Sequence variation of ookinete surface proteins Pvs25 and Pvs28 of Plasmodium vivax isolates from Southern Mexico and their association to local anophelines infectivity Lilia Gonza ´ lez-Cero ´n a, *, Alejandro Alvarado-Delgado b , Jesus Martı ´nez-Barnetche b , Mario H. Rodrı ´guez c , Marbella Ovilla-Mun ˜oz a , Fabia ´n Pe ´ rez a , Juan E. Hernandez-Avila d , Marco A. Sandoval a , Maria del Carmen Rodrı ´guez b , Cuauhte ´ moc Villarreal-Trevin ˜o a a Centro Regional de Investigacio ´n en Salud Pu ´blica, Instituto Nacional de Salud Pu ´blica, 4a Av. Norte and 19 calle Poniente, Tapachula, Chiapas, Mexico b Centro de Investigacio ´n en Enfermedades Infecciosas; Cuernavaca, Morelos, Mexico c Direccio ´n General, Cuernavaca, Morelos, Mexico d Direccio ´n de Informa ´tica: Instituto Nacional de Salud Pu ´blica, Av. Universidad 655, Cuernavaca, Morelos, Mexico 1. Introduction Plasmodium parasites are transmitted to their vertebrate host and mosquito vectors during the insect blood feeding. Mosquitoes obtain parasite sexual forms circulating in the blood of the vertebrate host, and after fecundation within the insect midgut, invasive forms (ookinetes) migrate to the external midgut surface, where sporozoites develop within oocysts. The sporozoites invade the mosquito salivary glands and are injected into new vertebrate hosts during the subsequent blood meals (Sinden, 1984). Several molecules that mediate the interactions of the different parasites stages with the mosquito organs have been identified (Vlachou et al., 2006); however, the molecular bases of the differences in infectivity of Plasmodia to different mosquito species remain unknown. The infectivity (measured by oocyst counts) of Southern Mexican Plasmodium vivax isolates to local Anopheles albimanus and Anopheles pseudopunctipennis was associated to the repeat central region sequence (CSPr) phenotypes (CSPrVk210 and Vk247) of their circumsporozoite proteins (Gonza ´ lez-Cero ´n Infection, Genetics and Evolution 10 (2010) 645–654 ARTICLE INFO Article history: Received 14 April 2009 Received in revised form 24 March 2010 Accepted 25 March 2010 Available online 2 April 2010 Keywords: Plasmodium vivax Pvs25 Pvs28 Ookinete surface molecules CSPr Polymorphism Haplotype Anopheles albimanus Anophelespseudopunctipennis ABSTRACT The polymorphism of Pvs25 and Pvs28 ookinete surface proteins, their association to circumsporozoite protein repeat (CSPr) genotypes (Vk210 and Vk247) and their infectivity to local Anopheles albimanus and Anopheles pseudopunctipennis were investigated in Plasmodium vivax-infected blood samples obtained from patients in Southern Mexico. The pvs25 and pvs28 complete genes were amplified, cloned and sequenced; and the CSPr genotype was determined by PCR amplification and hybridization. The amino acid Pvs25 and Pvs28 polymorphisms were mapped to their corresponding protein structure. Infected blood samples were simultaneously provided through artificial feeders to both mosquito species; the ratio of infected mosquitoes and oocyst numbers were recorded. The polymorphism of pvs25 and pvs28 was limited to few nucleotide positions, and produced three haplotypes: type A/A parasites presented Pvs25 and Pvs28 amino acid sequences identical to that of Sal I reference strain; parasites type B1 presented a mutation 130 Ile ! Thr in Pvs25, while type B2 presented 87 Gln ! Lys/130 Ile ! Thr in the same molecule. Both types B1 and B2 parasites presented changes in Pvs28 at 87 Asn ! Asp, 110 Tyr ! Asn and five GSGGE/D repeat sequences between the fourth EGF-like domain and the GPI. Most P. vivax parasites from the coastal plains and the overlapping region were Pvs25/28 A/A, CSPrVk210 and were infective only to An. albimanus (p  0.0001). Parasites originating in foothills were Pvs25/28 type B1/B or B2/B and CSPrVk210 or Vk247, and were more infective to An. pseudopunctipennis than to An. albimanus (p  0.001). These results and the analysis of Pvs25/28 from other parts of the world indicated that non-synonymous variations in these proteins occur in amino acid residues exposed on the surface of the proteins, and are likely to interact with midgut mosquito ligands. We hypothesize that these molecules have been shaped by co-evolutionary adaptations of parasites to their susceptible vectors. ß 2010 Elsevier B.V. All rights reserved. * Corresponding author at: ParasitologyDepartment, 4a Av. Norte y 19 calle Poniente, Tapachula, Chiapas 30700, Me ´ xico. Tel.: +52 962 6262219x112; fax: +52 962 6265782. E-mail address: lgonzal@insp.mx (L. Gonza ´ lez-Cero ´ n). Contents lists available at ScienceDirect Infection, Genetics and Evolution journal homepage: www.elsevier.com/locate/meegid 1567-1348/$ – see front matter ß 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.meegid.2010.03.014