Tinospora cordifolia Extract Enhances Dextromethorphan Bioavailability: Implications for Alzheimer's Disease.

Tinospora cordifolia (Willd.) Miers (Menispermaceae) is a traditional rejuvenator and a conventional medicine used to manage oxidative stress-related diseases, including those associated with the central nervous system. Decreased dextromethorphan (DEM) metabolism is necessary for high bioavailability and application against Alzheimer's disease (AD). Since T. cordifolia stem extract (TCE) can potentially inhibit several metabolic enzymes, it can also enhance dextromethorphan bioavailability. This study investigates the potential of TCE to improve DEM's bioavailability and efficacy for the management of AD. In silico analysis was carried out to find the inhibition potential of phytocomponents of T. cordifolia for CYP2D6 and CYP3A4. The LC-MS method was revalidated for the analysis of DEM and metabolite dextrorphan (DEX) in the presence of quinidine (QN). The ratio of DEM to DEX was estimated with varying doses of TCE following pharmacokinetic analysis. Network pharmacology analysis was carried out to understand the complementary potential of phytocomponents. This was further validated in the scopolamine-induced dementia model through behavioral and histopathological analyses. TCE (100 mg/kg) for 14 days increased the DEM to DEX ratio by 2.8-fold compared to QN treatment. While T max was comparable to that of QN treatment at this dose (100 mg/kg) of TCE, it increased significantly at the higher dose (400 mg/kg) of TCE pretreatment. All other pharmacokinetic parameters were also enhanced at this dose with a 4.7-fold increase in DEM/DEX compared with QN. Network pharmacology analysis indicated the ability of TCE to target multiple factors associated with AD. Furthermore, it improved spatial memory and reduced hyperactivity in rodents better than the combination of QN and DEM.

Tamarind seed polysaccharide-metformin insert: Higher ocular retention, slow-release, and efficacy against corneal burn.

Metformin (MET) can be an alternative therapeutic strategy for managing ocular burn primarily because of its pleiotropic mechanism. Longer retention on the ocular surface and sustained release are necessary to ensure the efficacy of MET for ocular application. Although the high aqueous solubility of MET is good for formulation and biocompatibility, it makes MET prone to high nasolacrimal drainage. This limits ocular residence and may be a challenge in its application. To address this, polymers approved for ophthalmic application with natural origin were analyzed through in silico methods to determine their ability to bind to mucin and interact with MET. An ocular insert of MET (3 mg/6 mm) was developed using a scalable solvent casting method without using preservatives. The relative composition of the insert was 58 ± 2.06 %w/w MET with approximately 14 %w/w tamarind seed polysaccharide (TSP), and 28 %w/w propylene glycol (PG). Its stability was demonstrated as per the ICH Q1A (R2) guidelines. Compatibility, ocular retention, drug release, and other functional parameters were evaluated. In rabbits, efficacy was demonstrated in the 'corneal alkali burn preclinical model'. TSP showed potential for mucoadhesion and interaction with MET. With adequate stability and sterility, the insert contributed to adequate retention of MET (10-12 h) in vivo and slow release (30 h) in vitro. This resulted in significant efficacy in vivo.

Molecular docking, molecular dynamics simulation, and MM/PBSA analysis of ginger phytocompounds as a potential inhibitor of AcrB for treating multidrug-resistant Klebsiella pneumoniae infections.

The emergence of Multidrug resistance (MDR) in human pathogens has defected the existing antibiotics and compelled us to understand more about the basic science behind alternate anti-infective drug discovery. Soon, proteome analysis identified AcrB efflux pump protein as a promising drug target using plant-driven phytocompounds used in traditional medicine systems with lesser side effects. Thus, the present study aims to explore the novel, less toxic, and natural inhibitors of Klebsiella pneumoniae AcrB pump protein from 69 Zingiber officinale phyto-molecules available in the SpiceRx database through computational-biology approaches. AcrB protein's homology-modelling was carried out to get a 3D structure. The multistep-docking (HTVS, SP, and XP) were employed to eliminate less-suitable compounds in each step based on the docking score. The chosen hit-compounds underwent induced-fit docking (IFD). Based on the XP GScore, the top three compounds, epicatechin (-10.78), 6-gingerol (-9.71), and quercetin (-9.09) kcal/mol, were selected for further calculation of binding free energy (MM/GBSA). Furthermore, the short-listed compounds were assessed for their drug-like properties based on in silico ADMET properties and Pa, Pi values. In addition, the molecular dynamics simulation (MDS) studies for 250 ns elucidated the binding mechanism of epicatechin, 6-gingerol, and quercetin to AcrB. From the dynamic binding free energy calculations using MM/PBSA, 6-gingerol exhibited a strong binding affinity towards AcrB. Further, the 6-gingerol complex's energy fluctuation was observed from the free energy landscape. In conclusion, 6-gingerol has a promising inhibiting potential against the AcrB efflux pump and thus necessitates further validation through in vitro and in vivo experiments.Communicated by Ramaswamy H. Sarma.

Development of pharmacophore models for AcrB protein and the identification of potential adjuvant candidates for overcoming efflux-mediated colistin resistance.

Growing multi-drug resistance (MDR) among ESKAPE pathogens is a huge challenge. Increased resistance to last-resort antibiotics, like colistin, has further aggravated this. Efflux is identified as a major route of colistin resistance. So, finding an FDA-approved efflux inhibitor for potential application as an adjuvant to colistin was the primary objective of this study. E. coli-AcrB pump inhibitors and substrates were used to develop and validate the pharmacophoric model. Drugs confirming this pharmacophore were subjected to molecular docking to identify hits for the AcrB binding pocket. The efflux inhibition potential of the top hit was validated through the in vitro evaluation of the minimum inhibitory concentration (MIC) in combination with colistin. The checkerboard assay was done to demonstrate synergism, which was further corroborated by the Time-kill assay. Ten common pharmacophore hypotheses were successfully generated using substrate/inhibitors. Following enrichment analysis, AHHNR.100 was identified as the top-ranked hypothesis, and 207 unique compounds were found to conform to this hypothesis. The multi-step docking of these compounds against the AcrB protein revealed argatroban as the top non-antibiotic hit. This significantly inhibited the efflux activity of colistin-resistant clinical isolates K. pneumoniae (n = 1) and M. morganii (n = 2). Further, their combination with colistin enhanced the susceptibility of these isolates, and the effect was found to be synergistic. Accordingly, the time-kill assay of this combination showed 8-log and 2-log reductions against K. pneumoniae and M. morganii, respectively. In conclusion, this study found argatroban as a bacterial efflux inhibitor that can be potentially used to overcome efflux-mediated resistance.

Investigating microbiome and transcriptome data to uncover the key microbial community involved in lignocellulose degradation within the Deulajhari hot spring consortium.

Geothermally heated spring water contaminated with decomposed leaf biomass creates unique hot spring ecosystems that support the recycling of diverse nutrients and harbor microbial consortia capable of degrading lignocellulose. We present microbiome and transcriptome data from the bacterial consortium of Deulajhari hot springs, characterized by a temperature of approximately 58 °C and surrounded by a dense population of pandanus plants in Angul, Odisha, India. Metagenomics and metatranscriptomics datasets were generated by extracting total DNA and RNA from the consortium sample of hotspring sediment, followed by shotgun sequencing using the Illumina HiSeq 2500 platform. The metagenomics dataset produced approximately 38,694 contigs, while the metatranscriptomics dataset yielded 9226 contigs, resulting in a total nucleotide size of 89,857,616 and 15,541,403 bps, respectively. Analysis using MEGAN6 against the NCBI "taxonomy" database revealed the presence of 18 and 12 phyla, including candidate phyla, in respective datasets. Proteobacteria exhibited the highest relative abundance in the metagenomics dataset, while Firmicutes was highly abundant in the metatranscriptomics dataset. At the genus level, a total of 92 and 25 genera were predicted in both datasets, with lignocellulose degrading Meiothermus being highly abundant in both metagenomics and metatranscriptomics datasets. We also observed that the unknown bacteria and unidentified sequences were found in significant proportion in the metatranscriptomics dataset. We assembled and functionally annotated approximately 23,960 contigs using the Prokka pipeline. Among the SEED category, the most expressed and annotated microbial genes fall under the unknown category as well as Biotin synthesis and their utilization. Furthermore, some of these genes were implicated in the degradation of aromatic amino acids, D-mannitol, and D-mannose. These findings contribute to our understanding of how the composition and abundance of bacterial communities facilitate lignocellulose degradation in extreme environments and biofuel generation.

Sequencing and Characterization of M. morganii Strain UM869: A Comprehensive Comparative Genomic Analysis of Virulence, Antibiotic Resistance, and Functional Pathways.

Morganella morganii is a Gram-negative opportunistic Enterobacteriaceae pathogen inherently resistant to colistin. This species causes various clinical and community-acquired infections. This study investigated the virulence factors, resistance mechanisms, functional pathways, and comparative genomic analysis of M. morganii strain UM869 with 79 publicly available genomes. The multidrug resistance strain UM869 harbored 65 genes associated with 30 virulence factors, including efflux pump, hemolysin, urease, adherence, toxin, and endotoxin. Additionally, this strain contained 11 genes related to target alteration, antibiotic inactivation, and efflux resistance mechanisms. Further, the comparative genomic study revealed a high genetic relatedness (98.37%) among the genomes, possibly due to the dissemination of genes between adjoining countries. The core proteome of 79 genomes contains the 2692 core, including 2447 single-copy orthologues. Among them, six were associated with resistance to major antibiotic classes manifested through antibiotic target alteration (PBP3, gyrB) and antibiotic efflux (kpnH, rsmA, qacG; rsmA; CRP). Similarly, 47 core orthologues were annotated to 27 virulence factors. Moreover, mostly core orthologues were mapped to transporters (n = 576), two-component systems (n = 148), transcription factors (n = 117), ribosomes (n = 114), and quorum sensing (n = 77). The presence of diversity in serotypes (type 2, 3, 6, 8, and 11) and variation in gene content adds to the pathogenicity, making them more difficult to treat. This study highlights the genetic similarity among the genomes of M. morganii and their restricted emergence, mostly in Asian countries, in addition to their growing pathogenicity and resistance. However, steps must be taken to undertake large-scale molecular surveillance and to direct suitable therapeutic interventions.

Unravelling the Evolutionary Dynamics of High-Risk Klebsiella pneumoniae ST147 Clones: Insights from Comparative Pangenome Analysis.

BACKGROUND: The high prevalence and rapid emergence of antibiotic resistance in high-risk Klebsiella pneumoniae (KP) ST147 clones is a global health concern and warrants molecular surveillance. METHODS: A pangenome analysis was performed using publicly available ST147 complete genomes. The characteristics and evolutionary relationships among ST147 members were investigated through a Bayesian phylogenetic analysis. RESULTS: The large number of accessory genes in the pangenome indicates genome plasticity and openness. Seventy-two antibiotic resistance genes were found to be linked with antibiotic inactivation, efflux, and target alteration. The exclusive detection of the blaOXA-232 gene within the ColKp3 plasmid of KP_SDL79 suggests its acquisition through horizontal gene transfer. The association of seventy-six virulence genes with the acrAB efflux pump, T6SS system and type I secretion system describes its pathogenicity. The presence of Tn6170, a putative Tn7-like transposon in KP_SDL79 with an insertion at the flanking region of the tnsB gene, establishes its transmission ability. The Bayesian phylogenetic analysis estimates ST147's initial divergence in 1951 and the most recent common ancestor for the entire KP population in 1621. CONCLUSIONS: Present study highlights the genetic diversity and evolutionary dynamics of high-risk clones of K. pneumoniae. Further inter-clonal diversity studies will help us understand its outbreak more precisely and pave the way for therapeutic interventions.

Identification of core therapeutic targets for Monkeypox virus and repurposing potential of drugs against them: An in silico approach.

Monkeypox virus (mpox virus) outbreak has rapidly spread to 82 non-endemic countries. Although it primarily causes skin lesions, secondary complications and high mortality (1-10%) in vulnerable populations have made it an emerging threat. Since there is no specific vaccine/antiviral, it is desirable to repurpose existing drugs against mpox virus. With little knowledge about the lifecycle of mpox virus, identifying potential inhibitors is a challenge. Nevertheless, the available genomes of mpox virus in public databases represent a goldmine of untapped possibilities to identify druggable targets for the structure-based identification of inhibitors. Leveraging this resource, we combined genomics and subtractive proteomics to identify highly druggable core proteins of mpox virus. This was followed by virtual screening to identify inhibitors with affinities for multiple targets. 125 publicly available genomes of mpox virus were mined to identify 69 highly conserved proteins. These proteins were then curated manually. These curated proteins were funnelled through a subtractive proteomics pipeline to identify 4 highly druggable, non-host homologous targets namely; A20R, I7L, Top1B and VETFS. High-throughput virtual screening of 5893 highly curated approved/investigational drugs led to the identification of common as well as unique potential inhibitors with high binding affinities. The common inhibitors, i.e., batefenterol, burixafor and eluxadoline were further validated by molecular dynamics simulation to identify their best potential binding modes. The affinity of these inhibitors suggests their repurposing potential. This work can encourage further experimental validation for possible therapeutic management of mpox.

In vitro synergistic interaction of colistin and other antimicrobials against intrinsic colistin-resistant Morganella morganii isolates.

Morganella morganii, a non-negligent opportunistic pathogen of the family Enterobacteriaceae, enlisted recently in the global priority pathogens by WHO for its swift propensity to acquire drug-resistant genes, engendering enhanced death rates. A combination of diverse antimicrobials could be recycled to overcome the ongoing acquisition of resistance mechanisms by M. morganii. Herein, we investigated the in vitro synergistic effect of colistin with meropenem, rifampicin, minocycline and linezolid against three intrinsic colistin-resistant M. morganii strains collected from critical departments of tertiary care hospitals. The strains were identified and tested for antimicrobial susceptibility by VITEK 2 automated system. The 16S rRNA sequencing was used to reconfirm the species identification. Minimum inhibitory concentrations (MICs) of colistin, meropenem, rifampicin, minocycline and linezolid were determined by the broth microdilution method. Synergistic interactions were studied by checkerboard and time-kill assay. The VITEK 2 identification and 16S rRNA sequencing confirmed that the strains were M. morganii. The automated antimicrobial susceptibility test revealed that all three isolates were multi-drug resistant. The checkerboard analysis demonstrated the synergy of all four combinations with FICI values ranging from 0.06 to 0.31 in all three isolates. These results suggest a potential role of meropenem as an adjuvant for treating M. morganii infections. The current work presented the first evidence of synergy between colistin and other antibiotics against M. morganii infection, which needs validation through in vitro and in vivo studies using a larger number of isolates. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03551-w.

Genomic insight of extremely drug-resistant Klebsiella pneumoniae ST5378 from a paediatric bloodstream infection.

OBJECTIVES: This study investigated the draft genome and phylogeny of an extremely drug-resistant and novel sequence type Klebsiella pneumoniae isolated from a paediatric bloodstream infection. METHODS: An isolate from a 7-year-old child with severe respiratory infection was identified, and the whole genome was sequenced using the Illumina MiSeq platform. High-quality reads were de novo assembled via Unicycler and annotated via PROKKA. Antimicrobial resistance genes, virulence factors, and plasmid and phage sequences were identified using the resistance gene identifier, VFanalyzer, Plasmidfinder, and PHASTER, respectively. Phylogenetics of closely related strains were inferred using core-genome multi-locus sequence typing and single nucleotide polymorphism. RESULTS: The draft genome of carbapenem-resistant K. pneumoniae RKS87 was 5 580 330 bp in size, with a GC content of 57.73%. The final assembly resulted in 38 contigs comprising 5075 CDS, 124 pseudo genes, 83 tRNA, 25 rRNA, and 10 ncRNA. The strain was assigned to a novel sequence type, ST5378, and harboured blaSHV-11, blaCTX-M-15, blaTEM-1, blaNDM-1, APH(3')-VI, OqxA, QnrS1, and fosA. We also identified the mutations in outer membrane porin (OmpK36 and OmpK37) and two-component system genes (PmrB and EptB). Three biomarkers (iroE, iroN, and iutA) associated with hypervirulent phenotype were also present in the genome. Phylogenetics of closely related strains revealed the clonal lineage of ST2938. CONCLUSIONS: The genome sequence and phylogenetics of the strain offer valuable insight into the clonal lineage, resistance genes, and pathogenicity of the novel sequence type ST5378.

Taxonomic Assignment-Based Genome Reconstruction from Apical Periodontal Metagenomes to Identify Antibiotic Resistance and Virulence Factors.

Primary apical periodontitis occurs due to various insults to the dental pulp including microbial infections, physical and iatrogenic trauma, whereas inadequate elimination of intraradicular infection during root canal treatment may lead to secondary apical periodontitis. We explored the complex intra-radicular microbial communities and their functional potential through genome reconstruction. We applied shotgun metagenomic sequencing, binning and functional profiling to identify the significant contributors to infection at the acute and chronic apical periodontal lesions. Our analysis revealed the five classified clusters representing Enterobacter, Enterococcus, Lacticaseibacillus, Pseudomonas, Streptococcus and one unclassified cluster of contigs at the genus level. Of them, the major contributors were Pseudomonas, with 90.61% abundance in acute conditions, whereas Enterobacter followed by Enterococcus with 69.88% and 15.42% abundance, respectively, in chronic conditions. Enterobacter actively participated in antibiotic target alteration following multidrug efflux-mediated resistance mechanisms, predominant in the chronic stage. The prediction of pathways involved in the destruction of the supportive tissues of the tooth in Enterobacter and Pseudomonas support their crucial role in the manifestation of respective disease conditions. This study provides information about the differential composition of the microbiome in chronic and acute apical periodontitis. It takes a step to interpret the role of a single pathogen, solely or predominantly, in establishing endodontic infection types through genome reconstruction following high throughput metagenomic DNA analysis. The resistome prediction sheds a new light on the therapeutic treatment guidelines for endodontists. However, it needs further conclusive research to support this outcome using a larger number of samples with similar etiological conditions, but different demographic origin.

Urban wastewater contributes to the emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) in an urban receiving river in eastern India.

The present study revealed the emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) and the associated driving factors in an urban river system surrounding Cuttack city, Odisha. The high contamination factor and contamination degree indicate poor water quality. The CRKP isolates showed 100% resistance against piperacillin, amoxicillin-clavulanic acid, piperacillin-tazobactam, ceftriaxone, ceftazidime, meropenem, and imipenem but less resistance to colistin (12.85%). Among the CRKP isolates, carbapenemase genes blaNDM, blaOXA-48-like, and blaKPC were detected in 94.28%, 35%, and 10% of isolates, respectively. The resistance genes (blaNDM, blaTEM, and blaCTX-M) were found to be significantly correlated with toxic metals (As, Cd, Co, Cu, Fe, Mn, Pb) (P < 0.05). Detection of virulence factors (yersiniabactin and aerobactin) and capsular serotypes (K1, K2, and K54 types) explain the pathogenicity of CRKP isolates. Enterobacterial repetitive intergenic consensus-PCR based molecular typing separated the CRKP strains into 13 clusters, of which VI and XI clusters showed similar resistance and virulence determinants, indicating the dissemination of clones from wastewater to the river system. Our results provide first-hand information on assessing risks to public health posed by the CRKP isolates and toxic metals in the Kathajodi River. Molecular surveillance of nearby hospitals for the prevalence of CRKP will help trace their transmission route.

Characterization and Comparative Genomic Analysis of a Highly Colistin-Resistant Chryseobacterium gallinarum: a Rare, Uncommon Pathogen.

For the first time, we describe the whole genome of a yellow-pigmented, capsule-producing, pathogenic, and colistin-resistant Chryseobacterium gallinarum strain MGC42 isolated from a patient with urinary tract infection in India. VITEK 2 automated system initially identified this isolate as C. indologenes. However, 16S rRNA gene sequencing revealed that MGC42 shared 99.67% sequence identity with C. gallinarum-type strain DSM 27622. The draft genome of the strain MGC42 was 4,455,926 bp long with 37.08% Guanine-Cytosine (GC) content and was devoid of any plasmid. Antibiotic resistance, virulence, and toxin genes were predicted by implementing a machine learning classifier. Potential homologs of 340 virulence genes including hemolysin secretion protein D, metalloprotease, catalase peroxidases and autotransporter adhesins, type VI secretion system (T6SS) spike proteins, and 27 toxin factors including a novel toxin domain Ntox23 were identified in the genome. Kyoto Encyclopedia of Genes and Genomes (KEGG) orthologs of 110 transporter proteins were predicted that were in agreement with moderate efflux activity. Twelve antibiotic resistance genes including two potentially novel putative ß-lactamase genes sharing low similarity with known ß-lactamase genes were also identified in the genome of this strain. The strain MGC42 was also resistant to several classes of antibiotics along with carbapenems and polymyxin. We also identified mutations in the orthologs of pmrB (M384T) and lpxD (I66V) that might be responsible for colistin resistance. The MGC42 strain shared 683 core genes with other environmental and clinical strains of Chryseobacterium species. Our findings suggest that the strain MGC42 is a multidrug-resistant, virulent pathogen and recommend 16S rRNA gene sequencing to identify clinical specimens of Chryseobacterium species.

Molecular prevalence of resistance determinants, virulence factors and capsular serotypes among colistin resistance carbapenemase producing Klebsiella pneumoniae: a multi-centric retrospective study.

The emergence of colistin-carbapenem-resistant Klebsiella pneumoniae (CCR-Kp) in bloodstream infection results in high mortality, and virulence factor contributes further to the difficulty of treatment. A total of 158 carbapenem-resistant K. pneumoniae (CRKP) isolates causing bloodstream infection were collected from three Indian tertiary care hospitals during the 9-month study period, of which 27 isolates exhibited resistance to both colistin and carbapenem antibiotics. In this study, all the strains were characterized for antimicrobial resistance, virulence factors and capsular serotypes that facilitate the development of colistin and carbapenem-resistant K.pneumoniae (CCR-Kp) in bloodstream infection. Fourteen isolates displayed extremely drug resistance (XDR), susceptible only to tigecycline, and the remaining 13 isolates displayed multidrug resistance (MDR). The gene prevalence analysis for CCR-Kp isolates showed the predominance of bla KPC (81.48%) followed by bla NDM (62.96%), bla VIM (37.03%) and bla IMP (18.51%) genes. The distribution of virulence genes was found to be fimH (81.48%), wabG (59.25%), mrkD (55.56%), entB (48.15%), irp1 (33.33%), and rmpA (18.52%). The capsular serotypes K1, K2, K5 and K54 have been identified in 16 isolates. The absence of plasmid-mediated colistin resistance (mcr) genes implies the involvement of other mechanisms. The ERIC and (GTG)5 molecular typing methods detected 18 and 22 distinct clustering patterns among the CCR-Kp isolates, respectively. A strong correlation between ERIC and (GTG)5 genotyping method was established with antimicrobial resistance patterns and virulence determinants at P < 0.05, while no correlation was found with capsular serotyping. Similar virulence and resistance typing among the isolates suggest hospital-acquired infection in a health care setup. These outcomes will advance our awareness of CCR-Kp outbreaks associated with tertiary care hospitals and help forecast their occurrence in the near future. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-03056-4.

Bacterial Diversity and CAZyme Potential Revealed in Pandanus Rich Thermal Spring Cluster of India: A Non-cultivable 16S rRNA Sequencing Approach.

In the present study, we explored four different geothermal spots of the Deulajhari spring cluster at a proximity of 10-20 meters with temperatures of 43 to 65°C to unravel their genesis, bacterial diversity and CAZyme potential. However, minor variations in physicochemical properties; TOC, sodium, chloride, zinc and nitrate were observed, including the pH of the spring openings. Illumina based amplicon sequencing revealed Firmicutes, Proteobacteria and Chloroflexi as the major bacterial phylum with higher abundance in the DJ04 sample. The alpha diversity of all the springs was almost same, whereas beta diversity revealed variations in the degree of uniqueness of OTUs at different temperatures. Statistical analysis established a positive correlation between sulfur content with Heliobacterium, Thermodesulfovibrio, Thermodesulfobacterium and Herpetosipho as well as TOC and HCO3 with Thermoanaerobacter, Desulfovibrio, Candidatus solibacter and Dehalogenimona. The major hydrocarbon family genes and Carbohydrate Active Enzyme pathways were predicted to be highest in DJ04 with elevated concentrations of HCO3 and TOC. Higher homogeneity in geo-physicochemical and microbial features direct the possibility of the common origin of these springs through plumbing systems. However, the minor variations in diversity and functionality were due to variations in temperature in spring openings through the mixing of subsurface water contaminated with carbohydrates from leaf biomass litter. Functional characterization of the thermophilic bacteria of this spring provides essential scope for further industrial applications. The biogeochemical reasons hypothesized for the genesis of unique multiple openings in the cluster are also of interest to conservation scientists for taking measures toward necessary laws and regulations to protect and preserve these springs.

Genomic characterization of XDR Klebsiella pneumoniae ST147 co-resistant to carbapenem and colistin - The first report in India.

OBJECTIVES: The emergence and outbreak of colistin-resistant CRKP (carbapenem-resistant Klebsiella pneumoniae) have been the major global public threat in recent years. Present study emphasized the genome-wide distribution, characterization of drug resistance virulence genes in an extremely drug-resistant (XDR) Klebsiella pneumoniae strain isolated from a patient with drug-induced hepatitis, hospitalized in a tertiary care facility in India. METHODS: The total genomic DNA was sequenced using the Illumina Hiseq platform. De novo assembly of reads was done using CLC genomics workbench. Genome annotation was performed using PROKKA. Sequence typing (ST), virulence-related genes and antimicrobial resistance genes were predicted from genome sequences. Phenotypic evaluation of efflux pump function was done in presence of colistin and efflux pump inhibitor (EPI). RESULT: Antibiogram analysis confirmed the isolate to be XDR. The number of contigs in assembly file was found to be 867 with a total of 6,060,836 bases and a total of 5547 coding sequences. The isolate exhibited high resistance to colistin due to mutations in two-component systems and predicted to be efflux mediated. The sequence typing of Klebsiella pneumoniae SDL79 is assigned to ST147. CONCLUSION: This is the first whole genome analysis of XDR Klebsiella pneumoniae ST147 from a hospital conferring co-resistance to last resort drugs. However, the detailed molecular mechanism behind the drug resistance will be carried out in our future endeavors.

Parameter optimization for thermostable lipase production and performance evaluation as prospective detergent additive.

Lipase based formulations has been a rising interest to laundry detergent industry for their eco-friendly property over phosphate-based counterparts and compatibility with chemical detergents ingredients. A thermo-stable Anoxybacillus sp. ARS-1 isolated from Taptapani Hotspring, India was characterized for optimum lipase production employing statistical model central composite design (CCD) under four independent variables (temperature, pH, % moisture and bio-surfactant) by solid substrate fermentation (SSF) using mustard cake. The output was utilized to find the effect of parameters and their interaction employing response surface methodology (RSM). A quadratic regression with R2 = 0.955 established the model to be statically best fitting and a predicted highest lipase production of 29.4 IU/g at an optimum temperature of 57.5 °C, pH 8.31, moisture 50% and 1.2 mg of bio-surfactant. Experimental production of 30.3 IU/g lipase at above conditions validated the fitness of model. Anoxybacillus sp. ARS-1 produced lipase was found to resist almost all chemical detergents as well as common laundry detergent, proving it to be a prospective additive for incorporation.

Cellulolytic thermophilic microorganisms in white biotechnology: a review.

Enzymes of microbial origin are of immense importance for organic material decomposition leading to bioremediation of organic waste, bioenergy generation, large-scale industrial bioprocesses, etc. The market demand for microbial cellulase enzyme is growing more rapidly which ultimately becomes the driving force towards research on this biocatalyst, widely used in various industrial activities. The use of novel cellulase genes obtained from various thermophiles through metagenomics and genetic engineering as well as following metabolic engineering pathways would be able to enhance the production of thermophilic cellulase at industrial scale. The present review is mainly focused on thermophilic cellulolytic bacteria, discoveries on cellulase gene, genetically modified cellulase, metabolic engineering, and their various industrial applications. A lot of lacunae are yet to overcome for thermophiles such as metagenome analysis, metabolic pathway modification study, search of heterologous hosts in gene expression system, and improved recombinant strain for better cellulase yield as well as value-added product formation.

Genotypic validation of extended-spectrum ß-lactamase and virulence factors in multidrug resistance Klebsiella pneumoniae in an Indian hospital.

The emergence of extended-spectrum ß-lactamase (ESBL)-producing Klebsiella pneumoniae has been increasing rapidly across the world. The presence of virulence factors in ESBL producers further adds to the pathogenicity and severity of infection, which often complicate empirical therapy and sometimes result in treatment failures. In the present study, 227 non-repeated clinical isolates of K. pneumoniae obtained from different clinical specimens from a tertiary care hospital in India were analyzed to detect the genes responsible for ESBL production (blaTEM, blaCTX-M, and blaSHV), virulence (fimH-1, mrkD, entB, irp-1), and capsule production (K1-K2). Phenotypically identified 72 ESBL producing K. pneumoniae isolates were further subjected to PCR based genotypic analysis but only 20 were found to have at least one of the ESBL producing genes. blaTEM was the most predominant gene (100%), followed by blaSHV (90%), and blaCTX-M (85%). Similarly, the most common virulence genes were fimH-1 (70%), entB (65%), markD (55%), irp-1 (25%), K1 (25%), and K2 (20%). REP-PCR profile separated them into five major clusters (I-V), indicating the existing heterogeneity among the isolates. The resistance profile data obtained from the present study can serve as the information base to understand the infection pattern prevailing in the hospital and for physicians to recommend suitable antibiotics for the patients.