Genetic Evidence of Drug-Resistant Malarial Strain from the Democratic Republic of the Congo Imported to Guatemala - Volume 20, Number 6—June 2014 - Emerging Infectious Disease journal - CDC

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Genetic Evidence of Drug-Resistant Malarial Strain from the Democratic Republic of the Congo Imported to Guatemala - Volume 20, Number 6—June 2014 - Emerging Infectious Disease journal - CDC

Volume 20, Number 6—June 2014

Research

Genetic Evidence of Drug-Resistant Malarial Strain from the Democratic Republic of the Congo Imported to Guatemala

Article Contents

• Methods
• Results
• Discussion
• Acknowledgments
• References
• Figure 1
• Figure 2
• Figure 3
• Figure 4
• Table
• Suggested Citation

Jaymin C. Patel , Steve M. Taylor, Patricia C. Juliao, Christian M. Parobek, Mark Janko, Luis Demetrio Gonzalez, Lucia Ortiz, Norma Padilla, Antoinette K. Tshefu, Michael Emch, Venkatachalam Udhayakumar, Kim Lindblade, and Steven R. Meshnick

Author affiliations: University of North Carolina, Chapel Hill, North Carolina, USA (J.C. Patel, S.M. Taylor, C.M. Parobek, M. Janko, M. Emch, S.R. Meshnick); Centers for Disease Control and Prevention, Atlanta, Georgia, USA (P.C. Juliao, V. Udhayakumar, K. Lindblade); Military Medical Center, Guatemala City, Guatemala (L.D. Gonzalez); Universidad de Valle de Guatemala, Guatemala City (L. Ortiz, N. Padilla); University of Kinshasa, Kinshasa, Democratic Republic of the Congo (A.K. Tshefu)

Suggested citation for this article

Abstract

Imported malaria threatens control and elimination efforts in countries that have low rates of transmission. In 2010, an outbreak of Plasmodium falciparum malaria was reported among United Nations peacekeeping soldiers from Guatemala who had recently returned from the Democratic Republic of the Congo (DRC). Epidemiologic evidence suggested that the soldiers were infected in the DRC, but local transmission could not be ruled out in all cases. We used population genetic analyses of neutral microsatellites to determine the outbreak source. Genetic relatedness was compared among parasites found in samples from the soldiers and parasite populations collected in the DRC and Guatemala; parasites identified in the soldiers were more closely related to those from the DRC. A phylogenetic clustering analysis confirms this identification with >99.9% confidence. Thus, results support the hypothesis that the soldiers likely imported malaria from the DRC. This study demonstrates the utility of molecular genotyping in outbreak investigations.

Imported malaria threatens control and elimination efforts in countries that report low malaria transmission rates (1–3). In Central America, malaria transmission decreased by >50% during 2000–2010 (4); in 2010, the Guatemala Ministerio de Salud Pública y Asistencia Social reported 31 confirmed cases of malaria, all caused by the species Plasmodium falciparum (5). Central America is unusual compared with other areas in which malaria is endemic because chloroquine remains an effective treatment option for P. falciparum infection there, but not in other parts of the world (6–8); the introduction of parasites harboring chloroquine-resistant genotypes could fuel a resurgence of clinical illness and transmission.

In 2010, an outbreak of malaria was reported among 12 soldiers from Guatemala shortly after they returned from a United Nations (UN) peacekeeping mission in the Democratic Republic of the Congo (DRC). Of the 12, 8 also reported visiting >1 area in Guatemala in which malaria is endemic when they returned but before the outbreak was identified. An outbreak investigation was undertaken after 1 of the infected soldiers died; laboratory tests of blood from this patient identified choroquine-resistant and -sensitive strains of P. falciparum.

The epidemiologic evidence suggested that the soldiers were infected while stationed in the DRC (9). Because the local acquisition of chloroquine-resistant parasites in Guatemala could necessitate a change in local treatment practices, it was vital to determine the origin of the soldiers’ infections.

Molecular markers have been used to assess the genetic relatedness of malarial parasites from different geographic regions (10,11). Accordingly, if the soldiers acquired P. falciparum in the DRC during their stay, the genotypes of the parasites isolated from the soldiers’ samples would be more closely related to parasites from the DRC than to parasites from Guatemala. To test this hypothesis, we used molecular methods from the field of population genetics to determine the source of the malaria outbreak among the soldiers who returned to Guatemala after being stationed in the DRC.

Acknowledgments

We thank Silvia Sosa for help in obtaining samples from Guatemala. We also thank Fabiola Enriquez, Mario Roberto Perez Franco, Harold Garcia Sergio Fernando, Nitsch Montiel, and Mario Luis Ovalle Figuero for their help in obtaining samples from the soldiers.

This work was supported by a Gillings Innovation Laboratory award from the UNC Gillings School of Global Public Health and by grants from the National Institutes of Health (NIH) (NIAID 1R56AI097609-01) and National Science Foundation (NSF) (BSC-13339949).

References

1. Feachem RG, Phillips AA, Hwang J, Cotter C, Wielgosz B, Greenwood BM, Shrinking the malaria map: progress and prospects. Lancet. 2010;376:1566–78. DOIPubMed
2. Martens P, Hall L. Malaria on the move: human population movement and malaria transmission. Emerg Infect Dis. 2000;6:103–9. DOIPubMed
3. Tatem AJ, Smith DL. International population movements and regional Plasmodium falciparum malaria elimination strategies. Proc Natl Acad Sci U S A. 2010;107:12222–7. DOIPubMed
4. World Health Organization. World malaria report 2011. Geneva: The Organization; 2011. [cited 2014 Mar 28].http://www.who.int/malaria/world_malaria_report_2011/en/
5. National Center for Epidemiology, Ministry of Public Health and Social Assistance. 2010 Vital statistics, Guatemala [in Spanish] 2010 [cited 2014 Mar 27]. Ciudad de Guatemala: The Ministry.http://epidemiologia.mspas.gob.gt/vigepi/2010/MEMORIA%20REPUBLICA%20DE%20GUATEMALA%202010.pdf 
6. World Health Organization. Guidelines for the treatment of malaria, 2nd ed. Geneva: The Organization; 2010 [modified 2011 June 8, cited 2014 Mar 28].http://whqlibdoc.who.int/publications/2010/9789241547925_eng.pdf?ua=1
7. World Health Organization. Global report on antimalarial efficacy and drug resistance: 2000–2010. Geneva: The Organization; 2010 [cited 2014 Mar 28].http://whqlibdoc.who.int/publications/2010/9789241500470_eng.pdf 
8. Mejia Torres RE, Banegas EI, Mendoza M, Diaz C, Bucheli STM, Fontecha GA, Efficacy of chloroquine for the treatment of uncomplicated Plasmodium falciparum malaria in Honduras. Am J Trop Med Hyg. 2013;88:850–4. DOIPubMed
9. Juliao PC, Sosa S, Gonzalez LD, Padilla N, Ortiz L, Goldman I, Importation of chloroquine-resistant Plasmodium falciparum by Guatemalan peacekeepers returning from the Democratic Republic of the Congo. Malar J. 2013;12:344. DOIPubMed
10. Rebaudet S, Bogreau H, Silai R, Lepere JF, Bertaux L, Pradines B, Genetic structure ofPlasmodium falciparum and elimination of malaria, Comoros archipelago. Emerg Infect Dis. 2010;16:1686–94. DOIPubMed
11. Schultz L, Wapling J, Mueller I, Ntsuke PO, Senn N, Nale J, Multilocus haplotypes reveal variable levels of diversity and population structure of Plasmodium falciparum in Papua New Guinea, a region of intense perennial transmission. Malar J. 2010;9:336. DOIPubMed
12. Chwalisz N, Kinosian S. Guatemala sends Special Forces to D. R. Congo for UN peacekeeping mission. 2013 [cited 2013 July 15].http://justf.org/blog/2013/08/29/guatemala-sends-special-forces-dr-congo-un-peacekeeping-mission
13. Taylor SM, Messina JP, Hand CC, Juliano JJ, Muwonga J, Tshefu AK, Molecular malaria epidemiology: mapping and burden estimates for the Democratic Republic of the Congo, 2007. PLoS ONE. 2011;6:e16420. DOIPubMed
14. Messina JP, Taylor SM, Meshnick SR, Linke AM, Tshefu AK, Atua B, Population, behavioural and environmental drivers of malaria prevalence in the Democratic Republic of Congo.Malar J. 2011;10:161. DOIPubMed
15. Taylor SM, van Eijk AM, Hand CC, Mwandagalirwa K, Messina JP, Tshefu AK,Quantification of the burden and consequences of pregnancy-associated malaria in the Democratic Republic of the Congo. J Infect Dis. 2011;204:1762–71. DOIPubMed
16. Taylor SM, Antonia AL, Parobek CM, Juliano JJ, Janko M, Emch M, Plasmodium falciparum sulfadoxine resistance is geographically and genetically clustered within the DR Congo. Scientific reports. 2013;3:1165.
17. Herderschee J, Kaiser KA, Samba DM. Resilience of an African giant: boosting growth and development in the Democratic Republic of Congo. Washington, (DC),The World Bank;2012. 116 p.
18. Nair S, Williams JT, Brockman A, Paiphun L, Mayxay M, Newton PN, A selective sweep driven by pyrimethamine treatment in southeast asian malaria parasites. Mol Biol Evol.2003;20:1526–36. DOIPubMed
19. Anderson TJ, Haubold B, Williams JT, Estrada-Franco JG, Richardson L, Mollinedo R,Microsatellite markers reveal a spectrum of population structures in the malaria parasitePlasmodium falciparum. Mol Biol Evol. 2000;17:1467–82. DOIPubMed
20. Anderson TJ, Su XZ, Bockarie M, Lagog M, Day KP. Twelve microsatellite markers for characterization of Plasmodium falciparum from finger-prick blood samples. Parasitology.1999;119:113–25. DOIPubMed
21. Roper C, Pearce R, Bredenkamp B, Gumede J, Drakeley C, Mosha F, Antifolate antimalarial resistance in southeast Africa: a population-based analysis. Lancet.2003;361:1174–81. DOIPubMed
22. Balloux F, Lugon-Moulin N. The estimation of population differentiation with microsatellite markers. Mol Ecol. 2002;11:155–65. DOIPubMed
23. Peakall ROD, Smouse PE. GENALEX 6: genetic analysis in Excel. Population genetic software for teaching and research. Mol Ecol Notes. 2006;6:288–95. DOI
24. Hardy OJ, Vekemans X. SPAGeDi: a versatile computer program to analyse spatial genetic structure at the individual or population levels. Mol Ecol Notes. 2002;2:618–20. DOI
25. Excoffier L, Laval G, Schneider S. Arlequin (version 3.0): an integrated software package for population genetics data analysis. Evol Bioinform Online. 2005;1:47–50http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2658868/.PubMed
26. Saitou N, Nei M. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol. 1987;4:406–25http://mbe.oxfordjournals.org/content/4/4/406.long.PubMed
27. Cavalli-Sforza LL, Edwards AW. Phylogenetic analysis. Models and estimation procedures. Am J Hum Genet. 1967;19:233–57http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1706274/.PubMed
28. Populations v. 1.2.31 [Internet]. Hudson (MA): Bioinformatics Organization [updated 2010 Oct 20; cited 2013 June 9].http://bioinformatics.org/~tryphon/populations/
29. Paradis E, Claude J, Strimmer K. APE: analyses of phylogenetics and evolution in R language. Bioinformatics. 2004;20:289–90. DOIPubMed
30. Hamady M, Lozupone C, Knight R. Fast UniFrac: facilitating high-throughput phylogenetic analyses of microbial communities including analysis of pyrosequencing and PhyloChip data. ISME J. 2010;4:17–27. DOIPubMed
31. Verhoef L, Vennema H, van Pelt W, Lees D, Boshuizen H, Henshilwood K, Use of norovirus genotype profiles to differentiate origins of foodborne outbreaks. Emerg Infect Dis. 2010;16:617–24 . DOIPubMed
32. Barry PM, Gardner TJ, Funk E, Oren E, Field K, Shaw T, Multistate outbreak of MDR TB identified by genotype cluster investigation. Emerg Infect Dis. 2012;18:113–6. DOIPubMed
33. Krause G, Schoneberg I, Altmann D, Stark K. Chemoprophylaxis and malaria death rates.Emerg Infect Dis. 2006;12:447–51. DOIPubMed
34. The Global Health Group and the Malaria Atlas Project. Atlas of Malaria-eliminating countries. San Francisco: The Global Health Group, Global Health Sciences, University of California, San Francisco; 2011 [cited 2014 Mar 28].http://www.malariaeliminationgroup.org/publications/atlas-of-malaria-eliminating-countries-2011
35. The Global Fund to Fight AIDS. Tuberculosis and Malaria News Releases [Internet]. Ten countries rally to eliminate malaria in Central America and the Caribbean. San Jose, Costa Rica: The Global Fund to Fight AIDS, Tuberculosis and Malaria; 2013 Jun 28 [cited 2014 Mar 28].http://www.theglobalfund.org/en/mediacenter/newsreleases/2013-06-28_Ten_Countries_Rally_to_Eliminate_Malaria_in_Central_America_and_the_Caribbean/
36. Warren M, Collins WE, Richardson BB, Skinner JC. Morphologic variants of Anopheles albimanus and susceptibility to Plasmodium vivax and P. falciparum. Am J Trop Med Hyg.1977;26:607–11http://www.ajtmh.org/content/26/4/607.long.PubMed
37. Baton LA, Ranford-Cartwright LC. Ookinete destruction within the mosquito midgut lumen explains Anopheles albimanus refractoriness to Plasmodium falciparum (3D7A) oocyst infection. Int J Parasitol. 2012;42:249–58. DOIPubMed
38. Wilson PE, Kazadi W, Kamwendo DD, Mwapasa V, Purfield A, Meshnick SR. Prevalence of pfcrt mutations in Congolese and Malawian Plasmodium falciparum isolates as determined by a new Taqman assay. Acta Trop. 2005;93:97–106. DOIPubMed
39. Gouandjika-Vasilache I, Mazitchi A, Gumede N, Manirakiza A, Manenegu C, KoyazegbeTD, Wild poliovirus importation, Central African Republic. Emerg Infect Dis.2013;19:1012–3. DOIPubMed
40. Odoom JK, Forrest L, Dunn G, Osei-Kwasi M, Obodai E, Arthur-Quarm J, Interruption of poliovirus transmission in Ghana: molecular epidemiology of wild-type 1 poliovirus isolated from 1995 to 2008. J Infect Dis. 2012;206:1111–20. DOIPubMed

Figures

• Figure 1. Location of the major cities, rivers, and the 7 Demographic Health Survey clusters (203, 81, 88, 183, 211, 164, and 29) within the Democratic Republic of the Congo (DRC)...
• Figure 2. Genetic relatedness (pairwise R_ST comparisons) among Plasmodium falciparumidentified in samples from the soldiers from Guatemala, persons in the Democratic Republic of the Congo, and persons in Guatemala.
• Figure 3. Neighbor-joining tree of 3 Plasmodium falciparum populationsPrefixes of genomes indicate parasite origins: Green text indicates parasite populations from the Democratic Republic of the Congo (DRC); orange indicates parasite populations...
• Figure 4. Phylogenetic tree showing predicted clustering between Plasmodium falciparumpopulations from the Democratic Republic of Congo (DRC), soldiers returning to Guatemala from the DRC, and GuatemalaThe predicted split between parasites...

Table

• Table. Neutral microsatellite loci and primer sequences used for PCR amplification of Plasmodiumfalciparum malaria genes to identify origin of drug-resistant genotype, Guatemala

Suggested citation for this article: Patel JC, Taylor SM, Juliao PC, Parobek CM, Janko M, Gonzalez LD, et al. Genetic evidence of drug-resistant malarial strain from the Democratic Republic of the Congo imported to Guatemala. Emerg Infect Dis. 2014 Jun [date cited].http://dx.doi.org/10.3201/eid2006.131204

DOI: 10.3201/eid2006.131204