Ostreid herpesvirus type 1 genomic diversity in wild populations of Pacific oyster Crassostrea gigas from Italian coasts.

Ostreid herpesvirus 1 (OsHV-1) is a significant pathogen affecting the young Pacific oyster Crassostrea gigas, worldwide. A new variant, OsHV-1 µVar, has been associated with recurrent mortality events in Europe since 2008. Epidemiological data collection is key for global risk assessment; however little is known about health status and genotypes present in European wild oyster beds. Most studies to date have involved only cultivated individuals during mortality events, and reported low genotype diversity. With this study, conducted along the Italian coasts, we investigated for the first time the presence of OsHV-1 in European natural oyster beds. Analysis of three genomic regions revealed the presence of at least nine different genotypes, including two variants close to the OsHV-1 reference, known since the early 1990s but with no European record reported since 2010, and highlights relevant genotype diversity in natural environment. Phylogenetic analysis distinguished two distinct clusters and geographical distribution of genotypes, with the exception of a variant very closely related to the µVar, which appeared the single genotype present in all the Adriatic sites. Interestingly, these wild symptom free populations could represent, in Europe, an accessible alternative to the import of OsHV-1-resistant oyster strains from the East Pacific, the native area of C. gigas, avoiding the high-risk of non-native marine species and new pathogen introductions.

Cryptosporidium parvum: From foal to veterinary students.

This paper describes the transmission of a zoonotic subtype of Cryptosporidium parvum between two foals hospitalized in an Equine Perinatology Unit (EPU) linked to an outbreak of cryptosporidiosis in veterinary students. Fecal specimens of 36 mares (105 samples) and 28 foals (122 samples) were subjected to Ziehl-Neelsen staining, nested PCR of 18S rDNA. Two foals tested positive for Cryptosporidium; PCR restriction fragment length polymorphism (PCR-RFLP) analysis and subtyping by nested PCR of the 60kDa glycoprotein (gp60) gene revealed C. parvum subtype IIdA23G1. The introduction of Cryptosporidium into the EPU is suspected to be in a foal showing no initial clinical signs that tested positive for C. parvum during an asymptomatic phase. A second foal, hospitalized afterwards for perinatal asphyxia syndrome complicated with failure of passive transfer and sepsis, showed severe watery diarrhea after 4 days of hospitalization and was positive for the same subtype. During this period, six students attending the EPU complained of abdominal pain and diarrhea and were positive for the same subtype of C. parvum. To the authors' knowledge, this is the first description of this subtype in foals and the first report of evidence of zoonotic transmission of cryptosporidiosis from foals to human.

Epidemiological survey on Cryptosporidium in an Equine Perinatology Unit.

The present study aims to evaluate the prevalence, pattern of spread and risk factors for the transmission of cryptosporidiosis in foals and mares hospitalized in a University Equine Perinatology Unit, where a new subtype family of Cryptosporidium horse genotype was described by Caffara et al. (2013). Mares (36) and foals (37) hospitalized during the 2012 foaling season were included. Multiple sampling from each animal was performed (a total of 305 stool samples were collected). One hundred and eleven environmental samples (gauze swabs) were also collected before and after the breeding season. Fourteen foals were found positive for Cryptosporidium spp. by PCR in at least one sample; a total of 35 foal stool specimens were confirmed for the presence of the protozoa. Instead none of the stool specimens from mares were found positive. PCR-RFLP analysis shows Cryptosporidium parvum in 5 stool samples and Cryptosporidium horse genotype in 21. In 9 specimens, from 4 different foals, the profile was suggestive for a mixed infection. The subtyping at gp60 locus showed 2 strains as members of the subtype family IId and six of the subfamily IIa of C. parvum. Twenty isolates were identified as Cryptosporidium horse genotype subtype VIaA15G4. Five gauze swabs collected from the walls of the boxes where the animals were hosted out of 111 environmental samples examined were PCR positive for Cryptosporidium spp. Cryptosporidium parvum was detected in one sample collected before the foaling season, while Cryptosporidium horse genotype profile was observed in 4 wall samples collected at the end of the 2012 foaling season. The prevalence observed in foals (37.8%) was higher than that reported in other studies. These features and the diffusion of the same genotype point out as the EPU, where critically ill foals are hospitalized, can support the spread of cryptosporidiosis. Therefore, the manual tasks and the activities carried out in these facilities are of great importance, as they might favor the diffusion of the infection.

Experimental and molecular study of cercariae of Clinostomum sp. (Trematoda: Clinostomidae) from Biomphalaria spp. (Mollusca: Planorbidae) in Brazil.

Despite the large number of reports of species of Clinostomum from vertebrate hosts in South America, studies evaluating the molluscan transmitters of these parasites are scarce. In the present study, clinostomatoid cercariae shed from 0.02% (4/17,485) specimens of Biomphalaria spp., collected at the Pampulha reservoir, Belo Horizonte, Minas Gerais, Brazil, were used for experimental infection of Poecilia reticulata . Samples of cercariae from molluscs and metacercariae experimentally obtained from fish were subjected to morphological and molecular analyses and compared with species of Clinostomum reported in the Americas. The cercariae and metacercariae, here identified as Clinostomum sp., present general morphology similar to that reported for Clinostomum marginatum , however, from molecular point of view, differ significantly from North American C. marginatum and other species of Clinostomum reported in South America. These results suggest that the diversity of Clinostomum found in Brazil may be underestimated. Additional studies aimed at molecular characterization of South American species of Clinostomum, including the finding of specimens with sequences similar to that reported for C. marginatum in North America are required.

Granulomatous lesions in a wild mullet population from the eastern Ligurian Sea (Italy): mycobacteriosis vs. pseudotuberculosis.

Mycobacterium spp. and Photobacterium damselae subsp. piscicida are recognized as the most frequent causative agents of granulomatous lesions in fish. Although frequent episodes of mycobacterial infections have been reported in wild fish worldwide, only sporadic cases have been documented to date in Italy. To investigate for the presence of lesions referable to mycobacteriosis and to identify the mycobacterial species involved, a total of 159 wild mullets were fished from the eastern coast of the Ligurian Sea, killed and necropsied. Liver and spleen samples were collected from all fish for histopathological and microbiological analyses. Molecular investigations for identification of Photobacterium damselae subsp. piscicida were performed. Gross examination revealed granulomatous lesions in one animal; microscopically, 42.14% of fish displayed granulomas with various histological features, 19.50% resulted positive at Ziehl-Neelsen staining, and were confirmed as mycobacterial lesions by culture. The identified colonies were characterized as M. fortuitum, M. abscessus, M. flavescens, M. chelonae, M. septicum and M. nonchromogenicum. In all, 35% of animals resulted positive for Photobacterium damselae subsp. piscicida. These data suggest widespread mycobacterial infection also by Photobacterium damselae subsp. piscicida infections in wild fish. Moreover, the pathogenicity of some mycobacterial species, previously considered as saprophytic, was demonstrated.

Metacercariae of Clinostomum complanatum (Trematoda: Digenea) in European newts Triturus carnifex and Lissotriton vulgaris (Caudata: Salamandridae).

Adults of Clinostomum spp. are digenetic trematodes found in fish-eating birds, reptiles and occasionally mammals, including humans. Freshwater snails serve as first intermediate hosts and many fish species and amphibians as second intermediate hosts. To date, amphibian hosts of Clinostomum metacercariae include members of urodele and anuran families in North America, but no data are available on infections of European amphibians, including newts. In this study, we characterize infections of Clinostomum complanatum metacercariae in four smooth (Lissotriton vulgaris) and 18 Italian crested newts (Triturus carnifex) from an artificial pond located in a protected area in Tuscany, Italy. Parasites were surgically removed from the infected newts and identified both morphologically and using sequences of a mitochondrial gene, cytochrome c oxidase I, and the ribosomal markers, internal transcribed spacers. This is the first record of C. complanatum in European newts and, more generally, in amphibians in Europe.

Infection dynamics of Cichlidogyrus tilapiae and C. sclerosus (Monogenea, Ancyrocephalinae) in Nile tilapia (Oreochromis niloticus L.) from Uganda.

The infection dynamics of the gill monogeneans Cichlidogyrus tilapiae and C. sclerosus on Oreochromis niloticus with respect to habitat type (reservoir, stream, ponds and cages), host sex, size and seasons was determined between January and November 2008. During the study period, 45.2% of the 650 fish examined were infected with Cichlidogyrus spp. The infected hosts harboured an average of 8.6 ± 3.4 parasites/fish. Across habitat types, the proportion of infected fish was not statistically different. In contrast, the number of parasites recorded on infected fish from different habitat types differed significantly. The highest parasite number was recorded in reservoir-dwelling fish and lowest in stream-dwelling hosts. Concerning sex, more female O. niloticus were infected and harboured a high number of parasites than male and sexually undifferentiated fish. A weak negative relationship was found between rainfall and monthly parasite infections. However, a higher number of parasites and proportion of infected hosts were found during dry than in wet seasons, except in ponds. Results of this study show that differential exposure due to changes in fish behaviour associated with habitat modification and sex may account for the infection difference across the sampled sites. Meanwhile, rainfall and the associated hydrological events are important factors regulating monogenean infections in tropical aquatic environments. The continuous presence of Cichlidogyrus spp. in fish provides evidence of possible parasite outbreaks, indicating the application of biosecurity measures as crucial for the success of intensive fish farming.

Viral encephalopathy and retinopathy outbreak in freshwater fish farmed in Italy.

Viral encephalopathy and retinopathy (VER), otherwise known as viral nervous necrosis (VNN), is a neuropathological condition affecting > 40 species of fish. Although VER affects mainly marine fish, the disease has also been detected in certain species reared in freshwater environments. There are relatively few reports concerning the disease in freshwater species, and there is not much information on clinical signs. Nevertheless, the most common clinical findings reported from affected freshwater species are consistent with the typical signs observed in marine species. In this paper we describe the main clinical signs and the laboratory results associated with the detection of a betanodavirus in hybrid striped bass x white bass (Morone saxatilis x Morone chrysops) and largemouth bass Micropterus salmoides, reared in a freshwater environment. We also detected the virus by real-time PCR and isolated it in cell culture from a batch of pike-perch Sander lucioperca farmed in the same system.

Sand flea (Tunga spp.) infections in humans and domestic animals: state of the art.

Tungiasis is a parasitic disease of humans and animals caused by fleas (Siphonaptera) belonging to the genus Tunga. Two species, Tunga penetrans (L.) and Tunga trimamillata, out of 10 described to date, are known to affect man or domestic animals; the other eight are exclusive to a few species of wild mammals. Tunga penetrans and T. trimamillata originated from Latin America, although the first species is also found in sub-Saharan Africa (between 20 degrees N and 25 degrees S). Hundreds of millions of people are at risk of infection in more than 70 nations, mostly in developing countries. The second species has been reported only in Ecuador and Peru. Males and non-fertilized females of Tunga are haematophagous ectoparasites; pregnant females penetrate the skin where, following dilatation of the abdomen, they increase enormously in size (neosomy) and cause inflammatory and ulcerative processes of varying severity. The importance of Tunga infection in humans concerns its frequent localization in the foot, which sometimes causes very serious difficulty in walking, thereby reducing the subject's ability to work and necessitating medical and surgical intervention. Tungiasis in domestic animals can be responsible for economic losses resulting from flea-induced lesions and secondary infections. Because tungiasis represents a serious problem for tropical public health and because of the recent description of a new species (Tunga trimamillata), it seems appropriate to review current knowledge of the morphology, molecular taxonomy, epidemiology, pathology, treatment and control of sand fleas of the genus Tunga.

Occurrence of Mycobacterium spp. in ornamental fish in Italy.

The occurrence of Mycobacterium spp. in freshwater and marine ornamental fish was studied in Italy from June 2002 to May 2005. Two surveys were carried out, one of aquarium fish sent to the Laboratory for diagnosis, and the other of prevalence of infection by mycobacteria in ornamental fish imported into Italy. Bacterial isolation was carried out from the spleen, kidney and liver, and the isolates were subsequently identified by biochemical tests. In the first survey, 387 fish were examined and Mycobacterium spp. were isolated from 181 (46.8%) fish. In the second survey 127 batches of ornamental fish from different countries were examined. Mycobacterium spp. were isolated from 38 (29.9%) batches. The following species were found: M. fortuitum, M. peregrinum, M. chelonae, M. abscessus, M. marinum, M. gordonae, M. nonchromogenicum and M. interjectum. There was a high prevalence of infection independent of the presence of macroscopic lesions. Mycobacterium fortuitum and M. chelonae were more prevalent than M. marinum in the samples examined.

Parasitic diseases of marine fish: epidemiological and sanitary considerations.

Over recent decades, parasitic diseases have been increasingly considered a sanitary and economic threat to Mediterranean aquaculture. In order to monitor the distribution of parasites in cultured marine fish from Italy and study their pathogenic effects on the host, a three-year survey based on parasitological and histopathological exams was carried out on 2141 subjects from eleven fish species and coming from different farming systems (extensive, intensive inland farms, inshore floating cages, offshore floating cages and submersible cages). A number of parasitic species was detected, mostly in European sea bass (Dicentrarchus labrax), gilthead sea bream (Sparus aurata), mullets (Chelon labrosus, Mugil cephalus, Liza ramada) and sharpsnout sea bream (Diplodus puntazzo), with distribution patterns and prevalence values varying in relation to the farming system, in-season period and size category. The epidemiology and pathological effects of the parasites found during the survey are discussed.

Spinal deformity in a sandtiger shark, Carcharias taurus Rafinesque: a clinical-pathological study.

The present study investigated spinal deformity in a captive sandtiger shark and describes the clinical and histopathological features of the lesion. Clinically, the shark presented a marked spinal curvature in the region between the pectoral girdle and cranial dorsal fin. Radiographic studies revealed scoliosis and kyphosis of the vertebral column with a modification of the basic structure of five vertebrae. Post-mortem examination confirmed the results of radiographic studies and histological examination showed remodelling and cartilaginous proliferation of the vertebral bodies together with necrosis, haemorrhage and signs of a local chronic inflammatory response extending to the surrounding muscle. Different aetiologies are proposed and discussed but nutritional imbalance (dietary vitamin deficiency) and/or traumatic injury are considered the most likely contributing factors.

Anatomy of Tunga trimamillata Pampiglione et al., 2002 (Insecta, Siphonaptera, Tungidae) and developmental phases of the gravid female.

This paper deals with some internal anatomical features observed in histological sections and freshly dissected mounts of Tunga trimamillata, a Siphonaptera recently discovered in Andean regions of Ecuador from several mammals, including man. It was possible to study in males and also non-gravid and gravid females, the location and anatomy of several organs not previously described for this species: the testes, epididymis, ganglia, Malpighian tubules, eyes, rectal ampulla with one of its pads and structures which could be interpreted as midgut diverticula, whose presence has not been recorded in the Siphonaptera. The process of neosomy in the female during pregnancy is illustrated by photographs of the consecutive developmental phases, taken at the stereomicroscope. Furthermore, some details of the exoskeleton, spermatheca during different phases of pregnancy of the gravid female and the presence of a foreign body (parasite?) within the haemocoel have been displayed in specimens cleared with Hoyer's medium.

Differential diagnosis between Tunga penetrans (L., 1758) and T. trimamillata Pampiglione et al., 2002 (Insecta, Siphonaptera), the two species of the genus Tunga parasitic in man.

Of the ten currently known species of sand fleas, only two, Tunga penetrans and Tunga trimamillata, are known to be parasites of man, besides other warm blooded animals, most of which are peridomestic. The hosts of the other eight are limited to a few genera of wild mammals. T. trimamillata was only recently identified and differentiated from T. penetrans by features of the gravid female phase. In the present paper the different morphological characters of both for non-gravid females and males of the two species are described. In the non-gravid-females the distinguishing characters are: a) differences in the length of the first two segments of the maxillary palps (the first is the longest in T. trimamillata, whereas the second is the longest in T. penetrans), this feature is associated with another character i.e. the presence of short, thick spines in addition to the bristles on the surface of the segments only in T. trimamillata; b) the presence, only in T. trimamillata, of a row of spines on the antero-medial surface of the libia of the 3rd pair of legs; c) the last abdominal spiracle protrudes in T. trimamillata but not in T. penetrans; d) the hood of the hilla in spermatheca is surrounded by a papilla only in T. penetrans. The following morphological characters differentiate males of T. trimamillata and T. penetrans: a) T. trimamillata has a row of spines on the antero-medial surface of the tibia of the 3rd pair of legs; b) the diameter of the abdominal spiracles of T. trimamillata is smaller than that of T. penetrans and the edges of the spiracles are more regular; c) the claspers and aedeagus of T. trimamillata are shorter than those of T. penetrans; d) the profile of the oedeagal apodeme differs between the two species.

Additional description of a new species of Tunga (Siphonaptera) from Ecuador.

A new species of chigoe flea belonging to the genus Tunga Jarocki, 1838 (Siphonaptera, Pulicidae, Tunginae), Tunga trimamillata, was recently described by Pampiglione et al. (2002). A better description and more details of the epidemiology of the flea are now presented. It is a species found in goats, pigs and cattle in Santa Isabel in Andean Ecuador. This new species differs from its most similar congener, Tunga penetrans (L., 1758), by several features, the most important of which are a) the presence on the anterior extremity of the gravid female of three rounded humps surrounding the head and thorax (which, however, are not visible if viewed in profile), b) slightly larger dimensions and c) the length of the first segment of the maxillary palpi which is longer than each of the other three.

[On a new species of sand flea from Ecuador and tungiasis, a problem of Public Health in many developing countries]. / Su di una nuova specie di pulce penetrante dell'Ecuador esulla tungiasi, problema di Sanità Pubblica in molti paesi in via di sviluppo.

The authors recently described a new species of sand flea (genus Tunga) occurred in goat, sheep, cow, pig and man, in the Andean village of S.ta Isabel (Ecuador) and named Tunga trimamillata. Its most important morphological characteristics are: slightly larger dimensions than T. penetrans, both in male and female; presence on the anterior extremity of gravid female of three rounded humps surrounding head and thorax; length of the first segment of maxillary palpi which is longer than each of the other three. The importance of this parasite is due to the fact that it can cause the same damages as T. penetrans both in humans and domestic animals, with conseguent walking difficulties and local or generalized infections. For breeders, economic losses can be remarkable. Tungiasis is a serious problem of Public Health in many countries of Latin America and Subsaharian Africa, and it can be considered in these localities as an indicator of underdevelopment and poor hygiene conditions.