We investigated the distribution of Cryptosporidium in pigs in Japan by immunofluorescence staining of fecal samples and characterization of isolates by multilocus sequencing. The 344 animals sampled on eight farms included pre-weaned piglets (<1 month old; n = 55), weaned piglets (1-2 months old; n = 65), finished pigs (2-4 months old, n = 105) and of 4-6 months old (n = 67), sows (n = 36), and boars (n = 16). Average prevalence of Cryptosporidium on farms was 32.6%, ranging from 4.9 to 58.1%, decreasing with animal age (prevalences of <1 month old, 1-2 months old, 2-4 months old, 4-6 months old, sows, and boars were 27.3, 47.7, 41.9, 22.4, 11.1, 18.8%, respectively). Piglets (<1 and 1-2 months old) showing signs of diarrhea shed relatively more oocysts (5.28 in average log scale of oocysts per gram) in feces than piglets with normal or loose stools (those of 4.90). Thirty seven successful sequencing of the 18S ribosomal RNA gene among 62 examined samples revealed that all of the identified isolates were Cryptosporidium suis or Cryptosporidium scrofarum, which are generally specific to pigs, and that other species, such as zoonotic Cryptosporidium parvum, were absent. Interestingly, C. suis was frequently found in piglets younger than 2 months old, while C. scrofarum infection was more prevalent in older pigs which also showed increased prevalence of mixed C. suis and C. scrofarum infections. Sequencing of actin gene loci revealed the existence of variants of both Cryptosporidium species in pigs in Japan. Although the number of pigs examined in this study was relatively low, our results suggest that Cryptosporidium infection is widespread among pigs in Japan. In addition, the possibility of age-related specificity and pathogenicity in pig infections is also suggested.

Protozoan parasites of the genus Entamoeba infect many classes of vertebrates and are primarily classified based on morphological criteria. To date, only a few species have been proven to cause disease. Here, we examined the pathology of infected pigs with hemorrhage and detected Entamoeba parasites. Isolates were characterized genetically and ultrastructurally to identify the species. Histopathologically, bleeding and thrombus formation were seen only in the large intestine mucosa, where a large number of trophozoites or some Entamoeba cysts were observed around breakdowns in the lamina propria. No screw-shaped bacteria were detected in the lesions, and no pathogenic bacteria such as Brachyspira spp. were detected in fecal cultures. Interestingly, electron microscopy revealed that the parasites possessed mitochondrial organelles, unlike other Entamoeba spp. The isolates were identified as Entamoeba suis by PCR analysis and sequencing of the small subunit ribosomal RNA (SSU rRNA) gene. In phylogenetic analyses based on the actin gene, the E. suis isolate formed a cluster with Entamoeba histolytica and Entamoeba invadens, as well as with other parasites of the Amoebidae. Whether the pathogenicity of the E. suis isolate is affected by the severity of infection or host health status remains unclear; however, our results suggest that E. suis could cause or exacerbate clinical symptoms such as hemorrhagic colitis or diarrhea.

Chordomas of the tip of the tail in 6 ferrets were examined using histopathological, histochemical and immunohistochemical procedures. Histopathologically, round neoplastic cells containing numerous cytoplasmic vacuoles of varying sizes, categorized as “physaliphorous cells”, were observed in the amorphous eosinophilic or pale basophilic myxoid stroma. Physaliphorous cells were arranged in lobules and in a “chordoid” or “cobblestone” manner. The neoplasms were diagnosed as benign chordoma without local invasion and metastasis. Histochemically, the cytoplasm of small neoplastic cells was positive for periodic acid-Schiff stain and alcian blue (AB) pH 2.5 and pH 1.0 stains, but negative for hyaluronidase digestion-AB pH 2.5 stain. All neoplastic cells were strongly stained with colloidal ion, negative for high iron diamine AB pH 2.5 and toluidine blue pH 2.5 stains, and positive for Mayer’s mucicarmine stain. Immunohistochemistry using antibodies directed against low-molecular-weight cytokeratins (CK18, CK19 and CK20), vimentin and mucin core protein (MUC5AC) revealed that neoplastic cells had both epithelial and mesenchymal elements. The expression of low-molecular-weight cytokeratins suggests that neoplastic cells acquired the properties of glandular epithelial cells and produced epithelial mucus. Furthermore, the expression of cytokeratins, vimentin, S100 protein, brachyury and epithelial membrane antigen indicates that the neoplasms were equivalent to the classic type of human chordoma. Therefore, immunohistochemistry using these antibodies can be useful for the characterization of ferret chordoma.

studies on the induction of diarrhea by protozoa 13,26 . In this study, we investigated swine parasitism by Cryptosporidium, Giardia, Balantidium, and coccidium by calculating the number of oocysts and cysts per gram of stools in 334 pigs and analyzing the relationships between the detection rate and age group. The stool condition score was also analyzed to investigate the relationship between infection and diarrhea. Using the results of this, the first systematically classified study of swine protozoan infection, we clarified the status of Cryptosporidium and other protozoan infections and investigated their relationships with diseases. IntroductionParasitic protozoa that inhabit the gastrointestinal tracts of pigs include Cryptosporidium, Giardia, Balantidium, coccidium, Chilomastix, Entamoeba, Iodamoeba, and Tritrichomonas 16 . Of coccidium species, Eimeria scabra 6 , Isospora suis 23, and Balantidium coli 25 are known to cause diarrhea in pigs. Cryptosporidium species (C. suis, C. pig genotype II) has been reported as having weakly pathogenic results in pigs 4,20,21,27 . However, the pathogenicity of Cryptosporidium species remains unclear, and there have been only a few pathological AbstractWe surveyed the rates of internal infection of swine with gastrointestinal tract protozoa by checking the stools of 334 pigs (suckling pigs, growing pigs aged 1 to 6 months and sows or sow candidates) on 8 hog farms in Saitama, Japan from September to November 2009. Oocysts and cysts per gram of stools in all pigs were calculated. Parasite detection rates were statistically analyzed for each of the farms, age groups, and stool condition scores, and the relationship between parasitic infection and diarrhea was investigated. Cryptosporidium was detected in 79 pigs (23.7%) of all pigs checked, Giardia was found in 53 (15.9%), Balantidium in 155 (46.4%), and coccidium in 20 (6.0%). The rate of Cryptosporidium infection was higher in the 2-and 3-month-old groups (55.6 to 60.0%) than in the others (2.5 to 27.6%) (P < 0.05): oocysts per gram of stools of 10 4 to 10 5 were detected in a total of 33 suckling pigs and 2-and 3-month-old pigs, including 10 of an outdoor farm. There was a correlation between the infection rate and diarrhea in five piglets aged 1 month or younger. In addition, the number of 6-month-old pigs infected with Giardia was 40.0%, as opposed to just 5.1% (P < 0.05) of suckling pigs. Detection rates differed among farms. The Balantidium infection rate was high in pigs older than 4 months (45.0 to 78.9%), as opposed to 15.4 to 17.2% (P < 0.05) in pigs younger than 1 month.