The authors have declared that no competing interests exist.
Multidrug-resistant (MDR)
Whole-genome sequencing was performed for 60 MDR
Genome comparison revealed the presence of ST-131 global MDR and ST410 as emerging-MDR clades of
This is the first report on the whole genome analysis of MDR
Virulence of
Among MDR
This current study was aimed at identifying the predominant virulent and AMR genes in MDR
A total of 99257 specimens were received at the Department of Clinical Microbiology, Christian Medical College, Vellore, India for routine screening from BSI during the year 2006 to 2016. Isolation and identification of the organism were carried out using a standard protocol as reported earlier [
AST testing was carried out using the Kirby-Bauer disk diffusion method. The antimicrobial agents tested were Amikacin (30 μg), netilmicin (30 μg), gentamycin (10 μg), chloramphenicol (30 μg), ciprofloxacin (5 μg), cefotaxime (30 μg), cefoxitin (30 μg), ceftazidime (30 μg), cefpodoxime (10 μg), piperacilllin-tazobactam (100/10 μg), cefoperazone-sulbactam (75/30), imipenem (10 μg) and meropenem (10 μg), tigecycline (15 μg) and tetracycline (30 μg) according to guidelines suggested by Clinical and Laboratory Standards Institute (CLSI) M100-S27, 2017. Quality control strains (
Colistin MICs for the studied isolates were determined by broth microdilution and interpreted using CLSI 2017 breakpoint recommendations.
Genomic DNA was extracted using a QIAamp DNA Mini Kit (QIAGEN, Hilden, Germany). Whole genome sequencing (WGS) was performed using an Ion Torrent™ Personal Genome Machine™ (PGM) sequencer (Life Technologies, Carlsbad, CA) with 400-bp read chemistry according to the manufacturer’s instructions. Data were assembled with reference
The assembled sequence was annotated using PATRIC, the bacterial bioinformatics database and analysis resource (
Sequence types (STs) were analysed using multi-locus sequence typing (MLST) 1.8 tool (
Gview, interactive genome viewer was used to compare the annotated
This Whole Genome Shotgun project has been deposited at GenBank under the accession numbers as mentioned in
The study was approved by the Institutional Review Board and Ethical committee, Christian Medical College, Vellore, India (IRB No.: 9540 dt 22-07-2015). All the samples were fully anonymized before processing and since our study only utilised isolates received from routine blood cultures, we did not require informed written consent from the patients.
All 60
MLSTFinder revealed the different sequence types of the isolates. The study isolates belonged to 6 clonal complexes with 14 different sequence types. Few of the sequence types were observed to share same founder types revealing the evolution of these strains. CC10 and CC 405 were the two major CCs observed with ST-167, ST-410 and ST-405 as the common STs. Interestingly, nine isolates belonging to CC/ST-131 were identified, of which, all were of H-30 clade, except the isolate BA9313 (H-24).
Whole genome composition of 60 MDR
Comparison between the core and pan genomes of 60 MDR
Serotypes were established from the whole genome data and O102:H6 was the most common serotype (18.75%), followed by O89:H9 (15.6%), O8:H9 (12.5%), O89:H5 (9.4%) and other serotypes (
Three common virulence gene profiles were observed among the isolates as follows, i)
The virulence gene profiles of the 60 isolates were compared to the FimH virulence types, MLST sequence type and SNP phylogeny. The isolates clustered in two distinct groups including ST-131(H-30 clade), based on the virulence genes identified (
Sequence types, virulence gene profile, O and H antigens and Fim-H types are shown next to the tree.
ResFinder revealed the presence of multiple AMR genes in each of the MDR
Depicting prevalence of
The increasing use of third-generation β-lactams and β-lactam inhibitors was accompanied with increases in prevalence of the MDR phenotype among
On genotypic characterization of MDR
The MDR
Globally,
There has been a global concern on aminoglycoside resistance in Gram-negatives. Acquired 16S-RMTases are known to confer extremely high level of aminoglycoside resistance, due to which key aminoglycosides including gentamicin, tobramycin, and amikacin are ineffective against carbapenem resistant strains [
Our study shows that, for cephalosporin resistance, the isolates from ST131 and ST405 clades carried
The antimicrobial susceptibility of
Though ST131 clones were predominantly reported worldwide, the STs observed in our study were striking for clustering in distinct phylogenetic lineages. ST167 was previously reported for its ability to carry
Virulence genes observed among the
FimH, that had been reported as a major candidate for the development of a vaccine against pathogenic
Our study shows that with a SNPs based phylogeny, higher discrimination between the clinical MDR
To the best of our knowledge, this is the first report on SNP phylogeny in comparison with AMR and virulence traits in
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The authors thank the Christian Medical College, Vellore, India for providing basic infrastructure required for this study.