Estuarine mangrove niches select cultivable heterotrophic diazotrophs with diverse metabolic potentials-a prospective cross-dialog for functional diazotrophy.

Introduction: Biological nitrogen fixation (BNF), an unparalleled metabolic novelty among living microorganisms on earth, globally contributes ~88-101 Tg N year-1 to natural ecosystems, ~56% sourced from symbiotic BNF while ~22-45% derived from free-living nitrogen fixers (FLNF). The success of symbiotic BNF is largely dependent on its interaction with host-plant, however ubiquitous environmental heterotrophic FLNFs face many limitations in their immediate ecological niches to sustain unhindered BNF. The autotrophic FLNFs like cyanobacteria and oceanic heterotrophic diazotrophs have been well studied about their contrivances acclimated/adapted by these organisms to outwit the environmental constraints for functional diazotrophy. However, FLNF heterotrophs face more adversity in executing BNF under stressful estuarine/marine/aquatic habitats. Methods: In this study a large-scale cultivation-dependent investigation was accomplished with 190 NCBI accessioned and 45 non-accessioned heterotrophic FLNF cultivable bacterial isolates (total 235) from halophilic estuarine intertidal mangrove niches of Indian Sundarbans, a Ramsar site and UNESCO proclaimed World Heritage Site. Assuming ~1% culturability of the microbial community, the respective niches were also studied for representing actual bacterial diversity via cultivation-independent next-generation sequencing of V3-V4 rRNA regions. Results: Both the studies revealed a higher abundance of culturable Gammaproteobacteria followed by Firmicutes, the majority of 235 FLNFs studied belonging to these two classes. The FLNFs displayed comparable selection potential in media for free nitrogen fixers and iron-oxidizing bacteria, linking diazotrophy with iron oxidation, siderophore production, phosphorus solubilization, phosphorus uptake and accumulation as well as denitrification. Discussion: This observation validated the hypothesis that under extreme estuarine mangrove niches, diazotrophs are naturally selected as a specialized multidimensional entity, to expedite BNF and survive. Earlier metagenome data from mangrove niches demonstrated a microbial metabolic coupling among C, N, P, S, and Fe cycling in mangrove sediments, as an adaptive trait, evident with the co-abundant respective functional genes, which corroborates our findings in cultivation mode for multiple interrelated metabolic potential facilitating BNF in a challenging intertidal mangrove environment.

Forecasting mangrove ecosystem degradation utilizing quantifiable eco-physiological resilience -A study from Indian Sundarbans.

Sundarbans mangrove forest, the world's largest continuous mangrove forests expanding across India and Bangladesh, in recent times, is immensely threatened by degradation stress due to natural stressors and anthropogenic disturbances. The degradation across the 19 mangrove forests in Indian Sundarbans was evaluated by eight environmental criteria typical to mangrove ecosystem. In an attempt to find competent predictors for mangrove ecosystem degradation, key eco-physiological resilience trait complex specific for mangroves from 4922 individuals for physiological analyses with gene expression and 603 individuals for leaf tissue distributions from 16 mangroves and 15 associate species was assessed along the degradation gradient. The degradation data was apparently categorized into four and CDFA discriminates 97% of the eco-physiological resilience data into corresponding four groups. Predictive Bayesian regression models and mixed effects models indicate osmolyte accumulation and thickness of water storage tissue as primary predictors of each of the degradation criteria that appraise the degradation status of mangrove ecosystem. RDA analyses well represented response variables of degradation explained by explanatory resilience variables. We hypothesize that with the help of our predictive models the policy makers could trace even the cryptic process of mangrove degradation and save the respective forests in time by proposing appropriate action plans.

Effect of Varying Nitrate Concentrations on Denitrifying Phosphorus Uptake by DPAOs With a Molecular Insight Into Pho Regulon Gene Expression.

Bacterial Pho regulon is a key regulator component in biological phosphorus-uptake. Poly-phosphate accumulating bacteria used in enhanced biological phosphorus removal (EBPR) system encounter negative regulation of the Pho regulon, resulting in reduced phosphorus-uptake from phosphorus-replete waste effluents. This study demonstrates possible trends of overcoming the PhoU negative regulation, resulting in excessive PO4 3--P uptake at varying concentrations of NO3 --N through denitrifying phosphorus removal process. We investigated the Pho regulon gene expression pattern and kinetic studies of P-removal by denitrifying phosphate accumulating organisms (DPAOs) which are able to remove both PO4 3--P and NO3 --N in single anoxic stage with the utilization of external carbon sources, without the use of stored polyhydroxyalkanoate (PHA) and without any anaerobic-aerobic or anaerobic-anoxic switches. Our study establishes that a minimum addition of 100 ppm NO3 --N leads to the withdrawal of the negative regulation of Pho regulon and results in ∼100% P-removal with concomitant escalated poly-phosphate accumulation by our established DPAO isolates and their artificially made consortium, isolated from sludge sample of PO4 3- -rich parboiled rice mill effluent, in a settling tank within 12 h of treatment. The same results were obtained when a phosphate rich effluent (stillage from distillery) mixed with a nitrate rich effluent (from explosive industry) was treated together in a single phase anoxic batch reactor, eliminating the need for alternating anaerobic/aerobic or anaerobic/anoxic switches for removing both the pollutants simultaneously. The highest poly-phosphate accumulation was observed to be more than 17% of cell dry weight. Our studies unequivocally establish that nitrate induction of Pho regulon is parallely associated with the repression of PhoU gene transcription, which is the negative regulator of Pho regulon. Based on earlier observations where similar nitrate mediated transcriptional repression was cited, we hypothesize the possible involvement of NarL/NarP transcriptional regulator proteins in PhoU repression. At present, we propose this denitrifying phosphorus removal endeavor as an innovative methodology to overcome the negative regulation of Pho regulon for accelerated unhindered phosphorus remediation from phosphate rich wastewater in India and the developing world where the stringency of EBPR and other reactors prevent their use due to financial reasons.