International Research Journal of Environment Sciences__________________________________ISSN 2319–1414Vol. 2(1), 25-30, January (2013) Int. Res. J. Environment Sci. International Science Congress Association 25 Microbial Diversity in the Surface Sediments and its Interaction with Nutrients of Mangroves of Gulf of Kachchh, Gujarat, India Kumar Goutam and Ramanathan Al School of Environmental Sciences, Jawaharlal Nehru University, New Delhi, INDIAAvailable online at: www.isca.in Received 01st December 2012, revised 31 December 2012, accepted 12 January 2012 Abstract The microbial population and nutrients status of the surface water indicates the nutrient dynamics in the study area. Nutrients in surface water directly provide the available source of nutrients to the microbes. Nutrients in the water such as NO (2.77-46.08mg/L), PO4 3-(0.32-2.81 mg/L), SO2-(1892-3839.98 mg/L) and dissolved silica (SiO) (3.32-16.98 mg/L) has been reported. Population of nitrate forming bacteria with higher population count at Old Bedi Port (M2), Narara (M5) and Mundra (M8) was reported while Phosphate solubilizing bacteria with higher count at Mundra (M8) and Jodiya (M10). The cellulose degrading bacteria was reported exceptionally high at Mundra, Narara and Old Bedi Port. The higher population count in the microbes synchronized with the nutrients availability in the surface water. Lack of significant correlations among the nutrients indicate the influx of anthropogenic inputs and waste discharges containing nutrients from river runoff into these environments. Keywords: Mangroves, sediment, microbes, nutrients. IntroductionMangrove ecosystems are unique environment, harboring diverse groups of microorganisms which perform an important role in nutrients cycling in the ecosystem1,2. Microbes play a vital role in the biogeochemical cycles of any ecosystem. Mangrove ecosystem is being very rich in organic matter, the presence of microbes especially bacteria are active participants in the ecosystem. Various groups of bacteria and fungi such as nitrogen fixers, phosphate solubilizers, cellulose decomposers, etc are prevalent in this ecosystem. Different groups of bacteria that get nourished by detritus and in turn, support the mangrove ecosystem in various ways. Distribution of bacteria depends on various physico-chemical parameters like water temperature, salinity, pH, available nutrients etc. Halophilic bacteria are believed to be predominant in the mangrove ecosystem as having high salinity. In one hand, microbes help in biogeochemical cycling of the nutrients in the ecosystem and on the other hand act as important source of food for a variety of marine organisms and maintain pristine nature of the environment. Mangroves are one of the most productive ecosystems contributing to the food chains of coastal oceanic areas and form a barrier against the waves and tides with their long spreading root system. Now a day, mangroves are among the most threatened habitats in the world disappearing at an accelerating rate, yet with little public notice. Biological diversity is a key issue of conservation and controlling the continuous degradation and destruction of mangroves, there is a critical need to understand them better. The microbial diversity and its distribution in a mangrove would improve our understanding of microbial functionality and their interactions found in that ecosystem7-9. In the present study microbial population and its interaction with nutrients have been investigated. Material and Methods Study Area: The Gulf of Kachchh, Gujarat, India lies approximately between latitudes 22° to 23°N and between longitudes 68° to 70° 30° E with an area of approximately 7300 Km. The climate is semi- arid and the maximum rainfall is of the order of 50 cm yr-1. There is no major river flow into it and hence little runoff has been observed. The sampling sites at various locations were chosen to get the broader picture of the interaction of various components (viz. anthropogenic input, natural impact etc.) and the sites were fixed by using GPS (figure 1). The major source of this sediment is considered to be the shore material and the load transported by the Indus River10. It is now recognized that the damming of the Indus River has drastically reduced the sediment delivery to the Arabian Sea11, 12. The reduction in sediment supply of the Indus River results changes in the Indus delta. The process of sediment supply to the Gulf of Kachchh via tidal erosion of the abandoned delta is still active. Many industries like metal smelting, cement, salt, textile, Ship dismantle and petrochemical refineries are situated nearby of towns like Jamnagar, Kandla, Mundra, Mandvi, Sikka and Jam-Salaya which are the potential source of pollution in the estuary of Gulf of Kachchh. The climate is semi-arid which influence the precipitation of air borne pollutants of the nearby cities/towns. International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 2(1), 25-30, January (2013) Int. Res. J. Environment Sci. International Science Congress Association 26 Figure-1 Showing study area in the Gulf of Kachchh, Gujarat, India Sample collection and preservation: The surface sediments from eleven different locations (upto10 cm depth) were collected from the inter-tidal regions of different mangroves at Mundra, Jodiya, Sachana, New Bedi Bandar, Old Bedi Bandar, Sikka, Jam-Salaya, Narara and Choreshwar during November 2008, taking consideration of anthropogenic input, mixing zones, natural weathering input zones, etc (figure 1). Since major part of the study area is under Marine National Park, conservation of mangroves was carried out at large scale by various governmental and non-governmental bodies. Sediment samples were collected in the pre-cleaned sterilized polythene bags and stored in the ice chest and transported to laboratory. Water samples were collected in pre-washed polythene bottles separately for nutrient analysis. The sediment samples were collected aseptically by removing off upper layer of the sediment. The samples were kept frozen in ice chest till the transfer in the laboratory where they were stored at 4o C. Nutrient analysis in surface water: The pH, temperature and total dissolved solids (TDS) in water were measured onsite using the Thermo-Orion water analysis kit (Model Beverly, MA, 01915). Salinity of the water samples were measured using refractometer. Nitrate (NO), sulphate (SO2-), bicarbonate (HCO), phosphate (PO3-) and inorganic silica (HSiO) were analyzed by using standard methods13. Microbial analysis: Sediments samples were processed for isolation of microbes. Sterilized Milli-Q water and lab wares were used during entire process of microbial analysis. One gram sediment sample was used for each inoculum. Serially diluted inoculums (10-8) were used for inoculums in pore-plating techniques for isolation of microbes14. The modified isolation media for bacteria contained Beef extract (3.0g), bacteriological Peptone (5.0g), NaNO3 (3.0g), KHPO4 (1.0g), MgSO.7HO (0.5 g), KCl (0.5 g), FeSO.7HO (0.01g), agar (15.0g), distilled water (1.0L), pH (6.8-7.0). Free living nitrogen fixer has been isolated in a selected medium, comprising mannitol (15.0g), KHPO (0.5g), MgSO.7HO (0.2g), CuSO (0.1g), NaCl (0.2g), CaCO3 (5.0g), agar (15.0g), distilled water (1.0L), pH maintained at 8.3. Phosphate solubilizing bacteria are isolated in the Pikovskayas as media, containing glucose (10.0g), Ca3 (PO (5.0 g), (NHSO (0.5g), KCl (0.2g), agar (20.0g), distilled water (1.0L), pH maintained at (6.8-7.0). Cellulose decomposers has been isolated in selective media containing KHPO (1.0g), CaCl (0.1g), MgSO.7HO (0.2g), CaSO4 (0.1g), NaCl (0.2g), NaNO3 (2.0g), Agar (12.0g), precipited cellulose (4.0g), distilled water (1.0L). Fungi isolated in the Czapedox agar media, which contained NaNO(3.0g), KHPO (1.0g), MgSO.7HO (0.5g), KCl (0.5g), FeSO.7HO (0.01g), sucrose (30g), agar (15.0g), ZnSO.7HO (0.05g), distilled water (1.0L). Isolated colonies were measured in colony formation unit (CFU per gram). CFU/g in original sample = No. of colonies counted/ {(dilution factor) x (volume plated, in ml)}. The total fungal load was calculated in terms of percentage occurrence, which may be expressed by the formula: Percentage occurrence = (No. of samples on which particular fungus is recorded x 100)/Total number of samples examined. International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 2(1), 25-30, January (2013) Int. Res. J. Environment Sci. International Science Congress Association 27 Table-1 Physical and chemical parameters of surface water and microbial load in the surface sediments Total bact. load Total fungal load Temp. C) pH Salinity TDS HCO NO PO3- SO2- HSiO Range 35-61 7-20 21-24 5.5-8.35 32-37 29.9-33.7 90-301.1 2.77-46.08 0.32-2.81 1892-3839.98 3.32-16.98 Mean± SD 47.7± 9.14 13± 4.02 22.8± 1.03 7.6± 0.77 33.9± 1.33 56.9± 81.63 173.2± 72.59 9.8± 14.44 1.3± 0.88 2635.6± 531.94 8.3± 4.88 Figure-2 Showing various parameters of surface water Results and Discussion Geochemical analysis of surface water: The environmental parameters showed wide variations at different locations in the study area depending upon the various factors such as topography, fresh water influx etc. The highest value of salinity was observed at Narara (35 ppt) and lowest at Mundra (32 ppt). Generally, estuarine mangrove waters have relatively low stocks of dissolved inorganic phosphorus and nitrogen15,16. High NO(46.08 mg/L) concentration was observed at Mundra (M8) indicates the impact of terrestrial runoff. The mean value and range of physico-chemical and microbial parameters of the study area are given in table-1. Water temperature ranged from 21C to 24C. Surface water temperature is influenced by the intensity of solar radiation, evaporation, fresh water influx and cooling and mixing up with ebb and flow from adjoining neritic waters17,18. This higher value of surface water temperature may be contributed by the low water level at Narara (M4), Jama-Salaya (M6) and Choreshwar (M7).The pH value of the surface water varied from 7.01 to 8.35 with maximum at Jam-Salaya (M6). Generally, variation in the pH value is attributed by factors like removal of CO by photosynthesis, sea water International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 2(1), 25-30, January (2013) Int. Res. J. Environment Sci. International Science Congress Association 28 dilution by fresh water influx, reduction in salinity and temperature and decomposition of organic matter19. The maximum value of bicarbonate (HCO) was reported at New Bedi Port (301 mg/L). This high value indicates the higher mixing of sea water (figure-2).Microbial population estimation: Population of the free living Nitrogen fixing, Phosphate solubilizing, Cellulose degrading bacteria have been analyzed in the surface sediments of various location (figure 1). Nitrogen fixing bacteria are known for improving nutrient status in sediments and hence to the mangrove plants20. Population of nitrate forming bacteria was reported highest at the locations like Old Bediport (M2), Narara (M4) and Mundra (M9) (4.0x103 cfu/g soil), indicate dominance of nitrogen fixing process which may be the reason for higher concentration of NOin the overlying water. Phosphate solubilizing bacteria may release phosphate ions from sparingly soluble inorganic and organic Phosphate compounds in the soil, which contribute with an increased phosphorus pool21. The Phosphate solubilizing bacteria varied from 3-17 x 10 cfu/g soil with highest at M6, M8 and M10 (17.0 x 10 cfu/g soil). The cellulose degrading bacterial population was also reported, which varied from 5-13 x10 cfu/g soils (figure-3). Besides assessment of microbial population dynamics, an attempt has also been made to identify four selected groups of bacteria such as nitrate forming, free living N2-fixing, phosphate solubilizing, and cellulose degrading. The microbial population in the sediment reflects the status of nutrient dynamics in the study area. Statistical analysis: Correlation analysis is a bivariate method commonly used to measure and establish the relationship between two variables and generally used to measure the degree of dependency of one variable to other. Strong correlation was observed between NO- and total fungal load (r=0.76) and between total bacterial load and PO3- (r=0.79) indicates these factors significantly influence the microbial population in the sediments (table 2). Lack of significant correlations among the nutrients indicate the influx of anthropogenic input and waste discharge containing nitrogen and phosphorous compounds from river runoff into these environments. Factor analysis is an important statistical method used to explain observed relationship among numerous variables. The mode of factor analysis is R-mode or Q-mode. Factor analysis is termed R-mode when the concern is interrelationships among the variables and Q-mode when concern is the interrelationships between samples22. R-mode factor analysis was used to identify major factors controlling the hydrochemistry of surface water of Gulf of Kachchh. Eigen value is greater than 1 reflects significant contribution of corresponding factor. Four factors with an Eigen value �1 were identified. These four factors explain about 86.73 % of the total variance. Factor 1 accounts for 26% variance in the dataset and shows high loading of PO3- and HSiO among nutrients and negative loading of SO4 2-. This factor explains contribution from silicate weathering, sulphate mineral dissolution, sulphide oxidation, phosphate mineral weathering or anthropogenic inputs. SO4 2- might have derived from sulphate mineral dissolution, hence showing negative loading in comparison to PO3- and HSiO. Factor 2 accounts for 22.4% variance in the dataset and shows strong loading of pH, HCO and HSiO, which describes contribution from weathering of carbonate silicate minerals. Factor 3 accounts for 21.4% variance in dataset and shows high loading of NO- among nutrients. This factor represents contribution from runoff and waste water discharge into the Gulf of Kachchh. Factor 4 contributes 16.7% variance in database and shows high loading of salinity and TDS (table 3). Figure-3 Microbial populations in the surface sediments of mangrove of Gulf of Kachchh, Gujarat International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 2(1), 25-30, January (2013) Int. Res. J. Environment Sci. International Science Congress Association 29 Table-2 Pearsons’ correlation matrix for total microbial load, nutrients in surface water and organic carbon content in the surface sediments Temp. (C) pH Salinity TDS HCO NO PO4 3- SO2- HSiOTotal bact. load Total fungal load Temp. ( o C) 1.00 pH -0.01 1.00 Salinity 0.50 0.10 1.00 TDS 0.08 0.09 -0.39 1.00 HCO 3 - 0.09 -0.73 -0.16 -0.06 1.00 NO 3 - -0.76 0.11 -0.46 -0.11 -0.22 1.00 PO 4 3 - 0.39 0.38 -0.11 0.28 -0.23 -0.04 1.00 SO 4 2 - -0.27 -0.11 0.11 0.08 -0.08 -0.06 -0.29 1.00 H 4 SiO 4 0.63 -0.49 0.11 0.05 0.70 -0.47 0.25 -0.49 1.00 Total bact. load 0.52 0.21 -0.08 0.11 -0.20 -0.04 0.79 -0.50 0.26 1.00 Total fungal load -0.49 0.22 -0.33 -0.20 -0.13 0.76 0.27 -0.49 -0.06 0.14 1 Table-3 Principal and varimax rotated R-mode factor loading matrix Variables Factor 1 Factor 2 Factor 3 Factor 4 Communities Temp o C 0.626 -0.199 -0.671 0.276 0.958 pH 0.298 0.810 - - 0.759 Salinity - - -0.286 0.937 0.963 TDS -0.236 0.213 - 0.871 0.867 HCO 3 -0.115 -0.909 - -0.157 0.872 NO 3 -0.164 0.234 0.868 -0.221 0.883 PO 4 0.818 0.277 - -0.186 0.783 SO 4 -0.724 0.301 -0.384 - 0.763 H 4 SiO 4 0.510 -0.809 -0.221 - 0.964 Total bacterial load 0.867 0.159 - - 0.785 Total fungal load 0.278 - 0.923 -0.114 0.944 Eigen value 3.196 2.868 2.428 1.048 % of variance 26.005 22.465 21.490 16.772 % of cumulative variance 26.005 48.471 69.961 86.733 Conclusion The source of nutrients in the mangroves of Gulf of Kachchh includes anthropogenic input and waste water discharge. A strong correlation between NO- and total fungal load (r=0.76) and between total bacterial load and PO3- (r=0.79) indicates these factors significantly influence the microbial population in the sediments. R-mode factor analysis indicates the anthropogenic input and various minerals dissolution contributed availability of nutrients in the study area. The available nutrients in the surface water directly influence the microbial status and nutrient dynamics in the sediment. Acknowledgement This research was supported by the University Grant Commission, India. The authors also thanks to the Forest Department of Gujarat for granting permission for current research work. Reference 1.Alongi D.M., Christoffersen P., Tirendi F., The influence of forest type on microbial-nutrient relationships in tropical mangrove sediments, J. Exp. Mar. Biol. Ecol.,171, 201-223(1993)2.Holguin G., Bashan Y., Mendoza-Salgado R.A., Amador E., Toledo G., Vazquez P., Amador A., La Microbiologia de los manglares. Bosques en la frontera entre el mar y la tierrra, Ciencia Desarrollo, 144, 26-35(1999)3.Holguin G., Vazquez P. and Bashan Y., The role of sediment microorganisms in the productivity, conservation and rehabitation mangrove ecosystems: an over view, Biol. Fertil. Soils, 33, 265-278 (2001)4.Capone D.G., Microbial nitrogen cycle, in: Manu- al of environmental microbiology (Ed: C.J. Aurstic and R.L. Crawford), ASM Press, Washington D.C., 439 (2002) International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 2(1), 25-30, January (2013) Int. Res. J. Environment Sci. International Science Congress Association 30 5.Alavandi S., Relationship between heterotrophic bacteria and suspended particulate matter in the Arabian Sea, Ind. J. Mar. Sci., 30, 89-92 (1990)6.Zaharan H.H., Moharram A.M. and Mohammad H.A., Some ecological and physiological studies on bacteria isolated from salt affected soils of Egypt, J. Basic Microbiol., 32, 405-413 (1992)7.Ozcelik R., Gul A.U., Merganic J. and Merganicova K., Tree species diversity its relationship to stand parameters and geomorphology features in the eastern black sea region forests of turkey, J. Environ. Biol., 29, 291-298 (2008)8.Kathiresan K., Why are mangroves degrading? Curr. Sci., 83, 1246-1254 (2002)9.Khan A.S. and Ali M.S., Mangroves- an ecosystem in peril, J. Curr. Sci., 10, 419-420 (2007)10.Zingde M.D., Pollution and its impact on ecology of the Gulf of Kachchh. Proceedings of the Workshop on Integrated Coastal and Marine Area Management Plan for the Gulf of Kachchh, Ahmadabad, 23–24 September 1999, organized by Dept of Ocean Development; Integrated Coastal and Marine Area Management Project Directorate; Chennai (India) and Space application Centre, Ahmadabad (New Delhi: Department of Ocean Development) 1–17 11.Giosan L., Clift P.D., Constantinescu S. and Tabrez A.R., Development of the Indus mega-delta. Abstracts of papers presented at the International Conference of Deltas (Mekong Venue) Geological Modeling and Management; Ho Chi Minh City, Vietnam Academy of Science and Technology 19, (2005)12.Giosan L., Constantinescu C., Clift P.D., Tabrez A.R., Danish M. and Inam A., Recent morphodynamics of the Indus delta shore and shelf, Mar. Geol.26 1668–1684(2006)13. APHA, AWWA and WEF, Standard methods for the Examination of Water and Wastewater, 21st ed. American Public Health Association, Washington, D.C (2005) 14.APHA, Standard methods for the Examination of Water and Wastewater, 20th edition, American Public Health Association, Washington, D.C (1998) 15.Alongi D.M., Boto K.G. and Robertson A.I., Nitrogen and phosphorus cycles. In:Tropical mangrove ecosystems. American Geophysical Union, Washington DC, 251-292 1992)16.Ramanathan A.L., Singh G., Majumdar J., Samal A.C., Chauhan R., Ranjan R.K., A study of microbial diversity and its interaction with nutrients in the sediments of Sundarban mangroves, Ind. J. Mar. Sci.37, 159-165 (2008) 17.Govindasamy C., Kannan L. and Azaiah J., Seasonal variation in physic-chemical properties and primary production in the coastal water biotopes of Coromandel Coast, India, J. Environ. Biol., 21, 1-7 (2000) 18.Arthur R., Coral bleaching and mortality in three Indian reef regions during an El Nino southern oscillation event, Curr. Sci., 79, 12 (2000)19.Chauhan R. and Ramanathan A.L., Evaluation of water quality of Bhitarkanika mangrove ecosystem, Orissa, east coast of India, Ind. J. Mar. Sci, 37 (2), 153-158 (2008)20.Robert C. R. and Virginia B. S., Evaluation of nutrient agar bile salts medium to selectively culture Vibrio cholera, Phil J Microbiol Infect Dis, 10(1), 18-20 (1981)21.Furumai H., Kondo T. and Ohgaki S., Phosphorus exchange kinetics and exchangeable phosphorus forms in sediments. Water Res., 21, 685-691(1989) 22.Singh V.B., Ramanathan A.L., Jose P.G., Sharma P., Linda A., Azam M.F. and Chatterjee C., Chemical characterisation of meltwater draining from Gangotri Glacier, Garhwal Himalaya, India. J. Earth Syst. Sci., 121(3), 625–636(2012)