Evaluation of Pathogenic Potentialities and Transmissibility of Aerobic and Anaerobic Bacteria in Sexually Transmitted Diseases.

Aims: Sexually transmitted diseases (STDs) caused by various aerobic and anaerobic bacteria have been reported from many developed and developing countries of the world. However, there is limited data available on the association of these pathogens with STDs on the Indian sub-continent. Therefore, the aim of this is to the presence of anaerobic and aerobic bacteria in sexually transmitted infections. Study Design: Patients attending the Department of Dermatology, Venereology & Leprology, Calcutta Medical College & Hospital, suspected to be suffering from STDs, were thoroughly examined and those having typical lesions of chancroid were excluded Original Research Article British Journal of Medicine & Medical Research, 4(27): 4533-4541, 2014 4534 from further work. The prevalence of different aerobic and anaerobic bacteria was determined from among the remaining patients after ruling out cases of chancres. Place and Duration of Study: This work was carried out in the Department of Oral Medicine, R. Ahmed Dental College; Department of Dermatalogy, Venereology & Leprology, Calcutta Medical College and Hospital; Division of Microbiology, Department of Pharmaceutical Technology, Jadavpur University and Department of Microbiology, Herbicure Healthcare Bio-Herbal Research Foundation, Kolkata, for a period of seven months from April 1 to October 31, 2012. Methodology: At least two smears were prepared from the infected ulcers of each of the patients who were not diagnosed as of chancroid. One smear was for dark ground microscopy and the other for Gram’s staining. Confirmation of chancre was by serological testing, while aerobes/anaerobes were identified following standard procedures. Results: The organisms isolated were Staphylococcus aureus, Shigella flexneri, Shigella sonnei, Gardenerella vaginalis, Actinomyces spp, Veillonella purvula, Peptococcus heliotrinreducens, Peptostreptococcus magnus and Peptostreptococcus hydrogenalis. These were subjected to tests for their antibiotic sensitivity pattern which was followed by successful specific therapy. Conclusion: Various Gram positive and Gram negative aerobes and anaerobes were found to be associated with STDs and these were transmissible among homosexual and heterosexual partners.

Possibilities of Developing New Formulations for Better Skin Protection from a Traditional Medicinal Plant Having Potent Practical Usage.

Aims: Natural products obtained from different types of plants have made an important impact since ancient times throughout Asia. Since wood of Hesperethusa crenulata (Roxb.) Roem has been used traditionally by women in Southeast Asia for protection against sunlight and other harmful environmental factors, present study was aimed to determine antioxidant and antimicrobial properties of water extract of H. crenulata wood. Methodology: Wood was powdered, soaked in water, macerated, extracted fluid was lyophilized and tested for antioxidant and antimicrobial properties following standard techniques. The IC50 values were calculated from regression equations prepared from concentrations of extract and inhibition percent of free radical formation. The extract was screened for antimicrobial action following international guidelines against 44 organisms. Results: In DPPH assay system IC50 value was 344.74 μg/ml, the same for superoxide scavenging assay was 326.13μg/ml, while in hydroxyl radical scavenging system it was 999.85 μg/ml. Among 32 strains of staphylococci 10 strains were inhibited at 25 -50 μg /ml level of the extract, 6 strains at 100 μg /ml, 4 at 200 μg /ml and the remaining 12 strains were resistant. Strains of Bacillus spp and Vibrio vulnificus were rather sensitive to the extract, while E. coli and Pseudomonas aeruginosa were resistant. Thus the extract was inhibitory for bacteria known to invade human skin. Conclusions: The distinctly beneficial properties of the wood of H. crenulata indicate its usefulness as a highly active agent for the protection of skin against invasion by microorganisms and free radicals including those that may come from the sunlight.

Evaluation of in vitro and in vivo antibacterial activity of dobutamine hydrochloride.

PURPOSE: To determine the in vitro and in vivo antibacterial activity of a cardiovascular drug dobutamine hydrochloride. METHODS: The minimum inhibitory concentration (MIC) of dobutamine was determined both by agar and broth dilution methods against 331 strains of bacteria from three gram positive and 13 gram negative genera. The antibacterial action of dobutamine was further tested in animal models. RESULTS: Dobutamine was seen to possess powerful inhibitory action (5-200mg/mL) against most test bacteria in in vitro studies. It was bacteriostatic in nature. In vivo studies showed that the drug offered significant protection (p< 0.001) to mice challenged with a virulent bacterium. CONCLUSION: Dobutamine showed remarkable antibacterial property against several pathogenic bacteria. Its potential as an antibacterial agent may be confirmed after further pharmacological studies.

Leprosy bacillus--possibly the first chemoautotrophic human pathogen cultivated in vitro and characterised.

Leprosy bacillus (LB) and leprosy derived in vitro culture forms, the chemoautotrophic nocardioform (CAN) bacteria, showed an extremely close homology and identity with each other as regards a chemoautotrophic nutritional pattern, a nocardioform morphology, a weak acid-fastness coupled with Gram and Gomori's stain positivity, an exclusive mycolate and lipid profile, a phenolic glycolipid (PGL-I) and a highly sequestrated DNA characteristic, namely, a unique small size, a low G+C % mole, an exceptionally high gamma and UV radiation resistance, and a high thermal resistance. LB/CAN bacteria (CANb) gave positive signals for 36 kDa protein PCR, as well as, for 65 kDa epitope, and hybridisation with two or more probes and also by RFLP-analysis. Both LB/and CAN bacteria exhibited bacillary multiplication in the mouse footpads (MFP), nerve infiltration and evidences for local pathogenicity associated with pronounced systemic invasion. A highly reproducible mutilation model could be established which enabled a successful application of the postulates of Koch. The proof of their total identity was their anergic reactions in LL cases counterpoised against Mitsuda type strong nodular responses, mirroring the reactions of leprosy bacilli in TT cases, in accordance with the dictum of XIth International Leprosy Congress (1978). Thus, the chemoautotrophic nutritional requirements of LB, entirely unsuspected for a medically important pathogenic bacterium, having dimorphic (both bacillary and mycelial) characters with spores, mycelia and granules and unique pathogenicity of multilation manifested through the virulence factor, the enzyme collagenase, made LB or M leprae the highly enigmatic bacterium for so long.

Validity of mechanism of gene transfer in the process called conjugation in bacteria.

We have attempted a new evaluation of the process of conjugation in bacteria, because of some basic dissimilarities observed between this and that of eukaryotes, or plants and animals. Reference donor and recipient strains, widely used to prove conjugation in bacteria, were chosen; addition of DNase during the conjugation process, led to an unexpected but highly reproducible increase in the transconjugant colony counts (TCC; ca. > or = 1 log), when compared with that of the controls without DNase. Transconjugants were also obtained when the same live donors were substituted with the UV-killed ones although the TCC was very low initially. Contrarily, donors treated with DNA-intercalating agents, e.g. acridine orange or ethidium bromide, resulted in a complete failure to produce transconjugants. There was a quantitative relationship between the DNase used on donors and levels of DNA sugars/nucleotides/DNA, which possibly resulted from interaction between the DNase and DNA being present/produced on the donor surface. This may be indicative of what may actually happen in the donor-recipient mixtures in the conjugation test proper, where the recipient DNase may activate a donor DNA production cycle. The evidences presented did not suggest that the donor DNA in the conjugation process is actually vestibuled through any intercellular conjugation passages, and is susceptible to the action of DNase or the intercalating dyes.

Potentiality of tricyclic compound thioridazine as an effective antibacterial and antiplasmid agent.

Thioridazine (Th), which is therapeutically used in psychiatric patients, was found to possess conspicuous antimicrobial activity when tested against 316 strains belonging to a number of Gram positive and Gram negative bacteria. Although Staphylococcus aureus, Vibrio chloerae and V. parahaemolyticus were found to be most sensitive, Th was highly bactericidal against S. aureus and bacteriostatic for vibrios and other Gram negative organisms. In the study of antiplasmid/curing effect of Th on twelve multiply antibiotic and Th resistant bacteria, it was observed that elimination of R plasmids was facilitated by choice of optimal concentration of Th. Significant elimination of single and combined antibiotic resistance occurred in E. coli and Shigella flexneri and not in S. aureus.

Antibacterial activity of the antiinflammatory agent diclofenac sodium.

Antimicrobial property of ten antiinflammatory drugs was tested with eleven sensitive bacteria belonging to both Gram positive and Gram negative types. Since most of the bacteria were moderate to highly sensitive to diclofenac (Dc), this compound was tested in vitro against 397 bacteria, most of which were inhibited by Dc at 50-100 micrograms/ml level. When tested in vivo, Dc at 1.5 and 3.0 micrograms/g body weight of a Swiss strain of white mice, could significantly protect the animals challenged with 50 MLD of Salmonella typhimurium NCTC 74. According to chi 2 test the in vivo data were highly significant (P < 0.001).

Antimicrobial potentiality of a new beta-lactum antibiotic fosfomycin.

Antimicrobial action of penicillin and some of its derivatives including fosfomycin was studied with respect to 225 strains of Gram-positive and Gram negative bacteria. Fosfomycin was found to possess somewhat less activity against Staphylococcus aureus compared with other penicillins; however, it showed powerful activity towards Escherichia coli, Klebsiella spp. and Proteus mirabilis.

In vitro & in vivo experimental susceptibility of Mycobacterium marinum to augmentin.

The effect of augmentin alone and in combination with various beta-lactam antibiotics was studied against a pathogenic Mycobacterium, M. marinum. The in vitro studies did not reveal any additional advantage over that found with augmentin alone and this antibiotic seemed considerably inhibitory to M. marinum at < 1 microgram/ml concentration. In vivo, the effects of augmentin on experimentally produced lesions in the mouse foot pads (MFPs) showed a significant regression of the lesions, which was compatible with an early disappearance of M. marinum from the MFP, in contrast with those of the untreated, control animals.

Electron microscopic characteristics of actinomycetic agents having aetiological association with human leprosy and epizootic ulcerative syndrome of fish.

The transmission electron microscopic (TEM) studies of the human leprosy derived chemoautotrophic nocardio-form (CAN) bacteria and EUS derived CAN bacteria showed presence of double contoured cell-walls consisting of an electron transparent and a dense layer. The fibrillar structures on the surface of these CAN bacterial cells also suggested their similarity to the human tissue derived Mycobacterium leprae cells. These EM studies further revealed mycelial and coccoid bodies in all these bacteria as was observed originally.

Propagation of Mycobacterium lepraemurium on supplemented minimal medium and its experimental pathogenesis.

The splenic tissue of a mouse experimentally infected with M. lepraemurium (Hawaiian strain, M-65) and developing 'rat leprosy', yielded a pure culture of an acid - fast bacterium having all the characteristics of M. lepraemurium on mineral salt minimal medium supplemented with simple sources of C and N, e.g., NH4 -salts, liquid paraffin, urea, gelatin etc. This could be maintained, by serial passages in vitro with good growth. Its indefinite propagation with tissue - free washed, small inoculum on complex media including Ogawa medium was difficult, and its serial sub-culture was practically impossible. The in vitro isolate from supplemented minimal medium could produce pathological lesions in mice typical of rat leprosy.

Phage-mediated conjugation in staphylococci as a means of transfer of drug-resistance.

Penicillin resistance plasmid was transferred from Staphylococcus aureus B4 (PcrKms, donor) to S. aureus ML351 (PcsKmr, recipient) by co-cultivation of the donor with the recipient in nutrient broth with or without the modifying effects of CaCl2 or sodium dodecyl sulfate. It was found that the transfer of drug-resistance occurred maximally between 6 and 18 hr postinoculation; however, addition of DNase (200 micrograms/ml) could totally prevent such a transfer up to 6 hr and significantly reduce it thereafter. Cell-free filtrate of the donor culture when mixed with the recipient was ineffective in bringing about the transfer of Pcr.

A pathogenicity model for the agents of epizootic ulcerative syndrome (EUS) in fish.

Chemoautotrophic nocardioform (CAN) bacteria had been repeatedly isolated from fish with ulcerative disease syndromes (EUS) from the massive epizootics that had repeatedly occurred since 1988 in eastern India as the major or only pathogenic agent in the background of distinctive environmental and epizootic data. Since these isolates bear significant similarity to the human and rat leprosy bacilli, attempts had been made to demonstrate the pathogenicity of this fish pathogen in the "Swiss" strain of mice as a convenient model. The studies reveal that the fish CAN bacteria could produce pathogenic effects in mice similar to that of the rat leprosy bacillus.

Plasmid-determined degradative metabolism and halophilism of Vibrio parahaemolyticus.

A set of 25 Kanagawa(+) and Kanagawa(-) strains of V. parahaemolyticus was studied for their ability to degrade hydrocarbons in minimal media. All strains gave positive results with respect to crystal violet (CV), methyl violet, liquid paraffin, benzene, naphthalene and phenol. The CV double ring (CVDR) response had earlier appeared to be a significant pathogenic marker [Chakrabarti et al, Indian J Med Res, 85 (1987) 508]. The CVDR response was found also to be a biodegradative marker, and correlates perfectly well with polymyxin resistance and low level of halophilism (4% NaCl). All these markers (characters) were found to be controlled by a single plasmid in the wild type. Elimination of the plasmid, as confirmed by gel electrophoresis studies, resulted in loss of CVDR response, polymyxin resistance, and acquisition of halophilism at a higher level (> 7%). The massive drainage of industrial effluents, rich in hydrocarbons, in the estuarine areas in many countries might have altered the ecosystem in favour of V. parahaemolyticus and its emergence as a new biodegradative and enterotoxigenic pathogen, contaminating fauna and flora in the littoral sea regions, with increased changes of communicability to humans.

Viability of acid-fast bacilli from gamma-& UV-irradiated lepromatous armadillo tissues infected with Mycobacterium leprae.

Gamma-irradiated splenic homogenates of armadillos infected with M. leprae proved sterile by conventional tests and media. However, on media for chemoautotrophy, these could repeatedly grow as a single type of acid-fast nocardioform bacterium like the unirradiated specimens, although with a much reduced count. In the slide culture, transition from the initial AFB/coccoid bodies, to sporulating mycelia and granules in the final stage, could be observed sequentially. The gamma-irradiated tissue specimens failed to yield any other mycobacterium/corynebacterium tested according to standard protocols.

Leprosy-derived chemoautotrophic nocardioform (CAN) bacteria closely resemble, or are identical with, Mycobacterium leprae on mycolate and other lipid profiles.

On the basis of thin layer chromatography and gas-chromatography-mass spectrometric studies, the lipid profiles of all the chemoautotrophic nocardioform (CAN) bacteria derived from human and animal leprosy tissues appear to be identical with each other, and closest to or identical with the most probable profile of M. leprae.

Metabolism of silicon as a probable pathogenicity factor for Mycobacterium & Nocardia spp.

The role of silicon (Si) in metabolism and growth of 22 strains of mycobacteria and 3 strains of nocardiae, which were mostly pathogenic, was studied on Kirchner's medium solidified with sodium metasilicate (KSM) and the C-free solidified metasilicate minimal medium (SMM) consisting of mineral salts only. On KSM, initial growth of mycobacteria appeared to be better, compared with that on Lowenstein-Jensen medium (LJM), although subsequent growth on the former was slower. On SMM lacking C, growth of mycobacteria and nocardiae could be achieved, only after repeated passages. These findings indicate that the mycobacteria and nocardiae are able to utilise Si at least to a limited extent, possibly as an alternative to C, with greater chances of survival.