Evolutionary and epidemiological analysis of Roka HIV outbreak
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Massive iatrogenic outbreak of human immunodeficiency virus type 1 in rural Cambodia, 2014-2015


François Rouet1 Janin Nouhin1, Du-Ping Zheng2, Benjamin Roche3, Allison Black4 Sophearot Prak1, Marie Leoz5, Catherine Gaudy-Graffin6, Laurent Ferradini7, Chandara Mom8, Sovatha Mam8, Charlotte Gautier1, Gérard Lesage6, Sreymom Ken1, Kerya Phon1, Alexandra Kerleguer1, Chunfu Yang2, William Killam9, Masami Fujita7, Chhivun Mean8, Didier Fontenille1, Francis Barin6, Jean-Christophe Plantier5, Trevor Bedford4, Artur Ramos9, Vonthanak Saphonn10

1Unité VIH/Hépatites, Institut Pasteur du Cambodge (IPC), Phnom Penh, Cambodia 2International Laboratory Branch (ILB), Centers for Disease Control and Prevention (CDC), Atlanta, USA 3UMR IRD 224-CNRS 5290-Université de Montpellier, Maladies Infectieuses et Vecteurs : Ecologie, Génétique, Evolution et Contrôle (MIVEGEC), Montpellier, France 4Fred Hutchinson Cancer Research Center, Seattle, USA 5Centre National de Référence (CNR) sur le VIH, Laboratoire Associé, EA2656, Rouen University Hospital, Rouen, France 6Centre National de Référence (CNR) sur le VIH and INSERM Unité 966, Tours, France 7World Health Organization (WHO), Phnom Penh, Cambodia 8National Center for HIV/AIDS, Dermatology and Sexual Transmitted Diseases (NCHADS), Phnom Penh, Cambodia 9Division of Global HIV/AIDS, Centers for Disease Control and Prevention (CDC), Phnom Penh, Cambodia 10University of Health Sciences (UHS), Phnom Penh, Cambodia


Background. In 2014-2015, 242 individuals aged 2–89 were newly HIV-1 diagnosed in Roka, a rural commune in Cambodia. A case-control study attributed the outbreak to unsafe injections. We aimed to reconstruct the likely transmission history of the outbreak.

Methods. We assessed in 209 (86.4%) HIV-infected cases the presence of hepatitis C and B viruses (HCV, HBV). We identified recent infections using antibody (Ab) avidity testing for HIV and HCV, and HBcIgM Ab for HBV. We performed evolutionary phylogenetic analyses of viral strains. Geographical coordinates and parenteral exposure through medical services provided by an unlicensed health care practitioner were obtained from 193 cases and 1499 controls during interviews.

Results. Cases were co-infected with HCV (78.5%) and HBV (12.9%). We identified 79 (37.8%) recent (<130 days) HIV infections. Phylogeny of 202 HIV env C2V3 sequences showed a 198-sample CRF01_AE strains cluster, with time to most recent common ancestor (tMRCA) in September 2013 (95% highest posterior density, August 2012–July 2014), and a peak of 15 infections/day in September 2014. Three geospatial HIV hotspots were discernible in Roka and correlated with high exposure to the practitioner (P=0.04). Fifty-nine (38.6%) of 153 tested cases showed recent (<180 days) HCV infections. Ninety HCV NS5B sequences formed three main clades, one containing 34 subtypes 1b with tMRCA in 2012, and two with 51 subtypes 6e and tMRCAs in 2002-2003.

Conclusions. Unsafe injections in Cambodia most likely led to an explosive iatrogenic spreading of HIV, associated with a long-standing and more genetically-diverse HCV propagation.


Rouet et al. 2017. Massive iatrogenic outbreak of human immunodeficiency virus type 1 in rural Cambodia, 2014-2015. Clin Infect Dis: cix1071. doi.org/10.1093/cid/cix1071

Data and analyses

All sequence data were either generated by or shared by the folks at Institut Pasteur.

We had the opportunity to perform some of the phylogenetic analysis for this project. Our goal was to explore the genomic data and improve our understanding the epidemiology of this outbreak. Some of the questions we were interested in were:

  • Are all infections that were found during contact tracing outbreak-related, or were additional non-outbreak related cases found because of enhanced surveillance?
  • When did the outbreak begin?
  • What was the growth rate of the epidemic and how many infections occurred?

Below are brief descriptions of the analyses we conducted for this project, with links out to the directories in this repo that are pertinent to each analysis. Within analysis-specific repos you’ll find READMEs that give a more in-depth explanation of the analysis, and that display the figures that were generated.

Supplemental analysis