SciELO - Scientific Electronic Library Online

 
vol.7 número3Perfil de los exámenes citopatológicos de cuello de útero realizados en la Casa da Mulher, Estado de Pará, BrasilVigilancia de ambientes de la fiebre maculosa: explorando las áreas silenciosas de Brasil índice de autoresíndice de materiabúsqueda de artículos
Home Pagelista alfabética de revistas  

Servicios Personalizados

Revista

Articulo

Indicadores

  • No hay articulos citadosCitado por SciELO

Links relacionados

  • No hay articulos similaresSimilares en SciELO

Compartir


Revista Pan-Amazônica de Saúde

versión impresa ISSN 2176-6215versión On-line ISSN 2176-6223

Rev Pan-Amaz Saude v.7 n.3 Ananindeua sep. 2016

http://dx.doi.org/10.5123/s2176-62232016000300007 

ORIGINAL ARTICLE

Frequency of Plasmodium vivax circumsporozoite protein genotypes in humans and anopheline mosquitoes in an endemic area of southeastern Pará State, Brazil

Frequência de genótipos da proteína circunsporozoíta de Plasmodium vivax em seres humanos e mosquitos anofelinos em área endêmica da região sudeste do Estado do Pará, Brasil

Frecuencia de genotipos de la proteína circunsporozoíta de Plasmodium vivax en seres humanos y mosquitos Anopheles en área endémica de la región sudeste del Estado de Pará, Brasil

Erian de Almeida Santos1  , Deocleciano Galiza Primo2  , Izis Mônica Carvalho Sucupira2  , Gustavo Capatti Cassiano3  , Danielle Regina Lima Barbosa2  , Giselle Maria Rachid Viana2  , Ricardo Luiz Dantas Machado1  2  3  , Marinete Marins Póvoa1  2 

1Universidade Federal do Pará, Belém, Pará, Brasil

2Instituto Evandro Chagas/SVS/MS, Ananindeua, Pará, Brasil

3Universidade Estadual Paulista, Botucatu, São Paulo, Brasil

ABSTRACT

The objective of this study was therefore to investigate the frequency of circumsporozoite protein (CSP) genotypes in human blood and their correlation with parasitemia, as well as to evaluate the presence of these genotypes in Anopheles in the Municipality of Goianésia do Pará, an endemic area of southeastern Pará State, Brazil from 2012-2013. Blood samples were collected from 118 patients with Plasmodium vivax and 369 anopheline mosquitoes. The CSP gene was genotyped using the polymerase chain reaction/restriction fragment length polymorphism technique, and the infectivity of the anophelines was determined using ELISA. Parasitemia ranged from 5-70,000 parasites/mm³, and the three genotypes (VK210, VK247, and P. vivax-like) were detected both in single and mixed infections. No sample exhibited mixed infection with all three genotypes. The most frequent genotype was VK210 followed by VK247 and the latter associated with the highest parasitemia values (p < 0.0001). Among the identified mosquitoes, only 11 specimens were infected; of the seven Anopheles darlingi specimens four were infected with Plasmodium falciparum, two with VK210, and one with VK247. The three Anopheles albitarsis specimens were infected with VK247, and one Anopheles nuneztovari specimen was infected with VK210. The VK210 genotype continues to be the most prevalent in southeastern Pará; however, a new evidence shows the adaptation of VK247. The species An. darlingi, An. albitarsis, and An. nuneztovari play an important role in the transmission of CSP genotypes in the study area. This finding may be a public health concern due to the possibility of resurgence of P. vivax malaria epidemics in susceptible communities.

Keywords: Malaria; Plasmodium vivax; Genotyping Techniques; Anopheles

RESUMO

O objetivo deste estudo foi investigar a frequência de genótipos da proteína circunsporozoíta (CSP) em sangue humano e sua correlação com a parasitemia, bem como avaliar a presença desses genótipos em Anopheles no Município de Goianésia do Pará, uma área endêmica do sudeste do Estado do Pará, Brasil, de 2012 a 2013. Amostras de sangue foram coletadas de 118 pacientes com Plasmodium vivax e 369 mosquitos anofelinos. O gene da CSP foi genotipado usando-se a reação em cadeia da polimerase/polimorfismo de comprimento de fragmento de restrição, e a infectividade dos anofelinos foi determinada pelo ELISA. A parasitemia variou de 5-70.000 parasitas/mm³, e os três genótipos (VK210, VK247 e P. vivax-like) foram detectados tanto em infecções simples quanto em mistas. Nenhuma amostra apresentou infecção mista com todos os três genótipos. O genótipo mais frequente foi o VK210, seguido pelo VK247 e o último associado com os valores mais altos de parasitemia (p < 0,0001). Entre os mosquitos identificados, somente 11 espécimes foram infectados; de sete espécimes Anopheles darlingi, quatro foram infectados por Plasmodium falciparum, dois por VK210 e um por VK247. Os três Anopheles albitarsis foram infectados por VK247 e um Anopheles nuneztovari por VK210. O genótipo VK210 continua sendo o mais prevalente no sudeste do Pará; entretanto, novas evidências indicam a adaptação do VK247. Os espécimes An. darlingi, An. albitarsis e An. nuneztovari desempenham um importante papel na transmissão dos genótipos CSP na área de estudo. Essa descoberta pode ser um problema de saúde pública devido à possibilidade de ressurgimento de epidemias de malária por P. vivax em comunidades suscetíveis.

Palavras-chave: Malária; Plasmodium vivax; Técnicas de Genotipagem; Anopheles

RESUMEN

El objetivo de este estudio fue de investigar la frecuencia de genotipos de la proteína circunsporozoíta (CSP) en sangre humana y su correlación con la parasitemia, bien como evaluar la presencia de esos genotipos en Anopheles en el Municipio de Goianésia do Pará, un área endémica del sudeste del Estado de Pará, Brasil, de 2012 a 2013. Se recolectaron muestras de sangre de 118 pacientes con Plasmodium vivax y 369 mosquitos Anopheles. El gen de la CSP fue genotipado usando la reacción en cadena de la polimerasa/polimorfismo de longitud de fragmento de restricción, y la infectividad de los Anopheles se determinó por el método ELISA. La parasitemia varió de 5-70.000 parásitos/mm³, y los tres genotipos (VK210, VK247 y P. vivax-like) fueron detectados tanto en infecciones simples como en mixtas. Ninguna muestra presentó infección mixta con todos los tres genotipos. El genotipo más frecuente fue el VK210, seguido por VK247 y el último asociado con los valores más altos de parasitemia (p < 0,0001). Entre los mosquitos identificados, solamente 11 especímenes fueron infectados; de siete especímenes Anopheles darlingi, cuatro fueron infectados por Plasmodium falciparum, dos por VK210 y uno por VK247. Los tres Anopheles albitarsis fueron infectados por VK247 y un Anopheles nuneztovari por VK210. El genotipo VK210 sigue siendo el más prevalente en el sudeste de Pará; mientras que nuevas evidencias indican la adaptación del VK247. Los especímenes An. darlingi, An. albitarsis y An. nuneztovari desempeñan un importante papel en la transmisión de los genotipos CSP en el área de estudio. Ese hallazgo puede ser un problema de salud pública debido a la posibilidad de resurgimiento de epidemias de malaria por P. vivax en comunidades susceptibles.

Palabras clave: Malaria; Plasmodium vivax; Técnicas de Genotipaje; Anopheles

INTRODUCTION

Plasmodium falciparum, Plasmodium malariae, and Plasmodium vivax are the main species that cause malaria in Brazil, and the latter parasite is responsible for 80% of the malaria cases registered. Furthermore, 99.7% of reported cases are detected in Brazilian Amazon1,2,3. P. vivax malaria rarely progresses to severe disease; however, studies reinforce the idea that this species may be involved in clinical complications and fatalities, making it a matter of public health concern4,5.

P. vivax sporozoites are covered by circumsporozoite protein (CSP), which is involved in hepatocyte invasion mechanisms and is highly immunogenic6. Genetic variants exist in the central repeat domain of the CSP gene that are characterized as three genotypes, termed VK210, VK247 and P. vivax-like7,8,9. Serological and molecular studies have shown that these genotypes circulate nationwide, mainly in Brazilian Amazon10,11,12,13,14. The VK210 genotype is the most common in the investigated areas, and the VK247 and P. vivax-like genotypes had been found only in mixed infections with other genotypes13,14. However, a decade after the first genotypic study in five different states of Brazilian Amazon, this distribution profile has changed, with the VK247 and P. vivax-like genotypes also detected as single infections in a single area of Pará State15.

Human malaria is transmitted by mosquitoes of the genus Anopheles, which includes 465 recognized species and more than 50 unidentified species16. Transmission occurs in a human-vector-human manner, and the main malaria-transmitting species in Brazil is Anopheles darlingi, although other anopheline species have been detected to be naturally infected with P. vivax and/or P. falciparum and can therefore act as secondary vectors17,18,19,20,21,22.

The sexual reproduction of the parasite occurs during the cycle of the vector, and genetic changes in the plasmodium may occur during this stage23. Additionally, the variability of the repeat region of the CSP gene has also been used to determine infectivity of Plasmodium species in Anopheles24,25. Previous studies using monoclonal antibodies against VK210 and VK247 CSP in non-endemic areas of the Atlantic Forest, São Paulo State, detected the presence of Anopheles infected with both genotypes26. In endemic areas of Brazilian Amazon, 15% of Anopheles were found to be infected with this genotype in Acre State using anti-P. vivax-like antibodies27. In Pará State, it has been observed that the Anopheles aquasalis and An. darlingi species are susceptible to infection by VK210 and VK24728. This demonstrates the importance of studying the genetic diversity of P. vivax in endemic areas. Thus, it is essential to investigate the epidemiology of P. vivax CSP genotypes and to detect naturally infected mosquitoes to understand immunity mechanisms as well as differences in parasite load, drug resistance, transmission dynamics, and natural selective pressure of the parasite among different local communities29,30. The objective of the current study was to investigate the frequency of CSP genotypes in human blood and its correlation with parasitemia, as well as to evaluate the presence of these genotypes in Anopheles in an endemic area of southeastern Pará State, Brazil.

MATERIALS AND METHODS

STUDY AREA

This descriptive epidemiological study was conducted in Municipality of Goianésia do Pará (03°50'33"S, 49°05'49"W) located in Pará State southeastern mesoregion and Paragominas microregion, bordering the Municipalities of Breu Branco, Novo Repartimento, Dom Eliseu, Ipixuna do Pará, Jacundá, and Rondon do Pará. This region has a land area of 7,021 km² and an estimated population of 29,161 inhabitants, of whom 52% are over 14 years old. This area has population density of 4.5 inhabitants/km², and its distance from Belém, the State capital, is approximately 350 km31.

STUDY POPULATION

Patients

The sample was formed by patients living in the localities of Santa Paula, Rouxinol, and Ararandeua that concentrate the highest number of malaria cases in Goianésia do Pará Municipality32 and which sought the healthcare services in their own localities during 2012-2013 and had a positive diagnosis by P. vivax. Thus, a total of 118 blood samples were collected from patients with P. vivax diagnosed by Thick Smear technique and confirmed by nested PCR as described by Kimura et al33. The following individuals were excluded from the study: pregnant women, children under 15 years old, and patients who were infected with other Plasmodium species, displayed mixed infection, or who did not agree to participate. This study was approved by the Ethics Committee for Research on Human Beings of Instituto Evandro Chagas under number 00141/10 - CAAE: 0014.0.0.072.000-10, and participants signed an informed consent form.

Mosquitoes

The adult mosquitoes were collected from the same localities as the patients, and three collections/year (one of 12 h and two of 4 h by locality) were performed. The mosquitoes were identified by dichotomous keys18,34, and their natural infectivity was determined using ELISA protocol35. This method detects natural infection in mosquitoes caused by P. falciparum, P. malariae, and the VK210 and VK247 genotypes of P. vivax.

P. vivax GENOTYPING

DNA was extracted from peripheral blood samples using the Easy-DNATM (Invitrogen, Carlsbad, California, USA) and the QIAamp® DNA Blood Kit (Qiagen, Inc., Chatsworth, California, USA) extraction/purification kits. The P. vivax CSP genotypes were determined using polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP)36.

STATISTICAL ANALYSIS

Differences between the P. vivax CSP genotypes were tested using the Mann-Whitney and Kruskal-Wallis tests. The significance level for statistical tests was set at p < 0.05.

RESULTS

P. vivax CSP GENOTYPES

Of the 118 blood samples collected and diagnosed as infected with P. vivax, 110 were successfully amplified and genotyped. The three genotypes were detected both as single and mixed infections (Table 1). There were no mixed infections containing all three genotypes. The most frequent genotype in Goianésia do Pará Municipality was VK210, followed by VK247. Genotyping revealed that 75.5% of the samples were single infections, with only 24.5% mixed infections.

Table 1 - Distribution of P. vivax genotypes in Municipality of Goianésia do Pará, Pará State, Brazil, 2012-2013 

*1: VK210; 2: VK247; 3: P. vivax-like. Conventional sign used: - Numeric data not equal to zero due to rounding.

RELATIONSHIP BETWEEN PARASITEMIA AND CSP GENOTYPES

Parasitemia ranged from 5-70,000 parasites/mm³ (geometric mean ± standard deviation: 1,299.95 ± 8:51 parasites/mm³). The mean parasitemia was 569.42 parasites/mm3 (± 10:52) for the VK210 genotype, 2,563.76 parasites/mm3 (± 4.13) for the VK247 genotype, and 3,307 parasites/mm3 (± 5.64) for mixed VK210 and VK247 infection.

The P. vivax-like genotype was excluded from the analysis due to its low frequency. A significant association was observed between the presence or absence of VK247 in the infections and parasitemia. Regardless of whether single or mixed infection was observed, individuals infected with this genotype were associated with the highest parasitemia values (Mann-Whitney, p < 0.001) compared with those without the VK247 genotype (Figure 1).

Figure 1 - Statistical association between the VK247 genotype and parasitemia significant difference (p < 0.001) between individuals with the VK247 (1) variant and those without the VK247 (0) genotype with parasitemia (number of parasites/mm3

NATURAL INFECTION OF MOSQUITOES

Of the 369 adult anopheline species found in that Municipality, namely An. darlingi, An. albitarsis s.l., Anopheles triannulatus, Anopheles strodei, Anopheles galvaoi, and Anopheles nuneztovari, only 11 specimens were infected: seven An. darlingi (four with P. falciparum, two with VK210, and one with VK247); three An. albitarsis s.l. (with VK247); and one An. nuneztovari (with VK210).

DISCUSSION

The municipal government of Goianésia do Pará has implemented diagnostic training measures and treatment adherence campaigns, and has introduced insecticide-treated nets to decrease the time between onset of symptoms and the start of treatment, and to reduce or prevent human-vector contact. Due to the significant drop in the number of cases reported in recent years (2011 - 2,856 registered cases, 2012 - 1,136, and 2013 - 192), it is believed that these measures are effective. However, it should be emphasized that clarification of the geographic distribution of genotypes and of mosquitoes naturally infected with different Plasmodium species can provide new opportunities for understanding vector/parasite interaction and the local epidemiology of malaria.

The detection of three P. vivax CSP genotypes circulating in the municipality confirms previous evaluations conducted in Brazilian Amazon in the States of Pará, Rondônia, Amapá, Acre, and Mato Grosso13,15. However, there was no evidence of mixed infections with the three CSP genotypes in Goianésia do Pará Municipality, which contrasts with previous data from Novo Repartimento in Pará State, a Municipality located in the same mesoregion of southeastern Pará15. The VK210 genotype remains the most prevalent, most likely because of the great susceptibility of the An. darlingi vector, which is the most abundant in the region, to this variant28. The P. vivax-like genotype had a frequency of only 2.7% of the genotyped samples; this low frequency could be due to its recent introduction into the region or due to differences in the development of this genotype in the vectors present in the area12,13. Interestingly, the VK247 genotype appeared for the first time as a single infection ten years ago in Municipality of Novo Repartimento15. In this study, this genotype was also found to have naturally infected humans in isolation and was observed in all mosquitoes of the species An. albitarsis and in one An. darlingi specimen.

Another relevant observation is the association of patients infected with the VK247 genotype and the high parasitemia. At the end of the 1990s, Machado and Póvoa13 observed that the VK210 genotype was associated with high levels of parasitemia in the City of Belém (350 km from Goianésia do Pará). This profile change may be related to the evolution of P. vivax. González-Cerón et al37 observed in samples of P. vivax from Mexico, Nicaragua, and Peru that the genetic diversity of the CSP gene is restricted mainly to the central repeat domain and 3'-terminal portion. These authors also stressed that this variation occurs due to changes in the type of nucleotides and number of repeats of the repeat region. The authors noted that the VK247 genotype displays high identity at the carboxy-terminal end with the reported sequence for Plasmodium cynomolgi CSP, and it is possible that the repeat region of VK247 is more stable than VK210. Despite of the association between high parasitemia and the VK247 genotype in this study can not be causal, the data obtained here contribute to the understanding of the molecular epidemiology of P. vivax in Brazil and suggest that the introduction of the VK247 and VK210 genotypes may have occurred at different times according to the endemic area of Brazilian Amazon.

Naranjo-Díaz et al38 showed evidence that the importance and distribution of human malaria vectors may vary depending on location. In fact, these authors observed An. nuneztovari infected with the VK210 and VK247 genotypes in Colombia, showing its importance for malaria transmission in areas with anthropic intervention38. This pattern was also observed in Brazil, where An. darlingi was found to be infected only with VK247 in District of Lourenço, Amapá State39. Moreover, in Municipality of Marabá, another mesoregion of southeastern Pará State, An. darlingi was found to be infected by P. falciparum and VK247, whereas An. albitarsis was infected with both VK210 and VK247 genotypes40. In turn, there was a 1.22% infection rate in Acre State for VK247 in the Anopheles oswaldoi mosquito24, the main malaria vector in the region41. In this descriptive study, the VK247 CSP genotype was detected in one mosquito An. darlingi and in three An. albitarsis, corresponding to 36.4% (4/11) of infection. As these two species play an important role in malaria transmission in Brazil and because this variant has been detected separately in different locations in this mesoregion of Pará, it was hypothesized in this study an increase in the number of cases of this genotype in the region. This new evidence should be investigated in other locations where different species of anopheline mosquitoes are the main vector, such as An. aquasalis in Amazon Region20 and Anopheles bellator and Anopheles cruzii in outside Amazon region3, to determine whether other species may facilitate the transmission process by carrying different CSP genotypes.

In general the distribution profile of CSP genotypes in other countries of Latin America is different from that observed in Brazil. Thus, the possibility of an outbreak of vivax malaria cases by VK247 in Brazil cannot be ruled out. This could lead to higher parasitemia as well as more severe clinical conditions. Although the VK247 variant is ancestral and its CSP repeat region is more stable than that of VK21042, the distribution of this genotype can be quite heterogeneous in other regions of Brazilian Amazon, most likely due to the absence or presence of VK247 polymorphisms in different locations and this fact may be related to changes in the geographical distribution profile of this variant37,43.

Another hypothesis that reinforces this idea is related to the immune response. The VK210 genotype is more immunogenic than VK247, which leads to high immune responses against this genotype and facilitates the selection process of VK247 sporozoites. The possible presence of anti-VK210 antibodies may therefore limit the production of VK210 sporozoites and result in a lower frequency of this genotype in some species of mosquito44. This may also impact the parasite load in infected individuals. These variations can occur due to physiological incompatibility between host/parasite, defense mechanisms of the mosquito such as the destruction of ookinetes blocking the development of oocysts in the mosquito, or ecological and evolutionary factors that can contribute to the divergence or restriction of gene flow among parasite strains adapted to different local vectors45,46,47. It is noteworthy that the detection of CSP in vectors only occurs when the parasite reaches the sporoblast stage48,49, and for this reason it is possible that the absence of CSP expression in a parasite or even the vector's immune response against this protein may have influenced the detection of genotypes in mosquitoes.

CONCLUSION

Although the VK210 genotype remains the most prevalent in Brazil, a new evidence reveals a strong adaptation of the VK247 variant in southeastern Pará, as well as the association of this genotype with high parasitemia. The species An. darlingi, An. albitarsis and An. nuneztovari play an important role in the transmission of these genotypes in the study area. However this is the second time that An. albitarsis has been found in a natural infection with the VK247 genotype in Pará State, and it may be the main vector in the spread/selection of this genotype. This may therefore present a public health concern because it raises the possibility of a resurgence of vivax malaria epidemics in susceptible communities.

ACKNOWLEDGMENTS

We thank the technicians from the Basic Malaria Research Laboratory of Instituto Evandro Chagas and from the Center of Microorganism Investigation of Faculdade de Medicina de São José do Rio Preto (FAMERP) for assistance in obtaining samples and laboratory support.

FINANCIAL SUPPORT

The study reported in this manuscript was funded by National Counsel of Technological and Scientific Development (CNPq - 555.654/2009-5) and Pará State Research Foundation (FAPESPA - ICAAF 005/2011) to MMP. The research was jointly sponsored by a grant from FAMERP/CNPq (471605/2011-5) to RLDM.

REFERENCES

1 Oliveira-Ferreira J, Lacerda MV, Brasil P, Ladislau JL, Tauil PL, Daniel-Ribeiro CT. Malaria in Brazil: an overview. Malar J. 2010 Apr;30(9):115. Doi: 10.1186/1475-2875-9-115 [Link] [ Links ]

2 Ministério da Saúde (BR). Secretaria de Vigilância em Saúde. Sistema de Informação e Vigilância Epidemiológica: Sivep-Malária [Internet]. Brasília: Ministério da Saúde; 2015 [citado 2015 dez 11]. Disponível em: Disponível em: http://portalweb04.saude.gov.br/sivep_malaria . [ Links ]

3 Pina-Costa A, Brasil P, Di Santi SM, Araujo MP, Suárez-Mutis MC, Santelli ACFS, et al. Malaria in Brazil: what happens outside the Amazonian endemic region. Mem Inst Oswaldo Cruz. 2014 Aug;109(5):618-33. Doi: 10.1590/0074-0276140228 [Link] [ Links ]

4 Lacerda MV, Hipólito JR, Passos LN. Chronic Plasmodium vivax infection in a patient with splenomegaly and severe thrombocytopenia. Rev Soc Bras Med Trop. 2008 Sep-Oct;41(5):522-3. Doi: 10.1590/S0037-86822008000500021 [Link] [ Links ]

5 Andrade BB, Reis-Filho A, Souza-Neto SM, Clarêncio J, Camargo LM, Barral A, et al. Severe Plasmodium vivax malaria exhibits marked inflammatory imbalance. Malar J. 2010;9(13):1-8. Doi: 10.1186/1475-2875-9-13 [Link] [ Links ]

6 Coppi A, Natarajan R, Pradel G, Bennett BL, James ER, Roggero MA, et al. The malaria circumsporozoite protein has two functional domains, each with distinct roles as sporozoites journey from mosquito to mammalian host. J Exp Med. 2011 Feb; 208(2):341-56. Doi: 10.1084/jem.20101488 [Link] [ Links ]

7 Arnot DE, Barnwell JW, Tam JP, Nussenzweig V, Nussenzweig RS, Enea V. Circumsporozoite protein of Plasmodium vivax: gene cloning and characterization of the immunodominant epitope. Science. 1985 Nov;230(4727):815-8. Doi: 10.1126/science.2414847 [Link] [ Links ]

8 Rosenberg R, Wirtz RA, Lanar DE, Sattabongkot J, Hall T, Waters AP, et al. Circumsporozoite protein heterogeneity in the human malaria parasite Plasmodium vivax. Science. 1989 Sep;245(4921):973-6. Doi: 10.1126/science.2672336 [Link] [ Links ]

9 Qari SH, Shi YP, Goldman IF, Udhayarkumar V, Alpers MP, Collins WE, et al. Identification of Plasmodium vivax-like human malaria parasite. Lancet. 1993 Mar;341(8848):780-3. Doi: 10.1016/0140-6736(93)90559-Y [Link] [ Links ]

10 Arruda ME, Aragaki C, Gagliardi F, Haile RW. A seroprevalence and descriptive epidemiological study of malaria among Indian tribes of the Amazon basin of Brazil. Ann Trop Med Parasitol. 1996 Apr;90(2):135-43. [ Links ]

11 Arruda ME, Souza RC, Veiga ME, Ferreira AF, Zimmerman RH. Prevalence of Plasmodium vivax variants VK247 and P. vivax-like human malaria: a retrospective study in indigenous Indian populations of the Amazon region of Brazil. Trans R Soc Trop Med Hyg. 1998 Nov-Dec;92(6):628. Doi: 10.1016/S0035-9203(98)90788-X [Link] [ Links ]

12 Arruda ME, Zimmerman RH, Souza RM, Oliveira-Ferreira J. Prevalence and level of antibodies to the circumsporozoite protein of human malaria parasites in five states of the Amazon region of Brazil. Mem Inst Oswaldo Cruz. 2007 Jun;102(3): 367-71. Doi: 10.1590/S0074-02762007005000041 [Link] [ Links ]

13 Machado RLD, Póvoa MM. Distribution of Plasmodium vivax variants (VK210, VK247 and P. vivax-like) in three endemic areas of the Amazon region of Brazil and their correlation with chloroquine treatment. Trans R Soc Trop Med Hyg. 2000 Jul-Aug;94(4):377-81. Doi: 10.1016/S0035-9203(00)90110-X [Link] [ Links ]

14 Machado RL, Figueiredo Filho AF, Calvosa VS, Figueredo MC, Nascimento JM, Póvoa MM. Correlation between Plasmodium vivax variants in Belém, Pará State, Brazil and symptoms and clearance of parasitaemia. Braz J Infect Dis. 2003 Jun;7(3):175-7. Doi: 10.1590/S1413-86702003000300002 [Link] [ Links ]

15 Storti-Melo LM, Souza-Neiras WC, Cassiano GC, Joazeiro ACP, Fontes CJ, Bonini-Domingos CR, et al. Plasmodium vivax circumsporozoite variants and Duffy blood group genotypes in the Brazilian Amazon region. Trans R Soc Trop Med Hyg. 2009 Jul;103(7):672-8. Doi: 10.1016/j.trstmh.2008.07.018 [Link] [ Links ]

16 Harbach RE. Genus Anopheles Meigen, 1818. Mosquito Taxonomic Inventory [Internet]. 2011 [cited 2015 Dec 14]. Available from: Available from: http://mosquito-taxonomic-inventory.info/genus-anopheles-meigen-1818/ . [ Links ]

17 Deane LM, Causey OR, Deane MP. Notas sobre a distribuição e a biologia dos anofelinos das regiões nordestina e amazônica do Brasil. Rev Serv Esp Saude Publica. 1948;1:827-965. [ Links ]

18 Consoli RA, Oliveira RL. Classificação e principais espécies de importância sanitária no Brasil. Fiocruz: Rio de Janeiro; 1994. [ Links ]

19 Marrelli MT, Honório NA, Flores-Mendoza C, Lourenço-de-Oliveira R, Marinotti O, Kloetzel JK. Comparative susceptibility of two members of the Anopheles oswaldoi complex, An. oswaldoi and An. konderi, to infection by Plasmodium vivax. Trans R Soc Trop Med Hyg. 1999 Jul-Aug;93(4):381-4. Doi: 10.1016/S0035-9203(99)90123-2 [Link] [ Links ]

20 Póvoa MM, Silva ANM, Santos CCB, Segura MNO, Machado RLD. Malaria transmission. Cienc Cult J Braz Assoc Adv Science. 2000;54(4/5):208-12. [ Links ]

21 Galardo AKR, Arruda M, Couto A, Wirtz R, Lounibos LP, Zimmerman RH. Malaria vector incrimination in three rural riverine villages in the Brazilian Amazon. Am J Trop Med Hyg. 2007 Mar;76(3):461-9. . [Link] [ Links ]

22 Foley DH, Linton YM, Ruiz-Lopez JF, Conn JE, Sallum MA, Póvoa MM, et al. Geographic distribution, evolution, and disease importance of species within the Neotropical Anopheles albitarsis Group (Diptera, Culicidae). J Vector Ecol. 2014 Jun;39(1):168-81. Doi: 10.1111/j.1948-7134.2014.12084.x [Link] [ Links ]

23 Mueller I, Galinski MR, Baird JK, Carlton JM, Kochar DK, Alonso PL, et al. Key gaps in the knowledge of Plasmodium vivax, a neglected human malaria parasite. Lancet Infect Dis. 2009 Sep;9(9): 555-66. Doi: 10.1016/S1473-3099(09)70177-X [Link] [ Links ]

24 Branquinho MS, Lagos CB, Rocha RM, Natal D, Barata JM, Cochrane AH, et al. Anophelines in the state of Acre, Brazil, infected with Plasmodium falciparum, P. vivax, the variant P. vivax VK247 and P. malariae. Trans R Soc Trop Med Hyg. 1993 Jul-Aug;87(4):391-4. Doi: 10.1016/0035-9203(93)90008-E [Link] [ Links ]

25 Curado I, Duarte AM, Lal AA, Oliveira SG, Kloetzel JK. Antibodies anti bloodstream and circumsporozoite antigens (Plasmodium vivax and Plasmodium malariae/P. brasilianum) in areas of very low malaria endemicity in Brazil. Mem Inst Oswaldo Cruz. 1997 Mar-Apr;92(2):235-43. Doi: 10.1590/S0074-02761997000200017 [Link] [ Links ]

26 Branquinho MS, Marrelli MT, Curado I, Natal D, Barata JM, Tubaki R, et al. Infection of Anopheles (Kerteszia) cruzii by Plasmodium vivax and Plasmodium vivax variant VK247 in the municipalities of São Vicente and Juquitiba, São Paulo. Rev Panam Salud Publica. 1997 Sep;2(3):189-93. Doi: 10.1590/S1020-49891997000900004 [Link] [ Links ]

27 Marrelli MT, Branquinho MS, Hoffmann EH, Taipe-Lagos CB, Natal D, Kloetzel JK. Correlation between positive serology for Plasmodium vivax-like/Plasmodium simiovale malaria parasites in the human and anopheline populations in the State of Acre, Brazil. Trans R Soc Trop Med Hyg. 1998 Mar-Apr;92(2):149-51. Doi: 10.1016/S0035-9203(98)90723-4 [Link] [ Links ]

28 Silva ANM, Santos CCB, Lacerda RN, Machado RLD, Póvoa MM. Susceptibility of Anopheles aquasalis and An. darlingi to Plasmodium vivax VK210 and VK247. Mem Inst Oswaldo Cruz. 2006 Aug;101(5):547-50. Doi: 10.1590/S0074-02762006000500011 [Link] [ Links ]

29 Kosek M, Yori PP, Gilman RH, Calderon M, Zimic M, Chuquiyauri R, et al. High degree of Plasmodium vivax diversity in the Peruvian Amazon demonstrated by tandem repeat polymorphism analysis. Am J Trop Med Hyg. 2012 Apr;86(4):580-6. Doi: 10.4269/ajtmh.2012.11-0627 [Link] [ Links ]

30 Raza A, Ghanchi NK, Thaver AM, Jafri S, Beg MA. Genetic diversity of Plasmodium vivax clinical isolates from southern Pakistan using pvcsp and pvmsp1 genetic markers. Malar J. 2013 Jan;12(16):1-5. Doi: 10.1186/1475-2875-12-16 [Link] [ Links ]

31 Secretaria de Estado de Planejamento, Orçamento e Finanças (Pará). Instituto de Desenvolvimento Econômico, Social e Ambiental do Pará. Estatística municipal: Goianésia do Pará. Belém: Idesp; 2011. [Link] [ Links ]

32 Ministério da Saúde (BR). Secretaria de Vigilância em Saúde. Sistema de Informação de Vigilância Epidemiológica - Sivep-Malária: incidência parasitária anual de malária no Estado do Pará [Internet]. Brasília: Ministério da Saúde ; 2009 [citado 2015 dez 15]. Disponível em: Disponível em: http://portalweb04.saude.gov.br/sivep_malaria/ . [ Links ]

33 Kimura M, Kneko O, Liu Q, Zhou M, Kawamoto F, Wataya Y, et al. Identification of the four species of human malaria parasites by nested PCR that targets variant sequences in the small subunit rRNA gene. Parasitol Intern. 1997 Jul;46(2):91-5. Doi: 10.1016/S1383-5769(97)00013-5 [Link] [ Links ]

34 Faran ME, Linthicum KJ. A handbook of the Amazonian species of Anopheles (Nyssorhynchus) (Diptera: Culicidae). Mosquito Systems. 1981;13(1): 1-81. [ Links ]

35 Wirtz RA, Burkot TR, Andre RG, Rosenberg R, Collins WE, Roberts DR. Identification of Plasmodium vivax sporozoites in mosquitoes using an enzyme linked immunosorbent assay. Am J Trop Med Hyg. 1985 Nov;34(6):1048-54. [ Links ]

36 Cassiano GC, Storti-Melo LM, Póvoa MM, Galardo AK, Rossit AR, Machado RLD. Development of PCR-RFLP assay for the discrimination of Plasmodium species and variants of P. vivax (VK210, VK247 and P. vivax-like) in Anopheles mosquitoes. Acta Trop. 2011 May;118(2):118-22. Doi: 10.1016/j.actatropica.2011.02.011 [Link] [ Links ]

37 González-Cerón L, Martinez-Barnetche J, Montero-Solis C, Santillán F, Soto AM, Rodriguez MH, et al. Molecular epidemiology of Plasmodium vivax in Latin America: polymorphism and evolutionary relationships of the circumsporozoite gene. Malaria J. 2013 Jul;12(243):1-19. Doi: 10.1186/1475-2875-12-243 [Link] [ Links ]

38 Naranjo-Díaz N, Altamiranda M, Luckhart S, Conn JE, Correa MM. Malaria vectors in ecologically heterogeneous localities of the Colombian Pacific region. PLoS One. 2014 Aug;9(8):e103769. Doi: 10.1371/journal.pone.0103769 [Link] [ Links ]

39 Couto AA, Calvosa VS, Lacerda R, Castro F, Santa Rosa E, Nascimento JM. Control of malaria transmission in a gold-mining area in Amapá State, Brazil, with participation by private enterprise. Cad Saude Publica. 2001 Jul-Aug;17(4):897-907. Doi: 10.1590/S0102-311X2001000400023 [Link] [ Links ]

40 Rocha JAM, Oliveira SB, Póvoa MM, Moreira LA, Krettli AU. Malaria vectors in areas of Plasmodium falciparum epidemic transmission in the Amazon region, Brazil. Am J Trop Med Hyg. 2008 Jun;78(6):872-7. [Link] [ Links ]

41 Branquinho MS, Araújo MS, Natal D, Marrelli MT, Rocha RM, Taveira FA, et al. Anopheles oswaldoi a potential malaria vector in Acre, Brazil. Trans R Soc Trop Med Hyg. 1996 May-Jun;90(3):233. Doi: 10.1016/S0035-9203(96)90225-4 [Link] [ Links ]

42 Hughes AL. The evolution of amino acid repeat arrays in Plasmodium and other organisms. J Mol Evol. 2004 Oct;59(4):528-35. Doi: 10.1007/s00239-004-2645-4 [Link] [ Links ]

43 Hernandez-Martinez MA, Escalante AA, Arevalo-Herrera M, Herrera S. Antigenic diversity of the Plasmodium vivax circumsporozoite protein in parasite isolates of Western Colombia. Am J Trop Med Hyg. 2011 Feb;84(2 Suppl):51-7. Doi: 10.4269/ajtmh.2011.09-0785 [Link] [ Links ]

44 González JM, Hurtado S, Arévalo-Herrera M, Herrera S. Variants of the Plasmodium vivax circumsporozoite protein (VK210 and VK247) in Colombia isolates. Mem Inst Oswaldo Cruz. 2001 Jul;96(5):709-12. Doi: 10.1590/S0074-02762001000500023 [Link] [ Links ]

45 Paskewitz SM, Brown MR, Collins FH, Lea AO. Ultrastructural localization of phenoloxidase in the midgut of refractory Anopheles gambiae and association of the enzyme with encapsulated Plasmodium cynomolgi. J Parasitol. 1989 Aug;75(4):594-600. [Link] [ Links ]

46 González-Céron L, Rodriguez MH, Santillan F, Chavez B, Nettle JA, Hernández-Avila JE, et al. Plasmodium vivax: ookinete destruction and oocyst development arrest are responsible for Anopheles albimanus resistance to circumsporozoite phenotype VK247 parasites. Exp Parasitol. 2001 Jul;98(3):152-61. Doi: 10.1006/expr.2001.4626 [Link] [ Links ]

47 Joy DA, Gonzalez-Ceron L, Carlton JM, Gueye A, Fay M, McCutchan TF, et al. Local adaptation and vector-mediated population structure in Plasmodium vivax malaria. Mol Biol Evol. 2008 Jun;25(6):1245-52. Doi: 10.1093/molbev/msn073 [Link] [ Links ]

48 Meis JFGM, Croes H, Mons B, Van Belkum A, Ponnudurai T. Localization of circumsporozoite protein in the sporogonic stages of Plasmodium vivax. Parasitol Res. 1992 Feb;78(2):165-7. Doi: 10.1007/BF00931660 [Link] [ Links ]

49 Menard R, Sultan AA, Cortes C, Altzuler R, Van Dijk MR, Janse CJ, et al. Circumsporozoite protein is required for development of malaria sporozoites in mosquitoes. Nature. 1997 Jan;385(6614): 336-40. Doi: 10.1038/385336a0 [Link] [ Links ]

Received: February 29, 2016; Accepted: June 02, 2016

Correspondence / Correspondência / Correspondencia: Erian de Almeida Santos, Cidade Nova VIII WE 42, 411. Bairro: Coqueiro, CEP: 67133-250, Ananindeua-Pará-Brazil. Phone #: +55 (91) 98140-2460. E-mail: eriansantos.bio@gmail.com

Creative Commons License This is an open-access article distributed under the terms of the Creative Commons Attribution License