Helicobacter pylori virulence regulatory network: insights into the host-environment and pathogen interactions

Helicobacter pylori [H. pylori] has involved significant regulatory mechanisms in order to acclimatize in extreme gastric environment of human beings. The virulence machinery of H. pylori is complicated as virulence factors of pathogen not only interact with transcription and translational machinery of host, but also are involved in the progression and development of the disease. The present study is an effort to model virulence mechanism in H. pylori, particularly ferric uptake regulator [FUR] under acidic and iron [Fe] depleted conditions, as well as its effects on the well known virulence factors cytotoxin-associated gene A [cagA] and vacuolating cytotoxin A [ vacA] gene. The virulence regulatory network of cagA and vacA is modeled based on an asynchronous kinetic logic formalism introduced by Rene Thomas. The cagA-vacA virulence regulatory network is then elaborated qualitatively to obtain insights into H. pylori induced pathogenesis. The findings have revealed the significant regulatory pathways through which H. pylori spreads infection to the gastric cells, and also verified that cagA is associated with acute gastritis while vacA is involved in vacuolation, apoptosis and atrophy. Interestingly, both cagA and vacA were found to modulate each other virulence potential which ultimately leads to the state of chronic gastritis; which in turn drives the pathway smoothly towards gastric adenocarcinoma via the formation of premalignant lesions. The proposed strategy can be extended to understand the mechanism of other similar bacterial infections and disease progression. It will also help in the prioritization of potential therapeutic targets to control such serious infections

In vitro efficacy of meropenem, colistin and tigecycline against the extended spectrum beta-lactamase producing gram negative bacilli

To compare the in vitro efficacy of meropenem, colistin and tigecycline against extended spectrum Beta-lactamase producing Gram negative bacilli by minimal inhibitory concentration. Cross-sectional descriptive study. Department of Microbiology, Army Medical College, National University of Sciences and Technology, Rawalpindi, from June to December 2010. Routine clinical specimens were subjected to standard microbiological procedures and the isolates were identified to species level. Extended spectrum beta-lactamase producing Gram negative bacilli were detected by Jarlier disc synergy method and confirmed by ceftazidime and ceftazidime-clavulanate Etest. Minimum Inhibitory Concentration [MIC[90]] of meropenem, colistin and tigecycline was determined by Etest [AB BIOMERIUX] and the results were interpreted according to the manufacturer's instructions and Clinical and Laboratory Standards Institute guidelines and Food and Drug Authority recommendations. Results were analyzed by using Statistical Package for the Social Sciences version 20. A total of 52 non-duplicate extended spectrum Beta-lactamase-producing Gram negative bacilli were included in the study. The MIC[90] of tigecycline [0.75 microg/ml] was lowest as compared to the meropenem [2 microg/ml] and colistin [3 microg/ml]. Tigecycline is superior in efficacy against the extended spectrum Beta-lactamase producing Gram negative bacilli as compared to colistin and meropenem

Evaluation of different phenotypic methods for detection of amp C beta-lactamase producing bacteria in clinical isolates

To compare the sensitivity and specificity of different phenotypic methods for detection of Amp C beta-lactamase producing bacteria. Analytical study. Department of Microbiology, Army Medical College / National University of Sciences and Technology [NUST], Islamabad, Pakistan, from June 2010 to December 2010. A total of 150 clinical isolates were screened for presence of Amp C beta-lactamase by using the cefoxitin disc. The confirmatory methods evaluated were inhibitor based assay [boronic acid], Amp C disc test and Amp C Etest. Three dimensional enzyme extract assay was used as the reference method for determining the sensitivity and specificity. Among the total isolates tested, 62.8% bacteria showed the presence of Amp C beta-lactamase by standard three dimensional enzyme extract assay. Among the three methods compared, boronic acid disk test found out to be highly sensitive [88%] and specific [92%] for the detection of Amp C beta-lactamase producing bacteria. Detection of Amp C production is crucial in order to establish the antibiotic therapy and to attain the favourable clinical outcomes. Implementation of simple tests like boronic acid disk tests in the laboratories will help to alleviate the spread of Amp C beta-lactamase harboring organisms

In vitro efficacy of Colistin against multi-drug resistant Pseudomonas aeruginosa by minimum inhibitory concentration

Multi-drug resistant bacteria are an important cause of mortality and morbidity. In the management of various infections, timely detection and appropriate treatment, in accordance with the culture and sensitivity reports can help improve the treatment outcome. Colistin is a bactericidal antibiotic which is emerging as a reliable solution for treating infections with multi-drug resistant Gram negative bacilli. The aim of this study was to find out the in-vitro efficacy of colistin against multidrug resistant Pseudomonas aeruginosa isolates by minimum inhibitory concentration. This cross sectional, descriptive study was conducted in the Department of Microbiology, Army Medical College, National University of Sciences and Technology, Islamabad from February 2010 to January 2011. Antimicrobial sensitivity testing was done on Pseudomonas aeruginosa isolated from routine clinical specimens received and the strains which appeared resistant to at least one antimicrobial agent in three or more anti-pseudomonal antimicrobial categories were subjected to the Colistin Etest. The MIC endpoint of colistin was read, as per manufacturer's instructions [AB Biodisk, Solna, Sweden]. The isolates showing MIC of 2micro.g/ml or less were considered sensitive, those with 4-6 micro.g/ml as intermediate and >/= 8 micro.gg/ml as resistant. MIC[50] and MIC[90] of colistin against MDR Pseudomonas aeruginosa was determined. A total of 52 MDR Pseudomonas aeruginosa strains were isolated during the period of the study. The highest percentage was isolated from urine [36%] followed by respiratory tract infections [18%] and pus specimens [20%]. The highest percentage of these isolates was found to be susceptible to colistin followed by piperacillin-tazobactam and cefoperazone-sulbactam. A total of 36 [69%] isolates were sensitive, 10 [20%] were intermediate and 6 [11%] were resistant to colistin by Kirby Bauer disc diffusion method. MIC[50] was found to be l.0 micro.gg/ml while MIC[90] was 3.0 micro.gg/ml. Colistin is a reliable solution in cases of infections with MDR, XDR or PDR Pseudomonas aeruginosa

Evaluation of combined disc method for the detection of metallo-β-lactamase producing Gram negative bacilli

Aims: Infections due to metallo-β-lactamase (MBL) producing Gram negative rods are a cause of high mortality and morbidity. Early detection by an economical and accurate method may improve patient outcome. This study was aimed to evaluate the diagnostic accuracy of combined disc method for MBL detection by comparing it with MBL-Etest. Methodology and Results: This cross-sectional, validation study was carried out in the Department of Microbiology, Army Medical College, National University of Sciences and Technology, Rawalpindi, over a period of six months. A total of 52 non-duplicate Gram-negative rods isolated from the routine clinical specimens and found resistant to meropenem/imipenem on Kirby Bauer Disc Diffusion method were subjected to two tests for metallo-β-lactamase detection. One was combined Disc test using imipenem with Ethylene Diamine Tetraacetic Acid (EDTA), where a strain showing an increase in zone of inhibition of combined disc of ≥ 7 mm as compared to imipenem alone, was considered as MBL producer and the other one was MBL-Etest for which results were interpreted as per manufacturer’s guidelines. Combined disc method for MBL detection was found to have a sensitivity, specificity, positive predictive value, negative predictive value and accuracy of 97.5%, 100%, 100%, 92% and 98%. Conclusion, Significance and Impact of study: Combined disc method is an economical and reliable method for metallo-β-lactamase detection which can be used routinely in any laboratory.

Detection and antibiotic susceptibility pattern of biofilm producing Gram positive and Gram negative bacteria isolated from a tertiary care hospital of Pakistan

Microorganisms adhere to non-living material or living tissue, and form biofilms made up of extracellular polymers/slime. Biofilm-associated microorganisms behave differently from free-floating bacteria with respect to growth rates and ability to resist antimicrobial treatments and therefore pose a public health problem. The objective of this study is to detect the prevalence of biofilm producers among Gram positive and Gram negative bacteria isolated from clinical specimens, and to study their antimicrobial susceptibility pattern. The study was carried out from October 2009 to March 2010, at the Department of Microbiology, Army Medical College/ National University of Sciences and Technology (NUST), Rawalpindi, Pakistan. Clinical specimens were received from various wards of a tertiary care hospital. These were dealt by standard microbiological procedures. Gram positive and Gram negative bacteria isolated were subjected to biofilm detection by congo red agar method (CRA). Antimicrobial susceptibility testing of those isolates, which showed positive results (slime production), was done according to the Kirby-Bauer disc diffusion technique. A total of 150 isolates were tested for the production of biofilm/slime. Among them, 81 isolates showed positive results. From these 81, 51 were Gram positive and 30 were Gram negative. All the 81(54%) slime producers showed reduced susceptibility to majority of antibiotics. Bacterial biofilms are an important virulence factor associated with chronic nosocomial infection. Detection of biofilm forming organisms can help in appropriate antibiotic choice.

Frequency and antibiogram of multi-drug resistant pseudomonas aeruginosa

To find out the frequency and susceptibility pattern of multi-drug resistant [MDR] Pseudomonas aeruginosa in clinical specimens. Cross-sectional observational study. Department of Microbiology, Army Medical College, National University of Sciences and Technology [NUST], Rawalpindi, from January to September 2010. Routine clinical specimens were subjected to standard microbiological procedures and the isolates were identified to the species level. The antibiotics susceptibility was determined by Kirby Bauer Disc diffusion method and the results were interpreted according to the Clinical and Laboratory Standards Institute [CLSI] guidelines. The frequency of MDR Pseudomonas aeruginosa among all the Pseudomonas aeruginosa strains isolated was found to be 22.7%. These isolates were most sensitive to Colistin followed by Piperacillin-Tazobactam and Cefoperazone-Sulbactum. Increasing fequency of infections due to MDR Pseudomonas aeruginosa is an emerging threat in our set up which can be prevented by prescribing antibiotics judiciously and by adopting proper disinfection measures

Frequency and antibiotic susceptibility pattern of Amp C beta-lactamase producing bacteria isolated from a tertiary care hospital of Rawalpindi, Pakistan

Amp C beta-lactamases are cephalosporinases which hydrolyze cephamycins and are poorly inhibited by clavulanic acid. Amp C beta-lactamases confer resistance to a wide variety of antibiotics and pose both diagnostic and therapeutic challenges. The objective was to detect the frequency and antibiotic susceptibility pattern of Amp C beta-lactamase producing bacteria isolated from a tertiary care hospital of Pakistan. Organisms were isolated from various clinical specimens. First, the screening of the isolates was done by using cefoxitin disc. Screen-positive organisms were subjected to three dimensional extract test for detection of Amp C beta-lactamases. From a total of 100 organism tested, 64 organisms were positive on cefoxitin screen test. Out of these 64, 40 [62.5%] showed the presence of Amp C beta-lactamase. [E.coli n=18, K.pneumoniae n=14, K.oxytoca n=1, Enterobacter species n=5, Citrobacter freundii n=2] by three dimensional extract test. The antibiotics found out to show good activity against these resistant bacteria include meropenem and tigecycline. This is the first study to determine the frequency of Amp C beta lactamases from Pakistan. This study shows a high frequency of Amp C beta-lactamase producing isolates from a hospital, which may lead to serious therapeutic problems

Susceptibility pattern and frequency of extended specturm beta-lactamase producers in a tertiary care hospital

Infections with extended spectrum beta-lactamase [ESBL] producing organisms continue to be associated with higher rates of mortality, morbidity and health care costs. This study was carried out to find out frequency and sensitivity pattern of ESBL producers among Gram negative rods [GNRs] from clinical isolates of a tertiary care Hospital. A total of 1430 GNRs were recovered from 2347 clinical samples received from admitted patients in Military Hospital Rawalpindi. All samples were dealt by standard microbiological methods. Extended spectrum beta-lactamase detection in these isolates was done by double disc approximation test of Jarlier et al, 1988. Frequency of GNRs among clinical isolates was 61% and about 33% of these were ESBL producers. Escherichia coli were the most frequent ESBL producers followed by Klebsiella pneumoniae. Most of the ESBL producers were isolated from urine followed by catheter tips and pus. Among all the antibiotics tested, ESBL producers showed highest susceptibility to carbapenems followed by amikacin. Organisms showing resistance pattern similar to ESBLs were also significant. Continued surveillance by clinical microbiology laboratories, judicious use of antimicrobial agents and implementation of infection control measures are recommended to reduce the frequency of ESBL isolates