Whole Genome Sequencing detects Inter-Facility Transmission of Carbapenem-resistant Klebsiella pneumoniae

Abstract

Objectives: To identify transmission patterns of Carbapenem-resistant Klebsiella pneumoniae infection during an outbreak at a large, tertiary care hospital and to detect whether the outbreak organisms spread to other facilities in the integrated healthcare network.

Methods: We analyzed 71 K. pneumoniae whole genome sequences collected from clinical specimens before, during and after the outbreak and reviewed corresponding patient medical records. Sequence and patient data were used to model probable transmissions and assess factors associated with the outbreak.

Results: We identified close genetic relationships among carbapenem-resistant K. pneumoniae isolates sampled during the study period. Transmission tree analysis combined with patient records uncovered extended periods of silent colonization in many study patients and transmission routes that were likely the result of asymptomatic patients transitioning between facilities.

Conclusions: Detecting how and where Carbapenem-resistant K. pneumoniae infections spread is challenging in an environment of rising prevalence, asymptomatic carriage and mobility of patients. Whole genome sequencing improved the precision of investigating inter-facility transmissions. Our results emphasize that containment of Carbapenem-resistant K. pneumoniae infections requires coordinated efforts between healthcare networks and settings of care that acknowledge and mitigate transmission risk conferred by undetected carriage and by patient transfers between facilities.

Keywords: Carbapenem; Epidemiology; Healthcare; Infection control; Klebsiella pneumoniae; Whole genome sequencing; transmission.

Fig. 1.. Increasing K. pneumoniae antibiotic resistance and CRKP outbreak.

(A). Heat map showing two-way hierarchical clustering of antibiotic resistance. Colors indicate the percentage of K. pneumoniae cultures collected at Atrium Health acute care hospitals that were resistant to specific antibiotics. Carbapenems are indicated in bold and italics. (B). Plot of CRKP infection rates over 12 months including the Hospital 2 outbreak. Each point represents the CRKP infection rate per 10,000 patient days for each month (M1–M12). Rates include all CRKP infection or colonization identified by clinical culture.

Fig. 2.. Plot identifying all healthcare visits for study patients with CRKP over the 2200-day study period.

Colors indicate the facility where the visit occurred and included 11 hospitals, 3 inpatient facilities and 29 outpatient facilities. Sequenced CRKP + cultures are represented by a black asterisk. Additional unsequenced CRKP + cultures are shown by a black dash.

Fig. 3.. Phylogenetic Tree identifying genomic relationships between CRKP ST258 isolates.

The tree was constructed using the core genome alignment of the 76 CRE genomes included in SNP distance calculation. Only the CRKP ST258 isolates are included in the tree construction to allow visualization of the primary endemic strain. Color gradient shows SNP distance ranges between individual isolate pairs.

Fig. 4.. Analysis of SNP distance relationships to collection time and facility reveals correspondence between genomics and transmissions.

(A). Comparison of collection times and SNP distances between pairs of isolates with 11 or fewer SNPs demonstrated a strong positive correlation (R2 = 0.125; P < 0.0001) despite variation. (B). Comparison of SNP distances between pairs collected during the outbreak period at the same facility (Mean 14.71; 10%ile 12.91, 90%ile 16.52) versus those collected at different facilities (Mean 35.74; 10%ile 33.08, 90%ile 38.41) showed that isolates from the same facility were much more closely related than those from different facilities (P < 0.0001). (C). Comparison of SNP distances between isolate pairs collected during the outbreak period where both were collected at Hospital 2 (Mean 12.22; 10%ile 9.72, 90%ile 14.71) versus isolate pairs where both were collected at the same facility, but not Hospital 2, (Mean 22.20, 10%ile 18.52, 90%ile 25.88) demonstrated that direct or close transmissions (defined by short core genome SNP differences) occurred significantly more frequently within Hospital 2 than at other facilities during the outbreak period.

Fig. 5.. Transmission tree combining genomics and culture dates indicates potential transmission between facilities.

SeqTrack analysis was conducted using culture date and core genome SNP distance to infer a probable transmission tree. Nodes represent sequenced isolates with each isolate ID displayed. Edges are likely transmission paths with the SNP distance between isolates shown in blue. Isolates without edges are >11 SNPs from ancestor isolate predicted by SeqTrack analysis. Node colors indicate the facility where the specimen was collected. Stacks of nodes above each numbered node were added to the tree to show additional CRKP + cultures collected during the study prior to the collection date of the sequenced isolate. Those below the main node are isolates that were collected after. Isolates contained within the red polygon were designated by SeqTrack as part of the primary outbreak at Hospital 2. Isolates determined to be outside the primary outbreak that were collected at Hospital 2 during the outbreak period have a heavy black circle surrounding the node.

Fig. 6.. Analysis of two isolate clusters beyond the outbreak identifies evidence that closely related isolates were shared between facilities.

Study dates of all CRKP + cultures for each patient in the 2 clusters are shown within the nodes with each sequenced culture indicated by an asterisk. The facilities where the cultures were collected are: Red = Rehab, Turquoise = Hospital 5, Gray = Outpatient and Yellow = Hospital 4, consistent with other figures. Edges show genetic connections between isolates with each the SNP distance between isolates displayed. Solid edges display the direct connections and SNP distances on the SeqTrack transmissions tree. Dashed edges show SNP distances and indirect connections on the tree. (A). Isolates 61, 63, 60, and 50 were closely related on the SeqTrack tree (0–3 SNPs), yet were collected at 3 different facilities. (B). Three of the isolates in the second cluster were collected at Hospital 4, as were their other unsequenced CRKP + cultures. Isolate 65 had the closest genetic relationship to 59 but was collected at Rehab.