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Skin cancer is a widespread malignancy that primarily affects light-skinned populations globally, categorized into melanoma and non-melanoma skin cancers (NMSCs). Basal cell carcinoma and squamous cell carcinoma are the most common subtypes within NMSCs, with the global incidence of NMSCs projected to reach 2–3 million cases annually across regions like Europe, Canada, the USA, and Australia. Despite this prevalence, the genetic mechanisms behind skin cancer remain poorly understood. This study presents a novel gene discovery approach, leveraging transcriptome data from Next-Generation Sequencing datasets sourced from the European Nucleotide Archive to uncover new genes and pathways linked to skin cancer. The novelty of this research lies in its comprehensive approach that combines differential gene expression analysis with gene network and pathway enrichment analysis to identify actionable therapeutic targets. By utilizing bioinformatics tools such as DESeq2, Gene Set Enrichment Analysis (GSEA), and Cytoscape, we revealed critical gene interactions and pathways that have been underexplored in the context of skin cancer. Following rigorous quality control using FastQC and transcriptome-seq data alignment to the human genome (hg38), we identified 19 differentially expressed genes, including 2 down-regulated and 17 up-regulated. Key genes such as IL6, CCND2, PLAUR, and CD44 were found to be involved in important pathways like IL6_JAK_STAT3_SIGNALING, ANGIOGENESIS, and APICAL_SURFACE. These findings provide valuable insights into skin cancer pathogenesis and offer potential therapeutic targets, laying the groundwork for future research aimed at improving treatment outcomes.

Context: Gastric cancer is one of the most prevalent types of cancer in developing countries and ranks fourth in terms of death causes. Helicobacter pylori infection is a significant contributor to the emergence of gastric cancer. Lack of early diagnosis of gastric cancer is a leading cause of death. Aims: To identify the key genes and pathways involved in gastric cancer. Methods: This study performed a comprehensive analysis of RNA-Seq data from human gastric cancer and adjacent normal tissues. Raw data passed quality checks with FastQC and were aligned to GRCh38 using HISAT2. Subread’s FeatureCounts handled transcript assembly and quantification. DESeq2 pinpointed significant genes, while ClueGO explored gene ontology and KEGG pathways. Protein-protein interaction networks, constructed with StringApp, aided in identifying hub genes through CytoHubba. This holistic approach yields insights into the molecular mechanisms underpinning gastric cancer. Results: This study detected 711 differentially expressed genes (DEGs) between normal and gastric samples. A total of 594 genes were identified as upregulated and 117 as downregulated. Major DEGs are enriched in signal transduction, stimulus-response regulation, transmembrane signaling receptor activity, and signal transduction pathways involving cytokines. In addition, 20 hub genes from the PPI network were identified based on MCC rank analysis from the CytoHubba plugin, contributing to the progression of gastric cancer. Conclusions: The top six hub genes, CD4, CTLA4, CD28, CD80, CD27, and SELL, are expected to regulate several pathways and may serve as potential biomarkers for the early detection and treatment of gastric cancer patients.

ABSTRACTColistin resistance in Acinetobacter baumannii is mediated by multiple mechanisms. Recently, mutations within pmrABC two-component system and overexpression of eptA gene due to upstream insertion of ISAba1 have been shown to play a major role. Thus, the aim of our study is to characterize colistin resistance mechanisms among the clinical isolates of A. baumannii in India. A total of 207 clinical isolates of A. baumannii collected from 2016 to 2019 were included in this study. Mutations within lipid A biosynthesis and pmrABC genes were characterized by whole-genome shotgun sequencing. Twenty-eight complete genomes were further characterized by hybrid assembly approach to study insertional inactivation of lpx genes and the association of ISAba1-eptA. Several single point mutations (SNPs), like M12I in pmrA, A138T and A444V in pmrB, and E117K in lpxD, were identified. We are the first to report two novel SNPs (T7I and V383I) in the pmrC gene. Among the five colistin-resistant A. baumannii isolates where complete genome was available, the analysis showed that three of the five isolates had ISAba1 insertion upstream of eptA. No mcr genes were identified among the isolates. We mapped the SNPs on the respective protein structures to understand the effect on the protein activity. We found that majority of the SNPs had little effect on the putative protein function; however, some SNPs might destabilize the local structure. Our study highlights the diversity of colistin resistance mechanisms occurring in A. baumannii, and ISAba1-driven eptA overexpression is responsible for colistin resistance among the Indian isolates.IMPORTANCEAcinetobacter baumannii is a Gram-negative, emerging and opportunistic bacterial pathogen that is often associated with a wide range of nosocomial infections. The treatment of these infections is hindered by increase in the occurrence of A. baumannii strains that are resistant to most of the existing antibiotics. The current drug of choice to treat the infection caused by A. baumannii is colistin, but unfortunately, the bacteria started to show resistance to the last-resort antibiotic. The loss of lipopolysaccharides and mutations in lipid A biosynthesis genes are the main reasons for the colistin resistance. The present study characterized 207 A. baumannii clinical isolates and constructed complete genomes of 28 isolates to recognize the mechanisms of colistin resistance. We showed the mutations in the colistin-resistant variants within genes essential for lipid A biosynthesis and that cause these isolates to lose the ability to produce lipopolysaccharides.

Aspartame, a widely used artificial sweetener, is present in many food products and beverages worldwide. It has been linked to potential neurotoxicity and developmental defects. However, its teratogenic effect on embryonic development and the underlying potential mechanisms need to be elucidated. We investigated the concentration- and time-dependent effects of aspartame on zebrafish development and teratogenicity. We focused on the role of sirtuin 1 (SIRT1) and Forkhead-box transcription factor (FOXO), two proteins that play key roles in neurodevelopment. It was found that aspartame exposure reduced the formation of larvae and the development of cartilage in zebrafish. It also delayed post-fertilization development by altering the head length and locomotor behavior of zebrafish. RNA-sequencing-based DEG analysis showed that SIRT1 and FOXO3a are involved in neurodevelopment. In silico and in vitro analyses showed that aspartame could target and reduce the expression of SIRT1 and FOXO3a proteins in neuron cells. Additionally, aspartame triggered the reduction of autophagy flux by inhibiting the nuclear translocation of SIRT1 in neuronal cells. The findings suggest that aspartame can cause developmental defects and teratogenicity in zebrafish embryos and reduce autophagy by impairing the SIRT1/FOXO3a axis in neuron cells.

Objectives: Recent advancement in understanding neurological disorders has revealed the involvement of dysbiosis of the gut microbiota in the pathophysiology of Parkinson's disease (PD). We sequenced microbial DNA using fecal samples collected from PD cases and healthy controls (HCs) to evaluate the role of gut microbiota. Methods: Full-length bacterial 16S rRNA gene sequencing of fecal samples was performed using amplified polymerase chain reaction (PCR) products on the GridION Nanopore sequencer. Sequenced data were analyzed using web-based tools BugSeq and MicrobiomeAnalyst. Results: We found that certain bacterial families like Clostridia UCG 014, Cristensenellaceae, and Oscillospiraceae are higher in abundance, and Lachinospiracea, Coriobacteriaceae and genera associated with short-chain fatty acid production, Faecalibacterium, Fusicatenibacter, Roseburia and Blautia, are lower in abundance among PD cases when compared with the HC. Genus Akkermansia, Dialister, Bacteroides, and Lachnospiraceae NK4A136 group positively correlated with constipation in PD. Conclusion: Observations from this study support the other global research on the PD gut microbiome background and provide fresh insight into the gut microbial composition of PD patients from a south Indian population. We report a higher abundance of Clostridia UCG 014 group, previously not linked to PD.

The O139 Vibrio cholerae serogroup emerged in the 1990s and spread rapidly but did not become globally dominant. Here, the authors describe the genomic epidemiology of this strain and identify changes in virulence and antimicrobial resistance characteristics that they hypothesise may have contributed to its decline.

Paratyphoid fever caused by S. Paratyphi A is endemic in parts of South Asia and Southeast Asia. The proportion of enteric fever cases caused by S. Paratyphi A has substantially increased, yet only limited data is available on the population structure and genetic diversity of this serovar. We examined the phylogenetic distribution and evolutionary trajectory of S. Paratyphi A isolates collected as part of the Indian enteric fever surveillance study "Surveillance of Enteric Fever in India (SEFI)." In the study period (2017-2020), S. Paratyphi A comprised 17.6% (441/2503) of total enteric fever cases in India, with the isolates highly susceptible to all the major antibiotics used for treatment except fluoroquinolones. Phylogenetic analysis clustered the global S. Paratyphi A collection into seven lineages (A-G), and the present study isolates were distributed in lineages A, C and F. Our analysis highlights that the genome degradation events and gene acquisitions or losses are key molecular events in the evolution of new S. Paratyphi A lineages/sub-lineages. A total of 10 hypothetically disrupted coding sequences (HDCS) or pseudogenes-forming mutations possibly associated with the emergence of lineages were identified. The pan-genome analysis identified the insertion of P2/PSP3 phage and acquisition of IncX1 plasmid during the selection in 2.3.2/2.3.3 and 1.2.2 genotypes, respectively. We have identified six characteristic missense mutations associated with lipopolysaccharide (LPS) biosynthesis genes of S. Paratyphi A, however, these mutations confer only a low structural impact and possibly have minimal impact on vaccine effectiveness. Since S. Paratyphi A is human-restricted, high levels of genetic drift are not expected unless these bacteria transmit to naive hosts. However, public-health investigation and monitoring by means of genomic surveillance would be constantly needed to avoid S. Paratyphi A serovar becoming a public health threat similar to the S. Typhi of today.

IntroductionOsteosarcoma is a rare disorder among cancer, but the most frequently occurring among sarcomas in children and adolescents. It has been reported to possess the relapsing capability as well as accompanying collateral adverse effects which hinder the development process of an effective treatment plan. Using networks of omics data to identify cancer biomarkers could revolutionize the field in understanding the cancer. Cancer biomarkers and the molecular mechanisms behind it can both be understood by studying the biological networks underpinning the etiology of the disease.MethodsIn our study, we aimed to highlight the hub genes involved in gene-gene interaction network to understand their interaction and how they affect the various biological processes and signaling pathways involved in Osteosarcoma. Gene interaction network provides a comprehensive overview of functional gene analysis by providing insight into how genes cooperatively interact to elicit a response. Because gene interaction networks serve as a nexus to many biological problems, their employment of it to identify the hub genes that can serve as potential biomarkers remain widely unexplored. A dynamic framework provides a clear understanding of biological complexity and a pathway from the gene level to interaction networks.ResultsOur study revealed various hub genes viz. TP53, CCND1, CDK4, STAT3, and VEGFA by analyzing various topological parameters of the network, such as highest number of interactions, average shortest path length, high cluster density, etc. Their involvement in key signaling pathways, such as the FOXM1 transcription factor network, FAK-mediated signaling events, and the ATM pathway, makes them significant candidates for studying the disease. The study also highlighted significant enrichment in GO terms (Biological Processes, Molecular Function, and Cellular Processes), such as cell cycle signal transduction, cell communication, kinase binding, transcription factor activity, nucleoplasm, PML body, nuclear body, etc.ConclusionTo develop better therapeutics, a specific approach toward the disease targeting the hub genes involved in various signaling pathways must have opted to unravel the complexity of the disease. Our study has highlighted the candidate hub genes viz. TP53, CCND1 CDK4, STAT3, VEGFA. Their involvement in the major signaling pathways of Osteosarcoma makes them potential candidates to be targeted for drug development. The highly enriched signaling pathways include FOXM1 transcription pathway, ATM signal-ling pathway, FAK mediated signaling events, Arf6 signaling events, mTOR signaling pathway, and Integrin family cell surface interactions. Targeting the hub genes and their associated functional partners which we have reported in our studies may be efficacious in developing novel therapeutic targets.

Emerging evidence from Alzheimer’s disease (AD) patients suggests that reducing tau pathology can restore cognitive and memory loss. To reduce tau pathology, it is critical to find brain-permeable tau-degrading small molecules that are safe and effective. HDAC6 inhibition has long been considered a safe and effective therapy for tau pathology. Recently, we identified protopine as a dibenzazecine alkaloid with anti-HDAC6 and anti-AD activities. In this study, we synthesized and tested novel protopine derivatives for their pharmacological action against AD. Among them, bromo-protopine (PRO-Br) demonstrated a two-fold increase in anti-HDAC6 activity and improved anti-tau activities compared to the parent compound in both in vitro and in vivo AD models. Furthermore, molecular docking results showed that PRO-Br binds to HDAC6, with a ∆G value of −8.4 kcal/mol and an IC50 value of 1.51 µM. In neuronal cell lines, PRO-Br reduced pathological tau by inducing chaperone-mediated autophagy (CMA). In 3xTg-AD and P301S tau mice models, PRO-Br specifically decreased the pathogenic hyperphosphorylated tau clumps and led to the restoration of memory functions. In addition, PRO-Br treatment promoted the clearance of pathogenic tau by enhancing the expression of molecular chaperones (HSC70) and lysosomal markers (LAMP2A) via CMA in AD models. Our data strongly suggest that administration of the brain-permeable protopine derivative PRO-Br, could be a viable anti-tau therapeutic strategy for AD.

Bacterial Extracellular Vesicles (BEVs) possess the capability of intracellular interactions with other cells, and, hence, can be utilized as an efficient cargo for worldwide delivery of therapeutic substances such as monoclonal antibodies, proteins, plasmids, siRNA, and small molecules for the treatment of neurodegenerative diseases (NDs). BEVs additionally possess a remarkable capacity for delivering these therapeutics across the blood–brain barrier to treat Alzheimer’s disease (AD). This review summarizes the role and advancement of BEVs for NDs, AD, and their treatment. Additionally, it investigates the critical BEV networks in the microbiome–gut–brain axis, their defensive and offensive roles in NDs, and their interaction with NDs. Furthermore, the part of BEVs in the neuroimmune system and their interference with ND, as well as the risk factors made by BEVs in the autophagy–lysosomal pathway and their potential outcomes on ND, are all discussed. To conclude, this review aims to gain a better understanding of the credentials of BEVs in NDs and possibly discover new therapeutic strategies.

ABSTRACT: Objectives: Linezolid resistance in Enterococcus faecium is emerging worldwide. In this study, we aimed to characterise two linezolid-resistant E. faecium isolates using whole-genome sequencing. Methods: Antimicrobial susceptibility testing was performed by the broth microdilution method. A hybrid assembly approach of IonTorrent and MinION sequencing reads was used to generate the complete genome of linezolid-resistant E. faecium isolates VB3025 and VB3240. Results: VB3025 and VB3240 had minimum inhibitory concentration (MICs) for linezolid of 1024 μg/mL and 512 μg/mL, respectively. In addition, VB3025 was found to be resistant to both vancomycin and teicoplanin, while VB3240 was susceptible to these antibiotics. A hybrid assembly approach was used to generate the complete genome of VB3025 and VB3240 isolates harbouring the optrA gene. Notably, VB3025 carried two copies of optrA (chromosomal and plasmid), while in VB3240 optrA was identified on the chromosome. Interestingly, the plasmid pVB3025_2 co-carried the resistance gene clusters aph(3)-IIIa–sat4–ant(6)-Ia–ermB, the vanHAX operon and a copy of the optrA gene. Moreover, the optrA gene inserted into a Tn554 transposon carrying the ermA gene was identified in both VB3025 and VB3240 isolates. Furthermore, mutation analysis revealed the presence of a G2592T mutation in the 23S rRNA of both isolates. Conclusion: This is the first study reporting optrA-positive linezolid-resistant E. faecium from India. A novel plasmid co-carrying vancomycin and linezolid resistance determinants highlights the threat for potential dissemination.

With the excessive genome plasticity, Acinetobacter baumannii can acquire and disseminate antimicrobial resistance (AMR) genes often associated with mobile genetic elements (MGEs). Analyzing the genetic environment of resistance genes often provides valuable information on the origin, emergence, evolution, and spread of resistance. Thus, we characterized the genomic features of some clinical isolates of carbapenem-resistant A. baumannii (CRAb) to understand the role of diverse MGEs and their genetic context responsible for disseminating carbapenem resistance genes. For this, 17 clinical isolates of A. baumannii obtained from multiple hospitals in India between 2018 and 2019 were analyzed. AMR determinants, the genetic context of resistance genes, and molecular epidemiology were studied using whole-genome sequencing. This study observed an increased prevalence of blaOXA–23 followed by dual carbapenemases, blaOXA–23, and blaNDM. This study identified three novel Oxford MLST sequence types. The majority of the isolates belonged to the dominant clone, IC2, followed by less prevalent clones such as IC7 and IC8. This study identified variations of AbaR4 and AbGRI belonging to the IC2 lineage. To the best of our knowledge, this is the first study that provides comprehensive profiling of resistance islands, their related MGEs, acquired AMR genes, and the distribution of clonal lineages of CRAb from India.

Cases of diphtheria have increased in recent years. Here, the authors analyse the genomes of 502 Corynebacterium diphtheriae isolates across 16 countries and territories over 122 years, describing an increase in antimicrobial resistance genes and identifying toxin variants.

BackgroundHypervirulent variants of Klebsiella pneumoniae (HvKp) were typically associated with a broadly antimicrobial susceptible clone of sequence type (ST) 23 at the time of its emergence. Concerningly, HvKp is now also emerging within multidrug-resistant (MDR) clones, including ST11, ST15, and ST147. MDR-HvKp either carry both the virulence and resistance plasmids or carry a large hybrid plasmid coding for both virulence and resistance determinants. Here, we aimed to genetically characterize a collection of MDR-HvKp ST2096 isolates haboring hybrid plasmids carrying both antimicrobial resistance (AMR) and virulence genes.MethodsNine K. pneumoniae ST2096 isolated over 1 year from the blood sample of hospitalized patients in southern India that were MDR and suspected to be HvKp were selected. All nine isolates were subjected to short-read whole-genome sequencing; a subset (n = 4) was additionally subjected to long-read sequencing to obtain complete genomes for characterization. Mucoviscosity assay was also performed for phenotypic assessment.ResultsAmong the nine isolates, seven were carbapenem-resistant, two of which carried blaNDM-5 on an IncFII plasmid and five carried blaOXA-232 on a ColKP3 plasmid. The organisms were confirmed as HvKp, with characteristic virulence genes (rmpA2, iutA, and iucABCD) carried on a large (~320 kbp) IncFIB–IncHI1B co-integrate. This hybrid plasmid also carried the aadA2, armA, blaOXA-1, msrE, mphE, sul1, and dfrA14 AMR genes in addition to the heavy-metal resistance genes. The hybrid plasmid showed about 60% similarity to the IncHI1B virulence plasmid of K. pneumoniae SGH10 and ~70% sequence identity with the first identified IncHI1B pNDM-MAR plasmid. Notably, the hybrid plasmid carried its type IV-A3 CRISPR-Cas system which harbored spacer regions against traL of IncF plasmids, thereby preventing their acquisition.ConclusionThe convergence of virulence and AMR is clinically concerning in K. pneumoniae. Our data highlight the role of hybrid plasmids carrying both AMR and virulence genes in K. pneumoniae ST2096, suggesting that MDR-HvKp is not confined to selected clones; we highlight the continued emergence of such genotypes across the species. The convergence is occurring globally amidst several clones and is of great concern to public health.

BackgroundThe incidence of hypervirulent (hv) carbapenem-resistant (CR) Klebsiella pneumoniae (Kp) is increasing globally among various clones and is also responsible for nosocomial infections. The CR-hvKp is formed by the uptake of a virulence plasmid by endemic high-risk clones or by the uptake of plasmids carrying antimicrobial resistance genes by the virulent clones. Here, we describe CR-hvKp from India belonging to high-risk clones that have acquired a virulence plasmid and are phenotypically unidentified due to lack of hypermucoviscosity.MethodsTwenty-seven CRKp isolates were identified to possess rmpA2 by whole-genome sequencing; and resistance and virulence determinants were characterized. By in silico protein modeling (and validation), protein backbone stability analysis, and coarse dynamics study, the fitness of RmpA, RmpA2, and aerobactin-associated proteins-IucA and IutA, were determined to establish a reliable marker for clinical identification of CR-hvKp.ResultsThe CR-hvKp belonged to multidrug-resistant (MDR) high-risk clones such as CG11, CG43, ST15, and ST231 and carried OXA-232 as the predominant carbapenemase followed by NDM. The virulence plasmid belonged to IncHI1B replicon type and carried frameshifted and truncated rmpA and rmpA2. This resulted in a lack of hypermucoviscous phenotype. However, functional aerobactin was expressed in all high-risk clones. In silico analysis portrayed that IucA and IutA were more stable than classical RmpA. Furthermore, IucA and IutA had lower conformational fluctuations in the functional domains than the non-functional RmpA2, which increases the fitness cost of the latter for its maintenance and expression among CR-hvKp. Hence, RmpA and RmpA2 are likely to be lost among CR-hvKp owing to the increased fitness cost while coding for essential antimicrobial resistance and virulence factors.ConclusionIncreasing incidence of convergence of AMR and virulence is observed among K. pneumoniae globally, which warrants the need for reliable markers for identifying CR-hvKp. The presence of non-functional RmpA2 among high-risk clones highlights the significance of molecular identification of CR-hvKp. The negative string test due to non-functional RmpA2 among CR-hvKp isolates challenges phenotypic screening and faster identification of this pathotype. This can potentially be counteracted by projecting aerobactin as a stable, constitutively expressed, and functional marker for rapidly evolving CR-hvKp.

ABSTRACT The currently ongoing COVID-19 pandemic caused by SARS-CoV-2 has accounted for millions of infections and deaths across the globe. Genome sequences of SARS-CoV-2 are being published daily in public databases and the availability of these genome data sets has allowed unprecedented access to the mutational patterns of SARS-CoV-2 evolution. We made use of the same genomic information for conducting phylogenetic analysis and identifying lineage-specific mutations. The catalogued lineage-defining mutations were analyzed for their stabilizing or destabilizing impact on viral proteins. We recorded persistence of D614G, S477N, A222V, and V1176F variants and a global expansion of the PANGOLIN variant B.1. In addition, a retention of Q57H (B.1.X), R203K/G204R (B.1.1.X), T85I (B.1.2-B.1.3), G15S+T428I (C.X), and I120F (D.X) variations was observed. Overall, we recorded a striking balance between stabilizing and destabilizing mutations, therefore leading to well-maintained protein structures. With selection pressures in the form of newly developed vaccines and therapeutics to mount in the coming months, the task of mapping viral mutations and recording their impact on key viral proteins should be crucial to preemptively catch any escape mechanism for which SARS-CoV-2 may evolve. IMPORTANCE Since its initial isolation in Wuhan, China, large numbers of SARS-CoV-2 genome sequences have been shared in publicly accessible repositories, thus enabling scientists to do detailed evolutionary analysis. We investigated the evolutionarily associated mutational diversity overlaid on the major phylogenetic lineages circulating globally, using 513 representative genomes. We detailed the phylogenetic persistence of key variants facilitating global expansion of the PANGOLIN variant B.1, including the recent, fast-expanding, B.1.1.7 lineage. The stabilizing or destabilizing impact of the catalogued lineage-defining mutations on viral proteins indicates their possible involvement in balancing the protein function and structure. A clear understanding of this mutational profile is of high clinical significance to catch any vaccine escape mechanism, as the same proteins make crucial components of vaccines that have recently been approved or are in development. In this vein, our study provides an imperative framework and baseline data upon which further analysis could be built as newer variants of SARS-CoV-2 continue to appear.

Objectives: Escherichia coli is regarded as one of the most commonly isolated Gram-negative pathogens from bloodstream infections. Increasing antimicrobial resistance (AMR) among E. coli is a threat to disease management as well as further dissemination of AMR genes to other clinically important pathogens. Here we report the genome of a multidrug-resistant (MDR) E. coli (BA22372) from a bloodstream infection belonging to ST410 B4/H24RxC subtype from India. Methods: Genomic DNA of E. coli BA22372 was sequenced using Ion Torrent™ PGM™ and MinION™ sequencing. Hybrid genome assembly was performed using short and long reads from both methods to achieve accurate and complete genome data. Results: Here we report the genome of MDR E. coli BA22372 harbouring blaOXA-181 and blaCTX-M-15 in two individual plasmids, namely pOXA181_22372 (IncX3) and pCTX-M-15_22372 (IncF). The pCTX-M-15 plasmid is well known to co-harbour blaNDM-5, which was not seen in the studied isolate here. Conclusion: To the best of our knowledge, this is the first report of B4/H24RxC MDR E. coli from India co-harbouring blaCTX-M-15 and blaOXA-181 along with other AMR genes. Information from this genome data revealed the possession of AMR genes in two individual plasmids and their potential for rapid dissemination. This isolate is of high health concern as it harbours a plasmid with replicatory mechanisms capable of acquiring blaNDM-5, which is a great threat for rapid dissemination of AMR. This study enhances our understanding of the AMR mechanisms among different clones of E. coli.

Abstract Shigella is ranked as the second leading cause of diarrheal disease worldwide. Though infection occurs in people of all ages, most of the disease burden constitutes among the children less than 5 years in low and middle income countries. Recent increasing incidence of drug resistant strains make this as a priority pathogen under the antimicrobial resistance surveillance by WHO. Despite this, only limited genomic studies on drug resistant Shigella exists. Here we report the first complete genome of clinical S. flexneri serotype 2a and S. sonnei strains using a hybrid approach of both long-read MinION (Oxford Nanopore Technologies) and short-read Ion Torrent 400 bp sequencing platforms. The utilization of this novel approach in the present study helped to identify the complete plasmid sequence of pSS1653 with structural genetic information of AMR genes such as sulII, tetA, tetR, aph(6)-Id and aph(3′’)-Ib. Identification of AMR genes in mobile elements in this human-restricted enteric pathogen is a potential threat for dissemination to other gut pathogens. The information on Shigella at genome level could help us to understand the genome dynamics of existing and emerging resistant clones.

BackgroundIn recent years, the emergence of multidrug resistant hypervirulent K. pneumoniae (MDR hvKp) isolates poses severe therapeutic challenge to global public health. The present study used the complete genome sequence of two MDR hvKp isolates belonging to ST23 to characterize the phylogenetic background and plasmid diversity.MethodsTwo hvKp isolates from patients with bacteremia were sequenced using Ion Torrent PGM and Oxford Nanopore MinION platforms and assembled by hybrid genome assembly approach. Comparative genomics approaches were used to investigate the population structure, evolution, virulence, and antimicrobial resistance of MDR hvKp strains.ResultsThe study isolates exhibited typical features of hvKp phenotypes associated with ST23. The convergence of multidrug resistance and hypervirulence were attributed by the presence of multiple plasmids including a 216 kb virulence plasmid and MDR plasmids belonging to IncA/C2, IncFIB, IncX3, and ColKP3 groups. The insertion of catA1 gene into virulence plasmid was observed along with genetic factors such as aerobactin, salmochelin, and rmpA2 that confer hvKp’s hypervirulent phenotype. The core genome single nucleotide polymorphism (SNP) phylogenetic analyses of the isolates showed the evolution of ST23 hvKp was predominantly driven by ICEKp acquisitions.ConclusionTo the best of our knowledge, this is the first report of MDR hvKp isolates of ST23 with insertion of catA1 gene into the virulence plasmid which presents the possibility of hotspot integration sites on the plasmids to aid acquisition of AMR genes. ST23 is no longer confined to susceptible strains of hvKp. Our findings emphasize the need for more studies on recombinant events, plasmid transmission dynamics and evolutionary process involving hvKp.

ABSTRACT Shigella is the second leading cause of bacterial diarrhea worldwide. Recently, Shigella sonnei seems to be replacing Shigella flexneri in low- and middle-income countries undergoing economic development. Despite this, studies focusing on these species at the genomic level remain largely unexplored. Here, we compared the genome sequences of S. flexneri and S. sonnei isolates from India with the publicly available genomes of global strains. Our analysis provides evidence for the long-term persistence of all phylogenetic groups (PGs) of S. flexneri and the recent dominance of the ciprofloxacin-resistant S. sonnei lineage in India. Within S. flexneri PGs, the majority of the study isolates belonged to PG3 within the predominance of serotype 2. For S. sonnei, the current pandemic involves globally distributed multidrug-resistant (MDR) clones that belong to Central Asia lineage III. The presence of such epidemiologically dominant lineages in association with stable antimicrobial resistance (AMR) determinants results in successful survival in the community. IMPORTANCE Shigella is the second leading cause of bacterial diarrhea worldwide. This has been categorized as a priority pathogen among enteric bacteria by the Global Antimicrobial Resistance Surveillance System (GLASS) of the World Health Organization (WHO). Recently, S. sonnei seems to be replacing S. flexneri in low- and middle-income countries undergoing economic development. Antimicrobial resistance in S. flexneri and S. sonnei is a growing international concern, specifically with the international dominance of the multidrug-resistant (MDR) lineage. Genomic studies focusing on S. flexneri and S. sonnei in India remain largely unexplored. This study provides information on the introduction and expansion of drug-resistant Shigella strains in India for the first time by comparing the genome sequences of S. flexneri and S. sonnei isolates from India with the publicly available genomes of global strains. The study discusses the key differences between the two dominant species of Shigella at the genomic level to understand the evolutionary trends and genome dynamics of emerging and existing resistance clones. The present work demonstrates evidence for the long-term persistence of all PGs of S. flexneri and the recent dominance of a ciprofloxacin-resistant S. sonnei lineage in India.

BACKGROUND:Multidrug-resistant (MDR) E. coli with extended-spectrum β-lactamases (ESBLs) is becoming endemic in health care settings around the world. Baseline data on virulence and antimicrobial resistance (AMR) of specific lineages of E. coli circulating in developing countries like India is currently lacking. METHODS:Whole-genome sequencing was performed for 60 MDR E. coli isolates. The analysis was performed at single nucleotide polymorphism (SNP) level resolution to identify the presence of their virulence and AMR genes. RESULTS:Genome comparison revealed the presence of ST-131 global MDR and ST410 as emerging-MDR clades of E. coli in India. AMR gene profile for cephalosporin and carbapenem resistance differed between the clades. Genotypes blaCTX-M-15 and blaNDM-5 were common among cephalosporinases and carbapenemases, respectively. For aminoglycoside resistance, rmtB was positive for 31.7% of the isolates, of which 95% were co-harboring carbapenemases. In addition, the FimH types and virulence gene profile positively correlated with the SNP based phylogeny, and also revealed the evolution of MDR clones among the study population with temporal accumulation of SNPs. The predominant clone was ST167 (blaNDM lineage) followed by ST405 (global clone ST131 equivalent) and ST410 (fast spreading high risk clone). CONCLUSIONS:This is the first report on the whole genome analysis of MDR E. coli lineages circulating in India. Data from this study will provide public health agencies with baseline information on AMR and virulent genes in pathogenic E. coli in the region.