Microscopic and molecular detection of Deraiophoronema evansi (Lewis, 1882) in domestic Bactrian camels (Camelus bactrianus) of Mongolia.

Cameline filarosis is an important parasitic disease having an economic impact on the camel industry around the world. However, there has been no study on filarosis in Bactrian camels of Mongolia. Therefore, the aim of the present study was to detect and identify microfilariae of Deraiophoronema evansi (D. evansi) in Bactrian camels from three provinces, located in southern and southwestern Mongolia. Blood samples were obtained from 400 healthy two-humped camels of different ages and both sexes. All blood samples were analysed using a variety of diagnostic techniques. Microfilariae were detected in 30 Bactrian camels (7.5%) by the Knott technique, while 13 Bactrian camels (3.3%) tested positive in a direct smear test. D. evansi was detected in 18 Bactrian camels (4.5%) by PCR assay. Prevalence was shown to be high among Bactrian camels in the age group up to 5 years, while the lowest positive results were obtained for Bactrian camels in the 5-10-year age group and the over 10-year age group. To confirm the morphological identification, D. evansi-COI gene sequences were subjected to phylogenetic analyses. The D. evansi-COI gene sequences from Mongolian two-humped camels were identical to sequences from Iranian one-humped camels and were clustered together with these sequences in the phylogeny. This is the first report of molecular detection and identification of microfilariae of D. evansi in Bactrian camels of Mongolia.

Dipetalonema graciliformis (Freitas, 1964) from the red-handed tamarins (Saguinus midas, Linnaeus, 1758) in French Guiana.

Six Dipetalonema species have been reported from Neotropical monkeys, Dipetalonema gracile, Dipetalonema graciliformis and Dipetalonema caudispina being the dominant species found in French Guiana primates. Adult filarioids isolated from the abdominal cavity of tamarins (Saguinus midas) in French Guiana were morphologically and molecularly identified as D. graciliformis. Phylogenetic analysis based on DNA and amino acid sequences of the cox1 gene as well as the concatenated sequences of the cox1 and the 18S genes indicated that D. graciliformis belongs to the clade 4 (ONC4) of Onchocercidae. Blast analysis of the 18S rDNA revealed that D. graciliformis in the studied tamarins is conspecific with the filarioid circulating in howler monkeys (Alouatta macconnelli) in French Guiana, previously referred to as unidentified Onchocercidae species.

Co-infection with filarial nematodes in Sapajus macrocephalus and Cebus albifrons (Primates: Cebidae) from the Peruvian Amazon.

Dipetalonema caudispina (Molin, 1858) and D. gracile (Rudolphi, 1809) (Filarioidea: Onchocercidae) are two of six known species of filarial nematodes that parasitize Neotropical non-human primates. Adult filariae were collected from the thoracic and abdominal cavities of 38 of 44 specimens of Sapajus macrocephalus (Spix, 1823) and nine of ten specimens of Cebus albifrons (Humboldt, 1812) (Primates: Cebidae), distributed in the Yavarí-Mirín river basin and used locally for human consumption. Co-occurrence of D. caudispina and D. gracile is reported for the first time, with a prevalence of 18.5% (10 of 54 hosts examined). Our finding of D. caudispina and D. gracile in cebids from the Peruvian Amazon constitutes a new geographical record for both filariae, two new host records for D. caudispina, and the first report of D. gracile in S. macrocephalus. In addition, we provide morphometric data for D. caudispina, complementing the original description, as well as scanning electron microscopy details on the structure of the area rugosa and number of caudal papillae in males.

The mitochondrial genome of Dipetalonema gracile from a squirrel monkey in China.

Dipetalonema gracile is a common parasite in squirrel monkeys (Saimiri sciureus), which can cause malnutrition and progressive wasting of the host, and lead to death in the case of massive infection. This study aimed to identify a suspected D. gracile worm from a dead squirrel monkey by means of molecular biology, and to amplify its complete mitochondrial genome by polymerase chain reaction (PCR) and sequence analysis. The results identified the worm as D. gracile, and the full length of its complete mitochondrial genome was 13,584 bp, which contained 22 tRNA genes, 12 protein-coding genes, two rRNA genes, one AT-rich region and one small non-coding region. The nucleotide composition included A (16.89%), G (20.19%), T (56.22%) and C (6.70%), among which A + T = 73.11%. The 12 protein-coding genes used TTG and ATT as start codons, and TAG and TAA as stop codons. Among the 22 tRNA genes, only trnS1AGN and trnS2UCN exhibited the TΨC-loop structure, while the other 20 tRNAs showed the TV-loop structure. The rrnL (986 bp) and rrnS (685 bp) genes were single-stranded and conserved in secondary structure. This study has enriched the mitochondrial gene database of Dipetalonema and laid a scientific basis for further study on classification, and genetic and evolutionary relationships of Dipetalonema nematodes.

Occurrence of Dipetalonema gracile in a wild population of woolly monkey Lagothrix poeppiigii in the northeastern Peruvian Amazon / Ocorrência de Dipetalonema gracile em uma população selvagem de macaco barrigudo Lagothrix poeppigii no nordeste da Amazônia peruana

Abstract Dipetalonema gracile (Rudolphi, 1809) (Filarioidea: Onchocercidae) is one of six species of cavities filarial parasites of Neotropical non-human primates. The present study recorded the occurrence of D. gracile, provides morphological and morphometric data and extends the geographical distribution. Adult filariae were obtained from the thoracic and abdominal cavities of 38 specimens of woolly monkey, which were used for local human consumption, in the northeastern Peruvian Amazon. Male and female filarids were processed and analysed using light and scanning electron microscopy. Details of the cephalic papillae, post-cloacal bands and papillae, vulva, phasmid position and lateral appendages are showed by scanning electron microscopy and is recorded the occurrencce of Lagothrix poeppigii monkey as a new host of this filaria in the Yavari-Mirin river basin, Peruvian Amazon.

Resumo Dipetalonema gracile (Rudolphi, 1809) (Filarioidea: Onchocercidae), é uma das seis espécies de filarias parasitas de primatas não humanos neotropicais. O presente trabalho registra a ocorrência de D. gracile , proporciona dados morfológicos e morfométricos e amplia sua distribuição geográfica. Filárias adultas foram obtidas da cavidade torácica e abdominal de 38 espécimes de macaco barrigudo, que foram utilizados para consumo humano local. Essa espécie de primatas não humanos neotropicais tem distribuição ao nordeste da Amazônia peruana. Filarídeos machos e fêmeas foram processados e analisados pela microscopia de luz e microscopia eletrônica de varredura. Detalhes das papilas cefálicas, bandas e papilas pós-cloacais, vulva, posição dos fasmídeos e apêndices laterais são mostradas pela microscopia eletrônica de varredura e registra-se a ocorrência do macaco Lagothrix poeppigii como novo hospedeiro desta filaria na bacia do rio Yavari-Mirin na Amazônia peruana.

Occurrence of Dipetalonema gracile in a wild population of woolly monkey Lagothrix poeppiigii in the northeastern Peruvian Amazon.

Dipetalonema gracile (Rudolphi, 1809) (Filarioidea: Onchocercidae) is one of six species of cavities filarial parasites of Neotropical non-human primates. The present study recorded the occurrence of D. gracile, provides morphological and morphometric data and extends the geographical distribution. Adult filariae were obtained from the thoracic and abdominal cavities of 38 specimens of woolly monkey, which were used for local human consumption, in the northeastern Peruvian Amazon. Male and female filarids were processed and analysed using light and scanning electron microscopy. Details of the cephalic papillae, post-cloacal bands and papillae, vulva, phasmid position and lateral appendages are showed by scanning electron microscopy and is recorded the occurrencce of Lagothrix poeppigii monkey as a new host of this filaria in the Yavari-Mirin river basin, Peruvian Amazon.

Molecular identification and phylogenetic analysis of Dipetalonema evansi (LEWIS, 1882) in camels (Camelus dromedarius) of Iran.

Despite the economic importance of camels, the parasites that affect them have not received adequate attention so far and molecular studies are scarce compared to other livestock. In this study, we characterized peripheral blood microfilariae in 200 healthy one-humped camels (Camelus dromedarius) from south-east Iran by microscopy and molecular tools to receive a more detailed insight into prevalence and species that affect them. Moreover, adult specimens of the filarial nematode Dipetalonema evansi were collected from the carcass of an infected animal. Microscopic examination was performed on Giemsa-stained blood smears, and blood was also spotted on Whatman FTA(®) cards for DNA analysis. Genomic DNA was extracted, and PCR was carried out for the detection of filaroid helminths, followed by sequence analysis of positive samples. Four samples were positive for microfilariae by microscopy, while 16 animals (8 %) were positive by PCR. Sequence analysis revealed D. evansi in all cases. Phylogenetic analysis of a cytochrome C oxidase subunit I (COI) sequence of filaroid nematodes showed that most species in a single genus cluster in the same clade; however, D. evansi and D. gracile are not monophyletic and branch rather at the base of the tree. Further studies on the life cycle of D. evansi, specifically the identification of intermediate host(s), have become feasible with the provision of the first specific COI sequences in this study.

Dipetalonema evansi infection in camels of Iran's central area.

Totally 294 dromedary camels of different ages and both sexes slaughtered at slaughterhouses in Yazd, Isfahan and Kerman provinces were inspected for infection with Dipetalonema evansi. Blood smears of all camels and carcasses of 125 of them (100 from Isfahan and 25 from Yazd) were studied for larva and adult forms of the parasite. Microfilariae were found in peripheral blood smears of 38 out of 294 (12.92%) tested camels, while 20 out of 125 camels (13.89%) harbored D. evansi adult worms in at least one region in their testicle, epididymis, spermatic cord, lung and heart. Two of infected males had adult forms of the parasite in all studied organs simultaneously. Pathological study of infected tissues revealed sections of parasite, severe acute and chronic inflammation, fibrosis and atrophy. D. evansi is endemic and constitutes an important health problem to camels in Iran's central desert, resulting in impaired working capacity and lowered productivity.

Multiple live subconjunctival dipetalonema: report of a case.

Parasitic infestations of the eye have been reported since centuries, affecting various parts of the eye. Some are subtle, coexisting with vision, while many others damage and destroy, in part or totally, the gift of sight. This report describes a patient with live subconjunctival dipetalonema infestation of the right eye, with 22 parasites removed live in one sitting from one eye.

Epidemiological aspects on vector-borne infections in stray and pet dogs from Romania and Hungary with focus on Babesia spp.

Canine arthropod-borne infections are of major interest in small animal practice and have been widely investigated in Central and Western Europe. However, only limited epidemiological data are available from South-Eastern European countries, although diseases including babesiosis or dirofilariosis are widely recognised as important canine infections in these countries. A steadily increasing number of dogs imported from South-Eastern Europe into Germany require particular attention by small animal practitioners. In this study, a total of 216 dogs [29 local Romanian pet dogs presented at Salvavet Veterinary Clinic in Bucharest, Romania, and 187 imported stray dogs from Romania (n = 109) and Hungary (n = 78) into Germany] were screened by molecular biological, serological and haematological methods for canine arthropod-borne infections. Eleven different parasitic and bacterial vector-borne pathogens-Babesia canis canis, Babesia canis vogeli, Babesia gibsoni, Babesia felis-like, Hepatozoon canis, Leishmania spp., Dirofilaria immitis, Dirofilaria repens, Acanthocheilonema reconditum, Anaplasma phagocytophilum and Mycoplasma haemocanis-were detected. Fifty-six percent of the dogs were positive by direct methods. B. canis canis was the most prevalent pathogen in dogs imported to Germany (42.8%) and dogs submitted for clinical consultation in Bucharest (44.8%). Our data strongly suggest the introduction of an adjusted screening panel in dogs from South-East Europe in view of increasing importation of dogs into Germany.

Hippobosca longipennis--a potential intermediate host of a species of Acanthocheilonema in dogs in northern India.

BACKGROUND: Hippobosca longipennis (the 'dog louse fly') is a blood sucking ectoparasite found on wild carnivores such as cheetahs and lions and domesticated and feral dogs in Africa, the Middle East and Asia, including China. Known as an intermediate host for Acanthocheilonema dracunculoides and a transport host for Cheyletiella yasguri, it has also been suggested that H. longipennis may be a vector for other pathogens, including Acanthocheilonema sp.? nov., which was recently reported to infect up to 48% of dogs in northern India where this species of fly is known to commonly infest dogs. To test this hypothesis, hippoboscid flies feeding on dogs in Ladakh in northern India were collected and subjected to microscopic dissection. RESULTS: A total of 12 infective larvae were found in 10 out of 65 flies dissected; 9 from the head, 2 from the thorax and 1 from the abdomen. The larvae averaged 2, 900 (± 60) µm in length and 34 (± 5) µm in width and possessed morphological features characteristic of the family Onchocercidae. Genetic analysis and comparison of the 18S, ITS-2, 12S and cox-1 genes confirmed the identity of the larvae as the Acanthocheilonema sp.? nov. reported in dogs in Ladakh. CONCLUSION: This study provides evidence for a potential intermediate host-parasite relationship between H. longipennis and the canine Acanthocheilonema sp.? nov. in northern India.

First report of Acanthocheilonema spirocauda in the Mediterranean monk seal (Monachus monachus).

The Mediterranean monk seal (Monachus monachus) is one of the world's most endangered marine mammals. The largest population is located mainly throughout the Aegean and Ionian islands and along the coastline of southern continental Greece. We report the findings of a necropsy and discuss their potential importance to the conservation of the species. The adult female monk seal appeared to be in a good nutritional state. The main necropsy findings were injuries consistent with a violent and sudden death, including three round wounds on the ventral surface of the body and several hematomas, as well as a decomposing male fetus in the uterus. Two nematodes were found in the right ventricle of the heart; no abnormalities were observed in the lungs, pulmonary vessels, or heart. The nematodes were identified as Acanthocheilonema spirocauda. This is the first report of infection of the Mediterranean monk seal with A. spirocauda, even though this is the most common heartworm found in most pinnipeds worldwide. This parasite should be considered in health care monitoring projects of this endangered species.

Characterization of the heartworm Acanthocheilonema spirocauda (Leidy, 1858) Anderson, 1992 (Nematoda: Onchocercidae) in Scandinavia.

The heartworm Acanthocheilonema spirocauda (Leidy, Proc Acad Nat Sci Philadelphia 10:110-112, 1858) Anderson, 1992 is described from material collected from harbour seals in Scandinavia and compared with types and other specimens described by Anderson (Can J Zool 37:481-493, 1959) from harbour seals in eastern USA. Most morphometric characters of the material from USA fall within the ranges established for the Scandinavian one. Some intraspecific variability in the organisation of papillae on the male tail was detected among the Scandinavian specimens. Differences between the specimens from Scandinavia and Eastern USA are also found in the organisation of papillae on the tail of males and females. An excretory pore was not discernible, but a clearly hemizonid-like structure is described. For the first time, scanning electron micrographs present external morphological structures of the species.

Prevalence and molecular phylogenetic characterization of Trypanosoma (Megatrypanum) minasense in the peripheral blood of small neotropical primates after a quarantine period.

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.

Heterodoxus spiniger (Enderlein, 1909) em cães domésticos (Canis familiaris, L. 1758) da cidade de Recife, Estado de Pernambuco, Brasil / Heterodoxus spiniger (Enderlein, 1909) on domestic dogs (Canis familiaris, L. 1758) from the city of Recife, Pernambuco State, Brazil

In the city of Recife, Pernambuco state, northeastern Brazil, domestic dogs are usually infected by ectoparasites of both veterinary and public health importance. Heterodoxus spiniger is a common dog chewing louse, which is known as an intermediate host of Dipetalonema reconditum. Despite of its wide geographical diffusion in Brazil, Heterodoxus spiniger was only recently recorded in Pernambuco. The aim of this paper was to verify the infestations by Heterodoxus spiniger on dogs from Recife. Three hundred and ten dogs were examined and 11 of these were parasitized by Heterodoxus spiniger, corresponding to a prevalence rate of 3.5%. This indicates that Heterodoxus spiniger can be found on dogs from Recife, but in low prevalence.

Na cidade de Recife, Estado de Pernambuco, nordeste do Brasil, os cães domésticos são freqüentemente infestados por ectoparasitos de interesse para medicina veterinária e para a saúde pública. Heterodoxus spiniger é um piolho mastigador comum em cães, apontado como hospedeiro intermediário de Dipetalonema reconditum. Apesar de sua ampla difusão geográfica no Brasil, apenas recentemente o piolho Heterodoxus spiniger foi registrado em Pernambuco. Neste sentido, o objetivo deste estudo foi verificar a presença de piolhos da referida espécie em cães de Recife. Dos 310 cães examinados, 11 estavam parasitados por Heterodoxus spiniger, correspondendo a uma taxa de prevalência de 3,5%. Este resultado indica que esta espécie de piolho mastigador pode ser encontrada em cães procedentes de Recife, porém com baixa prevalência.

A simple molecular method for discriminating common filarial nematodes of dogs (Canis familiaris).

Accurate diagnosis of canine filariosis is essential for choosing correct therapeutic approach. Therefore, reliable methods for discriminating among the different filarial infections in dogs are needed. The authors report simple and highly specific molecular methods that identify the three most common filarial nematodes of European dogs: Dirofilaria immitis, D. repens and Acanthocheilonema (syn. Dipetalonema) reconditum, based on (1) PCR amplifications of mitochondrial DNA (12S rDNA and coxI) with general filarial primers followed by digestion with restriction enzymes that generates band polymorphisms clearly discriminating the three species and (2) PCR amplifications with species-specific primers to support the restriction analysis, in particular in the case of multiple infections.

Concomitant immunity in a rodent model of filariasis: the infection of Meriones unguiculatus with Acanthocheilonema viteae.

In an attempt to study the occurrence of concomitant immunity in filarial infections, jirds (Meriones unguiculatus) were experimentally infected with Acanthocheilonema viteae, and patent animals were superinfected with a defined dose of A. viteae stage 3 larvae (L3). Infected animals harbored significantly less worms deriving from the superinfection than the control group (P < 0.05, 56.2%, and 63.4% protection), as shown by analysis of female worms 6 wk after superinfection on the basis of their developmental status and their length. This protection was not due to contact with L3 antigens because a significant reduction of worm burdens deriving of a superinfection was also observed after subcutaneous implantation of a single female worm (P < 0.05, 40.2% and 64.9% protection). The induced protective responses target L3 and restrict their migration because an established infection resulted in a reduction of L3 recovery (95.6% and 94.3%, P < 0.001) from tissues of jirds at day 5 after superinfection. Other data show that L3 from a superinfection are trapped within eosinophil-rich granulomas, which is likely to create unfavorable conditions for the worms and to lead to later death. Taken together, established A. viteae-infections partially protect hosts against homologous superinfection by an immune-mediated mechanism and, thus, regulate the population density of the parasites within the host by concomitant immunity.

First record of Acanthocheilonema dracunculoides from domestic dogs in Namibia.

Acanthocheilonema dracunculoides was diagnosed in 2 dogs from Windhoek, Namibia, by acid phosphatase staining of microfilariae. This is the 1st record of A. dracunculoides in Namibia.

Lethal LPS-independent side effects after microfilaricidal treatment in Acanthocheilonema viteae-infected rodents.

Mastomys coucha and jirds infected with Acanthocheilonema viteae, a filarial species free of endosymbiontic bacteria of the genus Wolbachia, suffer lethal side effects after effective microfilaricidal therapy with diethylcarbamazine and levamisole, whereas, M. coucha infected with the Wolbachia-infested species Brugia malayi or Litomosoides carinii tolerate corresponding treatment. Mortality in A. viteae infected, treated animals varied with microfilariae density in the blood. It was up to 100% in highly microfilaraemic M. coucha and jirds, but low or absent in animals with low microfilariae counts. Deaths occurred in most cases 5-24 h after treatment. Characteristic symptoms in animals, which died subsequently were a rapid drop in body temperature by 4-7 degrees C, an increase in hematokrit values by up to 10% and a moderate blood acidosis. Lethal effects in A. viteae infections did not depend on a particular status of hypersensitivity of the animals since desensitization procedures, which protected infected M. coucha against an otherwise lethal intravenous challenge with A. viteae homogenate did not protect against adverse reactions to a subsequent microfilaricidal treatment. The animals were protected from treatment induced death by injection of N-LMMA. Thus the final morbific agent seems NO. The data show that adverse effects after effective microfilaricidal therapy may be caused by microfilariae derived components different from Wolbachia-released LPS.