Mobilization of manganese by basalt associated Mn(II)-oxidizing bacteria from the Indian Ridge System.

The Indian Ridge System basalt bearing Mn-oxide coatings had todorokite as the major and birnesite as the minor mineral. We posit that microorganisms associated with these basalts participate in the oxidation of Mn and contribute to mineral deposition. We also hypothesized that, the Mn-oxidizing microbes may respond reversibly to pulses of fresh organic carbon introduced into the water column by mobilizing the Mn in Mn-oxides. To test these two hypotheses, we enumerated the number of Mn-oxidizers and -reducers and carried out studies on the mobilization of Mn by microbial communities associated with basalt. In medium containing 100 µM Mn(2+), 10(3) colony forming units (CFU) were recovered with undetectable number of reducers on Mn-oxide amended medium, suggesting that the community was more oxidative. Experiments were then conducted with basalt fragments at 4±2 °C in the presence 'G(+)' and absence 'G(-)' of glucose (0.1%). Controls included set-ups, some of which were poisoned with 15 mM azide and the others of which were heat-killed. The mobilization of Mn in the presence of glucose was 1.76 µg g(-1) d(-1) and in the absence, it was 0.17 µg g(-1) d(-1) after 150 d. Mn mobilization with and without added glucose was 13 and 4 times greater than the corresponding azide treated controls. However, rates in 'G(+)' were 16 times and 'G(-)' 24 times more than the respective heat killed controls. The corresponding total counts in the presence of added glucose increased from 1.63×10(6) to 6.71×10(7) cells g(-1) and from 1.41×10(7) to 3.52×10(7) cells g(-1) in its absence. Thus, the addition of glucose as a proxy for organic carbon changed the community's response from Mn(II)-oxidizing to Mn(IV)-reducing activity. The results confirm the participation of Mn oxidizing bacteria in the mobilization of Mn. Identification of culturable bacteria by 16S rRNA gene analysis showed taxonomic affiliations to Bacillus, Exiguobacterium, Staphylococcus, Brevibacterium and Alcanivorax sp.

Mixed response in bacterial and biochemical variables to simulated sand mining in placer-rich beach sediments, Ratnagiri, West coast of India.

We investigated the influence on bacterial community and biochemical variables through mechanical disturbance of sediment-akin to small-scale mining in Kalbadevi beach, Ratnagiri, a placer-rich beach ecosystem which is a potential mining site. Changes were investigated by comparing three periods, namely phase I before disturbance, phase II just after disturbance, and phase III 24 h after disturbance as the bacterial generation time is ≤7 h. Cores from dune, berm, high-, mid-, and low-tide were examined for changes in distribution of total bacterial abundance, total direct viability (counts under aerobic and anaerobic conditions), culturability and biochemical parameters up to 40 cm depth. Results showed that bacterial abundance decreased by an order from 10(6) cells g(-1) sediment, while, viability reduced marginally. Culturability on different-strength nutrient broth increased by 155% during phase II. Changes in sedimentary proteins, carbohydrates, and lipids were marked at berm and dune and masked at other levels by tidal influence. Sedimentary ATP reduced drastically. During phase III, Pearson's correlation between these variables evolved from non-significant to significant level. Thus, simulated disturbance had a mixed effect on bacterial and biochemical variables of the sediments. It had a negative impact on bacterial abundance, viability and ATP but positive impact on culturability. Viability, culturability, and ATP could act as important indicators reflecting the disturbance in the system at short time intervals. Culturability, which improved by an order, could perhaps be a fraction that contributes to restoration of the system at bacterial level. This baseline information about the potential mining site could help in developing rational approach towards sustainable harnessing of resources with minimum damage to the ecosystem.

Heavy metal pollution exerts reduction/adaptation in the diversity and enzyme expression profile of heterotrophic bacteria in Cochin estuary, India.

Over the past three decades heavy metal pollution has increased substantially in Cochin estuary, south west coast of India. Here we studied the distribution, diversity and enzyme expression profile of culturable microbial population along a pollution gradient. The distribution of resistance against 5 mM concentration of Zn, Co, Ni and Cu was observed among 90-100% of bacterial isolates retrieved from highly polluted Eloor, whereas it was less than 40% in Vypin and Munambam. Similarly, there was a difference in the distribution and diversity of bacterial phyla with predominance of Proteobacteria in Eloor and Firmicutes in Munambam and Vypin. We observed that 75-100% of the organisms retrieved from Eloor had low levels of expression for hydrolytic enzyme. In conclusion, the heavy metal pollution in Cochin estuary brought in reduction/adaptation in the distribution, diversity and enzyme expression profile of bacteria, which may impart adverse impacts on ecosystem functioning.

Diversity in transcripts and translational pattern of stress proteins in marine extremophiles.

Extremophiles occur in a diverse range of habitats, from the frigid waters of Antarctic to the superheated plumes of hydrothermal vents. Their in-depth study could provide important insights into the biochemical, ecological and evolutionary aspects of marine microbes. The cellular machinery of such extreme-lovers could be highly flexible to cope with such harsh environments. Extreme conditions of temperature, pressure, salinity, pH, oxidative stress, radiation, etc., above the physiological tolerance level can disrupt the natural conformation of proteins in the cell. The induction of stress proteins (heat/cold shock proteins/salt stress proteins/pressure-induced proteins) plays a vital role in the acclimatization of extremophiles. The present review focuses on the in vitro studies conducted on the transcripts and translational pattern of stress proteins in extremophiles. Though some proteins are unique, a commonality in stress resistance mechanism has been observed, for example, the universal occurrence of HSP60, 70 and the expression of metabolic and DNA repair proteins. The review highlights that among all the stressful conditions, salt/osmotic stress evokes the expression of highest number of transcripts/proteins while psychrophilic condition the least.

Water quality and bacteriology in an aquaculture facility equipped with a new aeration system.

The HOBAS aeration system was tested to compare changes in environmental and bacteriological parameters in ponds growing Penaeus monodon during a single production cycle. The stocking density in the aerated pond was doubled to 12 post-larvae (PL) m(-2) in contrast to the non-aerated pond with 6 (PL) m(-2). Microbial abundance in the ponds ranged between 10(5-6) cells ml(-1). Among the physiological groups of bacteria enumerated, the heterotrophs dominated with an abundance of 10(4) CFU ml(-1). Of the nitrogen and sulfur cycle bacteria, the nitrifiers flourished in the aerated pond and could maintain ammonia-N concentration within permissible levels. Bacterial activity also maintained sulfide concentrations at < 0.03 mg l(-1). Non-aerated conditions promoted denitrification maintaining nitrate concentration between 0.32 and 0.98 microM NO(3)(-)-N l(-1). However, a marked increase in ammonium content was observed in the non-aerated pond at the end of the culture period. Thus in high-density ponds, the aerators served to stimulate bacterial growth and activity which consequently maintained the quality of the water to match that of low-density ponds. Accordingly, these aerators could be effectively used to sustain higher yields. The effluent from the aerated pond is less likely to alter the redox balance of the receiving waters.

Anoxia over the western continental shelf of India: bacterial indications of intrinsic nitrification feeding denitrification.

Studies on the Arabian Sea coastal anoxia have been of immense interest, but despite its ecological significance there is sparse understanding of the microbes involved. Hence, observations were carried out off Goa (15 degrees 30'N, 72 degrees 40'E to 15 degrees 30'N, 72 degrees 59'E) to understand the processes that mediate the changes in various inorganic nitrogen species in the water column during anoxia. Water column chemistry showed a clear distinct oxic environment in the month of April and anoxic condition in October. Our study based on microbial signatures indicated that oxygen deficit appeared as a well-defined nucleus almost 40 km away from the coast during the oxic period (April) and spreads there after to the entire water column synchronizing with the water chemistry. Striking results of net changes in inorganic nitrogen species in nitrification blocked and unblocked experimental systems show that denitrification is the predominant process in the water column consuming available nitrate ( approximately 0.5 microM) to near zero levels within approximately 72 h of incubation. These observations have been supported by concomitant increase in nitrite concentration ( approximately 4 microM). Similar studies on denitrification-blocked incubations, demonstrate the potential of nitrification to feed denitrification. Nitrification could contribute almost 4.5 microM to the total nitrate pool. It was found that the relation between ammonium and total dissolved inorganic nitrogen (DIN) pool (r=0.98, p<0.001, n=122) was significant compared to the latter with nitrite and nitrate. The occurrence of high ammonium under low phosphate conditions corroborates our observations that ammonium does not appear to be locked under low oxygen regimes. It is suggested that ammonium actively produced by detrital breakdown (ammonification) is efficiently consumed through nitrification process. The three processes in concert viz. ammonification, nitrification and denitrification appear to operate in more temporal and spatial proximity than hitherto appreciated in these systems and this gives additional cues on the absence of measurable nitrate at surface waters, which was earlier attributed only to efficient algal uptake. Hence we hypothesize that the alarming nitrous oxide input into the atmosphere could be due to high productivity driven tighter nitrification-denitrification coupling, rather than denitrification driven by extraneous nitrate.

Bacterial contribution to mitigation of iron and manganese in mangrove sediments.

The Mandovi and Chapora are two tropical estuaries lying in close geographic proximity on the west coast of India. Seasonal changes in down core variation of Fe, Mn and Total Organic Carbon (TOC) in the mangrove sediments adjoining these estuaries were studied to assess their influence on some of the representative benthic bacteria belonging to heterotrophic and autotrophic groups. Heterotrophic bacteria (HB) cultured on different nutrient concentrations (0.01%, 0.1% and 25%) together with nitrifiers (NtB; representating autotroph) were chosen to assess the influence of the above-mentioned abiotic parameters on the former. The experimental site located along the Mandovi is under the influence of extensive ferromanganese ore mining, while the control site at Chapora is relatively free from such influences. Geoaccumulation index computed for Mandovi showed that sediments (0-10cm) were 'uncontaminated to moderately contaminated' by Fe during the pre monsoon and monsoon seasons, while in the post monsoon season the 4-10cm fraction was almost completely restored from contamination. Similar computations for Mn showed that in pre monsoon, sediments fell in the 'moderately contaminated' and 'moderately to strongly contaminated' categories, while in the monsoon and post monsoon seasons all the sections were 'Uncontaminated'. The difference observed in correlation between Fe and Mn with the various fractions of heterotrophs and nitrifiers indicated that though these two elements shared a similar chemistry in the environment, microbes involved in biogeochemical processes might prefer them differentially. The relationship between TOC and HB enumerated on 0.01% dilute nutrient agar remained at r=0.50, p<0.05 throughout the year. Hence, it could be apparently linked to their preferred concentration of organic carbon requirement. A relationship of r=0.61, p<0.01 between manganese concentration and heterotrophs recovered on different strengths of nutrient agar is suggestive of their response to the metal enrichment. They could thus contribute towards maintaining the level of Mn at par with reference levels at Chapora. A positive correlation between Mn with NtB (n=10, p<0.05, r=0.58) at the experimental site during the non-monsoon months is suggestive of the latter's contribution to regulation of the metal concentration in the sediment probably through anaerobic nitrification at the expense of manganese. The study therefore supports our hypothesis that both autochthonous autotrophs and heterotrophs work in tandem to mitigate concentration of Mn and related metals in mangrove sediments.

"Trade-off" in Antarctic bacteria: limnetic psychrotrophs concede multiple enzyme expressions for multiple metal resistance.

The present study examines the metal and antibiotic resistant bacteria in ice and water from lakes east and west of the Indian base camp (Maitri) in Antarctica. The isolates from western and eastern lakes showed distinct geographical differences in properties like metal resistance and enzyme expression. This may be attributed to high organic loading in the lakes on the west of Maitri. However, there was no marked geopraphical distinction in antibiotic resistance between the lakes. Bacteria from the lakes on the eastern side showed resistance to three or more metals including mercury while, those from the western were resistant to only 1-2 metals excluding mercury. Multiple enzyme expression was more pronounced in the lakes on the western side. On the eastern side multiple metal resistance was encountered in bacterial isolates associated with fewer enzyme expressions suggesting a "trade-off". Thus these Antarctic isolates from the east trade their ability to express multiple enzymes for developing resistance to multiple metals including mercury.

Metal and antibiotic-resistance in psychrotrophic bacteria from Antarctic Marine waters.

In the wake of the findings that Antarctic krills concentrate heavy metals at ppm level, (Yamamoto et al. 1987), the Antarctic waters from the Indian side were examined for the incidence of metal and antibiotic-resistant bacteria during the austral summer (13th Indian Antarctic expedition) along the cruise track extending from 50 degrees S and 18 degrees E to 65 degrees S and 30 degrees E. The bacterial isolates from these waters showed varying degrees of resistance to antibiotics (Chloramphenicol, ampicillin, streptomycin, tetracycline and kanamycin) and metals (K(2)CrO(4), CdCl(2), ZnCl(2) and HgCl(2)) tested. Of the isolates screened, about 29% and 16% were resistant to 100 ppm of cadmium and chromium salt respectively. Tolerance to lower concentration (10 ppm) of mercury (Hg) was observed in 68% of the isolates. Depending on the antibiotics the isolates showed different percentage of resistance. Multiple drug and metal-resistance were observed. High incidence of resistance to both antibiotics and metals were common among the pigmented bacterial isolates. Increased resistance decreased the ability of bacteria to express enzymes. The results reiterate previous findings by other researchers that the waters of southern ocean may not be exempt from the spread of metal and antibiotic-resistance.

Manganese oxidation by bacterial isolates from the Indian ridge system.

The abundance and activity of culturable manganese-oxidizing bacteria were assessed from near-bottom water samples of the tectonically active Carlsberg Ridge. Retrievable counts as colony forming units (CFU) on dilute nutrient agar medium (dilNA=2 gm l(-1) nutrient broth+2% agar) and on dilNA supplemented with 1, 2 and 3 mM MnCl(2).4H(2)O were in the order of 10(6) CFU l(-1). Retrievability of heterotrophs ranged from non-detectable levels (ND) to 2.82 x 10(6) CFU l(-1). The retrievable counts on Mn amended dilNA ranged from ND to 3.21 x 10(6), 1.47 x 10(6) and 1.45 x 10(6) CFU l(-1) on 1, 2 and 3 mM, respectively. About 87% of the Mn tolerant isolates (n=39) showed taxonomic affinities to Pseudomonas I and II sp. Two representative strains CR35 and CR48 (CR-Carlsberg Ridge) isolated on manganese-supplemented media were tested for their ability to tolerate a range of Mn amendments from 1 nM to 100 mM in terms of growth and respiration. CR35 represents 66% of the total CFU (3.04 x 10(6) CFU l(-1)), while CR48 represented only 6% of the total CFU (1.05 x 10(6) CFU l(-1)). The colonies of these two isolates were dark brown in color suggesting precipitation of Mn as oxide. Tests for the effect on growth and respiration were conducted in media simulating heterotrophic (amended with 0.01% glucose) and lithotrophic (unamended) conditions. Maximum stimulation in growth and respiration of CR35 occurred at 100 microM Mn both in unamended and amended media. At levels of Mn greater than 100 microM the counts decreased steadily. Total respiring cells of CR48 were stimulated to a maximum at 1 microM Mn in unamended medium and 1 nM in amended medium. Total cells counts for the same decreased beyond 100 microM Mn in unamended and 1 nM in amended medium. The isolates were tested for their ability to oxidize Mn amendments from 1 microM to 10 mM Mn. At the end of a 76-day incubation period, there was evidence of manganese oxide precipitation at high Mn concentrations (>or=1 mM) as a dark brown coloration on the sides of culture tubes. Highest Mn oxidation rates were observed at 10 mM Mn(II) concentration with CR35 oxidizing 27 and 25 microM Mn day(-1) in unamended and amended condition, respectively. CR48 oxidized Mn at the rate of 26 microM Mn day(-1) in unamended medium and 35 microM Mn day(-1) in amended medium. Scanning electron microscope (SEM) observations of both isolates revealed free-living cells in clustered matrices approximately 2 microm diameter. Energy dispersive spectrum of the cell matrix of CR35 cultured in 1 mM Mn detected 30% Mn, while the cell aggregates of CR48 harbored 7-10% Mn. The relatively high specific activity of these mixotrophic bacteria under relatively oligotrophic conditions suggests that they may be responsible for scavenging dissolved Mn from the Carlsberg Ridge waters and could potentially participate in oxidation.

Representation, dispersion, and variation of bacterial indicators in the coastal waters of Nagore (east coast of India).

The occurrence of bacterial indicators in the coastal waters of Nagore on the east coast of India was studied during two monsoon and nonmonsoon cycles from 1995 to1996. Proteus/Klebsiella, Vibrio parahaemolyticus, and total coliforms were typically present at a mean level of 10(9) colony-forming units/g dry sediment, whereas other indicators such as Salmonella, Shigella, and V. cholerae were at the 10(5) level. In the water column, total coliforms and Salmonella were 2 to 3 orders of magnitude lower than in sediment. Higher numbers of indicators were retrieved at 10- and 15-m depth lines than at a 5-m depth line. While southwest monsoon caused increased bacterial input to the coastal sediment of Nagore, cyclonic effect on indicators was observed only in the water samples. Although physicochemical parameters did not influence the variation in coliform population significantly, most of the indicators in the sediment showed significant negative correlation with chlorophyll a, indicating an inverse relationship between them. With ever increasing development of Nagore, nonpoint sources of pollution may become a serious problem. Continued monitoring of these groups of bacteria over longer periods of time would help develop models for their regulation and may suggest remedial measures.

Mercury and lead tolerance in hypersaline sulfate-reducing bacteria.

Sulfate-reducing bacteria (SRB) HSR1, HSR4, and HSR14 isolated from the salt pans of Goa grew best at 90-100/1000 salinity on substrates like formate, acetate, lactate, butyrate, ethanol and benzoate. They were gram negative, non-sporulating, non-motile rods lacking in desulfoviridin and cytochromes. Examination of these isolates for heavy metal tolerance and response studies in terms of growth and sulfate-reducing activity (SRA) were carried out using HgCl2 and Pb(NO3)2 at final concentration of 50, 100, and 200 and 100, 200 and 500 microg ml(-1) respectively. With Hg, HSR1 showed approximately 80% of the control's growth at 100 and 200 microg ml(-1) but SRA reached only 60% of the control values at the end of 14 days. HSR14 could reach >100% of the control's growth at 200 microg ml(-1) but the SRA reached only up to 60% of the control without metal at 100 microg ml(-1). Though the concentration of Pb was double that of Hg, HSR4 could grow and respire better than the control, the growth being stimulated by 160% and respiration by 170% in the presence of 500 microg ml(-1) of Pb(NO3)2. It is probable that some hypersaline SRB are more tolerant to heavy metals than the mesohaline counterparts and could be more effectively used for precipitating these metals in bioremediatory measures. Further examination of their responses to varied concentration of metals under different salinities would indicate their range of applicability.

Effect of lead, mercury and cadmium on a sulphate-reducing bacterium.

A sulphate-reducing bacterial strain isolated from the south-west coast of India resembling Desulfosarcina in its physiology was tested for its behaviour towards HgCl(2), CdSO(4) and Pb(NO(3))(2). The order of toxicity to growth of these metal salts in a lactate-based medium at 50 microg ml(-1) concentrations was Cd>Pb>Hg and to respiration Pb>Cd>Hg. Inhibitory concentrations (viz. 100 microg ml(-1) of HgCl(2) and 200 microg ml(-1) of Pb(NO(3)(2)) had a stimulatory effect when the substrate was changed to acetate. With sodium acetate at 0.1% concentration, Hg and Pb had maximum stimulatory effect for growth and sulphide production. Experiments conducted directly with sediment slurries amended with lactate showed that all three metals (at levels below their inhibitory concentrations, i.e. 50 microg ml(-1) of metal salt for Cd and Hg and 100 microg ml(-1) for Pb) inhibited sulphate-reducing activity (SRA) with Pb decreasing the peak production by 68%. The order of toxicity in both lactate and acetate-amended slurry was Pb>Cd>Hg and Pb>Hg>Cd, respectively. With acetate, SRA in the presence of Cd and Hg was stimulated 110% and 27%, respectively. Pb inhibited SRA by 11%. There is a general reduction in the inhibition of sulphide production in slurries as compared with pure culture of the isolate.