Serum Expression Level of MicroRNA-122 and its Significance in different Stages of Hepatitis B Virus Infection.

This study aims to determine the serum expression level of miRNA-122 and its significance in the different stages of Hepatitis B virus infection. The study subjects were recruited and grouped for Hepatitis B associated with Chronic Hepatitis B infection, hepatic sclerosis, hepatocellular carcinoma, and healthy controls were also considered. Venous blood was collected from the participants including the controls and routine blood tests and quantification of miRNA-122 were done and analyzed in each case of hepatitis B infection and compared with that of healthy controls. The miRNA-122 was determined, which came to be highest in patients with Chronic Hepatitis B while patients with hepatic sclerosis and patients with hepatocellular carcinoma showed a subsequent number of copies. The number of copies of miRNA-122 in the CHB, hepatic sclerosis, and HCC group was significantly higher than in the healthy control. The quantification of miRNA-122 and subsequently plotting the ROC curve has shown that miRNA-122 can be considered as a biomarker of hepatitis B for screening and diagnosis purposes.

Gut milieu shapes the bacterial communities of invasive silver carp.

Silver carp is an invasive fish present in the Gobindsagar reservoir, India and has a profound impact on aquaculture. Understanding taxonomic diversity and functional attributes of gut microbiota will provide insights into the important role of bacteria in metabolism of silver carp that facilitated invasion of this exotic species. Microbial composition in foregut, midgut, hindgut and water samples was analysed using 16S rRNA gene amplicon sequencing. The bacterial communities of water samples were distinct from gut microbiota, and unique microbial assemblages were present in different regions of gut depicting profound impact of gut environment on microflora. Proteobacteria was the most abundant phyla across all samples. Ecological network analysis showed dominance of competitive interactions within posteriors region of the gut, promoting niche specialization. Predictive functional profiling revealed the microbiota specialized in digestive functions in different regions of the gut, which also reflects the dietary profile of silver carp.

Genome based reclassification of Deinococcus swuensis as a heterotypic synonym of Deinococcus radiopugnans.

Deinococcus species are widely studied due to their utility in bioremediation of sites contaminated with radioactive elements. In the present study, we re-evaluated the taxonomic placement of two species of the genus Deinococcus namely D. swuensis DY59T and D. radiopugnans ATCC 19172T based on whole genome analyses. The 16S rRNA gene analysis revealed a 99.58% sequence similarity between this species pair that is above the recommended threshold value for species delineation. These two species also clustered together in both the 16S rRNA gene and core genome based phylogenies depicting their close relatedness. Furthermore, more than 98% of genes were shared between D. swuensis DY59T and D. radiopugnans ATCC 19172T. Interestingly, D. swuensis DY59T and D. radiopugnans ATCC 19172T shared high genome similarity in different genomic indices. They displayed an average nucleotide identity value of 97.63%, an average amino acid identity value of 97% and a digital DNA-DNA hybridization value equal to 79.50%, all of which are well above the cut-off for species delineation. Altogether, based on these evidences, D. swuensis DY59T and D. radiopugnans ATCC 19172T constitute a single species. Hence, as per the priority of publication, we propose that Deinococcus swuensis Lee et al. 2015 should be reclassified as a later heterotypic synonym of Deinococcus radiopugnans.

Perspectives on emerging pressures and their integrated impact on large river systems: An insight from the Yellow River basin.

The Yellow River, with a developmental and historical significance to China, is now facing several emerging pressures, which are degrading the river status and creating challenges for high-quality development in the basin. Numerous studies on such emerging pressures, present scattered outcomes, and trigger uncertainties and deficient assumptions on the river's problems. This review integrated such scattered information and investigated the emerging pressures, their drivers and integrated impacts at the basin level. The study intended to prioritize those pressures needing expeditious consideration, and carried a discussion on the alternative pathways to the solution. To determine the critical emerging pressures, a literature review was conducted and experts' opinion was sought. The outcome further led to a comprehensive review, data collection, and analysis of three groups of emerging pressures. The review recognized 'Water Stress' in the lower reach, primarily caused by an abated flow, as the most distressing emerging pressure inflicting social, ecological, and economic consequences. Such decline in flow was mostly induced by a recent increase in 'Anthropogenic activities', such as intensive water withdrawal for irrigation (≥27 BCM), and construction of check dams in the Loess Plateau region (trapping~5 BCM water). The increasing 'Pollution' in the river, besides threatening public health and ecology, also contributed to the water stress by rendering certain stretches of the river biologically dead and unsuitable for any use. The 'Climate Change', with its key negative effect on precipitation in the middle sub-basin, overall contributed small (8-11 %) to the observed reduction in river flow. With increasing challenges for the adopted engineering solutions tackling the water stress, the study suggested the use of a demand management approach, employing adaptive policy measures, as an alternative or supplementary solution to the current approach. In addition, the study highlights that regular reviewing and reforming the key decisions based on evidence and updated information, and taking a participatory approach, may offer a sustainable pathway to the environment as well as socio-economic goals.

The genus Sphingopyxis: Systematics, ecology, and bioremediation potential - A review.

The genus Sphingopyxis was first reported in the year 2001. Phylogenetically, Sphingopyxis is well delineated from other genera Sphingobium, Sphingomonas and Novosphingobium of sphingomonads group, family Sphingomonadaceae of Proteobacteria. To date (at the time of writing), the genus Sphingopyxis comprises of twenty validly published species available in List of Prokaryotic Names with Standing in Nomenclature. Sphingopyxis spp. have been isolated from diverse niches including, agricultural soil, marine and fresh water, caves, activated sludge, thermal spring, oil and pesticide contaminated soil, and heavy metal contaminated sites. Sphingopyxis species have drawn considerable attention not only for their ability to survive under extreme environments, but also for their potential to degrade number of xenobiotics and other environmental contaminants that impose serious threat to human health. At present, genome sequence of both cultivable and non-cultivable strains (metagenome assembled genome) are available in the public databases (NCBI) and genome wide studies confirms the presence of mobile genetic elements and plethora of degradation genes and pathways making them a potential candidate for bioremediation. Beside genome wide predictions there are number of experimental evidences confirm the degradation potential of bacteria belonging to genus Sphingopyxis and also the production of different secondary metabolites that help them interact and survive in their ecological niches. This review provides detailed information on ecology, general characteristic and the significant implications of Sphingopyxis species in environmental management along with the bio-synthetic potential.

Gut microbiome of endangered Tor putitora (Ham.) as a reservoir of antibiotic resistance genes and pathogens associated with fish health.

BACKGROUND: Tor putitora, the largest freshwater fish of the Indian subcontinent, is an endangered species. Several factors have been attributed towards its continuous population decrease, but very little is known about the gut microbiome of this fish. Also, the fish gut microbiome serves as a reservoir of virulence factors and antibiotic resistance determinants. Therefore, the shotgun metagenomic approach was employed to investigate the taxonomic composition and functional potential of microbial communities present in the gut of Tor putitora, as well as the detection of virulence and antibiotic resistance genes in the microbiome. RESULTS: The analysis of bacterial diversity showed that Proteobacteria was predominant phylum, followed by Chloroflexi, Bacteroidetes, and Actinobacteria. Within Proteobacteria, Aeromonas and Caulobacter were chiefly present; also, Klebsiella, Escherichia, and plant symbionts were noticeably detected. Functional characterization of gut microbes endowed the virulence determinants, while surveillance of antibiotic resistance genes showed the dominance of ß-lactamase variants. The antibiotic-resistant Klebsiella pneumoniae and Escherichia coli pathovars were also detected. Microbial genome reconstruction and comparative genomics confirmed the presence of Aeromonads, the predominant fish pathogens. CONCLUSIONS: Gut microbiome of endangered Tor putitora consisted of both commensals and opportunistic pathogens, implying that factors adversely affecting the non-pathogenic population would allow colonization and proliferation of pathogens causing diseased state in asymptomatic Tor putitora. The presence of virulence factors and antibiotic resistance genes suggested the potential risk of dissemination to other bacteria due to horizontal gene transfer, thereby posing a threat to fish and human health. The preservation of healthy gut microflora and limited use of antibiotics are some of the prerequisites for the conservation of this imperilled species.

Dicarboxylate Transporters of Azospirillum brasilense Sp7 Play an Important Role in the Colonization of Finger Millet (Eleusine coracana) Roots.

Azospirillum brasilense is a plant growth-promoting bacterium that colonizes the roots of a large number of plants, including C3 and C4 grasses. Malate has been used as a preferred source of carbon for the enrichment and isolation Azospirillum spp., but the genes involved in their transport and utilization are not yet characterized. In this study, we investigated the role of the two types of dicarboxylate transporters (DctP and DctA) of A. brasilense in their ability to colonize and promote growth of the roots of a C4 grass. We found that DctP protein was distinctly upregulated in A. brasilense grown with malate as sole carbon source. Inactivation of dctP in A. brasilense led to a drastic reduction in its ability to grow on dicarboxylates and form cell aggregates. Inactivation of dctA, however, showed a marginal reduction in growth and flocculation. The growth and nitrogen fixation of a dctP and dctA double mutant of A. brasilense were severely compromised. We have shown here that DctPQM and DctA transporters play a major and a minor role in the transport of C4-dicarboxylates in A. brasilense, respectively. Studies on inoculation of the seedlings of a C4 grass, Eleusine corcana, with A. brasilense and its dicarboxylate transport mutants revealed that dicarboxylate transporters are required by A. brasilense for an efficient colonization of plant roots and their growth.

Microbial Inoculants Assisted Growth of Chrysopogon zizanioides Promotes Phytoremediation of Salt Affected Soil.

Restoration of salt-affected soil through cultivation Chrysopogon zizanioides is a promising approach. The two way benefit of such an approach is that reclamation of salt-affected soil concomitant to improve plant growth and increased yield of essential oil produced in the plants roots. Earlier studies showed physiological changes and reduced growth of C. zizanioides under salinity. In the present study, plant growth promoting microorganisms viz. Pseudomonas monteilii, Bacillus megaterium, Azotobacter chroococcum and Rhizophagus intraradices were used as bio-inoculants for cultivation of C. zizanioides under salt-affected soil. Bio-inoculants in combination with vermicompost significantly increased the growth and productivity of C. zizanioides under salt-affected soil, and simultaneously improved soil health. When compared to control, the soil physico-chemical and biological properties of bio-inoculants treated plants was significantly improved. The reclamation of salt-affected soil was evident by the significant decrease in the level of soil pH (11.0%), electrical conductivity (23.5%), sodium adsorption ratio (15.3%), and exchangeable sodium percent (12.4%) of bio-inoculants treated plants. The improvement of soil cation exchange capacity indicated the decrease in soil salinity. Whereas increase in the microbial count (four-fold), AMF spores (447 spores), dehydrogenase (six-fold), acid (two-fold) and alkaline phosphatase (five-fold) activities in rhizosphere soil of bio-inoculant treated plants indicated the improved biological properties. A positive correlation of plant biomass production to soil organic carbon, total Kjeldahl nitrogen, available phosphorus and cation exchange capacity depicted improved nutrients content in rhizosphere soil of bio-inoculant treated plants. The findings of this study suggest that P. monteilii and R. intraradices with vermicompost can be effectively used as bio-inoculants for encouragement of phytoremediation in salt-affected soil.

Genome Sequencing Revealed the Biotechnological Potential of an Obligate Thermophile Geobacillus thermoleovorans Strain RL Isolated from Hot Water Spring.

In the present study, we report the draft genome sequence of an obligate thermophile Geobacillus thermoleovorans strain RL isolated from Manikaran hot water spring located atop the Himalayan ranges, India. Strain RL grew optimally at 70 °C but not below 45 °C. The draft genome (3.39 Mb) obtained by Illumina sequencing contains 138 contigs with an average G + C content of 52.30%. RAST annotation showed that amino acid metabolism pathways were most dominant followed by carbohydrate metabolism. Genome-wide analysis using NCBI's Prokaryotic Genome Annotation Pipeline revealed that strain RL encodes for a cocktail of industrially important hydrolytic enzymes glycoside hydrolase, α-and ß-glucosidase, xylanase, amylase, neopullulanase, pullulanase and lipases required for white biotechnology. In addition, the presence of genes encoding green biocatalyst multicopper polyphenol oxidase (laccase) and an anticancer enzyme l-glutaminase reflects the significance of strain RL in gray and red biotechnology, respectively. Strain RL is a thermophilic multi-enzyme encoding bacterium which could be the source for the recombinant production of biotechnologically significant enzymes. In, addition whole cells of strain RL may be used in bioremediation studies.

SRC family kinases in hamster spermatozoa: evidence for the presence of LCK.

Sperm capacitation is a prerequisite for successful fertilization. Increase in tyrosine phosphorylation is considered the hallmark of capacitation and attempts to understand its regulation are ongoing. In this regard, we attempted to study the role of SRC family kinases (SFKs) in the hamster sperm functions. Interestingly, we found the presence of the lymphocyte-specific protein tyrosine kinase, LCK, in mammalian spermatozoa and further characterized it in terms of its localization and function. LCK was found in spermatozoa of several species, and its transcript was identified in the hamster testis. Autophosphorylation of LCK at the Y394 residue increased as capacitation progressed, indicating an upregulation of LCK activity during capacitation. Inhibition of LCK (and perhaps the other SFKs) with the use of a specific inhibitor showed a significant decrease in protein tyrosine phosphorylation of several proteins, implying LCK/SFKs as key tyrosine kinase(s) regulating tyrosine phosphorylation during hamster sperm capacitation. Dihydrolipoamide dehydrogenase was identified as a substrate for LCK/SFK. LCK/SFKs inhibition significantly reduced the percentage fertilization (in vitro) but had no effect on sperm motility, hyperactivation and acrosome reaction. In summary, this is the first report on the presence of LCK, an SFK of hematopoietic lineage in spermatozoa besides being the first study on the role of SFKs in the spermatozoa of Syrian hamsters.

Enhancement of the photocatalytic activity of rGO/NiO/Ag nanocomposite for degradation of methylene blue dye.

The current study focuses on boosting the photocatalytic ability of reduced graphene oxide (rGO) by decorating the rGO nano-sheets with nickel oxide (NiOx) and silver (Ag) nanomaterials. The developed ternary nanomaterials were investigated using FTIR, XRD, FESEM, TEM, Raman, and UV-vis to evaluate the photo-degradation process. The rGO/NiOx/Ag ternary system showed promising photocatalytic dye degradation under simulated sunlight irradiance. The addition of NiOx and Ag nanomaterials widened the catalytic activity spectrum from the visible region to the UV-region. Besides, these materials hindered the electron-hole recombination, boosting the catalytic activity. The reusability results also clearly showed that the synthesized ternary nanomaterials have good reproducibility and stability for photocatalytic degradation of industrial wastewater.

Cultivation-independent analysis of archaeal and bacterial communities of the formation water in an Indian coal bed to enhance biotransformation of coal into methane.

Biogenic origin of the significant proportion of coal bed methane has indicated the role of microbial communities in methanogenesis. By using cultivation-independent approach, we have analysed the archaeal and bacterial community present in the formation water of an Indian coal bed at 600-700 m depth to understand their role in methanogenesis. Presence of methanogens in the formation water was inferred by epifluorescence microscopy and PCR amplification of mcrA gene. Archaeal 16S rRNA gene clone library from the formation water metagenome was dominated by methanogens showing similarity to Methanobacterium, Methanothermobacter and Methanolinea whereas the clones of bacterial 16S rRNA gene library were closely related to Azonexus, Azospira, Dechloromonas and Thauera. Thus, microbial community of the formation water consisted of predominantly hydrogenotrophic methanogens and the proteobacteria capable of nitrogen fixation, nitrate reduction and polyaromatic compound degradation. Methanogenic potential of the microbial community present in the formation water was elucidated by the production of methane in the enrichment culture, which contained 16S rRNA gene sequences showing close relatedness to the genus Methanobacterium. Microcosm using formation water as medium as well as a source of inoculum and coal as carbon source produced significant amount of methane which increased considerably by the addition of nitrite. The dominance of Diaphorobacter sp. in nitrite amended microcosm indicated their important role in supporting methanogenesis in the coal bed. This is the first study indicating existence of methanogenic and bacterial community in an Indian coal bed that is capable of in situ biotransformation of coal into methane.

Catalytic C-H Bond Activation and Knoevenagel Condensation Using Pyridine-2,3-Dicarboxylate-Based Metal-Organic Frameworks.

Three 1D coordination polymers (CPs) [M(pdca)(H2O)2] n (M = Zn, Cd, and Co; 1-3), and a 3D coordination framework {[(CH3)2NH2][CuK(2,3-pdca)(pa)(NO3)2]} n (4) (2,3-pdca = pyridine-2,3-dicarboxylate and pa = picolinic acid), have been synthesized adopting a solvothermal reaction strategy. The CPs have been thoroughly characterized using various spectral techniques, that is, elemental analyses, FT-IR, TGA, DSC, UV/vis, and luminescence. Structural information on 1-4 was obtained by PXRD and X-ray single-crystal analyses, whereas morphological insights were attained through FESEM, AFM, EDX, HRTEM, and BET surface area analyses. Roughness parameters were calculated from AFM analysis, whereas dimensions of small domains and interplanar spacing were defined with the aid of HRTEM. CPs 1-3 are 1D isostructural networks, whereas 4 is a 3D framework. Moreover, 1-4 display moderate luminescence at rt. In addition, 1-4 have been applied as economic and efficient porous catalysts for the Knoevenagel condensation reaction and C-H bond activation under mild conditions with good yields (95-98 and 97-99%), respectively. Notably, 1-3 can be reused up to seven cycles, whereas 4 can be reused up to five catalytic cycles with retained catalytic efficiency. Relative catalytic efficacy toward the Knoevenagel condensation reaction follows in the order 2 > 1 > 3 > 4, whereas 2 > 4 > 1 > 3 for C-H activation. The present result demonstrates synthetic, structural, optical, morphological, and catalytic aspects of 1-4.

Comparative proteomics unravelled the hexachlorocyclohexane (HCH) isomers specific responses in an archetypical HCH degrading bacterium Sphingobium indicum B90A.

Hexachlorocyclohexane (HCH) is a persistent organochlorine pesticide that poses threat to different life forms. Sphingobium indicum B90A that belong to sphingomonad is well-known for its ability to degrade HCH isomers (α-, ß-, γ-, δ-), but effects of HCH isomers and adaptive mechanisms of strain B90A under HCH load remain obscure. To investigate the responses of strain B90A to HCH isomers, we followed the proteomics approach as this technique is considered as the powerful tool to study the microbial response to environmental stress. Strain B90A culture was exposed to α-, ß-, γ-, δ-HCH (5 mgL-1) and control (without HCH) taken for comparison and changes in whole cell proteome were analyzed. In ß- and δ-HCH-treated cultures growth decreased significantly when compared to control, α-, and γ-HCH-treated cultures. HCH residue analysis corroborated previous observations depicting the complete depletion of α- and γ-HCH, while only 66% ß-HCH and 34% δ-HCH were depleted from culture broth. Comparative proteome analyses showed that ß- and δ-HCH induced utmost systemic changes in strain B90A proteome, wherein stress-alleviating proteins such as histidine kinases, molecular chaperons, DNA binding proteins, ABC transporters, TonB proteins, antioxidant enzymes, and transcriptional regulators were significantly affected. Besides study confirmed constitutive expression of linA, linB, and linC genes that are crucial for the initiation of HCH isomers degradation, while increased abundance of LinM and LinN in presence of ß- and δ-HCH suggested the important role of ABC transporter in depletion of these isomers. These results will help to understand the HCH-induced damages and adaptive strategies of strain B90A under HCH load which remained unravelled to date.

Rapid and solitary production of mono-rhamnolipid biosurfactant and biofilm inhibiting pyocyanin by a taxonomic outlier Pseudomonas aeruginosa strain CR1.

Biosurfactant - Rhamnolipids (RLs) and antibacterial toxin - pyocyanin (PYO) produced by Pseudomonas aeruginosa strains have great potential for biotechnological applications. Generally, RLs are produced as a mixture of di-rhamnolipids (di-RLs) and mono-rhamnolipids (mono-RLs). Mono-RLs possess superior emulsification and antimicrobial properties and are costlier than di-RLs. In this study, a taxonomic outlier P. aeruginosa strain CR1 isolated from rhizosphere soil was explored for mono-RLs and PYO production. Phylogenetically strain CR1 resembles avirulent outlier P. aeruginosa strain ATCC9027, lacks archetypical virulence genes and harbors unique pathways for the synthesis of solely mono-RLs and PYO. Strain CR1 produced RL biosurfactant which efficiently emulsified hydrocarbons, showed hemolysis and inhibited Bacillus subtilis. At 37 °C, strain CR1 exclusively produced 21.77 g L-1 and 19.22 g L-1 rhamnolipid in glycerol amended Luria Bertani (LB) medium and basal medium amended with rice bran oil, respectively after 54 h growth. Besides RL production was unaffected under varying nitrogen sources. Structural characterization using FTIR, TLC, and LC-MS confirmed that strain CR1 exclusively produced mono-RLs, majorly dominated by Rha-C10-C10, Rha-C10-C8, and CH3-Rha-C12:2-C10:1. The compound was stable over a wide pH range (4-12), salinity (25%) and 100 °C indicating its applicability under harsh environmental conditions. In addition, strain CR1 produced 4.5 µg mL-1 PYO, which could efficiently inhibit biofilm formation by Bacillus species. The environmental outlier strain CR1 can be used for the industrial production of biotechnologically important mono-RLs and PYO.

Draft Genome Sequence of Deinococcus sp. Strain S9, Isolated from Microbial Mat Deposits of Hot Springs Located atop the Himalayan Ranges at Manikaran, India.

Here, we present the draft genome sequence of Deinococcus sp. strain S9, a red-pigmented and moderately thermophilic bacterium isolated from microbial mat deposits around the hot springs at Manikaran, Himachal Pradesh, India. The draft genome (3.34 Mb) contains 101 contigs with an average GC content of 66.4%.

Whole-Genome Sequences of Four Indian Isolates of Azospirillum brasilense.

Azospirillum brasilense is used worldwide as a plant growth-promoting inoculant for agricultural crops. To understand how the genomes of Indian strains of A. brasilense compare with their South American counterparts, we determined the whole-genome sequences of four strains of A. brasilense isolated from the rhizosphere of grasses from India.

Spin reorientation and disordered rare earth magnetism in Ho2FeCoO6.

We report the experimental observation of spin reorientation in the double perovskite Ho2FeCoO6. The magnetic phase transitions in this compound are characterized and studied through magnetization and specific heat, and the magnetic structures are elucidated through neutron powder diffraction. Two magnetic phase transitions are observed in this compound-one at [Formula: see text] K, from paramagnetic to antiferromagnetic, and the other at [Formula: see text] K, from a phase with mixed magnetic structures to a single phase through a spin reorientation process. The magnetic structure in the temperature range 200-45 K is a mixed phase of the irreducible representations [Formula: see text] and [Formula: see text], both of which are antiferromagnetic. The phase with mixed magnetic structures that exists in Ho2FeCoO6 gives rise to a large thermal hysteresis in magnetization that extends from 200 K down to the spin reorientation temperature. At T N2, the magnetic structure transforms to [Formula: see text]. Though long-range magnetic order is established in the transition metal lattice, it is seen that only short-range magnetic order prevails in the Ho3+ lattice. Our results should motivate further detailed studies on single crystals in order to explore the spin reorientation process, spin switching and the possibility of anisotropic magnetic interactions giving rise to electric polarization in Ho2FeCoO6.

Identification and characterization of a novel phosphodiesterase from the metagenome of an Indian coalbed.

Phosphoesterases are involved in the degradation of organophosphorus compounds. Although phosphomonoesterases and phosphotriesterases have been studied in detail, studies on phosphodiesterases are rather limited. In our search to find novel phosphodiesterases using metagenomic approach, we cloned a gene encoding a putative phosphodiesterase (PdeM) from the metagenome of the formation water collected from an Indian coal bed. Bioinformatic analysis showed that PdeM sequence possessed the characteristic signature motifs of the class III phosphodiesterases and phylogenetic study of PdeM enabled us to identify three distinct subclasses (A, B, and C) within class III phosphodiesterases, PdeM clustering in new subclass IIIB. Bioinformatic, biochemical and biophysical characterization of PdeM further revealed some of the characteristic features of the phosphodiesterases belonging to newly described subclass IIIB. PdeM is a monomer of 29.3 kDa, which exhibits optimum activity at 25°C and pH 8.5, but low affinity for bis(pNPP) as well as pNPPP. The recombinant PdeM possessed phosphodiesterase, phosphonate-ester hydrolase and nuclease activity. It lacked phosphomonoesterase, phosphotriesterase, and RNAse activities. Overexpression of PdeM in E.coli neither affected catabolite respression nor did the recombinant protein hydrolyzed cAMP in vitro, indicating its inability to hydrolyze cAMP. Although Mn2+ was required for the activity of PdeM, but addition of metals (Mn2+ or Fe3+) did not induce oligomerization. Further increase in concentration of Mn2+ upto 3 mM, increased α-helical content as well as the phosphodiesterase activity. Structural comparison of PdeM with its homologs showed that it lacked critical residues required for dimerization, cAMP hydrolysis, and for the high affinity binding of bis(pNPP). PdeM, thus, is a novel representative of new subclass of class III phosphodiesterases.

Inhibiting sperm pyruvate dehydrogenase complex and its E3 subunit, dihydrolipoamide dehydrogenase affects fertilization in Syrian hamsters.

BACKGROUND/AIMS: The importance of sperm capacitation for mammalian fertilization has been confirmed in the present study via sperm metabolism. Involvement of the metabolic enzymes pyruvate dehydrogenase complex (PDHc) and its E3 subunit, dihydrolipoamide dehydrogenase (DLD) in hamster in vitro fertilization (IVF) via in vitro sperm capacitation is being proposed through regulation of sperm intracellular lactate, pH and calcium. METHODOLOGY AND PRINCIPAL FINDINGS: Capacitated hamster spermatozoa were allowed to fertilize hamster oocytes in vitro which were then assessed for fertilization, microscopically. PDHc/DLD was inhibited by the use of the specific DLD-inhibitor, MICA (5-methoxyindole-2-carboxylic acid). Oocytes fertilized with MICA-treated (MT) [and thus PDHc/DLD-inhibited] spermatozoa showed defective fertilization where 2nd polar body release and pronuclei formation were not observed. Defective fertilization was attributable to capacitation failure owing to high lactate and low intracellular pH and calcium in MT-spermatozoa during capacitation. Moreover, this defect could be overcome by alkalinizing spermatozoa, before fertilization. Increasing intracellular calcium in spermatozoa pre-IVF and in defectively-fertilized oocytes, post-fertilization rescued the arrest seen, suggesting the role of intracellular calcium from either of the gametes in fertilization. Parallel experiments carried out with control spermatozoa capacitated in medium with low extracellular pH or high lactate substantiated the necessity of optimal sperm intracellular lactate levels, intracellular pH and calcium during sperm capacitation, for proper fertilization. CONCLUSIONS: This study confirms the importance of pyruvate/lactate metabolism in capacitating spermatozoa for successful fertilization, besides revealing for the first time the importance of sperm PDHc/ DLD in fertilization, via the modulation of sperm intracellular lactate, pH and calcium during capacitation. In addition, the observations made in the IVF studies in hamsters suggest that capacitation failures could be a plausible cause of unsuccessful fertilization encountered during human assisted reproductive technologies, like IVF and ICSI. Our studies indicate a role of sperm capacitation in the post-penetration events during fertilization.