Utility of Whole-Genome Sequencing in Characterizing Acinetobacter Epidemiology and Analyzing Hospital Outbreaks

Acinetobacter baumanniifrequently causes nosocomial infections and outbreaks. Whole-genome sequencing (WGS) is a promising technique for strain typing and outbreak investigations. We compared the performance of conventional methods with WGS for strain typing clinicalAcinetobacterisolates and analyzing a carbapenem-resistantA. baumannii(CRAB) outbreak. We performed two band-based typing techniques (pulsed-field gel electrophoresis and repetitive extragenic palindromic-PCR), multilocus sequence type (MLST) analysis, and WGS on 148Acinetobacter calcoaceticus-A. baumanniicomplex bloodstream isolates collected from a single hospital from 2005 to 2012. Phylogenetic trees inferred from core-genome single nucleotide polymorphisms (SNPs) confirmed threeAcinetobacterspecies within this collection. Four majorA. baumanniiclonal lineages (as defined by MLST) circulated during the study, three of which are globally distributed and one of which is novel. WGS indicated that a threshold of 2,500 core SNPs accurately distinguishedA. baumanniiisolates from different clonal lineages. The band-based techniques performed poorly in assigning isolates to clonal lineages and exhibited little agreement with sequence-based techniques. After applying WGS to a CRAB outbreak that occurred during the study, we identified a threshold of 2.5 core SNPs that distinguished nonoutbreak from outbreak strains. WGS was more discriminatory than the band-based techniques and was used to construct a more accurate transmission map that resolved many of the plausible transmission routes suggested by epidemiologic links. Our study demonstrates that WGS is superior to conventional techniques forA. baumanniistrain typing and outbreak analysis. These findings support the incorporation of WGS into health care infection prevention efforts.

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Genomically Informed Surveillance for Carbapenem-Resistant Enterobacteriaceae in a Health Care System

mBio ◽

10.1128/mbio.01030-15 ◽

2015 ◽

Vol 6 (4) ◽

Cited By ~ 60

Author(s):

Nicole D. Pecora ◽

Ning Li ◽

Marc Allard ◽

Cong Li ◽

Esperanza Albano ◽

...

Keyword(s):

Health Care ◽

Drug Resistance ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Resistance Genes ◽

Enterobacter Cloacae ◽

Mobile Elements ◽

Whole Genome ◽

Content Type ◽

Carbapenem Resistant

ABSTRACT Carbapenem-resistant Enterobacteriaceae (CRE) are an urgent public health concern. Rapid identification of the resistance genes, their mobilization capacity, and strains carrying them is essential to direct hospital resources to prevent spread and improve patient outcomes. Whole-genome sequencing allows refined tracking of both chromosomal traits and associated mobile genetic elements that harbor resistance genes. To enhance surveillance of CREs, clinical isolates with phenotypic resistance to carbapenem antibiotics underwent whole-genome sequencing. Analysis of 41 isolates of Klebsiella pneumoniae and Enterobacter cloacae, collected over a 3-year period, identified K. pneumoniae carbapenemase (KPC) genes encoding KPC-2, −3, and −4 and OXA-48 carbapenemases. All occurred within transposons, including multiple Tn4401 transposon isoforms, embedded within more than 10 distinct plasmids representing incompatibility (Inc) groups IncR, -N, -A/C, -H, and -X. Using short-read sequencing, draft maps were generated of new KPC-carrying vectors, several of which were derivatives of the IncN plasmid pBK31551. Two strains also had Tn4401 chromosomal insertions. Integrated analyses of plasmid profiles and chromosomal single-nucleotide polymorphism (SNP) profiles refined the strain patterns and provided a baseline hospital mobilome to facilitate analysis of new isolates. When incorporated with patient epidemiological data, the findings identified limited outbreaks against a broader 3-year period of sporadic external entry of many different strains and resistance vectors into the hospital. These findings highlight the utility of genomic analyses in internal and external surveillance efforts to stem the transmission of drug-resistant strains within and across health care institutions. IMPORTANCE We demonstrate how detection of resistance genes within mobile elements and resistance-carrying strains furthers active surveillance efforts for drug resistance. Whole-genome sequencing is increasingly available in hospital laboratories and provides a powerful and nuanced means to define the local landscape of drug resistance. In this study, isolates of Klebsiella pneumoniae and Enterobacter cloacae with resistance to carbapenem antibiotics were sequenced. Multiple carbapenemase genes were identified that resided in distinct transposons and plasmids. This mobilome, or population of mobile elements capable of mobilizing drug resistance, further highlighted the degree of strain heterogeneity while providing a detailed timeline of carbapenemase entry into the hospital over a 3-year period. These surveillance efforts support effective targeting of infection control resources and the development of institution-specific repositories of resistance genes and the mobile elements that carry them.

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MCR-1 and OXA-48 In Vivo Acquisition in KPC-Producing Escherichia coli after Colistin Treatment

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02540-16 ◽

2017 ◽

Vol 61 (8) ◽

Cited By ~ 32

Author(s):

Racha Beyrouthy ◽

Frederic Robin ◽

Aude Lessene ◽

Igor Lacombat ◽

Laurent Dortet ◽

...

Keyword(s):

Escherichia Coli ◽

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Whole Genome ◽

Content Type ◽

First Report ◽

E Coli ◽

Carbapenem Resistant

ABSTRACT The spread of mcr-1-encoding plasmids into carbapenem-resistant Enterobacteriaceae raises concerns about the emergence of untreatable bacteria. We report the acquisition of mcr-1 in a carbapenem-resistant Escherichia coli strain after a 3-week course of colistin in a patient repatriated to France from Portugal. Whole-genome sequencing revealed that the Klebsiella pneumoniae carbapenemase-producing E. coli strain acquired two plasmids, an IncL OXA-48-encoding plasmid and an IncX4 mcr-1-encoding plasmid. This is the first report of mcr-1 in carbapenemase-encoding bacteria in France.

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Loss and Gain in the Evolution of theSalmonella entericaSerovar Gallinarum Biovar Pullorum Genome

mSphere ◽

10.1128/msphere.00627-18 ◽

2019 ◽

Vol 4 (2) ◽

Cited By ~ 6

Author(s):

Yachen Hu ◽

Zhenyu Wang ◽

Bin Qiang ◽

Yaohui Xu ◽

Xiang Chen ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Genomic Analysis ◽

Developed Countries ◽

Epidemiological Surveillance ◽

Recent Common Ancestor ◽

Whole Genome ◽

Nucleotide Polymorphisms ◽

Content Type ◽

Most Recent Common Ancestor

ABSTRACTSalmonella entericasubspeciesentericaserovar Gallinarum biovar Pullorum (S. Pullorum) is the etiological agent of pullorum disease, causing white diarrhea with high mortality in chickens. There are many unsolved issues surrounding the epidemiology ofS. Pullorum, including its origin and transmission history as well as the discordance between its phenotypic heterogeneity and genetic monomorphism. In this paper, we report the results of whole-genome sequencing of a panel of 97S. Pullorum strains isolated between 1962 and 2014 from four countries across three continents. We utilized 6,795 core genome single nucleotide polymorphisms (SNPs) to reconstruct a phylogenetic tree within a spatiotemporal Bayesian framework, estimating that the most recent common ancestor ofS. Pullorum emerged in ∼914 CE (95% confidence interval [95%CI], 565 to 1273 CE). The extantS. Pullorum strains can be divided into four distinct lineages, each of which is significantly associated with geographical distribution. The intercontinental transmissions of lineages III and IV can be traced to the mid-19th century and are probably related to the “Hen Fever” prevalent at that time. Further genomic analysis indicated that the loss or pseudogenization of functional genes involved in metabolism and virulence inS. Pullorum has been ongoing since before and after divergence from the ancestor. In contrast, multiple prophages and plasmids have been acquired byS. Pullorum, and these have endowed it with new characteristics, especially the multidrug resistance conferred by two large plasmids in lineage I. The results of this study provide insight into the evolution ofS. Pullorum and prove the efficiency of whole-genome sequencing in epidemiological surveillance of pullorum disease.IMPORTANCEPullorum disease, an acute poultry septicemia caused bySalmonellaGallinarum biovar Pullorum, is fatal for young chickens and is a heavy burden on poultry industry. The pathogen is rare in most developed countries but still extremely difficult to eliminate in China. Efficient epidemiological surveillance necessitates clarifying the origin of the isolates from different regions and their phylogenic relationships. Genomic epidemiological analysis of 97S. Pullorum strains was carried out to reconstruct the phylogeny and transmission history ofS. Pullorum. Further analysis demonstrated that functional gene loss and acquisition occurred simultaneously throughout the evolution ofS. Pullorum, both of which reflected adaptation to the changing environment. The result of our study will be helpful in surveillance and prevention of pullorum disease.

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Salmonella entericaPhylogeny Based on Whole-Genome Sequencing Reveals Two New Clades and Novel Patterns of Horizontally Acquired Genetic Elements

mBio ◽

10.1128/mbio.02303-18 ◽

2018 ◽

Vol 9 (6) ◽

Cited By ~ 24

Author(s):

Jay Worley ◽

Jianghong Meng ◽

Marc W. Allard ◽

Eric W. Brown ◽

Ruth E. Timme

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Foodborne Pathogens ◽

Bacterial Species ◽

Whole Genome Sequence ◽

Nucleotide Sequencing ◽

Whole Genome ◽

Nucleotide Polymorphisms ◽

Content Type ◽

Genetic Elements

ABSTRACTUsing whole-genome sequence (WGS) data from the GenomeTrakr network, a globally distributed network of laboratories sequencing foodborne pathogens, we present a new phylogeny ofSalmonella entericacomprising 445 isolates from 266 distinct serovars and originating from 52 countries. This phylogeny includes two previously unidentifiedS. entericasubsp.entericaclades. Serovar Typhi is shown to be nested within clade A. Our findings are supported by both phylogenetic support, based on a core genome alignment, and Bayesian approaches, based on single-nucleotide polymorphisms. Serovar assignments were refined byin silicoanalysis using SeqSero. More than 10% of serovars were either polyphyletic or paraphyletic. We found variable genetic content in these isolates relating to gene mobilization and virulence factors which have different distributions within clades. Gifsy-1- and Gifsy-2-like phages appear more prevalent in clade A; other viruses are more evenly distributed. Our analyses reveal IncFII is the predominant plasmid replicon inS. enterica. Few core or clade-defining virulence genes are observed, and their distributions appear probabilistic in nature. Together, these patterns demonstrate that genetic exchange withinS. entericais more extensive and frequent than previously realized, which significantly alters how we view the genetic structure of the bacterial species.IMPORTANCERapid improvements in nucleotide sequencing access and affordability have led to a drastic increase in availability of genetic information. This information will improve the accuracy of molecular descriptions, including serovars, withinS. enterica. Although the concept of serovars continues to be useful, it may have more significant limitations than previously understood. Furthermore, the discrete absence or presence of specific genes can be an unstable indicator of phylogenetic identity. Whole-genome sequencing provides more rigorous tools for assessing the distributions of these genes. Our phylogenetic and genetic content analyses reveal how active genetic elements are dynamically distributed within a species, allowing us to better understand genetic reservoirs and underlying bacterial evolution.

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Whole-Genome Sequencing for Characterization of Capsule Locus and Prediction of Serogroup of Invasive Meningococcal Isolates

Journal of Clinical Microbiology ◽

10.1128/jcm.01609-18 ◽

2018 ◽

Vol 57 (3) ◽

Cited By ~ 3

Author(s):

Henju Marjuki ◽

Nadav Topaz ◽

Lorraine D. Rodriguez-Rivera ◽

Edward Ramos ◽

Caelin C. Potts ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Whole Genome ◽

Phase Variable ◽

Rt Pcr ◽

Content Type ◽

Strain Collection ◽

Capsule Locus ◽

Outbreak Investigations

ABSTRACTInvasive meningococcal disease is mainly caused byNeisseria meningitidisserogroups A, B, C, X, W, and Y. The serogroup is typically determined by slide agglutination serogrouping (SASG) and real-time PCR (RT-PCR). We describe a whole-genome sequencing (WGS)-based method to characterize the capsule polysaccharide synthesis (cps) locus, classifyN. meningitidisserogroups, and identify mechanisms for nongroupability using 453 isolates from a global strain collection. We identified novel genomic organizations within functionalcpsloci, consisting of insertion sequence (IS) elements in unique positions that did not disrupt the coding sequence. Genetic mutations (partial gene deletion, missing genes, IS insertion, internal stop, and phase-variable off) that led to nongroupability were identified. The results of WGS and SASG were in 91% to 100% agreement for all serogroups, while the results of WGS and RT-PCR showed 99% to 100% agreement. Among isolates determined to be nongroupable by WGS (31 of 453), the results of all three methods agreed 100% for those without a capsule polymerase gene. However, 61% (WGS versus SASG) and 36% (WGS versus RT-PCR) agreements were observed for the isolates, particularly those with phase variations or internal stops incpsloci, which warrant further characterization by additional tests. Our WGS-based serogrouping method provides comprehensive characterization of theN. meningitidiscapsule, which is critical for meningococcal surveillance and outbreak investigations.

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Characterization of an Enterobacter cloacae Strain Producing both KPC and NDM Carbapenemases by Whole-Genome Sequencing

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01275-15 ◽

2015 ◽

Vol 59 (10) ◽

pp. 6625-6628 ◽

Cited By ~ 27

Author(s):

Wenjing Wu ◽

Yu Feng ◽

Alessandra Carattoli ◽

Zhiyong Zong

Keyword(s):

Whole Genome Sequencing ◽

Bloodstream Infection ◽

Genome Sequencing ◽

Carbapenem Resistance ◽

Enterobacter Cloacae ◽

Whole Genome ◽

Content Type ◽

Carbapenem Resistant

ABSTRACTA carbapenem-resistantEnterobacter cloacaestrain, WCHECl-14653, causing a fatal bloodstream infection, was characterized by genome sequencing and conjugation experiments. The strain carried two carbapenemase genes,blaNDM-1andblaKPC-2, on separate IncF plasmids. The coexistence ofblaNDM-1andblaKPC-2conferred slightly higher-level carbapenem resistance compared with that ofblaNDM-1orblaKPC-2alone, and the coexistence of two IncF plasmids may generate new platforms for spreading carbapenemase genes.

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Whole-Genome Sequencing To Identify Drivers of Carbapenem-Resistant Klebsiella pneumoniae Transmission within and between Regional Long-Term Acute-Care Hospitals

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01622-19 ◽

2019 ◽

Vol 63 (11) ◽

Cited By ~ 3

Author(s):

Jennifer H. Han ◽

Zena Lapp ◽

Frederic Bushman ◽

Ebbing Lautenbach ◽

Ellie J. C. Goldstein ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Treatment Options ◽

Health Care Facilities ◽

Resistant Organism ◽

Whole Genome ◽

Content Type ◽

Transmission Rates ◽

Carbapenem Resistant

ABSTRACT Carbapenem-resistant Klebsiella pneumoniae (CRKP) is an antibiotic resistance threat of the highest priority. Given the limited treatment options for this multidrug-resistant organism (MDRO), there is an urgent need for targeted strategies to prevent transmission. Here, we applied whole-genome sequencing to a comprehensive collection of clinical isolates to reconstruct regional transmission pathways and analyzed this transmission network in the context of statewide patient transfer data and patient-level clinical data to identify drivers of regional transmission. We found that high regional CRKP burdens were due to a small number of regional introductions, with subsequent regional proliferation occurring via patient transfers among health care facilities. While CRKP was predicted to have been imported into each facility multiple times, there was substantial variation in the ratio of intrafacility transmission events per importation, indicating that amplification occurs unevenly across regional facilities. While myriad factors likely influence intrafacility transmission rates, an understudied one is the potential for clinical characteristics of colonized and infected patients to influence their propensity for transmission. Supporting the contribution of high-risk patients to elevated transmission rates, we observed that patients colonized and infected with CRKP in high-transmission facilities had higher rates of carbapenem use, malnutrition, and dialysis and were older. This report highlights the potential for regional infection prevention efforts that are grounded in genomic epidemiology to identify the patients and facilities that make the greatest contribution to regional MDRO prevalence, thereby facilitating the design of precision interventions of maximal impact.

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Whole-Genome Sequencing in Epidemiology of Campylobacter jejuni Infections

Journal of Clinical Microbiology ◽

10.1128/jcm.00017-17 ◽

2017 ◽

Vol 55 (5) ◽

pp. 1269-1275 ◽

Cited By ~ 35

Author(s):

Ann-Katrin Llarena ◽

Eduardo Taboada ◽

Mirko Rossi

Keyword(s):

Whole Genome Sequencing ◽

Campylobacter Jejuni ◽

Genome Sequencing ◽

Outbreak Detection ◽

Whole Genome ◽

Ample Evidence ◽

Content Type ◽

Routine Monitoring ◽

Outbreak Investigations ◽

Bacterial Gastroenteritis

ABSTRACT This review describes the current state of knowledge regarding the application of whole-genome sequencing (WGS) in the epidemiology of Campylobacter jejuni , the leading cause of bacterial gastroenteritis worldwide. We describe how WGS has increased our understanding of the evolutionary and epidemiological dynamics of this pathogen and how WGS has the potential to improve surveillance and outbreak detection. We have identified hurdles to the full implementation of WGS in public health settings. Despite these challenges, we think that ample evidence is available to support the benefits of integrating WGS into the routine monitoring of C. jejuni infections and outbreak investigations.

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mSphere of Influence: Whole-Genome Sequencing, a Vital Tool for the Interruption of Nosocomial Transmission

mSphere ◽

10.1128/msphere.00230-21 ◽

2021 ◽

Vol 6 (2) ◽

Author(s):

Ahmed Babiker

Keyword(s):

Health Care ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Multidrug Resistant ◽

Nosocomial Transmission ◽

Whole Genome ◽

Content Type ◽

Genomic Epidemiology ◽

Hospital Outbreak ◽

Carbapenem Resistant

ABSTRACT Ahmed Babiker’s work focuses on the clinical and genomic epidemiology of multidrug-resistant health care-associated pathogens and other high-consequence pathogens. In this mSphere of Influence article, he reflects on how the paper “Tracking a Hospital Outbreak of Carbapenem-Resistant Klebsiella pneumoniae with Whole-Genome Sequencing” by Evan S. Snitkin et al. (Sci Transl Med 4:148ra116, 2012, https://doi.org/10.1126/scitranslmed.3004129) impacted his thinking on the use of whole-genome sequencing for nosocomial transmission investigation.

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Population Dynamics of Staphylococcus aureus in Cystic Fibrosis Patients To Determine Transmission Events by Use of Whole-Genome Sequencing

Journal of Clinical Microbiology ◽

10.1128/jcm.00164-17 ◽

2017 ◽

Vol 55 (7) ◽

pp. 2143-2152 ◽

Cited By ~ 13

Author(s):

Andrea Ankrum ◽

Barry G. Hall

Keyword(s):

Cystic Fibrosis ◽

Staphylococcus Aureus ◽

Population Dynamics ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Phylogenetic Trees ◽

Hospital Environment ◽

Snp Analysis ◽

Whole Genome ◽

Content Type

ABSTRACT Strict infection control practices have been implemented for health care visits by cystic fibrosis (CF) patients in an attempt to prevent transmission of important pathogens. This study used whole-genome sequencing (WGS) to determine strain relatedness and assess population dynamics of Staphylococcus aureus isolates from a cohort of CF patients as assessed by strain relatedness. A total of 311 S. aureus isolates were collected from respiratory cultures of 115 CF patients during a 22-month study period. Whole-genome sequencing was performed, and using single nucleotide polymorphism (SNP) analysis, phylogenetic trees were assembled to determine relatedness between isolates. Methicillin-resistant Staphylococcus aureus (MRSA) phenotypes were predicted using PPFS2 and compared to the observed phenotype. The accumulation of SNPs in multiple isolates obtained over time from the same patient was examined to determine if a genomic molecular clock could be calculated. Pairs of isolates with ≤71 SNP differences were considered to be the “same” strain. All of the “same” strain isolates were either from the same patient or siblings pairs. There were 47 examples of patients being superinfected with an unrelated strain. The predicted MRSA phenotype was accurate in all but three isolates. Mutation rates were unable to be determined because the branching order in the phylogenetic tree was inconsistent with the order of isolation. The observation that transmissions were identified between sibling patients shows that WGS is an effective tool for determining transmission between patients. The observation that transmission only occurred between siblings suggests that Staphylococcus aureus acquisition in our CF population occurred outside the hospital environment and indicates that current infection prevention efforts appear effective.

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