REVIEW OF LITERATURE Chapter 2 Review of Literature

Facts are many, but the truth is one. -Rabindranath Tagore

2.1 Features of metabolic and physiological properties, such as the formation of a wide variety The phylum Actinobacteria represents of secondary metabolites and one of the largest taxonomic units production of extracellular enzymes among major lineages currently (Schrempf et al. 2001). Some of these recognized within secondary metabolites especially (Stackebrandt et al. 2000, Stackebrandt produced by Streptomyces species have et al. 1997) in terms of number and been proved to be potent antibiotics variety of identified species. It (Lechevalier et al. 1967). This trait comprises gram-positive high G+C turned Streptomyces into primary content bacteria. The G+C content antibiotic-producing organisms generally ranges from 51% (some exploited by the pharmaceutical ) to more than 70% in industry (Bérdy et al. 2005).Moreover, Streptomyces and Frankia. An diverse forms of lifestyles are exception is the genome obligate encountered among Actinobacteria.For pathogen , whose instance pathogens (, G+C content is less than 50%. N o c a r d i a , T r o p h e r y m a , Actinobacteria exhibit a broad range of Corynebacterium, morphologies, from rod-coccoid etc.), plant commensals (Leifsonia), ( Arthrobacter ) or coccoid nitrogen-fixing symbionts (Frankia), (Micrococcus) to fragmented hyphal soil inhabitants (Streptomyces), thermal forms (Nocardia spp.) or highly (Acidothermus), stone dwellers differentiated branched mycelium ( Geodermatophilus ) a n d (Streptomyces spp.) (Atlas et al. gastrointestinal tract (GIT) inhabitants 1997).They also exhibit diverse (Bifidobacterium spp.).

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Abnormal developmental features are biodiversity. also displayed by many actinobacterial 2.2 Evolution and Dynamics of genera, for example,the continual non- Bacterial Genomes replicating state exhibited by certain The primary genetic proceedings that mycobacteria or sporulating aerial establish genome shape and structure mycelium formation in Streptomyces are believed to be (1)horizontal gene species. Actinobacteria are extensively transfer (HGT), (2) gene duplication, distributed equally in terrestrial and (3) chromosomal rearrangements and aquatic (including marine) ecosystems, (4) gene loss. Despite several efforts to specially in soil, where they play a vital quantify the relative contribution of role in the recycling of unruly each of these processes, no consistent biomaterials by decomposition as well model can so far explain and outline as humus formation (Goodfellow et al. the evolutionary development of 1983, Stach et al. 2005). Furthermore, bacteria based on their existing genome some Bifidobacterium species are used structure (Andersson et al. 2000, Hong as active component in a variety of so- et al. 2004, Kunin et al. 2003, Snel et called functional foods due to their al. 2002). A brief review regarding all apparent health-promoting and the aforementioned processes could be probiotic properties. They have been helpful in reconsidering the genomic reported to take part inbile salt aspects of organisms. hydrolase activity, protection against pathogens mediated through the 2.2.1 HGT: process of competitive exclusion, The introduction of alien or novel immune modulation along with the genes by HGT permits for quick niche- ability to stick to mucus or intestinal specific adaptation, which may lead to epithelium (Lievin et al. 2000, bacterial diversification and speciation Ouwehand et al. 2002, Stanton et al. (Cohan et al. 2001). Bacterial genome 2005). Thus actinobacteria may be evolution is dependent upon the relevant to human and veterinary combined outcome of acquired genes medicine, biotechnology, and ecology. through cell division (vertically Moreover the genomic heterogeneity inherited) and by HGT (Woese et al. observed from already sequenced 2000). Taking this perception to its actibacterial genomes reflects their extreme, one can argue that two

REVIEW OF LITERATURE 15 bacterial taxa are more linked to each fitness of the infecting mycobacterium other than to a third one not because was acquired by HGT (Pethe et al. they allocateto a more recent ancestor 2004). Other genetic studies of the but because they swap over genes more ancestral M. prototuberculosis species frequently (Gogarten et al. 2002). HGT have specified that various HGT events is responsible for increasing the happened before the evolutionary competitiveness among bacteria in bottleneck that led to the appearance of their natural habitats. For instance, in the M. complex (Gutierrez some , DNA et al. 2005), perhaps from the Indian segments with many virulence genes subcontinent (Filliol et al. 2006). and gene clusters (pathogenicity Bioinformatic methods to recognize islands), appear to be acquired by HGT HGT events are based predominantly (Ochman et al. 2000). Actinobacterial on the analysis of deviation in the GC examples of virulence genes content (GC deviation), four-letter transmission through HGT are rare genomic signatures, dinucleotide (Rosas-Magallanes et al. 2006). Of differences, and/or codon usage. Some these, three cases showed to signify geneticists often proposed HGT as the obvious HGT events: (i) presence of explanation for genes present in only major toxin gene in phages one organism which would mean that of Corynebacterium diphtheria, (ii) HGT frequency is pretty low (below carries the 10% of the total gene complement) macrolide toxin genes liable for the (Kunin et al. 2003, Snel et al. 2002). ulceration in their linear plasmid Interestingly, a recent analysis revealed (Stinear et al. 2004), (iii) a large that many actinobacteria specific chromosomal segment of Streptomyces proteins were also encoded by turgidiscabies associated with causing Magnetospirillum magnetotacticum potato scab can be transferred via which is an alpha-proteobacterium, but conjugation (Loria et al. 2006). In not by any other alpha-proteobacterial addition, it has been argued that the strains. This proposes that M. virulence operonof Mycobacterium magnetotacticum may have acquired tuberculosis Rv0986-8 which plays a those genes from actinobacterial vital role in host phagocytic cells species by HGT (Gao et al. 2006). parasitism by enhancing the ecological An interesting case of HGT between

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Chlamydia and some Actinobacterial may be either large deletions that span genera (e.g., Streptomyces, multiple loci or deletions of one or a Tropheryma, Bifidobacterium, few nucleotides (Andersson et al. Leifsonia, Arthrobacter, a nd 2001). The influence of these altered Brevibacterium) have already been routes is erratic among bacterial described (Griffiths et al. 2006). In lineages (Mira et al. 2001). Inactivation serine hydroxymethyltransferase and deleterious mutations in genes with enzyme (GlyA protein), two conserved little involvement to fitness can be inserts of 3 and 31 amino acids remain transmitted to progeny and mount up in present in some along with populations, ultimately leading to gene the aforementioned subset of loss; whereas such mutations in genes Actinobacteria. Likewise, these that are lethal will prevent the bacteria also contain a preserved 16- production of progeny and so will be amino-acid insert in the enzyme UDP- eradicated from populations, resulting N-acetylglucosamine enolpyruvyl in the conservation of the functional transferases (MurA) responsible for gene (Ochman et al. 2006). peptidoglycan biosynthesis. However, Gene inactivation and loss are mainly the physiological and functional apparent in several bacterial groups significance of these HGT events with host-associated lifestyle, in which between Actinobacteria and the host provides many of the Chlamydiae is presently unclear. metabolic intermediates, thereby 2.2.2 Gene decay: preventing the requirement to preserve many biosynthetic genes. In Bacterial genome size is dogged by the endosymbiotic bacteria, such as outcome of numerous opposing forces. Buchnera and Rickettsia, loss of Genome contraction occurs by deletion individual loci or operons is the only bias and genetic drift whereas, a resource of discrepancy in the gene prominent selection pressure on the inventories between species (McLeod functional genes encourages the et al. 2004, Tamas et al. 2002). A clear preservation of DNA.Genome example of genome reduction is augmentation depends on both provided by , acquisitions of alien DNA and gene which has lost more than 1,000 genes duplications, coupled to adaptive compared with M. tuberculosis (Cole et benefits (Mira et al. 2001). DNA loss

REVIEW OF LITERATURE 17 al. 2001). Moreover, the presence of quickly, and thus only rarely n o n - functional genes, i.e., persevered in bacterial genomes pseudogenes, in M. leprae suggests (Ochman et al. 2006). Other bacteria that this genome loss is still in reveal a lower level of gene loss: in progress. Although the principles for Rickettsia prowazekii, anobligate pseudogenes identification differ intracellular pathogen where only 12 among studies, the overall rationale is pseudogenes were identified quite similar: the predicted protein (Andersson et al. 1998); and a current must be distorted to degree that genome analysis of two Streptococcus abolishes its function. The thresholds thermophilus strains (Bolotin et al. applied for identifying pseudogene are 2004) found that pseudogenes based on the organization and known constituted only 10% of the total size of functional domains within genome, reflecting adaptation of S. proteins, the observed length deviation thermophilus to their specific within individual gene families, and environment which is milk (Bolotin et available data on experimentally al. 2004). disrupted proteins (Ochman et al. 2.2.3 Gene duplications: 2006). Generally, pseudogenes include Earlier it was thought that bacterial cases where deletion of functional part genomes evolved from a much smaller or insertion of stop codons have ancestral genome through a number of resulted in distorted protein with very gene duplication events and the small gene length (less than 80% of resulting generation of paralogs original length) or where a frameshift (Kunisawa et al. 1995). However, some or insertion has altered more than 20% analysis based on currently available of the amino acid sequence of the data does not support this theory and protein (Lerat et al. 2004). Majority of reveals that gene duplications the annotated bacterial pseudogenes are contribute only diffidently to genome among the open reading frames (ORFs) evolution (Coenye et al. 2005). Despite with unknown functionality. The lack this, it has been noted that genes of common pseudogenes among participated in a precise adaptation different strains of same species have been conserved after duplications indicates that pseudogenes are signifying that gene duplication does produced continually, are eliminated have an evolutionary role (Coenye et

REVIEW OF LITERATURE 18 al. 2005). This is nicely exemplified by recombination between these repeated the mycobacterial paranome, sequences catalyze genetic corresponding to a functional class of rearrangement as well as gene loss genes concerned with fatty acid leading to taxonomic diversification. metabolism, in accordance to the These recombination events probably composite nature of the mycobacterial have endorsed in T. whipplei speciation cell wall and possibly reflecting the (Raoult et al. 2003). Moreover, adaptive evolution of microbial cellular evolution of chromosome is influenced structure (Coenye et al. 2005, Tekaia et by bulky chromosomal rearrangements, al. 1999). e.g., large inversions (Eisen et al. 2000). 2.2.4 Genome rearrangements: 2.3 Taxonomic reclassification of Along with the afore-mentioned Actinobacteria factors, chromosome rearrangements can also participate in genome Ray fungi, also called as Strahlenpilze arrangements. Synteny, a term used for by Lieske (1921), or actinobacteria indicating the preservation of gene with a tricky (Krassilnikov order between genomes, can be used as 1941) were historically considered as a phylogenetic tool to examine „intermediary‟ between bacteria and relationships among species, since the fungi. The name itself comes from the degree of genome rearrangements colony morphology on agar exhibiting enhances linearly in relation to the radial growth, a characteristic common bacterial divergent time (Korbel et al. to fungi. The mycelial shape and musty 2002, Wolf et al. 2001). Chromosomal smell of actinobacteria is also similar rearrangement is chiefly dependent on to fungi. Moreover, the peptidoglycan the repeated and mobile elements based cell wall of this group is also a activity for example functionality of property shared to . The very transposons, insertion sequences (ISs), primitive taxonomical treatment of prophage sequences, and plasmids actinobacteria was proposed by (Kolsto et al. 1997). Bacterial genomes Buchanan (1917) who proposed an with a higher density repeat sequence order name . Some have higher rearrangement rates taxonomical works have been based on leading to a rapid loss of gene order the perception of evolutionary trend (Rocha et al. 2003). Homologous from a simple form to more

REVIEW OF LITERATURE 19 complicated features like hyphae and lineages (Goodfellow 1967), especially sporangia (Kluyver & van Niel 1936). the genera Streptomyces (Williams et However most of the taxonomists were al. 1983) and Mycobacterium not satisfied since morphological (Tsukamura et al. 1969). features were not considered which The arrival of sequencing of 16S may result in faulty positioning, rRNA, 16S rRNA gene classification especially in classifications specifically (Stackebrandt & Woese 1981) and dependent upon a dichotomous advanced molecular phylogenetic tools scheme. have advnced the search for a Chemical criteria were initially very molecular clock that is required to be few and could not provide a solid base present in all lineages. These for taxonomy, yet they have gathered a techniques presented actinobacteria as solid set of data compensating for few a consistent subdivision of „Gram- lost functions. Now-a-days, to explain Positive bacteria‟ along with the a species, it is compulsory to afford a firmicutes (low G+ C), Bacillus and measure G+C content of DNA which is Clostridium (Woese 1987). This a characteristic of the genome; confirmation supported Actinobacteria phospholipids analysis which is an as coherent, with little genetic important feature of membrane; the di- distances. amino acids characterising cell wall; or Actinobacteria are generally aerobic quinones which represent the and posses filaments (in branching respiratory chain of organisms. None pattern) except Bifidobacteriales of these elements taken independently (Woese 1987). This treatment was is adequate to recognize a microbe; reserved in many consecutive editions however, taken as a whole they can of Bergey‟s Manual and the class name give way a solid taxonomic basis in was available in 1997 (Stackebrandt et amalgamation with growth al. 1997) although untenably because characteristics and morphology. Sokal no type order was proposed at the time & Sneath (1963) used a mixture of (Euze´by & Tindall 2001). However, such characters as the basis of 16S rRNA genes sequences and numerical taxonomical study. These phylogenetic reconstructions may not approaches have allowed the be considered as the golden benchmark classification of several actinobacterial in bacterial taxonomy for several

REVIEW OF LITERATURE 20 reasons. First, there are examples of the 16S rRNA gene phylogeny, has species with more than one copy of the noticeably tailored all levels of their 16S rRNA gene differing by taxonomy. There are six classes approximately 6%, as in comprising of five basal ones Thermomonospora chromogena (Yap (Acidimicrobiia, Coriobacteriia, et al. 1999), which was taken as Nitriliruptoria, Rubrobacteria and support of lateral transfer. Secondly, Thermoleophilia) each with one or two the 16S rRNA gene may be plasmid- orders. The main class Actinobacteria borne as in Bacillus megaterium comprises of 15 orders (Kunnimalaiyaan et al. 2001), further (Actinomycetales, Actinopolysporales, following the idea of lateral transfer. Bifidobacteriales, Catenulisporales, Finally, the 16S rRNA gene is a single Corynebacteriales, Frankiales, marker that thus disobeys one basic Glycomycetales, Jiangellales’ principle of biology, that any Kineosporales, Micrococcales, investigation should be considered for Micromonosporales , reproducibility. This constraint led to Propionibacteriales , multi-locus sequence analysis (MLSA) Pseudonocardiales, Streptomycetales developed originally for Neisseria Streptosporangiales). (Maiden et al. 1998), where five or Gao and Gupta (2012) proposed an more preserved genes were sequenced approach of concatenating 35 proteins and analysed. This approach is now to re-classify actinobacterial commonly used, mainly to distinguish phylogeny, with an idea of indels to species within a genus. It helped the re- corroborate the major lines of that characterization of the genera Nocardia classification (Wu et al. 2009). Study (McTaggart et al. 2010) and of Verma et al. (2013) revealed the Streptomyces (Doroghazi & Buckley need of reassessment in the orders 2010), with some noteworthy Frankiales and Micrococcales. Sen et deviations from the topological pattern al. (2014) performed a detailed obtained by 16S rRNA gene sequences, phylogenetic reconstruction indicating extensive recombination. exclusively for Actinobacteria using The most current taxonomical both bootstrapping (Felsenstein 1985) treatment of Actinobacteria (Ludwig et MLSA and Prunier approach (Abby et al. 2012a, b), based fundamentally on al. 2010) and spitted Frankiales into

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F r a n k i a l e s ord. nov., Elaeaginus allowing their growth in Geodermatophilales ord. nov., nitrogen-poor condition (Benson and Acidothermales ord. nov. and Silvester 1993; Chaia et al. 2010; Nakamurellales ord. nov.. Moreover, Schwencke and Carú 2001). Micrococcales was also divided into Elucidation of the Frankia genomes several monophyletic orders – has revealed novel possibilities in Micrococcales (Micrococcus, Kocuria, metabolic diversity, stress tolerance Rothia, Arthrobacter, Tropheryma, and natural product biosynthesis, Microbacterium, Clavibacter and which aid in establishment of Frankia- Leifsonia); Cellulomonales actinorhizal symbiosis (Schwencke and (Beutenbergia, Cellulomonas, Caru 2001; Benson and Dawson 2007). Xylanimonas, Jonesia a n d Depending upon the host specificity, Sanguibacter); Brachybacteriales Frankia has been clustered in four (Brachybacterium) and Dermacoccales major groups (Ghodbhane-Gtari et al. (Kytococcus, Intrasporangium). 2013, Nouioui et al. 2011; Normand et al. 1996; Cournoyer and Lavire 1999, 2.4 Interaction with niche Ghodhbane-Gtari et al. 2010). Actinobacteria being one of the major Members of lineage I infect plants of representatives of rhizospheric the Betulaceae, Myricaceae, and microbial flora play diverse roles in Casuarinaceae families (except p l a n t - associated microbial Gymnostoma). Dryadoideae (all communities. Some genera are actinorhizal Rosaceae), Coriariaceae, predominant soil saprophytes while Datiscaceae, and the genus Ceanothus others are either symbionts or (Rhamnaceae) are infected by strains endophytes or plant pathogens. of lineage II. The lineage III stains are 2.4.1 Actinobacteria as nitrogen fixers most promiscuous infecting Elaeagnaceae, Colletieae (all Frankia, a genus with the ability to fix actinorhizal Rhamnaceae except nitrogen, can live both as soil Ceanothus), Myricaceae, saprophytes as well as endophytic Gynmmostomam (Casuarinaceae), and symbionts and are competent to occasionally Alnus. On the contrary, establish mutualistic symbiotic Frankia of lineage IV are „atypical‟ association with non-leguminous plants strains either unable to infect plant or notably Alnus, Casuarina and

REVIEW OF LITERATURE 22 produce ineffective root nodules which inactivation (Murry et al. 1985). A do not participate in nitrogen fixation natural pentacyclic triterpenoid lipid (Tisa et al. 2016). called hopanoid formed a multilayed envelop surrponding the vesicle Several genes have been reported to be structure which act as the oxygen involved in nitrogen fixation. Among barrier (Berry et al. 1991, 1993). In them, nitrogen fixing (nif) genes are Frankia hopanoid remain as bacterioho most important (Normand et al. 2007). -panetetrol and bacterio-hopanotetrol Strains of cluster I, II and III own their phenylacetate monoester derivatives. nif genes however, in lineage IV this Generally, there are two pathways for gene is totally absent which probably hopanoid biosynthesis viz. mevalonate lead to their inability to fix nitrogen. pathway and methyl erythritol Eleven- twelve nif genes remain phosphate (MEP) pathway. However, clustered together as an operon on 11.6 in Frankia, only MEP pathway has -14 kb region and generally one copy been identified (Rohmer 2008). of each nif gene is present in that Surprisingly, hopanoid biosynthetic cluster (Tisa et al. 2016). However, gene cluster have been identified in all exceptions are there. For instance, nifV Frankia strains including non nitrogen gene of lineage III strains has been fixer lineage IV which indicates that reported to be present at a distant from their functional implication is nitrogen the nif cluster and two copies of nifU independent (Dobritsa et al. 2001; were reported in lineage II. Udwary et al. 2011; Richau et al., Interestingly, six non-nif genes (orf1, 2013, Tisa et al. 2016). Hence, these orf2, hesA orf3, orf4 and fdx) were genes are not only crucial for nitrogen also found to be located between nif fixation besides, they are also genes (Tisa et al. 2016). Another important as primary cell wall important part of frankial N2 fixation is component conferring the membrane the presence of specialized thick- integrity. walled vesicles (Noridge and Benson 1986; Tisa and Ensign 1987; Normand Concurrently, Ni-Fe hydrogenase or et al. 2014). Their formation is uptake hydrogenase (hup) operons are controlled by partial pressure of also imperative for Frankia life style. oxygen. Vesicles play a pivotal role in There are two hup operons in lineage I protecting nitrogenase from oxygen and II strains Among them, operon I is

REVIEW OF LITERATURE 23 expressed under free living condition (Ghodhbane-Gtari and Tisa 2014). whereas, operon II is expressed under However, Chabaud et al. 2015 has symbiotic condition (Leul et al. 2007; revealed a novel signalling mechanism Richau et al. 2013). However, all hup for symbiosis in Frankia. Host plant genes were found to be clustered in one induced Frankia have been reported to operon for lineage III and IV strains. exert a signal triggering the expression Truncated hemoglobin genes, which of reinfection genes in host plant as are broadly classified into three groups well as Ca2+ level. Their additive - group I (HbN), group II (HbO), and effect results in biologically active group III (HbP) act either as NO symbiotic signal. Moreover, next- scavengers or protect nitrogenase from generation proteomic analysis has also oxygen inactivation by buffering the provided plethora of information oxygen level (Wittenberg et al. 2002). regarding the proteome footprinting of Blastp analysis revealed the complete frankial metabolomics altered by root absence of canonical nodABC gene exudates of compatible and non- (NodA-acyl transferase, NodB-chitin compatible host plants. This study deacetylase, NodC-chitin synthase) revealed that, proteins having cluster among Frankia strains (Tisa et functionality in the early steps of host al. 2016) except Candidatus Frankia recognition and infection were Datisca Dg1(Persson et al. 2015). expressed highly in Frankia induced There prevail two hypotheses behind by root exudate of compatible host this finding. First one is, Dg1 follow an plant (Ktari et al. 2017). identical symbiotic pathway to legume- Bioinformatic analysis has also rhizobium relation and second one is, explored a relation between the while sequencing, Dg1 has genome size and niche specificity as incorporated genes from other well as biogeographic ranges among organisms as a part of contamination. this genus. For instance, the genome Actinorhizal nodules are well known to size of narrow-host-range- Casuarina contain several bacteria within them strain Frankia sp. CcI3 is 5.4 Mb hence, genes of other microbes can be whereas, the size expands to 8.9 Mb accidentally mixed to the genome of for Frankia sp. Ean1pec which is a Dg1 resulting the exceptional presence broad-host-range Elaeagnus strain as of nod genes within the genome well as soil cosmopolitan. The

REVIEW OF LITERATURE 24 infectivity of F. alni ACN14a is iron and thus deprive other microbes restricted mainly to Alnus and from this essential nutrient (Jog et al. Myricaceae hence, they possess a 2014). Some water soluble chitinolytic genome with intermediate size (Tisa et enzymes produced from soil al. 2016). Thus a hypothesis prevailed actinobacteria are also effective against that, the genome size of Frankia fungal infection and soil borne changes with their host range and it pathogens. Broad range of antibiotics held true with all the strains of this produced by this particular microbial genus sequenced so far. Furthermore, population is well recognized plant study with horizontal gene transfer protectors against pathogenic bacteria, (HGT) and insertion elements (IS) in oomocytes, nematodes and fungi (Van Frankia has availed us with et al. 2008). Moreover, production of information regarding their genomic nitrous oxide by some Streptomyces plasticity (Tisa et al. 2016). Frankia sp. strains elicits the plant defense Eul1c appeared to be the most stable mechanism. Streptomycetes can also genome with a very few HGT and IS degrade a signaling molecule elements. Thus, our understanding responsible for pathogenicity in regarding the infection process of Pectobacterium carotovorum (Baz et Frankia could be enhanced through a al. 2012). Thus, actinobacteria can act detailed comparative study. as a soldier of plant defense mechanism. 2.4.2 Actinobacteria as plant protector and plant defense elicitor 2.4.3 Actinobacteria help in plant growth: Actinomycetes are well recognized for their capacity to produce a broad range Several nitrogen fixing actinobacteria of metabolites which can act against have been found to be associated with plant pathogens (Van et al. 2008). The the plant root system. Majority of them mechanisms by which actinobacteria aid in the development of plant (Qin et inhibit the pathogens include al. 2011). For example, antibiosis, nutrient competition, Corynebacterium sp. AN1, which was quorum quenching, nitrous oxide and isolated from forest phyllosphere can degradative enzyme production (Conn reduce acetylene thus substituting the et al. 2008). Some strains also produce role of nitrogenous fertilisers and siderophores with the ability to chelate helping in plant growth promotion

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(Nimaichand, 2016). Pseudonocardia others. Most well known dioxanivorans CB1190 has also been actinobacterial phyto-pathogenicity is reported to fix dinitrogen (Mahendra et the potato scab disease caused by al. 2005). Above all, Streptomyces, Streptomyces scabiei. S. turgidiscabies accounting an abundant percentage of and S. acidiscabies also causes scab soil microflora and an effective plant disease. “Common” scab is caused by colonizer is well recognized as source S. scabiei and S. turgidiscabies of plant growth promoters and other whereas; S. acidiscabies is responsible biocontrol agents (Igarashi et al. 2002). for “acid scab” (Loria et al. 1997). The Besides, endophytic (microbes remain stains of these two types of scab differ within the plant host for a long time from each other in pigmentation without causing any harm to host) pattern, pH resistant capacity and streptomycetes can aid in their host raffinose utilization capacity. plant growth by auxin production Furthermore, S. scabiei and S. (Sadeghi et al. 2012). turgidiscabies can produce melanin however, S. acidiscabies is unable to 2.4.4 Actinobacteria- as pathogen produce the same (Nahar et al. 2018). Plant pathogenesis is a very complex Another form of scab called Russet multi step process. The pathogens have scab is restricted to potato skin and to infect the plant system, break the quality of the crop. This disease is cell wall and win over the plant again of two types: American rudest defense mechanism (Qin et al. 2011). scab and European russet scab. Though Although, the number of they are also caused by Streptomyces actinobacterial plant pathogen is less however, the causative strains are however, sometimes they can cause a phytopathologically different from huge agricultural and economical loss each other. Further, some by destroying huge amount of crops streptomycetes (S. flavovirens) through (Qin et al. 2011). Clavibacter, their ligninocellulose degrding Leifsonia, Rhodococcus a nd enzymes can harm the intact phloem Streptomyces are the most popular cell wall of hosts and cause harm to plant pathogenic actinomycetes. Most them (Locci et al. 1994). Rhodococcus importantly, Rhodococcus and fascians have been reported to infect Streptomyces have ride range of host both monocot and dicot plants which and thus are more deleterious than

REVIEW OF LITERATURE 26 include economically important crops Abscisic acid which inhibit the plant too (Cornelis et al. 2001). Upon growth. One interesting fact regarding infection, excess intracellular epiphytic Leifsonia infection is, they are very growth through stomata leads to leaf much xylem limited (Young et al. deformation, fasciations, leafy galls 2006). Moreover, members of and witched broom formation. Shoots Mycobacterium, Corynebacterium , are hyper-induced by dormant auxillary Propionebacterium and Tropherymae meristem activation and de novo are popular as animal pathogen. meristem formation (Cornelis et al. 2.4.5 Actinobacteria- as a source of 2001). Manipulation of host hormonal natural products balance by the pathogen derived auxins Actinomycin was the first discovered and cytokines is the main reason antibiotic produced from Streptomyces behind the aforementioned symptoms antibioticus in 1940 (Katz 1967). (Cornelis et al. 2001). Later, streptothricin and streptomycin Clavibacter michiganensis is the only were also produced from S. lavendulae reported plant pathogen of its genus a nd S. griseus respectively (Daferera et al. 2003). All the five sub- (Handelsman et al. 1998). With strains of this species are potent plant advancement of research, pathogen colonizing in their host actinobacteria proved itself as a great xylem. The severity of C . source and reservoir of natural michiganensis infection and difficulty products. Thus, this group has become in controlling them has recognize this important from the biotechnological species as quarantine organism under perspective (Strobel et al. 2004). the European Union Plant Health Aminoglycosides (neomycin, Legislation and laws of many other kanamycin, streptomycin), beta- countries (Bentley et al. 2008). Ratoon lactum, anthracyclines, stunting of sugarcane is a disease chloramphenicol, glycopeptides, caused by Leifsonia xyli subsp. xyli lantibiotics, streptogramins, (Monteiro-Vitorello et al. 2004). This tetracyclins etc. are some other major is a systematic disease where plant antibiotics produced from growth is retarded. Putative fatty acid actinomycetes. The antibiotic desaturation modifies the carotenoid production capacity of each microbe biosynthesis pathway and produce also varies significantly. For instance,

REVIEW OF LITERATURE 27 some Streptomyces strains produce Since, vertebrates do not depend upon single antibiotic whereas, some are the breakdown (hydrolysis) of capable of producing a broad range of disaccharide trehalose, the antibiotics (Strobel et al. 2003). functionality of validamycin is Concurrently, actinomycetes are also biologically selective. Furthermore, popular as source of other secondary Polyoxin B act as antifungal agent in metabolites such as, herbicides, friuts, vegetables and ornamental insectisides, immune-suppresors, plants whereas, polyoxin D can control antifungal agents etc. S. aureus S-3466 the growth of Rhizoctonia solani which has been reported to biosynthesise a causes therice sheath blight by mixture of tetranactin and dinactin inhibiting the microbial protein which are used for commercial biosynthesis (Handelsman et al. 1998). production of macrotetrolides Kasugamycin is active against rice (Handelsman et al. 1998). This blast (Pyricularia oryzae cavara) and compound act as as insecticides and Pseudomonas disease in several crops immuno-suparresive agents. Another (Rossman et al. 1990). example is Mildiomycin which act as Recent genomic analysis of an antifungal agent. Their primary sites Actinobacteria reported that, some of actions are locations of cell wall secondary metabolite biosynthetic chitin synthesis, inositol biosynthesis machineries regulated by selected as well as sites of protein and DNA genetic loci may become active under synthesis (Handelsman et al. 1998). precise lab conditions which may Similarly, validamycin, kasugamycin, further enhance the commercial polyoxinin B and D are also potent importance of these microbes as a natural antifungal agents synthesised natural resource of biologically active from different strains of Streptomyces. compounds. Validimycin A, produced from S. 2.4.6 Probiotic Actinobacteria hygroscopicus var. limoneus is converted into validoxylamine A Advancement of research has proved within fungal cell. This compound the vital role played by gut microbiome inhibits the breakdown of trehalose and in various aspects of human health. thus suppresses the supply of Bifidobacterium, is one of the major carbohydrate to fungi and insects. constituent of gut microflora which is

REVIEW OF LITERATURE 28 ubiquitous in every human (Ishibashi et respectively (Ishibashi et al. 1997, Wei al. 1997). The population of these et al. 2012). Further, formula fed microbe changes with age. In babies do not develop B. adolescentis childhood they are enormous and in their adulthood. Thus, breast milk unstable whereas, in adulthood their does have a bifidogenic effect since; population comparatively reduces and selected glycans present in breast milk become stable. Bifidobacterium can be utilized by Bifidobacterium longum, B. breve, and B. bifidum (Wei et al. 2012). This report was also predominate in infants. On contrary, B. supported by another finding that, catenulatum, B. adolescentis and B. breast milk of mother with mutation in longum are more prevalent in Fucosyl transferase 2 gene (FUT2) adulthood (Lau and Liong 2014). delays the colonization of gut Several extrinsic factors are important Bifidobacteria since, the mentioned for the initial colonization of this genus mutation inhibit the conversion of in intestine. Infants can also acquire fucose to glycans (Ishibashi et al. this microbial population from mother 1997). through mother‟s vaginal tract, GIT, Bifidobacterium composition in human breast milk, placenta and amniotic fluid intestine also fluctuates with different (Wei et al. 2012). Further, birth mode disease condition. Obesity, one of the is another important factor in this case. major condition affecting children and Infant born vaginally are more adults has been shown to have abundant with bifidobacterial correlation with alterations in gut population than infant born by cesarean microbiome. A relatively lower set of section. The gut microbiome may vary Bifidobacterium and higher set of with the feeding regime too. For Staphylococcus have been reported example, Bifidobacterium breve, B. previously in obese children (Million et bifidum, B. longum ssp. longum, and B. al. 2012). Those mothers who during longum spp. infantis have been their pregnancy period gain over reported to be present in both breast- weight supply lower amount of and formula-fed infants. B. longum ssp. Bifidobacterium to their babies than infantis and B. longum ssp. longum are that of healthy normal weight mother. commonly associated with breast-fed Besides, patients with short or long infants and bottle-fed infants term asthma also bear lower amount of

REVIEW OF LITERATURE 29 aforementioned strains.B. adolescentis also be useful in restoration of has been found to be dominant in lipoprotein imbalance in children asthma patients. This strain is generally having dyslipidemia (Sharma et al. found in adulthood however, if a 2016). Concurrently, the probiotic mother has allergy, the baby may get strains also deal with some brain gut the strain through breast feeding. On disorders like anxiety, depression, contrary, reports are there suggesting stress etc. Interestingly, Bifidobacteria the probiotic and anti allergic effect of is now being used in diagnosis of B. longum in children (Ouwehand et al. tumor since; they have the ability to 2001). Further, during irritable bowel colonize at tumor sites. They can also syndrome, the population of inhibit the growth of tumor by tumor Bifidobacterium also decreases. Thus, specific T-cell responses, CD8+ T cell the variety and population of responses and dendritic function Bifidobacteria has a relation with modulation (Kim et al. 2007). Thus, different syndrome, suggesting their Bifidobacteria with their probiotic role in regulating that activities have become some of the pathophysiological condition major genera of Actinobacteria. (Shirakawa et al. 2018). Moreover, 2.5 Extremotolerant Actinobacteria their capacity to stimulate the immune Actionobacteria can be found both in system has recognized them as normal as well as in extreme nutrient- probiotics. However, this capacity has deprived environments characterized been found to be specie specific. by low or high temperatures, acidic/ Systematic and intestinal immunity is alkaline pH, high radiation, salinity, very much elevated by B. bifidum, B. low levels of available nutrients, and dentium, and B. longum. Members of moisture (Zenova et al., 2011). The these genera are also used in treatment varied physiology as well as metabolic against allergies, celiac disease, elasticity of extremotolerant obesity, diarrheas, colic, infections or actinobacteria enables them to endure necrotizing enterocolitis (Vazquez et unfavourable and hostile al. 2010). The bile sale hydrolase circumstances. Bull (2010) recorded activity of some selected strains the high abundance of actinobacteria reduces the blood cholesterol and under extreme conditions which broke lipoprotein cholesterol level hence, can the conventional concept of restricted

REVIEW OF LITERATURE 30 preponderance of actinobacteria in soil different metabolic modes (i.e., and fresh water environment. Several heterotrophy, autotrophy, and studies have been carried out on saprobes) antibiosis and fabrication of actinobacteria isolated from normal specific enzymes which may help them environments (neutral pH and to survive under harsh conditions (high temperature ranging 20–40◦ C). temperature, saline and alkaline). The However, only a few reports are thermo-tolerance is accredited to the available to understand the diversity of presence of elevated hydrophobic and actinobacteria present in extreme electrostatic interactions as well as environments, their adaptation and disulfide bonds in the proteins of ecological role. Poly-extremotolerant thermophilic strains (Ladenstein and and poly- extremophiles actinobacteria Ren, 2006). They have definite unusual are also capable to exist in proteins called chaperones which help environments with more than two in refolding the proteins with partial extreme conditions. Poly - denaturation (Singh et al., 2010). extremophiles are able to live in Several other proteins are also present environments having multiple stresses which bind to DNA and avoid their (Gupta et al., 2014). Thermo- denaturation at high temperatures. A acidophilic, alkali-thermophilic, thermophilic Streptomyces sp., haloalkaliphilic, thermophilic obtained from desolated place was radiotolerant, and thermo-alkalitolerant reported to produce autotrophic actinobacteria are some examples of metabolic pathway enzymes like Poly-extremophiles. Their occurrence carbon monoxide dehydrogenase has been recognized in different (CODH) (Gadkari et al., 1990). This extreme geographical locations enzyme facilitates the growth of including the Arctic (Augustine et al., microbes in nutrient deprived 2012) and Antarctic (Gousterova et al., conditions via oxidizing the accessible 2014) regions, hot springs (Chitte and inorganic compound such as carbon

Dey, 2002), oceans (Raut et al., 2013), monoxide into CO2 which can further and deserts (Kurapova et al., 2012). fixed by the enzyme RuBisCO into microbial biomass by Calvin–Benson The extremophilic actinobacteria show cycle (King and Weber, 2007). The numerous adaptive schemes for chemolitho-autotroph thermophilic, example- switching between two

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Acidithiomicrobium sp., isolated from taiwanensis (Chen et al., 2004) were geothermal environment, exploits reported to be radio-tolerant. The sulphur as their energy source (Norris resistance machinery has not been et al., 2011).The antibiosis is an sufficiently understood, but the whole additional strategy through which genome analysis of R. radiotolerans actinobacteria keep going by killing RSPS-4 exposed the presence of genes other microbial flora under nutrient responsible for encoding proteins those restricted conditions. Alkaliphiles and are involved in oxidative stress acidophiles regulate H+ concentrations response, DNA repair system and inside and outside the cell through biosynthetic pathways of proton pumps for maintaining their companionable sugars (mannosyl physiological pH inside (Kumar et al., glycerate and trehalose) playing a vital 2011). Alkaliphiles have negatively role in extenuating the damage caused charged cell wall polymers which can via radiation (Egas et al., 2014). In stabilize cell membrane by charge current years, some more radio tolerant density reduction at the cell surface and alkali tolerant actinobacterial (Wiegel and Kevbrin, 2004). The species such as Microbacterium adaptive strategy of halo-alkaliphiles radiodurans GIMN 1.002T (Zhang et embraces excess tolerance to salty al., 2010), Microbacterium environment by producing and maritypicum (Williams et al., 2007), accumulating higher amount of attuned Cellulosimicrobium cellulans UVP1 solutes (Roberts, 2005) which can (Gabani et al., 2012), Kocuria sp. avoid desiccation through osmo- ASB107 (Asgarani et al., 2012) and regulation. Additionally, they have Kocuria rosea strain MG2 (Gholami et Na+/H+ antiporter system to exclude al., 2015) have been reported. The two undue salt from inside of the cell. alkali tolerant Kocuria strains were. The Kocuria sp. ASB107 isolated from Some actinobacteria also have the Ab-e-Siah radioactive spring of Iran is capacity to live in various radioactive a psychrotrophic strain showing sites and tolerate extremely harmful tolerance to ionizing radiation (upto radiations like UV and gamma rays. 90% lethal doses) such as corona Three thermophiles of Rubrobacter discharge and ultraviolet (256nm-UV). species - R. xylanophilus (Ferreira et The Kocuria rosea strain MG2 was al., 1999), R. radiotolerans and R.

REVIEW OF LITERATURE 32 revealed to tolerate the high quantity of industrially costly compounds (Singh damaging UV-C radiation. This strain et al., 2013). The manufacture of can grow in a ample pH range (5–11 biomolecules through extremophiles with optimum growth at pH 9.2) along alleviates the risks of other bacterial with salt concentration (0– 15%). Cell- contaminations, besides providing halo viability analysis on Kocuria rosea -tolerant, alkali-stable and thermo- strain MG2 under multiple stresses was stable compounds. Enzymes produced reported by Gholami et al. (2015). It through these extremophilic/ was revealed that, after 28 days of extremotolerant actinobacteria are also incubation under desiccation condition, useful under extreme environment, the Kocuria strain cells were viable and thus, making them appropriate demonstrated high tolerance to the candidates for industrial processes with strong oxidant such as H2O2 (1–4%) harsh treatment methods. and radiation. H O is a well accepted 2 2 2.5.1 Thermophilic and antimicrobial compound damaging Thermotolerant Actinobacteria biological membranes through Thermophilic actinobacteria flourish at hydroxyl radical generation. Hence, the comparatively elevated temperature aforementioned Kocuria strains seem ranging from 40 - 80◦C (Tortora et al., to display both non-enzymatic anti 2007). They are generally found in oxidant defense systems (carotenoids) moldy silage (Corbaz et al., 1963), self and enzymatic (catalase and -heating residues of plant, sugar cane peroxidase) diminishing the bagasse, cereal grains (Suihko et al., consequence of radiation, strong 2006), compost heaps, decaying oxidants and other stresses (Gholami et vegetable materials (Shivlata and al., 2015). Tulasi 2015).These are 2 types: The flexibility and adaptability of stringently thermophilic which can extremophilic/ extremotolerant grow in the temperature range between actinobacteria award them a 37 and 65◦Cwith optimum proliferation competitive benefit over other 55–60◦Cand reasonably thermophilic microbes. Besides helping them to which thrive at 28–60◦C and need 45– endure extreme conditions, the 55◦C for their optimum growth (Jiang physiology as well as metabolic and Xu, 1993). Another collection elasticity also prompts them to create called thermo-tolerant actinobacteria is

REVIEW OF LITERATURE 33 able to endure upto 50◦C temperature During the 1970s and 1980s, it helped (Lengeler et al., 1999). to produce new and powerful scientific techniques, specifically recombinant Thus, actinobacteria being a diverse DNA research, genetic engineering, group of microbes are present in all rapid gene sequencing, etc. Around the probable habitats and are important same time, the term “bioinformatics” from medicinal, ecological and was coined by Ben Hesper and Paulien environmental aspects. In spite of all Hogeweg (Hesper and Hogeweg, diversities, there remain some unities 1970). Bioinformatics is by nature a among these organisms in both cross-disciplinary field that began in genomics and proteomic level. Those the 1970s with the efforts of Margaret signatures, if identified, will definitely O. Dayhoff, Walter M. Fitch, Russell help us to fill up the pit holes F. Doolittle and others and has regarding our understanding about gradually matured into a fully actinobacteria. In this study, we are developed discipline. Initially, it was going to utilize the power of referred as „the study of information bioinformatics to analyze the massive- processes in biotic systems‟ (Hesper data generated from a mass of and Hogeweg, 1970). However, its actinobacteria to obtain an overall primary use since at least the late “birds‟-eye” view of these microbes. 1980s has been to describe the 2.6 Bioinformatics as tool to explore application of computer science and the complexities microbial world: information sciences to the analyses of 2.6.1 Dawn of bioinformatics: biological data, particularly in the areas of genomics involving large-scale It was in the year 1953 when James DNA sequencing (Luscombe et al., Watson and Francis Crick proposed the 2001). The arrival of the INTERNET is twisted-ladder (double helix) structure another important milestone in the of deoxyribonucleic acid (DNA). This development of bioinformatics as a full particular event forever changed the -fledged discipline. This discipline history of biological science and gave represents the convergence of rise to modern molecular biology. In genomics, biotechnology and short order, their discovery yielded information technology, and ground-breaking insights into the encompasses analysis and genetic code and protein synthesis.

REVIEW OF LITERATURE 34 interpretation of data, modeling of 1998), Protein DataBank (PDB) biological phenomena, and (Bernstein et al., 1977) and several development of algorithms and others. After the formation of the statistics (Fenstermacher, 2005). The databases, tools became available to need for bioinformatics was further execute various analyses. Two accelerated when the Human Genome programs, which greatly facilitated the Project (HGP) was launched in 1990. similarity search, were FASTA The aim of the project was to sequence (Pearson and Lipman, 1988) and the entire human genome. Information BLAST (Altschul et al., 1990). Many gleaned from the HGP is not very programs have been further developed useful until the huge data is managed since then. Accessibility of free and and interpreted in a proper way by the open source software has taken computational tools leading to the bioinformatics and its application to all materialization of bioinformatics. The -together new heights. success of HGP opened the flood-gates 2.6.2 Bioinformatics based platforms for other genome sequencing projects. suited for microbial research: Gradually genome sequences of Several „omics‟ based research mouse, rat, worms, yeast and plants disciplines i.e., genomics (for genome like rice, Arabidopsis were completed. data), proteomics (for protein data), The publication of huge amount of transcriptomics (for gene transcription sequence data were greatly supported data), etc., have emerged with the by development of high end computers, enormous advancement of smart computational tools for large- bioinformatics. The „cause and effect‟ scale annotation, functional relationships of biological systems are classification of the proteins (Searls, being frequently employed by 2000) and development of specific mathematical models and databases (Birney et al., 2002). computational simulations for proper Availability of complete genome elucidation of biological complexities. sequences of different organisms lead Proper blend of knowledge-driven to the development of public computational simulations and data- repositories of gene data like GenBank oriented bioinformatics holds the key (Benson et al., 2000), EMBL (Baker et in apt realization of the riddles of host- al., 2000), DDBJ (Okayama et al., microbe interactions. Meaningful

REVIEW OF LITERATURE 35 analysis of high-throughput sequencing popular for the chore but nowadays, it data promises to unravel the enigma of has been replaced by Illumina the complex associations. Specific sequencing which has been reported to biological databases, providing access produce equivalent results with a better to annotated genomic data of bacterial coverage (Luo et al., 2012). QIIME populations, like IMG/M (Markowitz (Navas-Molina et al., 2013) and et al., 2014), SEED (Overbeek et al., MEGAN (Huson et al., 2011) have 2005) and Greengenes (DeSantis et al., been efficient pipelines for analyzing 2006) have opened up revenues to data extracted via marker gene extract significant information. profiling. PICRUSt effectively predicts Annotation pipelines like RAST (Aziz the functional potentials of host et al., 2008) and MG-RAST (Meyer et associated microbes by analyzing the al., 2008) have been instrumental for metagenome from the marker gene the wholesome purpose of data (Langille et al., 2013). Marker characterizing bacterial masses. gene profiling does not involve high Systems Biology Markup Language cost and thus, aptly suits the purpose of (SBML) (Hucka et al., 2003) and large projects concerned with data BioModels repository (Le Novere et extraction. Pathway enrichment al., 2006) have been very useful for followed by metabolic reconstruction proper standardization of biochemical has been suggested to be fruitful reaction models. Various pertaining to proper analysis of bioinformatics based platforms suited metagenomic data (Abubucker et al., for genomic investigations are as 2012; Sharon et al., 2011). Robust follows: machine learning algorithms have been suggested to be useful for proper 2.6.2.1 Marker Gene Profiling predictive analysis of data produced by Marker gene profiling (also known as marker gene profiling (Nakano et al., gene amplicon sequencing) involves 2014; Statnikov et al., 2013). The extraction of DNA from different major limitation of the technique is that samples. DNA extraction is followed it largely depends on pre-existing by amplification using primers curated rRNA databases for taxonomic specifically designed against ribosomal profiling which highlights the RNA genes. Initially, 454 impotence of the method to pyrosequencing technology was pretty

REVIEW OF LITERATURE 36 successfully characterize novel generation sequencing (NGS) bacterial species present in the samples technologies has been a boost for the (Dicksved, 2008). of metagenomics. NGS based techniques allow in-depth profiling and 2.6.2.2 Metagenomics: analysis of microbiome without the Metagenomics is the branch of limitations of selection bias and genomics that performs investigations constraints, associated with cultivation by direct extraction and cloning of methods. NGS-based methods rely DNA from assembly of organisms greatly on sophisticated bioinformatics (Handelsman, 2004). Comprehensive based tools, regularly updated data sequencing is necessary to pursue repositories and functional know-how. metagenomics based investigations The main motif of bacterial (Fig 2.1). It is taken into presumption metagenomic analysis has been that human gut microbiome contains a proficient characterization of and large number of uncultured species and profiling of the microbial members therefore, thorough sequencing is framing the diverse microflora. mandatory to proceed with Similarity based alignment tools like metagenomics of gut samples BLAST serve the preliminary purpose (Emerson et al., 2012; Hess et al., of finding out regions of similarity 2011; Narasingarao et al., 2012). between the raw or assembled Sophisticated genome assembly sequences against a reference database protocols (Bashir et al., 2012; and thus, provide an initial clue about Goldberg et al., 2006) and efficient the functionalities of the query single-cell sequencing technologies samples. MG-RAST pipeline employs (Lasken, 2012; Shapiro et al., 2013) the M5nr database that is a large have been suggested to aid proper repository of non-redundant protein metagenomic analysis. Due to sequences from multiple sources extensive focus on host associated (Weinstock, 2012; Wilke et al., 2012). microbiome and availability of QIIME pipeline facilitates taxonomic numerous bacterial genomes, assembly classifications. Functional and -free methods tend to be useful in metabolic databases like KEGG thorough scrutiny of metagenomic Orthology (Kanehisa and Goto, 2000) samples (Carr et al., 2013; Luo and or SEED subsystems (Overbeek et al., Moran, 2013). Rapid progress of next-

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Fig 2.1 Schematic diagram for Metagenomics data analysis. The scheme was adopted from Prakash, T. and Taylor, T.D., 2012. Functional assignment of metagenomic data: challenges and applications. Briefings in bioinformatics, 13(6), pp.711-727.

2005) assist investigations pertaining to in apt execution of function oriented functionalities. Sophisticated statistical studies (Abubucker et al., 2012; Jiao et techniques are also frequently al., 2013; Levy and Borenstein, 2013). employed for extensive functional 2.6.2.3 Metatranscriptomics: analysis (Kristiansson et al., 2009). Metatranscriptomics involves Metabolic modeling tends to be useful

REVIEW OF LITERATURE 38 extraction of RNA from samples of Another crucial step is the removal of interest and is much more complex ribosomal RNA from bacterial RNA in than DNA extraction technologies. The order to avoid misinterpretations while major concern of metatranscriptomics analyzing metatranscriptomic data. is to achieve good quality sequence and 2.6.2.4 Metabolomics: sufficient yield. Fast alignment tools Metabolomes refer to the complete set like bowtie (Langmead et al., 2009) of metabolites present in a cell/ and SSAHA (Ning et al., 2001) find organism or a community of organisms applications in proper alignment of (Jordan et al., 2009). Research transcriptomic data, retrieved from the pertaining to metabolome of human gut host-microbiota sample, with a set of microbiota has been an exciting representative bacterial genomes challenge these days. Metabolomes (McNulty et al., 2011; Turnbaugh et have been suggested to be crucial al., 2010; Xiong et al., 2012). Simple players of proper interactions and cross BLAST has also been employed for the -talks between gut associated microbes purpose (Gosalbes et al., 2011; Xiong and human host (Marcobal et al., 2013; et al., 2012) but the task requires Nicholson and Lindon, 2008). extensive and rigorous computational Extensive profiling of metabolomes support to align millions of sequence promise to confer large body of reads. Current pipelines use variations information regarding the complex of this basic approach (Leimena et al., interactions and also the health benefits 2013; Xu et al., 2014). and hazards associated with the Metatranscriptomic data promise to be concerned microbes of human gut handy in exploring the intricacies of (Larsen and Dai, 2015). gut microbiota and provide vivid depiction of the host-microbiota 2.6.2.5 Computational Modeling and interactome (Westermann et al., 2012; Simulation: Xu et al., 2014). However, retrieval of Availability of complete genome sufficient bacterial RNA from sequences has made it feasible to combined host-microbiota sample has execute computational and metabolic been a daunting task as host RNA modeling of concerned prevails over bacterial RNA due to low microorganisms (Oberhardt et al., biomass of the microbial members. 2009; Thiele and Palsson, 2010). Huge

REVIEW OF LITERATURE 39 data surging out of the various efficiently for a community of metagenomic projects also provide organisms displaying significant ample scope for apt simulation and interactions (Borenstein, 2012). This metabolic modeling (Henry et al., technique also finds its applications in 2010). Computational simulations do supra-organism where there is employ simplified techniques based on significant assemblage of metabolic abstractions. Computational models not pathway associated genes reflecting all only prove fruitful for a single involved species (Borenstein, 2012). organism but also can be employed