Molecular detection of virulence markers to identify diarrhoeagenic Escherichia coli isolated from Mula-Mutha river, Pune District, India.

In this study presence of virulence genes in multidrug resistant Escherichia coli isolated from Mula-Mutha river, Pune, India was undertaken. The objective was to understand whether the isolates were of diarrhoeagenic or of environmental origin. This was essential since the river flows through urban and rural parts of Pune and its water is used not only for industrial and agricultural purposes but also for domestic usage. One hundred and two multidrug E. coli isolates were selected from our previous study which detected genes coding for antibiotic resistance as well as identified integrons associated with multidrug resistance. Isolates were subjected to multiplex PCR to detect presence of virulence genes, set1A, set1B, sen astA, aggA, aafA, pet, stx1 and stx. Sequencing was performed to confirm the amplified PCR product. Seven of the 102 E. coli isolates showed gene set1A alone identifying them as Enteroaggregative E. coli. Thus, the findings revealed that majority of drug resistant E. coli were environmental in origin. The presence of antibiotic resistant genes, integrons in the environment as well as diarrhoeagenic E. coli isolates is a warning and calls for efficient public health measures to ensure that untreated sewage and industrial waste does not enter the Mula-Mutha river.

1H nuclear magnetic resonance-based metabolite profiling of guava leaf extract: an attempt to develop a prototype for standardization of plant extracts.

BACKGROUND: Herbal medicines are fast gaining popularity. However, their acceptability by modern practitioners is low which is often due to lack of standardization. Several approaches towards standardization of herbals have been employed. The current study attempted to recognize key peaks from 1H NMR spectra which together would comprise of a spectral fingerprint relating to efficacy of Psidium guajava (guava) leaf extract as an antidiarrhoeal when a number of unidentified active principles are involved. METHODS: Ninety samples of guava leaves were collected from three locations over three seasons. Hydroalcoholic (water and ethanol, 50:50) extracts of these samples were prepared and their 1H NMR spectra were acquired. Spectra were also obtained for quercetin, ferulic acid and gallic acid as standards. Eight bioassays reflecting different stages of diarrhoeal pathogenesis were undertaken and based on pre-decided cut-offs, the extracts were classified as 'good' or 'poor' extracts. The bioactivity data was then correlated with the 1H NMR profiles using Regression or Orthogonal Partial Least Square-Discriminant Analysis (OPLS-DA). RESULTS: OPLS-DA showed seasonal and regional segregation of extracts. Significant models were established for seven bioassays, namely those for anti-bacterial activity against Shigella flexneri and Vibrio cholerae, adherence of E. coli, invasion of E. coli and S. flexneri and production and binding of toxin produced by V. cholerae. It was observed that none of the extracts were good or bad across all the bioassays. The spectral analysis showed multiple peaks correlating with a particular activity. Based on NMR and LC-MS/MS, it was noted that the extracts contained quercetin, ferulic acid and gallic acid. However, they did not correlate with the peaks that segregated extracts with good and poor activity. CONCLUSIONS: The current study identified key peaks in 1H NMR spectra contributing to the anti-diarrhoeal activity of guava leaf extracts. The approach of using spectral fingerprinting employed in the present study can thus be used as a prototype towards standardization of plant extracts with respect to efficacy.

Psidium guajava: A Single Plant for Multiple Health Problems of Rural Indian Population.

The rural population in India faces a number of health problems and often has to rely on local remedies. Psidium guajava Linn. (guava), a tropical plant which is used as food and medicine can be used by rural communities due to its several medicinal properties. A literature search was undertaken to gauge the rural health scenario in India and compile the available literature on guava so as to reflect its usage in the treatment of multiple health conditions prevalent in rural communities. Towards this, electronic databases such as Pubmed, Science Direct, google scholar were scanned. Information on clinical trials on guava was obtained from Cochrane Central Register of Controlled Trials and Clinicaltrial.gov. The literature survey revealed that guava possesses various medicinal properties which have been reported from across the globe in the form of ethnobotanical/ethnopharmacological surveys, laboratory investigations and clinical trials. Besides documenting the safety of guava, the available literature shows that guava is efficacious against the following conditions which rural communities would encounter. (a) Gastrointestinal infections; (b) Malaria; (c)Respiratory infections; (d) Oral/dental infections; (e) Skin infections; (f) Diabetes; (g) Cardiovascular/hypertension; (h) Cancer; (i) Malnutrition; (j) Women problems; (k) Pain; (l) Fever; (m) Liver problems; (n) Kidney problems. In addition, guava can also be useful for treatment of animals and explored for its commercial applications. In conclusion, popularization of guava, can have multiple applications for rural communities.

Development of botanicals to combat antibiotic resistance.

The discovery of antibiotics in the previous century lead to reduction in mortality and morbidity due to infectious diseases but their inappropriate and irrational use has resulted in emergence of resistant microbial populations. Alteration of target sites, active efflux of drugs and enzymatic degradations are the strategies employed by the pathogenic bacteria to develop intrinsic resistance to antibiotics. This has led to an increased interest in medicinal plants since 25-50% of current pharmaceuticals are plant derived. Crude extracts of medicinal plants could serve as an alternate source of resistance modifying agents owing to the wide variety of secondary metabolites. These metabolites (alkaloids, tannins, polyphenols etc.) could act as potentials for antimicrobials and resistance modifiers. Plant extracts have the ability to bind to protein domains leading to modification or inhibition protein-protein interactions. This enables the herbals to also present themselves as effective modulators of host related cellular processes viz immune response, mitosis, apoptosis and signal transduction. Thus they may exert their activity not only by killing the microorganism but by affecting key events in the pathogenic process, thereby, the bacteria, fungi and viruses may have a reduced ability to develop resistance to botanicals. The article is meant to stimulate research wherein the cidal activity of the extract is not the only parameter considered but other mechanism of action by which plants can combat drug resistant microbes are investigated. The present article emphasizes on mechanisms involved in countering multi drug resistance.

Activity of Medicinal Plant Extracts on Multiplication of Mycobacterium tuberculosis under Reduced Oxygen Conditions Using Intracellular and Axenic Assays.

Aim. Test the activity of selected medicinal plant extracts on multiplication of Mycobacterium tuberculosis under reduced oxygen concentration which represents nonreplicating conditions. Material and Methods. Acetone, ethanol and aqueous extracts of the plants Acorus calamus L. (rhizome), Ocimum sanctum L. (leaf), Piper nigrum L. (seed), and Pueraria tuberosa DC. (tuber) were tested on Mycobacterium tuberculosis H37Rv intracellularly using an epithelial cell (A549) infection model. The extracts found to be active intracellularly were further studied axenically under reducing oxygen concentrations. Results and Conclusions. Intracellular multiplication was inhibited ≥60% by five of the twelve extracts. Amongst these 5 extracts, in axenic culture, P. nigrum (acetone) was active under aerobic, microaerophilic, and anaerobic conditions indicating presence of multiple components acting at different levels and P. tuberosa (aqueous) showed bactericidal activity under microaerophilic and anaerobic conditions implying the influence of anaerobiosis on its efficacy. P. nigrum (aqueous) and A. calamus (aqueous and ethanol) extracts were not active under axenic conditions but only inhibited intracellular growth of Mycobacterium tuberculosis, suggesting activation of host defense mechanisms to mediate bacterial killing rather than direct bactericidal activity.

Implementing Perennial Kitchen Garden Model to Improve Diet Diversity in Melghat, India.

Lack of diet diversity causing micronutrient deficiency is common in developing countries and is gaining attention due to the hidden consequences of impaired physical and cognitive development. This paper describes the propagation of a sustainable perennial kitchen garden (KG) model to address household (HH) diet diversity in Melghat. Nutrient dense plants, comprising of minimum one tree (perennial) and one green leafy vegetable (GLV) were given to participating HHs along with qualitative interventions. Baseline survey was conducted in winter 2011 followed by seasonal surveys over 2 years to record changes in KG practices, dietary intake and childcare practices. Marked increase from 4% at baseline to 95% at endline was seen in the KG maintainance. Increased diversity was seen in all food categories other than cereals and pulses. Variety of GLVs consumed increased over the two winters as well as the 2 summers. However, no change in the quantity of GLV consumed was noted which was attributed to the duration of the study period being insufficient for the trees to grow and provide adequate leaves for consumption. Notably, livelihood component was not promoted and HHs were encouraged to harvest and distribute excess seeds to relatives and neighbours. The study generated huge demand from HHs within the intervention and neighbouring villages. It concludes that a well designed perennial KG along with imparting adequate knowledge can be a sustainable practice to increase diet diversity and GLV intake which would help address micronutrient deficiencies in the community.

Approaches in fostering quality parameters for medicinal botanicals in the Indian context.

India is among the important megabiodiversity centers of the world with nearly 45,000 known plant species. This diversity coupled with a rich heritage of traditional knowledge has made India a home to several important time-honored systems of health care such as Ayurveda, Siddha and Unani. Herbal medicines, however, are associated with a number of shortcomings including uniform efficacy and lack of appropriate quality control measures at various stages of product development. The review intends to outline the importance of fostering quality parameters towards standardization and manufacturing of botanicals for India to emerge as a leader in global market of herbal products. Literature survey was carried out on important parameters for processing and manufacturing of botanicals. The review highlights that there have been constant efforts for developing state of the art technologies in the field of herbal research. It also reflects that Government authorities have also taken a number of initiatives to formulate appropriate guidelines from standardization of raw materials to obtaining botanical products. However, in the Indian context, there exist certain lacunae in the current regulatory mechanisms which need to be strengthened and stringently implemented to ensure safety, purity and efficacy of herbal medicines. Towards this the approaches being developed globally can be adopted. Based on the literature reviewed, in our opinion, four areas viz., benefit sharing, investment by industry, standardization and national/international networking structure need immediate attention for strengthening Traditional Systems of Medicine in India.

Possible causes of malnutrition in Melghat, a tribal region of Maharashtra, India.

Melghat, situated in Amravati District of Maharashtra, India is a tribal region with amongst the highest numbers of malnutrition cases. This paper focuses on possible causes of malnutrition in the Dharni block of Melghat. Quantitative survey recorded the existing burden of malnutrition, kitchen garden (KG) practices, Public Distribution System, food provisioning, Anganwadi services and hygiene/sanitation in the community. Additionally a qualitative study was undertaken to understand the community's perspective on nutrition, cultural beliefs, spending habits and other factors contributing to malnutrition. Malnutrition was found to be highly prevalent amongst all age groups with 54% children aged 1-5 years and 43% adults aged ≥ 20 years being severe to moderately underweight. A major cause for malnutrition in children was faulty child care practices. Data on food provisioning revealed that while the caloric needs of the community were substantially met by consumption of cereals and pulses, minimal consumption of green leafy vegetables (GLVs) could lead to micronutrient deficiency in the community. KGs, which provide GLVs, were mainly cultivated in monsoon (98%) which declined to merely 4% in summer. The benefits of government schemes though targeted at malnourished children were often shared by the entire household and thus got diluted. Key finding was that nutrition interventions should be designed to address the entire household and emphasis should be given to appropriate nutrition education, without which distributing food or increasing income would have minimal effect.

Bactericidal Effect of Selected Antidiarrhoeal Medicinal Plants on Intracellular Heat-Stable Enterotoxin-Producing Escherichia coli.

Diarrhoeal diseases due to enterotoxigenic Escherichia coli continue to be a cause of global concern. Medicinal plants have been gaining popularity as promising antidiarrhoeal agents. In the present study, four antidiarrhoeal plants, viz. Aegle marmelos, Cyperus rotundus, Psidium guajava and Zingiber officinale were screened against a heat-stable toxin-producing enterotoxigenic E. coli strain. Decoctions of these plants were studied for their effect on intracellular killing of the bacterial strain using murine monocytic cell line, J774. [(3)H] thymidine release assay was used to evaluate the apoptotic/necrotic effect. All plants at concentrations <1% enhanced intracellular killing of the bacteria by J774 cells. However, at higher concentrations, the decoctions induced apoptosis in J774 cells. The study demonstrates that these plants could control diarrhoea caused by heat-stable toxin-producing enterotoxigenic E. coli through their immunomodulatory effect.

Studies on the activity of Cyperus rotundus Linn. tubers against infectious diarrhea.

To study the antidiarrheal activity of the decoction of Cyperus rotundus Linn. tubers using representative assays of diarrheal pathogenesis and understand its mechanism of action.Antibacterial, antigiardial and antirotaviral activities were studied. Effect on adherence of enteropathogenic Escherichia coli (EPEC) and invasion of enteroinvasive E. coli (EIEC) and Shigella flexneri to HEp-2 cells was evaluated as a measure of effect on colonization. Effect on enterotoxins such as enterotoxigenic E. coli (ETEC) heat labile toxin (LT), heat stable toxin (ST) and cholera toxin (CT) was also assessed. The decoction showed antigiardial activity, reduced bacterial adherence to and invasion of HEp-2 cells and affected production of CT and action of LT. The decoction of C. rotundus does not have marked antimicrobial activity and exerts its antidiarrheal action by mechanisms other than direct killing of the pathogen.

Sacred Groves of Parinche Valley of Pune District of Maharashtra, India and their Importance.

Sacred groves are protected areas of forests because of religious beliefs and constitute an important aspect of the cultural life of various communities throughout the world. The source of this culture can be traced back to prehistoric times. The sacred groves of Parinche valley, Pune district of Maharashtra, India were studied to understand their status and importance in relation to common natural resource bases and cultural aspects. Through formal interviews and non-participatory observation undertaken in all seasons between October 2002 and February 2004, 14 sacred groves were documented. Both EMIC and ETIC dimensions were incorporated in the choice of subjects. The results show that they are important for various communities as a common natural resource base. They promote cultural bonding among various communities and maintain harmony in social life. This can be evidenced during annual village festivals and family celebrations, which invariably take place within the premises of sacred groves. They are also important for biodiversity as many groves function like mini sanctuaries and harbour endemic and threatened plant species. The natural vegetation of the groves comprises many medicinal plants that are regularly harvested by local medicine men ( Vaidus ). Water is a precious resource, especially during summer months, and the perennial water sources that are present in the sacred groves are the only remaining water sources for seven villages. There is a compunction, on the other hand, of the sacred groves being influenced by urbanization, resulting in the building of concrete temples by clearing natural vegetation and planting ornamental and other economically useful trees in the cleared areas of the groves. Such actions alter these fragile ecosystems and deplete or destroy the natural resource base for which this ancient culture is believed to have been originally evolved. We conclude here that sacred groves were originally a common natural resource which were assimilated into the social and cultural life of villages in the Parinche valley. Hence their protection and enhancement should become part of village and forest planning.

Mechanisms involved in peripheral nerve damage in leprosy with special reference to insights obtained from in vitro studies and the experimental mouse model.

The histopathological observations of Khanolkar and Iyer, that M. leprae has a predeliction for nerves, first highlighted the central role of peripheral nerves in the pathology of leprosy. It is now well recognized that nerve damage in leprosy will still continue to be an important problem in control and rehabilitation despite the presence of more efficient therapy. The multiplicity of mechanisms postulated, identified, and demonstrated in the last three decades has received little recognition from the scientific community at large. This review is therefore an attempt to collate these multiple studies on mechanisms of nerve damage into a cohesive analysis, which would facilitate the design of future studies. The objective of this review is to focus therefore only on studies which serve to illustrate mechanisms of nerve damage.