Presence of a cryptic Onchocerca species in black flies of northern California, USA.

BACKGROUND: Black flies (Diptera: Simuliidae) serve as arthropod vectors for various species of Onchocerca (Nematoda: Onchocercidae) that may be associated with disease in humans, domestic animals, and wildlife. The emergence of zoonotic Onchocerca lupi in North America and reports of cervid-associated zoonotic onchocerciasis by Onchocerca jakutensis highlight the need for increased entomological surveillance. In addition, there is mounting evidence that Onchocerca diversity in North America is far greater than previously thought, currently regarded as Onchocerca cervipedis species complex. This study reports new geographic records and black fly vector associations of an uncharacterized Onchocerca species. METHODS: To better understand the biodiversity and geographic distribution of Onchocerca, 485 female black flies (2015: 150, 2016: 335) were collected using CO2-baited traps from February to October 2015-2016 in Lake County, northern California, USA. Individual flies were morphologically identified and pooled (≤ 10 individuals) by species, collection date, and trap location. Black fly pools were processed for DNA extraction, and subsequent PCR and sequencing targeting of the NADH dehydrogenase subunit 5 gene of filarioids. RESULTS: Among the pools of black flies, there were 158 individuals of Simulium tescorum (2015: 57, 2016: 101), 302 individuals of Simulium vittatum (sensu lato [s.l.]) (2015: 82, 2016: 220), 16 individuals of Simulium clarum "black" phenotype (2015: 5, 2016: 11), and 13 individuals of S. clarum "orange" phenotype (2015: 6, 2016: 7). PCR analysis revealed the percentage of filarioid-positive pools were 7.50% (n = 3) for S. tescorum, 3.75% (n = 3) for S. vittatum (s.l., likely S. tribulatum), 7.69% (n = 1) for S. clarum "black" phenotype, and no positives for S. clarum "orange" phenotype. Genetic distance and phylogenetic analyses suggest that the northern California Onchocerca isolates belong to the same species reported in black flies from southern California (average pairwise comparison: 0.32%), and seem closely related to Onchocerca isolates of white-tailed deer from upstate New York (average pairwise comparison: 2.31%). CONCLUSION: A cryptic Onchocerca species was found in Lake County, California, and may be a part of a larger, continentally distributed species complex rather than a single described species of North America. In addition, there are at least three putative vectors of black flies (S. clarum, S. tescorum, S. vittatum) associated with this cryptic Onchocerca species. A comprehensive reassessment of North American Onchocerca biodiversity, host, and geographic range is necessary.

Confirmed cases of human Onchocerca lupi infection: a systematic review of an emerging threat.

Diverse Onchocerca species are present mostly parasitizing ungulates, with the exception of Onchocerca volvulus (O. volvulus) in humans and O. lupi in canids and cats. The human cases due to the O. lupi have been more highlighted during last years. So, the present review was performed to determine the detailed characteristics of confirmed human O. lupi case reports documented worldwide. Hence, a systematic search was done using English international databases (Scopus, PubMed, Web of Science, Embase, ProQuest, and Google Scholar). Totally, 14 confirmed human cases were documented during the last decade, mostly from the USA and Turkey with 7 and 3 cases, respectively. Most cases (7 individuals) were male with the age range of 22-month-old to 54-year-old. The parasite was frequently isolated from the right eye (5 cases), followed by the left eye (4 cases), cervical spinal canal (3 cases), scalp, and right forearm (one case each). Molecular identification of the isolated agent was the preferred way of diagnosis in most cases (9 records). In conclusion, human O. lupi cases have been more highlighted in recent years, whether due to the improved diagnostics and/or host-switching phenomenon, and both veterinarians and healthcare authorities should be alerted.

Autochthonous, zoonotic Onchocerca lupi in a South Texas dog, United States.

BACKGROUND: Onchocerca lupi is an emerging, zoonotic filarioid nematode associated with ocular disease in companion animals in North America and the Old World. The areas where this parasite is assumed to be endemic in the USA comprise southwestern states. Thus far, all cases reported outside of the southwest are associated with travel or animal movement. METHODS: An 11-year-old, castrated male Pitbull dog from McAllen, Hidalgo County, southern Texas, with no travel history, was diagnosed with a perforating corneal ulceration of the right eye. Enucleation was performed and tissues submitted for histopathology. RESULTS: Histologically, sections of two filarioid nematodes were observed. DNA was extracted from formalin-fixed paraffin-embedded tissue using a commercial kit. We performed PCR targeting the cox1 gene of the mitochondrial DNA, followed by sequencing and phylogenetic analysis. Altogether, these results confirmed the identification of the nematode specimens as O. lupi, phylogenetically belonging to haplotype 1. CONCLUSION: We report the first autochthonous case of O. lupi in a dog from Hidalgo County, southern Texas, USA. Our finding suggests Texas as an additional state where this zoonotic nematode is endemic. Further investigations are required to understand the epidemiology of this parasite along the USA/Mexico border.

Prevalence of Onchocerca japonica and O. takaokai infections in the Japanese wild boar, Sus scrofa leucomystax, and the Ryukyu wild boar, S. s. riukiuanus, in Japan.

Reports of zoonotic infections with Onchocerca japonica (Nematoda: Filarioidea), which parasitizes the Japanese wild boar, Sus scrofa leucomystax, have recently increased in Japan. To predict the occurrence of infection in humans, it is necessary to determine the prevalence of O. japonica infection in the natural host animals. We investigated the presence of adult worms in the footpads, and of microfilariae in skin snips, taken from the host animals, between 2000 and 2018. Onchocerca japonica was found in 165 of 223 (74%) Japanese wild boars in Honshu and Kyushu. Among the nine regions studied, the highest prevalence of O. japonica infection was found in Oita, Kyushu, where 47 of 52 (90.4%) animals were infected. The ears were the predilection sites for O. japonica microfilariae. Adult worms of O. japonica were found more frequently in the hindlimbs than in the forelimbs of the host animals. Onchocerca takaokai was found in 14 of 52 (26.9%) Japanese wild boars in Oita. In Kakeroma Island among the Nansei Islands, both O. japonica and O. takaokai were isolated from the Ryukyu wild boar, S. s. riukiuanus. These observations could help predict future occurrences of human zoonotic onchocercosis in Japan.

Major antigen and paramyosin proteins as candidate biomarkers for serodiagnosis of canine infection by zoonotic Onchocerca lupi.

Onchocerca lupi (Spirurida: Onchocercidae) is a filarial worm parasitizing domestic carnivores and humans. Adult nematodes usually localize beneath in the sclera or in the ocular retrobulbar of infected animals, whilst microfilariae are found in the skin. Therefore, diagnosis of O. lupi is achieved by microscopic and/or molecular detection of microfilariae from skin biopsy and/or surgical removal of adults from ocular tissues of infected hosts. An urgent non-invasive diagnostic tool for the diagnosis of O. lupi in dog is mandatory. In this study, an immunoproteomic analyses was performed using a combination of immunoblotting and mass spectrometry techniques. Onchocerca lupi major antigen (Ol-MJA) and paramyosin (Ol-PARA) proteins were identified as potential biomarkers for serodiagnosis. Linear epitopes were herein scanned for both proteins using high-density peptide microarray. Sera collected from dog infected with O. lupi and healthy animal controls led to the identification of 11 immunodominant antigenic peptides (n = 7 for Ol-MJA; n = 4 for Ol-PARA). These peptides were validated using sera of dogs uniquely infected with the most important filarioids infesting dogs either zoonotic (Dirofilaria repens, Dirofilaria immitis) or not (Acanthocheilonema reconditum and Cercopithifilaria bainae). Overall, six antigenic peptides, three for Ol-MJA and for Ol-PARA, respectively, were selected as potential antigens for the serological detection of canine O. lupi infection. The molecular and proteomic dataset herein reported should provide a useful resource for studies on O. lupi toward supporting the development of new interventions (drugs, vaccines and diagnostics) against canine onchocercosis.

The Mbam drainage system and onchocerciasis transmission post ivermectin mass drug administration (MDA) campaign, Cameroon.

BACKGROUND: The impact of large scale Mass Drug Adminstration (MDA) of ivermectin on active onchocerciasis transmission by Simulium damnosum, which transmits the parasite O. volvulus is of great importance for onchocerciasis control programmes. We investigated in the Mbam river system area, the impact of MDA of ivermectin on entomological indices and also verify if there are river system factors that could have favoured the transmission of onchocerciasis in this area and contribute to the persistence of disease. We compared three independent techniques to detect Onchocerca larvae in blackflies and also analyzed the river system within 9 months post-MDA of ivermectin. METHOD: Simulium flies were captured before and after 1, 3, 6 and 9months of ivermectin-MDA. The biting rate was determined and 41% of the flies dissected while the rest were grouped into pools of 100 flies for DNA extraction. The extracted DNA was then subjected to O-150 LAMP and real-time PCR for the detection of infection by Onchocerca species using pool screening. The river system was analysed and the water discharge compared between rainy and dry seasons. PRINCIPAL FINDINGS: We used human landing collection method (previously called human bait) to collect 22,274 adult female Simulium flies from Mbam River System. Of this number, 9,134 were dissected while 129 pools constituted for molecular screening. Overall biting and parous rates of 1113 flies/man/day and 24.7%, respectively, were observed. All diagnostic techniques detected similar rates of O. volvulus infection (P = 0.9252) and infectivity (P = 0.4825) at all monitoring time points. Onchocerca ochengi larvae were only detected in 2 of the 129 pools. Analysis of the river drainage revealed two hydroelectric dams constructed on the tributaries of the Mbam river were the key contributing factor to the high-water discharge during both rainy and dry seasons. CONCLUSION: Results from fly dissection (Microscopy), real-time PCR and LAMP revealed the same trends pre- and post-MDA. The infection rate with animal Onchocerca sp was exceptionally low. The dense river system generate important breeding sites that govern the abundance of Simulium during both dry and rainy seasons.

Duplex Real-Time PCR Assay for Clinical Differentiation of Onchocerca lupi and Onchocerca volvulus.

In the United States and Europe, human onchocerciasis is a rare disease caused by zoonotic or anthropophilic parasites in the genus Onchocerca. The zoonotic species identified in focal areas of Europe and United States is Onchocerca lupi, and Onchocerca volvulus, the anthroponotic species, may be found among people who had lived in endemic areas of Africa, the Arabian Peninsula, or Latin America. Onchocerciasis due to O. lupi is an emergent parasitic disease, with limited diagnostic methods, in addition to the lack of information on its biology, transmission, and epidemiology. Cutaneous nodules are the disease's most prevalent manifestation but lack diagnostic specificity. To address the diagnosis of onchocerciasis at reference laboratories, we developed a duplex TaqMan real-time PCR (qPCR) method, targeting the cytochrome oxidase subunit I locus which has species-specific probes to identify and differentiate O. lupi from O. volvulus. We determined the performance of the duplex with a panel of 45 samples: 11 positives for O. lupi, six for O. volvulus, five samples with negative results for Onchocerca spp., and 23 non-Onchocerca nematodes. The duplex qPCR correctly detected 10 of 11 O. lupi- and six of six O. volvulus-positive specimens. The new duplex assay allowed the simultaneous detection and discrimination of O. lupi and O. volvulus in clinical specimens, expediting and facilitating the clinical diagnosis of O. lupi in non-endemic settings where the disease is an infrequent finding.

First report of Onchocerca lupi from Israel and confirmation of two genotypes circulating among canine, feline and human hosts.

Onchocerca lupi is a parasitic filarioid and the causative agent of canine ocular onchocercosis, a zoonotic disease of domestic dogs with sporadic reports in humans. A 13-year-old dog with no travel history outside of Israel was presented to an ophthalmology veterinary clinic in Israel with severe right ocular and periocular disease. After surgical exploration, thin helminths were removed from the dorsal sclera of the eye and identified as Onchocerca lupi by polymerase chain reaction according to the cytochrome c oxidase subunit I (cox1), reduced nicotinamide adenine dinucleotide dehydrogenase subunit 5 (nad5) and 12S rRNA genes. Phylogenetic trees and haplotype networks of the cox1 and nad5 genes confirmed the circulation of two genotypes: genotype 1 with worms from dogs, cats and humans from both the Old and New Worlds, and genotype 2 with specimens from Portugal and Spain. The Israeli sequences clustered in genotype 1 and were identical to O. lupi from the USA. Evidence of two genotypes separated geographically sheds light on the phylogeography and evolution of this zoonotic pathogen, and suggests a diverse pathology observed in different regions of the world.

First molecular detection of Onchocerca flexuosa (Wedl, 1856) in red deer in Slovakia.

The present paper deals with the post-mortem diagnostics of onchocerciasis and the molecular detection of causative agents of this disease in wild ruminant ungulates (Cervus elaphus, Dama dama and Capreolus capreolus). The animals were shot in hunting seasons 2017 and 2018, in two regions of the Eastern Slovakia. The total number of examined skins was fifty-eight. The presence of subcutaneous nodules was confirmed in 27.59% (95% CI 16-39) of animals. All positive skins belonged to red deer individuals (47.06%; 95% CI 30-64). The nodules were present mainly in the back area and in the lumbar area, and their sizes ranged from 2.9 to 24.1 mm, with the average count of 10 nodules per animal. Thirteen worms, isolated from the nodules collected from 13 animals, were subjected to molecular identification. Applying the standard PCR method, targeting the mitochondrial 12S rRNA, 16S rRNA and NADH-dehydrogenase gene, and subsequent sequencing, all the worms were identified as Onchocerca flexuosa Wedl, 1856. The sequences were submitted to GenBank under specific accession numbers. Two samples were identified as Onchocerca flexuosa haplotype B, in which T176A and A177T were present. Despite the presence of mutations in the 12S rRNA of the Onchocerca flexuosa, the standardized PCR remains to be a very specific and sensitive method that uses this fragment as a selectable marker for the detection of the studied parasite.

Human case of Onchocerca dewittei japonica infection in Fukushima, Northeastern Honshu, Japan.

A 73-year-old man living in Kawamata-machi, Fukushima Prefecture, Northeastern Honshu, Japan, visited a hospital with complaints of a subcutaneous swelling that had developed on the back of his left hand. The nodule was surgically removed from the vagina fibrosa tendinis of his left forefinger. Based on the histopathological characteristics, the causative agent of this nodule was identified as a female Onchocerca dewittei japonica (Spirurida: Onchocercidae). The species identification was confirmed by cox1 gene sequencing of the worm tissues from paraffin-embedded sections of the nodule. Although 11 cases of zoonotic onchocercosis have previously been recorded in Kyushu and Western Honshu, Japan, the present findings represent the first human case of infection with O. dewittei japonica in Northeastern Honshu, Japan.

Filarial worm circulation by mosquitoes along an urbanization gradient in southern Spain.

Mosquitoes are the main vectors of pathogens affecting wild animals, livestock and humans. Here, we used molecular tools to assess the local circulation of filarial parasites in mosquitoes collected during 2013 from natural, rural and urban habitats from southern Spain. We screened parasites in 22,791 female mosquitoes of the genera Aedes, Culex and Culiseta. Filarial worms were only detected in two mosquito pools. An Ae. caspius pool was positive for Setaria equina and an unidentified worm related to Onchocerca was detected in a Cx. pipiens pool. None of the mosquito pools were positive for Dirofilaria. These results underlay the role of Ae. caspius in the transmission of Setaria parasites among livestock and/or wildlife to humans in southern Spain.

A cryptic species of Onchocerca (Nematoda: Onchocercidae) in blackflies (Simulium spp.) from southern California, USA.

BACKGROUND: Entomological surveillance for pathogens based on molecular screening of putative arthropod vectors such as blackflies (Diptera: Simuliidae) is becoming increasingly important. Surveillance provides a means to understand host and geographical patterns of underestimated biodiversity among North American species of Onchocerca and a pathway to identify and track expanding emergence of the zoonotic Onchocerca lupi. Herein, we have screened two blackfly species, Simulium tescorum and Simulium vittatum (s.l.), from Los Angeles County, southern California, USA for DNA of filarioid nematodes to better understand species richness and limits within the genus Onchocerca. METHODS: A total of 1056 and 378 female blackflies was collected using CO2-baited mosquito traps from March to November of 2015 and 2016, respectively. All blackflies during 2015 were individually processed for DNA extraction and PCR targeting of the cytochrome c oxidase subunit 1 (cox1) of the mitochondrial DNA (mtDNA). Specimens of S. tescorum collected in 2016 were processed individually with heads and bodies extracted separately, whereas those of S. vittatum (s.l.) were processed in pooled samples with heads and bodies extracted separately. A subset of filarioid-positive samples from 2015 and all samples from 2016 were screened using a PCR targeting the NADH dehydrogenase subunit 5 (nad5) gene (mtDNA). RESULTS: In 2015, 356 S. tescorum (33.7%) and 683 S. vittatum (s.l.) (64.7%) were collected, and an additional 17 specimens were not assessed morphologically. In 2016, a total of 378 blackflies was collected. Of these, 43 (11.6%) were S. tescorum and 327 (88.4%) were S. vittatum (s.l.), and an additional 8 specimens were not assessed morphologically. In 2015, Onchocerca sequences were detected in 4.8% (n = 17) of S. tescorum samples, and only one S. vittatum (0.15%). In 2016, only a single S. vittatum pool was positive for the same cryptic Onchocerca species. In phylogenetic comparisons based on nad5, the Onchocerca sequences from California formed a clade with those isolates in white-tailed deer from upstate New York, suggesting these belong to a single widespread cryptic species. CONCLUSIONS: An uncharacterized species of Onchocerca associated with cervid hosts was found in blackflies from southern California. Sequence data demonstrated it is likely conspecific with an unnamed species of Onchocerca previously found in white-tailed deer from upstate New York. Current data support recognition of a broad geographical distribution across North America for an apparently cryptic species of Onchocerca that is discrete from O. cervipedis, considered to be a typical filarioid among cervids. Our data suggest that this cryptic species of Onchocerca may infect subspecies of white-tailed deer (Odocoileus virginianus), and mule and black-tailed deer (Odocoileus hemionus) at temporal latitudes. The blackflies Simulium tescorum and S. vittatum (s.l.) (presumably, S. tribulatum) are putative vectors. Discovery of a cryptic complex indicates that species diversity and putative associations for definitive hosts and vectors of Onchocerca species in North America must be reassessed.

Morphological and molecular characterization of Onchocerca fasciata (Nematoda, Onchocercidae) from dromedary camels (Camelus dromedarius) in Iran.

Skin nodules of Onchocerca fasciata Railliet and Henry, 1910 (Spirurida, Onchocercidae) are a common finding in dromedary camels, though with a minimal clinical impact. There is little information about the morphology, molecular make-up and pathological impact of this parasite. Onchocerca fasciata nodules (1.3-2.1 cm in diameter and 509-841 mg in weight) were detected on the neck region in 31.5% of dromedary camels examined in Kerman province, southeastern Iran. Of 38 isolated nodules, only 23 (60.5%) contained viable worms. Measurement and morphological analyses were performed on isolated female worms by light microscopy. The identification of O. fasciata specimens was confirmed by sequence analysis of two mitochondrial genes (12S rDNA and cox1), which showed 0.4% divergence from available O. fasciata sequences. In addition, a phylogeny of filarial nematodes was constructed, based on these two mitochondrial genes and five nuclear genes (18S rDNA, 28S rDNA, MyoHC, rbp1, hsp70); this indicated that O. fasciata belongs to clade ONC3 of Onchocercidae, with representatives of the genera Onchocerca and Dirofilaria. Within the genus Onchocerca, O. fasciata is grouped with bovine parasitic species and the human parasitic Onchocerca volvulus, which suggests an impact of domestication on the radiation of the genus. Data provided here on the distribution and morphology of O. fasciata contribute to the molecular identification and phylogenetic position of the species.

Microfilariae Onchocerca alcis Bain et Rehbinder, 1986 ­ a new parasite of moose Alces alces (L.) in Poland

Onchocerca alcis Bain et Rehbinder, 1986 belongs to the subfamily Onchocercinae. Mature nematodes of O. alcis are located on the surface of hindlimb tendons. The aim of this article was to describe the occurrence of microfilariae of O. alcis in the skin of moose from Kampinos Forest. This is the first report of O. alcis in moose from Poland and the third finding of this rare species in the world.

Filaricidal properties of Lantana camara and Tamarindus indica extracts, and Lantadene A from L. camara against Onchocerca ochengi and Loa loa.

BACKGROUND: Ivermectin is the only drug currently recommended for the treatment of onchocerciasis, the second leading infectious cause of blindness in the world. This drug kills only the first stage larvae-microfilariae (mf) of Onchocerca volvulus and is to be used cautiously in areas where Loa loa is prevalent because of severe adverse events observed with coinfected patients. METHODOLOGY/PRINCIPAL FINDINGS: This study investigated the anti-filarial activities of two Cameroonian medicinal plants, Lantana camara and Tamarindus indica locally used to treat onchocerciasis. Twelve (12) extracts were prepared and tested in vitro on the bovine model parasite, O. ochengi as well as L. loa mf. Both mf and adult male worm viabilities were assessed by motility scoring, while adult female worm viability was determined biochemically by standard MTT/formazan colorimetry. Cytotoxicity and acute toxicity were determined respectively, in monkey kidney epithelial cells and in BALB/c mice. Pure compounds were isolated by LC/MS using a bio-assay guided strategy. All the extracts showed 100% activity at 500 µg/mL against O. ochengi adult worms and mf. The highest activity against O. ochengi was observed with the hexane extract of L. camara leaves (LCLhex), with IC50 of 35.1 µg/mL for adult females and 3.8 µg/mL for the mf. Interestingly, this extract was more active against O. ochengi mf than L. loa mf. Further studies on the extracts led to the isolation of lantadene A from the methylene chloride extract of L. camara leaves, with IC50s of 7.85 µg/mL for adult males, 10.38 µg/mL for adult females, 10.84 µg/mL for O. ochengi mf and 20.13 µg/mL for L. loa mf. CONCLUSIONS/SIGNIFICANCE: We report for the first time the anti-onchocercal activities of these locally consumed medicinal plants and lantadene A, a potential lead for further development as an onchocerciasis cure.

A real-time PCR tool for the surveillance of zoonotic Onchocerca lupi in dogs, cats and potential vectors.

The ocular onchocercosis is caused by the zoonotic parasite Onchocerca lupi (Spirurida: Onchocercidae). A major hindrance to scientific progress is the absence of a reliable diagnostic test in affected individuals. Microscopic examination of skin snip sediments and the identification of adults embedded in ocular nodules are seldom performed and labour-intensive. A quantitative real-time PCR (qPCR) assay was herein standardized for the detection of O. lupi DNA and the results compared with microscopic examination and conventional PCR (cPCR). The specificity of qPCR and cPCR was assessed by processing the most common filarial nematodes infecting dogs, skin samples from O. lupi infected (n = 35 dogs) or uninfected animals (n = 21 dogs; n = 152 cats) and specimens of potential insect vector (n = 93 blackflies; n = 59 mosquitoes/midges). The analytical sensitivity of both assays was assessed using 10-fold serial dilutions of DNA from adult specimen and from a pool of microfilariae. The qPCR on skin samples revealed an analytical specificity of 100% and a sensitivity up to 8 x 10-1 fg/2µl O. lupi adult-DNA and up to 3.6 x 10-1 pg/2µl of mfs-DNA (corresponding to 1 x 10-2 mfs/2µl). Only 9.5% O. lupi-infected skin samples were positive for cPCR with a sensitivity of 8 x 10-1 pg/2µl of DNA. Out of 152 blackflies and mosquitoes/midges, eight specimens experimentally infected (n = 1 S. erythrocephalum; n = 1 S. ornatum; n = 6 Simulium sp.) were positive by qPCR. The qPCR assay herein standardized represents an important step forward in the diagnosis of zoonotic onchocercosis caused by O. lupi, especially for the detection and quantification of low number of mfs. This assay provides a fundamental contribution for the establishment of surveillance strategies aiming at assessing the presence of O. lupi in carnivores and in insect species acting as potential intermediate hosts. The O. lupi qPCR assay will enable disease progress monitoring as well as the diagnosis of apparently clinical healthy dogs and cats.

International dog travelling and risk for zoonotic Onchocerca lupi.

Onchocerca lupi is a recently recognized threat for the health of animals and humans in European, American, African and Middle Eastern countries. We describe a case of imported O. lupi infection in Italy and report the lifespan of this parasite in a non-endemic area, to advocate increased awareness of the veterinary community for this zoonotic parasitosis.

Full mitochondrial and nuclear genome comparison confirms that Onchocerca sp. "Siisa" is Onchocerca ochengi.

Onchocerca ochengi is a nodule-forming filarial nematode parasite of cattle. It is the closest known relative of the human parasite Onchocerca volvulus, with which it shares the black fly vector Simulium damnosum. Onchocerca sp. "Siisa" was described in black flies and in cattle and, based on limited mitochondrial sequence information, appeared to be about equally phylogenetically distant from O. ochengi and O. volvulus. Based on molecular genetic markers and apparent interbreeding, we later proposed that O. sp. "Siisa" belongs to the species O. ochengi. However, we did not demonstrate directly that the hybrids were fertile, and we were still unable to resolve the phylogenetic relationship of O. ochengi, O. sp. "Siisa," and O. volvulus, leaving some concerns with the conclusion mentioned above. Here, we present fully assembled, manually curated mitochondrial genomes of O. ochengi and O. sp. "Siisa," and we compare multiple individuals of these two taxa with respect to their whole mitochondrial and nuclear genomes. Based on the mitochondrial genomes, O. ochengi and O. sp. "Siisa" are phylogenetically much closer to each other than to O. volvulus. The differences between them are well within the range of what is expected for within-species variation. The nuclear genome comparison provided no indication of genetic separation of O. ochengi and O. sp. "Siisa." From this, in combination with the earlier literature, we conclude that O. ochengi and O. sp. "Siisa" should be considered one species.

A case of ocular infection with Onchocerca lupi in a dog from Germany.

Onchocerca lupi is an emerging zoonotic parasite infecting the ocular connective tissue of dogs, cats and humans. The only known case of canine ocular onchocerciasis in Germany was documented in 2002 in a shelter dog. However, the species of Onchocerca causing the infection could not be identified. Here, we report a case of the ocular infection with O. lupi in a dog, confirmed by PCR and sequencing of the cox1 gene. Further investigations are required to assess the risk factors for transmission and spread of the parasite in Germany.

Molecular Identification of Onchocerca spp. Larvae in Simulium damnosum sensu lato Collected in Northern Uganda.

Previous studies have demonstrated that the presence of larvae of other filarial species in Simulium damnosum sensu lato can distort estimates of transmission potential for Onchocerca volvulus in West Africa. However, studies conducted in foci of onchocerciasis in West Central Uganda indicated that larvae other than O. volvulus were not common in vectors collected there. Recent data collected in Northern Uganda revealed a striking discordance between estimates of the prevalence of flies carrying O. volvulus infective larvae obtained from molecular pool screening and dissection methods. To resolve this discrepancy, sequences from three mitochondrially encoded genes were analyzed from the larvae collected by dissection. All larvae analyzed were Onchocerca ochengi v. Siisa, a parasite of cattle, or Onchocerca ramachandrini, a parasite of warthogs. These results suggest that nonhuman parasite larvae are common in vectors in Northern Uganda, underscoring the necessity for molecular identification methods to accurately estimate O. volvulus transmission.