Genome Dynamics and Evolution of Multiple-Drug-Resistant Bacteria: Implications for Global Infection Control Priorities.

Genomics-driven molecular epidemiology of pathogenic bacteria has largely been carried out through functionally neutral/inert sequences, mostly entailing polymorphic gene loci or repetitive tracts. However, it is very important to harness phenotypically relevant markers to assign a valid functional epidemiological context to tracking of pathogens. These should include microbial acumen to acquire multiple drug resistance (MDR), their physiological coordinates with reference to clinical or community-level dynamics of incidence/transmission, and their response or refractoriness to the activated immune system. We propose that multidimensional and multicentric approaches, based on diverse data integration coupled with comparative genomics and functional molecular infection epidemiology, would likely be successful in tracking the emergence and spread of MDR pathogens and thereby guiding the global infection control strategies in a highly informed manner.

Evolutionary Dynamics Based on Comparative Genomics of Pathogenic Escherichia coli Lineages Harboring Polyketide Synthase (pks) Island.

The genotoxin colibactin is a secondary metabolite produced by the polyketide synthase (pks) island harbored by extraintestinal pathogenic E. coli (ExPEC) and other members of the Enterobacteriaceae that has been increasingly reported to have critical implications in human health. The present study entails a high-throughput whole-genome comparison and phylogenetic analysis of such pathogenic E. coli isolates to gain insights into the patterns of distribution, horizontal transmission, and evolution of the island. For the current study, 23 pks-positive ExPEC genomes were newly sequenced, and their virulome and resistome profiles indicated a preponderance of virulence encoding genes and a reduced number of genes for antimicrobial resistance. In addition, 4,090 E. coli genomes from the public domain were also analyzed for large-scale screening for pks-positive genomes, out of which a total of 530 pks-positive genomes were studied to understand the subtype-based distribution pattern(s). The pks island showed a significant association with the B2 phylogroup (82.2%) and a high prevalence in sequence type 73 (ST73; n = 179) and ST95 (n = 110) and the O6:H1 (n = 110) serotype. Maximum-likelihood (ML) phylogeny of the core genome and intergenic regions (IGRs) of the ST95 model data set, which was selected because it had both pks-positive and pks-negative genomes, displayed clustering in relation to their carriage of the pks island. Prevalence patterns of genes encoding RM systems in the pks-positive and pks-negative genomes were also analyzed to determine their potential role in pks island acquisition and the maintenance capability of the genomes. Further, the maximum-likelihood phylogeny based on the core genome and pks island sequences from 247 genomes with an intact pks island demonstrated horizontal gene transfer of the island across sequence types and serotypes, with few exceptions. This study vitally contributes to understanding of the lineages and subtypes that have a higher propensity to harbor the pks island-encoded genotoxin with possible clinical implications.IMPORTANCE Extraintestinal pathologies caused by highly virulent strains of E. coli amount to clinical implications with high morbidity and mortality rates. Pathogenic E. coli strains are evolving with the horizontal acquisition of mobile genetic elements, including pathogenicity islands such as the pks island, which produces the genotoxin colibactin, resulting in severe clinical outcomes, including colorectal cancer progression. The current study encompasses high-throughput comparative genomics and phylogenetic analyses to address the questions pertaining to the acquisition and evolution pattern of the genomic island in different E. coli subtypes. It is crucial to gain insights into the distribution, transfer, and maintenance of pathogenic islands, as they harbor multiple virulence genes involved in pathogenesis and clinical implications of the infection.

Genome Informatics and Machine Learning-Based Identification of Antimicrobial Resistance-Encoding Features and Virulence Attributes in Escherichia coli Genomes Representing Globally Prevalent Lineages, Including High-Risk Clonal Complexes.

Escherichia coli, a ubiquitous commensal/pathogenic member from the Enterobacteriaceae family, accounts for high infection burden, morbidity, and mortality throughout the world. With emerging multidrug resistance (MDR) on a massive scale, E. coli has been listed as one of the Global Antimicrobial Resistance and Use Surveillance System (GLASS) priority pathogens. Understanding the resistance mechanisms and underlying genomic features appears to be of utmost importance to tackle further spread of these multidrug-resistant superbugs. While a few of the globally prevalent sequence types (STs) of E. coli, such as ST131, ST69, ST405, and ST648, have been previously reported to be highly virulent and harboring MDR, there is no clarity if certain ST lineages have a greater propensity to acquire MDR. In this study, large-scale comparative genomics of a total of 5,653 E. coli genomes from 19 ST lineages revealed ST-wide prevalence patterns of genomic features, such as antimicrobial resistance (AMR)-encoding genes/mutations, virulence genes, integrons, and transposons. Interpretation of the importance of these features using a Random Forest Classifier trained with 11,988 genomic features from whole-genome sequence data identified ST-specific or phylogroup-specific signature proteins mostly belonging to different protein superfamilies, including the toxin-antitoxin systems. Our study provides a comprehensive understanding of a myriad of genomic features, ST-specific proteins, and resistance mechanisms entailing different lineages of E. coli at the level of genomes; this could be of significant downstream importance in understanding the mechanisms of AMR, in clinical discovery, in epidemiology, and in devising control strategies. IMPORTANCE With the leap in whole-genome data being generated, the application of relevant methods to mine biologically significant information from microbial genomes is of utmost importance to public health genomics. Machine-learning methods have been used not only to mine, curate, or classify the data but also to identify the relevant features that could be linked to a particular class/target. This is perhaps one of the pioneering studies that has attempted to classify a large repertoire of E. coli genome data sets (5,653 genomes) belonging to 19 different STs (including well-studied as well as understudied STs) using machine learning approaches. Important features identified by these approaches have revealed ST-specific signature proteins, which could be further studied to predict possible associations with the phenotypic profiles, thereby providing a better understanding of virulence and the resistance mechanisms among different clonal lineages of E. coli.

Survey to assess knowledge, attitude and practices for safe and efficient vein-to-vein transfusion practices and effect of educational intervention in a hospital setting.

OBJECTIVES: The primary aim was to assess the knowledge, attitude and practices (KAP) of employees (medical, paramedical and other hospital staff) towards blood donation and transfusion practices in a tertiary care hospital. A secondary objective was to assess and interpret the effect of an educational module on improvement in the KAP of employees in a hospital setting. BACKGROUND: For satisfactory practices, it is essential to initiate KAP studies. METHODS/MATERIALS: This was a prospective, observational study conducted among hospital staff (clinical and non-clinical) from January to December 2019. The study was divided into two phases: pre-educational module (P1) and post-educational module (P2). In both phases, a questionnaire was distributed. Knowledge was assessed by 30 questions, attitude by 20 questions and practice by 30 questions. If any individual had unsatisfactory scores in both the P1 and P2 phases (scores <40; 50%), they had to participate in a mandatory certificate course. RESULTS: A total of 180 individuals participated in the P1 and 172 participated in the P2 phase. Mean score for practice (0.471) was better than that for attitude (0.447) and knowledge (0.43). Factors associated with good scores were younger age group, more than 5 years of employment and clinical field of study. The total score for KAP increased, and a statistically significant difference (P-value < .05) was observed between P1 and P2 scores. Of 172 participants, 27 were asked to attend the 2-week certificate course due to unsatisfactory scores (score < 40) in both P1 and P2 phases. These 27 participants required attending this certificate course a mean of 1.67 ± 0.83 times to obtain satisfactory scores. CONCLUSION: Educational intervention is an important tool for improving KAP among not only physicians but hospital staff as well.

Genomic and Functional Characterization of Poultry Escherichia coli From India Revealed Diverse Extended-Spectrum ß-Lactamase-Producing Lineages With Shared Virulence Profiles.

Extended-spectrum ß-lactamases (ESBLs) form the most important resistance determinants prevalent worldwide. Data on ESBL-producing Escherichia coli from poultry and livestock are scarce in India. We present data on the functional and genomic characterization of ESBL-producing E. coli obtained from poultry in India. The whole genome sequences of 28 ESBL-producing E. coli were analyzed comprising of 12 broiler chicken E. coli isolates, 11 free-range chicken E. coli isolates, and 5 human extraintestinal pathogenic E. coli. All of the 28 ESBL-producing E. coli isolates were tested for antibiotic susceptibilities, in vitro conjugation, and virulence-associated phenotypic characteristics. A total of 13 sequence types were identified from the poultry E. coli, which included globally successful sequence types such as ST117 (9%), ST131 (4.3%), and ST10 (4.3%). The most common ESBL gene detected in poultry E. coli genomes was bla CTX-M-15 (17%). Also, FIB (73%) and FII (73%) were the most common plasmid replicons identified. Conjugation experiments demonstrated 54 (7/13), 30 (3/10), and 40% (2/5) of broiler, free-range, and human ExPEC E. coli to be able to transfer their ESBL genes, respectively. The in vitro virulence-associated phenotypic tests revealed the broiler, free-range, and human ExPEC isolates to be comparable in biofilm formation, resistance to serum bactericidal activity, adherence, and invasion capabilities. Our overall results showed prevalence of virulence phenotypes among the diverse ESBL-producing E. coli from poultry; while certain E. coli clones from broiler-poultry may indeed have the potential to cause infection in humans.

Molecular Genetic and Functional Analysis of pks-Harboring, Extra-Intestinal Pathogenic Escherichia coli From India.

Colibactin, a genotoxin, encoded by the pks pathogenicity island of Escherichia coli belonging to the B2 phylogroup has been reported as a determinant of bacterial pathogenicity. The present study was carried out to detect the pks pathogenicity island in extraintestinal pathogenic E. coli (ExPEC) isolated from a tertiary hospital in Pune, India. Of 462 isolates analyzed, the pks genomic island was detected in 35 (7.6%) isolates, which predominantly belonged to pathogenic phylogroup B2 (97%), and harbored virulence genes such as fimH, sfaD/E, and usp. Biofilm formation assay revealed 21 of the 35 pks-carrying isolates to be strong (SBF > 1.0), 10 isolates to be moderate (SBF = 0.5-1.0), and 4 as weak (SBF < 0.5) biofilm formers. All of the pks-carrying isolates proved resistant against bactericidal activity of human serum. Assays carried out to detect antimicrobial susceptibility revealed 11% of these isolates to be multidrug resistant, 37% producing ESBL and 25% were positive for bla CTX-M-15. The observed prevalence of multidrug resistance and colibactin producing characteristics among pathogenic E. coli belonging to phylogenetic group B2 advocate urgent need for broader surveillance in order to understand and prevent transmission of these ExPEC in community and hospital settings.