G. Di Guardo

Pathologic and Immunohistochemical Findings in Naturally Occurring West Nile Virus Infection in Horses

The pathologic and peroxidase immunohistochemical features of West Nile flavivirus (WNV) infection were compared in four horses from the northeastern United States and six horses from central Italy. In all 10 animals, there were mild to severe polioencephalomyelitis with small T lymphocyte and lesser macrophage perivascular infiltrate, multifocal glial nodules, neutrophils, and occasional neuronophagia. Perivascular hemorrhages, also noted macroscopically in two animals, were observed in 50% of the horses. In the four American horses, lesions extended from the basal nuclei through the brain stem and to the sacral spinal cord and were more severe than the lesions observed in the six Italian horses, which had moderate to severe lesions mainly in the thoracolumbar spinal cord and mild rhombencephalic lesions. WNV antigen was scant and was identified within the cytoplasm of a few neurons, fibers, glial cells, and macrophages. WNV infection in horses is characterized by lesions with little associated antigen when compared with WNV infection in birds and some fatal human infections and with other important viral encephalitides of horses, such as alphavirus infections and rabies.

PrP genotype in Sarda breed sheep and its relevance to scrapie

Summary. Several PrP gene polymorphisms modulate sheep scrapie susceptibility. Recently, an increase of scrapie outbreaks has been reported in Italy. A vaccine containing sheep brain homogenate was used in most of the outbreaks. We investigated PrP gene polymorphisms in scrapie-affected and clinically healthy Sarda breed sheep from a flock exposed to the aforementioned vaccine, and in affected Sarda sheep from unexposed flocks. All affected animals were (Gln/Gln)171 homozygous. Moreover, we observed no variation for Ala136 and a new polymorphism (Lys to Asn) at codon 176. Our findings confirm the correlation between scrapie and (Gln/Gln)171 in breeds with no variation for Ala136.

Animal transmissible spongiform encephalopathies and genetics

The genotype of the host plays a crucial role in the pathogenesis of transmissible spongiform encephalopathies (TSEs). In this respect, the most important factor is represented by the gene of the prion protein (PrP). The present work summarizes the currently available knowledge on the genetic basis of TSEs focusing, in particular, on sheep scrapie. Interest in this disease has grown markedly following the discovery of bovine spongiform encephalopathy, both for scientific and health reasons. In Italy, specific research grants from the Ministry of Health and the National Research Council (CNR), together with cooperation between the Istituto Superiore di Sanità and the Istituti Zooprofilattici Sperimentali, have allowed us to study the PrP genotype and to investigate the genetic susceptibility to scrapie in the most important Italian sheep breeds, with special reference to Sarda, Comisana and Massese. The PrP genotype in relation to scrapie susceptibility was also studied in goats of Ionica breed.

Cerebral Toxoplasmosis in Striped Dolphins (Stenella coeruleoalba) Stranded Along the Ligurian Sea Coast of Italy

This article reports the results of necropsy, parasitologic, microbiologic, histopathologic, immunohistochemical, indirect immunofluorescence, biomolecular, and serologic investigations on 8 striped dolphins (Stenella coeruleoalba) found stranded from August to December 2007 on the Ligurian Sea coast of Italy. Severe, nonsuppurative meningoencephalitis was found in 4 animals, as characterized by prominent perivascular mononuclear cell cuffing and macrophage accumulations in neuropil. These lesions were associated with mild lymphocytic–plasmacytic infiltration of choroid plexuses in 1 dolphin. Toxoplasma gondii cysts and zoites, confirmed by immunohistochemical labeling, were scattered throughout the brain parenchyma of 2 of the 4 dolphins. No viral inclusions were seen in the brain of any animal. Other findings included severe bronchointerstitial pneumonia and pulmonary atelectasis, consolidation, and emphysema. Parasites were identified in a variety of organs, including lung (Halocerchus lagenorhynchi). Microbiologic and serologic examinations for Brucella spp were negative on all 8 dolphins. The 4 animals with meningoencephalitis had serum antibodies against T gondii (titers ranging from 1:80 to 1:320) but not against morbillivirus. In contrast, the other 4 dolphins were seropositive for morbillivirus (with titers ranging from 1:10 to 1:40) but seronegative for T gondii. No morbillivirus antigen or nucleic acid was detected in the tissues of any dolphin. It is concluded that the severe lung and brain lesions were the cause of death and that T gondii was the likely etiologic agent of the cerebral lesions. Morbillivirus infection was not considered to have contributed to death of these animals.

PrP Sc deposition in nervous tissues without lymphoid tissue involvement is frequently found in ARQ/ARQ Sarda breed sheep preclinically affected with natural scrapie

Summary.The pathogenesis of natural scrapie in Sarda breed sheep was investigated in 1050 asymptomatic and 49 sick sheep from scrapie-affected flocks. Central and peripheral nervous system, along with lymphoreticular system (LRS) tissues, were subjected to immunohistochemistry (IHC) and Western-blotting (WB) for detection of pathological isoform of the prion protein (PrPSc). A total of 69 of the 1050 clinically healthy sheep were found to be infected with scrapie, with PrPSc being detected in both the central nervous system (CNS) and enteric nervous system (ENS) plexuses of 60 of the sheep, while IHC and WB yielded evidence of (PrPSc) deposition only in lymphoid tissues of the remaining 9 clinically healthy sheep. PrPSc was also detected in the CNS, as well as in ENS plexuses from all of the 49 clinically affected sheep. Nevertheless, 18 of the 69 clinically healthy animals (26%, 17 ARQ/ARQ and 1 ARQ/AHQ sheep), along with 3 ARQ/ARQ sheep (6%) of the clinically affected group, showed no IHC or WB evidence of PrPSc in lymphoid tissues, but PrPSc could be still detected in their CNS and ENS plexuses. The study demonstrates dual CNS and ENS PrPSc deposition in Sarda sheep with scrapie, in spite of an apparent lack of lymphoid tissue involvement in a number of cases.

Number and size of silver-stained nucleoli (Ag-NOR clusters) in canine seminomas: correlation with histological features and tumour behaviour.

Twenty spontaneous canine seminomas were examined. A statistically significant difference (P < 0.01) was detected in both the mean silver-stained nucleolar area/cell and the mean silver-stained nucleolar number/cell between (1) two intraductal seminomas with no signs of invasion, and (2) three intraductal ones with signs of invasion and 15 diffuse-type seminomas (two of which metastasized 3 and 12 months after surgical treatment). Furthermore, the metastasizing seminomas showed a mean silver-stained nucleolar area/cell which was significantly greater than that observed in all the other seminomas examined (P < 0.01). It is concluded that quantitative studies of nucleolar changes are useful in evaluating the biological behaviour of spontaneous canine seminomas.

Lipofuscin, lipofuscin-like pigments and autofluorescence.

A brief overview is here provided on lipofuscin and lipofuscin-like substances, with particular reference to their biological significance as well as to their cellular origin and pathophysiological role. Special emphasis is also placed on the mutual relationships between lipofuscin and lipofuscin-like lipopigments on one side, and cell autofluorescence on the other.

Fatal bronchopneumonia in a Metastrongylus elongatus and Porcine circovirus type 2 co-infected pig

Porcine circovirus type 2 (PCV2) infection is distributed worldwide and PCV2-associated disease (PCVAD) is considered among the most economically relevant ones to the global swine industry. PCV2 is known to play a causal role in the porcine respiratory disease complex, usually in close association with a large plethora of other biologic agents. We describe herein a case of fatal parasitic bronchopneumonia by Metastrongylus elongatus in a PCV2-infected pig. Metastrongylosis may still represent a major concern for outdoor herds. Our recent experience suggests that a concurrent PCVAD condition may trigger metastrongylosis, which may subsequently result, at its turn, in severe, sometimes fatal, pulmonary disease.

Ileal tract and Peyer's patch innervation in scrapie-free versus scrapie-affected ovines

Ileal Peyer’s patches (PPs) are involved early during sheep scrapie infection. This study qualitatively and semi-quantitatively evaluated ileal tract and PP innervation in 29 Sarda ovines of different age, PrP genotype and scrapie status. A prominent network of fibres was detected within PPs, mainly located in interfollicular lymphoid and stromal components. Intrafollicular fibres were rarely observed, with no apparent differences between scrapie-free and scrapie-affected animals, or among ovines carrying different PrP genotypes. In adult sheep, independent of their scrapie status, nerve fibres could be detected infrequently, close to the follicle-associated epithelium. Fibres were also detected within newly formed follicles and intrafollicular microgranulomas.

Toxoplasma gondii: Clues From Stranded Dolphins

An article recently published by Dr Nicole Andenmatten and coworkers in Nature Methods reveals the existence of an intriguing pathway of host-cell invasion used by Toxoplasma gondii, an apicomplexan protozoan parasite infecting a range of hosts worldwide, including several aquatic mammal species, in which it may cause abortion and lethal systemic disease. Free-ranging bottlenose dolphins (Tursiops truncatus) and striped dolphins (Stenella coeruleoalba) rank among T. gondii– susceptible hosts, with both cetacean species inhabiting the Mediterranean Sea and many other marine ecosystems worldwide. More in detail, T. gondii is believed to be a pathogen of concern for the 2 aforementioned species, with a documented potential to affect their already threatened health and conservation status, as clearly highlighted by the prominent subacute-to-chronic, nonsuppurative meningoencephalitis lesions reported in several striped dolphins found stranded between 2007 and 2008 along the Ligurian Sea coast of Italy. The brain tissue from these animals showed immunohistochemical and biomolecular evidence of T. gondii in close association with the above inflammatory lesions, with the brain parenchyma from 3 dolphins also scoring positive for the protozoan B1, uprt, and gra6 genes, which subsequently allowed classification of the concerned T. gondii isolates as a genotype II and as 2 ‘‘atypical’’ genotype II strains, respectively. Interestingly enough, although the bottlenose dolphin—a coastal species—is believed to acquire T. gondii infection through fecal oocyst contamination flowing from land to sea, this appears much less plausible for the striped dolphin, a typically pelagic species. Indeed, the biological, eco-pathological, and epidemiological features of T. gondii infection among striped dolphins (as well as in other susceptible cetacean species living offshore) could not entirely reflect those characterizing T. gondii infection among bottlenose dolphins and, more in general, in terrestrial mammals. Therefore, an ‘‘open sea T. gondii life cycle’’ involving striped dolphins and other pelagic cetaceans, and thereby existing independently from the land and the benthic protozoan cycle(s), should not be ruled out. Furthermore, T. gondii strains infecting striped dolphins and other susceptible cetacean hosts living in the open sea could display a pathogenetic behavior not completely overlapping that of the protozoan isolates infecting bottlenose dolphins as well as terrestrial mammals. In this respect, differences in genes coding for virulence factors, such as those crucially involved in host-cell colonization, may account for a pathogenetic behavior of T. gondii among striped dolphins distinct from that of the protozoan isolates infecting bottlenose dolphins and land mammals. On the basis of what has been described above, T. gondii strains obtained from stranded cetaceans, with special emphasis on those recovered from striped dolphins and other offshore species, should be considered a potentially valuable source of information for comparative studies addressing, in the future, the protozoan genes and factors involved in host-parasite interaction and host-cell invasion.