Onchocerca volvulus: genetic diversity of parasite isolates from Sudan

Onchocerciasis in Sudan exists in three distinct foci which exhibit differing clinical presentations. Previous studies have demonstrated that a tandemly repeated Onchocerca sequence family with a unit repeat length of 150 bp (the O-150 family) is a useful marker for deducing relationships among different O. volvulus populations. In the current study, the O-150 repeat families of O. volvulus from Sudan were analyzed and compared to each other and to those of parasites from West Africa. Similar to West African and American O. volvulus, the O-150 families of the Sudanese parasites could be divided into clusters within which little or no intracluster variation was evident, suggesting that the O-150 family in these parasites was subject to the forces of concerted evolution. Statistical analysis of the O-150 families from the different Sudanese parasite isolates, employing a nested algorithm based on an analysis of variance, revealed that O. volvulus endemic to the northern focus at Abu Hamed were significantly different from all other O. volvulus populations examined to date. In contrast, parasites from the southern and eastern foci of Sudan were indistinguishable from those endemic to the West African savanna. The significance of these data are discussed in light of knowledge of the biogeography and biology of transmission of O. volvulus in Africa.     

Onchocerca volvulus: application of the polymerase chain reaction to identification and strain differentiation of the parasite

Previous studies have demonstrated that the genome of Onchocerca volvulus contains a variable tandemly repeated DNA sequence family with a unit length of 150 bp. The variability of the 150-bp family has been exploited to develop O. volvulus strain and species specific DNA probes. Application of these DNA probes to the study of the epidemiologically most significant life cycle stages of the parasite has been confounded by several obstacles. These include the relative insensitivity of some of the DNA probes and the difficulty in releasing genomic DNA from infective larvae and skin microfilariae in a form that may be directly detected by hybridization to the probes. DNA sequence comparison of 18 known examples of the 150-bp repeat has been used to develop two populations of degenerate oligonucleotides. These oligonucleotides have been shown to support the amplification of the 150-bp repeat family from Onchocerca DNA, using the polymerase chain reaction. Both strain and species specific members of the repeat family are faithfully amplified, allowing characterization of a parasite on the basis of hybridization of the PCR amplification products to the previously developed DNA probes. This method is shown to be applicable to all diagnostically important forms of the parasite, including adults, infective larvae, and skin microfilariae. In addition, the method is capable of detecting O. volvulus infective larvae directly in extracts of blackfly vectors.     

Origin and diversification of the human parasite Schistosoma mansoni

Schistosoma mansoni is the most widespread of the human-infecting schistosomes, present in 54 countries, predominantly in Africa, but also in Madagascar, the Arabian Peninsula, and the Neotropics. Adult-stage parasites that infect humans are also occasionally recovered from baboons, rodents, and other mammals. Larval stages of the parasite are dependent upon certain species of freshwater snails in the genus Biomphalaria, which largely determine the parasite's geographical range. How S. mansoni genetic diversity is distributed geographically and among isolates using different hosts has never been examined with DNA sequence data. Here we describe the global phylogeography of S. mansoni using more than 2500 bp of mitochondrial DNA (mtDNA) from 143 parasites collected in 53 geographically widespread localities. Considerable within-species mtDNA diversity was found, with 85 unique haplotypes grouping into five distinct lineages. Geographical separation, and not host use, appears to be the most important factor in the diversification of the parasite. East African specimens showed a remarkable amount of variation, comprising three clades and basal members of a fourth, strongly suggesting an East African origin for the parasite 0.30-0.43 million years ago, a time frame that follows the arrival of its snail host. Less but still substantial variation was found in the rest of Africa. A recent colonization of the New World is supported by finding only seven closely related New World haplotypes which have West African affinities. All Brazilian isolates have nearly identical mtDNA haplotypes, suggesting a founder effect from the establishment and spread of the parasite in this large country.     

The relationships between the burden of adult parasites, host age and the microfilarial density in human onchocerciasis

We investigate the relationship between the microfilarial density in the skin and the burden of adult female Onchocerca volvulus by analysing pre-control nodulectomy data which allow for a direct approach, independent of exposure. The data of 169 patients in Burkina Faso and 182 patients in Liberia represent savannah and forest onchocerciasis in West Africa, respectively. Whereas in Burkina Faso, a saturating relationship between microfilarial density and worm burden suggests the operation of density-dependent processes within human hosts, the Liberian data show a linear relationship implying no density dependence. The differences may derive from differences between both parasite strains, i.e. the savannah or the forest strain of O. volvulus. Consistently for both parasite strains and independent of the worm burden, the microfilarial density increases with host age emphasising the concept of the acquisition of immunological tolerance. In male hosts in Liberia, the microfilarial density increases stronger with the worm burden than in female hosts, whereas such sex-specific differences cannot be found in Burkina Faso. In the methodological part of this investigation, we suggest the beta-distribution to be most appropriate for describing variability in microfilarial densities and we present an approach to consider the uncertainty in the adult parasite burden which cannot be determined precisely in helminth infections. Implications of density dependence are discussed with respect to immunological processes in the human host and with respect to the success of control programs. The relationships described show that regulatory processes between the parasite and the human host are multi-dimensional, operating within a high degree of biological variability.     

Infection of Aotus and Saimiri monkeys with Plasmodium gonderi

Attempts were made to infect 4 species of New World monkeys (Saimiri boliviensis, Aotus nancymai, A. vociferans, A. azarae boliviensis) with Plasmodium gonderi, a malaria parasite of African monkeys. Sporozoites were obtained from Anopheles dirus or A. stephensi mosquitoes that fed on an infected rhesus monkey (Macaca mulatta). Inoculation of sporozoites was by injection of dissected sporozoites by either the intravenous or intrahepatic routes, or by mosquito bite. Liver biopsies done 7 or 8 days after sporozoite inoculation showed that hepatocytes of all 4 species of these New World monkeys supported exoerythrocytic stages of P. gonderi, but daily blood film examination during a 60-day observation period failed to detect blood stages of the parasite.     

A preliminary analysis of the population genetics and molecular phylogenetics of Onchocerca volvulus (Nematoda: Filarioidea) using nuclear ribosomal second internal transcribed spacer sequences

Nuclear internal transcribed spacer 2 (ITS2) rDNA sequences were used for a molecular phylogenetics analysis of five Onchocerca species. The sister species of the human parasite O. volvulus was found to be the cattle parasite O. ochengi and not O. gibsoni, contrary to chromosomal evidence. The genetic differentiation of two African populations (representing the two African strains) and a Brazilian population of O. volvulus was also studied. Phylogenetic and network reconstruction did not show any clustering of ITS2 alleles on geographic or strain grounds. Furthermore, population genetics tests showed no indication of population differentiation but suggested gene flow among the three populations.     

Onchocerca volvulus: limited heterogeneity in the nuclear and mitochondrial genomes

West African populations of Onchocerca volvulus endemic to the rain forest and savanna bioclimes of West Africa differ in their ability to induce ocular disease in infected individuals. In recent years, both clinical- and animal-model-based studies have implicated particular parasite antigens in the development of ocular onchocerciasis. To test the hypothesis that the difference in pathogenic potential of blinding and nonblinding parasites might be reflected in qualitative differences in antigens that have been implicated in the development of ocular onchocerciasis, we compared the sequences of two parasite antigens implicated in the development of ocular disease in blinding- and nonblinding-strain parasites. The results demonstrated a high level of homogeneity between the parasite strains in these genes. The study was extended to include additional nuclear genes encoding antigens that are commonly recognized by individuals infected with O. volvulus and to the mitochondrial genome of the parasite. The results demonstrate a high degree of homogeneity in both the nuclear and the mitochondrial genomes among O. volvulus isolates collected from several different sites in Africa and in the Americas. This high degree of genetic homogeneity may reflect the passage of the parasite through a recent genetic bottleneck.     

Macrofilaricidal activity of tetracycline against the filarial nematode Onchocerca ochengi: elimination of Wolbachia precedes worm death and suggests a dependent relationship

Filarial nematodes are important and widespread parasites of animals and humans. We have been using the African bovine parasite Onchocerca ochengi as a chemotherapeutic model for O. volvulus, the causal organism of 'river blindness' in humans, for which there is no safe and effective drug lethal to adult worms. Here we report that the antibiotic, oxytetracycline is macrofilaricidal against O. ochengi. In a controlled trial in Cameroon, all adult worms (as well as microfilariae) were killed, and O. ochengi intradermal nodules resolved, by nine months' post-treatment in cattle treated intermittently for six months. Adult worms removed from concurrent controls remained fully viable and reproductively active. By serial electron-microscopic examination, the macrofilaricidal effects were related to the elimination of intracellular micro-organisms, initially abundant. Analysis of a fragment of the 16S rRNA gene from the O. ochengi micro-organisms confirmed them to be Wolbachia organisms of the order Rickettsiales, and showed that the sequence differed in only one nucleotide in 858 from the homologous sequence of the Wolbachia organisms of O. volvulus. These data are, to our knowledge, the first to show that antibiotic therapy can be lethal to adult filariae. They suggest that tetracycline therapy is likely to be macrofilaricidal against O. volvulus infections in humans and, since similar Wolbachia organisms occur in a number of other filarial nematodes, against those infections too. In that the elimination of Wolbachia preceded the resolution of the filarial infections, they suggest that in O. ochengi at least, the Wolbachia organisms play an essential role in the biology and metabolism of the filarial worm.     

Analysis of Wuchereria bancrofti infections in a village community in northern Nigeria: increased prevalence in individuals infected with Onchocerca volvulus

Infections with Wuchereria bancrofti causing lymphatic filariasis still represent one of the major health problems in the tropics, with 120 million people infected and over 750 million exposed to this filarial parasite. We have studied lymphatic filariasis infections as part of a multi-parasite survey in a village community in the savannah of northern Nigeria. We analysed serum samples from 341 individuals aged 5-70 years, detecting a W. bancrofti circulating antigen using the commercially available ICT Filariasis card test. The prevalence of infections was 10% and clearly age-dependent, increasing from below 2% in children to over 20% in subjects older than 40 years. Measuring IgG4 antibodies against the recombinant W. bancrofti antigen SXP1 showed that 36% of all tested individuals had been at least exposed to the parasite. Antibody levels also increased very significantly with age. A further analysis measuring Onchocerca volvulus-specific IgG4 antibodies showed a very significant association between infections with O. volvulus and those with W. bancrofti. Our data show that infections with W. bancrofti in Nigeria are still a frequently occurring health problem, since they are more prevalent than previously reported, and that individuals with an O. volvulus infection are more often infected with W. bancrofti than expected statistically.     

Sorting out the Snakebirds: The species status,phylogeny, and biogeography of the Darters (Aves: Anhingidae)

Although the avian family Anhingidae is unequivocally monophyletic, the number and relationships of the component species within the single genus (Anhinga) have long remained unclear. Here, we use extensive mitochondrial and nuclear DNA sequence data (8,878 bp) to show that four species should be recognized. Our fully resolved and well‐supported tree shows that the American Anhinga (Anhinga anhinga) is sister to the three Old World species, with the Oriental (A. melanogaster) and African (A. rufa) Darters sister within the Old World clade, which also includes the Australian Darter (A. novaehollandiae). We estimate that the divergence between the New World and Old World branches occurred 19–22 mya, with the Australian Darter separating from its Old World congeners 14–16 mya and the Oriental and African species splitting ~10 mya. The genus is yet another example of osteological conservatism in the Suliformes, which is comparable to that shown by the cormorants and shags. Nevertheless, the relationships we infer are congruent with recent plumage studies and are biogeographically plausible. We suggest that further investigation of the variation within the African and Australian Darters would be of interest.     

Phylogenetics of barbets (Aves: Piciformes) based on nuclear and mitochondrial DNA sequence data

A combination of nuclear (beta-fibrinogen intron 7; 938 bp) and mitochondrial (cytochrome b; 1045 bp) DNA sequence data analyzed with model-based phylogenetic methods yields a hypothesis of barbet and toucan relationships supported by high Bayesian posterior probabilities and several synapomorphic indels in the nuclear intron data. The basal topology differs from previous morphology and mitochondrial DNA sequence based hypotheses, but is consistent with DNA-DNA hybridization results. The mitochondrial DNA sequence data provide resolution at the tips of the tree, but when analyzed alone, yield a different basal topology than the combined data. However, the basal nodes in the mitochondrial-based tree have little statistical support. Combined data analysis produced strong statistical support at basal nodes and a relatively simple geographic structure. Barbets from each of the three tropical regions are monophyletic, but the Old World barbets are paraphyletic. The African and New World clades are sister taxa, and the Asian clade is basal. This geographic structure indicates that similarities in plumage, voice, and behavior between Gymnobucco and Calorhamphus are convergent. The data are not conclusive, but suggest paraphyly of the New World barbets. Only 4% of the Bayesian posterior distribution unites Semnornis with the other New World barbets.     

Mitochondrial DNA phylogeny of the Old-World monkey tribe Papionini.

The evolution of the Old World monkey tribe Papionini, composed of macaques, baboons, mandrills, drills, and mangabeys, was examined using mitochondrial DNA (mtDNA) sequence data on the cytochrome oxidase subunit II gene. When analyzed cladistically, these data support a baboon clade of savannah (Papio) plus gelada (Theropithecus) baboons, as well as a clade containing drill (Mandrillus) plus mangabey (Cerocebus) genera. This result stands in opposition to most morphological phylogenies, which break up the baboon clade by placing Papio and Mandrillus as sister taxa and Theropithecus as a more distantly related lineage. Analyses of COII gene sequences also suggest that the papionin ancestral stock divided into two lineages, one leading to macaques and the other to the purely African genera. From a molecular evolutionary perspective, the papionin COII gene sequences reveal a pattern of amino acid replacements concentrated in the regions spanning the mitochondrial membrane.     

JC virus as a marker of human migration to the Americas

JC virus is a ubiquitous human polyomavirus present in populations worldwide. Seven genotypes differing in DNA sequence by approximately 1-3% characterize three Old World population groups (African, European and Asian) as well as Oceania. It is possible to follow Old World populations into the New World by the JC virus genotypes they carried. The first population to settle in the Americas, the Native Americans, brought with them type 2A from northeast Asia. European settlers arriving after Columbus carried primarily type 1 and type 4. Africans brought by the slave trade carried type 3 and type 6.     

Molecular phylogenetics of blackflies of the Simulium damnosum complex and cytophylogenetic implications

The molecular phylogenetics of the Simulium damnosum complex, including vectors of the human parasite Onchocerca volvulus, from various parts of Africa was studied and compared with results of cytogenetic analyses. The sequence data of mitochondrial 16s and nuclear ITS2 rDNA revealed that the complex comprises two main clades, roughly covering the more easterly and westerly African taxa, respectively. However, striking inconsistencies in the tree topologies turned up between the DNA fragments regarding the position of certain subcomplexes and species. The cytophylogenetic relationships are better reflected in the ITS2 tree where Simulium pandanophilum and Simulium mengense constitute a basal, Central African clade of the entire complex and are therefore suggested to be the chromosomal roots too. Further divisions and the corresponding biogeographic interpretations are discussed. Several species and cytoforms are placed within the system for the first time. The phylogenetic relationships within the complex do hardly correlate with host preferences or other behavioral and ecological characteristics.     

Sequence of eruption of permanent teeth and epiphyseal union in three species of African monkeys.

Sequences of eruption of teeth and union of epiphyses determined for three species of African Cercopithecinae. These sequences are highly scalable and are highly comparable between among the species. Some sex differences in sequence were found within each species. The sequences of epiphyseal union in these species of Old World monkeys are very similar to published sequences in New World monkeys and other groups of higher primates. It appears that there is a single pattern common to the Anthropoidea as a whole.     

Phylogeny and biogeography of the genus Lycium (Solanaceae): inferences from chloroplast DNA sequences

Lycium comprises approximately 70 species and is disjunctly distributed in temperate to subtropical regions in South America, North America, southern Africa, Eurasia, and Australia. Among them, only Lycium sandwicense A. Gray sporadically occurs widely on oceanic islands in the Pacific Ocean. To investigate phylogenetic and biogeographic relationships of the genus with emphasis on L. sandwicense, the coding region of matK, the two intergenic spacers trnT (UGU)-trnL (UAA) and trnL (UAA)-trnF (GAA), and the trnL (UAA) intron of chloroplast DNA (cpDNA) were sequenced. A strict consensus tree resulting from the phylogenetic analysis indicates the following: (1) New World species comprise a potentially paraphyletic assemblage; (2) southern African, Australian, and Eurasian species together are monophyletic; (3) southern African species are a paraphyletic assemblage; and (4) L. sandwicense is in a clade with certain New World species. The estimated biogeographic events based on the cpDNA analysis indicate that (1) Lycium originated in the New World, (2) all southern African, Australian, and Eurasian species have a common ancestor from the New World, (3) Australian and Eurasian species originated once from a southern African progenitor, and (4) L. sandwicense differentiated from the New World species.     

A comparison of sexual dimorphism and range of variation inPapio cynocephalus andGorilla gorilla dentition

The dentitions of 48 baboons (Papio cynocephalus) and 242 gorillas (Gorilla gorilla) are compared metrically and the baboons are found to have a greater range of variation, and greater sexual dimorphism than the gorillas. This is explained in terms of the different ecologies of these species: life on the African savannah, with its sharp seasonal changes in available food, seems to have given selective advantage to broader niches than life in the rain forest. Further, the historic continuity of the savannah has provided fewer chances for allopatric speciation than the rain forest. These contrasts between forest and savannah speciation should provide insights into hominid evolution. In trying to judge whether australopithecines, probable savannah residents, can be lumped into one or several species, based upon dental variability, a comparison with baboons should be more informative than the now frequently used contrast with gorillas.