SEROVAR PROFILE AND DETECTION OF INVA VIRULENCE GENE AMONG NON-TYPHOIDAL SALMONELLAE SEROVARS ISOLATED FROM ACUTE GASTROENTERITIS CASES IN COASTAL KARNATAKA, SOUTHERN INDIA

  Objective: Non-typhoidal salmonellosis is one of the leading zoonosis in the world caused by non-typhoidal Salmonella (NTS). Invasive infections with NTS serovars occurs due to the presence of virulence genes like invA along with the immunosuppressive conditions of the patient. The study was conducted to isolate and identify the NTS serovars and their antimicrobial resistance profile from patients with diarrhea and also to detect the virulence marker – invA gene among these NTS serovars.Methods: A prospective cross-sectional study was conducted from January 2015 to December 2016 at the Enteric Diseases Division, Kasturba Medical College, Manipal. 1218 fecal specimens were collected from patients with diarrhea and before antibiotic treatment. NTS serovars were identified, serotyped and then screened for the presence of invA virulence gene.Results: A total of 33 (2.7%) NTS was isolated. Salmonella typhimurium (33.34%) was predominant followed by Salmonella oslo (30.3%). Out of 33 NTS, invA was positive for 28 isolates (84.8%) of which 25 (89.3%) patients were febrile which was statistically significant (p=0.000).Conclusion: Non-typhoidal salmonellosis is an emerging global infection among immunocompromised patients. Our study showed an association between the invA gene and febrile illness among the patients suffering. Thus, this study highlights the importance of invA as a significant marker for bloodstream invasion

ANTIBIOGRAM PROFILING OF HELICOBACTER PYLORI STRAINS AND THE EFFICACY OF BRASSICA CAPITATA AGAINST RESISTANT STRAINS ISOLATED FROM THE PATIENTS SUFFERING FROM GASTRODUODENAL DISEASES IN GUWAHATI, ASSAM

  Objective: Helicobacter pylori resistance toward commonly used antibiotics is increasing leading to the treatment failure; hence, our aim is to determine the antibiogram susceptibility pattern of H. pylori strains isolated from Guwahati, Assam (Northeast India) and also to test the efficacy of the Brassica capitata against the multi and dual drug-resistant strains of North and Northeast India.Methods: Minimum inhibitory concentration of different antibiotics was determined by agar dilution method. Disc diffusion method was used to check the efficacy of B. capitata against clarithromycin (CLR), metronidazole (MTZ), and levofloxacin (LEV)-resistant H. pylori strains.Results: All the H. pylori strains were 100% sensitive to CLR, tetracycline, amoxicillin, and furazolidone. 72.8% of the strains were sensitive toward MTZ and 54.5% were sensitive toward LEV. B. capitata showed good efficacy against the resistant strains of H. pylori of North and Northeast India.Conclusion: Most of the H. pylori strains from Northeast India were sensitive toward the commonly used antibiotics for the treatment regime. B. capitata is effective against H. pylori infection, suggesting its potential as an alternative therapy, and opens the way for further studies on identification of novel antimicrobial targets of B. capitata

Characterization of Vibrio cholerae O1 strains that trace the origin of Haitian-like genetic traits

Vibrio cholerae O1 is the etiological agent of the severe diarrheal disease cholera. The bacterium has recently been causing outbreaks in Haiti with catastrophic effects. Numerous mutations have been reported in V. cholerae O1 strains associated with the Haitian outbreak. These mutations encompass among other the genes encoding virulence factors such as the pilin subunit of the toxin-co-regulated pilus (tcpA), cholera toxin B subunit (ctxB), repeat in toxins (rtxA), and other genes such as the quinolone resistance-determining region (QRDR) of gyrase A (gyrA), rstB of RS element along with the alteration in the number of repeat sequences at the promoter region of ctxAB. Given the numerous genetic changes in those Haitian isolates, we decided to investigate the possible origins of those variations in the Indian subcontinent. Thus, we determined the genetic traits among V. cholerae O1 strains in Delhi, India. A total of 175 strains isolated from cholera patients during 2004 to 2012 were analysed in the present study. Our results showed that all the tested strains carried Haitian type tcpA (tcpACIRS) and variant gyrA indicating their first appearance before 2004 in Delhi. The Haitian variant rtxA and ctxB7 were first detected in Delhi during 2004 and 2006, respectively. Interestingly, not a single strain with the combination of El Tor rtxA and ctxB7 was detected in this study. The Delhi strains carried four heptad repeats (TTTTGAT) in the CT promoter region whereas Haitian strains carried 5 such repeats. Delhi strains did not have any deletion mutations in the rstB like Haitian strains. Overall, our study demonstrates the sequential accumulation of Haitian-like genetic traits among V. cholerae O1 strains in Delhi at different time points prior to the Haitian cholera outbreak

Majority-layered T hybridization using quantum-dot cellular automata

The atomistic quantum-dot cellular automata (QCA) based implementations of the reversible circuits have got tremendous exposures in the last few days, due to “room-temperature workability” of the QCA. The researchers are in serious need of a methodology that can realize the area-efficient QCA counterparts of reversible benchmark circuits. In this work, a novel methodology named majority-layered T hybridization is proposed to synthesize the reversible circuits using QCA. Firstly the reversible library consisting of CNTS Gates have been generated to validate the usability of the proposed methodology. Then, an elementary QCA module of 3×3 Toffoli Gate have been proposed and extended in the realization of 4×4, 5×5 and 6×6 Toffoli Gates (multi-control Toffoli Gates). The proper mathematical modelling of the several QCA design metrics like effective area, delay and O-cost has been established. The QCA counterpart of 3×3 Toffoli Gate reports 18.61% less effective area and 8.33% less O-cost compared to the previous Toffoli Gate designs. Moreover, the QCA layout of rd-32 reversible benchmark using multi-control Toffoli Gate has been employed to verify the scalability and reproducibility of the proposed methodology. The QCA layouts are generated, tested and simulated by renowned computer aided design tool QCADesigner 2.0.3

Characterization of an N6 adenine methyltransferase from Helicobacter pylori strain 26695 which methylates adjacent adenines on the same strand

Genomic sequences of Helicobacter pylori strains 26695, J99, HPAGI and G27 have revealed an abundance of restriction and modification genes. hp0050, which encodes an N6 adenine DNA methyltransferase, was cloned, overexpressed and purified to near homogeneity. It recognizes the sequence 5'-GRRG-3' (where R is A or G) and, most intriguingly, methylates both adenines when R is A (5'-GAAG-3'). Kinetic analysis suggests a nonprocessive (repeated-hit) mechanism of methylation in which HP0050 methyltransferase methylates one adenine at a time in the sequence 5'-GAAG-3'. This is the first report of an N6 adenine DNA methyltransferase that methylates two adjacent residues on the same strand. Interestingly, HP0050 homologs from two clinical strains of H. pylori (PG227 and 128) methylate only 5'-GAGG-3' compared with 5'-GRRG-3' in strain 26695. HP0050 methyltransferase is highly conserved as it is present in more than 90% of H. pylori strains. Inactivation of hp0050 in strain PG227 resulted in poor growth, suggesting its role in the biology of H. pylori. Collectively, these findings provide impetus for exploring the role(s) of this conserved DNA methyltransferase in the cellular processes of H. pylori

Comparative Transcriptomics of H. pylori Strains AM5, SS1 and Their hpyAVIBM Deletion Mutants: Possible Roles of Cytosine Methylation

Helicobacter pylori is an important human pathogen and one of the most successful chronic colonizers of the human body. H. pylori uses diverse mechanisms to modulate its interaction with the host in order to promote chronic infection and overcome host immune response. Restriction-modification genes are a major part of strain-specific genes present in H. pylori. The role of N-6 -adenine methylation in bacterial gene regulation and virulence is well established but not much is known about the effect of C-5 -cytosine methylation on gene expression in prokaryotes. In this study, it was observed by microarray analysis and RT-PCR, that deletion of an orphan C-5 -cytosine methyltransferase, hpyAVIBM in H. pylori strains AM5and SS1 has a significant effect on the expression of number of genes belonging to motility, adhesion and virulence. AM Delta DhpyAVIBM mutant strain has a different LPS profile and is able to induce high IL-8 production compared to wild-type. hpyAVIBM from strain 26695 is able to complement mutant SS1 and AM5 strains. This study highlights a possible significance of cytosine methylation in the physiology of H. pylori

Characterization of the clonal profile of methicillin resistant Staphylococcus aureus isolated from patients with early post-operative orthopedic implant based infections

Abstract Background To analyze the molecular epidemiology and to compare between the major methicillin resistant Staphylococcus aureus biotypes for association with patient characteristics who had an implant for closed fracture and developed early post-operative wound infections (POWI) in a tertiary care hospital of India. Methods Pulsed-field gel electrophoresis (PFGE), antimicrobial resistance, accessory gene regulator (agr) and staphylococcal cassette chromosome mec (SCCmec) types, Paton–Valentine leukocidin (PVL) gene, toxin gene profiling, biofilm formation and patient demographics were correlated with MLST clonal complexes (CC). Findings Overall eight different sequence types (STs) were detected with a predominance of ST239 (66%), ST22 (18%) and some minor types ST772, ST30 (4% each) ST1, ST642, ST6, ST107 (2% each). All ST239 isolates belong to CC239 and SCCmec III whereas ST22 isolates belong to CC22 and SCCmec IV. The isolates varied in the distribution of various toxin genes. With 63.63% biofilm formers ST239 were all multidrug resistant with frequent resistance to erythromycin, clindamycin, gentamicin, cefuroxime, amoxyclav and ciprofloxacin indicating doxycycline, amikacin, vancomycin and linezolid can be the drug of choice. Conclusion This study shows that ST239 MRSA is still most prevalent strain with new emergence of ST642 and ST107 isolates in association with orthopedic implant based POWI. As compare to other ST types ST239 strain was associated with adverse treatment outcomes. This highlights the importance of improving nosocomial infection control measures in this unit

Mutations in hpyAVIBM, C5 cytosine DNA methyltransferase from Helicobacter pylori result in relaxed specificity

The genome of Helicobacter pylori is rich in restrictionmodification (RM) systems. Approximately 4% of the genome codes for components of RM systems. hpyAVIBM, which codes for a phase-variable C5 cytosine methyltransferase (MTase) from H. pylori, lacks a cognate restriction enzyme. Over-expression of M.HpyAVIB in Escherichia coli enhances the rate of mutations. However, when the catalytically inactive F9N or C82W mutants of M.HpyAVIB were expressed in E. coli, mutations were not observed. The M.HpyAVIB gene itself was mutated to give rise to different variants of the MTase. M.HpyAVIB variants were purified and differences in kinetic properties and specificity were observed. Intriguingly, purified MTase variants showed relaxed substrate specificity. Homologues of hpyAVIBM homologues amplified and sequenced from different clinical isolates showed similar variations in sequence. Thus, hpyAVIBM presents an interesting example of allelic variations in H. pylori where changes in the nucleotide sequence result in proteins with new properties