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Mansonella streptocerca is a species of neglected skin-dwelling filarial nematode parasite with scarce epidemiologic data from Central Africa. We conducted a cross-sectional survey of 1,007 adults from 51 rural and semiurban communities in Gabon to update prevalence estimates and identify risk factors. Molecular analyses by quantitative PCR detected filarial DNA in 18.3% of skin snips; M. streptocerca predominated (14.2%), and Onchocerca volvulus (3.4%) occurred focally in a single rural area. Blood-dwelling parasite species such as Loa loa, M. perstans, and Mansonella sp. \"DEUX\" were rarely detected. M. streptocerca infection was 4 times more frequent in rural areas than in semiurban areas and independently associated with male sex, urticaria, and poor housing conditions. Wolbachia DNA occurred in 28% of M. streptocerca-positive samples, suggesting endosymbiosis. Our findings reveal a substantial but overlooked burden of M. streptocerca nematodes in Gabon and emphasize the need for integrated surveillance of skin-dwelling filarial infections in Central Africa.

Background Though being prevalent worldwide, Mansonella parasites are among the most neglected filarial nematodes. The true prevalence and genetic diversity of this genus have yet to be fully understood. Mansonella sp. “DEUX” is a recently described filarial nematode infecting humans and other primates in Gabon and Cameroon, although its status as distinct species has been controversial. We investigated four different polymorphic regions to further explore the genetic differences between Mansonella species and to support their status as distinct species. Methods We screened whole blood samples collected in EDTA tubes from individuals from rural areas in Gabon for mono-infections with only one Mansonella species, either Mansonella sp. “DEUX” or Mansonella perstans , as determined by quantitative polymerase chain reaction (qPCR) targeting the ITS1 region. We also included nine blood samples from Togo that had been collected as dried blood spots on 903™ protein saver cards and identified as M. perstans mono-infection. We further amplified, sequenced, and analyzed three molecular marker regions cox1 , 12S rDNA, and 28S rDNA for their potential to discriminate between the two Mansonella species. Results In total, 93 mono-infected blood samples were identified. Distinct single-nucleotide polymorphism (SNP) patterns for the two investigated Mansonella species were consistently detected in all four loci. The observed nucleotide divergences were comparable to other Onchocercidae family members. Species identification based on the ITS1 marker region was fully concordant with the SNP patterns in all samples. A complete genetic dimorphism could be observed in each of the four marker regions investigated. Conclusions The four polymorphic markers, ITS1, cox1 , 12S rDNA, and 28S rDNA, consistently demonstrated clear dimorphism between the two Mansonella species. Our results support the classification of Mansonella sp. “DEUX” as a distinct, nonrecombining Mansonella species within the Onchocercidae family. Graphical Abstract

Despite the broad distribution of M. ozzardi in Latin America and the Caribbean, there is still very little DNA sequence data available to study this neglected parasite’s epidemiology. Mitochondrial DNA (mtDNA) sequences, especially the cytochrome oxidase (CO1) gene’s barcoding region, have been targeted successfully for filarial diagnostics and for epidemiological, ecological and evolutionary studies. MtDNA-based studies can, however, be compromised by unrecognised mitochondrial pseudogenes, such as Numts. Here, we have used shot-gun Illumina-HiSeq sequencing to recover the first complete Mansonella genus mitogenome and to identify several mitochondrial-origin pseudogenes. Mitogenome phylogenetic analysis placed M. ozzardi in the Onchocercidae “ONC5” clade and suggested that Mansonella parasites are more closely related to Wuchereria and Brugia genera parasites than they are to Loa genus parasites. DNA sequence alignments, BLAST searches and conceptual translations have been used to compliment phylogenetic analysis showing that M. ozzardi from the Amazon and Caribbean regions are near-identical and that previously reported Peruvian M. ozzardi CO1 reference sequences are probably of pseudogene origin. In addition to adding a much-needed resource to the Mansonella genus’s molecular tool-kit and providing evidence that some M. ozzardi CO1 sequence deposits are pseudogenes, our results suggest that all Neotropical M. ozzardi parasites are closely related.

INTRODUCTION Mansonella ozzardi is a widely distributed filaria worm in the Amazon region. This study aimed to determine the prevalence of M. ozzardi infection in riverine communities of Lábrea municipality, Amazonas State, Brazil. METHODS A diagnostic blood filtration method in a polycarbonate membrane was used. RESULTS M. ozzardi was found in 50.3% of the sample, with the highest prevalence in farmers/fishermen (69.4%; χ 2 = -19.14, p<0.001). The prevalence was higher in longer-term residents (≥11 years; 60.2%). CONCLUSIONS M. ozzardi infection rates are high near the Purus River, much greater than those previously reported based on diagnosis using thick blood smears.

This study assessed the prevalence of Mansonella ozzardi in riverine communities of the Tefé River, Amazonas, Brazil. The prevalence of M. ozzardi was estimated by microscopic examination of thick blood smears. The M. ozzardi prevalence rate was 6.3% (19/300). Filarial infection was found in 8 of the 11 communities surveyed, with prevalence rates varying from 2.5% to 22.2%. Tefé is a region of oil and natural gas exploration, in which there is a high turnover of workers. Migration patterns may facilitate the dissemination of mansonelliasis to other regions.

Neotropical primates of the Cebidae and Callitrichidae, in their natural habitats, are frequently infected with a variety of trypanosomes including Trypanosoma cruzi, which causes a serious zoonosis, Chagas' disease. The state of trypanosome infection after a 30-day quarantine period was assessed in 85 squirrel monkeys (Saimiri sciureus) and 15 red-handed tamarins (Saguinus midas), that were wild-caught and exported to Japan as companion animals or laboratory animals, for biomedical research, respectively. In addition to many microfilariae of Mansonella (Tetrapetalonema) mariae at a prevalence of 25.9%, and Dipetalonema caudispina at a prevalence of 3.5%, a few trypomastigotes of Trypanosoma (Megatrypanum) minasense were detected in Giemsa-stained thin films of blood from 20 squirrel monkeys at a prevalence of 23.5%. Although few T. minasense trypomastigotes were found in Giemsa-stained blood films from tamarins, a buffy-coat examination detected trypanosomes in 12 red-handed tamarins (80.0%), and PCR amplification of a highly variable region of the small subunit ribosomal RNA genes (SSU rDNA) for Trypanosoma spp. detected the infection in 14 of the 15 tamarins (93.3%). Nucleotide sequences of the amplicons were identical for trypanosomes from tamarins and squirrel monkeys, indicating a high prevalence but low parasitemia of T. minasense in imported Neotropical nonhuman primates. Based on the SSU rDNA and 5.8S rDNA, the molecular phylogenetic characterization of T. minasense indicated that T. minasense is closely related to trypanosomes with Trypanosoma theileri-like morphology and is distinct from Trypanosoma (Tejeraia) rangeli, as well as from T. cruzi. Using some blood samples from these monkeys, amplification and subsequent sequencing of the glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) gene fragments detected 4 trypanosome genotypes, including 2 types of T. cruzi clade, 1 type of T. rangeli clade, and 1 T. rangeli-related type, but failed to indicate its phylogenetic position based on the gGAPDH gene. Furthermore, species ordinarily classified in the Megatrypanum by morphological criteria do not form a clade in any molecular phylogenetic trees based on rDNA or gGAPDH genes.

An unindentified microfilaria sharing characteristics with Mansonella ozzardi and Onchocerca volvulus was detected in blood samples from seven human volunteers, inhabitants of a community in the border of Amazonas and Acre State. They were detected during epidemiological studies carried out in some communities along Antimary, Acre, and Purus Rivers in the Brazilian Amazon. The most striking difference was presented in the shape of the cephalic space from this microfilaria which was different from those of M. ozzardi and with similarities to O. volvulus in this region, but no remarkable differences were observed at the caudal region. More accurate studies are being carried out in order to provide additional data and supporting evidences before establishment of a new species can be done.

Background The Onchocercidae is a family of filarial nematodes with several species of medical or veterinary importance. Microfilariae are found in the blood and/or the dermis and are usually diagnosed in humans by microscopy examination of a blood sample or skin biopsy. The main objectives of this study were to evaluate whether filariae DNA can be detected in faecal samples of wild non-human primates (NHPs), whether the detected parasites were closely related to those infecting humans and whether filarial DNA detection in faeces is associated with co-infections with nematodes ( Oesophagostumum sp. and Necato r sp.) known to cause blood loss while feeding on the host intestinal mucosa. Methods A total of 315 faecal samples from 6 species of NHPs from Cameroon and Gabon were analysed. PCRs targeted DNA fragments of cox 1 and 12S rDNA genes, to detect the presence of filariae, and the internal transcribed spacer 2 (ITS2), to detect the presence of Oesophagostomum sp. and Necator sp. infections. Results Among the 315 samples analysed, 121 produced sequences with > 90% homology with Onchocercidae reference sequences. However, 63% of the 12S rDNA and 78% of the cox 1 gene sequences were exploitable for phylogenetic analyses and the amplification of the 12S rDNA gene showed less discriminating power than the amplification of the cox 1 fragment. Phylogenetic analyses showed that the cox 1 sequences obtained from five chimpanzee DNA faecal samples from Gabon and two from Cameroon cluster together with Mansonella perstans with high bootstrap support. Most of the remaining sequences clustered together within the genus Mansonella , but the species could not be resolved. Among the NHP species investigated, a significant association between filarial DNA detection and Oesophagostomum sp. and Necator sp. infection was observed only in gorillas. Conclusions To our knowledge, this is the first study reporting DNA from Mansonella spp. in faecal samples. Our results raise questions about the diversity and abundance of these parasites in wildlife, their role as sylvatic reservoirs and their potential for zoonotic transmission. Future studies should focus on detecting variants circulating in both human and NHPs, and improve the molecular information to resolve or support taxonomy classification based on morphological descriptions.

Background Mansonella perstans is a vector-borne filarial parasite widely endemic in sub-Saharan Africa, with sporadic cases in Latin America. Infection is often overlooked; treatment is not standardized, and effectiveness of common regimes is difficult to ascertain. Anti- Wolbachia macrofilaricidal treatment with doxycycline has been applied, but there are scant and contrasting reports about the presence of Wolbachia in M. perstans isolates from different geographical locations. Taking advantage of a network of European centres expert in traveller and migrant health, we aimed to expand the knowledge concerning the distribution of Wolbachia in M. perstans to contribute to the design of optimal treatment approaches. Methods We analysed 19 samples of concentrated microfilariae or whole blood from M. perstans -infected patients who reported having resided or travelled in one or more of 10 West African countries. Wolbachia was detected by PCR targeting 16S and ftsZ genes and phylogenetic analysis of M. perstans was performed based on COX1 gene sequencing. Results Wolbachia was identified in 14/19 (74%) samples. With the possible inaccuracy deriving from potential origin of infection being identified retrospectively from routine clinical visit’s documents, this study identified Wolbachia in M. perstans from Burkina Faso, Equatorial Guinea, Republic of Guinea and Senegal for the first time to our knowledge. Furthermore, Wolbachia might also be present in M. perstans from Democratic Republic of the Congo, Mali, Niger and Nigeria. Conclusions The retrieval of Wolbachia -positive and Wolbachia -negative M. perstans samples can either be explained by technical limitations or reflect the real existence of Wolbachia -positive and Wolbachia -negative M. perstans populations. However, this latter hypothesis was not supported by our phylogenetic analysis. Our results suggest that doxycycline could be used for the treatment of M. perstans infection upfront or, if possible, after ascertaining the presence of Wolbachia by PCR performed on concentrated microfilariae using two targets to avoid false-negative results. Graphical Abstract

Background Nematodes of the Ascarididae, Ancylostomatidae and Onchocercidae families are parasites of human and veterinary importance causing infections with high prevalence worldwide. Molecular tools have significantly improved the diagnosis of these helminthiases, but the selection of genetic markers for PCR or metabarcoding purposes is often challenging because of the resolution these may show. Methods Nuclear 18S rRNA, internal transcribed spacers 1 (ITS-1) and 2 (ITS-2), mitochondrial gene cytochrome oxidase 1 ( cox 1) and mitochondrial rRNA genes 12S and 16S loci were studied for 30 species of the mentioned families. Accordingly, their phylogenetic interspecies resolution, pairwise nucleotide p-distances and sequence availability in GenBank were analyzed. Results The 18S rRNA showed the least interspecies resolution since separate species of the Ascaris , Mansonella , Toxocara or Ancylostoma genus were intermixed in phylogenetic trees as opposed to the ITS-1, ITS-2, cox 1, 12S and 16S loci. Moreover, pairwise nucleotide p-distances were significantly different in the 18S compared to the other loci, with an average of 99.1 ± 0.1%, 99.8 ± 0.1% and 98.8 ± 0.9% for the Ascarididae, Ancylostomatidae and Onchocercidae families, respectively. However, ITS-1 and ITS-2 average pairwise nucleotide p-distances in the three families ranged from 72.7% to 87.3%, and the cox 1, 12S and 16S ranged from 86.4% to 90.4%. Additionally, 2491 cox 1 sequences were retrieved from the 30 analyzed species in GenBank, whereas 212, 1082, 994, 428 and 143 sequences could be obtained from the 18S, ITS-1, ITS-2, 12S and 16S markers, respectively. Conclusions The use of the cox 1 gene is recommended because of the high interspecies resolution and the large number of sequences available in databases. Importantly, confirmation of the identity of an unknown specimen should always be complemented with the careful morphological examination of worms and the analysis of other markers used for specific parasitic groups. Graphical Abstract