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International Journal of Life Sciences • ISSN 2091–0525 • Year 2012 • Volume 6 • Issue 1

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International Journal of Life Sciences • ISSN 2091–0525 • Year 2012 • Volume 6 • Issue 1 ResearchAAnn IInnddee particlepeennddee n n tt , , O O pp e e n n A A c c c c ee s s s s , , P P e e ee r r R R ee v v i i e e ww e e d d ,, N N o o nn - - P P rr o o f f i iDesaitt JJoo uandurrnn aaBhamrell (2012) IInntteerrnnaattiioonnaall JJoouurrnnaall ooff LLiiffee SScciieenncceess YY eeaarr 22001122 VVoollluummee 66,,, IIIssssuuee 11 Reserach Article Novel gut bacterial fauna of Gryllotalpa africana Beau. (Orthoptera : Gryllotalpidae) Ashok E. Desai 1, * and Pramila R. Bhamre 1 1 Zoology Research Laboratory, P.G.Department of Zoology, K.T.H.M. College, Nashik-422 002. (M.S.) India. Corresponding author ABSTRACT Email: [email protected] Gryllotalpa africana (Beauvois) is a successful group of detritus feeder therefore it is an important both for their roles in carbon cycling in the environment and converting organic decaying material Phone No: in to biofuel. Hence, bacterial diversity and relationship were explored from the mole cricket 091-9422751115 G.africana (Beau) to show ideal symbiotic association. The analysis and characterization of gut micro biota of G.africana (Beau) revealed the presence of 10 species and two genera of bacteria viz. Fax No: Acidiphilium rubrum, Dienococcus proteolyticus, Sporosarcina ureae, Micrococcus varians, 091-253-2571376 Micrococcus kristinae, Acetobactrium, Alcaligenes eutrophus, Micrococcus roseus, Micrococcus lylae, Sporosarcina, Citrobacter amalonaticus and Corynebacterium xerosis. All the species and genera Editor were confirmed by observing their morphological features of the colony and various biochemical Randeep Rakwal tests. These results indicate that the presence of mole cricket specific bacterial lineages implies relationship of gut microbes and their host mole cricket. The outcome of this work indicted that DOI the gut bacterial fauna of G.africana shows the species-rich with greater diverse communities dx.doi.org/10.3126/ijls.v6 which might be providing resistance to invasion of pathogenic bacteria. i1.5949 Key words: Gryllotalpa africana (Beau); Gut; Bacterial species; Genera; Diversity. INTRODUCTION polysaccharides (xylans, pectins and gums ) and oligosaccharides. It is known that insects inhabiting in moist soil get infection of Gram-negative, small rod shaped, Insect gut microbial communities play coliform bacteria followed by micrococci and important role in processes linked to global carbon sporeforming bacilli micro-organisms through their cycling and production of green house gases. Review of literature reveals that only a studies predominantly food and thus have a constant association of micro- with termites and cockroaches have been carried out organisms in gut or in various tissues intracellularly ( to study gut micro biota (Gijzen et al, 1994). Similarly Lysenko, 1985; Ohkuma and Kudo, 1996;). Hence, gut the phylogenetic diversity and physiology of termite microflora of insects reflects the local environment gut Spirocheats were carried out (Breznak, 2002). (Eutick et al, 1978).Usually insects with straight Shifting of Peridroma saucia (Hub.) from field to alimentary canal have few micro-organisms than laboratory caused changes in the quantity of facultative and aerobic bacteria in gut (Bruce those with complicated gut with a variety of pH range Lighthart, 1988) and gut dependent effects of gut providing number of different niches (Bignell, 1984; bacteria on their insects host Erwinia sp. (De Vries et Tanada and Kaya, 1993). The association of micro- al, 2004).The hindgut microbiota of cricket have been organisms in gut or other tissues is important with characterized by using fluorescently labelled rRNA respect to nutrition as the micro-organisms provide targeted oligonucleiotide probes (Santo Domingo et essential dietary factors like vitamins, sugar, amino al., 1998). While studies of Rajgopal (2009) found that acids etc. to the host or help in digestion of plant gut micro biota have beneficial role in successful establishment of insects. Zurek et al. (2000) studied 50 | International Journal of Life Sciences • ISSN 2091–0525 • Year 2012 • Volume 6 • Issue 1 Research article Desai and Bhamre (2012) the role of diversity and contribution of the intestinal Morphological and biochemical test of isolated bacterial community to the development of larvae of colonies Musca domestica (Linn.). The antimicrobial phenolics derived from the bacteria are widely present in locust The various bacterial colonies was observed and grasshopper of different stages and nutritional by microscopic examinations for shape,size, colour, state (Dillon,R. and Charnley,A;1995) and Yunhua Zhu elevation, motility characteristics and were subjected et al.(2012) reported phylogenetic analysis of the gut for Gram staining for primary identification. Isolated bacterial microflora of Macrotermes barneyi. The colonies were subjected to various biochemical tests review of literature reveals that only a few studies viz: indole production, methyl red, Voges-Proskauer predominantly with termites and cockroaches, and citrate utilization tests as well as other enzyme collembola, silkworm larvae and locust have been detection tests in triplicates and identification of carried out to study their gut microbiota (Dillon,R. bacteria was carried out following Holt (1994). and Charnly,2002; Dillon et al.,2002). There is no information on gut or tissue microbiota of Gryllotalpa RESULTS africana (Beau), a species inhabiting in most soil or The results of the present findings are mud. Hence, the present study was undertaken to find out the type of micro-organisms inhabiting the gut of presented in tabular form (Table 1, 2 and 3) Table 1 this insect which is detritus feeder. depicts the shape, size, colour, margin , elevation and motility of the different microbial colonies. Table 2 represent the results of indole,methyl red, Voges MATERIALS AND METHODS Proskauer, citrate, tributyrin and different enzyme tests, while Table 3 shows the identified colonies to Collection of Gryllotalpa africana (Beau) Genus and species. Thus the results of various The adults of mole cricket Gryllotalpa africana biochemical tests indicates that the midgut of (Beau.) were collected from different localities around Gryllotalpa africana (Beau.) contains 12 types of Nashik city (M.S.) and brought to the laboratory in bacteria viz. Acidiphilium rubrum, Dienococcus polypropylene containers (Fowler &Justi, 1987; Short proteolyticus, Sporosarcina ureae, Micrococcus varians, &Koehler, 1979). They were maintained in glass Micrococcus kristinae, Acetobacterium, Alcaligenes troughs containing moist soil with decaying debris of eutrophus, Micrococcus roseus, Micrococcus lylae, plant leaves at temp. 22±2o C, L: D- 12:12 h and RH 65 Sporosarcina, Citrobacter amalonaticus and %. These specimens used for further studies. Corynebacterium xerosis. Isolation of Bacteria and their cultivation DISCUSSION The adult mole crickets were dissected in petriplates containing 0.65% saline (Lum, 1961). The insects gut bacteria represents a dynamic midgut region of alimentary canal was dissociated food assimilation system. The bacterial fauna can and transferred into test tube containing 0.85% saline thrive in the gut environment, withstanding the buffer then homogenized in sterile glass homogenizer and system of pH, ionic composition and steep redox centrifuged at 3000 rpm. The supernatant suspension gradations, various micro-organisms have distinct was used as sample for cultivation of bacteria and for niches within the gut, the reasons of this are not yet further tests. The supernatant suspension was then clear (Cash, 2005).Studies on diversity and density of inoculated by sterile nichrome wire loop on small bacterial communities of gut provide an ideal tool in area of nutrient agar forming zone of inoculation molecular biology techniques like sequencing under sterile conditions (laminar airflow filter). The genomes etc. (Natarajan et al., 2012). The insect nutrient agar plates were incubated at 37oc for 24 hrs. contains varied types of nonpathogenic microbial After incubation period the microbial colonies on communities inhabiting the gut which provide ideal nutrient agar plates were observed for their different conditions for bacterial conjugation suggesting that types. Some of these colonies were then transferred the gut is a ‘hot spot for gene transfer’ ( Dillon and to the nutrient agar slants for the preservation. 51 | International Journal of Life Sciences • ISSN 2091–0525 • Year 2012 • Volume 6 • Issue 1 Research article Desai and Bhamre (2012) Dillon ; 2004). The presence of bacteria in some studies on gut of Lymantria dispar (Linn.) revealed insects which are localized in special portion of gut that some acid tolerant bacteria Streptococci create have been well documented. The presence of their own low pH environment by producing lactic Klebsiella pneumoniae and Enterococcus casseli in the acid (Kodama and Nakasujii, 1971). Studies observed gut of locust Pantoea agglomeran have been that diet rich in cellulose induced increase in demonstrated (Dillon and Charnley;2002). The protozoan population in hindgut of Periplaneta phylogenetic diversity of intestinal community and Americana (Linn) thereby increasing cellulolytic physiology of gut spirochaetes in the termite gut activity for cellulose digestion (Gijzen et al., 1994). observed (Breznak, 2002 and Ohkuma & Kudo ,1996). Steinhaus (1949) stated that the microbes are often Further studies have also reported that the insects associated with gut. Backhed et al. (2005) reported with simple straight
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