging as a critical human health issue. There is an urgent need to resolve the issue by taking some preventive measures. Combined efforts of health care professionals and researchers are required to educate people about the proper use of antibiotics and other infection control measures.

Chemical and antimicrobial studies on the essential oil from Salvia santolinifolia Boiss.

In view of the reputation of genus Salvia in folklore medicine and its abundance in our region, the chemical composition and antimicrobial activity of essential oil from S. santolinifolia Boiss. was analyzed. Chemical analysis, using gas chromatography and gas chromatography mass spectrometry, retention indices and C-13 nuclear magnetic resonance spectroscopy has resulted in identification of 116 constituents, comprising about 97% of the total constituents. Out of these 116, 78 constituents are hitherto unreported from this source. The species belongs to α-pinene chemotype. In antibacterial assay, gram negative gastropathogens (Shigella boydii, S. flexneri, S. dysenteriae, Vibrio cholerae); causative agent of urinary tract infection (Proteus mirabilis and P. vulgaris) and pneumonia (Klebsiella pneumoniae) were found sensitive to this essential oil while Corynebacteria species and Staphylococcus epidermidis were significantly inhibited in antibacterial assay against gram positive bacteria. Clinical and Laboratory Standards Institute protocol was used for determining antimicrobial activity. Thus the essential oil from this species can be utilized as potential chemotherapeutic agent.

Thin Layer Immunoassay: An Economical Approach to Diagnose Helicobacter pylori Infection in Gastroduodenal Ulcer Disease Patients of Pakistan, a Comparative Analysis.

Helicobacter pylori is a causative agent of gastritis, gastroduodenal ulcers and gastric adenocarcinoma. The majority of H. pylori-associated patients live in underdeveloped areas, facing the problem of lack of proper diagnostic facility. Hence, a simple and economical assay is required to handle the majority of gastric patients. Serum samples from gastroduodenal ulcers and gastritis patients were screened for H. pylori infection by thin layer immunoassay. A polystyrene plate coated with H. pylori sonicate whole cell antigen (10 µg/mL). Two-fold diluted patient's serum was allowed to react at 37 °C, incubated at 60 °C for 1 min over a water bath and the water condensation pattern for the H. pylori antibody was recorded. ELISAs were used as reference assays to evaluate the efficacy of the developed thin layer immunoassay (TLI). Gastric patients' blood samples (62% male and 6% female) tested positive for H. pylori, while age-wise, 15-25-year-old males (36%) and 65-75-year-old females (50%) showed the highest number of H. pylori infections. TLI showed sensitivity (72-67%), specificity (100%), accuracy (94-69%) and κ value (0.493-0.357) in comparison with wELISA (Surface whole cell ELISA), sELISA (sonicate whole cell ELISA) and kELISA (commercial KIT ELISA). We conclude that thin layer immunoassay is a low cost, fast, simple and clinically reliable method for H. pylori diagnosis at initial stages in patients in under-developed countries.

Identification of immune biomarkers in recent active pulmonary tuberculosis.

Tuberculosis (TB) has remained an unsolved problem and a major public health issue, particularly in developing countries. Pakistan is one of the countries with the highest tuberculosis infection rates globally. However, methods or biomarkers to detect early signs of TB infection are limited. Here, we characterized the mRNA profiles of immune responses in unstimulated Peripheral blood mononuclear cells obtained from treatment naïve patients with early signs of active pulmonary tuberculosis without previous history of clinical TB. We identified a unique mRNA profile in active TB compared to uninfected controls, including cytokines such as IL-27, IL-15, IL-2RA, IL-24, and TGFß, transcription factors such as STAT1 and NFATC1 and immune markers/receptors such as TLR4, IRF1, CD80, CD28, and PTGDR2 from an overall 84 different transcripts analyzed. Among 12 significant differentially expressed transcripts, we identified five gene signatures which included three upregulated IL-27, STAT1, TLR4 and two downregulated IL-24 and CD80 that best discriminate between active pulmonary TB and uninfected controls with AUC ranging from 0.9 to 1. Our data identified a molecular immune signature associated with the early stages of active pulmonary tuberculosis and it could be further investigated as a potential biomarker of pulmonary TB.

Temporal expression of extracellular products of Staphylococcus aureus in vivo mouse cage model.

OBJECTIVE: To test the hypothesis that Staphylococcus aureus genome has regulatory genes which coordinate the expression of extracellular products, and particular genes not expressed in vitro conditions may be turned on in a vivo environment. METHODS: The study was conducted at the Immunology and Infectious Disease Research Laboratory (IIDRL), Microbiology Department, Husein Ebrahim Jamal Research Institute (HEJ), Animal House, Karachi University, from July to December 2009. Micro pore Teflon cages using a mouse cage model were fixed into the subcutaneous tissue in Albino mice (BALB/c) on their dorsal surface. After 15 days, the holes closed down with healthy tissue. Three staphylococcal isolates from clinical samples confirmed by DNA sequencing of 16s ribosomal RNA were tested for expression of extracellular protein in vitro and were later injected into the cages. After the institution of infection, the fluid aspirated from the cages was analysed by Sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE). This was done to test for possible induction of additional extracellular proteins in vivo. RESULTS: The appearance of enhanced extracellular products was observed in the fluid recovered from the cages of two mice on days 5 and 7 subsequent to the institution of infection, suggesting a turn-on of particular genes which were not expressed in vitro conditions. CONCLUSIONS: In-vivo host and environmental signals contribute to the induction of genes for the production of extracellular proteins.

Diversity of Rotavirus Strains among Children with Acute Diarrhea in Karachi, Pakistan.

One of the common viral pathogens in infectious diarrhea is Rotavirus; in developing countries, it is a primary cause of deaths in children less than five years of age. This study was planned to find out the etiologic agents of acute watery diarrhea. In this study, 1465 stool samples were analyzed with the symptoms of acute diarrhea. Demographic data analysis showed no. of episodes of diarrhea, vomiting, and fever. All samples were checked by ELISA technique for the presence of Rotavirus circulating strains. More than 6% patients were found to be positive with Rotavirus. Common Rotavirus genotypes, including G2P4, G2P6, G3P4, G8P4, G8P6, G9P4, and G10P4, were detected in patients through RT-PCR. This study concluded that detection of rotavirus strain diversity and management of diarrheal patients may identify assortment of emerging strains and reduce emergence of antimicrobial resistance and repeated episodes of diarrhea, which may also help to avoid and manage the essential nutrients lost leading to malnutrition and stunted growth, as well as to reduce high mortality rate in young children less than five years.

Antibacterial activity in spices and local medicinal plants against clinical isolates of Karachi, Pakistan.

CONTEXT: Development of resistance in human pathogens against conventional antibiotic necessitates searching indigenous medicinal plants having antibacterial property. Twenty-seven medicinal plants used actively in folklore, ayurvedic and traditional system of medicine were selected for the evaluation of their antimicrobial activity for this study. Eleven plants chosen from these 27 are used as spices in local cuisine. OBJECTIVE: Evaluation of the effectiveness of some medicinal plant extracts against clinical isolates. MATERIAL AND METHODS: Nonedible plant parts were extracted with methanol and evaporated in vacuo to obtain residue. Powdered edible parts were boiled three times and cooled in sterile distilled water for 2 min each and filtrate collected. The minimum inhibitory concentration (MIC) of plant extracts and filtrates/antibiotics was evaluated against clinical isolates by microbroth dilution method. RESULTS: Water extract of Syzygium aromaticum L. (Myrtaceae) buds, methanol extracts of Ficus carica L. (Moraceae) and Olea europaea L. (Oleaceae) leaves and Peganum harmala L. (Nitrariaceae) seeds had MIC ranges of 31.25-250 µg/ml. S. aromaticum inhibited growth of Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pyogenes, Salmonella enterica serovar Typhi and Pseudomonas aeruginosa. F. carica and O. europaea inhibited growth of S. aureus, S. epidermidis, and S. pyogenes whereas P. harmala was effective against S. aureus, Acinetobacter calcoaceticus and Candida albicans. Ampicillin, velosef, sulfamethoxazole, tetracycline and ceftazidime, cefotaxime, cefepime, which are used as control, had MIC ≥ 50 and 1.5 µg/ml, respectively, for organisms sensitive to extracts. DISCUSSION AND CONCLUSION: Mono/multiextract from identified plants will provide an array of safe antimicrobial agents to control infections by drug-resistant bacteria.

Structural determination of daphnecin, a new coumarinolignan from Daphne mucronata.

Daphnecin (1), a new coumarinolignan, has been isolated from the ethyl acetate-soluble fraction of Daphne mucronata along with three known compounds, aquillochin (2), umbelliferone (3), and coumarin (4). Their structures were determined by spectroscopic studies.

Current pattern of antibiotic resistance in Staphylococcus aureus clinical isolates and the emergence of vancomycin resistance.

OBJECTIVE: To determine the pattern of antibiotic resistance in the clinical isolates of Staphylococcus (S.) aureus, Methicillin Resistant S. aureus (MRSA) and define the possible emergence of Vancomycin resistant S. aureus (VRSA) in Karachi. STUDY DESIGN: An observational study. PLACE AND DURATION OF STUDY: Essa Laboratories and Department of Molecular Genetics, Ziauddin Hospital, from January to December 2009. METHODOLOGY: Staphylococcal isolates from different clinical specimens, pus, urine, blood, high vaginal swab and other secretions received at Ziauddun laboratories and Dr.Essa laboratories were collected. The specimens were inoculated on blood agar, MacConkey agar and Chrom agar. Antibiotic susceptibility to conventional antibiotics was done by disc diffusion, and E-test. Methicillin resistance was tested by using Oxacillin and Methicillin disks and confirmed by gold standard PCR for presence of mecA gene. All MRSA strains were subjected in addition to Vancomycin screen agar test. RESULTS: Out of the 450 S. aureus isolates 174 (38.6%) were found to be MRSA. In those isolates, high resistance was found to Cefixime (100%) Doxicycline (100%) Oxacillin (96.5%) Gentamicin, (96.3%), Timethoprim/Sulfametoxazole (95.6%) Chloramphenicol (93%) Tobramicin (81.03%), Ofloxacin (72.4%) and Ciprofloxacin (63.7%). Low resistance was found to Ceftazidine (36%), Amoxicillin/Clavulanate (32.7%), Fosfomycin (31%), Cefroxime (24%), Amikacin (17.2%) and Meropenem (13%). One isolate was found to be Vancomycin resistant (MIC 32 µg/ml). Four isolates had intermediate resistance, with two strains having MIC of 16µg/ml and two having MIC of 8µg/ml. These strains were also resistant to all the other tested antibiotics except Linezolid to which all isolates were susceptible. CONCLUSION: Antibiotic resistance to all the conventionally used antibiotics was high in the tested isolates. All the strains were susceptible to Linezolid which is an expensive alternative with adverse side effects. Judicious use of antibiotics focused on the compliance and formation of antibiotic policy guide lines is highly recommended.

Helicobacter pylori severely reduces expression of DNA repair proteins PMS2 and ERCC1 in gastritis and gastric cancer.

Gastric cancers are the third leading cause of cancer mortality in the world. Helicobacter pylori causes over 60 % of all stomach cancers. Colonization of the gastric mucosa by H. pylori results in increased DNA damage. Repair of DNA damage may also be reduced by H. pylori infection. Reduced DNA repair in combination with increased DNA damage can cause carcinogenic mutations. During progression to gastric cancer, gastric epithelium goes through stages of increasing pathology. Determining the levels of DNA repair enzymes during progression to gastric cancer could illuminate treatment approaches. Our aim is to determine the level of gastric expression of DNA repair proteins ERCC1 (a nucleotide excision repair enzyme) and PMS2 (a mismatch repair enzyme) in the presence of H. pylori infection at successive stages of gastric pathology and in gastric cancers. We analyzed gastric tissues of 300 individuals, including 30 without dyspepsia, 200 with dyspepsia and 70 with gastric cancers. The presence of H. pylori, gastric pathology and expression of DNA repair proteins ERCC1 and PMS2 were evaluated. Infection by H. pylori carrying the common cagA gene reduced median nuclear expression of ERCC1 and PMS2 to less than 20 % and 15 % of normal, respectively, in all pathologic stages preceding cancer. ERCC1 and PMS2 nuclear expression was 0-5 % of normal in gastric cancers. H. pylori can cause deficiency of ERCC1 and PMS2 protein expression. These deficiencies are associated with gastric pathology and cancer. This reduction in DNA repair likely causes carcinogenic mutations. Substantially reduced ERCC1 and PMS2 expression appears to be an early step in progression to H. pylori-induced gastric cancer.

Molecular characterization of Pakistani field isolates of infectious bursal disease virus.

The reverse transcriptase-polymerase chain reaction followed by restriction fragment length polymorphism (RT-PCR/RFLP) technique was used to identify and characterize Pakistani field isolates of infectious bursal disease virus (IBDV). These isolates have caused heavy losses to the poultry industry (mortality up to 60%) during the period between 1999 and 2005. Ten samples (five local isolates and five commercial vaccines) were examined for IBDV. Nine samples were positive for IBDV as evidenced by the amplification of the 743-bp region of the VP2 gene by RT-PCR. The RT-PCR products were subjected to restriction enzyme digestion with BstNI, MboI, and SspI. The RFLP profiles of all samples on digestion with the MboI enzyme yielded a fragment size of 229 and 362 bp except for vaccine strain Bursine Plus, which yielded a profile of 229 and 480 bp. However, digestion with BstNI yielded two distinct RFLP patterns. The first profile was detected in field isolates ML-1/SPVC/2001 and NP2/SPVC/2002 with four fragments of 119, 154, 172, and 209 bp, resembling RFLP profiles of molecular group 4 isolates. NL-3/SPVC/2003, NK-4/SPVC/2004, and NPK-5/SPVC/2005 generated a different RFLP profile with fragments of 119, 172, and 424 bp, resembling the profiles of molecular group 6 isolates. However, all the field and vaccine strains showed the absence of SspI restriction sites in their genome. It can be concluded that the Pakistani isolates can be grouped in molecular groups 4 and 6 of IBDV.