The first report of Angiostrongylus vasorum (Nematoda; Metastrongyloidea) in Poland,in red foxes (Vulpes vulpes)

Angiostrongylus vasorum belongs to the superfamily of Metastrongyloidea. This nematode occurs in foxes, dogs and other predators. The Nematode A. vasorum place themselves in the pulmonary artery and its branches, and in the right ventricle and atrium of the heart. Numerous species of land snails are the intermediate hosts of the parasite. In 2013, lungs and hearts of 76 foxes shot in the Forest District G??boki Bród in Augustowska Primeval Forest were parasitologically necropsied. Four of the examined foxes were infected with the nematode A. vasorum, a prevalence of 5.2%. In one fox pericardium there were 6 male and 6 female nematodes. In the remaining three foxes nematodes were localized in the pulmonary artery. In two foxes 2 specimens of nematodes were detected (male and female, and two females) while 1 female was detected in the other fox. This is the first report of the presence of the nematode A. vasorum in fox in Poland.     

Disseminated angiostrongylosis with massive cardiac and cerebral involvement in a dog from Italy

A case of disseminated angiostrongylosis caused by Angiostrongylus vasorum in a dog living in Italy is here described. The dog was referred for severe respiratory distress and epileptic seizures; clinicopathological findings were consistent with severe pneumonia associated with right-sided heart failure and multifocal involvement of the brain. Bronchoalveolar fluid analysis identified a multitude of nematode larvae, identified as A. vasorum by conventional and biomolecular (PCR) methods. The major anatomo-histopathological lesions were chronic granulomatous pneumonia, a severe multifocal granulomatous myocarditis and multifocal mild vascular and inflammatory disease in the brain. A. vasorum should be included among the differentials of dogs with cardiovascular and neurologic disease.     

Pathology of naturally acquired Angiostrongylus vasorum infection in the red fox (Vulpes vulpes)

199 out of 509 foxes culled in a rabies control scheme, were shown to be infected with the metastrongylid parasite Angiostrongylus vasorum. Pathological lesions associated with angiostrongylosis in the fox are described and compared to reported lesions of A. vasorum infection in the dog. The lesions observed in the foxes seemed to be less severe and mainly confined to the ventral parts of the lung lobes which showed granulomatous pneumonia. Right ventricular hypertrophy of the heart was present in all infected foxes. It is believed that the fox population represents an important reservoir of A. vasorum.     

Infections with cardiopulmonary and intestinal helminths and sarcoptic mange in red foxes from two different localities in Denmark

Monitoring parasitic infections in the red fox is essential for obtaining baseline knowledge on the spread of diseases of veterinary and medical importance. In this study, screening for cardiopulmonary and intestinal helminths and sarcoptic mange (Sarcoptes scabiei) was done on 118 foxes originating from two distinct localities in Denmark, (Copenhagen) greater area and southern Jutland. Fifteen parasite species were recorded in 116 foxes (98.3%), nine parasitic species are of zoonotic potential. Parasite diversity was greater in foxes of Copenhagen in terms of overall parasite species richness and species richness of all helminth groups individually: trematodes; cestodes; and nematodes. Six parasite species were recovered from foxes of Copenhagen, but not from foxes of Southern Jutland: Echinochasmus perfoliatus; Echinostoma sp.; Pseudamphistomum truncatum; Dipylidium caninum; Angiostrongylus vasorum; and Sarcoptes scabiei, but Toxascaris leonina was only recorded in foxes of southern Jutland. A high prevalence and abundance of A. vasorum in foxes of Copenhagen was observed. The prevalence of four nematode species; Eucoleus (Capillaria) aerophilus, Uncinaria stenocephala, Toxocara canis, and Crenosoma vulpis, in foxes of both localities were comparable and ranging from 22.9% to 89%. The prevalence of Mesocestoides sp. was significantly higher in foxes of Copenhagen. Taenia spp. were detected using morphological and molecular analysis, which revealed the dominance of T. polyacantha in foxes of both localities. Infections with sarcoptic mange were evident only among foxes of Copenhagen (44.9%), which significantly affected the average weight of the infected animals. Further remarks on the zoonotic and veterinary implications of the parasites recovered are given.     

PCR detection of Angiostrongylus vasorum in faecal samples of dogs and foxes

The cardiovascular nematode Angiostrongylus vasorum is spreading in the fox and dog populations of northern Europe. A. vasorum can result in severe clinical manifestations in dogs; therefore, specific diagnosis is crucial for assessing its prevalence. In the present study, faecal samples from foxes and domestic dogs were tested by a new polymerase chain reaction (PCR) targeting the second internal transcribed region of the ribosomal DNA (ITS2) of A. vasorum. Initial isolation of faecal larvae by sieving facilitated the processing of larger sample volumes and allowed for the recovery of dead larvae from frozen samples. The sieve-PCR method enabled the identification of a single larva per 2 g of faecal sample and did not amplify DNA of a range of canine helminths, thus presenting a non-invasive tool for wildlife surveillance and for confirmative diagnosis in dogs.     

The common frog (Rana temporaria) as a potential paratenic and intermediate host forAngiostrongylus vasorum

Common frogs (Rana temporaria) were exposed either to third-stage larvae (L3) or to first-stage larvae (L1) ofAngiostrongylus vasorum. Following exposure to L3, viable larvae could be detected in the frogs for at least 2 weeks. Following exposure to L1, the frogs developed viable L3 in their tissues within 30 days. In a test of the infectivity of these larvae, one fox was fed frogs previously infected with L3 and another fox was fed frogs previously infected with L1. On autopsy it was found that adultA. vasorum populations had been established in both foxes. Thus, it could be concluded that frogs can act not only as paratenic hosts but also as intermediate hosts forA. vasorum.     

Muscular sarcocystosis in two arctic foxes (Vulpes lagopus) due to Sarcocystis arctica n. sp.: sarcocyst morphology,molecular characteristics and phylogeny

The arctic fox (Vulpes lagopus) is a critically endangered species in Norway, and therefore, the small population is closely monitored, and most foxes found dead are subjected to necropsy. In two deceased foxes, thin-walled muscular sarcocysts were first detected in histological sections, and numerous sarcocysts were later found in frozen and thawed muscle samples from Fox 1. These sarcocysts measured 1–12?×?0.1–0.25 mm and had closely spaced, short, knob-like protrusions, giving the cysts a serrated outline. Genomic DNA was extracted from eight isolated sarcocysts (Fox 1) and two muscle samples (Fox 2) and subjected to polymerase chain reaction amplification at four loci: the nuclear 18S and 28S ribosomal RNA genes and internal transcribed spacer 1 region and the mitochondrial cytochrome c oxidase subunit 1 gene (cox1). Both foxes were infected by the same Sarcocystis sp., which displayed little or no genetic variation at the three nuclear loci (99.9–100 % identity) and slightly more variation at cox1 (99.4–100 % identity). Sequence comparisons and phylogenetic analyses revealed that this species was distinct from other named Sarcocystis spp. but was closely related to various species using avian intermediate hosts and possibly identical to an unnamed species reported from two American dogs. The species described from the two arctic foxes was named Sarcocystis arctica n. sp.     

New insights onto cardiopulmonary nematodes of dogs in Sardinia,Italy

Dog heartworms Angiostrongylus vasorum and Dirofilaria immitis cause severe parasitological diseases; the importance of these parasitosis is growing due to their health impact on animals, the possible zoonotic implications and the recent spreading across several European countries and previously non-endemic areas. The aim of this study is to update the epidemiological scenario of cardiopulmonary nematodes A. vasorum and D. immitis in dogs of Sardinia island and to perform a morphological identification of larvae by the use of the Baermann and Knott techniques respectively and the molecular characterization of the mitochondrial cytochrome c oxidase subunit I (cox1) and the second ribosomal transcribed spacer region (ITS-2) of larvae L1 of A. vasorum. In the present study, 3.4 % (5/146) of dogs resulted positive at Baermann technique for A. vasorum while 8.9 % (61/684) to D. immitis. If on one side A. vasorum can be considered an emerging parasite in Sardinia, the parasitic pressure and the risk of infection for D. immitis in the island seems to be increased compared with the recent past.     

Spirocerca lupi infection in a dog from southern Italy: an “old fashioned” disease?

Canine spirocercosis caused by Spirocerca lupi is a life-threatening helminthic disease featured by severe clinical signs and potential development of oesophageal neoplasia. This infection is considered fairly frequent in Europe but almost unknown in Italy, from where only few reports have been published in local journals at the beginning of the XXI century. In the present study, an autochthonous case of canine spirocercosis in a 2-year-old dog from southern Italy is described. The animal was admitted to a private veterinary clinic in the municipality of Potenza (Basilicata region) due to persistent dyspnoea, vomiting, and regurgitation. At the abdominal ultrasound, a mass (1.3?×?2 cm) was observed and, thereafter, surgically removed from the apical part of the stomach. A female specimen of S. lupi was morphologically identified during the histological examination of the nodule and its eggs were detected in the faeces. In addition, the morphological identification was confirmed by molecular amplification and sequencing of partial cox2 gene sequence of S. lupi. Veterinarians should be aware of the potential risks derived from infection with this spirurid, which requires specific diagnostic and preventive measures.     

Molecular identification of novel intermediate host species of Angiostrongylus vasorum in Greater London

Angiostrongylus vasorum is a parasitic nematode that can cause serious and potentially fatal disease in dogs and other canids. The aim of this study was to determine the intermediate slug species infected in nature by sampling sites in Greater London and Hertfordshire located within a known hyperendemic region. Overall, A. vasorum larvae were recovered from 6/381 slugs (1.6 %) by tissue digestion, and their identity was confirmed by PCR. Infected slugs originated from three different sites in the Greater London area: one in Waltham Forest and two in Bromley. Slugs parasitised by A. vasorum were identified by a combination of external morphological characteristics and molecular techniques and belonged to three different families: the Arionidae, the Milacidae and the Limacidae. This includes two new host records for the parasite: Arion distinctus and Tandonia sowerbyi. This is the first record of A. vasorum in the family Milacidae, indicating that the parasite has a broader intermediate host range than previously recognised.     

Epidemiological studies on Echinococcus in Pampas fox (Lycalopex gymnocercus) and European hare (Lepus europaeus) in Buenos Aires province, Argentina

In Argentina, hydatid disease caused by Echinococcus granulosus is widespread. The south of Buenos Aires province, Argentina, is one of the three regions where hydatidosis is endemic. Although domestic dogs and sheep are considered to be the main hosts for E. granulosus, the potential role of wildlife in the local transmission of E. granulosus has not been investigated. The aim of this study was to estimate the hydatidosis/echinococcosis prevalence in European hare (Lepus europaeus) and Pampas fox (Lycalopex gymnocercus), two abundant species with a strong predator–prey relationship in rural areas of Buenos Aires province using different diagnostic tests. A total of 61 fox intestines were examined, finding that 52 (85.2 %) harbored at least one helminth species. However, no adult or immature form of Echinococcus sp. was found in the intestinal contents. Coproparasitological analysis and Copro–ELISA followed by Copro–PCR were used as supplementary diagnostic tests. Only one (1.7 %) of 59 fecal samples was positive to Taeniidae eggs by coproparasitological analysis, but this same sample was negative by the Copro–ELISA test. The analysis by Copro–ELISA showed 6 of 57 (10.6 %) positive samples, but the Copro–PCR tests carried out on these samples were negative to E. granulosus. A total of 6,808 lungs, 3,576 livers, and 3,542 hearts of hunted hares were examined and palpated, but no structure resembling hydatid cysts were detected. Our results suggest that hares and Pampas foxes are not currently important wild reservoirs of E. granulosus in the studied area.     

Histopathologic and ultrastructural studies of disseminated cytomegalovirus infection in strain 2 guinea pigs

Inbred strain 2 guinea pigs developed severe disseminated disease during acute experimental guinea pig cytomegalovirus (GPCMV) infection. A high mortality rate (100%) resulted, with most animals dying between 10 and 14 days after high dose (7.5 X 10(5) TCID50) virus inoculation. Infectious virus was recovered from many tissues, including spleen, lungs, liver, pancreas, heart, adrenals, kidneys, and salivary glands. The rate of GPCMV isolation from these tissues ranged from 50 to 100%. Gross lesions were observed in the spleen, liver, and lungs. On histologic examination, lesions were also seen in many other organs, including heart, pancreas, kidneys, adrenals, brain, intestines, and salivary glands. Intranuclear viral inclusions were present in many cell types of various organs. Under electron microscopic examination, cells with viral inclusions were easily found in the spleen, and liver, but less readily in the lungs, kidneys, salivary glands, and other organs. Most of the intranuclear inclusions consisted of electron-dense fibrils (10 nm diameter), viral nucleocapsids (100 nm), and tubular structures (60 nm diameter). Dense bodies and enveloped dense virions containing single or multiple capsids were present in the cytoplasm of many infected cells. The morphologic developments of GPCMV in these visceral tissues of strain 2 guinea pigs resembled those seen in GPCMV-infected cultured guinea pig cells but differed from those observed in the infected salivary gland duct cells. Strain 2 guinea pigs are a useful animal model for studying disseminated infection in CMV-associated human diseases.     

Some new aspects of the pathology,pathogenesis, and aetiology of disseminated lung lesions in slaughter pigs

From 40 pigs rejected for human consumption at slaughter due to an apparent presence of pyemic lung lesions (defined as disseminated processes containing pus and/or necrotic material), the lungs, spleen, liver, and kidneys were subjected to an extended macroscopic examination. Several lung lesions were sampled from each animal for histological and bacteriological examination. Samples from the kidneys and spleens were also subjected to bacteriological examination. At gross level, four groups of lung lesions were identified: 1) disseminated foci with contents of pus and/or necrotic material (n=26); 2) disseminated or multifocally located ecchymoses with a central area of fibroplasia (n=9); 3) non-pneumonic lesions, i.e., disseminated areas of atelectasis (n=1) or haemorrhagic areas developing due to the process of slaughter (n=1); and 4) suppurative lesions without a disseminated distribution pattern (n=3). Histologically, the disseminated suppurative/necrotic foci were identified as: A) abscesses (n=10); B) necrotic lesions (n=6); and C) ectatic or ectatic-like bronchioles with contents of pus and necrotic material (n=10). The macroscopic observation of disseminated centres of fibroplasia with peripheral ecchymoses (n=9) was confirmed histopathologically. The livers of five pigs contained multiple areas of chronic interstitial fibrosis related to migration of Ascaris suum larvae ("milk spotted liver"). Such hepatic lesions were significantly (p<0.01) related to the simultaneous occurrence of disseminated pulmonary ecchymoses with a central area of fibroplasia. Generally, all lung lesions of each individual animal contained identical monocultures of bacteria following this pattern: Staphylococcus aureus (abscesses); Actinomyces hyovaginalis (necroses); S. aureus, A. hyovaginalis, and Arcanobacterium pyogenes (ectatic and ectatic-like bronchioles). Areas with fibrosis were sterile or contained bacteria considered to be a result of contamination. Apart from one kidney, from which S. aureus was cultured, all other organs were sterile. It is concluded that difficulties exist in differentiating pulmonary pyemic lesions from non-pyemic lesions at the gross level. Thus, it was not possible to distinguish between abscesses/necroses and ectatic bronchioles, the pathogenesis of the latter being uncertain. However, the chronic non-pyemic lesions related to the migration of A. suum larvae should be identified by the absence of pus/necrosis. S. aureus was predominantly isolated from abscesses, whereas, and most surprisingly, A. hyovaginalis was the dominant bacterium isolated from the pulmonary necroses.     

Fatal case of disseminated infection with the turtle bacillus Mycobacterium chelonae.

An apparently immunocompetent 78 year old woman presented with confusion, subcutaneous abscesses, and lesions of the nasopharynx. Gram positive, acid fast bacilli were isolated from her blood after 10 days' incubation. She was treated with trimethoprim-sulphamethoxazole for presumed disseminated nocardiasis but deteriorated and died. A post mortem examination showed skin and pulmonary lesions and endomyocardial fibrous plaques. Organisms isolated from the skin and lung were indistinguishable from those cultured from the blood. The organism was subsequently identified as Mycobacterium chelonae. Primary pulmonary infection and disseminated disease are rarely caused by this organism and bacteraemia is seldom documented. The clinical presentation and bacteriological and histological findings are difficult to differentiate from those of disseminated nocardiasis. Isolation of the organism may fail without prolonged incubation of initial cultures and there is a danger of its being dismissed as medically unimportant. Diagnosis is further hampered because large pulmonary foci may be poorly revealed by conventional radiological examination of the chest.     

First autochthonous case of canine Angiostrongylus vasorum in Slovakia

Angiostrongylus vasorum is a metastrongyloid nematode that may cause cardiopulmonary disease, neurological signs and coagulopathies in dogs. The parasite has an indirect life cycle with molluscs as intermediate hosts, in which the infective third larval stage develops. Recently, A. vasorum has been repeatedly reported in dogs outside the endemic areas, indicating that this parasite is widely distributed over Europe. This is the first record of an autochthonous infection in a dog from Slovakia that was casually diagnosed during routine preventive parasitological examination. A. vasorum first-stage larvae were recovered using the Baermann technique and identified by length and characteristic tail morphology. The animal originated from Slovakia and had not travelled abroad. The dog had been regularly walked on grass fields with a concentrated presence of common species of Gastropoda and frogs. The owner reported that the dog had been licking and eating grass and it had shown curiosity for molluscs and frogs. The first finding of A. vasorum-infected dog in Slovakia has confirmed that the parasite is spreading beyond the traditional hyperendemic foci, which accentuates the need for monitoring and increasing of disease awareness in primary care clinical practice.     

Cat-scratch disease in Northern Italy: atypical clinical manifestations in humans and prevalence of Bartonella infection in cats

In this paper, we report an investigation on cat-scratch disease (CSD) in Northern Italy. Seventy-four cases of CSD were diagnosed at the San Matteo hospital, Pavia, during the period 2005–2010. Of these 74 patients, 18 (24.3 %) reported atypical clinical manifestations such as ocular papillitis, maculopapular eruptions, vertebral infection, pulmonary infiltrates, and granulomatous hepatitis. Contact with cats was documented for 61 patients (82.4 %), while cat-related trauma was reported for 49 patients (66.2 %). We subsequently investigated the presence of Bartonella infection in cats belonging to the above patients and in other domestic and stray cats from three provinces of Northern Italy. Among the 27 domestic cats tested, nine of the 11 belonging to the CSD patients and two of the remaining 16 were infected by B. henselae (81.8 % vs. 12.5 %). Out of over 1,300 stray cats examined, 23.1 % were seropositive for B. henselae; after culturing and genotyping, 17 % were found to be infected by B. henselae (15.5 %) or B. clarridgeiae (1.5 %).     

Differential susceptibility to Mycoplasma pulmonis intranasal infection in X-linked immunodeficient (xid), severe combined immunodeficient (scid), and immunocompetent mice

Mice with the scid mutation are highly susceptible to Mycoplasma pulmonis infection and develop a disseminated disease. In order to study the contribution of humoral immunity to the immune response, M. pulmonis was inoculated intranasally to X-linked immunodeficient (xid) mice. Severe combined immunodeficient (scid) and immunocompetent CBA mice were used as controls. The mice were killed and necropsied at day 30 or 37 post-infection. Samples from the nose, lungs and joints were taken for bacteriological and histological examination. Rhinitis was observed in all mouse strains. Chronic purulent bronchopneumonia was diagnosed in some of the CBA mice. Xid mice did not show severe lung lesions, despite the presence of numerous mycoplasma organisms in the lungs, in contrast to immunocompetent mice, which developed lung pathology. Scid mice showed less signs of pneumonia, but unlike in xid and CBA mice, there was spread of mycoplasmas from the respiratory tract and severe pathological changes in the joints. Our results indicate that B and/or T lymphocytes protect against dissemination of M. pulmonis from the airways. Innate immune reactions and/or bacterial virulence factors seem to contribute to the development of joint lesions, whereas IgG3 and IgM antibodies might be involved in lung pathology.     

Pathogenesis of a Chinese strain of bovine adenovirus type 3 infection in albino guinea pigs

Bovine adenovirus type 3 (BAV-3) is considered one of the most important respiratory tract agents of cattle and is widespread among cattle around the world. A BAV-3 strain was isolated from a bovine nasal swab for the first time in China in 2009 and named HLJ0955. Subsequently, BAV-3 has frequently been isolated from calves with respiratory diseases in China. To date, only limited study on the pathogenesis of BAV-3 infection in cotton rats has been conducted, and the pathogenesis of BAV-3 infection in guinea pigs has not been reported. Therefore, sixteen albino guinea pigs were inoculated intranasally with HLJ0955. All of the infected guinea pigs had apparently elevated rectal temperatures (39.2 °C–39.9 °C) at 2–7 days post-inoculation (PI). Consolidation and petechial hemorrhage were also observed in guinea pigs experimentally infected with HLJ0955. Viral replication was detectable by virus isolation and titration and by immunohistochemistry in the lungs of guinea pigs as early as 24 h PI. Viral DNA was detectable in the lungs of infected guinea pigs during 11 days of observation by real-time PCR. Virus-neutralizing antibodies against BAV-3 were detectable from 11 days PI and reached a peak titer at 15 days PI. Histopathological changes mainly occurred in the lungs of infected guinea pigs and were characterized by thickening of alveolar septa, mononuclear cell infiltration, hemorrhage and alveolar epithelial necrosis. These results indicate that HLJ0955 can replicate in the lungs of guinea pigs and cause fever and gross and histological lesions. The guinea pig infection model of BAV-3 would serve as a useful system for monitoring the infection process and pathogenesis of the Chinese BAV-3 strain HLJ0955, as well as immune responses to BAV-3 vaccines.     

Feline lungworm Oslerus rostratus (Strongylida: Filaridae) in Italy: first case report and histopathological findings

Oslerus rostratus syn. Anafilaroides rostratus (Strongylida: Filaroididae) is a metastrongyloid transmitted by snails, which localizes in peri-bronchial tissues and in the lung parenchyma of wild as well as domestic cats. In Europe, this nematode has been reported only on two occasions, being diagnosed in cats from Majorca Island and in northern Spain. Here, we describe a case of O. rostratus infection in a necropsied 4-year-old cat in Sicily (southern Italy). At the inspection of lungs, slender and greyish nematodes (four females and two males) were found embedded in the peri-bronchial tissues and in the bronchial walls. Parasites were morphological and molecularly identified as O. rostratus, with their 18S sequences being identical among them and showing a high homology (99 %) with those available in public databases. At the histology, nematodes were encapsulated in a pseudo-cystic formation surrounded by an interstitial inflammatory process and fibrous tissue. Lung lesions were mainly represented by peri-luminal fibrosis, hyperplasia and hypertrophy of the bronchial mucosa and glands, respectively. This first record of O. rostratus infection from Italy indicates that this parasite should be included in the differential diagnosis of feline of lungworm infection.