Cryptosporidiosis in two foals

Cryptosporidium oocysts were identified by phase contrast microscopy on smears from flotations of greenish-yellow pasty feces obtained from two foals. One foal, a one week old Percheron was recumbent, anorectic and lethargic, believed to be the result of a septicemia of undetermined etiology. Despite therapy and nursing care the animal died. Using light and electron microscopy, numerous stages of Cryptosporidium sp. were seen protruding from the surface of epithelial cells of intestinal villi. The other foal, a six week old Arabian had a mild diarrhea. The diarrhea and passage of oocysts eventually ceased. Immunological tests on sera of both these foals provided no evidence of abnormal immune function. This report is the first to describe cryptosporidiosis in apparently immunocompetent horses.     

Molecular characterization of Cryptosporidium parvum detected in Japanese black and Holstein calves in Iwate Prefecture and Tanegashima Island,Kagoshima Prefecture,Japan

Cryptosporidium oocysts were found in 43 out of 77 calves from two farms in Iwate Prefecture and nine farms on Tanegashima Island, Kagoshima Prefecture, Japan. The DNA fragments of 18S ribosomal RNA (18S rRNA) gene were amplified by a nested PCR from 43 oocyst-positive as well as one oocyst-negative samples. All of them were precisely identified as C. parvum by analyzing the nucleotide sequences of the 18S rRNA gene. C. parvum oocyst-positive calves ranged in age from 6 to 13 days old and significantly have watery diarrhea (P<0.05). Sequences of the gene encoding the 60-kDa glycoprotein (GP60) in 43 Cryptosporidium oocyst-positive samples were identical to that of the zoonotic IIaA15G2R1 subtype. We therefore suggest that calves could be potential sources of C. parvum infections in humans.     

Prevalence and age-related infection of Cryptosporidium suis, C. muris and Cryptosporidium pig genotype II in pigs on a farm complex in the Czech Republic

A total of 413 pig faecal samples were collected from pre-weaners (119), starters (131), pre-growers (123) and sows (40) from a farm with a closed breeding system segmented into two breeding complexes and a growing complex in the region of Vysočina, Czech Republic and screened for the presence of Cryptosporidium using staining methods and genotyping (SSU rRNA). Cryptosporidium oocysts were detected by microscopy in the faeces of 21.1% of the samples (87/413). Sequence analyses and RFLP identified C. suis in 44, Cryptosporidium pig genotype II in 23 and C. muris in 2 samples. No mixed infections were found.Pigs under 7 weeks of age were infected with C. suis only. Cryptosporidium pig genotype II was found in animals from 7 weeks of age. No relationship was found between diarrhoea and any Cryptosporidium infection in any of the different age groups (P < 0.05). The pre-weaned pigs shed significantly more Cryptosporidium oocysts than older pigs and it was associated with C. suis infection.     

Comparative evaluation of Cryptosporidium infection in malnourished and well-nourished children: Parasitic infections are affected by the interaction of nutritional status and socio-demographic characteristics

Cryptosporidium, as a small protozoan parasite, is a leading cause of persistent diarrhea in children in developing countries and has both a short and long-term impact on the growth of children. In the present study, Cryptosporidium infection was compared in malnourished and well-nourished children by modified acid-fast staining, nested-polymerase chain reaction (nested-PCR) and loop-mediated isothermal amplification (LAMP) methods. As a case-control study, Cryptosporidium infection in 94 malnourished children was evaluated and compared with those of 188 age and gender-matched well-nourished children. Oocysts of Cryptosporidium were detected by modified acid-fast staining method. The extracted DNA was amplified by nested-PCR and LAMP techniques. In addition, positive amplicons were directly sequenced for phylogenetic analysis. Cryptosporidium oocysts were found in the stools of two (2.12 %) children who were hospitalized and had diarrhea by nested-PCR while three isolates (3.2 %) were found by LAMP. Cryptosporidium-positive children were more malnourished compared to those who were negative for Cryptosporidium infection but this important finding was not statistically significant. C. parvum was the main species of Cryptosporidium detected in malnourished children in northwest Iran. LAMP can be considered as a sensitive field monitoring assay in patients with low parasite burden. Nutritional status and socio-demographic factors may have interactive effects on the incidence and severity of parasitic diseases.     

Captive Agamid lizards in Germany: Prevalence,pathogenicity and therapy of gastrointestinal protozoan and helminth infections

Reptiles are becoming popular pets in many parts of the world. They are also known to harbor numerous gastrointestinal parasites. We used faecal smears to examine 748 stool samples from 14 different agamid lizard species. In addition, we used coproantigen ELISA tests (11 samples) and immunofluorescence assays (IFA) (19 samples) to detect reptile Cryptosporidium infections. In 28 cases, veterinarians requested therapy to treat oxyurid- and/or Isospora amphiboluri-infections and resent fecal samples after proposed therapy and anti-parasitic treatments had been applied. We also performed complete dissections of 24 deceased agamas in order to specify protozoan and helminth parasite infections.Overall, the examined fecal samples contained 6 different taxa. Oxyurids (Pharyngodonidae) were the most prevalent nematodes (41.2%), followed by I. amphiboluri (17.0%), Entamoeba spp. (0.8%), Choleoeimeria spp. (0.5%), Trichomonas spp. (0.3%), Cryptosporidium spp. (0.3%) and Strongyloides-like nematodes (0.1%). I. amphiboluri infections were significantly more prevalent (Chi-square test: χ² = 21,5, df = 1, P < 0.001) in juvenile agamid lizards (31.9%) than in adults (14.2%). One of 11 (9.1%) coproantigen ELISA-examined samples was positive for Cryptosporidium. In 10.5% of the samples we found oocysts of Cryptosporidium. Thirteen (54.2%) of necropsied agamid lizards were infected with endoparasites and it is likely that three (12.5%) of them died due to severe parasitic infections. 74.0% of the samples that were submitted after therapy had been applied were negative. The high prevalences and pathological findings of several clinical parasitoses observed in these exotic reptiles calls for more detailed investigations on agamid gastrointestinal parasite fauna.     

Prevalence of Cryptosporidium species isolated from HIV/AIDS patients in southwest of Iran

This study aimed to determine the prevalence and species of Cryptosporidium among HIV/AIDS patients in southwest of Iran. Two hundred fifty faecal samples from HIV patients were examined for the presence of Cryptosporidium oocysts using a conventional coproscopic approach. Such oocysts were detected in 18 (7.2%) out of 250 faecal samples. Genomic DNAs from 250 samples were then subjected to a nested-PCR-RFLP technique targeting different loci of 18S rRNA gene for species identification. Out of 250 samples, 27 (10.8%) were positive for different Cryptosporidium spp; Restriction patterns resulting from the digestion of the nested amplicon with restriction endonucleases VspI and SspI showed that C. parvum (70.38%) was the most prevalent species, followed by C. hominis (25.92%) and C. meleagridis (3.7%), respectively. The mean CD4+ T-cell count was 215 cells/μL. There was a strong association between cryptosporidiosis and CD4+ T-cell count (P = 0.000) with the highest prevalence recorded among patients with CD4+ T-cell count < 200 cells/μL. This confirms that there is a low opportunity for this parasite to get established as the patients CD4+ T-cell count increases. Also HIV infection increased the risk of having Cryptosporidium. Our epidemiological findings are useful for any preventive intervention to control disease diffusion.     

An outbreak of massive mortality among farm rabbits associated with Cryptosporidium infection

Cryptosporidium in farm rabbits is not often recognised due to a low prevalence and asymptomatic course of infection. Nonetheless, incidences of fatal diarrhoeic diseases are frequently noticed in the rabbitries. In this article, we report an outbreak where there was massive mortality among farm rabbits associated with Cryptosporidium infection. The disease was characterised by profuse diarrhoea resulting in the death of rabbits. A pooled faecal sample was screened for a presence of parasites using microscopy methods. In the tested sample no other parasites other than Cryptosporidium oocysts were found. Further identification of the parasite species was performed at a molecular level, using the 18 SSU rRNA, COWP and LIB13 PCR followed by a subtyping at the GP60 gene locus. Sequence analysis of GP60 gene fragment revealed the presence of a novel subtype VbA24 of Cryptosporidium cuniculus. In this outbreak a Cryptosporidium protozoan parasite played a major role in the etiology of the gastrointestinal disorders in rabbits resulting in massive mortality of the infected animals.     

Molecular characterisation of Cryptosporidium isolates from rivers,water treatment plants and abattoirs in Ibadan,Nigeria

To understand the molecular characteristics of Cryptosporidium species contaminating rivers, water treatment plants and abattoirs in Ibadan Nigeria, water samples were obtained from ten rivers used for household and agricultural purposes, three major functional water treatment plants and three major abattoirs located within Ibadan metropolis during dry and rainy seasons between November, 2016 to October, 2017. Obtained samples were examined for Cryptosporidium oocysts using microscopy after using modified formalin–ether concentration method and modified acid-fast staining. Cryptosporidium oocysts were detected in samples from five rivers with mean oocyst count/field ranging from 7.70 ± 0.57–1.34 ± 0.57, oocysts were also detected in samples from two abattoirs with mean oocyst count/field ranging from 4.60 ± 0.33–2.50 ± 0.33. Genomic DNA were extracted from microscopy positive river and abattoir samples using sucrose gradient purification method and genotypes and subtypes of parasites were detected by nested PCR amplification and nucleotide sequence analysis of both 18S rRNA and 60-kDa glycoprotein (gp60) genes. Cryptosporidium parvum, C. muris and C. fragile were the only genotypes detected in some river samples, while gp60 gene sequence analysis showed that the C. parvum strain detected was subtype IIa. This study provides evidence that rivers used for household and agricultural purposes in studied area may be potential reservoirs and infection sources for Cryptosporidium species and zoonotic subtypes of public health importance.     

The prevalence of <Emphasis Type="Italic">Cryptosporidium</Emphasis> species in diarrhoeic lambs in Kars province and potential risk factors

This study was carried out to determine the prevalence of Cryptosporidium species in diarrhoeic lambs and investigate some risk factors in Kars province (Northeastern region of Anatolia) in Turkey. Four hundred faecal samples were taken from the rectums of clinically diarrhoeic and aged to 1-month-old lambs from 34 sheep farms in 20 villages in March-April 2007 and examined by using the modified acid-fast staining technique. The prevalence of Cryptosporidium species was found as 38.8% (155/400). Cryptosporidium oocysts were detected in 90.0% (18/20) of villages and in 76.5% (26/34) of the sheep farms. Infection rates were detected as: 44.4% (67/151) in 1-week-old lambs, 37.5% (39/104) in 2-week-old lambs, 40.0% (38/95) in 3-week-old lambs, and 22.0% (11/50) in 4-week-old lambs. Farms classified according to their zoohygienic conditions and fine, average and bad conditioned farms were contaminated with Cryptosporidium with the percentages of 14.7%, 20.6% and 41.2%, respectively. Clinical cryptosporidiosis was determined in 35.0% of the villages (7/20) and in 29.4% of the sheep farms (10/34), Cryptosporidium oocysts were found in 81.3% of the lambs (91/112) in these farms. Cryptosporidiosis may be a major epidemiological significance in lambs in Kars province, and suggests that naturally infected lambs may be reservoirs of Cryptosporidiosis infections for calves even for humans too.     

Occurrence of Cryptosporidium and Giardia on beef farms and water sources within the vicinity of the farms on Prince Edward Island, Canada

The objectives of this study were to determine the prevalence and assemblages of Giardia and species of Cryptosporidium on beef farms in Prince Edward Island (PEI), Canada, including the water sources associated with the farms, and to determine risk factors for infection of cattle with these parasites. Twenty beef farms were selected based on the presence of surface water < 500 m from the barn. Prevalence was determined by direct immunofluorescence microscopy, while genotyping and species determination were performed by nested-PCR and DNA sequencing. Giardia was detected in 42% (95% CI: 38-46%) of fecal samples from 100% farms while Cryptosporidium was detected in 17% (95% CI: 14-19%) of fecal samples from 80% of farms. The most predominant Giardia assemblage isolated was the livestock specific assemblage E (89%). The zoonotic assemblages A and B were found in 4 and 7% of the Giardia isolates that were genotyped, respectively. The Giardia assemblages were detected equally between the cows and calves examined. Overall, the most common Cryptosporidium species detected in this study was Cryptosporidium andersoni (49%), predominantly found in cattle >6 mo of age, while most Cryptosporidium bovis and Cryptosporidium pestis (previously Cryptosporidium parvum ‘bovine genotype’) isolates were detected in calves ≤ 6 mo of age. All Cryptosporidium ryanae isolates (four) were found in calves. Giardia cysts and Cryptosporidium oocysts were detected in 14 and 93% of surface water samples of 14 farms, respectively. Cryptosporidium oocysts were detected in three (15%) ground water samples of 20 farms. One Cryptosporidium-positive water sample, which was the only surface water sample amenable to genotyping, contained C. parvum. The farm-level risk factors investigated in this study, age of animals and location of the farm, were not associated with the risk of infection in cattle with either Cryptosporidium spp. or Giardia duodenalis.We conclude that beef cattle are a potential reservoir of Cryptosporidium spp. and G. duodenalis that could contaminate source water. There is the possibility of further transmission to humans on PEI if the source water is not properly treated prior to consumption.     

Identity and public health potential of Cryptosporidium spp. in water buffalo calves in Egypt

Little is known about the diversity and public health significance of Cryptosporidium species in water buffaloes. In this study, we examined the distribution of Cryptosporidium spp. in water buffalo calves in Egypt. Rectal fecal specimens from 179 calves and 359 adults were screened microscopically for Cryptosporidium oocysts using modified Ziehl–Neelsen stain. Cryptosporidium spp. in 17 microscopy-positive specimens from calves were genotyped by DNA sequence analysis of the small-subunit rRNA gene, and Cryptosporidium parvum was subtyped by sequence analysis of the 60 kDa glycoprotein gene. Cryptosporidium ryanae was found in 10 specimens and C. parvum in 7 specimens, with the former belonging to the newly identified C. ryanae buffalo variant and the latter belonging to the subtypes IIdA20G1 (in 5 specimens) and IIaA15G1R1 (in 2 specimens). The prevailing occurrence of C. ryanae and the subtype family IId of C. parvum and the absence of C. bovis and C. andersoni represent some features of Cryptosporidium transmission in water buffaloes in Egypt.     

Prevalence and molecular characterization of Cryptosporidium spp. in dairy cattle from farms in China

Fecal samples of 2,056 dairy cattle from 14 farms were collected in three geographical regions of China and stained using a modified acid-fast staining technique to identify Cryptosporidium oocysts. A total of 387 (18.82%) positive samples were identified and further analyzed by polymerase chain reaction (PCR) using primers designed to amplify DNA fragments from the small subunit ribosomal RNA. The PCR products were sequenced and the sequences were deposited in the GenBank database under accession numbers EU369377-84 and GU070730-33. Phylogenetic analysis was performed and a distances matrix generated from these sequences confirmed the existence of Cryptosporidium (C.) parvum ''mouse'' genotype, C. bovis, C. andersoni, C. hominis, and C. serpentis in cattle. These results represent the first report on the prevalence and genetic identification of Cryptosporidium species, and may contribute to a better understanding of the epidemiology of Cryptosporidium in cattle in China.     

Detection of a novel genotype of Cryptosporidium in Antarctic pinnipeds

A study was conducted to investigate the presence of Cryptosporidium and Giardia in Antarctic marine mammals. A total of 270 faecal samples from different species of pinnipeds from different locations in the South Shetland Islands and Antarctic Peninsula were analysed by immunofluorescence microscopy and PCR. Cryptosporidium was detected by PCR in three samples from Southern elephant seals (Mirounga leonina) and 2 Weddell seals (Leptonychotes weddellii). However, no oocysts were observed in any of the samples by immunofluorescence microscopy. Molecular characterisation of the isolates, using the 18S rDNA, the HSP70 and the COWP loci, revealed the presence of a Cryptosporidium sp., previously reported from an Antarctic Southern elephant seal, in the elephant seals and a novel genotype in Weddell seals. Giardia could not be detected in any of the samples analysed.     

Molecular detection of Cryptosporidium spp. infections in water buffaloes from northeast Thailand

The objectives of this study were to determine the individual and herd-level prevalence and genotype of Cryptosporidium and to identify putative risk factors associated with Cryptosporidium spp. infections in water buffaloes in northeast Thailand. Fecal samples from 600 water buffaloes of 287 farms in six provinces were collected and tested using DMSO-modified acid-fast staining and polymerase chain reaction. The overall prevalence of Cryptosporidium infections in buffaloes was 5.7 and 8.7 % among individual animals and herds, respectively. The provinces with highest infected Cryptosporidium were located in the Sakon Nakhon Basin in the northern part of the region. In addition, higher herd prevalence was observed among farms with more than five buffaloes (30 %) than those with five or less animals (16.2 %). Thirty (88.2 %) of the 34 Cryptosporidium-positive samples were Cryptosporidium parvum and four (11.8 %) were Cryptosporidium ryanae.     

Seroprevalence of Toxoplasma gondii and Neospora caninum in feral cats (Felis silvestris catus) in Majorca, Balearic Islands, Spain

Cryptosporidium spp. are common intestinal protozoan parasites that infect a wide range of hosts, including humans and livestock, worldwide. The objective of this study was to determine the prevalence of Cryptosporidium spp. in dairy calves in Prince Edward Island, Canada, and the potential for transmission of this parasite between dairy calves and humans. Fecal samples were collected from 183 dairy calves from 11 farms in Prince Edward Island. The prevalence of Cryptosporidium spp. infections in these animals was determined by examining for the presence of oocysts in the fecal samples, using immunofluorescence microscopy. Molecular characterization was done using a nested-PCR protocol to amplify fragments of the Cryptosporidium heat-shock protein 70 gene, followed by DNA sequencing. Ten calves (6.2%), representing 4 out of 11 farms tested, were positive for Cryptosporidium spp. DNA sequence analysis on five PCR positive samples demonstrated that Cryptosporidium parvum was the only species present in the calves tested, suggesting that there is a potential risk of zoonotic transmission between dairy calves and humans in this region.     

Cryptosporidium Recovered from Musca domestica,Musca sorbens and Mango Juice Accessed by Synanthropic Flies in Bahirdar,Ethiopia

The study was conducted to determine the role of house flies, Musca domestica and Musca sorbens to carry Cryptosporidium species in natural environment and filth flies potential for contamination of food item they visited using acid‐fast stain technique. Cryptosporidium was identified from flies collected in dairy cow barns, butchery, market and defecating grounds. Musca domestica captured from dairy cow barns and M. sorbens from defecating ground were found carrying more oocyst of Cryptosporidium parvum. Oocyst load per fly for M. domestica and M. sorbens was 5.84 and 3.42, respectively. Flies’ population dynamics in each month had little relation to the monthly oocyst frequency, r = 0.06 and 0.02 for M. domestica and M. sorbens, respectively. Cryptosporidium species oocysts were isolated from frozen mango juice, which filth flies visited in dairy farm barn. Load of oocysts in the mango juice was dependent on time contact of flies with mango juice and more oocysts were recovered (P < 0.05) in mango juice samples accessed by filth flies for longer period. Role of filth flies to carry and deposit Cryptosporidium species oocyst for development of food‐borne cryptosporidiosis is signified.     

A study of neonatal cryptosporidosis of foals in New Zealand

Abstract

AIM: To assess the occurrence of Cryptosporidium oocysts in faecal specimens from foals, and investigate an outbreak of neonatal cryptosporidiosis in foals revealed in the course of the study.

METHODS: Faecal specimens from foals received by a diagnostic veterinary laboratory in New Zealand between 2006 and 2007 were submitted to Massey University and tested microscopically for the presence of Cryptosporidium oocysts. The Cryptosporidium isolates in the oocyst-positive specimens were genetically identified to species level. In addition, specimen submission data from the participating laboratory for 2005–2007 were examined. In the course of the study, the identification of one Cryptosporidium-positive specimen triggered an on-farm investigation.

RESULTS: Twelve faecal specimens submitted by the participating laboratory between 2006 and 2007 were tested further, and three were positive for C. parvum. Specimen submission records indicated a total of 67 faecal specimens were tested for Cryptosporidium by the participating laboratory between 2005 and 2007; 12 (18%) were positive. The on-farm investigation on a broodmare farm revealed a high incidence of neonatal diarrhoea in foals; C. parvum was the only enteropathogen found in the faeces of 4/4 affected foals examined. The diarrhoea in all those foals was self-limiting, manifesting during the second week of life, resembling foal heat diarrhoea, and accompanied by a short but intense period of shedding oocysts.

CONCLUSIONS AND CLINICAL RELEVANCE: The fact that Cryptosporidium parasites were identified in 18% of faecal specimens from foals analysed for this agent in 2005–2007 by the participating laboratory indicated that infection with this agent in foals is not uncommon.

Collectively, the results of this and previous studies performed in New Zealand indicate C. parvum is a cause of diarrhoea in newborn foals, potentially accounting for a proportion of cases empirically diagnosed as foal heat diarrhoea. It is therefore advisable to take precautions when handling diarrhoeic foals, until this potentially zoonotic agent is ruled out in the laboratory.     

Zoonotic Cryptosporidium parvum in Romanian newborn lambs (Ovis aries)

This study was undertaken to investigate the occurrence and public health significance of Cryptosporidium species/genotypes and subtypes in a newborn lambs. A total of 175 diarrheic fecal samples from lambs (younger than 21 days) were collected in seven sheep flocks located in western Romania, and were microscopically examined for the presence of Cryptosporidium oocysts after staining with modified Ziehl–Neelsen technique. Twenty-four (13.7%) fecal samples were tested Cryptosporidium positive by microscopy and were subjected for molecular characterization. All positive samples were successfully amplified through a nested polymerase chain reaction (PCR) of the small subunit (SSU) rRNA gene (18S). Cryptosporidium species were determined by restriction fragment length polymorphism (RFLP) analysis of the secondary PCR products using the conventional SspI and VspI restriction enzymes. The identified species were: Cryptosporidium parvum (20/24), C. ubiquitum (2/24) and C. xiaoi (2/24), respectively. PCR-RFLP results for C. ubiquitum and C. xiaoi isolates were confirmed by DNA sequencing. Subsequently, subtyping of seven randomly selected C. parvum isolates, based on sequence analysis of the GP60 gene, revealed the presence of five different subtypes (IIaA17G1R1, IIaA16G1R1, IIdA20G1, IIdA24G1 and IIdA22G2R1) belonging in two zoonotic subtype families (IIa and IId). These findings may suggest the potential role of the newborn lambs as a source for human cryptosporidiosis. This is the first published report about the presence of C. ubiquitum and C. xiaoi in lambs from Romania.     

Circulating antibody response to Eimeria tenella oral and subcutaneous infections in chickens

Immune responses in chickens to Eimeria tenella using oral and subcutaneous routes of infection were investigated. The results obtained indicated that sporulated oocysts inoculated subcutaneously in doses up to 50 000 oocysts per bird were not fatal to 21-day-old chicks. Subcutaneous inoculation of oocysts was found to be less immunogenic than oral administration. The dynamics of the antibody responses were different for the two routes of infection. Orally administered oocysts stimulated a dramatic primary increase in the serum antibody titre with a tendency towards a decrease in the titre 14 days post infection irrespective of second infections at that time. However, a third oral dose of oocysts stimulated a slight increase in antibody titre. Two doses of oocysts injected subcutaneously induced only a slight increase in serum antibody titre. Such a low titre was dramatically increased following a subsequent oral dose of oocysts. Antibodies specific to E. tenella are IgM and IgG immunoglobulins. IgA immunoglobulin was not investigated.     

The prepatent and patent periods of Eimeria alabamensis and further description of the exogenous stages

Eimeria alabamensis infections were established in calves 5 to 6 weeks of age by adminestering 10 million, 80 million, or 100 million sporulated oocysts. The prepatent period was 6 to 8 days (mean 6.6). Oocyst discharges usually lasted for 2 to 3 days although a few calves passed oocysts throughout the rest of the 3-week observation period. Calves with oocyst discharge exceeding 1 million oocysts per g of feces had a moderate diarrhea at the time of peak oocyst discharge. No other clinical signs were observed in any of the infected calves. Reinoculations with 100 million sporulated oocysts given 3 weeks after the initial inoculations of 10 million or 80 million oocysts resulted in infections characterized by greatly reduced oocyst discharges. Sporulated oocysts of E. alabamensis were 16 to 24 μ by 12 to 16 μ and were usually ovoid. The oocyst wals consisted of two layers. Sporocysts were elongate-ellipsoid, had a distinct Stieda body, and were 10 to 12 μ by 4 to 6 μ. Completely sporulated oocysts were first observed after 5 days at 25 °C, and most were sporulated after 8 days. Oocysts did not sporulate at 4 °C, 33 °C, and 37 °C.