KEYNOTE & LEAD LECTURES
METABOLOMICS IN MICROBIOLOGY
1Ch. Venkata Ramana and 2Ch. Sasikala
1Department of Plant Sciences, School of Life Sciences, University of Hyderabad, Hyderabad 500046, Telangana, India.
Mail: cvr449@gamil.com
2Bacterial Discover Laboratory, Centre for Environment, Institute for Science & Technology, Jawahar Lal Nehru Technological University Hyderabad, Kukatpally, Hyderabad 500085 Telangana, India.
Mail: sasikala.ch@gmail.com
Metabolomics is a rapidly growing area in the field of modern biology dealing with organic biomolecules (metabolites) with about 1 Kda. Small organic molecules play key role in the function of a microorganism and are greatly influenced by external factors. Though most of the metabolites are the product of metabolism brought about through gene coded reactions (specific and non-specific enzymes), a large number of reactions are non-enzymatic transformations which are much more important than the enzymatic reactions. Metabolomics is useful to all branches of microbiologists who are dealing with taxonomy, physiology, biochemistry, pathology, industrial microbiologists, biodegradations, biotransformations and all other branches of microbiology. The lecture focuses microbiological metabolomics in some of the above areas in which our lab is focusing.
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IDENTIFICATION AND DESCRIPTION OF NEW TAXA OF PROKARYOTES
1Ch. Sasikala and 2Ch. V. Ramana
1Centre for Environment, IST, JNTU Hyderabad, Kukatpally, Hyderabad
2Department of Plant Sciences, Uiversity of Hyderabad, Gachchi Bowli, Hyderabad.
Prokaryotic systematics/taxonomy deals with the inter-related areas of identification, nomenclature and classification of Bactria and Archaea. Identification of prokaryotes is necessary even for those whose main area of research lies outside the bacterial taxonomy. Every scientist who works on the bacterial biotechnology needs to give a name (at least till genus level) to the isolate/ culture on hand. It is also necessary for quality check, to make sure that the culture you are working with is indeed what one supposes it to be. The various levels of prokaryotic taxa are Domain, Phylum, Class, Order, Family, and Species. While only small number of characters is studied for identification, more comprehensive, polyphasic characterization is a must for classification of taxa.
Identification scheme: A scheme of characteristics helping in identification of an isolate as a member of an established taxon or as a member of a previously unidentified species. For identification, pure culture is not absolutely essential. However, it is essential that the bacterium is already classified, only small number of characters studied and particular pattern of characters is used. 16S r RNA gene sequence analysis FAME (cellular fatty acid analysis) and MALDI-TOF MS analysis are the Universal methods of identification of Prokaryotes.
Classification scheme: Arrangement of organisms into taxa/groups on basis of similarities or relationships, based on polyphasic characterization involving Ecological, morphological, physiological, biochemical, molecular, genetic, antigenic, serological characters etc. The gold standard for species delineation is genome relatedness determined through whole genome DNA-DNA hybridization. With the availability of whole genome sequences of a large number of prokaryotes, it is now possible to do in-silico hybridizations as well.
Description of novel taxa: Minimal standards are prescribed for description of different groups of bacteria through polyphasic taxonomy. Various characters to be determined and their relevance and limitations will be discussed taking anoxygenic phototrophic bacteria as an example. Deposition in two culture collections belonging to two different countries is absolutely essential for valid publication of description of novel taxa. Accession numbers and certificates of deposition has to be obtained indicating the culture is available and is publicly accessible, without any restrictions. No conditions can be laid on distribution. Comparative taxonomic analysis with closely related taxa in author's laboratory is increasing the difficulty in description of novel taxa. Already a number of descriptions of novel taxa is including genomic data and soon it will become mandatory to include genome sequences for description of novel taxa.
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Microbial-derived silver nanoparticles for application in drug delivery systems
C. Ganesh Kumar
Biotechnology Laboratory, Organic Synthesis and Process Chemistry Division, CSIR-Indian Institute of Chemical Technology, Uppal Road, Hyderabad 500007, India.
Email: cgkumar@iict.res.in
Antimicrobial therapy has played a significant role for treatment of microbial infections and has saved millions of lives but also increased the threat of antibiotic resistance which has become an alarming issue from a global perspective. In the post-antibiotics era, the consequences due to bacterial resistance have led to higher rates of mortality and increased health-care expenditure. In the recent past, microbial derived silver nanoparticles (Ag-NPs) have gained renewed interest for potential application in antimicrobial therapy and other biomedical domains in view of their unique physiochemical and biological properties. The recent strategy to improve the drug efficacy is by combining them with metal nanoparticles for the control of microbial infections. Considering these facts, we developed a facile and biodirected synthetic approach using cell free supernatant of Delftia sp. strain KCM-006 for preparation of Ag-NPs and its application as an antifungal drug carrier. They were characterized using UV-visible, TEM, DLS, zeta potential, XRD and FT-IR techniques. Transmission electron microscopy (TEM) analysis revealed the formation of spherical and monodispersed Ag-NPs with an average size of 9.8 nm. The synthesized Ag-NPs were found to be highly stable and crystalline in nature having a zeta potential of -31 mV. XRD studies revealed that the Ag-NPs exhibited a face-centered cubic lattice. The Ag-NPs exhibited very good antifungal activity against various pathogenic Candida strains. Furthermore, the efficacy of nanoparticles was increased by conjugating the antifungal drug miconazole to Ag-NPs which exhibited significant fungicidal activity, inhibited ergosterol biosynthesis and biofilm inhibition by increasing ROS levels. In addition, the cell viability and immunocytochemistry analysis against different normal cell lines including Chinese hamster ovary cells (CHO), human lung cell line (MRC5) and human vascular endothelial cells (HUVEC) demonstrated that these nanoparticles were non-toxic up to a concentration of 20 µM. In conclusion, these results suggest that the synthesized Ag-NPs find application as both antifungal agents and drug delivery vehicles.
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Deciphering molecular intricacies of protein structure and function with the help of fluorescence spectroscopy
Dr. Priyankar Sen
School of Bio Sciences and Technology, Vellore Institute of Technology, Vellore - 632 014, Tamil Nadu, India.
Email: Priyankar.sen@vit.ac.in
Abstract
Fluorescence spectroscopy becomes a widely used tool at the interface of biology, chemistry and physics, because of its precise sensitivity and recent technical advancements. The measurements can provide information on a wide range of molecular processes including the interactions of solvent molecules with fluorophores, rotational diffraction of biomolecules, distances between sites of biomolecules, conformational changes and binding interactions. These advances in fluorescence technology are decreasing the cost and complexity of previously complex processes. Fluorescence spectroscopy is a highly developed and non-invasive technique that enables the on-line measurements of substrate and product concentrations or the identification of characteristic process states.
Fluorescence measurements can provide information not only on the specific molecular makeup of a sample but also on the local environment surrounding the fluorescence molecule, or fluorophore. The absorption and emission characteristics of these fluorophores have been extensively studied and reviewed in numerous publications. Our lab has worked extensively on fluorescence spectroscopy to study protein-ligand interactions, Folding/refolding and molecular aggregation studies. It helped us to study unfolding, refolding and aggregation of albumins in co-solvent like alcohol, detergents and denaturants. A cosolvent; TFE, was found to induce α-helical aggregation at highly alkaline condition. In contrary, physiological concentration of urea was found to enhance helical structure in albumin. Many clinically important dyes like thioflavinT, curcumin, fluorescein and an insulin analogue were found to exhibit moderate binding affinity towards albumin, and specific binding sites were predicted with domain-specific unfolding and in-silico studies.
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MICROBIAL SYSTEMATICS - WITH SPECIAL REFERENCE TO DIVERSITY AND BIOPROSPECTIVE ATTRIBUTES OF ACTINOBACTERIA
Dayanand Agsar, Raghavendra H, Umesh M. K., Mohan Reddy K., S. S. Deepthi,
Sudarshan A, Neelam Mishra, Reshma S and Shilanjali Bhalerao
A-DBT Research Laboratory, Department of Microbiology,
Gulbarga University, Kalaburagi-585 106 Karnataka India
dayanandagsar@gmail.com and sivadeepthi218@gmail.com
Systematics conventionally referred as classification or taxonomy and is not being accepted as an attractive area of research by majority of biologists. Recently, it is being realized that, without biosystematics of potential microorganisms, Research and Development in microbial biotechnology may not march ahead and progress with desired applications. It is expected that, hardly about 6% of the total microbial world has been put under biosystematics and biotechnological exploration. In the past few years, the knowledge of microbial systematics has increased enormously due to the advances made in basic, molecular and chemo-systematics with analytical techniques and computational biology. The biodiversity of Indian subcontinent is the richest in the world vowing to its vast geographical area, varied topology and climate. Increased knowledge of microbial ecology and physiology with an echo physiological perspective focused to explore neglected habitats and more intensive investigation of the better studied microorganisms. Current paradigm shift in biotechnology points to the importance of systematic studies of the novel microorganisms shown to be producing new bioactive molecules. Among microorganisms, especially in comparison with bacteria and fungi, actinomycetes have been given not much attention, despite their greater genomic diversity. The bio-prospecting strategies are based upon the promise that, new secondary metabolites can be found by screening small numbers of novel actinobacteria isolated from the unique habitat.
Efforts were made by several researchers in our Actinomycetes-Diversity and Bioprocess Technology (A-DBT) Research laboratory since 2001 to explore various unique habitats of the deccan trap, mainly focusing lime stone quarries for the detection of novel and potential actinobacteria. Six new species and a new genus of actinobacteria have been reported. Further, the detected novel isolates of actinobacteria were explored for few potential bioactive molecules including enzymes-tyrosinase and cellulase, and also water soluble pigment-melanin. Novel isolates of actinobacteria detected from the lime stone quarries and the bioprospective attributes of those isolates will be discussed in detail during the presentation.
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Survival Games that Bacteria Play
Dr. C. S. Srinandan
Biofilm Biology Lab, Centre for Research in Infectious Diseases, School of Chemical and Biotechnology, SASTRA Deemed University, Thanjavur 613406, Tamil Nadu, India
Email: srinandan@gmail.com; srinandan@scbt.sastra.edu
Bacteria are ubiquitously found on our planet and it is amazing to see that they grow at extreme as well as moderate habitats. They have been surviving from a very ancient period, at least 3.5 billion years ago or even before. It has been estimated that the total bacterial numbers on the planet earth could be around a whopping five nonillion (5 x 1030). Moreover, previously it was estimated that there could be around one trillion species of bacteria on our earth, but the new tree of life suggests that the number could be an underestimate. It makes anyone wonder about the survival strategies that bacteria follow in-spite of these astonishing huge numbers. A predominant form of survival strategy of bacteria in their natural environment is the biofilm, wherein the bacteria aggregate and encase themselves in an exopolymeric matrix. A gamut of survival games are being played by the bacteria in their biofilm lifestyle. For example, chemical warfares like the production of antibiotics or bacteriocins, specific conflict by secretion of Type Six Secretion System, public good cooperation by the production of exoproducts, and also specific sharing of resources via secretion of nanotubes. In this talk, I discuss the results of my lab on the intra-population conflict and cooperation that we observe in related bacterial biofilms of Salmonella and E. coli. The public good cooperation among Salmonella and how they discriminate the defectors, as well as the bet-hedging strategy used by E. coli for their population survival, will be discussed.
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SYNTHESIS OF SILVER NANOPARTICLES FROM WHITE ROT FUNGI: THEIR CHARACTERIZATION AND BIOLOGICAL APPLICATIONS
Prof. M.A. Singara Charya,
BSR Faculty Fellow UGC, Department of Microbiology, Kakatiya University, Warangal -506 009
E-mail: singaracharyam@gmail.com
The field of nanotechnology is an tremendously developing field as a result of its wide-ranging applications in different areas of science and technology. The term nanotechnology is defined as the creation, exploitation and synthesis of materials at a scale smaller than 1 mm. Nanomaterials like copper, zinc, titanium, magnesium, gold, alginate and silver have come up but among them silver nanoparticles have proved to be most effective and predominant as it has good antimicrobial efficacy against bacteria, viruses and other eukaryotic microorganisms. Silver is a naturally occurring precious metal, most often as a mineral ore in association with other elements. It has been positioned as the 47th element in the periodic table, having an atomic weight of 107.8 and two natural isotopes 106.90 and 108.90 Ag with abundance 52 and 48%. It has been used in a wide variety of applications. Of all the different type of nanoparticle, silver nanoparticles are playing a major role in the field of nanotechnology and nanomedicine.
Fungi in the form of fruit bodies were collected in Eturnagaram forest of Warangal district. A total of fifty five strains of basidiomycetes were collected and cultured. The isolated fungi were subjected for silver nanoparticles synthesis. Out of 55 pure cultures 10 cultures showed the color change.
Two fungal cultures were identified by Classical and Molecular identification: GK-1 -Ganoderma enigmaticum , GK-14- Trametes ljubarskyi. In Ganoderma enigmaticum the spectral data revealed two types of vibrations (i.e. stretching and bending) in the wavelength range of 3,500-500 cm-1. In the above figure it is evident that the presence of an amine vibration band at 3,385cm-1 represents a primary amine (N-H) stretching and amide (N-H) bending vibration at 1,638 cm-1. In Trametes ljubarskyi the spectral data revealed two types of vibrations (i.e. stretching and bending) in the wavelength range of 3,500-500 cm-1 . It is evident that the presence of an amine vibration band at 3,396cm-1 represents a primary amine (N-H) stretching and amide (N-H) bending vibration at 1,638 cm-1. The AgNPs of two selected white rot fungi at 80µl/well showed good anti candidal activity ranging from 10 to 25 mm. Antimicrobial properties of mycosynthesized AgNPs treated cotton textiles were examined against some pathogens. The anti-bacterial activity of mycosynthesized AgNPs treated cotton materials was examined using agar based diffusion technique against both Gram positive and Gram negative bacteria and the outcomes presented. Among different types of metal nanoparticles AgNps have proven to be most effective due to its wide range of applications such as antimicrobial agent, in medical devices, dental materials, textiles fabrics, Agriculture, cosmetics, and also found to posses anti-inflammatory, anti-cancer, anti-angiogenesis and anti-platelet activity etc. The formation of AgNps mediated by biological route is considered as better method than the other conventional methods such as chemical, physical, and mechanical methods which are capital intensive, toxic and non-ecofriendly. Hence, "Green Process" is the best option to opt for the synthesis of AgNps using microbes.
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MIMICRY OF VIRUSES
Dr. D.V.R. Sai Gopal
Professor, Department of Virology, Sri Venkateswara University, Tirupati, Andhra Pradesh 517502, India
Email: dvrsaigopal@gmail.com
(This talk is dedicated to Prof. B. Rajasekhar Reddy, Dept. of Microbiology, SK University, Anantapuram on the eve of superannuation)
Viruses had an unique feature is the genome diversity, this won't be seen any other organisms. Viruses are the cleverest organisms on the globe. Depending on the environment and the type of host available they may change their behavior and propensity. Viruses had form simple structure to complex structure. Another important feature is the virus and host- specificity. Some of the viruses show pleomorphic character. Antigenic shift and drift are the phenomenon showed in certain viruses such as Influenza viruses and HIV. This shift and drift had the limitations in the change of their antigenic surface proteins- Hemaggultinin (HA) and Neuraminidase (N) example Influenza virus. The HA is responsible for binding and entry of the virion into host epithelial cells and N is involved in the processing of new virions budding out from the host cells, the sites recognized on the H and N proteins by host immune systems are under constant selective pressure. Antigenic drift allows for evasion of these host immune systems by small mutations in the H and N genes that make the protein unrecognizable to pre-existing host immunity. The antigenic drift is this continuous process of genetic and antigenic change among flu strains. Antigenic shift phenomenon is contrasted with antigenic drift, which it is the natural mutations of the strains of Influenza, it may lead to a loss of immunity or in vaccine mismatch, it occurs only in Influenza virus A. Antigenic drift occurs in all types of Influenza viruses- A,B,C. Due to antigenic shift new viral pathogens are evolved in the society. But this antigenic shift and drift phenomenon in HIV is different, which is an unlimited, so this is called as mimicry, it lead variable strains of HIV in the society. This molecular mimicry described the antigen sharing between host and the virus, because of the immunodominant specificity of the antibody chose to examine whether molecular mimicry might be the mechanism for HIV-1-ITP in the patients. Molecular mimicry of HIV involved various viral peptides occurred, however the extent and specificity are not easy to establish, by the nature of the problem, short, typically hydrophilic and sequences are involved. At a minimum, sequence analysts must be their claims with respect to compositional factors, statistical significance and sequence degeneracy and other important considerations include secondary and tertiary structure, amino acid modifications and insignificant cross-reactivities.
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Exploration of microbial contaminations in Potable water and their adverse effects on children's health: Molecular detection and prevention
Pavan Kumar Pindi
Professor, Department of Microbiology, Palamuru University, Mahabubnagar, Telangana State, India-509 001
E-mail: pavankumarpindi@gmail.com, +91-98493-27029
Globally 160 million children suffer from stunting and chronic malnutrition linked to water. Lack of access to safe water by school-going children is likely to contract various emerging diseases like typhoid, cholera, diarrhea, etc. Henceforth, there is an utmost need for the exploitation of uncultivable pathogens and rapid detection. Furthermore, fetal exposure to emerging microbial and chemical contaminations increases the risk of child's brain damage, lower IQ and immunity, learning disabilities or developmental problems. All the primary government schools of 64 mandals in Mahabubnagar district in Telangana state were surveyed for water quality and hygiene. The main sources of supply of water to these schools being Ramanpadu and Koilsagar reservoirs were studied further. Evaluation of coliforms and non-coliforms contamination of these two reservoirs both by cultivable and uncultivable methods was done which included standardization of DNA isolation protocols for DGGE and generation of clone/metagenomic libraries. Ramanpadu yielded more genera of bacterial pathogens than Koilsagar in both cases DGGE (37 species belonging to 5 coliform and 3 non-coliform genera EMBL-RP; LT592288- LT59327) and clone libraries (53 species belonging to 5 coliform genera and 7 non coliform genera NCBI-RP; KR612007-KR612048) which further proved for the necessity of water treatment. In cultivable assessment, 23 species belonging to 5 non-coliform and 3 coliform genera (NCBI; GU566304-GU566358) were reported. It was found that the most predominant coliform pathogens belonged to genus Enterobacter followed by Citrobacter and the predominant non-coliforms belonged to the genus Salmonella in both the reservoirs. It was found that more than 70% of the schools were found to be not meeting the standards of the WHO. These contaminations showed adverse effects on the school children exposing them to various health problems ultimately leading to drop outs. The contaminations in these reservoirs persisted even after many treatments, insisting on more innovative treatment techniques. In this study, the treatment at large-scale-level was done by a technique, rotating biological contactor (RBC) which proved efficient, removed BOD and chemical contaminants along with the microorganisms onto the biofilms with low power consumption and high water flow system. A novel house-hold treatment was developed by using Mentha arvensis (Mint) and Pimpinella thirupathensis (Adavi kothimera) leaves' extracts showed adverse effects on the growth of coliform pathogens and minimized chemical contaminants, proving this a simple, swift and cost-effective method for water treatment at house-hold-level. Further, molecular markers were developed for the rapid detection of coliforms in potable water within 3-4 hr before supplying to the house-holds.
In spite of the shooting measures taken, water quality is always a pertinent issues prevailing in the drinking water space. This may be attributed not only to the severe water contamination, threat and depletion of water reservoirs, but also lack of awareness among people. Treating water with simple edible leaves like mint leaves and adivi kothimeera leaves, and existing technologies can be adopted to save millions of lives.
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Why the influence of a good teacher can never be erased? : An empirical analysis, with the teaching methods of Prof. B.R. Reddy for the progress of Microbiology by enhancing student learning
(A tribute to Prof. B. Rajasekhar Reddy on his superannuation)
Buddolla Viswanath
Department of Bionanotechnology, Gachon University, Seongnam si, Gyeonggi do 461 701, Republic of Korea.
E-mail: buddolla@gmail.com
"Few teachers are great; probably none is great at all times. Yet, many teachers are great occasionally." (Anonymous)
The discipline of microbiology is both a stand-alone discipline with many sub-disciplines, as well as a discipline integrated within other disciplines. Microbiology education within a classroom is substantially enhanced by good and motivated teachers and by hands-on laboratory exercises. In this learning environment, students learn standard procedures and protocols that are routinely practiced in a microbiology laboratory. In my presentation I want to discuss my more than 15 years relationship with Prof. B.R. Reddy as an amazing teacher of microbiology and as research mentor. Particularly, in great appreciation for his years of dedication to education, and especially for all the lives that he have impacted over the years.