Prevalence and pathogenicity of Cryptosporidium suis in pre- and post-weaned pigs

A total of 4338 faecal samples, 135 of sows, 3368 of pre-weaned and 835 of post-weaned piglets from eight farms in South Bohemia, Czech Republic were collected and examined for Cryptosporidium infection. No sow, but 5.7% pre-weaned and 24.1% post-weaned piglets were positive for Cryptosporidium infection. No relationship was found between diarrhoea and Cryptosporidium infection in any of the different age groups (pre- and post-weaned piglets). Four piglets, which were sporadically shedding cryptosporidia in faeces, were necropsied. Neither clinical signs of diarrhoea nor macroscopical changes were found. Histologically, a moderate infection of cryptosporidia was detected in the glandular epithelium along the large intestine, with predisposition to the ansa centralis of the colon. No inflammatory response in the lamina propria was observed. Cryptosporidia were also commonly found in the glandular epithelium of submucosal lymphoglandular complexes in the colon. Cryptosporidium isolates from all farms were identified as Cryptosporidium suis using molecular markers (SSU rRNA). All of the C. suis strains obtained were larger [6.2 (6.0-6.8) x 5.5 (5.3-5.7) microm] than any isolate described so far [4.6 (4.4-4.9) x 4.2 (4.0-4.3) microm] and did not appear to be infective for neonatal BALB/c mice.     

Cloaca prolapse and cystitis in green iguana (Iguana iguana) caused by a novel Cryptosporidium species

Cryptosporidium infection was associated with colitis and cystitis in 2 green iguanas (Iguana iguana). The disease was characterized by a chronic clinical course of cloacal prolapses and cystitis. Histological examination of the gut and urinary bladder showed numerous Cryptosporidium developmental stages on the surface of the epithelium with mixed inflammatory response in the lamina propria. Cryptosporidium oocysts were visualised in a cytological preparation of the faeces. Based on the small subunit ribosomal RNA gene the cryptosporidia were characterized as belonging to the intestinal cryptosporidial lineage, but not to Cryptosporidium saurophilum or Cryptosporidium serpentis species.     

灌南县域肉羊源人兽共患隐孢子虫的流行病学调查及其类型分析

以隐孢子虫(Cryptosporidium spp.)18S rRNA为靶基因,通过巢氏PCR检测发现在采集的101份新鲜粪便样品中18份样本为阳性。不同地区羊场的隐孢子虫感染率分别为：李集镇36%(18/50),新安镇0%(0/30)、堆沟港镇0%(0/21)未检出隐孢子虫。但通过对4羊场的分析发现,有2个羊场(50%)为隐孢子虫感染阳性,且不同的羊场感染率差异显著,因此单纯的以地区来评价隐孢子虫的感染率,是值得商榷的。山羊隐孢子虫的感染率为33.3%,湖羊隐孢子虫的感染率为2%。2～6月龄的育肥羊隐孢子虫的感染率为36%,6～10月龄的育成羊(0%)。对检测为阳性的样品进行了隐孢子虫18S rRNA基因片段序列分析,发现18个样品全部为肖氏隐孢子虫(Cryptosporidium xiaoi),不存在泛在隐孢子虫(Cryptosporidium ubiquitum)。在检测隐孢子虫感染阳性的1个山羊场和1个羊湖羊场均存在肖氏隐孢子虫感染,不存在泛在隐孢子虫,更未发现肖氏隐孢子虫和泛在隐孢子虫的混合感染。目前的数据提示肖氏隐孢子虫对2～6月龄山羊(33.3%)和湖羊(2%)具有更高的...     

Aural-pharyngeal polyps associated with Cryptosporidium infection in three iguanas (Iguana iguana)

Cryptosporidium spp. infection was associated with aural-pharyngeal polyps in three iguanas (Iguana iguana). All iguanas were presented for masses protruding from the ear canal, and the disease was characterized by a chronic clinical course. The masses consisted of nests of cystic glands surrounded by abundant fibrous connective tissue and lined by hyperplastic cuboidal to pseudostratified columnar epithelium that was moderately to heavily colonized by cryptosporidial organisms. Electron microscopy revealed that the majority of organisms were trophozoites.     

Abomasal cryptosporidiosis in cattle

A 6-week-old calf and nine feedlot steers shed oocysts similar to Cryptosporidium muris-like oocysts. There were massive populations of this Cryptosporidium in the peptic glands of most of these animals. The oocysts were larger and more oval than the frequently reported type which is generated in the intestines of many animal species and thought to be similar to Cryptosporidium parvum. The pattern of shedding of this newly discovered Cryptosporidium in the steers was continuous over a period of months whereas the C. parvum-like oocysts cease to be shed 1 to several weeks after onset. The nature of the host-parasite interaction in abomasal cryptosporidiosis is yet to be determined. Morphologic changes that resulted from the interaction were an approximate 10% increase in abomasal mucosal thickness, widening of gland lumens in the middle region, and atrophy of epithelium in the same region.     

Genotypic characterization of cryptosporidium oocysts isolated from healthy people in three different counties of Korea

To investigate Cryptosporidium infection among healthy people, we collected stool samples from 150 healthy individuals in Gokseong, Muan, and Imshil Counties, southwest Korea, where neighbors on both an animal farm and a river respectively. In 12 of 150 samples, Cryptosporidium oocysts were detected by means of modified acid-fast staining. The bovine genotype, Cryptosporidium parvum, was identified by PCR/RFLP and 18S rRNA sequencing. C. parvum existed endemically in these areas, and the residents showed a relatively higher infection rate for C. parvum than that for C. hominis. Our results indicate that countermeasures against Cryptosporidium infection must be taken in these areas to ensure human health.     

Investigation of cryptosporidial infection in calves in San Bernardino County, California

Cryptosporidium is a protozoan parasite that inhabits the intestinal epithelium of calves, lambs, foals, pigs, and poultry. Of the 500 calves studied, 95 calves had diarrhea and 20 (21.05%) of those were infected with Cryptosporidium; only 8 (1.9%) of the remaining 405 nondiarrheic calves had Cryptosporidium oocysts in their feces.     

A novel multiplex polymerase chain reaction approach for detection of four human infective Cryptosporidium isolates: Cryptosporidium parvum, types H and C, Cryptosporidium canis, and Cryptosporidium felis in fecal and soil samples.

A nested multiplex polymerase chain reaction (PCR) approach was adopted for the simultaneous detection of 4 human infective genotypes of the protozoan parasite Cryptosporidium. Specific PCR primers were designed for the heat shock protein 70 gene of 2 genotypes of Cryptosporidium parvum (human and bovine types), Cryptosporidium canis, and Cryptosporidium felis. These 4 genotypes have all been found in human fecal samples. The primers amplified DNA fragments of specific sizes, each representing a unique genotype. The limit of detection of the method was found to vary between 10 and 100 oocysts per 1 ml fecal material. There appeared to be no cross-reactivity with other organisms commonly present in feces and soil, and the approach has a high specificity. The rapid identification of various human infective Cryptosporidium isolates is a part of the authors' long-term aim of determining the routes of infection with oocysts and thereby increase their epidemiological understanding of Cryptosporidium infection in humans and animals.     

Prevalence of Cryptosporidium spp. in pigs in Shanghai, China

A survey on prevalence of Cryptosporidium spp. in pigs at 12 farms in 8 suburban districts and 1 county of Shanghai was conducted under Sheather's sucrose flotation protocol and modified acid-fast stain methods from 2006 to 2009. A total of 2323 faecal samples were collected and Cryptosporidium spp. oocysts were detected in 800 samples (34.4%). Cryptosporidium was found in all 12 pig farms. Significant variations of prevalence were observed in different farms ranging from 14.1% to 90.6%. A follow-up survey on a positive pig farm for 13 consecutive months revealed that the most serious infection of Cryptosporidium spp. in pigs happened in winter and spring, and the lowest infection season was summer. Cryptosporidium spp. infection was mainly found in piglets within 2 months and no infection was found among those pigs of 90-180 days of age. The genotype analyses were carried out through PCR-RFLP and partial sequences analysis of small subunit ribosomal RNA (SSU rRNA) in some of the positive samples. Cryptosporidium suis (57/69, 82.6%), Cryptosporidium pig genotype II (6/69, 8.7%) and mixed infection of above two species/genotype (6/69, 8.7%) were found to be the main species/genotype in pigs in Shanghai area.     

Prevalence and age-related variation of Cryptosporidium species and genotypes in dairy calves

Fifteen dairy farms in seven states on the east coast of the US were each visited on two consecutive years to determinate the prevalence of Cryptosporidium species in pre-weaned (5 days to 2 months) and post-weaned calves (3-11 months), respectively. After each of 971 fecal specimens collected directly from each calf was sieved and subjected to density gradient centrifugation to remove debris and concentrate oocysts, specimens were examined by immunofluorescence microscopy, and polymerase chain reaction (PCR). For all PCR-positive specimens the 18S rRNA gene of Cryptosporidium was sequenced. Cryptosporidium was identified from all farms. Types of housing appeared to have no influence with regard to prevalence of infection. Of 971 calves, 345 were infected with Cryptosporidium (35.5%), but more pre-weaned calves (253 of 503; 50.3%) than post-weaned calves (92 of 468; 19.7%) were found to be infected. A total of 278 PCR-positive specimens characterized by gene sequencing revealed Cryptosporidium parvum, Cryptosporidium andersoni, and two unnamed Cryptosporidium genotypes Bovine B (AY120911) and deer-like genotype (AY120910). The prevalence of these Cryptosporidium species and genotypes appeared to be age related between pre- and post-weaned calves. C. parvum, the only zoonotic species/genotype, constituted 85% of the Cryptosporidium infections in pre-weaned calves but only 1% of the Cryptosporidium infections in post-weaned calves. These findings clearly demonstrate that earlier reports on the presence and prevalence of C. parvum in post-weaned cattle that were based solely on oocyst morphology must be reassessed using molecular methods to validate species and genotype. This finding also indicates that persons handling or otherwise exposed to calves under 2 months of age are at greater risk of zoonotic infection from Cryptosporidium than the risk of infection from exposure to older calves.     

Cryptosporidium infection in dogs in Osaka,Japan

Cryptosporidium parvum is a zoonotic pathogen composed of genetically distinct but morphologically identical genotypes. Recent molecular study indicates that dogs may transmit the cattle genotype, which is known to be pathogenic to humans. Although large-scale studies of Cryptosporidium infection in dogs have been performed in several countries, the isolates were not accurately identified because of the lack of a method for molecular analysis. It is important to identify the isolates harbored in dogs, which come in close contact with humans, in order to control human cryptosporidiosis. The aim of the present study was to calculate the prevalence of Cryptosporidium infection in dogs in Osaka city, Japan, and to characterize the isolates molecularly. The prevalence was determined to be 9.3% (13/140) by PCR. All isolates were found to be Cryptosporidium canis (previously known as the dog genotype), which is thought to be non-pathogenic in humans, based on the sequencing of diagnostic fragments. These results indicate that PCR-based diagnostic methods are a useful tool for the diagnosis and molecular epidemiology of Cryptosporidium infection in dogs, and that dogs living in Osaka are not a significant reservoir for human cryptosporidiosis. It is unclear why C. canis is dominant in dogs. Further study is required to understand this partial parasitism.     

The first report on Cryptosporidium suis and Cryptosporidium pig genotype II in Eurasian wild boars (Sus scrofa) (Czech Republic)

A total of 193 faecal samples of adult Eurasian wild boars were collected at 12 enclosures across the Czech Republic and examined for Cryptosporidium infection using both microscopic and molecular tools. Cryptosporidium oocysts were not detected in any of the 193 faecal samples examined using the aniline-carbol-methyl violet staining method. Thirty-two positive cases of Cryptosporidium infection were detected using either genus- or species-specific nested PCR. Mono-infection with Cryptosporidium suis and Cryptosporidium pig genotype II were found in 13 and 7 cases, respectively. Five mixed infections of C. suis and Cryptosporidium pig genotype II were detected using PCR/RFLP with genus specific primers. The number of detected mixed infections increased 2.4 fold when a species-specific PCR was employed. No other Cryptosporidium spp. was detected. Unlike cryptosporidiosis of domestic pigs, C. suis was detected as a dominant species infecting adult Eurasian wild boars. There was no association between diarrhoea and the presence of Cryptosporidium infection in the Eurasian wild boars studied. This is the first report on the Cryptosporidium infection caused by C. suis and Cryptosporidium pig genotype II in Eurasian wild boars (Sus scrofa).     

伊犁河谷新疆褐牛隐孢子虫感染情况调查

为了解伊犁河谷区域新疆褐牛隐孢子虫病流行情况,从该地区的伊宁市和察布查尔锡伯自治县的4个养殖场采集356份粪便样品,提取基因组DNA,以隐孢子虫小亚基核糖体DNA(SSU rDNA)为靶基因,进行套式PCR扩增,并对阳性样品进行序列分析.结果显示:伊宁市和察布查尔锡伯自治县各有1个养殖场的新疆褐牛发现隐孢子虫感染,总的...     

我国羊隐孢子虫感染情况及危险因素

隐孢子虫是可以感染小反刍动物的肠道原生生物,具有潜在的公共卫生学问题。隐孢子虫可引起人和动物的腹泻和肠道疾病,严重情况下会出现死亡。被感染的动物可能是人畜共患型隐孢子虫的宿主,可引起公共卫生风险、农场利润减少和动物福利等问题。隐孢子虫病已被报道是新生牛、羊等反刍动物腹泻和死亡的一个重要原因,认为是新生羔羊腹泻的第二大诱因,仅次于轮状病毒。羊隐孢子虫病在世界范围内都有发现,不同程度地威胁着人和动物的身体健康,基于此,现就隐孢子虫对羊感染的危险因素和常用检测方法,以及我国羊感染的现状进行综述,为其防控提供参考。     

Prevalence and pathogenicity of Cryptosporidium andersoni in one herd of beef cattle

Over a 35-week period from January to July 2002, a breed of Hereford beef cattle (H) and their hybrids were monitored. Five hundred and ninety-nine individual fecal samples from calves and 96 samples from their mothers were examined. First excretion of Cryptosporidium andersoni oocysts in calves was found in the 9th week after the start of calving (a calf 63-day old). The prevalence of C. andersoni in calves ranged from 11.1 to 92.9% depending on age. The mean prevalence in their mothers was found to be 43.8%. The size of oocysts was 8.48 +/- 0.78 x 6.41 +/- 0.59 microm. Infection intensity in calves ranged from 32 000 to 4 375 000 oocysts per gram (OPG) and in mothers from 78 000 to 2 552 000 OPG. Three cases of abomasal cryptosporidiosis slaughtered at the age of 81, 157 and 236 days were examined histologically and ultrastructurally [transmission electron microscopy (TEM) and scanning electron microscopy (SEM)]. Cryptosporidium infection of the abomasum was located in the upper half of the mucosal glands in the plicae spirales of the fundus, corpus and near the ostium omasoabomasicum. Cryptosporidia were not located in the glandular epithelium of the pars pylorica in the abomasum minimally 10 cm from pylorus. Histopathological changes in the site of cryptosporidial infection in the abomasum had a non-inflammatory character and included distinctive dilatation of infected parts of the glands with atrophy and metaplasia of the glandular epithelial cells, goblet cell activation and mucus hyperproduction. The TEM revealed a relatively small number of Cryptosporidium life cycle stages attached to glandular epithelial cells. In SEM the inner mucosal abomasal surface appeared swollen but was never infected by cryptosporidia.     

Natural infection with zoonotic subtype of Cryptosporidium parvum in Capybara (Hydrochoerus hydrochaeris) from Brazil

A total of 145 capybara (Hydrochoerus hydrochaeris) fecal samples from the state of S?o Paulo, Brazil, were screened for Cryptosporidium spp. oocysts using the malachite green method. Eight samples (5.52%) showed positive results and were further submitted to nested PCR reaction for amplification of fragments of 18S rRNA gene and 60-kDa glycoprotein gene for determination of species, alleles and subtypes of Cryptosporidium. Sequencing of the PCR products of the 18S rRNA gene fragments and 60-kDa glycoprotein gene fragments showed that for both genes all Cryptosporidium isolates from capybara were respectively 100% genetically similar to a bovine isolate of C. parvum and to C. parvum subtype IIaA15G2R1. To the best of our knowledge this is the first report of Cryptosporidium infection in this rodent. The finding of zoonotic C. parvum infection in a semi-aquatic mammal that inhabits anthroponotic habitats raises the concern that human water supplies may be contaminated with zoonotic Cryptosporidium oocysts from wildlife.     

分子生物学技术在隐孢子虫研究中的应用

隐孢子虫是顶复门原虫，主要寄生于人或动物的消化道上皮并引起腹泻，是寄生虫学、环境微生物学研究热点之一。分子生物学技术给隐孢子虫研究注入了强大动力，本文就其在隐孢子虫检测、分子流行病学调查、分类学以及药物设计等研究领域的应用作一综述。     

Molecular characterization of Cryptosporidium spp. in native breeds of cattle in Kaduna State, Nigeria

Despite numerous molecular epidemiologic studies of cryptosporidiosis in dairy cattle in industrialized countries, there are very few studies on the diversity and public health significance of Cryptosporidium species in native cattle in developing countries. In this study, a polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis of the small-subunit (SSU) rRNA gene was used to detect and identify Cryptosporidium spp. in 194 fecal specimens from 2 to 365 days old calves in 20 White Fulani and Sokoto Gudali herds in Nigeria. Thirty one (16.0%) of the specimens were positive for Cryptosporidium. Restriction digestion of the PCR products showed the presence of Cryptosporidium bovis (7.2%), Cryptosporidium ryanae (4.1%), Cryptosporidium andersoni (2.5%), and concurrent occurrence of C. bovis and C. ryanae (1.5%), and C. bovis and C. andersoni (0.5%). There were no significant differences (p>0.05) in Cryptosporidium infection rates by sex, herd location, management system, breed of calves, or fecal consistency. However, calves 180 days or younger had a higher infection rate of Cryptosporidium than older calves (p=0.034). Likewise, younger calves also had higher occurrence of C. bovis and C. ryanae (p=0.022). The absence of zoonotic Cryptosporidium parvum in the calves studied suggests that native breeds of cattle may not be important in the transmission of human cryptosporidiosis in Kaduna State, Nigeria.     

Morphological and immunohistochemical features of Cryptosporidium andersoni in cattle

Light and electron microscopic features and immunohistochemical features of Cryptosporidium andersoni (C. andersoni) and host reaction in the mucosa were studied. Although the affected cattle demonstrated no apparent clinical signs, a severe infection of C. andersoni was observed in the abomasum. C. andersoni were round in shape, measured 6-8 microm in size and were mainly observed to be freely located in the gastric pits, being attached in occasional cases to the surface of the abomasum epithelium. Frequent inflammatory cells had infiltrated the lamina propria of the affected mucosa, and frequent mitotic figures were observed in epithelial cells at the dilated isthmus. To access the cell kinetics, the number of epithelial cells infected with C. andersoni were counted and compared with noninfected cattle. The number of gastric pit cells in infected cattle was significantly higher than that in the controls. The number of proliferative cells determined by the Ki-67 antigen in C. andersoni infected cattle was also significantly higher than that in the controls. Transmission electron microscopy and scanning electron microscopy revealed that the morphology of the C. andersoni organism was common to those of other Cryptosporidium spp. Immunohistochemically, several commercial antibodies against Cryptosporidium spp. showed positive reactions at the wall of these oocysts or parasitophorous vacuoles. This report is possibly the first to discuss the prominent hyperplasia of the abomasum mucosa, as well as morphologic features of C. andersoni in cattle.