skip to main content


Title: Using Whole-Genome Sequencing to Improve Surveillance Measures: Case Study of Methicillin-Resistant Staphylococcus aureus (MRSA) in a Florida Hospital
Background: The CDC considers methicillin-resistant Staphylococcus aureus (MRSA) one of the most important hospital-acquired infections (HAIs) in the United States. However, infection control departments (ICDs) often rely on subjective data to determine whether multiple MRSA cases are a true outbreak and whether the hospital is responsible (community- vs hospital-acquired). Objective: Our objective was to determine whether whole-genome sequencing (WGS) of MRSA provided new insights into on transmission dynamics at large, inner-city hospital in Jacksonville, Florida. Methods: Over a 4-month period, MRSA samples were obtained from 44 infected patients at 3 campuses within a single hospital system. Limited nonpatient identifying information was obtained, including date of collection, campus, unit, reason for admission, and days post admission (DPA) of MRSA diagnosis. Whole-genome sequences were generated using the Illumina platform. Raw reads were processed, and genetic distances were calculated and used to identify genetically linked bacterial infections using FoxSeq version 1.0 software. Results: Based on their length of stay, 10 patients were reported by the ICD as obtaining an HAI. Three distinct “episodes” were evident in which >5 MRSA cases were observed within a 3–5-day period. Genomic analysis identified 5 clusters of linked infections: 4 clusters contained 2 patients and another contained 3. Of these clusters, only 1 contained multiple cases that were reported as HAIs; however, because these case were separated by 2 weeks, it is unlikely that they came from a source in the hospital. The results suggest that HAIs were overreported and that most MRSA present in the hospital likely came from community sources. Conclusions: WGS provided clear evidence that temporally clustered MRSA cases do not imply an outbreak is occurring. Furthermore, ongoing detection of the same community-acquired infections over several months is indicative of a shared source outside of the hospital, which could be uncovered through examination of clinical records. Considering the implications of HAIs, best approaches to combat them should include identifying their sources. As molecular surveillance approaches to infection control are rapidly becoming easier and less expensive, the methods can be used to bring objective clarity to the ICD. Funding: None Disclosures: Susanna L. Lamers reports salary from BioInfoExperts and contract research for the NIH, the University of California - San Francisco, and UMASS Medical School.  more » « less
Award ID(s):
1830867
NSF-PAR ID:
10218303
Author(s) / Creator(s):
; ; ; ; ; ;
Date Published:
Journal Name:
Infection Control & Hospital Epidemiology
Volume:
41
Issue:
S1
ISSN:
0899-823X
Page Range / eLocation ID:
s505 to s506
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. null (Ed.)
    Abstract Background VREfm is a major cause of Hospital Acquired Infection in the United States. We analyzed all the VREfm infections that occurred in our institution between 2018 and 2019 using Whole Genome Sequencing (WGS) to understand epidemiological relationship between previously unidentified clusters. In this study we describe a cluster in our hematology oncology unit. Methods A total of 109 discrete VREfm isolates from 66 patients were analyzed. VREfm isolates used in this study were identified from positive blood and urine cultures. Genomic deoxyribonucleic acid (DNA) was extracted from pure cultures. The purity and integrity of extracted DNA were determined using appropriate assays. Library construction and sequencing were conducted and Multi Locust Sequence Typing (MLST) obtained (image 1). Phylogenomic tree was plotted using the Interactive Tree of Life (image 2). Image 1 - methods Image 2 - Tree of Life Results Total of 7 clusters were identified. Here we describe one cluster (image 3) with the highest genetic similarity which showed maximum difference of 5 Single Nucleotide Polymorphisms (zero between patient 1 and 2, image 4). The cluster is composed of 24 clinical strains of VREfm from 6 patients, over a 9 month time period (Image 5). All patients had hematologic malignancies; 4/6 patients had received recent chemotherapy and 5/6 patients were neutropenic. 4 patients were admitted in a single unit (labelled E7), 1 patient was on a sister unit (labelled F7); and 1 patient was in the cancer infusion center. All patients had central venous access placed by radiology at the time of diagnosis of infection and had visited our outpatient infusion center multiple times during this time frame. Image 3 - Close look at cluster 1 Image 4 - Dendrogram of 106 isolates performed with coreSNP(Single Nucleotide Polymorphisms) pairwise distances. • Dendogram shows different patients (same color for isolates that belong to the same patient) and the patient numbers. • Besides the patient number, the number of largest number SNPS that separate those isolates is shown. • Branches represent the number of coreSNPs that differ strains from that branch. As you see isolates from cluster 1 differ in a maximum of 5 SNPs but isolates of patient 1 and patient 2 differ in 0 SNPs between them. Cluster 1 is represented by a green square. Image 5 - Time period of infections Conclusion The prolonged period in our cluster argues in favor of an environmental niche in the hospital unit. We are unable to elucidate pattern of transmission in a cluster of infections without knowing patient colonization of VREfm; because we are likely looking at the tip of the iceberg when analyzing infected cases. It is difficult to ascribe causality to any one of these exposures without concomitant surveillance cultures of environment and personnel. Retrospective WGS is of limited value in infection control. We now have third generation sequencing with the MinION device to do real time sequencing with which we also validated some of our samples. Disclosures Atul Kothari, MD, Ansun Biopharma (Consultant) 
    more » « less
  2. null (Ed.)
    Background: In spring of 2019, 2 positive sputum cases of Pseudomonas aeruginosa in the cardiac critical care unit (CCU) were reported to the UFHJ infection prevention (IP) department. The initial 2 cases, detected within 3 days of each other, were followed shortly by a third case. Epidemiological evidence was initially consistent with a hospital-acquired infection (HAI): 2 of the 3 patients roomed next to each other, and all 3 patients were ventilated, 2 of whom shared the same respiratory therapist. However, no other changes in routine or equipment were noted. The samples were cultured and processed using Illumina NGS technology, generating 1–2 million short (ie, 250-bp) reads across the P. aeruginosa genome. As an additional positive control, 8 P . aeruginosa NGS data sets, previously shown to be from a single outbreak in a UK facility, were included. Reads were mapped back to a reference sequence, and single-nucleotide polymorphisms (SNPs) between each sample and the reference were extracted. Genetic distances (ie, the number of unshared SNPs) between all UFHJ and UK samples were calculated. Genetic linkage was determined using hierarchical clustering, based on a commonly used threshold of 40 SNPs. All UFHJ patient samples were separated by >18,000 SNPs, indicating genetically distinct samples from separate sources. In contrast, UK samples were separated from each other by <16 SNPs, consistent with genetic linkage and a single outbreak. Furthermore, the UFHJ samples were separated from the UK samples by >17,000 SNPs, indicating a lack of geographical distinction of the UFHJ samples (Fig. 1). These results demonstrated that while the initial epidemiological evidence pointed towards a single HAI, the high-precision and relatively inexpensive ( more » « less
  3. Background: Carbapenem-resistant Enterobacteriaceae (CRE) are a global threat. Here, we describe the clinical and molecular characteristics of Centers for Disease Control and Prevention (CDC)-defined CRE in the US. Methods: The second Consortium on Resistance Against Carbapenems in Klebsiella and other Enterobacteriaceae (CRACKLE-2, ClinicalTrials.gov: NCT03646227) is a prospective, multicenter, cohort study. Patients hospitalized in 49 US hospitals, with clinical cultures positive for CDC-defined CRE between 30 April 2016 and 31 August 2017 were included. Primary outcome was desirability of outcome ranking (DOOR) at 30 days. Clinical data and bacteria were collected, and whole genome sequencing (WGS) was performed. Findings: 1,040 patients with unique isolates were included; 449 (43%) with infection and 591 (57%) with colonization. CDC-defined CRE admission rate was 57 CDC-defined CRE admissions/100,000 admissions (95% CI: 45–71). Three subsets of CDC-defined CRE were identified: carbapenemase-producing Enterobacteriaceae (618/1,040, 59%); non-carbapenemase-producing CRE (194/1,040, 19%); and unconfirmed CRE (228/1,040, 22%; initially reported as CRE, but susceptible to carbapenems in two central laboratories). Klebsiella pneumoniae carbapenemase (KPC)-producing clonal group 258 K. pneumoniae was the most common carbapenemase-producing Enterobacteriaceae. In 449 patients with CDC-defined CRE infections, DOOR outcomes were not significantly different in patients with carbapenemase-producing Enterobacteriaceae, non-carbapenemase-producing CRE, and unconfirmed CRE. At 30 days 107/449 (24%, 95% CI 20–28%) patients had died. Interpretation: Among patients with CDC-defined CRE, similar outcomes were observed among three subgroups, including the novel unconfirmed CRE group. CDC-defined CRE represent diverse bacteria, whose spread may not respond to interventions directed to carbapenemase-producing Enterobacteriaceae. 
    more » « less
  4. Abstract Background

    Four severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants predominated in the United States since 2021. Understanding disease severity related to different SARS-CoV-2 variants remains limited.

    Method

    Viral genome analysis was performed on SARS-CoV-2 clinical isolates circulating March 2021 through March 2022 in Cleveland, Ohio. Major variants were correlated with disease severity and patient outcomes.

    Results

    In total 2779 patients identified with either Alpha (n = 1153), Gamma (n = 122), Delta (n = 808), or Omicron variants (n = 696) were selected for analysis. No difference in frequency of hospitalization, intensive care unit (ICU) admission, and death were found among Alpha, Gamma, and Delta variants. However, patients with Omicron infection were significantly less likely to be admitted to the hospital, require oxygen, or admission to the ICU (χ2 = 12.8, P < .001; χ2 = 21.6, P < .002; χ2 = 9.6, P = .01, respectively). In patients whose vaccination status was known, a substantial number had breakthrough infections with Delta or Omicron variants (218/808 [26.9%] and 513/696 [73.7%], respectively). In breakthrough infections, hospitalization rate was similar regardless of variant by multivariate analysis. No difference in disease severity was identified between Omicron subvariants BA.1 and BA.2.

    Conclusions

    Disease severity associated with Alpha, Gamma, and Delta variants is comparable while Omicron infections are significantly less severe. Breakthrough disease is significantly more common in patients with Omicron infection.

     
    more » « less
  5. Abd El-Aty, A. M. (Ed.)
    Background Higher viral loads in SARS-CoV-2 infections may be linked to more rapid spread of emerging variants of concern (VOC). Rapid detection and isolation of cases with highest viral loads, even in pre- or asymptomatic individuals, is essential for the mitigation of community outbreaks. Methods and findings In this study, we analyze Ct values from 1297 SARS-CoV-2 positive patient saliva samples collected at the Clemson University testing lab in upstate South Carolina. Samples were identified as positive using RT-qPCR, and clade information was determined via whole genome sequencing at nearby commercial labs. We also obtained patient-reported information on symptoms and exposures at the time of testing. The lowest Ct values were observed among those infected with Delta (median: 22.61, IQR: 16.72–28.51), followed by Alpha (23.93, 18.36–28.49), Gamma (24.74, 18.84–30.64), and the more historic clade 20G (25.21, 20.50–29.916). There was a statistically significant difference in Ct value between Delta and all other clades (all p.adj<0.01), as well as between Alpha and 20G (p.adj<0.05). Additionally, pre- or asymptomatic patients (n = 1093) showed the same statistical differences between Delta and all other clades (all p.adj<0.01); however, symptomatic patients (n = 167) did not show any significant differences between clades. Our weekly testing strategy ensures that cases are caught earlier in the infection cycle, often before symptoms are present, reducing this sample size in our population. Conclusions COVID-19 variants Alpha and Delta have substantially higher viral loads in saliva compared to more historic clades. This trend is especially observed in individuals who are pre- or asymptomatic, which provides evidence supporting higher transmissibility and more rapid spread of emerging variants. Understanding the viral load of variants spreading within a community can inform public policy and clinical decision making. 
    more » « less