Exploring the gut microbiome of tribal population of West Bengal to identify the diversity of the microflora and possibly identify uniquely conserved genes
The gastrointestinal tract of human is inhabited by trillions of bacteria of diverse nature, composition of which varies among individuals within and between communities. The initial inoculumsare acquired maternally during birth or even in uteri. Subsequent colonisation of the gut depends upon several factors including diet, age and diseases. Although not properly understood, the bacterial communities exert phenotypic traits to the host by a complex network of interactions among them. Many of such interactions arising from the altered gut bacterial profiles (GBP) have been implicated to cause diseases in human. Moreover, modern lifestyle with altered GBP has been thought to cause inflammatory disorders in the western countries. To understand GBP of the diseased state, knowledge on GBP in the healthy state is also important. The GBP of several populations of the world, both with modern and traditional lifestyles have been studied in America, Europe, Africa, Korea and China. With its largest tribal population, India offers a unique scenario for such studies as diverse communities are still dependent on hunting, agriculture and fishing with their own culture, tradition, dietary habits, language and genetic make-up. Till now there is no such report on GBP of the ethnic tribes of India. With the rapid economic development coupled with modernization of the lifestyle, the tribal population of India may eventually undergo alteration in GBP causing a gap in our knowledge on GBP across the globe. The need of data on GBP of ethnic tribes has earlier been emphasised. Till date no study has focussed on the gut microbiome of West Bengal tribal population despite the fact that there are 40identified tribes in the state comprising of 5.8 per cent of the total population of the state as per 2011 Census. This project shall aim to identify the GBP of representative members of two tribes of West Bengal to generate an insight into their food habits and possibly identify unique genes in the bacterial members.
Virtual Screening of Natural Compounds against Shigella Invasion proteins for identifying potential leads for drug and vaccine design
Shigellosis is a type of food poisoning caused by infection with the Shigella species. It is a major public health problem in developing countries where sanitation is poor. Shigellosis is spread by means of fecal-oral transmission. It is one of the most common disorders specially affecting children in West Bengal. Infection is initiated by ingestion of shigellae.An early symptom, diarrhoea (possibly elicited by enterotoxins and/or cytotoxin), may occur as the organisms pass through the small intestine. The hallmarks of shigellosis are bacterial invasion of the colonic epithelium and inflammatory colitis. These are interdependent processes amplified by local release of cytokines and by the infiltration of inflammatory elements. Colitis in the rectosigmoid mucosa, with concomitant malabsorption, results in the characteristic sign of bacillary dysentery: scanty, unformed stools tinged with blood and mucus.Shigellosis occurs worldwide, and it tends to occur whenever war, natural calamities (eg, earthquakes, floods), or unhygienic living conditions result in overcrowding and poor sanitation. S boydii and S dysenteriae occur more commonly internationally. Disease from Shigella species causes an estimated 1 million deaths and 165 million cases of diarrhea annually worldwide. Severe dysentery is treated with ampicillin, trimethoprim-sulfamethoxazole, or, in patients over 17 years old, a 4-fluorquinolone such as ciprofloxacin. Vaccines are not currently available, but some promising candidates are being developed.This project aims to identify potential natural compounds as leads for drug discovery trials through the process of virtual screening; as a method for management of the disease and arm the human race with a potential cure for this fatal disease.
Prediction and Design of Antisense Oligonucleotides against tRNA derived fragments (tRFs) and small RNAs implicated in various diseases and development of a Antisense Oligonucleotide repository- “ASObase”
The relevance of the non-coding genome to human disease has mainly been studied in the context of the widespread disruption of microRNA (miRNA) expression and function that is seen in human cancer. However, we are only beginning to understand the nature and extent of the involvement of non-coding RNAs (ncRNAs) in disease. Other ncRNAs, such as PIWI-interacting RNAs (piRNAs), small nucleolar RNAs (snoRNAs), tRNA derived microRNAs and long non coding RNAs are emerging as key elements of cellular homeostasis. Along with microRNAs, dysregulation of these ncRNAs is being found to have relevance not only to tumorigenesis, but also to neurological, cardiovascular, developmental and other diseases. tRNAs are characterized by their evolutionarily conserved secondary and tertiary structures, extensive post-transcriptional processing and modifications (up to 100 nucleoside modifications have been described), extreme stability and resistance to nucleases. Moreover, the human genome contains more than 500 genes encoding over 40 different tRNAs. Identification of novel ncRNAs derived from tRNAs, known as tRNA-derived RNA fragments, has recently gained significant attention. Some of these fragments are derived from precursor tRNA molecules, others from mature cytoplasmic tRNAs. The list of possible functions of tRNA-derived fragments is growing. The recent discovery that the abundance of 5'-tRNA halves found in sperm and oocytes rapidly decreases upon fertilization suggests that this class of molecules can be physiologically regulated. Similarly, 5'-tRNA halves and 5'-tRFs are found in exosomes within semen. tRNAs and tRNA fragments has been reported as novel sources of piRNAs (PIWI-interacting RNAs) in both mouse gametes and zygotes as well as in human somatic cells. The functions of these tRNA-derived piRNAs are not known but may be connected to epigenetic inheritance, silencing of retrotransposons and other genetic elements or post-transcriptional regulation of mRNAs. Finally, tRNA fragments are also induced by viral infections such as by Respiratory Syncytial Virus (RSV).
Thus as we attempt to quantify the functions of these small molecules in diseases, it is imperative to devise plan of actions to regulate, control and suppress their activities, such that the onset of diseases can be prevented. This project aims to explore the efficacy of antisense oligonucleotides as a mechanism for controlling the function of these non coding small molecules. Both wet lab and computational techniques would be explored for mining, predicting and validating the interactions of the designed antisense oligonucleotides to their respective targets as well as identify the efficacy of these interactions through RNA sequencing.
Metagenomic Analyses for understanding the soil – plant continuum by exploring the rhizospheric and root endophytic microflora of Heritiera fomes and Nypa fruticans from Indian Sunderbans
Over the years numerous studies have made an effort to understand the causes behind the depletion of the mangrove cover along the coastal lines. However, very less emphasis has been given to the microbial population and the alterations in the metapopulation dynamics of the microbial communities prevalentalong the mangrove forest areas. Metagenomics allows the easy identification of the various microbial communities. A metagenomic study of the mangrove region with its rhizospheric and endophytic communitieswould be undertaken to study the soil – plant continuum prevalent in the mangrove forests of the eastern coasts focussing on Heritiera fomes (Sundari) and Nypa fruticans (Golpata); the two of the most abundant and signature species of the Indian Sunderbans. RNA would be isolated from the soil samples and Illumina sequencing chemistry would be used to amplify the V3 - V4 region leading to taxonomic identification of the bacterial members. Corresponding root samples would also be collected to explore the endophytic members as well. Hence understanding of their physiological continuum would enable proper conservation strategies to be developed. The project shall attempt to identifythe total microflora that constitute the rhizospheric and root endophytic strata of the plants under study to predict prospective biomarkers and assess the alterations in microbial community pattern from the sampling sites.
Identification of major pathogenic microbes in hospital effluents using metagenomic screening and design of potent inhibitors using virtual screening of medicinal plant library against them
The occurrence of emerging biological contaminants including antibiotic resistance genes (ARGs) and Faecal Indicator Bacteria (FIB) is still little investigated in developing countries under tropical conditions. Antibiotic resistance has become a serious public health concern with economic and social implications throughout the world, Community acquired infections like streptococcal infections, pneumonia, typhoid fever, etc., or hospital acquired infections due to methicillin resistant Staphylococcus aureus, vancomycin resistant enterococci, vancomycin intermediate S. aureus or extended spectrum beta-lactamase enzyme producing Gram negative bacteria are getting prevalent. These infections lead to higher rates of hospitalization, longer hospital stay, leading to pressure ulcers (bedsore) and increase in the cost of treatment and thus increased economic burden on the community. The most common organisms identified in pressure ulcers are Staphylococcus aureus, Proteus mirabilis, Pseudomonas aeruginosa, and Enterococcus faecalis. In general, wounds are poorly characterized with minimal to no physical description and/or location provided.The emergence of pathogenic bacteria resistant to most, if not all, currently available antimicrobial agents has become a critical problem in modern medicine, particularly because of the concomitant increase in immunosuppressed patients . In June 2000, the WHO warned that the level of resistance to drugs used to treat common infectious diseases is reaching a crisis point. Resistant and even multi-resistant pathogenic bacteria have been detected in wastewater and sewage treatment plants, as well as in other environmental compartments [5–8]. Furthermore, in arid regions, wastewater containing resistant bacteria and antibiotics is used for irrigation, and sewage sludge serves as a fertilizer. This allows resistant bacteria to enter the food chain directly. Hospitals are one, albeit not the only source of antibiotic input into the environment . This investigation aims to explore the profile of antimicrobial resistance in hospital effluents and map the approximate pathogenic load that is being released into the environment using metagenomic approaches, which not only provides a faster and wider method for mapping the hospital effluent microbiome, but also provides us with an efficient tool for creating a database of antibiotic resistant microbes in the state. Cheaper alternatives using established medicinal plant extracts would also be explored since inhibition of multidrug resistant bacteria currently requires expensive therapeutic interventions.
Comparative Metagenomic Profiling of Pasteurized Milk for assessment of microbial content and setting up of a standard for milk production
As milk is consumed by people of all the age groups, the wholesomeness and quality of milk is of great importance. The health benefits of milk and dairy products are known to humanity since medieval times and may be attributed to the biological active components that are present in milk and also, due to their suitably modulated activities produced through the action of probiotics. We Indians generally consume milk in daily basis and mainly in boiled form. The urban population generally consumes pasteurized and packaged milk. Pasteurization is not a process of sterilization. Microorganisms which are not pathogenic when consumed may be retained in pasteurized milk. As with the milk we take up these organisms, their genes can be transferred to the host, which is in this case the human consumer. Moreover, different gene products both beneficial as well as harmful may be present in the milk sample. Therefore, we should have an overview of the different types of microorganisms, their different genes and the pathways which are more active of these microorganisms. To achieve these goals we have a great scientific tool in our hand, which is metagenomics. Metagenomics profiling of the microbiome of milk should provide us with a standardized bench-mark regarding the overall quality of the boiled and unboiled pasteurized milk that is being used commercially in the state of West Bengal. The completion of the project would enable the complete documentation of the available microbiome of milk that is consumed on a daily basis in the state of West Bengal and in and around Kolkata. The microbiome content would provide us with quantifiable data regarding the beneficial and harmful microorganisms that exists in the pasteurized packaged milk of the state. This would enable us to develop a parametric standard for safe and nutritious milk which can be consumed hassle free by consumers from all age groups.
Identification of potential lead compounds as inhibitors and convoy drugs against p - glycoprotein from Medicinally important plants Foeniculum vulgare (Fennel) and Trigonella foenum-graecum (Fenugreek) using De novo transcriptome analyses and virtual screening
Despite the available knowledge of the important chemical constituents and their possible health benefits as well as the components being a suitable convoy drugs; no reports are available regarding the testing of the active constituents either invite or computationally to understand their effects on p - glycoprotein inhibition and identification of the enriched pathways which result in the production of these chemicals. This project attempts to investigate this gap in linking the traditional knowledge and identifying suitable phytochemicals for proposal as suitable convoy drugs effective against p - glycoprotein. The results of the analyses would enable the identification of a proper set of lead molecules against p - glycoprotein and also shed light on the actual pathways that regulate the production of the target phytochemicals of plant origin from Fenugreek and Fennel.
Development of a Computational Framework for Integrating transcriptome and metagenome sequencing, annotation and pathway prediction to identify key stress and disease association networks in plants
Two most important and established techniques which utilises the Next Generation Sequencing Based methods for biological data generation are transcriptome and Metagenome sequencing. While the former provides insights into the exact transcriptome dynamics of a particular organism in a particular timeframe, the latter is used for the analyses of the microbial communities in a particular region in an either time or experimental condition specific manner. The basic steps following the sequencing run is more or less similar involving assembly, annotation, prediction of repetitive sequences and pathway analyses. However, the target reference databases and genomes differ and in case of transcriptome analyses, for organisms who do not have their complete genome sequenced as yet, de novo transcriptome analyses is performed for proper elucidation of the RNAome. However, if we consider the soil plant continuum concept, then an integration of these two techniques might enable us to properly understand the genetic responses of plants in context of the rhizospheric microorganisms, soil profiles and abiotic stress parameters such as salinity or heavy metal enrichment.
This work focusses on the proposal of a computational framework where data from transcriptome and Metagenome (both endophytic and rhizospheric) can be compared to formulate a biological model of plant function under a particular condition. Prediction would be made based on neural network based models using machine learning classifiers such as support vector machines to generate a network of the uniquely and combinatorial enriched pathways which would not only enable us to predict response to biotic and abiotic stress in the plant under study but would also provide insights on the successful amelioration strategies for sustainable agriculture.
Analyses and Identification of the transitional gut microbiome in ASD for the formulating a standardized human gut microbiota (SHGM) consortium with insights from unadulterated guts of ethnic tribes and development of ASD-Gut a open source platform for exchange of scientific and social findings on autism spectrum disorders
Autism spectrum disorder (ASD) is a developmental disorder that affects communication and behavior. Although autism can be diagnosed at any age, it is said to be a “developmental disorder” because symptoms generally appear in the first two years of life. Over the years social scientist and medical researchers worldwide have attempted to dissect the possible causes to this condition and come up with methods by which the people affected can be counselled into healthy adults. However, apart from some sparsely successful dietary interventions and special cases of good behavioural practice based improvements, the inabilities of the affected individuals have remained or worsened. Very recently alterations and supplementations of the gut microflora have produced exciting results which show the ability to alter the neurological signalling cascade by altering the gut. However, till date this approach has not been explored with Indians suffering from this spectrum disorder. Thus the need of the hour is two fold. First is to compare the normal and affected gut microbiome to understand the differences between the two and secondly to study the transitional gut microbiome from the mother to the child in a case control setting to understand what part of the mother’s gut is transmitted to the child and what is the contribution of that transitional microflora towards the normal development of the child. This project shall attempt to compare the normal and ASD gut as well as the transitional microbiome from mother to the child in a case control setting. We believe that the commonalities and the differences would enable us to propose a consortium of standardised human gut microbiota (SHGM) which can be then taken up by suitable authorities towards the formulation of a probiotic which may serve as a non invasive therapeutic intervention and indirect neurostimulant. This project also shall develop an open source exchange platform available in the form of a backend database supported mobile app ASDGut which would enable the exchange the issues from numerous communities from any social structure and background having access to a smartphone. This app would not only have video support for identification of possible behavioural symptoms from diverse patient groups but shall also have a comparative gut microbiome profile available for comparison.
Rhizospheric metagenome of the terrestrial mangrove Fern Acrostichum from Indian Sunderbans
In this study we report the sequencing and analyses of the rhizospheric microbiome of the terrestrial mangrove fern Acrostichum aureum Linn from the Indian Sunderbans. Samples were collected using standard protocols and 16rRNA gene amplicon sequencing was performed to identify the microbial communities prevalent in the fern rhizosphere. 19,31,252 raw reads were assembled into 2,04,818 contigs and were analysed using QIIME to reveal the abundance of Proteobacteria, Acidobacteria and Planctomycetes. The data is available at the NCBI - Sequence Read Archive with accession number: SRX2660456. This is the first report of the rhizospheric microbiome belonging to a Fern species.