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.

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.

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.

Characterization of novel metagenomic-derived lipase from Indian hot spring.

Extreme environments are the main source of industrially suitable biocatalysts. The non-cultivable approach of searching enzymes is known to provide ample scope to accomplish novelty for their industrial applications. Lip479 clone out of seven lipase-producing clones obtained from Taptapani hot spring was found to be optimally active at pH 8.0 and temperature 65 °C. The recombinant Lip479 was highly stable in organic solvents, methanol, DMF, DMSO, acetone, and dichloromethane. Lip479 lipase activity was enhanced in the presence of K+, Mn2+, Na+, Zn2+, and Ca2+ except for Fe3+. The ability of Lip479 lipase to act on long carbon chain of 4-nitrophenyl myristate suggests it might be a true lipase. Lip479 clone was found to have ORF of 1251 bp encoding 416 amino acid residues of 42.57 KDa size (theoretically calculated). The presence of conserved motif Ala-His-Ser-Gln-Gly and Zn2+-binding consensus sequence (GAAHAAKH) of the clone assigns the protein to lipase family 1.5. Phylogenetic lineage of the protein sequence of Lip479 was traced to family 1.5 as it was clubbed up with those of reported lipases of the same family. The above biochemical features indicated that Lip479 lipase can be a potential biocatalyst for its use in various industries.