Interaction of Malaysian sera with Plasmodium vivax sporozoite antigen

A seroepidemiologic survey of Plasmodium vivax and Plasmodium falciparum transmission was conducted in 94 Orang Asli children and adults. The prevalence of malaria was 46% in this population, and infections due to P. vivax and P. falciparum occurred with equal frequency. Multi-species infection was common, particularly in children less than 10 years of age. Circumsporozoite (CS) antibodies to P. vivax were detected by ELISA, using the recombinant protein NS181V20, in sera from 53-95% of all subjects in this study. The specificity of reactivity to NS181V20 was confirmed by immunofluorescence using air-dried sporozoites. CS antibodies to P. falciparum were present in less than 50% of the population less than 30 years of age. These data support further testing of this protein as a candidate vivax vaccine.     

Cross-reactive cellular immune response to circumsporozoite proteins of Plasmodium vivax and P. falciparum in malaria-exposed individuals

Acquired immunity against the recombinant circumsporozoite protein of P. falciparum (rPfCS) or P. vivax (rPvCS) was studied in two malarious areas of the Brazilian Amazon. Cellular responsiveness, evaluated by proliferative assays, was detected in about 45% of individuals who had recovered from recent acute malaria infections. Peripheral blood mononuclear cells of individuals whose last malaria infection was by P. vivax responded more to the rCS proteins than those who had P. falciparum . Since in P. vivax infections hypnozoites in the liver retain CS antigen, this stage may have contributed to the increased cellular response. The unexpected result was that in primoinfections by P. falciparum or P. vivax the proliferative response did not correspond to the rPfCS and rPvCS, respectively. Furthermore, among the malaria-exposed individuals, there was a positive correlation between the intensity of the responses to the two rCS proteins. Our results suggest that cross-reactive epitopes exist in the CS protein of P. falciparum and P. vivax . In the areas studied, the frequency of antibodies against rPvCS and/or rPfCS ranged from 43% to 11%. Species-specific antibodies against the CS protein were detected in the primoinfected individuals. Some individuals living in the endemic area but with no clinical history of malaria were positive by serology (8%) or by in vitro proliferation (21%). However, antibodies and cellular responses against rCS were detected only in malaria-exposed individuals, since those living outside the endemic area were all negative.     

Circumsporozoite antibody as a serologic marker of Plasmodium falciparum transmission.

In a longitudinal study of a malaria-endemic village in southeastern Thailand, circumsporozoite (CS) antibody to sporozoites of Plasmodium falciparum was measured by an enzyme-linked immunosorbent assay to determine its usefulness as a seroepidemiologic marker of malaria transmission. The CS anti-(NANP)n antibody level and prevalence during a 25-month period paralleled the pattern of seasonal transmission consistent with conventional parasitologic and entomologic measurements. The prevalence and level of antibody decreased during the non-transmission wet season, and increased over a 1-2-month transition period between the end of monsoon rains and the onset of dry conditions, an interval of maximum vector activity. Antibody increased with age in the population. The prevalence of antibody to the asexual blood stage as measured by conventional indirect fluorescent antibody assay did not coincide with changes in transmission and was sustained throughout the study period. Thus, CS antibody appeared to reflect the relative population exposure to mosquito inoculation of P. falciparum sporozoites and provided a useful measure of malaria transmission dynamics.     

High prevalence of malaria infection in Amazonas State, Venezuela

This study was carried out to determine the incidence of malaria in an endemic region of Amazonas State, Venezuela. For this, 200 random samples were collected from symptomatic and asymptomatic individuals from San Fernando de Atabapo and Santa Barbara. Epidemiological factors were related to malaria infection, which was diagnosed by microscopy observation and amplification of the 18S rDNA sequence by PCR. Malaria prevalence in these populations was 28.5%, whilst P. vivax and P. falciparum prevalences were 12 and 17%, respectively. No infection by P. malariae was found. A mixed infection was found on an asymptomatic individual. Prevalence patterns differed between age groups depending on the Plasmodium species. We found that 34.8% of the P. vivax and 15.2% of the P. falciparum infections were asymptomatic. The use of nets was helpful to prevent P. vivax infection, but did not protect against P. falciparum infection. The results suggest the presence of more than one mosquito vector in the area, displaying a differential pattern of infection for each Plasmodium species. There appear to be risk factors associated with malaria infections in some individuals. The population based approach and PCR diagnosis improved the accuracy of the statistical analysis in the study.     

The epidemiology of malaria in a population surrounding Madang, Papua New Guinea

Malaria is prevalent throughout coastal and lowland Papua New Guinea. Recent changes, including a shift from predominance of Plasmodium vivax to Plasmodium falciparum, appearance of chloroquine-resistant P. falciparum and decreased effectiveness of vector control programs have been observed. Epidemiological features of malaria were studied through four six-month surveys of a population of 16,500 in Madang Province from 1981-1983. Baseline data on parasitology, splenic enlargement, serology, hemoglobin levels, prevalence of 4-aminoquinolines, utilization of mosquito nets and incidence of fever were collected for use in future evaluation of malaria control measures including possible field trials of an antimalarial vaccine. Prevalence of parasitemia (all species, all ages) varied from 35.0% to 42.7% over the four surveys each of which covered a random sample of 25% of the population. The ratio of parasite species was: P. falciparum 70:P. vivax 25:P. malariae 5 in the dry seasons, shifting slightly in favor of P. falciparum during the wet seasons. Intense year-round transmission was indicated by decreasing parasite prevalence and splenic enlargement with age, low density asymptomatic parasitemias and high prevalence of antimalarial antibodies (i.e., greater than 80% of the population over five years of age was ELISA-positive). Levels of endemicity varied geographically, presence of 4-aminoquinolines in urine samples was relatively common (12.7% positive) and chloroquine resistance was widespread (81.6% in vitro, 46.6% in vivo).     

Infectious reservoir of Plasmodium vivax and Plasmodium falciparum malaria in an endemic region of Sri Lanka

The infectious reservoir of Plasmodium vivax and P. falciparum in a malaria endemic region in Sri Lanka was defined in a population of 3,625 by directly feeding mosquitoes on a sample of infected individuals during a period of 17 months. The malaria case incidence in this population was concurrently monitored. P. vivax gametocyte densities were highest in the youngest age groups, and decreased steadily with increasing age. However, the infectivity per gametocyte appeared to be lower in the younger age groups than in the older ones. There was no significant correlation between the age of patients and their gametocyte densities for P. falciparum, to which this population was only recently exposed, nor was there a discernible trend in the infectivity per gametocyte in different age groups. The average infectivity of patients was lowest in the youngest (0-5 years) and the oldest (greater than 50) age groups. The contribution made by P. vivax patients in the different age groups to the reservoir of infection was estimated. Patients in the 6-25 year age groups made the largest contribution to the reservoir, followed by those in the 26-50 year age group. Patients in the youngest and the oldest age groups contributed least to the infectious reservoir. When population sizes in the different age groups were taken into consideration, the age groups between 6 and 50 years contributed almost equally to approximately 87% of the infectious reservoir. The reservoir of P. falciparum malaria was very small, being confined to 9% of the patients, and this appears to be a characteristic of epidemic malaria, as was the case with P. falciparum.     

Naturally acquired circumsporozoite antibodies and their role in protection in endemic falciparum and vivax malaria

The role of naturally acquired circumsporozoite (CS) antibodies in protection against falciparum and vivax malaria was evaluated in a group of Thai endemic villagers using a prospective cohort and a case-control study design. There was no evidence of protection by either the presence of positive CS antibody levels at the presumed time of sporozoite exposure or in individuals who persistently had measurable levels of the antibodies. The study defined levels of CS antibodies that were not protective in natural infection.     

Life-spans of human T-cell responses to determinants from the circumsporozoite proteins of Plasmodium falciparum and Plasmodium vivax.

The longevity of specific human memory T-cell responses is largely unknown. However, a knowledge of the duration of memory is important for understanding immunity to an organism and for planning vaccine intervention. To address this, we have examined T-cell memory to malaria by determining T-cell responses by subjects recently exposed to peptides spanning the circumsporozoite (CS) proteins of two species of malaria-causing organisms, Plasmodium falciparum and Plasmodium vivax. Responses to vivax CS peptides by exposed Thai subjects were more frequent than responses by nonexposed individuals, permitting identification of determinants seen by vivax-induced responses. At the population level, there appears to be life-long memory, as the time since individuals were exposed did not diminish responsiveness to these determinants. In contrast, falciparum-exposed subjects were largely indistinguishable from nonexposed controls in responsiveness to falciparum CS determinants. However, a single peptide (F16: DNEKLRKPKHKKLKQPGDGN) was recognized significantly more frequently by P. falciparum-exposed than nonexposed Thai subjects. T cells responsive to this peptide were CD450+ and produced gamma-interferon. In contrast to the response to the vivax determinants and the other falciparum determinants, responsiveness to F16 was undetectable or minimal 2 years after exposure. Our data provide the average life-spans of certain malaria-specific T cells and are consistent with, but do not prove, the hypothesis that antigenic persistence (in the form of P. vivax hypnozoites) correlates with persistence of human T-cell memory.     

Epidemiological and ecological aspects related to malaria in the area of influence of the lake at Porto Primavera dam, in western São Paulo State, Brazil

A study was carried out in the area of influence of the Porto Primavera Hydroelectric Power Station, in western S?o Paulo State, to investigate ecological and epidemiological aspects of malaria in the area and monitor the profile of the anopheline populations following the environmental changes brought about by the construction of the lake. Mosquitoes captured were analyzed by standardized indicator species analysis (ISA) before and during different flooding phases (253 m and 257 m elevations). The local human population was studied by means of parasitological (thin/thick blood smears), molecular (PCR) and serological tests. Serological tests consisted of Enzyme Linked Immunosorbent Assay (ELISA) with synthetic peptides of the circumsporozoite protein (CSP) from classic Plasmodium vivax, P. vivax variants (VK247 and "vivax-like"), P. malariae and P. falciparum and Indirect Immunofluorescence Assay (IFA) with asexual forms of P. vivax, P. malariae and P. falciparum. The results of the entomological survey indicated that, although the Anopheles darlingi population increased after the flooding, the population density remained very low. No malaria, parasite infection or DNA was detected in the inhabitants of the study area. However, there was a low frequency of antibodies against asexual forms and a significant prevalence of antibodies against P. vivax, P. vivax variants, P. falciparum and P. malariae; the presence of these antibodies may result from recent or less recent contact with human or simian Plasmodium (a parallel study in the same area revealed the existence of a sylvatic cycle). Nevertheless, these results suggest that, as in other places where malaria is present and potential vectors circulate, the local epidemiological conditions observed could potentially support the transmission of malaria in Porto Primavera Lake if infected individuals are introduced in sufficient numbers. Further studies are required to elucidate the phenomena described in this paper.     

Characteristics of malaria transmission in Kataragama, Sri Lanka: a focus for immuno-epidemiological studies

Parasitological and entomological parameters of malaria transmission were monitored for 17 months in 3,625 residents in a Plasmodium vivax malaria endemic region in southern Sri Lanka; the study area consisted of 7 contiguous villages where routine national malaria control operations were being conducted. Malaria was monitored in every resident; fever patients were screened and 4 periodical mass blood surveys were conducted. An annual malaria incidence rate of 23.1% was reported during the period: 9.3% was due to P. vivax and 13.8% was due to P. falciparum; there had been a recent epidemic of the latter in this region, whereas the P. falciparum incidence rate in the previous 10 years had been negligible. There was a wide seasonal fluctuation in the malaria incidence, with the peak incidence closely following the monsoon rains. The prevalence of malaria due to both species detected at the 4 mass blood surveys ranged from 0.98% (at low transmission) to 2.35% (at peak transmission periods). Adults and children developed acute clinical manifestations of malaria. Entomological measurements confirmed a low degree of endemicity with estimated inoculation rates of 0.0029 and 0.0109 (infectious bites/man/night) for P. vivax and P. falciparum, respectively. Several anopheline species contributed to the transmission, and the overall man biting rates (MBR) showed a marked seasonal variation. Malaria at Kataragama, typical of endemic areas of Sri Lanka, thus presents characteristics of "unstable" transmission. Malaria was clustered in the population. There was a low clinical tolerance to P. falciparum malaria, to which most had only been at risk, compared to P. vivax, to which most had had a life-long exposure.     

Significance of circumsporozoite-specific antibody in the natural transmission of Plasmodium falciparum, Plasmodium vivax, and Plasmodium malariae in an aboriginal (Orang Asli) population of central peninsula Malaysia

Two hundred and seventy-five Orang Asli volunteers living in nine villages in the Pos Legap Valley of Perak State, peninsular Malaysia, participated in a prospective study designed to characterize the epidemiological, parasitological, and entomological characteristics of Plasmodium falciparum, P. vivax, and P. malariae malaria transmission. Prevalence rates for the three plasmodial species at initiation of the study ranged from 56% in the 0-4-year-old age group to 0% in individuals over the age of 40. Entomological surveys were conducted, enabling us to determine mosquito salivary gland-positive rates and entomological inoculation rates of 1.2 infectious mosquito bites per person per month for P. falciparum, 2.4 for P. vivax, and 0.3 for P. malariae. Cumulative incidence rates over the 16 weeks of the study, following radical cure of all volunteers, were 22.5% for P. falciparum, 12.7% for P. vivax, and 1.5% for P. malariae. The median baseline antibody titer against the immunodominant repetitive B cell epitope of P. falciparum or P. vivax circumsporozoite protein was significantly higher for volunteers who did not become parasitemic. Volunteers were selected for further study if they had evidence of being challenged with P. falciparum sporozoites during the study, based on a two-fold or greater increase in antibody titer against the immunodominant repetitive B cell epitope of the circumsporozoite protein. Resistance to infection was seen in six of 10 individuals who had high (greater than 25 OD units) baseline ELISA titers, compared with only three of 24 individuals who had low baseline ELISA titers (chi 2 P less than 0.02). A similar analysis for P. vivax did not show a significant correlation.     

Antibodies against malaria sporozoites in patients with acute uncomplicated malaria and patients with cerebral malaria

Serum samples from 95 patients with acute uncomplicated falciparum malaria (AM) and 95 patients with cerebral malaria (CM) were tested by the indirect immunofluorescent assay (IFA) for IgG and IgM antibodies against Plasmodium falciparum and P. vivax sporozoites. Forty-six (48%) CM patients were positive for antibodies against P. falciparum sporozoites whereas only 23 (24%) were positive for antibodies against P. vivax sporozoites (P less than 0.002). A similar result was obtained in AM patients. However, CM patients had significantly lower mean IgG anti-sporozoite titer for P. falciparum than did AM patients (P less than 0.05), especially when only anti-sporozoite antibody-positive CM and AM patients were compared (P less than 0.0005), suggesting that CM patients had relatively less exposure and were probably less immune to malaria than were AM patients. The persistence of anti-sporozoite antibodies also was investigated in paired sera taken 63 days apart from 108 patients with acute falciparum malaria. There were significant decreases in the mean antibody titers in the follow-up sera during the period of stay in the malaria-free area. It was proposed that determination of anti-sporozoite antibody be made as a substitute for, or in addition to, anti-blood stage antibody for seroepidemiological study of malaria, especially in the monitoring of the success of the malaria control program.     

Malaria during pregnancy and infancy, in an area of intense malaria transmission in central India

The clinico-epidemiological pattern of malarial infection in a cohort of pregnant women and infants was analysed during a malaria epidemic (1997-1998). The subjects were all members of tribal communities in an isolated and almost inaccessible area of central India. Overall, 151 (55%) of the 274 pregnant women investigated were found to have malarial infections at some time during the study, with Plasmodium falciparum predominating (88% of infections). All of the women investigated, whether primigravidae (42% found infected), secundigravidae (68%) or multigravidae (54%), were at great risk of developing severe malaria. When trimesters were compared, the highest prevalence of P. falciparum infection was recorded in the second (59% infected), irrespective of parity. Of the women found infected with P. falciparum, 3% had abortions, 4% stillbirths and 2% had babies who died while neonates. The small number of P. vivax infections observed prevented similar analyses for this species of parasite. Malarial infection was also seen in 218 (41%) of the 535 infants investigated. The values of age-specific prevalences revealed that > 30% of the infants examined at 2 months of age were then found to have P. vivax and/or P. falciparum parasitaemias. At 1 year of age, overall malaria prevalence was 50%, with P. vivax representing 25% of the infections and P. falciparum the rest. Subsequent follow-up revealed that three of the infants investigated, each of whom had had P. falciparum infections previously, died before their first birthdays. Re-infections (or treatment failures) were found to be common, both in the infants and the pregnant women. Pregnant women and infants from the study area clearly require systematic intervention to reduce their malaria-attributable morbidity.     

Epidemiology of malaria in a hypoendemic Brazilian Amazon migrant population: a cohort study

The present study describes aspects of the epidemiology of malaria in a migrant population living in a hypoendemic area in Brazil using an open cohort study design. Rural settlement residents in Leonislandia, Peixoto de Azevedo, Mato Grosso, Brazil were followed from September 1996 to April 1997. At baseline, an interview and malaria diagnoses were carried out and spleen size was measured. Incident cases were detected through follow-up visits and laboratory records. Cox regression was used to assess risk factors for time to malaria onset. Eighty percent (n = 414) of the study population (n = 521) contributed follow-up data. Overall, malaria prevalence during any study visit ranged from 0.3% to 5.4% and the malaria incidence rate (IR) was 4.49 (95% confidence interval = 3.66, 5.46) per 100 person-months. The IR of Plasmodium vivax malaria was approximately four times higher than the IR for P. falciparum malaria during follow-up. Among individuals who had had malaria during his or her lifetime, 14.03% reported hospitalization (median duration = 3 days) and 70.1% reported days of work lost (median duration = 4 days for P. falciparum malaria and 3 days for P. vivax malaria) related to the last malaria episode. No important risk factor was associated with the malaria IR. The fact that neither work-related factors nor age was associated with the risk of malaria indicates that indoor/peri-domiciliary transmission by the local vector is more important or as important as workplace-related transmission.     

Antibodies to Plasmodium falciparum and Plasmodium vivax merozoite surface protein 5 in Indonesia: species-specific and cross-reactive responses

BACKGROUND: Merozoite surface protein (MSP) 5 is a candidate antigen for a malaria vaccine. In cross-sectional and longitudinal studies, we measured MSP5 antibody responses in Papuans with acute Plasmodium falciparum malaria, Plasmodium vivax malaria, and mixed P. falciparum and P. vivax malaria and in those with past exposure. METHODS: Enzyme-linked immunosorbant assay (ELISA) was used to quantitate antibody responses to P. falciparum MSP5 (PfMSP5) and P. vivax MSP5 (PvMSP5) in 82 subjects with P. falciparum infection, 86 subjects with P. vivax infection, 85 subjects with mixed infection, and 87 asymptomatic individuals. Longitudinal responses through day 28 were tested in 20 persons. Cross-reactivity was tested by competition ELISA. RESULTS: PfMSP5 or PvMSP5 immunoglobulin (Ig)Gwas detected in 39%-52% of subjects, and IgM was detected in 44%-72%. IgG responses were distributed equally between IgG3 and IgG1 for PfMSP5 but were predominantly IgG3 for PvMSP5. Although IgG responses were generally specific for PfMSP5 or PvMSP5, cross-species reactivity was found in 7 of 107 dual-positive responders. No significant difference was seen in the magnitude, frequency, or subclass of PfMSP5 or PvMSP5 IgG antibodies between groups. There was no significant association between antibody responses and therapeutic response. CONCLUSION: PfMSP5 and PvMSP5 were frequently recognized by short-lived, species-specific antibodies. Although infrequent, the cross-reactive MSP5 antibodies indicate that an appropriately formulated vaccine may elicit and/or enhance cross-species recognition, which may be very useful in areas where both parasites are endemic.     

Characteristics of natural antibody responses to the circumsporozoite protein of Plasmodium vivax

The antibody response to the prototype circumsporozoite (CS) protein of Plasmodium vivax (CSPV) was studied in Thai soldiers experiencing occupational malaria. Seventy-four (65%) of 114 men followed during assignment to a malaria transmission area developed blood-stage infection with P. vivax. IgG antibodies against the central repeat region of the CSPV protein were quantitated by ELISA using the recombinant protein, NS181V20, as the capture antigen. One quarter of the subjects had detectable anti-CSPV antibodies at the beginning of the study. CSPV antibody seroconversion was documented in 16 of 26 subjects assessed during their first observed episodes of vivax malaria. This antibody response was of moderate magnitude, fell off after the first week post-diagnosis and appeared, at the low levels observed, to be unassociated with protection. Continued assessment of anti-CSPV antibody after subjects left the transmission area found no increase associated with release of P. vivax. These findings indicate that CS antibody responses to P. vivax during occupational malaria share many characteristics with responses to P. falciparum.     

Seroepidemiology of malaria in northern Thailand: II. Focal and temporal variations in endemicity

Four population groups from regions of Northern Thailand were surveyed for the presence of antibodies to Plasmodium falciparum using the indirect immunofluorescent antibody (IFA) test. Each of the four populations was selected from areas known to represent different patterns of malaria transmission. Group 1 was from an area where there had been no malaria transmission for approximately 30 years. Individuals in this group below age 40 showed an extremely low prevalence of malaria antibodies. Group 2 was chosen from an area where low levels of transmission have continued despite more than 30 years of DDT spraying. In this group the age related pattern of malaria antibodies varied from village to village but in all villages there was a sharp increase in the prevalence of IFA positive tests in individuals over 30. The third group has had continuously high levels of transmission. Although there are differences in the age related prevalence of IFA positives when individual villages are compared, there is a greater prevalence at all ages than in the first two groups. The fourth group was selected from an area where transmission had recently resumed after freedom from indigenous cases for approximately six years. There was little difference in the prevalence of IFA positive individuals below the age of 25 but above that there was a steady increase in prevalence with age. The correlation of IFA antibody positives with known patterns of malaria transmission in these four areas demonstrates the usefulness of this serological technique in assessing malaria endemicity and the effectiveness of control measures as well as in the interpretation of other malaria statistics.     

Antibodies against circumsporozoite proteins of Plasmodium falciparum induced by natural infection

Sera from 10 individuals who lived in a malaria endemic area, 10 patients with acute uncomplicated falciparum malaria and 10 patients with cerebral malaria and hyperimmune mouse serum were tested for their reactivities against Plasmodium falciparum sporozoite antigens by Western blot analysis using 125I-labeled staphylococcal protein A as the detecting reagent. These sera were shown by indirect immunofluorescence and/or circumsporozoite precipitation test to have antibodies reacting against the parasites. It was found that all serum antibodies from the three groups of individuals and the mouse serum reacted in a similar pattern with circumsporozoite (CS) proteins of P. falciparum. Ten sera from normal individuals were negative in all reactions. Monoclonal antibody (MAB) specific against CS proteins of the parasites showed that the proteins exhibited as four different molecular weight (MW) polypeptides, i.e., 67,000, 65,000, 60,000, and 58,000 daltons. These CS proteins of P. falciparum were found to be species and stage specific. Radioimmunoprecipitation using 35S-methionine-labeled parasites and sera of individuals from the various categories or MABs gave a similar result. Another protein antigen of P. falciparum sporozoites had a MW of 80,000 daltons. This antigen was not species specific, probably not membrane associated and was present in a minute quantity in the parasite's extract.     

Low CD8+ T lymphocyte response to P. falciparum circumsporozoite protein in naturally-exposed malaria endemic populations in Thailand

Cytotoxic T lymphocytes (CTLs) specific for epitope(s) within the circumsporozoite (CS) protein of malaria sporozoite have been shown to play an important role in protective immunity against malaria. Human CTLs against the potential epitope at the carboxy terminal region of CS protein of Plasmodium falciparum 7G8 strain (Pf7G8CS 368-390) were determined in thirty-six falciparum malaria patients and ten healthy controls. Four of 36 individuals and none of the healthy controls developed Pf7G8CS 368-390 specific CTL activity. The CTL activity was antigen specific and CD8+ T cell dependent. Although low CTL response has been determined, the study suggested that there was a correlation between initial parasitemia and the specific Pf7G8CS 368-390 CTL activity. A correlation between such CTL activity and anti-R32tet32 antibody levels among individuals with previous malaria experience was found, which was in contrast to those among individuals with recent malaria infection. All these 4 CTL positive individuals had at least two episodes of clinical malaria experience while all 25 individuals who were exposed to malaria for the first time did not have such a specific CTL response. These results showed that individuals with a history of natural endemic exposure to P. falciparum sporozoite developed low specific CTL responses to Pf7G8CS 368-390, so that previous but recent sporozoite exposure might be a prerequisite for generation of such CS protein specific CTL response.     

Identification of HLA-A2 restricted CD8(+) T-lymphocyte responses to Plasmodium vivax circumsporozoite protein in individuals naturally exposed to malaria

Specific CD8(-) T-lymphocyte (CTL) activity against Plasmodium pre-erythrocytic stages (P-ES) derived antigens is considered one of the most important mechanisms for malaria protection. Plasmodium vivax is the second most prevalent human malaria parasite species distributed worldwide. Although several CTL epitopes have been identified in Plasmodium falciparum P-ES derived antigens, none has been described for P. vivax to date. In this study, we analysed HLA-A*0201 specific CD8(-) T-lymphocyte responses to the P. vivax circumsporozoite (CS) protein in both malaria exposed and non-exposed populations from the Colombian Pacific Coast. First, we analysed the prevalence of HLA-A2 allele in the study populations and found that approximately 38 of the individuals expressed this molecule and that 50 of them were HLA-A*0201. We then selected, on the P. vivax CS, five peptide sequences containing the HLA-A*0201 binding motifs and used the corresponding synthetic peptides to evaluate the CD8(-) T-lymphocyte interferon (IFN)-gamma response. Peripheral blood mononuclear cells from the HLA-A*0201 donors were in vitro stimulated with these peptides and IFN-gamma production was determined by an ELISPOT assay. Specific CD8(-) T-lymphocyte responses were detected for three peptides located in the C-terminal region of the protein. Specific responses to these peptides were also detected in several individuals expressing different HLA-A*02 subtypes. The potential of these peptides to induce specific cytolysis and that of long synthetic peptides comprising these epitopes as P. vivax malaria vaccine subunits are being studied.