Whole-genome sequencing to determine transmission of Neisseria gonorrhoeae: an observational study  Dilrini De Silva, PhD, Joanna Peters, MBBS, Kevin.

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Whole-genome sequencing to determine transmission of Neisseria gonorrhoeae: an observational study  Dilrini De Silva, PhD, Joanna Peters, MBBS, Kevin Cole, BSc, Michelle J Cole, DBMS, Fiona Cresswell, MBChB, Gillian Dean, MBChB, Jayshree Dave, MD, Daniel Rh Thomas, Kirsty Foster, MSc, Alison Waldram, PhD, Daniel J Wilson, DPhil, Xavier Didelot, DPhil, Yonatan H Grad, MD, Prof Derrick W Crook, FRCPath, Prof Tim E A Peto, FRCP, Prof A Sarah Walker, PhD, John Paul, MD, Dr David W Eyre, DPhil  The Lancet Infectious Diseases  Volume 16, Issue 11, Pages 1295-1303 (November 2016) DOI: 10.1016/S1473-3099(16)30157-8 Copyright © 2016 Elsevier Ltd Terms and Conditions

Figure 1 Transmission calibration sampling frames showing SNPs in Neisseria gonorrhoeae between colony picks from the same clinical sample (A), isolates from different anatomical sites within the same patient (B), isolates from contact pairs (C), isolates from the same patient over time (D), and first isolates from different patients in Brighton (E) In (A) six clinical samples were randomly chosen, and 12–14 colonies were sequenced independently. Within each clinical sample, sequences from the first colony chosen were compared with all other colonies sequenced. On the right-hand side, each colour represents a different clinical sample. The area of the circles is proportional to the number of colonies with identical genome sequences. Lines between circles represent the numbers of SNPs between colonies. In five samples all sequences were identical, which is shown as a single circle. In (E), all first samples from each infection in each patient were compared pairwise. 4929 pairs were related by 0–10 SNPs, which is too small a number to be accurately plotted given the scale. SNPs=single nucleotide polymorphisms. The Lancet Infectious Diseases 2016 16, 1295-1303DOI: (10.1016/S1473-3099(16)30157-8) Copyright © 2016 Elsevier Ltd Terms and Conditions

Figure 2 Transmission nomogram of expected SNPs between Neisseria gonnorrhoeae samples over years (A) and within a year (B) SNPs expected between direct or indirect transmission pairs for varying time between samples are shaded (99% prediction interval). (A) shows expected numbers of SNPs over the longest interval possible between samples in the study. Of 1061 distinct infections, only two (0·2%) had a potential source with lower than the expected number of SNPs—no SNPs after 466 days, and one SNP after 686 days. SNPs=single nucleotide polymorphisms. The Lancet Infectious Diseases 2016 16, 1295-1303DOI: (10.1016/S1473-3099(16)30157-8) Copyright © 2016 Elsevier Ltd Terms and Conditions

Figure 3 Proportion of Brighton gonorrhoea infections genetically linked to another sampled case For the Brighton versus Brighton comparison, we compared cases in Brighton (2011–15) with all previous Brighton cases (from 2004 onwards). To avoid double counting of cases, cases were only compared with previous cases, accepting that sampling dates might not indicate the direction of transmission. In the Brighton versus UK and Brighton versus USA plots, we compared all cases from Brighton (2004–15) to all cases from the rest of the UK or USA respectively, independent of the order of sampling. The Lancet Infectious Diseases 2016 16, 1295-1303DOI: (10.1016/S1473-3099(16)30157-8) Copyright © 2016 Elsevier Ltd Terms and Conditions

Figure 4 Brighton clusters of genetically linked cases of gonorrhoea from January, 2011, to March, 2015 (A); and when sampling of consecutive cases occurred within 30 days (B) Cases within Brighton were clustered on the basis of single nucleotide polymorphism distances and time compatible with transmission. The Lancet Infectious Diseases 2016 16, 1295-1303DOI: (10.1016/S1473-3099(16)30157-8) Copyright © 2016 Elsevier Ltd Terms and Conditions

Figure 5 Genetic clusters of gonorrhoea within Brighton, UK, and USA identified from 2004 onwards (A) and 2011 onwards (B) Each genetic cluster contains all cases related by single nucleotide polymorphisms and time compatible with transmission. Each genetic cluster is plotted on its own horizontal line, with individual cases shown as dots. For ease of visualisation, clusters arising from January, 2011, are shown separately in (B). Samples obtained in Brighton in 2004 and 2005 were collected within a 2 month period, but the exact collection dates were not available. These samples have been randomly distributed throughout the 2 months of sampling. Similarly, only the month and year of collection was known for the USA samples, and a random day has been assigned. The Lancet Infectious Diseases 2016 16, 1295-1303DOI: (10.1016/S1473-3099(16)30157-8) Copyright © 2016 Elsevier Ltd Terms and Conditions