Contrasting Distribution of SARS-CoV-2 Lineages across Multiple Rounds of Pandemic Waves in West Bengal, the Gateway of East and North-East States of India.

The evolution of viral variants and their impact on viral transmission have been an area of considerable importance in this pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We analyzed the viral variants in different phases of the pandemic in West Bengal, a state in India that is important geographically, and compared the variants with other states like Delhi, Maharashtra, and Karnataka, located in other regions of the country. We have identified 57 pango-lineages in 3,198 SARS-CoV-2 genomes, alteration in their distribution, as well as contrasting profiles of amino acid mutational dynamics across different waves in different states. The evolving characteristics of Delta (B.1.617.2) sublineages and alterations in hydrophobicity profiles of the viral proteins caused by these mutations were also studied. Additionally, implications of predictive host miRNA binding/unbinding to emerging spike or nucleocapsid mutations were highlighted. Our results throw considerable light on interesting aspects of the viral genomic variation and provide valuable information for improved understanding of wave-defining mutations in unfolding the pandemic. IMPORTANCE Multiple waves of infection were observed in many states in India during the coronavirus disease 2019 (COVID19) pandemic. Fine-scale evolution of major SARS-CoV-2 lineages and sublineages during four wave-window categories: Pre-Wave 1, Wave 1, Pre-Wave 2, and Wave 2 in four major states of India: Delhi (North), Maharashtra (West), Karnataka (South), and West Bengal (East) was studied using large-scale virus genome sequencing data. Our comprehensive analysis reveals contrasting molecular profiles of the wave-defining mutations and their implications in host miRNA binding/unbinding of the lineages in the major states of India.

Structure and Function of N-Acetylmannosamine Kinases from Pathogenic Bacteria.

Several pathogenic bacteria import and catabolize sialic acids as a source of carbon and nitrogen. Within the sialic acid catabolic pathway, the enzyme N-acetylmannosamine kinase (NanK) catalyzes the phosphorylation of N-acetylmannosamine to N-acetylmannosamine-6-phosphate. This kinase belongs to the ROK superfamily of enzymes, which generally contain a conserved zinc-finger (ZnF) motif that is important for their structure and function. Previous structural studies have shown that the ZnF motif is absent in NanK of Fusobacterium nucleatum (Fn-NanK), a Gram-negative bacterium that causes the gum disease gingivitis. However, the effect in loss of the ZnF motif on the kinase activity is unknown. Using kinetic and thermodynamic studies, we have studied the functional properties of Fn-NanK to its substrates ManNAc and ATP, compared its activity with other ZnF motif-containing NanK enzymes from closely related Gram-negative pathogenic bacteria Haemophilus influenzae (Hi-NanK), Pasteurella multocida (Pm-NanK), and Vibrio cholerae (Vc-NanK). Our studies show a 10-fold decrease in substrate binding affinity between Fn-NanK (apparent KM ≈ 700 µM) and ZnF motif-containing NanKs (apparent KM ≈ 60 µM). To understand the structural features that combat the loss of the ZnF motif in Fn-NanK, we solved the crystal structures of functionally homologous ZnF motif-containing NanKs from P. multocida and H. influenzae. Here, we report Pm-NanK:unliganded, Pm-NanK:AMPPNP, Pm-NanK:ManNAc, Hi-NanK:ManNAc, and Hi-NanK:ManNAc-6P:ADP crystal structures. Structural comparisons of Fn-NanK with Hi-NanK, Pm-NanK, and hMNK (human N-acetylmannosamine kinase domain of UDP-N-acetylglucosamine-2-epimerase/N-acetylmannosamine kinase, GNE) show that even though there is less sequence identity, they have high degree of structural similarity. Furthermore, our structural analyses highlight that the ZnF motif of Fn-NanK is substituted by a set of hydrophobic residues, which forms a hydrophobic cluster that helps the proper orientation of ManNac in the active site. In summary, ZnF-containing and ZnF-lacking NanK enzymes from different Gram-negative pathogenic bacteria are functionally very similar but differ in their metal requirement. Our structural studies unveil the structural modifications in Fn-NanK that compensate the loss of the ZnF motif in comparison to other NanK enzymes.

High prevalence of carbapenemase, AmpC ß-lactamase and aminoglycoside resistance genes in extended-spectrum ß-lactamase-positive uropathogens from Northern India.

OBJECTIVES: This study investigated the occurrence of extended-spectrum ß-lactamase (ESBL) genes coexisting with carbapenemase, AmpC and aminoglycoside resistance gene in uropathogens in India. METHODS: Antimicrobial susceptibility testing was performed by disk diffusion. Antimicrobial resistance genes were detected by multiplex PCR. RESULTS: Of 1516 consecutive urine samples, 454 (29.9%) showed significant bacteriuria with a single micro-organism, predominantly Escherichia coli (n=343), followed by Klebsiella pneumoniae (n=92), Pseudomonas aeruginosa (n=10) and Proteus mirabilis (n=9). Among the uropathogens, 61 ESBL-producers were identified containing blaCTX-M-15 (n=32), blaCTX-M-15+blaOXA-2 (n=15), blaCTX-M-15+blaOXA-2+blaTEM-1 (n=6), blaOXA-2 (n=5), blaOXA-2+blaSHV-76 (n=1), blaTEM-1+blaSHV-76 (n=1) and blaTEM-1 (n=1). All ESBL genes were located on horizontally transferable plasmids of incompatibility types HI1, I1, FIA+FIB, FIA and Y. Among the 61 ESBL-producers, 59 harboured carbapenemase genes, including blaNDM-5 (n=48), blaNDM-5+blaOXA-48 (n=5), blaNDM-5+blaIMP (n=5) and blaNDM-5+blaIMP+blaVIM (n=1). ESBL-producing uropathogens also harboured 16S rRNA methylase genes, including rmtB (n=9), rmtA (n=4), rmtC (n=1) and armA (n=1). ESBL-positive isolates also contained AmpC genes, including blaCIT (n=8) and blaDHA-1 (n=1). Imipenem and gentamicin had the lowest resistance rates against the uropathogens. CONCLUSION: This is the first report showing the high prevalence of carbapenemases in ESBL-positive isolates in this area. Regular surveillance for such resistance mechanisms will be useful for health personnel to treat infections by these multidrug-resistant pathogens.

Serotonin is essential for eye regeneration in planaria Schmidtea mediterranea.

Planaria is an ideal system to study factors involved in regeneration and tissue homeostasis. Little is known about the role of metabolites and small molecules in stem cell maintenance and lineage specification in planarians. Using liquid chromatography and mass spectrometry (LC-MS)-based quantitative metabolomics, we determined the relative levels of metabolites in stem cells, progenitors, and differentiated cells of the planarian Schmidtea mediterranea. Tryptophan and its metabolic product serotonin are significantly enriched in stem cells and progenitor population. Serotonin biosynthesis in these cells is brought about by a noncanonical enzyme, phenylalanine hydroxylase. Knockdown of Smed-pah leads to complete disappearance of eyes in regenerating planaria, while exogenous supply of serotonin and its precursor rescues the eyeless phenotype. Our results demonstrate a key role for serotonin in eye regeneration.

The First Report of Phenotypic and Molecular Characterization of Extended-Spectrum Beta-Lactamase-Producing Uropathogens in Sikkim and Darjeeling Hills of India.

Extended-spectrum ß-lactamase (ESBL)-producing bacteria are a global health threat both in hospital and in community settings. The emergence of these organisms poses major difficulty in treating infections. This study was carried out to assess major ESBL-producing uropathogens in female patients of Sikkim and Darjeeling by phenotypic and genotypic methods. Out of 1,516 urine samples, 454 uropathogens were isolated with a prevalence rate of 29.94%. Among them, Escherichia coli (74.3%) was the predominant type followed by Klebsiella pneumoniae (20.1%), Pseudomonas aeruginosa (2.4%), and Proteus mirabilis (1.98%). Four different ESBL genes were detected in 63 isolates, which included CTX-M (n = 32), CTX-M+OXA-2 (n = 15), CTX-M-15+OXA-2+TEM (n = 6), OXA-2 (n = 5), TEM+CTX-M-15 (n = 2), TEM+OXA-2+SHV-76 (n = 2), and TEM (n = 1). All ESBL genes (bla genes) were found on a plasmid, which was mostly of HI1, I1, FIA+FIB, FIA, and Y types and was horizontally transferable. Among all ESBL genes, blaCTX-M-I5 group was the most prevalent. The study of urinary tract infection (UTI) caused by ESBL-producing bacteria needs to be studied in other high-altitude parts of India to understand the actual burden of UTI in the female.