Lemurinus Griseimembra Research Articles

Night Monkey Hybrids Exhibit De Novo Genomic and Karyotypic Alterations: The First Such Case in Primates.

Using molecular chromosomal analyses, we discovered night monkey hybrids produced in captivity from matings between a female Aotus azarae boliviensis (2n = 50) and a male Aotus lemurinus griseimembra (2n = 53). The parents produced seven offspring in total, including one male and six females—a pattern consistent with Haldane’s rule. Chromosomal studies were conducted on four of the hybrid offspring. Two of them showed relatively “simple” mixture karyotypes, including different chromosome numbers (2n = 51, 52), which were formed because of a heteromorphic autosome pair in the father (n = 26, 27). The other two hybrid monkeys exhibited de novo genomic and karyotypic alterations. Detailed analysis of the alterations revealed that one individual carried a mixture karyotype of the two parental species and an X chromosome trisomy (53,XXX). The second individual displayed trisomy of chromosome 18 (52,XX,+18) and a reciprocal translocation between autosomes 21 and 23 (52,XX,+18,t(21;23)). Interestingly, the second monkey exhibited mosaicism among blood cells (mos52,XX,+18[87]/52,XX,+18,t(21;23)[85]), but only a single karyotype (52,XX,+18) in skin fibroblast cells. The X- and 18-trisomies were derived from a doubling of the mother’s chromosomes in early embryonic cell division, and the reciprocal translocation likely developed in the bone marrow of the offspring, considering that it was observed only in blood cells. Such occurrence of trisomies in hybrid individuals is a unique finding in placental mammals.

Antibody responses to a novel Plasmodium falciparum merozoite surface protein vaccine correlate with protection against experimental malaria infection in Aotus monkeys.

The Block 2 region of the merozoite surface protein-1 (MSP-1) of Plasmodium falciparum has been identified as a target of protective immunity by a combination of seroepidemiology and parasite population genetics. Immunogenicity studies in small animals and Aotus monkeys were used to determine the efficacy of recombinant antigens derived from this region of MSP-1 as a potential vaccine antigen. Aotus lemurinus griseimembra monkeys were immunized three times with a recombinant antigen derived from the Block 2 region of MSP-1 of the monkey-adapted challenge strain, FVO of Plasmodium falciparum, using an adjuvant suitable for use in humans. Immunofluorescent antibody assays (IFA) against erythrocytes infected with P. falciparum using sera from the immunized monkeys showed that the MSP-1 Block 2 antigen induced significant antibody responses to whole malaria parasites. MSP-1 Block 2 antigen-specific enzyme-linked immunosorbent assays (ELISA) showed no significant differences in antibody titers between immunized animals. Immunized animals were challenged with the virulent P. falciparum FVO isolate and monitored for 21 days. Two out of four immunized animals were able to control their parasitaemia during the follow-up period, whereas two out of two controls developed fulminating parasitemia. Parasite-specific serum antibody titers measured by IFA were four-fold higher in protected animals than in unprotected animals. In addition, peptide-based epitope mapping of serum antibodies from immunized Aotus showed distinct differences in epitope specificities between protected and unprotected animals.

Observations on the Uganda I Strain of Plasmodium malariae and Plasmodium brasilianum in Aotus and Saimiri Monkeys and Anopheles Mosquitoes

Splenectomized Aotus lemurinus griseimembra, A. azarae boliviensis, A. nancymaae, A. vociferans, and Saimiri boliviensis monkeys were infected with the Uganda I/CDC strain of Plasmodium malariae. The maximum parasite counts were lower if the animals had been previously infected with Plasmodium vivax. Mosquito infection was concentrated in the 12 days following the rise in count above 1,000/microl. Mosquito infection and parasite counts were highest with A. l. griseimembra. Anopheles freeborni was more readily infected than An. gambiae, which was more readily infected than An. stephensi. Parasite counts and mosquito infection with P. brasilianum were much higher in S. boliviensis monkeys than with the Uganda I strain of P. malariae in this host, suggesting marked differences between the host-parasite-vector relationships and indicating that P. brasilianum in S. boliviensis monkeys may be a better reflection of the relationship of P. malariae in the human host.

Infection of Mosquitoes with Plasmodium falciparum by Feeding on Humans and on Aotus Monkeys

Of 1,004 positive lots of mosquitoes fed on 229 humans infected with Plasmodium falciparum, 46.2% had 1-10 oocysts/(+)gut, 21.2% had 10-30 oocysts/(+)gut, 22.2% had 30-100 oocysts/(+)gut, and 10.4% had > 100 oocysts/(+) gut. The highest levels of infection occurred between 6 and 15 days after the peak in the asexual parasite count. Of 2,281 lots of Anopheles freeborni mosquitoes fed on splenectomized Aotus monkeys infected with the Santa Lucia strain of P. falciparum, 1,191 were infected (52.2%). The highest intensity infections ranged from 2.78 oocysts per positive gut in mosquitoes fed on Aotus vociferans to 6.08 oocysts per positive gut for those fed on A. lemurinus griseimembra to 10.4 oocysts per positive gut for those fed on A. nancymaae. The pattern of infection for mosquitoes fed on splenectomized Aotus monkeys was similar to that obtained by feeding on humans, but the intensity, based on oocyst/(+)gut, was much lower.

The Santa Lucia Strain of Plasmodium falciparum in Aotus Monkeys

The Santa Lucia strain of Plasmodium falciparum was studied in 150 Aotus lemurinus griseimembra, 30 A. azarae boliviensis, 103 A. nancymaae, and 121 A. vociferans monkeys. All four of these splenectomized hosts supported the production of gametocytes infective to Anopheles freeborni mosquitoes. Transmission through sporozoites from An. freeborni, An. stephensi, An. maculatus, and An. albimanus mosquitoes was successful to all four species of Aotus on a total of 100 occasions with a median pre-patent period of 21 days. For the production of infective mosquitoes for vaccine challenge studies, A. l. griseimembra and A. vociferans were the most predictable hosts.

Studies on the Salvador I Strain of Plasmodium vivax in Non-human Primates and Anopheline Mosquitoes

A review is presented on studies conducted in New World monkeys and chimpanzees with the Salvador I strain of Plasmodium vivax. This isolate has been adapted to Aotus and Saimiri (squirrel) monkeys and developed as a model for the testing of antimalarial vaccines. After the injection of 10,000 sporozoites, the median prepatent period in S. boliviensis monkeys was 21.5 days. In 103 sporozoite-induced infections in splenectomized monkeys, the median maximum parasite count ranged from 2,139 to 202,368/microL, with a median maximum parasite count of 48,174/microL. Median maximum parasite counts in Aotus lemurinus griseimembra, A. nancymaae, A. azarae boliviensis, and A. vociferans monkeys were 19,902, 18,390, 21,420, and 18,210/microL, respectively and ranged from 124 to 156,000/microL. Mosquito infections were readily obtained in different species of Anopheles mosquitoes. The S. boliviensis monkey and Salvador I strain seems suitable for the testing of sporozoite and liver stage vaccines but not for blood-stage vaccines against P. vivax unless adapted further in spleen-intact Saimiri boliviensis monkeys.

The Chesson Strain of Plasmodium vivax in Humans and Different Species of Aotus Monkeys

Comparison was made between the parasitemia of Chesson strain Plasmodium vivax in humans and in splenectomized Aotus lemurinus griseimembra, A. nancymaae, A. vociferans, and A. azarae boliviensis monkeys. In the monkeys, 56.3% of the animals had maximum counts > 25,000/muL and in humans 59.6% were above this peak parasitemia. In humans, it took an average of 9.3 days to reach the maximum parasite count. In monkeys with no previous infections, it took an average of 18.9 days to reach the maximum parasite count; for those with previous infections, it took an average of 15 days. Human and nonhuman primate data on this parasite suggest that splenectomized Aotus monkeys, particularly A. lemurinus griseimembra, and to a somewhat lesser extent A. vociferans, can mimic the course of Chesson malaria in humans regarding parasitemia and mosquito infection.

Plasmodium inui shortii: Studies in Old World and New World Monkeys

Plasmodium inui shortti was studied in monkeys (66 Macaca mulatta, 2 M. fascicularis, 12 Saimiri boliviensis, 4 Aotus lemurinus griseimembra, and 1 A. nancymaae). Prepatent periods for 30 sporozoite transmissions by Anopheles stephensi, An. dirus, and An. maculatus mosquitoes ranged from 10 to 48 days with a median of 15.5 days. In rhesus monkeys, mean maximum parasite counts for intact animals were 181,970/muL; for splenectomized animals, the mean maximum parasite count was 1,167,890/muL.

STUDIES ON SPOROZOITE-INDUCED AND CHRONIC INFECTIONS WITH PLASMODIUM FRAGILE IN MACACA MULATTA AND NEW WORLD MONKEYS

Plasmodium fragile continues to be investigated because of its biologic similarities to the human malaria parasite, Plasmodium falciparum. Two strains of P. fragile are available for study; one strain is able to infect mosquitoes, whereas the other strain is transmissible only by blood inoculation. The Sri Lanka strain of P. fragile was transmitted to Macaca mulatta, Macaca fascicularis, Aotus lemurinus griseimembra, Aotus nancymaae, Aotus vociferans, and Saimiri boliviensis monkeys via sporozoites that developed to maturity only in Anopheles dirus mosquitoes. The prepatent periods ranged from 12 to 35 days for macaques and from 15 to 30 days for New World monkeys after intravenous injection of sporozoites. Eight rhesus monkeys were infected with the Nilgiri strain and followed for 482 days. Parasitemia in 6 animals persisted at relatively high density through the period of observation. Erythrocyte, hematocrit, and hemoglobin values reached their lowest levels 3 wk after infection and slowly recovered; however, the values did not approach preinfection levels as long as parasitemia persisted in the monkeys. The mean corpuscular volume and corpuscular hemoglobin concentration reached their peak and lowest values, respectively, at day 38 and then returned to the preinfection level. The mean corpuscular hemoglobin value decreased to its lowest level at day 87 and then returned to preinfection level.

Surgical bone marrow aspiration in Aotus lemurinus griseimembra

Aotus lemurinus griseimembra are highly susceptible to infection by human malaria parasites and reproduce some of its clinical manifestations, including anemia. We developed a new surgical technique to obtain bone marrow samples from Aotus by surgical aspiration of the femur. First, we determined that the femur offered advantages over other bones, primarily due to lower fracture vulnerability. We tested a surgical technique using 20 G IV catheters in formaldehyde-preserved animals, then conducted the procedure on 27 live animals. This technique provided easy, quick surgical access to adequate volumes of bone marrow and was safe for almost all animals: only one died; another developed nervous impairment of the lower limb. Adequate cell samples were obtained in all animals and allowed cytological studies. This procedure offers a useful tool for bone marrow research in Aotus and helps overcome current limitations of such research in human where these studies are limited by ethical and technical issues.

AOTUS NANCYMAAE AS A POTENTIAL MODEL FOR THE TESTING OF ANTI-SPOROZOITE AND LIVER STAGE VACCINES AGAINST PLASMODIUM FALCIPARUM

The Santa Lucia strain of Plasmodium falciparum was transmitted to Aotus lemurinus griseimembra, A. azarae boliviensis, A. vociferans, and A. nancymaae monkeys by bite and by intravenous inoculation of sporozoites dissected from Anopheles freeborni, An. stephensi, An. gambiae, An. albimanus, and An. maculatus mosquitoes. The data obtained from these infections indicate that A. nancymaae can be considered a suitable host model when combined with the Santa Lucia strain of P. falciparum for the testing of candidate anti-sporozoite and liver stage vaccines.

AOTUS LEMURINUS GRISEIMEMBRA MONKEYS: A SUITABLE MODEL FOR PLASMODIUM VIVAX SPOROZOITE INFECTION

This study describes a successful Plasmodium vivax sporozoite infection in Aotus lemurinus griseimembra. Twenty-eight naive or previously infected monkeys, either splenectomized or spleen intact, were inoculated intravenously or subcutaneously with Plasmodium vivax sporozoites of the Salvador I strain or with two wild isolates (VCC-4 and VCC-5; Vivax-Cali-Colombia). The monkeys were successfully infected regardless of the parasite strain, spleen presence, or inoculation route and showed prepatent periods that ranged from 16 to 89 days. Only one monkey inoculated intravenously failed to develop parasitemia. Since immune protection against malaria pre-erythrocytic forms is mediated by both helper and cytolytic T cells that may home in the spleen and P. vivax cultures are not yet available; the use of spleen-intact A. lemurinus griseimembra, susceptible to both adapted and non-adapted strains of P. vivax sporozoites, is a valuable model for evaluation of pre-erythrocytic vaccine candidates.

Coupling and decoupling of evolutionary mode between X- and Y-chromosomal red-green opsin genes in owl monkeys

We previously discovered Y-chromosomal red-green opsin genes in two types of owl monkeys with different chromosomal characteristics. In one type, the Y-linked opsin gene is a single-copy intact gene and in the other, the genes exist as multiple pseudogenes on a Y/autosome fusion chromosome. In the present study, we first distinguished the two types of monkeys as distinct allopatric species on the basis of karyotypic characteristics: Aotus lemurinus griseimembra (Karyotype III, diploid chromosome number [2 n] = 53) and Aotus azarae boliviensis (Karyotype VI; male 2 n = 49; female 2 n = 50), belonging to the northern and southern species groups, respectively, separated by the Amazon River system. Our sequence analysis revealed a common L1-Alu-Alu insertion between the two species in the 3′-flanking region of the X-linked opsin genes. The insertion was absent in the Y-linked opsin genes and in the human red and green opsin genes, indicating that it occurred in the X copy before the split into northern and southern species and after the X to Y duplication, i.e. duplication preceded speciation. We also show that in the northern species, the Y-linked opsin gene has evolved concomitantly with the X-linked copy whereas in the southern species, the Y-autosome fusion possibly led to decoupling evolutionary processes between X- and Y-linked copies and subsequent degeneration and duplications of the Y-linked opsin gene.

Observations on the Vietnam Palo Alto Strain of Plasmodium vivax in Two Species of Aotus Monkeys

Thirty-three splenectomized Aotus lemurinus griseimembra monkeys with no previous experience with malaria were infected with the Vietnam Palo Alto strain of Plasmodium vivax. The median maximum parasite count was 280,000/microl. Nine splenectomized monkeys with previous infection with Plasmodium falciparum had median maximum parasite counts of 120,000/microl. Splenectomized Aotus nancymai monkeys supported infections at a lower level. Transmission via the bites of Anopheles dirus mosquitoes was obtained in a splenectomized A. lemurinus griseimembra, with a prepatent period of 31 days. It is estimated that between 1.5 x 10(8) and 1.6 x 10(9) parasites can be removed from an infected animal for molecular or diagnostic antigenic studies.

STUDIES ON TWO STRAINS OF PLASMODIUM CYNOMOLGI IN NEW WORLD AND OLD WORLD MONKEYS AND MOSQUITOES

Infections that cause the Gombak and Smithsonian strains of Plasmodium cynomolgi were induced in Macaca mulatta, Aotus lemurinus griseimembra, Aotus nancymai, and Saimiri boliviensis monkeys. Transmission of the Gombak strain to Aotus spp. monkeys was obtained by the injection of sporozoites dissected from the salivary glands of experimentally infected Anopheles dirus and by the bites of infected An. dirus and Anopheles farauti mosquitoes. Two S. boliviensis monkeys were infected via the injection of sporozoites dissected from An. dirus. Prepatent periods in New World monkeys ranged from 14 to 44 days, with a median of 18 days. The Smithsonian strain was transmitted via sporozoites to 1 A. lemurinus griseimembra and 9 A. nancymai monkeys. Prepatent periods ranged from 12 to 31 days.

Rio Meta strain of Plasmodium vivax in New World monkeys and anopheline mosquitoes.

An archived strain of Plasmodium vivax, isolated from Rio Meta, northern Colombia, in 1972 was adapted to grow in splenectomized Aotus lemurinus griseimembra and A. nancymai monkeys. Anopheles freeborni, An. maculatus, An. dirus, An. culicifacies, and An. albimanus were shown to be susceptible to infection by feeding on infected monkeys. Infections were more readily obtained by feeding on A. L. griseimembra than on A. nancymai. Transmission through sporozoites was obtained in an A. l. griseimembra monkey after a prepatent period of 24 days.

ADAPTATION OF A STRAIN OF PLASMODIUM FALCIPARUM FROM GHANA TO AOTUS LEMURINUS GRISEIMEMBRA, A. NANCYMAI, AND A. VOCIFERANS MONKEYS

A strain of Plasmodium falciparum from Ghana was adapted to Aotus lemurinus griseimembra, A. nancymai, and A. vociferans monkeys. Gametocytes in splenectomized A. nancymai were infective to Anopheles freeborni mosquitoes. Sporozoite transmission was accomplished in two splenectomized A. nancymai with prepatent periods of 22 and 25 days. The Ghana III/CDC strain of P. falciparum is susceptible to treatment with chloroquine and mefloquine.

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.

Taxonomic status of <i>Aotus hershkovitzi</i>: its relationship to <i>Aotus lemurinus lemurinus</i>


 
 
 We analyze various taxonomic problems of northern Aotus and describe the phenotype and karyotype of specimens, which have been called Aotus hershkovitzi since their discovery. Karyotypes from Panama with a diploid number of 2n = 55–56 had been referred to as Aotus lemurinus lemurinus but actually belong to Aotus lemurinus zonalis, a taxon found throughout the Pacific lowlands of Colombia, extending to Panamá and the Río Sinú on the Caribbean coast of Colombia. Aotus lemurinus zonalis is different phenotypically and karyologically from that of the Colombian Andes population (Aotus lemurinus lemurinus). We correct Hershkovitz’ designation of the Quindío Pass as an amended type locality for Aotus lemurinus. There are no solid grounds to reject the vicinity of Santa Fe de Bogotá as type locality, and this should be listed as the correct type locality. We then point out that as the karyotype of Aotus lemurinus lemurinus has not been described and as the specimens called Aotus hershkovitzi are phenotypically within the range of other Aotus lemurinus lemurinus, that Aotus hershkovitzi is a synonym for A. lemurinus lemurinus, which should now be recognized as having a diploid karyotype of 58 as well as the highest Fundamental Number (76) known for the genus. This interpretation is supported by the discovery of an Aotus (the Murrillo specimen) with 2n = 58 from the Cordillera Central. We suggest further that the karyotypes published for Aotus lemurinus griseimembra need to be confirmed with specimens from known collection sites and that, in fact, Aotus from lowland sites of the Caribbean coast may be another taxon, yet to be described. Furthermore, we clarify the unwarranted assumption that the origin of the “Quindío specimen” described karyologically by Torres et al. (1998) is from this region, but may represent an undescribed species from an as yet unidentified locality. Finally we suggest that in fact the “subspecies” of Aotus lemurinus may in fact be full species in light of their karyological differences. In order to clarify the Aotus species it is important to do specific systematic research, including the collection of specimens from specific localities.
 
 

Adaptation of a chloroquine-resistant strain of Plasmodium vivax from Indonesia to New World monkeys.

The spread of chloroquine-resistant Plasmodium vivax from Papua New Guinea and Indonesia poses a serious health threat to areas of Southeast Asia where this species of malaria parasite is endemic. A strain of P. vivax from Indonesia was adapted to develop in splenectomized Aotus lemurinus griseimembra, Aotus vociferans, Aotus nancymai, and Saimiri boliviensis monkeys. Transmission to splenectomized Saimiri monkeys was obtained via sporozoites. Chemotherapeutic studies indicated that the strain was resistant to chloroquine and amodiaquine while sensitive to mefloquine. Infections of chloroquine-resistant P.vivax in New World monkeys should be useful for the development of alternative treatments.