Adaptation of a strain of Plasmodium vivax from India to New World monkeys, chimpanzees, and anopheline mosquitoes.

A strain of Plasmodium vivax from India was adapted to develop in splenectomized Saimiri boliviensis, Aotus lemurinus griseimembra, A vociferans, A. nancymai, A. azarae boliviensis, hybrid Aotus monkeys, and splenectomized chimpanzees. Infections were induced via the inoculation of sporozoites dissected from the salivary glands of Anopheles stephensi and An. dirus mosquitoes to 12 Aotus and 8 Saimiri monkeys; transmission via the bites of infected An. stephensi was made to 1 Aotus monkey and 1 chimpanzee. The intravenous passage of infected erythrocytes was made to 9 Aotus monkeys and 4 chimpanzees. Gametocytes in 13 Aotus monkeys and 4 chimpanzees were infectious to mosquitoes. Infection rates were markedly higher in mosquitoes fed on chimpanzees. PCR studies on 10 monkeys injected with sporozoites revealed the presence of parasites before their detection by microscopic examination. The India VII strain of P. vivax develops in Aotus and Saimiri monkeys and chimpanzees following the injection of parasitized erythrocytes, or sporozoites, or both. The transmission rate via sporozoites to New World monkeys of approximately 50% may be too low for the testing of sporozoite vaccines or drugs directed against the exoerythrocytic stages. However, the strain is highly infectious to commonly available laboratory-maintained anopheline mosquitoes. Mosquito infection is especially high when feedings are made with gametocytes from splenectomized chimpanzees.

Adaptation of the AMRU-1 strain of Plasmodium vivax to Aotus and Saimiri monkeys and to four species of anopheline mosquitoes.

A chloroquine-resistant strain of Plasmodium vivax (AMRU-1) from Papua New Guinea has been adapted to grow in 4 species of Aotus monkeys (Aotus lemurinus griseimembra, Aotus vaciferans, Aotus nancymai, and Aotus azarae boliviensis), hybrid Aotus monkeys, and Saimiri boliviensis monkeys. Whereas it was possible to infect Saimiri monkeys with this parasite by inoculation of parasitized erythrocytes, only 42% of Saimiri monkeys became infected, compared to 92% of Aotus monkeys attempted. Comparative mosquito feedings showed that only A. vociferans, A. l. griseimembra, and Saimiri boliviensis monkeys produced infections in mosquitoes. Oocysts were observed on the guts of the 4 species of mosquitoes used (Anopheles gambiae, Anopheles stephensi, Anopheles freeborni, and Anopheles dirus), but sporozoite transmission was effected only with the intravenous inoculation of sporozoites from An. dirus into an A. l. griseimembra monkey.

Salvador II strain of Plasmodium vivax in Aotus monkeys and mosquitoes for transmission-blocking vaccine trials.

Infections with the Salvador II strain of Plasmodium vivax in Aotus lemurinus griseimambra monkeys were fed upon by Anopheles freeborni mosquitoes. Periods of mosquito infectivity were determined to establish a model system for the testing of transmission-blocking vaccines. The highest levels of mosquito infection were associated with the ascending asexual parasitemia after reaching 1,000/microl, and before the peak asexual parasite count. Sporozoite-induced infections were more infectious than were trophozoite-induced infections. Secondary episodes of parasitemia were also infectious, indicating the lack of development of naturally developing transmission-blocking immunity to this strain of P. vivax in splenectomized Aotus monkeys following single infections.

Adaptation of a strain of Plasmodium vivax from Mauritania to New World monkeys and anopheline mosquitoes.

A strain of Plasmodium vivax from Mauritania was adapted to develop in Aotus lemurinus griseimembra, Aotus nancymai, Saimiri boliviensis, and hybrid Aotus monkeys. Infections were induced via the inoculation of sporozoites dissected from the salivary glands of Anopheles gambiae, Anopheles freeborni, and Anopheles stephensi mosquitoes or the intravenous passage of infected erythrocytes. Infections in 3 A. lemurinus griseimembra monkeys readily infected mosquitoes. Four lines of the Mauritania parasites have been stored frozen for further reference.

Attempts to transmit the N-3 strain of Plasmodium fieldi to Aotus monkeys.

Aotus lemurinus griseimembra monkeys inoculated with parasitized erythrocytes of the N-3 strain of Plasmodiumfieldi had transient low-density parasitemia. Exoerythrocytic stages of this strain of parasite were demonstrated in sections of liver from Aotus vociferans monkeys taken 8 days after the intravenous inoculation of sporozoites dissected from the salivary glands of Anopheles dirus mosquitoes; no blood-stage infections were observed.

Infection of Anopheles freeborni mosquitoes on New World monkeys infected with the Uganda I/CDC strain of Plasmodium malariae.

Anopheles freeborni mosquitoes fed during 85 primary and 26 recrudescent infections of the Uganda I/CDC strain of Plasmodium malariae in Saimiri and Aotus monkeys were examined for the presence of oocysts. Of these, 42 primary and 14 recrudescent infections were infective. Mosquitoes were more frequently infected when fed upon A. lemurinus griseimembra animals. A retrospective examination indicated the greatest mosquito infectivity occurred before the maximum parasite count. Mosquito infection was highest 4, 5, and 6 days after the parasite count exceeded 1,000/microl. Overall, 98 of 304 positive lots (32.2%) had > or = 50% of the individual mosquitoes infected. In addition, lots of An. freeborni were fed through membranes on the blood of 34 monkeys. During the days following the parasite count reaching > or = 1,000/microl, feedings on the animals resulted in lower levels of infection than membrane feeding, thus extending the period of mosquito infection.

Adaptation of a strain of Plasmodium falciparum from a Montagnard refugee to Aotus monkeys.

A strain of Plasmodium falciparum from a Montagnard refugee was shown to produce large numbers of gametocytes in culture. Attempts were made to establish this strain in Aotus monkeys via trophozoite and sporozoite inoculation. The Montagnard S-1 strain was readily adapted to A. l. griseimembra monkeys via trophozoite inoculation. Other species of Aotus failed to support the development of high density parasitemia. None of 12 attempts to transmit the infection via sporozoites from Anopheles freeborni or An. dirus mosquitoes was successful; however, developing exoerythrocytic stages were demonstrated in hepatocytes of an A. lemurinus griseimembra monkey.

Studies on a primaquine-tolerant strain of Plasmodium vivax from Brazil in Aotus and Saimiri monkeys.

A nonimmune American acquired an infection of Plasmodium vivax Type 1 malaria in Brazil in 1994. After returning to the U.S.A., he had a primary attack followed by 3 relapses. The primary attack and first 2 relapses were treated with a standard regimen of chloroquine, followed by 14 days of primaquine (15 mg/day). Following the third relapse, the primaquine treatment was extended to 28 days. No further relapses occurred. The lack of response to primaquine by this strain may recommend it as a suitable candidate for chemotherapeutic study if it can be adapted to an animal model. Anopheles quadrimaculatus mosquitoes infected by feeding on the patient during the first relapse were used to establish the strain in Aotus and Saimiri monkeys. Monkeys supported well the development of long-lasting parasitemia. Anopheles freeborni, Anopheles stephensi, and Anopheles gambiae mosquitoes were readily infected by feeding on the monkeys and by membrane feeding on diluted blood. Monkey-to-monkey transmission was obtained via the bites of infected mosquitoes and the intravenous injection of sporozoites dissected from salivary glands. This parasite is designated as the Brazil I/CDC strain of P. vivax.

The Malayan IV strain of Plasmodium falciparum in Aotus monkeys.

Forty-nine infections with the Malayan IV strain of Plasmodium falciparum were induced in different species of Aotus monkeys. The parasite was shown to be infective to four different species of Aotus monkeys via the inoculation of parasitized erythrocytes. Sporozoite transmission was obtained to A. lemurinus griseimembra, A. vociferans, and hybrid monkeys with A. azarae boliviensis x A. nancymai and A. lemurinus griseimembra x A. nancymai parentage. Anopheles freeborni and An. stephensi mosquitoes fed readily on animals and through membranes; both supported the development of infective sporozoites. Markedly increased levels of mosquito infection were routinely obtained by membrane feeding, indicating the presence of serum factors inhibitory to infection. The Malayan IV strain appears suitable for blood-stage and transmission-blocking vaccine trials.

Plasmodium vivax infections in chimpanzees for sporozoite challenge studies in monkeys.

The development and testing of vaccines directed against Plasmodium vivax has relied on Saimiri and Aotus monkeys as the animal test system and on chimpanzees to provide infective gametocytes to produce sporozoites for monkey challenge studies and vaccine development. One sporozoite-induced and 29 blood-induced infections with the Salvador I strain of P. vivax were studied in splenectomized chimpanzees. Eighteen primary infections with P. vivax resulted in maximum parasite counts ranging from 1,519 to 81,810/ microliters (median 29,100/microliters). Twelve infections induced in animals previously infected with the homologous or heterologous strains of P. vivax had maximum parasite counts ranging from 155 to 14,136/microliters (median 1,736/microliters). A total of 202 of 237 lots containing a total of 293,175 Anopheles freeborni, An. stephensi, An. gambiae, An. dirus, An. quadrimaculatus, and An. maculatus mosquitoes were infected by membrane feeding on gametocytes from chimpanzees. Despite lower levels of parasitemia during secondary (reinfection) parasitemia, 66 of 70 lots of mosquitoes (94.3%) were infected. Based on the mean number of oocysts per positive mosquito gut, An. freeborni was more heavily infected than An. stephensi; An. stephensi was more heavily infected than An. gambiae; there was no significant difference between An. stephensi and An. dirus. Sporozoites from An. stephensi, An. gambiae, An. dirus, and An. freeborni infected with the Salvador I strain of P. vivax produced in chimpanzees were used to infect 193 Saimiri and six Aotus monkeys as well as one chimpanzee.

The Nigerian I/CDC strain of Plasmodium ovale in chimpanzees.

The chimpanzee is the only animal host currently available that can support the development of the human malaria parasite Plasmodium ovale. Thirty-one infections with the Nigerian I/CDC strain were induced in splenectomized chimpanzees. Maximum parasite counts ranged from 1,240 to 127,224/microliters. Infections were transient and unpredictable. Anopheles stephensi, Anopheles gambiae, Anopheles freeborni, and Anopheles dirus mosquitoes were infected by feeding through parafilm membranes on heparinized blood containing gametocytes; each species supported development to sporozoites in the salivary glands. Mean oocyst counts per infected mosquito ranged from 1 to 85.1; 21.7% of infected lots of mosquitoes averaged > 20 oocysts per positive mosquito gut. One infection was induced via the bites of infected An. gambiae. The prepatent period was 16 days.

The Santa Lucia strain of Plasmodium falciparum as a model for vaccine studies. I. Development in Aotus lemurinus griseimembra monkeys.

The Santa Lucia strain of Plasmodium falciparum and the Aotus lemurinus griseimembra monkey are proposed as models for the testing of sporozoite vaccines and transmission-blocking vaccines. Approximately 85% of splenectomized monkeys were infected when fed upon by 10 or more heavily infected Anopheles freeborni mosquitoes. Sporozoite-induced infections in monkeys with or without previous infection with P. vivax readily infected mosquitoes, thus making them candidates for testing transmission-blocking vaccines.

The Santa Lucia strain of Plasmodium falciparum as a model for vaccine studies. II. Development of Aotus vociferans as a model for testing transmission-blocking vaccines.

The Santa Lucia strain of Plasmodium falciparum and the Aotus vociferans monkey were studied as models for the testing of transmission-blocking vaccines. Virulence developed early in the passage history. Despite the use of only small quantities of chlorguanide and/or quinine to control infection coupled with the use of small inocula and delays in splenectomy, mosquito infection was markedly reduced from that seen during primary passage to this species of Aotus. It appears that the model may be most useful during its initial passage from the primary species, Aotus lemurinus griseimembra.

Observations on the biological nature of Plasmodium vivax sporozoites.

The relapsing malaria parasites are characterized by the production of sporozoites with varying potential for exoerythrocytic development. Some sporozoites develop soon after introduction to produce mature schizonts and merozoites that initiate the erythrocytic stage infection. Relapsing hypnozoite forms are characteristic of some strains of Plasmodium vivax and are more apt to develop late than early with many time intervals in between. Studies in Saimiri monkeys suggest another type of sporozoite-induced infection. With the Salvador I strain of P. vivax, early developing exoerythrocytic schizonts apparently release parasites with different levels of virulence for these monkeys, ranging from those producing high-level parasitemia to a more abundant avirulent form. The induction of low-density avirulent infections requires the development of more sensitive detection methods for the evaluation of sporozoite vaccines.

Sporozoite transmission of three strains of Plasmodium knowlesi to Aotus and Saimiri monkeys.

Attempts were made to infect Aotus and Saimiri monkeys with sporozoites of 3 strains of Plasmodium knowlesi to determine the potential of these animals in a monkey/malaria model. Splenectomized Saimiri and Aotus monkeys were infected with the H strain of P. knowlesi via sporozoites from Anopheles dirus mosquitoes. Prepatent periods ranged from 5 to 16 days. Saimiri monkeys infected with the Philippine strain had prepatent periods ranging from 6 to 8 days. Saimiri monkeys infected with the Hackeri strain had prepatent periods ranging from 6 to 11 days. Exoerythrocytic (EE) stages of the Philippine strain were readily demonstrated; EE stages of the H strain were less abundant. Results indicate that the Philippine strain of P. knowlesi in Saimiri monkeys has a course of parasitemia and EE stages similar to those previously seen in macaques and could serve as a reproducible model for biologic and immunologic studies.

Use of the rhesus monkey as an experimental model to test the degree of efficacy of an anti-sporozoite peptide malaria vaccine candidate combined with copolymer-based adjuvants.

Humoral response against sporozoites is not effective in protecting individuals from getting malaria. Reduction in the infectivity of sporozoites has not been quantified for most anti-sporozoite vaccines tested. Quantification requires animal models providing predictable prepatent periods, e.g., time elapsed between sporozoite inoculation and detection of parasitemia, to be used as an indicator of activity against sporozoites. A delay in prepatent period from vaccinated animals would therefore reflect a protective effect in reducing the number of parasites. We report the vaccination of rhesus monkeys with a synthetic peptide reproducing part of the repeated region of the circumsporozoite protein of Plasmodium cynomolgi. This peptide was conjugated to the carrier protein diphtheria toxoid and injected with four adjuvant formulations that differed only by the type of emulsion or immunomodulator. Because all five control animals had a synchronous prepatent period after challenge with live sporozoites, it was possible to quantify the protective efficacy for each vaccine formulation, even though all monkeys developed parasitemia. Sporozoite elimination correlated with the immunomodulator and the type of emulsion. Such elimination was related neither to antibody titer against the immunizing peptide or the whole sporozoite, nor to antibody isotype induced by the vaccine formulation.

Further studies on the sporozoite transmission of the Salvador I strain of Plasmodium vivax.

Different species of Saimiri and Aotus monkeys were inoculated with sporozoites of the Salvador I strain of Plasmodium vivax. Of 58 Saimiri inoculated, 45 developed parasitemia (4 following bites and 41 following intravenous inoculation). Prepatent periods ranged from 10 to 63 days. Twelve of 19 monkeys inoculated with sporozoites that had been stored frozen developed patent parasitemia after 16-53 days. Of 41 Aotus monkeys inoculated, only 10 (2 via bites and 8 via intravenous inoculation) developed parasitemia. One of 7 Aotus inoculated with sporozoites that had been frozen developed parasitemia with a prepatent period of 26 days. Mosquitoes were infected by feeding on gametocytes from Aotus and Saimiri monkeys, chimpanzees, and a human. Sporozoites from Anopheles stephensi, Anopheles freeborni, Anopheles dirus, and Anopheles gambiae induced infection.

Plasmodium ovale: observations on the parasite development in Saimiri monkey hepatocytes in vivo and in vitro in contrast with its inability to induce parasitemia.

Exoerythrocytic stage parasites of the human malaria parasite Plasmodium ovale were cultured in vitro by inoculating primary cultures of hepatocytes from Saimiri sciureus boliviensis monkeys with sporozoites. Morphology and size of the liver stages were similar to previous in vivo descriptions in humans and chimpanzees. Saimiri monkeys did not develop parasitemia after repeated inoculations with P. ovale sporozoites. However, liver-stage parasites were observed in liver biopsies performed 7 days after sporozoite inoculation. Together with observations on other parasite development, these results demonstrate that host specificity for many malaria parasites occurs at the blood-stage level. Lack of host specificity of primary malaria parasite species for the liver forms the basis for the close relationship existing between human and nonhuman primate malaria species.

Experimental infection of Anopheles gambiae s.s., Anopheles freeborni and Anopheles stephensi with Plasmodium malariae and Plasmodium brasilianum.

Susceptibility to infection of 2 strains of Anopheles gambiae s.s., An. freeborni and An. stephensi, was determined for 2 closely related malaria parasites, Plasmodium malariae and P. brasilianum. Neither strain of An. gambiae supported development of oocyst densities as great as the other 2 anopheline mosquitoes. The ZAN strain of An. gambiae s.s. from Zanzibar was more susceptible to infection with the strain of P. malariae from Uganda than the G-3 strain of An. gambiae s.s. from The Gambia. All species and strains of mosquitoes supported complete development to the presence of sporozoites in the salivary glands.

Development of a polymorphic strain of Plasmodium vivax in monkeys.

A strain of Plasmodium vivax from Thailand with a polymorphic repeat unit of the circumsporozoite protein was established in Saimiri sciureus boliviensis and 3 species of Aotus monkeys. All 11 attempts to transmit infection via sporozoite inoculation, 4 times to splenectomized S. sciureus boliviensis, 2 times to splenectomized Aotus nancymai, and 5 times to intact Saimiri monkeys, were successful. Anopheles freeborni, Anopheles stephensi, Anopheles dirus, and Anopheles gambiae mosquitoes were infected by feeding on parasitemic blood from a chimpanzee and an Aotus azarae boliviensis monkey. Our results indicate that this strain may be useful in antisporozoite vaccine trials.