Debjani Mandal

Arsenic (As) contamination of groundwater is a common problem in BDP. The alkaline pH of the groundwater provides favourable condition for geomicrobiochemical processes, where organic matter also plays an important role in mobilization of As. The total arsenic content (AsT) of the groundwater is in the range of 69-178 µgL-1 with mean value of 118.58 µgL-1. The groundwater quality of monitored drinking water wells are related to local land–use pattern and rural sanitation installation and practices (which can have a strong influence on microbial contamination). The study indicates that As heterogeneity, probably a consequence of microbial heterogeneity is a feature of the study area. The excessive total and fecal coliform (TC and FC) count of the ponds possibly suggest uncontrolled surface run offs and sewage leakage, notably from nearby pit-latrine, which increased the microbial contamination in these water sources. The tube wells located in close proximity (~ 2-3 m) of the surveyed ponds are also contaminated with high TC and FC values, due to faulty water supply system, pond water infiltration and cracked tube well platforms. Coliforms may influence geomicrobiological processes in groundwater and thereby, contribute in As release and mobilization. The TC and FC values of monitored tube wells and ponds show seasonal variation, with high TC and FC values during wet season. Attempt has been made to focus on sanitation risk of the monitored wells (based on sanitation and fecal coliform scores) which is usually less explored. The sanitary risk score reveal the need of immediate attention to all the sites, as 25.0 %, 58.3 %, 11.1 % and 5 % of tube wells of the six sites fell under very high risk (sanitary risk score ≥ 9), high risk (sanitary risk score 6-8), intermediate risk (sanitary risk score 3-5) and low risk category (sanitary risk score 0-2), respectively. Fecal coliform score classified all the sites under intermediate risk category (FC count 10-100 per 100 ml of water). The predominance of As heterogeneity particularly in areas under high risk category (showing high TC and FC contamination) is observed along with the similar sinusoidal pattern of TC, FC and AsT content. Finally, this study shed light on the factors responsible for unsafe nature of monitored tube wells and the influence of anthropogenic activities on the microenvironment of the groundwater, which could be a probable reason behind the As heterogeneity in the groundwater of the study area.

YspD is a hydrophilic translocator forming the platform for assemblage of functional translocon. Exposure to the extra-cellular milieu makes YspD a potential therapeutic target. DoGSiteScorer predicted best druggable pocket (P0) within YspD, encompassing predominantly the C-terminal helical bundles and the long helices-9 & 5. COACH metaserver also identified ligand binding residues within the aforementioned druggable pocket mapping to helix-9. Amino acids of helix-9 are involved in oligomerization of YspD. Interaction of helix-9 and parts of C-terminal of YspD with hydrophobic translocator protein (YspB), is essential for translocation of bacterial effectors to initiate an infection. Helices-9 & 5 form an intramolecular coiled-coil structure, required for protein–protein interaction. Targeting intramolecular coiled-coil and parts of C-terminal would be important for functional inactivation of YspD. Solvent exposed surface in YspD, particularly in P0, enhances its accessibility to ligands. Nine small molecular inhibitors of TIIISS were identified and retrieved from ZINC15 database (drug-library) as putative drug candidates. Molecular docking of potential ligands with P0 was done using SwissDock server and Achilles Blind Docking server. Considering the “Significance” threshold of binding score and region of interaction, Salicylidene Acyl Hydrazide derivatives (INP0400) and Phenoxyacetamide derivative (MBX1641) were found to bind effectively with YspD. These potential ligands interact with functional domains of YspD including parts of C-terminal and the intramolecular coiled-coil, which may affect the oligomerization of YspD and disrupt the interaction of YspD with YspB, inhibiting formation of functional translocon. The identified small molecular antimicrobial ligands of YspD could be tested in vivo to attenuate Y. enterocolitica infection by deregulation of Ysa-Ysp TIIISS.

The soil and groundwater of the Bhagobangola I block of Murshidabad district, West Bengal, India is severely arsenic-contaminated. A bacterium was isolated from the garden soil of the Mahishasthali village, which could tolerate 36.49 mM arsenic (III), 280.44 mM arsenic (V) and 63 mM chromium (III), which makes it arsenic (III and V) and chromium (III) hyper-tolerant bacterium. The growth pattern of this bacterium does not show much alteration in the presence of 10 mM arsenic (III) and chromium (III), emphasizing its resistance to these heavy metals. Scanning electron microscopic analysis depicted this bacterium to be rod-shaped with a size of ~1.45 µm. 16S rDNA sequencing, followed by subsequent phylogenetic analysis, established the identity of this bacterium as Microbacterium paraoxydans. This bacterium is capable of bioremediation of arsenic and showed 30.8% and 35.2% of bioremediation for 1mM and 22.6%, and 30.5% of bioremediation for 4mM arsenite, over a period of 24 and 48 h, respectively. Microbacterium paraoxydans also exhibits potential plant growth-promoting properties such as nitrogen fixation, phosphate solubilization, indole-3-acetic acid production and production of siderophores. Therefore, the heavy metal resistance, bioremediation potential and plant growth-promoting potential of the bacterium could be utilized not only for reduction in arsenic toxicity in soil and groundwater but also for plant growth promotion.

The soil and groundwater of Bhagobangola I block of Murshidabad district, West Bengal, India is severely arsenic contaminated. A bacterium was isolated from the garden soil of Mahishasthali village, which could tolerate 36.49 mM arsenic (III), 280.44 mM arsenic (V) and 63 mM chromium (III), which makes it arsenic (III & V) and chromium (III) hyper-tolerant bacterium. The growth pattern of this bacterium does not show much alteration in presence of 10 mM arsenic (III) and chromium (III), emphasizing its resistance to these heavy metals. Scanning electron microscopy depicted the size of this bacterium to be ~1.45 µm. 16S rDNA sequencing followed by subsequent phylogenetic analysis established the identity of this bacterium as Microbacterium paraoxydans. The bacterium is capable of bioremediation of arsenic and showed 30.8% & 22.6%, and 35.2% and 30.5% of bioremediation over a period of 24 and 48 hours in 1 mM and 4 mM arsenite, respectively. Microbacterium paraoxydans also exhibit plant growth promoting properties like nitrogen fixation, phosphate solubilization, indole-3-acetic acid production and production of siderophores. Therefore, the heavy metal resistance, bioremediation potential and plant growth promoting potential of the bacterium could be utilized not only for reduction of arsenic toxicity in soil and groundwater but also for plant growth promotion by using it as a biofertilizer.

Introduction:Yersinia enterocolitica is an opportunistic pathogen which causes enteric diseases like gastrointestinal and mesenteric adenitis in immune-compromised individuals. The gastrointestinal phase of Y. enterocolitica infection is mediated by Yersinia secretion apparatus- Yersinia secretion protein (Ysa-Ysp) Type III Secretion System (T3SS). Certain serotypes of Y. enterocolitica Biovar 1B exhibit enhanced virulence due to presence of Ysa-Ysp T3SS. Translocator proteins form the translocation apparatus (translocon) at the tip of the needle complex of the T3SS. YspC is a minor hydrophobic translocator protein which is sequestered in the bacterial cytoplasm by its cognate chaperone SycB. SycB plays the dual role of a class II chaperone and a regulator of Ysa-Ysp T3SS.Experiments and key results findings: Homology model of SycB depicts a structure with a concave core formed by tetratricopeptide repeats (TPRs) and a flexible N- terminal helix. Deletion mutants of SycB showed that the N-terminal helix of SycB is responsible for its dimerization, which is further corroborated by molecular docking analysis. The dimeric state of SycB dissociates during the interaction with YspC due to steric hindrances. And it forms a 1:1 heterodimeric YspC-SycB complex as confirmed by size-exclusion chromatography, chemical cross-linking and molecular docking studies. FRET analysis indicated that the tyrosine residues present in first two TPRs of SycB is responsible for its interaction with YspC. Deletion mutants of SycB possessing the first two TPR regions interacted with YspC, as also depicted by the YspC-SycB interaction model. YspC is a unique translocator protein with high stability, rigid tertiary structure and monomeric form unlike any other translocator proteins. It shows structural alteration in the complex formed with SycB as shown by spectroscopic data and proteolytic digestion. YspC has a Y-shaped three dimensional structure and SycB completely localizes within the fork formed by the two arms of Y-shaped YspC. This structural analysis and mechanism of interaction between YspC and SycB would be potentially beneficial for designing drugs to deregulate the Ysa-Ysp T3SS and attenuate the virulence of Yersinia enterocolitica. Introduction:Yersinia enterocolitica is an opportunistic pathogen which causes enteric diseases like gastrointestinal and mesenteric adenitis in immune-compromised individuals. The gastrointestinal phase of Y. enterocolitica infection is mediated by Yersinia secretion apparatus- Yersinia secretion protein (Ysa-Ysp) Type III Secretion System (T3SS). Certain serotypes of Y. enterocolitica Biovar 1B exhibit enhanced virulence due to presence of Ysa-Ysp T3SS. Translocator proteins form the translocation apparatus (translocon) at the tip of the needle complex of the T3SS. YspC is a minor hydrophobic translocator protein which is sequestered in the bacterial cytoplasm by its cognate chaperone SycB. SycB plays the dual role of a class II chaperone and a regulator of Ysa-Ysp T3SS. Experiments and key results findings: Homology model of SycB depicts a structure with a concave core formed by tetratricopeptide repeats (TPRs) and a flexible N- terminal helix. Deletion mutants of SycB showed that the N-terminal helix of SycB is responsible for its dimerization, which is further corroborated by molecular docking analysis. The dimeric state of SycB dissociates during the interaction with YspC due to steric hindrances. And it forms a 1:1 heterodimeric YspC-SycB complex as confirmed by size-exclusion chromatography, chemical cross-linking and molecular docking studies. FRET analysis indicated that the tyrosine residues present in first two TPRs of SycB is responsible for its interaction with YspC. Deletion mutants of SycB possessing the first two TPR regions interacted with YspC, as also depicted by the YspC-SycB interaction model. YspC is a unique translocator protein with high stability, rigid tertiary structure and monomeric form unlike any other translocator proteins. It shows structural alteration in the complex formed with SycB as shown by spectroscopic data and proteolytic digestion. YspC has a Y-shaped three dimensional structure and SycB completely localizes within the fork formed by the two arms of Y-shaped YspC. This structural analysis and mechanism of interaction between YspC and SycB would be potentially beneficial for designing drugs to deregulate the Ysa-Ysp T3SS and attenuate the virulence of Yersinia enterocolitica.