Bacteriophage therapy of human-restricted Salmonella species-a study in a surrogate bacterial and animal model.

Salmonella Typhi has been an important human-restricted pathogen from time immemorial, and unfortunately, the indiscriminate use of antibiotics has induced the emergence of multidrug resistance in S. Typhi. Bacteriophage therapy may be a possible alternative in countering antimicrobial resistance. Therefore, this study was planned to assess the efficacy of bacteriophages in treating acute and chronic S. Typhimurium infection in the mouse as a surrogate model. We isolated bacteriophages against S. Typhimurium and selected three different bacteriophages for the in vivo experiments. The lethal dose of S. Typhimurium was decided for Swiss albino mice, and acute infection was developed. Further, bacteriophage therapy by daily intraperitoneal injection of phage cocktail was given for 14 days. While the chronic carrier state of S. Typhimurium in Swiss albino mice was developed by inoculating intraperitoneally sequential 10-fold increasing doses of the bacterium. On the successful establishment of carrier state, oral feeding of phage cocktail at a high count was given, which completely cured the carrier state within 7 days of feeding. These experiments confirmed that the phage cocktail could eradicate the S. Typhimurium from the mice in both types of infections, that is acute and chronic.

Bacteriophages: A possible solution to combat enteropathogenic Escherichia coli infections in neonatal goats.

Due to awareness and benefits of goat rearing in developing economies, goats' significance is increasing. Unfortunately, these ruminants are threatened via multiple bacterial pathogens such as enteropathogenic Escherichia coli (EPEC). In goat kids and lambs, EPEC causes gastrointestinal disease leading to substantial economic losses for farmers and may also pose a threat to public health via the spread of zoonotic diseases. Management of infection is primarily based on antibiotics, but the need for new therapeutic measures as an alternative to antibiotics is becoming vital because of the advent of antimicrobial resistance (AMR). The prevalence of EPEC was established using bfpA gene, uspA gene and Stx1 gene, followed by phylogenetic analysis using Stx1 gene. The lytic activity of the isolated putative coliphages was tested on multi-drug resistant strains of EPEC. It was observed that a PCR based approach is more effective and rapid as compared to phenotypic tests of Escherichia coli virulence. It was also established that the isolated bacteriophages exhibited potent antibacterial efficacy in vitro, with some of the isolates (16%) detected as T4 and T4-like phages based on gp23 gene. Hence, bacteriophages as therapeutic agents may be explored as an alternative to antibiotics in managing public, livestock and environmental health in this era of AMR.

Evaluation of highly conserved hsp65-specific nested PCR primers for diagnosing Mycobacterium tuberculosis.

OBJECTIVE: To evaluate the sensitivity and specificity of a new nested set of primers designed for the detection of Mycobacterium tuberculosis complex targeting a highly conserved heat shock protein gene (hsp65). DESIGN: The nested primers were designed using multiple sequence alignment assuming the nucleotide sequence of the M. tuberculosis H37Rv hsp65 genome as base. Multidrug-resistant Mycobacterium species along with other non-mycobacterial and fungal species were included to evaluate the specificity of M. tuberculosis hsp65 gene-specific primers. RESULTS: The sensitivity of the primers was determined using serial 10-fold dilutions, and was 100% as shown by the bands in the case of M. tuberculosis complex. None of the other non M. tuberculosis complex bacterial and fungal species yielded any band on nested polymerase chain reaction (PCR). The first round of amplification could amplify 0.3 ng of the template DNA, while nested PCR could detect 0.3 pg. CONCLUSION: The present hsp65-specific primers have been observed to be sensitive, specific and cost-effective, without requiring interpretation of biochemical tests, real-time PCR, sequencing or high-performance liquid chromatography. These primer sets do not have the drawbacks associated with those protocols that target insertion sequence 6110, 16S rDNA, rpoB, recA and MPT 64.

Antimicrobial assay and genetic screening of selected freshwater Cyanobacteria and identification of a biomolecule dihydro-2H-pyran-2-one derivative.

AIMS: Explorations of freshwater Cyanobacteria as antimicrobial (bacteria, fungi and methicillin-resistant Staphylococcus aureus (MRSA) strains) drug resource using bioassay, NRPS (non-ribosomal polypeptide synthetase) and PKS (polyketide synthase) genes, as well as in silico approach. METHODS AND RESULTS: We have bioassayed the extracts of Phormidium CCC727, Geitlerinema CCC728, Arthrospira CCC729, Leptolyngbya CCC732, Phormidium CCC730, Phormidium CCC731 against six pathogenic bacteria comprising Gram (+ve): S. aureus including seven clinical MRSA and Enterococcus faecalis, Gram (-ve): Escherichia coli, Salmonella Typhimurium, Klebsiella pneumoniae and Shigella boydii along with non-pathogenic Enterobacter aerogenes as well as fungal strains (Cryptococcus neoformans and Candida albicans, C. krusei, C. tropicalis and Aspergillus niger) exhibiting antimicrobial potential. The NRPS and PKS genes of the target strains were also amplified and sequenced. The putative protein structures were predicted using bioinformatics approach. CONCLUSION: PKS gene expression indicated ß keto-acyl synthase as one of the important active domains in the biomolecules related to antitumour and antifungal group. The simultaneous identification of the biomolecule (dihydro-2H-pyran-2-one derivative) was also inferred spectroscopically. SIGNIFICANCE AND IMPACT OF THE STUDY: Freshwater Cyanobacteria are prolific producers of secondary metabolite(s) that may act as the antimicrobial drug resource in addition to their much explored marine counterpart.

Flow behind an exponential shock wave in a rotational axisymmetric perfect gas with magnetic field and variable density.

A self-similar model for one-dimensional unsteady isothermal and adiabatic flows behind a strong exponential shock wave driven out by a cylindrical piston moving with time according to an exponential law in an ideal gas in the presence of azimuthal magnetic field and variable density is discussed in a rotating atmosphere. The ambient medium is assumed to possess radial, axial and azimuthal component of fluid velocities. The initial density, the fluid velocities and magnetic field of the ambient medium are assumed to be varying with time according to an exponential law. The gas is taken to be non-viscous having infinite electrical conductivity. Solutions are obtained, in both the cases, when the flow between the shock and the piston is isothermal or adiabatic by taking into account the components of vorticity vector. The effects of the variation of the initial density index, adiabatic exponent of the gas and the Alfven-Mach number on the flow-field behind the shock wave are investigated. It is found that the presence of the magnetic field have decaying effects on the shock wave. Also, it is observed that the effect of an increase in the magnetic field strength is more impressive in the case of adiabatic flow than in the case of isothermal flow. The assumption of zero temperature gradient brings a profound change in the density, non-dimensional azimuthal and axial components of vorticity vector distributions in comparison to those in the case of adiabatic flow. A comparison is made between isothermal and adiabatic flows. It is obtained that an increase in the initial density variation index, adiabatic exponent and strength of the magnetic field decrease the shock strength.

Curative effect of Terminalia chebula extract on acetic acid-induced experimental colitis: role of antioxidants, free radicals and acute inflammatory marker.

The present study has evaluated the healing effects of extract of dried fruit pulp of Terminalia chebula (TCE) on acetic acid (AA)-induced colitis in rats. TCE (600 mg/kg) showed healing effects against AA-induced colonic damage score and weight when administered orally daily for 14 days. TCE was further studied for its effects on various physical (mucus/blood in stool and stool frequency, food and water intake and body weight changes), histology, antibacterial activity and free radicals (NO and LPO), antioxidants (SOD, CAT and GSH) and myeloperoxidase in colonic tissue. Intra-colonic AA administration increased colonic mucosal damage and inflammation, mucus/bloody diarrhoea, stool frequency, but decreased body weight which were reversed by TCE and sulfasalazine (SS, positive control) treatments. TCE showed antibacterial activity and both TCE and SS enhanced the antioxidants, but decreased free radicals and myeloperoxidase activities affected in acetic acid-induced colitis. TCE indicated the presence of active principles with proven antioxidants, anti-inflammatory, immunomodulatory, and free radical scavenging and healing properties. Thus, TCE seemed to be safe and effective in healing experimental colitis.

Observation on integron carriage among clinical isolates of Klebsiella pneumoniae producing extended-spectrum beta-lactamases.

PURPOSE: Klebsiella pneumoniae is considered an important pathogen causing nosocomial and community-acquired infections and is often associated with the production of extended-spectrum beta-lactamases (ESBL) belonging to SHV and CTX-M families, which are frequently described as a part of complex integrons, facilitate their horizontal transfer to other related as well as unrelated microbes. The present study was undertaken to investigate the occurrence and characterization of integrons among K pneumoniae isolates producing ESBL in a tertiary referral hospital. MATERIALS AND METHODS: A total of 136 clinical isolates of K pneumoniae were investigated for the presence of ESBL. Their ESBL genes were characterized by multiplex polymerase chain reaction (PCR). Integrase gene PCR was performed to detect the presence of integron. The isolates were further typed by random amplification of polymorphic DNA (RAPD). RESULT: Out of 136 K pneumoniae isolates, 63 (46%) were confirmed to be ESBL producers. SHV (68%) and CTX-M (67%) ESBL genes were the most common in our study. Of the 63 ESBL-positive isolates, 58 (92%) strains carried integrons; 52 strains (82%) carried only class 1 integron, whereas 6 (9%) isolates harboured both class 2 integrons and the class 1 gene. However, in ESBL negatives, only 29 (40%) strains were positive for class 1 integron and none for class 2 integron. CONCLUSION: The presence of class 2 integron amongst ESBL-producing K pneumoniae is being described for the first time in this part of the world. The findings of this study strongly suggest that integrons have a role in the dissemination of ESBL-mediated resistance among the nosocomial isolates of K pneumonia.

Preparation and in vitro antibacterial evaluation of gatifloxacin mucoadhesive gellan system.

BACKGROUND AND THE PURPOSE OF THE STUDY: The poor bioavailability and therapeutic response exhibited by the conventional ophthalmic solutions due to precorneal elimination of the drug may be overcome by the use of mucoadhesive in situ gel forming systems that are instilled as drops into the eye and undergo a sol-gel transition in the cul-de-sac and have good mucoadhesion with ocular mucus layers. The objective of this study was to formulate ophthalmic mucoadhesive system of gatifloxacin (GTN) and to evaluate its in vitro antibacterial potential against, Staphylococcus aureus and Escherichia coli. METHODS: : Mucoadhesive systems were prepared using gellan combined with sodium carboxymethylcellulose (NaCMC) or sodium alginate to enhance the gel bioadhesion properties. The prepared formulations were evaluated for their gelation, and rheological behaviors, mucoadhesion force, in vitro drug release, and antibacterial activity. RESULTS: All formulations in non-physiological or physiological conditions showed pseudoplastic behaviors. Increase in the concentration of mucoadhesive agent enhanced the mucoadhesive force significantly. In vitro release of gatifloxacin from the mucoadhesive system in simulated tear fluid (STF, pH of 7.4) was influenced significantly by the properties and concentration of gellan, sodium carboxymethyl cellulose and sodium alginate. Significant reduction in the total bacterial count was observed between drug solution (control) and mucoadhesive batches against both tested organisms. MAJOR CONCLUSION: The developed mucoadhesive system is a viable alternative to conventional eye drops of GTN due to its ability to enhance bioavailability through its longer precorneal residence time and ability to sustain the release of the drug.

Synthesis, anti-bacterial and anti-fungal activities of some novel Schiff bases containing 2,4-disubstituted thiazole ring.

A series of arylidene-2-(4-(4-methoxy/bromophenyl) thiazol-2-yl) hydrazines (4a-z) and 1-(4-(4-methoxy/bromophenyl) thiazol-2-yl)-2-cyclohexylidene/cyclopentylidene hydrazines (5a-b/6a-b) were synthesized, characterized and screened for their antimicrobial activities. The structures of synthesized compounds were established by spectroscopic (FT-IR, (1)H NMR, (13)C NMR, Mass) and elemental analyses. Both the anti-bacterial and anti-fungal activities with MIC values of compounds were evaluated. The results of anti-bacterial screening reveal that among all the compounds screened eight compounds showed moderate to good anti-bacterial activity while ten of the newly synthesized compounds displayed good to excellent anti-fungal activity. Among the tested compounds, the most effective compounds with MIC value in the range of 6.25-25 microg/ml are 4a, 4n, 4z, 5a, 5b, 6a and 6b against three fungal strains viz. Candida albicans, Cryptococcus neoformans and Aspergillus flavus.

Evaluation of polymerase chain reaction and adenosine deaminase assay for the diagnosis of tuberculous effusions in children.

AIM: To evaluate and compare the utility of polymerase chain reaction (PCR) for the diagnosis of tuberculous effusions in children. METHODS: PCR, adenosine deaminase (ADA) activity and absolute lymphocyte count (ALC) were evaluated in the fluid of 31 tuberculous (20 pleural, 8 ascites and 3 pericardial) and 24 non-tuberculous (10 transudtative ascites, 8 empyema thoracis, 3 malignant pleural and 3 pyopericardium) effusions. RESULTS: Fluid PCR for Mycobacterium tuberculosis was positive in 74% of tuberculous effusions, whereas it was falsely positive in 13% of the non-tuberculous group. The mean fluid ADA and ALC values were significantly higher in tuberculous effusions than in non-tuberculous effusions (p<0.001). The sensitivity and specificity of PCR, ADA (> or =38 IU/l) and ALC (> or =275/mm3) were 74% and 88%, 81% and 75%, and 90% and 83%, respectively, in diagnosing tuberculous effusions. The sensitivity of PCR, ADA and ALC was 100%, 100% and 88%, respectively, for confirmed tuberculous effusions. When the two tests were combined (either/or positive), the sensitivity increased (90-100%) at the expense of specificity. When both the tests were positive, then the specificity markedly increased (92-96%), but sensitivity of the tests decreased. CONCLUSION: Fluid PCR alone should not be relied on as a single test; rather, combined analysis with either ADA or ALC could be more useful in the diagnosis of tuberculous effusions in children.

Unprecedented control over polymerization of n-hexyl isocyanate using an anionic initiator having synchronized function of chain-end protection.

The living anionic polymerization of isocyanates carried out using conventional initiators is associated with side reactions due to rapid initiation rates as well as back-biting by the growing chain, resulting in a lack of control on the molecular weight (MW) and molecular weight distribution (MWD) of the polymers. Successful control over the reaction was possible by using additives that could prevent back-biting. We find an initiator in sodium benzanilide (Na-BA), which has a slow initiation rate combined with additive function, so that use of an external additive is eliminated. The initiator has resulted in polymers with high yields and an unprecedented control over the MW and MWD. It is possible to introduce a number of functionalities at the termini of the polymer by using Na-BA derivatives as well as suitable terminating agents, leading to macromonomer, reactive and chiral polymers, and chiral macromonomer in approximately 100% yields. In the process, the finding has expanded the scope of polyisocyanates in diverse applications.

Immune status with empyema thoracis.

OBJECTIVE: To evaluate the humoral and cell mediated immune status of children with empyema thoracis. METHODS: Serum IgG, IgA, IgM, Complement C3 assay and cell mediated immunity (CMI) tests were performed in 33 patients of empyema thoracis, and 14 healthy age matched controls. RESULTS: The mean serum IgG and IgA levels in empyema thoracis and its subgroups were significantly raised as compared to controls. The overall values of IgG and IgA were 104% (p<0.001) and 114% (p<0.01) of normal mean, respectively. The mean serum IgM and complement C3 levels did not differ significantly in both the groups. The frequency of negative skin reaction to purified protein derivative (PPD) was significantly higher in children with empyema thoracis as compared to controls (p<0.05). The mean absolute lymphocyte count (ALC) was significantly decreased and serum adenosine deaminase (ADA) activity was significantly raised in empyema thoracis in comparison to controls. The overall ALC was 76.1% (p<0.01) and serum ADA activity was 169.4% (p<0.001) of normal mean, respectively. No significant differences were observed in the mean levels of immunoglobulins, complement C3 and CMI tests between pyothorax and pyopneumothorax and pleural fluid culture positive and negative cases. CONCLUSIONS: Thus, both humoral and cell mediated immunity were affected in empyema thoracis patients. However, CMI demonstrated more pronounced change in comparison to humoral immunity.

Helicobacter pylori infection and recurrent abdominal pain in children.

The association of Helicobacter pylori infection with recurrent abdominal pain (RAP) was studied in 65 children aged between 3 and 12 years. Cases with a specific cause for abdominal pain were excluded from the study. All the patients were subjected to upper gastrointestinal endoscopy and antral biopsy. The biopsy specimens were subjected to rapid urease test, histopathological examination, and culture to document the presence of H. pylori in the gastric mucosa. All the patients were put on H. pylori eradication drugs for 10 days. About 69 per cent of the children had histopathological evidence of antral inflammation. Helicobacter pylori detection rate was 77 per cent. About 83 per cent of patients had complete symptomatic relief when followed over a period of 6 months. The study concludes a possible association of H. pylori with recurrent abdominal pain in children and all these patients should be considered for H. pylori eradication.

Significance of Cryptosporidium as an aetiology of acute infectious diarrhoea in elderly Indians.

OBJECTIVE: To explore the significance of Cryptosporidium isolation in elderly Indian with acute infectious diarrhoea and its clinical correlates. DESIGN: A hospital-based clinico-aetiological study of 120 patients aged 60 years or older, 25 adults younger than 60 years and 25 children up to age 14 with acute diarrhoea, and 57 apparently healthy elderly individuals. RESULTS: Cryptosporidium was isolated in 22 older patients with diarrhoea (18.3%) and was highly significant (P < 0.01) compared with healthy age-matched controls. Of these patients 66% had a history of close contact with animals. Most (68%) Cryptosporidium infections occurred during the rainy season. Among the elderly patients 17% suffered from vomiting and abdominal pain, 31% were febrile; none were severely dehydrated. Stools numbered three to nine per day with duration of 5-17 days. Stool leucocytes were <6/hpf and no RBCs were seen. Isolation of Cryptosporidium in older persons was associated with diabetes mellitus (22.7%), tuberculosis (9.0%), malignancy (4.5%) and coronary artery disease (4.5%). CONCLUSION: Cryptosporidium is an important cause of diarrhoea in elderly Indians, especially those with close contact with animals. The infection has a mild clinical course, is self-limiting and does not cause dysenteric stool. Its effect on nutrition requires further study.

Prevalence of methicillin resistant Staphylococcus aureus in a tertiary referral hospital in eastern Uttar Pradesh.

We report the prevalence of methicillin resistant Staphylococcus aureus (MRSA) infections and their antibiotic susceptibility pattern in our hospital located in eastern Uttar Pradesh. Out of total 549 strains of Staphylococcus aureus isolated from different clinical specimens 301 (54.85%) were found to be methicillin resistant. More than 80% of MRSA were found to be resistant to penicillin, cotrimoxazole, ciprofloxacin, gentamicin, erythromycin, tetracycline, 60.5% to amikacin and 47.5% to netilmicin. However, no strains were resistant to vancomycin. Many MRSA strains (32.0%) were multi-drug resistant. To reduce the prevalence of MRSA, the regular surveillance of hospital associated infection, monitoring of antibiotic sensitivity pattern and formulation of definite antibiotic policy may be helpful.

Correlation of Helicobacter pylori and gastric carcinoma.

BACKGROUND: Difference of opinion about the prevalence of H. pylori association with gastric cancer exists in the literature. AIMS: To study the correlation of Helicobacter pylori (H. pylori) to gastric carcinoma. METHODS: 50 proved cases of gastric cancer were studied by rapid urease test, culture, histopathology and ELISA test for H. pylori IgG. RESULTS: 68% of cases of gastric cancer were found to be positive for H. pylori infection as compared to 74% of healthy controls. CONCLUSIONS: The prevalence rate of H. pylori infection in our patients of gastric cancer was lower than in the control population though statistically not significant, suggesting that H. pylori may not be responsible for gastric carcinogenesis in this population.