Diversity, functional classification and genotyping of SHV β-lactamases in <i>Klebsiella pneumoniae</i>
Bibliographic record
Abstract
Abstract Interpreting phenotypes of bla SHV alleles in Klebsiella pneumoniae genomes is complex. While all strains are expected to carry a chromosomal copy conferring resistance to ampicillin, they may also carry mutations in chromosomal bla SHV alleles or additional plasmid-borne bla SHV alleles that have extended-spectrum β-lactamase (ESBL) activity and/or β-lactamase inhibitor (BLI) resistance activity. In addition, the role of individual mutations/amino acid changes is not completely documented or understood. This has led to confusion in the literature and in antimicrobial resistance (AMR) gene databases (e.g., NCBI’s Reference Gene Catalog and the β-lactamase database (BLDB)) over the specific functionality of individual SHV protein variants. Therefore, identification of ESBL-producing strains from K. pneumoniae genome data is complicated. Here, we reviewed the experimental evidence for the expansion of SHV enzyme function associated with specific amino-acid substitutions. We then systematically assigned SHV alleles to functional classes (wildtype, ESBL, BLI-resistant) based on the presence of these mutations. This resulted in the re-classification of 37 SHV alleles compared with current assignments in NCBI’s Reference Gene Catalog and/or BLDB (21 to wildtype, 12 to ESBL, 4 to BLI-resistant). Phylogenetic and comparative genomic analyses support that; i) SHV-1 (encoded by bla SHV-1 ) is the ancestral chromosomal variant; ii) ESBL and BLI-resistant variants have evolved multiple times through parallel substitution mutations; iii) ESBL variants are mostly mobilised to plasmids; iv) BLI-resistant variants mostly result from mutations in chromosomal bla SHV . We used matched genome-phenotype data from the KlebNET-GSP Genotype-Phenotype Group to identify 3,999 K. pneumoniae isolates carrying one or more bla SHV alleles but no other acquired β-lactamases, with which we assessed genotype-phenotype relationships for bla SHV . This collection includes human, animal, and environmental isolates collected between 2001 to 2021 from 24 countries across six continents. Our analysis supports that mutations at Ambler sites 238 and 179 confer ESBL activity, while most omega-loop substitutions do not. Our data also provide direct support for wildtype assignment of 67 protein variants, including eight that were noted in public databases as ESBL. We reclassified these eight variants as wildtype, because they lack ESBL-associated mutations, and our phenotype data support susceptibility to 3GCs (SHV-27, SHV-38, SHV-40, SHV-41, SHV-42, SHV-65, SHV-164, SHV-187). The approach and results outlined here have been implemented in Kleborate v2.4.1 (a software tool for genotyping K. pneumoniae from genome assemblies), whereby known and novel bla SHV alleles are classified based on causative mutations. Kleborate v2.4.1 was also updated to include ten novel protein variants from the KlebNET-GSP dataset and all alleles in public databases as of November 2023. This study demonstrates the power of sharing AMR phenotypes alongside genome data to improve understanding of resistance mechanisms. Impact statement Since every K. pneumoniae genome has an intrinsic SHV β-lactamase and may also carry additional mobile forms, the correct interpretation of bla SHV genes detected in genome data can be challenging and can lead to K. pneumoniae being misclassified as ESBL-producing. Here, we use matched K. pneumoniae genome and drug susceptibility data contributed from dozens of studies, together with systematic literature review of experimental evidence, to improve our understanding of bla SHV allele variation and mapping of genotype to phenotype. This study shows the value of coordinated data sharing, in this case via the KlebNET-GSP Genotype-Phenotype Group, to improve our understanding of the evolutionary history and functionality of bla SHV genes. The results are captured in an open-source AMR dictionary utilised by the Kleborate genotyping tool, that could easily be incorporated into or used to update other tools and AMR gene databases. This work is part of the wider efforts of the KlebNET-GSP group to develop and support a unified platform tailored for the analysis and interpretation of K. pneumoniae genomes by a wide range of stakeholders. Data summary Bla SHV allele sequences and class assignments are distributed with Kleborate, v2.4.1, DOI:10.5281/zenodo.10469001. Table S1 provides a summary of bla SHV alleles, including primary accessions, class-modifying mutations, and supporting evidence for class assignments that differ from NCBI’s Reference Gene Catalog or BLDB. Whole genome sequence data are publicly available as reads and/or assemblies, individual accessions are given in Table S2 ; corresponding genotypes and antibiotic susceptibility phenotypes and measurements are available in Tables S3 and S4 , respectively.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.002 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.002 | 0.002 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.001 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".