The lungworm, Halocercus brasiliensis (Nematoda: Pseudaliidae), from Guiana Dolphins Sotalia guianensis from Brazil with Pathological Findings.

The Guiana dolphin, Sotalia guianensis, is a small cetacean species found on the Atlantic coast of Central and South America from Honduras to Santa Catarina State, Brazil. The nematode Halocercus brasiliensis infects this cetacean, resulting in lung pathologies and death. The present study aimed to conduct a morphological and morphometric study of specimens of H. brasiliensis collected from S. guianensis from the coast of the state of Espírito Santo, Brazil. For this study, 7 dolphins were collected and examined for the presence of lung parasites. Nematodes were collected and processed for light microscopy, and lung fragments were processed for histological analysis. The nematodes were identified as H. brasiliensis according to their morphology and morphometry. The histopathological analysis revealed mineralization of the bronchiolar cartilage and inflammatory process. The parasitic infections by H. brasiliensis in S. guianensis can contribute to the debilitating status of these cetaceans, resulting in their stranding and accidental capture in fishing nets.

Temperature-dependent development and freezing survival of protostrongylid nematodes of Arctic ungulates: implications for transmission.

BACKGROUND: Umingmakstrongylus pallikuukensis and Varestrongylus eleguneniensis are two potentially pathogenic lungworms of caribou and muskoxen in the Canadian Arctic. These parasites are currently undergoing northward range expansion at differential rates. It is hypothesized that their invasion and spread to the Canadian Arctic Archipelago are in part driven by climate warming. However, very little is known regarding their physiological ecology, limiting our ability to parameterize ecological models to test these hypotheses and make meaningful predictions. In this study, the developmental parameters of V. eleguneniensis inside a gastropod intermediate host were determined and freezing survival of U. pallikuukensis and V. eleguneniensis were compared. METHODS: Slug intermediate hosts, Deroceras laeve, were collected from their natural habitat and experimentally infected with first-stage larvae (L1) of V. eleguneniensis. Development of L1 to third-stage larvae (L3) in D. laeve was studied at constant temperature treatments from 8.5 to 24 °C. To determine freezing survival, freshly collected L1 of both parasite species were held in water at subzero temperatures from -10 to -80 °C, and the number of L1 surviving were counted at 2, 7, 30, 90 and 180 days. RESULTS: The lower threshold temperature (T0) below which the larvae of V. eleguneniensis did not develop into L3 was 9.54 °C and the degree-days required for development (DD) was 171.25. Both U. pallikuukensis and V. eleguneniensis showed remarkable freeze tolerance: more than 80% of L1 survived across all temperatures and durations. Larval survival decreased with freezing duration but did not differ between the two species. CONCLUSION: Both U. pallikuukensis and V. eleguneniensis have high freezing survival that allows them to survive severe Arctic winters. The higher T0 and DD of V. eleguneniensis compared to U. pallikuukensis may contribute to the comparatively slower range expansion of the former. Our study advances knowledge of Arctic parasitology and provides ecological and physiological data that can be useful for parameterizing ecological models.

Feline lungworms unlock a novel mode of parasite transmission.

Snail-borne lungworms exert an enormous toll on the health and welfare of animals and humans. Of these parasites, Aelurostrongylus abstrusus and Troglostrongylus brevior affect the respiratory tract of felids. These lungworms share both the ecological niche and the species of snail (Helix aspersa) acting as intermediate host. Recently, the ability of H. aspersa to shed infective third-stage larvae (L3s) of A. abstrusus and T. brevior in the environment has been demonstrated, matching previous knowledge of mode of transmission of zoonotic lungworms. Here, we evaluated, for the first time, the ability of A. abstrusus and T. brevior L3s to infect new, susceptible snail hosts following their release from experimentally infected molluscs, and refer to this novel route of parasite transmission as intermediesis. The implications of snail-to-snail transmission in the epidemiology of snail-borne diseases are also discussed.

Lesions associated with Halocercus brasiliensis Lins de Almeida, 1933 in the lungs of dolphins stranded in the Northeast of Brazil.

The parasitic fauna of cetaceans is an important tool for ecological studies, including analyses on the causes of death. Halocercus brasiliensis is a nematode frequently found in the bronchi and bronchioles of some cetaceans, and it is commonly associated with focal inflammation of the respiratory tract leading to bacterial pneumonia and septicemia and, sometimes, to death. The objective of this study was to report infections by H. brasiliensis in the respiratory tract of Delphinidae stranded on the northern seaside of Bahia, Sergipe, and south of Alagoas, all states in the northeast region of Brazil. A total of 30 individuals, 1 Feresa attenuate (pygmy killer whale), 9 Stenella clymene (Clymene dolphin), and 20 Sotalia guianensis (Guiana dolphin) were studied. In 16 of them, the presence of H. brasiliensis was observed with a mean intensity of 3.5 ± 0.6 (range 1-9) in the hosts. Macroscopically, parasitic calcified nodules, lung congestion, edema, and emphysema were observed. Histopathological examination showed interstitial and granulomatous pneumonia with multifocal infiltrates, discrete to moderate edema, congestion, diffuse hemorrhage, and foci of calcification. We conclude that parasitic pneumonia in the sampled individuals may have directly contributed to stranding and death of the animals.

Elaphostrongylus cervi in a population of red deer (Cervus elaphus) and evidence of cerebrospinal nematodiasis in small ruminants in the province of Varese, Italy.

Thirty-one faecal samples were collected from red deer in the northern area of Varese, in the Italian region of Lombardy, between August and October 2008. The animals had either been hunted or accidently killed. Examination for internal parasites showed a prevalence of 45.2% for Elaphostrongylus cervi larvae and species identification was confirmed by polymerase chain reaction (PCR). Ninety-seven faecal samples were also collected from two goat flocks grazing in the same area between December 2007 and May 2008. These showed a prevalence of 74.7% for lungworms. Furthermore, the central nervous systems from five goats and one sheep from this area with a history of neurologically related lameness were examined. Histopathology confirmed E. cervi cerebro-spinal nematodiasis in five cases out of six. This study demonstrates E. cervi transmission from wild to domestic ruminants when the animals graze in the same area, and the possible occurrence of clinical disease in infected goats and sheep associated with high prevalence in deer.

Construction of a complementary DNA library for Parelaphostrongylus tenuis and identification of a potentially sero-diagnostic recombinant antigen.

Newly developed serological tests for diagnosing parelaphostrongylosis in cervids, using the excretory-secretory products (ES) of the infective larvae of Parelaphostrongylus tenuis in enzyme-linked immunosorbent assays (ELISAs), have demonstrable superiority over the traditional method of larval recovery and microscopic identification. To generate a source of ELISA antigen by genetic engineering, we created a complementary DNA (cDNA) expression library by the reverse transcription of mRNA of P. tenuis adult worms, and ligation with the vector lambda-ZAP II. The library was screened using antisera produced in mice by immunization with a somatic antigen preparation of adult worms. Seventeen clones were isolated, sequenced, and checked for similarity to other DNA sequences in GenBank. A previously identified parasite gene encoding an aspartyl protease inhibitor (API) was isolated from the cDNA library, subcloned and expressed using the pET expression vector to produce a glutathione S transferase (GST)-His-S.Tag-P. tenuis API fusion protein (molecular weight = 63 kDa). An enzyme-linked immunosorbent assay utilizing the API fusion protein as the coating antigen was used to serologically diagnose all white-tailed deer (WTD, 10 out of 10) that had been inoculated with 6 - 150 L3 P. tenuis, indicating that the antigen may be a useful serodiagnostic antigen for P. tenuis infection in this cervid species.

First record of a nematode Metastrongyloidea (Aelurostrongylus abstrusus larvae) in Achatina (Lissachatina) fulica (Mollusca, Achatinidae) in Brazil.

Achatina (Lissachatina) fulica was introduced in Brazil in the 1980s for commercial purposes ("escargot" farming) and nowadays, mainly by human activity, it is widespread in at least 23 out of 26 Brazilian states and Brasília, including the Amazonian region and natural reserves, where besides a general nuisance for people it is a pest and also a public health concern, since it is one of the natural intermediate host of Angiostrongylus cantonensis, ethiological agent of the meningoencephalitis in Asia. As Brazil is experiencing the explosive phase of the invasion, the Laboratório de Malacologia do Instituto Oswaldo Cruz/Fiocruz has been receiving samples of these molluscs for identification and search for Angiostrongylus cantonensis and Angiostrongylus costaricensis larvae. While examining samples of A. fulica different nematode larvae were obtained, including Aelurostrongylus, whose different species are parasites of felids, dogs, primates, and badger. Morphological and morphometric analyses presented herein indicated the species Aelurostrongylus abstrusus, as well as the occurrence of other nematode larvae (Strongyluris-like) found in the interior of the pallial cavity of A. fulica. This is the first report in Brazil of the development of A. abstrusus infective larvae in A. fulica evidencing the veterinary importance of this mollusc in the transmission of A. abstrusus to domestic cats. Since the spread of A. fulica is pointed out in the literature as one of the main causative spread of the meningoencephalitis caused by A. cantonensis the authors emphasize the need of sanitary vigilance of snails and rats from vulnerable areas for A. cantonensis introduction as the port side areas.

The first record of Elaphostrongylus alces larvae in moose in Poland and their development to the invasive stage.

BACKGROUND: This is the first record of the first stage larvae of Elaphostrongylus alces in moose in Poland. MATERIAL AND METHODS: The 75 faecal samples (5 g) from moose in Kampinos National Park were examined using Baermann method. RESULTS: The prevalence of E. alces larvae in feaces samples was 37%. The maximal intensity reached 294 larvae. 20 larvae of the first stage and 27 larvae of the third stage were measured. The comparison of their dimensions with the results obtained by Lankester, shows that they represent E. alces.

Acquired cervical scoliosis attributed to Parelaphostrongylus tenuis infection in an alpaca.

CASE DESCRIPTION: A 2-year-old alpaca was evaluated because of acute onset of cervical scoliosis. CLINICAL FINDINGS: Physical examination revealed severe scoliosis of the caudal portion of the cervical vertebral column with a C-shaped curvature to the right side. No gait deficits were observed. Cervical radiography confirmed severe curvature of C4 to C6 but did not reveal any bony changes. Cerebrospinal fluid had high total protein concentration and extremely high nucleated cell count with a high proportion of eosinophils, suggesting parasitic infection. TREATMENT AND OUTCOME: The alpaca was treated for suspected parelaphostrongylosis with ivermectin, fenbendazole, flunixin, vitamin E, thiamine, physical therapy, and a custom-made neck brace. The alpaca's condition continued to deteriorate, and it developed tetraparesis and ataxia and was euthanized after approximately 1 month. Microscopic evaluation of the cervical spinal cord revealed marked vacuolar changes in the left medial portion of the ventral funiculus, mild lymphoplasmacytic infiltration, and multifocal granulomas. The lesions were continuous from C1 to C7 and were compatible with parasite migration. CLINICAL RELEVANCE: To the authors' knowledge, this is the first report of acquired scoliosis in an alpaca, which appears to represent an unusual manifestation of parelaphostrongylosis that was reported in horses.

Cathepsin B homologue at the interface between a parasitic nematode and its intermediate host.

Parelaphostrongylus tenuis is a parasitic nematode that causes a debilitating neurologic disease in many North American cervids and domestic livestock species. We produced a PCR-based cDNA library from infective larvae (L3) in order to identify molecules that mediate parasitism. A dominant 1,250-bp amplicon encoded a homologue of cathepsin B cysteine proteases. The sequence incorporated a C29G substitution in the putative active site. Antibodies generated against a recombinant form detected the native protein (PtCPR-1) in Western blot assays of L3, but not adult worm, extracts. Immunohistochemical methods revealed that PtCPR-1 synthesis was restricted to larval stages within the snail intermediate host (Triodopsis sp.), beginning as early as 2 days postinfection (dpi) of snails. The protein was present in the intestine and luminal contents and was lost from larvae over time. Concurrent studies showed that larvae induced an immune response in snails beginning at 1 dpi. Layers of hemocytes encapsulated larvae immediately after infection, and granuloma-like structures formed around parasites in chronic infections. Loss of PtCPR-1 from L3 and its accumulation in host tissues coincided with degeneration of granuloma architecture 90 to 105 dpi. Fully developed L3 emerged from the snail at this time. Our data implicate PtCPR-1 in larval development and possibly in the emergence of P. tenuis from the intermediate host. Emerged L3 survived desiccation and cold stress, suggesting that they could remain infectious in the environment. Molecules promoting emergence would facilitate dispersal of L3 and increase the likelihood of transmission to definitive hosts.

Mortality of Rocky Mountain elk in Michigan due to meningeal worm.

Mortality from cerebrospinal parelaphostrongylosis caused by the meningeal worm (Parelaphostrongylus tenuis) has been hypothesized to limit elk (Cervus elaphus nelsoni) populations in areas where elk are conspecific with white-tailed deer (Odocoileus virginianus). Elk were reintroduced into Michigan (USA) in the early 1900s and subsequently greatly increased population size and distribution despite sympatric high-density (>or=12/km2) white-tailed deer populations. We monitored 100 radio-collared elk of all age and sex classes from 1981-94, during which time we documented 76 mortalities. Meningeal worm was a minor mortality factor for elk in Michigan and accounted for only 3% of mortalities, fewer than legal harvest (58%), illegal kills (22%), other diseases (7%), and malnutrition (4%). Across years, annual cause-specific mortality rates due to cerebrospinal parelaphostrongylosis were 0.033 (SE=0.006), 0.029 (SE=0.005), 0.000 (SE=0.000), and 0.000 (SE=0.000) for calves, 1-yr-old, 2-yr-old, and >or=3-yr-old, respectively. The overall population-level mortality rate due to cerebrospinal parelaphostrongylosis was 0.009 (SE=0.001). Thus, meningeal worm had little impact on elk in Michigan during our study despite greater than normal precipitation (favoring gastropods) and record (>or=14 km2) deer densities. Further, elk in Michigan have shown sustained population rates-of-increase of >or=18%/yr and among the highest levels of juvenile production and survival recorded for elk in North America, indicating that elk can persist in areas with meningeal worm at high levels of population productivity. It is likely that local ecologic characteristics among elk, white-tailed deer, and gastropods, and degree of exposure, age of elk, individual and population experience with meningeal worm, overall population vigor, and moisture determine the effects of meningeal worm on elk populations.

Immunoglobulin responses of northern elephant and Pacific harbor seals naturally infected with Otostrongylus circumlitus.

Immunoglobulin (Ig) binding patterns of Pacific harbor seals (PHS, Phoca vitulina richardsi) and northern elephant seals (NES, Mirounga angustirostris) to tissues of adult Otostrongylus circumlitus were examined by immunoblotting to investigate the role of age in the unusual response of juvenile NES to infection with O. circumlitus. Serum was taken from NES between March 1997 and March 2001 and from PHS between May 1996 and August 1999. The serum of seals infected with O. circumlitus contained antibodies that bound to all nematode tissues examined. Intensity of band staining on Western blots suggested that there were higher levels of antibody recognizing the excretory-secretory (ES) glands in the serum of NES that were 1 yr and older and in the majority of PHS compared with that in 2- to 9-mo-old NES. All juvenile NES infected with O. circumlitus and a proportion of the PHS and older NES infected with O. circumlitus contained Ig specific to a 28 kDa protein band that was dominant in the female reproductive tract of the nematode. The Ig binding patterns of NES and PHS to adult Parafilaroides sp., larval Pseudoterranova sp., and larval and adult Anisakis sp. differed sufficiently from that of O. circumlitus that immunoblotting for the 28 kDa protein could be useful for diagnosis of this parasite in juvenile NES. The banding patterns suggest that O. circumlitus nematodes die and disintegrate in PHS and NES and that NES of 1 yr and older and most PHS respond differently to the ES glands than 2- to 9-mo-old NES.

Muskox lungworm (Umingmakstrongylus pallikuukensis) does not establish in experimentally exposed thinhorn sheep (Ovis dalli).

Muskoxen (Ovibos moschlatus moschatus) on the northwestern mainland of Nunavut and Northwest Territories, Canada, are infected with the protostrongylid lungworm, Umingmaksrongylus pallikuuhkensis. The geographic range of this muskox population is expanding to the south and west, and it is anticipated that these animals will eventually become sympatric with Dall's sheep (Ovis dalli dalli) in the Mackenzie and Richardson Mountains. To address the concern of wildlife managers that U. pallikulkensis may infect and adversely affect Dalls sheep, four Dalls/Stone's (Ovis dalli stonei) hybrid lambs and one adult muskox (Ovibos moschatus wardi) were each given 100 third-stage larvae of U. pallikuukensis. All animals were intensively monitored for 9 mo postinfection (PI) using clinical examinations, fecal analyses, hematology, blood chiemistry and medical imaging. No first-stage larvae of U. pallikuinkensis were recovered from the lambs and monitoring revealed nio evidence that the parasite had established in any of these animals. First-stage larvae were found in the feces of the muskox beginning at 94 days PI, and typical parasite cysts were visible in lung radiographs at 188 days PI. This study addresses an important management and wildlife health issue associated with the potential for host-switching of pathogens and indicates that it is improbable that thinhorn sheep are suitable hosts for U. pallikuukensis.

Meningeal worm in a reintroduced elk population in Kentucky.

Meningeal worm (Parelaphostrongylus tenuis) has been implicated in the failure of several elk (Cervus elaphus) restoration attempts in the eastern United States. However, limited post-release monitoring and a paucity of published literature prevents a clear understanding of this parasite's role in past failures. During winters of 1997-2001, the Kentucky Department of Fish and Wildlife Resources translocated 1,044 elk from western states to eastern Kentucky (USA) in an effort to restore a free-ranging population. We monitored 521 radio-collared elk over 4 yr to determine the impact meningeal worm had on population establishment. Thirty (23%) of 129 non-capture related mortalities were attributed to meningeal worm. Twenty-two (73%) of these meningeal worm-caused mortalities were animals < 3 yr old. If younger elk born in Kentucky suffer higher mortality rates than older translocated elk, the population growth observed during the initial years of restoration may be temporary. Additional research is necessary to determine the influence meningeal worm will have on elk population growth in Kentucky.

Low-dose meningeal worm (Parelaphostrongylus tenuis) infections in moose (Alces alces).

Parelaphostrongylosis has a rapid onset and is lethal in neonatal moose (Alces alces) when large numbers of third-stage Parelaphostrongylus tenuis larvae (L3) are given experimentally. Little is known, however, about the severity and prognosis of infections acquired naturally by accidentally ingesting terrestrial gastropods which are rarely infected and have few larvae. To investigate the relationship between infecting dose, age of moose, and severity of disease, five calves were given low doses of three to 10 L3 when five (n = 2) or 9.5 mo old (n = 3). Each of two animals initially given low doses were later challenged with a dose of 15 L3. As positive controls, two calves were given doses of 15 and 30 L3, considered to be high. All five calves given low doses showed abnormal locomotory signs at 20-28 days postinoculation (DPI) that progressively became more pronounced with hind quarter weakness and front lameness. However, after 77-130 DPI, signs diminished markedly in two of these animals and disappeared in another two. Challenge infections of 15 L3 given 199 days after initial infections had no noticeable effects although an immature worm, probably resulting from the challenge, was found in the spinal cord of one animal killed 51 days later. Two positive control animals given the high doses of 15 and 30 L3 showed moderate to severe, non-resolving, locomotory signs and had to be euthanized. Results demonstrate that single, low doses of three to 10 P. tenuis L3 cause moderate disease in moose calves but over time, some worms die and animals can recover. A degree of protection may develop against future infection.

Aspects of the life cycle and pathogenesis of Elaphostrongylus cervi in red deer (Cervus elaphus).

Aspects of the migratory life cycle and pathogenesis of Elaphostrongylus cervi were studied in red deer (Cervus elaphus) using 2 farmed calves experimentally infected with 450 third-stage larvae killed 40 and 45 days postinfection and using 3 wild calves and 3 wild yearlings with natural infections killed during autumn hunting. A full necropsy was carried out on the experimental calves, but only the head, eviscerated carcass, and lungs were examined from the naturally infected animals. Histological examination included extensive studies of the central nervous system (CNS), spinal nerve roots, and lungs. The experimental calves had prepatent infections, with many immature adult nematodes in the CNS, whereas the wild calves showed CNS lesions indicating a very recent E. cervi infection. The yearlings had patent infections, with many mature E. cervi in their skeletal muscles, reflecting acquisition of infection during the previous summer. Our findings showed that E. cervi develop to the adult stage in the CNS (subarachnoid spaces) and subsequently migrate into the skeletal muscles, where the mature nematodes live in reproductive pairs and groups. In the nervous system, the nematode caused encephalomyelitis, focal encephalomalacia and gliosis, meningitis, radiculitis, ganglionitis, and perineuritis.

Experimental Elaphostrongylus cervi infection in sheep and goats.

The pathogenesis and migratory life cycle of Elaphostrongylus cervi were studied in four sheep and six goats killed and examined 6 days to 5 months after inoculation with infective third-stage larvae (L3). Detailed histological studies demonstrated that the L3 followed a porto-hepatic, and probably also a secondary lymphatic, migratory route from the abomasum and small intestine to the lungs, with subsequent spread via the general circulation to the central nervous system (CNS) and other tissues. In addition, the results suggested that haematogenously spread L3, arrested in arterial vessels outside the spinal cord, migrated into the cord along the spinal nerves. During migration, the L3 caused focal inflammation and necrosis in the organs and along the spinal nerve roots, and infarcts occurred in the myocardium, kidneys and CNS. Nematode development took place in the CNS. During development, there was a gradual die-off of nematodes and patent infections were not observed. However, in one animal many mature nematodes were demonstrated in the CNS. In the nervous system, the nematodes caused encephalomyelitis, focal traumatic encephalomalacia, gliosis, meningitis, choroiditis, radiculitis and perineuritis. Two goats and one sheep displayed long-lasting paraparesis starting 6 weeks after inoculation. The signs apparently resulted from nematode-induced spinal nerve root lesions. From 19 weeks after inoculation the sheep also showed signs of severe brain disturbances due to traumatic and inflammatory lesions caused by adult E. cervi in the cerebral parenchyma. We conclude that E. cervi represents a potential cause of neurological disease in small ruminants grazing areas inhabited by red deer. This is the first report confirming the infectivity of E. cervi for domestic ruminants.

Susceptibility of mule deer (Odocoileus hemionus) and two species of North American molluscs to Elaphostrongylus cervi (Nematoda: Metastrongyloidea).

An experimental study was done to determine whether Elaphostrongylus cervi can be transmitted to common intermediate and definitive hosts indigenous to North America. First-stage larvae of E. cervi obtained from red deer (Cervus elaphus elaphus) in New Zealand developed to the infective third stage in snails (Triodopis multilineata) and slugs (Deroceros reticulatum). Two mule deer (Odocoileus hemionus) were inoculated orally with 102 or 406 third-stage larvae. One animal developed a patent infection on day 121 postinoculation (PI), and the other had numerous first-stage larvae in its lungs and colonic feces on 128 days PI when it was killed. A control red deer inoculated with 100 larvae began passing larvae on 119 days PI. Larval excretion was low and intermittent in the mule deer and during the first 4 wk of patency in the red deer. Both mule deer developed progressive neurological disease beginning on day 104 PI and had to be killed 4 or 7 wk later, respectively. The red deer remained clinically normal.