Parvum Infection Research Articles

Prevalence of Cryptosporidium parvum in dairy calves and GP60 subtyping of diarrheic calves in central Argentina.

Cryptosporidiosis of calves is caused by the enteroprotozoan Cryptosporidium spp. The disease results in intense diarrhea of calves associated with substantial economic losses in dairy farming worldwide. The aim of this study was to determine calf, herd, and within-herd Cryptosporidium prevalence and identify Cryptosporidium species and subtypes in calves with diarrhea in intensive dairy herds in central Argentina. A total of 1073 fecal samples were collected from 54 randomly selected dairy herds. Cryptosporidium-oocysts were isolated and concentrated from fecal samples using formol-ether and detected by light microscopy with the modified Ziehl-Neelsen technique. Overall prevalence of oocyst-excreting calves was found to be 25.5% (274/1073) (95% C.I. 22.9; 28.1%). Of the herds studied, 89% (48/54) included at least one infected calf, whereas within-herd prevalence ranged from the absence of infection to 57% (20/35). A highly significant association was found between the presence of diarrhea and C. parvum infection (χ2 = 55.89, p < 0.001). For species determination, genomic DNA isolated from oocyst-positive fecal samples was subjected to PCR-RFLP of the 18S rRNA gene resulting exclusively in Cryptosporidium parvum identification. C. parvum isolates of calves displaying diarrhea and high rate of excretion of oocysts were subtyped by PCR amplification and direct sequencing of the 60 kDa glycoprotein (GP60) gene. Altogether five GP60 subtypes, designated IIaA18G1R1, IIaA20G1R1, IIaA21G1R1, IIaA22G1R1, and IIaA24G1R1 were identified. Interestingly, IIaA18G1R1 and IIaA20G1R1 were predominant in calves with diarrhea and high infection intensity. Notably, IIaA24G1R1 represents a novel, previously unrecognized C. parvum subtype. The subtype IIaA18G1R1, frequently found in this study, is strongly implicated in zoonotic transmission. These results suggest that calves might be an important source for human cryptosporidiosis in Argentina.Electronic supplementary materialThe online version of this article (10.1007/s00436-019-06366-y) contains supplementary material, which is available to authorized users.

Delivery of SA35 and SA40 peptides in mice enhances humoral and cellular immune responses and confers protection against Cryptosporidium parvum infection

BackgroundCryptosporidium parvum is a major cause of diarrhea in children and ruminants at the earliest stages of life. Maternal antibodies represent the main shield of neonate mammals for most of the infections. Two recombinant antigens (SA35 and SA40), portions of two C. parvum proteins, were tested for their ability to induce immune responses in adult mice and for protection on neonate BALB/c mice born from females immunised by mucosal delivery of both peptides.MethodsAdult BALB/c mice were intraperitoneally immunised with SA35 and SA40, separately or mixed, and their immune response was characterised. Furthermore, BALB/c pregnant mice were immunised by mucosal delivery with an SA35/40 mix, before and during pregnancy. Soon after birth, their offspring were infected with two doses (1 × 105 and 5 × 103) of C. parvum oocysts and the parasitic burden was determined at 5 and 9 days post-infection.ResultsIntraperitoneal immunisation with SA35 and SA40 induced specific IgG and IgG1 in serum, specific IgA in the intestinal mucosa, increase of CD3+/CD4+ and CD30+ cells in splenocytes, which produced IFN-γ. Neonates born from immunised mice and infected with 1 × 105 oocysts showed a significant reduction of oocysts and intestinal forms (23 and 42%, respectively). A reduction of all parasitic forms (96%; P < 0.05) was observed when neonates were infected with 5 × 103 oocysts.ConclusionsSA35 and SA40 peptides induce specific humoral and cell-mediated immune responses to C. parvum in adult mice. Moreover, mucosal administration of the SA35/40 mix in pregnant mice reduces C. parvum burden in their litters.

Outbreak of cryptosporidiosis due to Cryptosporidium parvum subtype IIdA19G1 in neonatal calves on a dairy farm in China

Neonatal diarrhea is one of the most important syndromes in dairy cattle. Among enteropathogens, Cryptosporidium spp. are primary causes of diarrhea, but outbreaks due to cryptosporidiosis are rarely reported in cattle. From January to April in 2016, severe diarrhea was observed in over 400 neonatal dairy calves on a large dairy farm in Jiangsu Province of East China. Approximately 360 calves died due to watery diarrhea despite antibiotic therapy. In this study, 18 fecal specimens were collected from seriously ill calves on this farm during the diarrhea outbreak, and analysed for common enteropathogens by enzymatic immunoassay (EIA). In a post-outbreak investigation, 418 and 1372 specimens collected from animals of various age groups were further analysed for rotavirus and Cryptosporidium spp. by EIA and PCR, respectively, to assess their roles in the occurrence of diarrhea on the farm. Cryptosporidium spp. were genotyped using established techniques. Initial EIA tests showed that 15/18 seriously ill calves during the outbreak were positive for Cryptosporidium parvum, while 8/18 were positive for rotavirus. The overall infection rate of Cryptosporidium in pre-weaned calves on the farm was 22.7%, with odds of the Cryptosporidium infection during the outbreak 4.4–23.5 times higher than after the outbreak. Four Cryptosporidium spp. were identified after the outbreak including C. parvum (n = 79), Cryptosporidium ryanae (n = 48), Cryptosporidium bovis (n = 31), and Cryptosporidium andersoni (n = 3), with co-infections of multiple species being detected in 34 animals. Infection with C. parvum (73/79) was found in the majority of calves aged ≤3 weeks, consistent with the age of ill calves during the outbreak. All C. parvum isolates were identified as subtype IIdA19G1. In the post-outbreak investigation, C. parvum infection was associated with the occurrence of watery diarrhea in pre-weaned calves, C. ryanae infection was associated with moderate diarrhea in both pre- and post-weaned calves, while no association was identified between rotavirus infection and the occurrence of diarrhea. Results of logistic regression analysis further suggested that C. bovis infection might also be a risk factor for moderate diarrhea in calves. Thus, we believe this is the first report of a major outbreak of severe diarrhea caused by C. parvum IIdA19G1 in dairy calves. More attention should be directed toward preventing the dissemination of this virulent subtype in China.

Induction of inflammatory responses in splenocytes by exosomes released from intestinal epithelial cells following Cryptosporidium parvum infection

Abstract Cryptosporidium, a protozoan parasite that infects the gastrointestinal epithelium and other mucosal surfaces in humans and animals, is an important opportunistic pathogen in AIDS patients and one of the most common enteric pathogens affecting young children in developing regions. We previously demonstrated that C. parvum infection stimulates host epithelial cells to release exosomes and these released exosomes shuttle several antimicrobial peptides to carry out anti-C. parvum activity. Murine intestinal epithelial (IEC4.1) cells and neonatal mice were applied for infection. Primary splenocytes were exposed to exosomes isolated from infected cells or un-infected controls. Inflammatory response in splenocytes were measured. We detected upregulation of inflammatory genes in the liver and spleen following C. parvum intestinal infection in neonatal mice. Interestingly, exosomes released from intestinal epithelial cells following C. parvum infection could activate the nuclear factor kappa B signaling pathway and trigger inflammatory gene transcription in isolated primary splenocytes. Several epithelial cell-derived proteins and a subset of parasite RNAs were detected in the exosomes released from C. parvum-infected intestinal epithelial cells. Shuttling of these effector molecules, including the high mobility group box 1 protein, was involved in the induction of inflammatory responses in splenocytes induced by the released exosomes from infected cells. Our data indicate that exosomes released from intestinal epithelial cells upon C. parvum infection can activate immune cells through shuttling various effector molecules, a process that may be relevant to host systemic responses to Cryptosporidium infection.

Induction of a long non-coding RNA, lncRNA-Chr1:1226, by Cryptosporidium infection primes intestinal epithelial cells for IFN-γ-mediated host antimicrobial gene transcription

Abstract Cryptosporidium, a protozoan parasite that infects the intestinal epithelium and other mucosal surfaces in animals and humans, is an important opportunistic pathogen in AIDS patients and a common cause of diarrhea in young children in developing countries. Intestinal epithelial cellular defense is key to innate mucosal anti-Cryptosporidium defense but underlying molecular mechanisms are still obscure. Here, we identified several long non-coding RNAs (lncRNAs) that are predominantly expressed in intestinal epithelial cells. Several of such epithelial-enriched lncRNAs, such as lncRNA-Chr1:1226, were upregulated in cells following C. parvum infection. Induction of lncRNA-Chr1:1226 in infected intestinal epithelial cells was controlled by TLR4/NF-κB/Cdc42 signaling and epithelial specific transcription factor Eif3. Induction of lncRNA-Chr1:1226 promoted IFN-γ-mediated epithelial antimicrobial defense, through facilitating STAT1/SWI/SNF-associated chromatin remodeling to promote IFN-γ-mediated transcription of defense genes in intestinal epithelial cells. We observed that IFN-γ-mediated antimicrobial defense was suppressed in neonatal intestinal epithelium. Expression of PRDM1 in the neonatal intestinal epithelium might contribute to suppression of IFN-γ-mediated antimicrobial gene transcription. Furthermore, PRDM1 interacted with lncRNA-Chr1:1226 and PIAS1 to attenuate SWI/SNF-mediated antimicrobial transcription induced by IFN-γ in intestinal epithelium of neonates. Our data demonstrate that lncRNAs, particularly epithelial lncRNAs, may be key regulators in IFN-γ-mediated epithelial defense.

OR22: Altered fetal hemodynamic status and neuroinflammation in a nonhuman primate model of intrauterine Ureaplasma parvum infection

American Journal of Reproductive ImmunologyVolume 81, Issue S1 p. 37-37 ABSTRACTS OR22: Altered fetal hemodynamic status and neuroinflammation in a nonhuman primate model of intrauterine Ureaplasma parvum infection First published: 31 May 2019 https://doi.org/10.1111/aji.21_13119Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume81, IssueS1Special Issue: The 39th Annual Meeting of the American Society for Reproductive Immunology, 12-15 June 2019, Grand Rapids, Michigan, USAMay 2019Pages 37-37 RelatedInformation

Long-term use of yeast fermentation products in comparison to halofuginone for the control of cryptosporidiosis in neonatal calves

The objective of this study was to compare the effect of non-GMO Saccharomyces cerevisiae fermentation products (SCFP) with that of a halofuginone treatment against Cryptosporidium parvum infection in pre-weaned calves on a commercial dairy farm. A total of 123 neonatal female calves, housed in individual hutches, were enrolled sequentially based on date of birth in 41 blocks of 3 animals each. Calves within each block were allocated to one of 3 treatments: remaining untreated, fed with SCFP (Diamond V SmartCare® at 1 g/d in milk and NutriTek® at 5 g/d in starter grain) for the first 63 days of life, or treated with halofuginone (0.1 mg/kg/d) for the first 7 days of life. Fecal samples collected on days 4–21 post-partum were examined for both Cryptosporidium oocysts and coproantigen. The presence and intensity of diarrhea were monitored by scoring daily for the first 4 weeks of life. Calves were weighed at 0, 21, 42 and 63 days of age. Almost all calves were Cryptosporidium-positive at least once during the study. Halofuginone significantly reduced the number of Cryptosporidium-positive fecal samples as compared to the two other groups. Based on the coproantigen scores, both halofuginone and SCFP feeding significantly reduced the intensity of Cryptosporidium infection as compared to the untreated group. Diarrhea was recorded in almost all calves at least once. Neither the proportion of diarrheic calves nor the intensity and duration of diarrhea differed among the 3 treatment groups significantly. The mean daily weight gain during the first 3 weeks of life was significantly lower in halofuginone treated calves than in both other groups; however, at the end of the study period the total weight gain did not significantly differ among the 3 treatment groups. In conclusion, the clinical results and weight gains of pre-weaning supplementation with the SCFP were neither better nor worse than the 7-day halofuginone treatment suggesting that the SCFP feeding may be from the clinical point of view a natural alternative measure, instead of halofuginone treatment, in bovine cryptosporidiosis.

Cryptosporidium parvum-Infected Neonatal Mice Show Gut Microbiota Remodelling Using High-Throughput Sequencing Analysis: Preliminary Results.

During the last decade, the scientific community has begun to investigate the composition and role of gut microbiota in normal health and disease. These studies have provided crucial information on the relationship between gut microflora composition and intestinal parasitic infection, and have demonstrated that many enteric pathogen infections are associated with altered gut microflora composition. In this study, we investigated the effects of Cryptosporidium parvum infection (zoonotic protozoan affecting a large range of vertebrates) on both qualitative and quantitative composition of gut microbiota in a CD-1 neonatal mouse model. 5-day-old neonate mice were experimentally infected with 105Cryptosporidium parvum Iowa oocysts by oesophageal gavage. The intestinal microbiota of both infected (Cp+) and uninfected (Cp-) mice groups was examined by high-throughput sequencing of the bacterial 16S rDNA gene V3-V4 hypervariable region. The most consistent change in the microbiota composition of Cp+ mice was the increased proportion of bacterial communities belonging to the Phylum Bacteroidetes. In contrast, the microbiota of Cp- mice was associated with increased proportions of several Firmicutes and Actinobacteria phyla members. For the first time, our study provides evidence of an association between cryptosporidial infection and gut dysbiosis, thus contributing valuable knowledge to the as-yet little-explored field of Cryptosporidium-microbiota interactions in a neonatal mouse model.

Induction of Inflammatory Responses in Splenocytes by Exosomes Released from Intestinal Epithelial Cells following Cryptosporidium parvum Infection.

Cryptosporidium, a protozoan parasite that infects the gastrointestinal epithelium and other mucosal surfaces in humans and animals, is an important opportunistic pathogen in AIDS patients and one of the most common enteric pathogens affecting young children in developing regions. This parasite is referred to as a "minimally invasive" mucosal pathogen, and epithelial cells play a central role in activating and orchestrating host immune responses. We previously demonstrated that Cryptosporidium parvum infection stimulates host epithelial cells to release exosomes, and these released exosomes shuttle several antimicrobial peptides to carry out anti-C. parvum activity. In this study, we detected the upregulation of inflammatory genes in the liver and spleen following C. parvum intestinal infection in neonatal mice. Interestingly, exosomes released from intestinal epithelial cells following C. parvum infection could activate the nuclear factor kappa B signaling pathway and trigger inflammatory gene transcription in isolated primary splenocytes. Several epithelial cell-derived proteins and a subset of parasite RNAs were detected in the exosomes released from C. parvum-infected intestinal epithelial cells. Shuttling of these effector molecules, including the high mobility group box 1 protein, was involved in the induction of inflammatory responses in splenocytes induced by the exosomes released from infected cells. Our data indicate that exosomes released from intestinal epithelial cells upon C. parvum infection can activate immune cells by shuttling various effector molecules, a process that may be relevant to host systemic responses to Cryptosporidium infection.

A protocol to count Cryptosporidium oocysts by flow cytometry without antibody staining.

Cryptosporidiosis caused by the protozoan parasites Cryptosporidium hominis and C. parvum, threatens the lives of young children in developing countries. In veterinary medicine, C. parvum causes life-threatening diarrhea and dehydration in newborn dairy calves. Protocols to detect Cryptosporidium spp. oocysts using flow cytometry have been reported; however, these protocols use antibodies against the parasite and typically focus on detection of oocysts, not quantification. These techniques are not well-suited for studies that generate large variations in oocyst burdens because the amount of antibody required is proportional to the number of oocysts expected in samples. Also, oocysts are lost in washes in the staining protocol, reducing accuracy of oocyst counts. Moreover, these protocols require costly fluorochrome-conjugated monoclonal antibodies and are not optimal for studies involving large numbers of samples. Here we present an optimized protocol for purifying oocysts from mouse stool and intestine samples combined with a reliable method to quantify oocysts in a relatively pure population without the need for antibody staining. We used morphology (SSC-A vs FSC-A) and the innate characteristics of C. parvum oocysts compared to fecal and intestinal contaminants to develop a two-step gating strategy that can differentiate oocysts from debris. This method is a fast, reliable, and high-throughput technique to promote research projects on C. parvum infections in mice and potentially other animal hosts.

High frequency and molecular characterization of porcine hemotrophic mycoplasmas in Brazil

Mycoplasma suis and Mycoplasma parvum are the two hemotrophic mycoplasmas species described in pigs. M. suis is involved in infectious anemia, while M parvum infection is commonly subclinical. The objectives of this study were twofold: (i) to investigate the prevalence of porcine hemotrophic mycoplasmas in sows from the southern region of Brazil by quantitative real-time PCR (qPCR) and (ii) to genetically characterize a subset of the samples based on the 16S rRNA gene. A total of 429 blood samples were evaluated from 53 different farm sites. Porcine hemoplasmas was detected at all the 53 tested sites and in 79.72% of the samples (342/429). Two sequences were obtained for Mycoplasma spp. The phylogenetic analysis based on the 16S rRNA gene (900 bp) showed that the Mycoplasma sequences were closely related to the M. suis cluster and that one sequence was positioned in the M. parvum cluster. In conclusion, porcine hemoplasmas have a high rate of prevalence in sows from commercial farms in the southern region of Brazil. This study demonstrated the first molecular detection and characterization of partial 16S rRNA gene of M. parvum in Brazil.

Assessment of selenium supplementation by systemic injection at birth on pre-weaning calf health

The objective of this randomized clinical trial was to evaluate the effect of selenium and vitamin E supplementation by systemic injection on dairy calf health and growth during the pre-weaning period. The study was conducted at 39 dairy farms in Ontario, selected through a convenience sample of farms in close proximity to either Guelph or Kemptville. A total of 835 Holstein heifer calves were enrolled in this study. At birth, calves were randomly allocated to receive selenium and vitamin E supplementation by injection (3 mg sodium selenite and 136 IU dl-a-tocopherol acetate) or a placebo solution. At enrollment, producers recorded time of birth, calving ease, and colostrum feeding status. At weekly visits to the individual dairy farms, trained technicians collected measurements and samples from enrolled calves. Blood was collected from calves that were up to 8 d of age to assess the concentration of serum total protein and selenium. Between 8 and 15 d of age, fecal samples were collected to identify the presence of rotavirus and Cryptosporidium parvum (C. parvum) using a commercially +available lateral immuno-chromatography antigen detection kit. Each enrolled calf was also weighed and assessed for health scores during the first, second, and seventh weeks of life. The average serum selenium concentration (SD) in treated calves was 0.08 μg/mL (0.02), versus 0.06 μg/mL (0.01) in control calves. The incidence of failure of passive transfer (FPT) among study calves was 21%, and did not differ between treatment groups. The mean average daily gain (ADG) for the study period was 1.43 lb (0.65 kg)/day and was not associated with selenium and vitamin E supplement injection. Of the 761 fecal samples tested, 272 (36%) and 118 (16%) tested positive for C. parvum and rotavirus antigen, respectively. Selenium and vitamin E treatment had a protective effect against rotavirus infection. However, there was no effect of experimental treatment on C. parvum infection status. Reduced odds of treatment for diarrhea was also seen in the selenium and vitamin E treatment group. This study suggests that selenium and vitamin E injection at birth could improve pre-weaning health by reducing rotavirus infection and diarrhea.

MicroRNA expression profile of HCT-8 cells in the early phase of Cryptosporidium parvum infection

BackgroundCryptosporidium parvum is an important zoonotic parasitic disease worldwide, but the molecular mechanisms of the host–parasite interaction are not fully understood. Noncoding microRNAs (miRNAs) are considered key regulators of parasitic diseases. Therefore, we used microarray, qPCR, and bioinformatic analyses to investigate the intestinal epithelial miRNA expression profile after Cryptosporidium parvum infection.ResultsTwenty miRNAs were differentially expressed after infection (four upregulated and 16 downregulated). Further analysis of the differentially expressed miRNAs revealed that many important cellular responses were triggered by Cryptosporidium parvum infection, including cell apoptosis and the inflammatory and immune responses.ConclusionsThis study demonstrates for the first time that the miRNA expression profile of human intestinal epithelium cells is altered by C. parvum infection. This dysregulation of miRNA expression may contribute to the regulation of host biological processes in response to C. parvum infection, including cell apoptosis and the immune responses. These results provide new insight into the regulatory mechanisms of host miRNAs during cryptosporidiosis, which may offer potential targets for future C. parvum control strategies.

Interferon-λ3 Promotes Epithelial Defense and Barrier Function Against Cryptosporidium parvum Infection.

Background & AimsThe epithelial response is critical for intestinal defense against Cryptosporidium, but is poorly understood. To uncover the host strategy for defense against Cryptosporidium, we examined the transcriptional response of intestinal epithelial cells (IECs) to C parvum in experimentally infected piglets by microarray. Up-regulated genes were dominated by targets of interferon (IFN) and IFN-λ3 was up-regulated significantly in infected piglet mucosa. Although IFN-λ has been described as a mediator of epithelial defense against viral pathogens, there is limited knowledge of any role against nonviral pathogens. Accordingly, the aim of the study was to determine the significance of IFN-λ3 to epithelial defense and barrier function during C parvum infection.MethodsThe significance of C parvum–induced IFN-λ3 expression was determined using an immunoneutralization approach in neonatal C57BL/6 mice. The ability of the intestinal epithelium to up-regulate IFN-λ2/3 expression in response to C parvum infection and the influence of IFN-λ2/3 on epithelial defense against C parvum invasion, intracellular development, and loss of barrier function was examined using polarized monolayers of a nontransformed porcine-derived small intestinal epithelial cell line (IPEC-J2). Specifically, changes in barrier function were quantified by measurement of transepithelial electrical resistance and transepithelial flux studies.ResultsImmunoneutralization of IFN-λ2/3 in C parvum–infected neonatal mice resulted in a significantly increased parasite burden, fecal shedding, and villus blunting with crypt hyperplasia during peak infection. In vitro, C parvum was sufficient to induce autonomous IFN-λ3 and interferon-stimulated gene 15 expression by IECs. Priming of IECs with recombinant human IFN-λ3 promoted cellular defense against C parvum infection and abrogated C parvum–induced loss of barrier function by decreasing paracellular permeability to sodium.ConclusionsThese studies identify IFN-λ3 as a key epithelial defense mechanism against C parvum infection.

Induction of a Long Noncoding RNA Transcript, NR_045064, Promotes Defense Gene Transcription and Facilitates Intestinal Epithelial Cell Responses against Cryptosporidium Infection.

Cryptosporidium is an important opportunistic intestinal pathogen for immunocompromised individuals and a common cause of diarrhea in young children in developing countries. Gastrointestinal epithelial cells play a central role in activating and orchestrating host immune responses against Cryptosporidium infection, but underlying molecular mechanisms are not fully understood. We report in this paper that C. parvum infection causes significant alterations in long noncoding RNA (lncRNA) expression profiles in murine intestinal epithelial cells. Transcription of a panel of lncRNA genes, including NR_045064, in infected cells is controlled by the NF-κB signaling. Functionally, inhibition of NR_045064 induction increases parasite burden in intestinal epithelial cells. Induction of NR_045064 enhances the transcription of selected defense genes in host cells following C. parvum infection. Epigenetic histone modifications are involved in NR_045064-mediated transcription of associated defense genes in infected host cells. Moreover, the p300/MLL-associated chromatin remodeling is involved in NR_045064-mediated transcription of associated defense genes in intestinal epithelial cells following C. parvum infection. Expression of NR_045064 and associated genes is also identified in intestinal epithelium in C57BL/6J mice following phosphorothioate oligodeoxynucleotide or LPS stimulation. Our data demonstrate that lncRNAs, such as NR_045064, play a role in regulating epithelial defense against microbial infection.

A novel detection method of Cryptosporidium parvum infection in cattle based on Cryptosporidium parvum virus 1.

Cryptosporidium parvum is an important zoonotic parasite that causes significant economic loss in the animal husbandry industry, especially the cattle industry. As there is no specific vaccine or drug against Cryptosporidium, a rapid and accurate method for the detection of C. parvum is of great significance. In this study, colloidal gold strips were developed based on Cryptosporidium parvum virus 1 (CSpV1) for the detection of C. parvum infection in cattle fecal samples. The colloidal gold solution was prepared by reducing trisodium citrate and the CSpV1 #5 monoclonal antibody was labeled with colloidal gold. A polyclonal antibody against the CSpV1 capsid protein and an anti-mouse IgG antibody were coated on the colloidal gold strips for use in the test and control lines, respectively. Our results showed that the detection sensitivity in fecal samples was up to a 1:64 dilution. There was no cross-reaction with Cryptosporidium andersoni or Giardia in the fecal samples. The different preservation conditions (room temperature, 4°C, and 37°C) and preservation time (7, 30, 60, and 90 days) were analyzed. The data showed that the strips could be preserved for 90 days at 4°C and for 60 days at room temperature or 37°C. The colloidal gold strips were used to detect the samples of 120 clinical fecal in Changchun, China. The results indicated that the rate of a positive test was 5% (6/120). This study provides a rapid and accurate method for detecting C. parvum infection in cattle and humans.

Disinfection with hydrated lime may help manage cryptosporidiosis in calves

Diarrhea is common in young calves and is often caused by Cryptosporidium parvum infection. The aim of this study was to investigate if disinfection of calf pens with hydrated lime would reduce contamination of C. parvum oocysts and improve calf health in herds with C. parvum associated diarrhea problems. Four dairy herds with ongoing C. parvum associated calf diarrhea problems each participated in the study over six to seven months. During the study period, all pens/huts for young calves were cleaned according to the usual farm routine before a new calf entered. Hydrated lime was then used to disinfect half of the pens/huts. Diarrhea incidence was recorded by the farmers and by veterinarians, who clinically examined the calves every second month. In total, 402 calves participated in the study. The farmers detected diarrhea in 214 (53%) calves, with similar proportions in calves kept in lime disinfected and control pens. Age at diarrhea onset was significantly higher in lime disinfected pens than in control pens, 9.0 days and 7.6 days, respectively. There was no difference between the groups regarding duration or severity index recorded by the farmers. The body condition score in 6–8 week old calves was significantly higher in calves that had been kept in lime disinfected pens during their first weeks of life, indicating that calves in disinfected pens/huts were less affected by their infections.Faecal samples from 5 to 21 day old calves, were collected on four occasions at each farm (n = 95). Cryptosporidium positive samples were found at all samplings in all four herds. Cryptosporidium spp. was detected in 79 (83%) samples with no difference between lime disinfected and control pens. C. parvum was the dominant species. Two different C. parvum subtypes were found; IIaA16G1R1b in three herds and IIaA16G1R1b_variant in one herd. Only one subtype was found in each herd.Disinfection of calf pens with slaked lime delayed onset of diarrhea and improved the body condition in the calves, but did not affect diarrhea incidence or duration. Although lime disinfection alone will not be sufficient to control Cryptosporidium associated diarrhea in herds with extensive calf diarrhea problems, these results suggest that it can be a valuable complement to other measures.

Development of CpGP15 recombinant antigen of Cryptosporidium parvum for detection of the specific antibodies in cattle

The infection of neonatal calves with Cryptosporidium parvum can have a huge economic impact because diarrhea caused by the parasite sometimes results in death. A serodiagnostic system will be helpful in the diagnosis of C. parvum infection. CpP23 is commonly used as an antigen for enzyme-linked immunosorbent assay (ELISA); however, some positive sera show low reactivities, as shown in this study. Herein, we focused on three other antigens, CpGP15, CpP2 and CpGP60, in addition to CpP23, to detect C. parvum-specific antibodies in cattle sera. CpP23 and CpGP15 showed substantial ability to discriminate between positive (n = 10) and negative (n = 10) control cattle sera. Unlike our previous report, both the sensitivity and the specificity were 100% when the two antigens were employed for the ELISA. The newly developed ELISA was applied to a total of 344 sera obtained from 9 cattle farms. Two farms among them had suffered from C. parvum infections before, and were regarded as the C. parvum-positive farms. The positive rates of antibodies against CpP23 and CpGP15 in the C. parvum-positive farms were 42.7% and 49.8%, respectively, whereas the positive rate for either of the antigens was 63.0% in the farms. In contrast, 14.3% and 9.8% were positive for CpP23 and CpGP15 in the C. parvum-negative farms, respectively, whereas 18.8% was positive for either of the antigens. This study revealed that the ELISAs employing both of CpP23 and CpGP15 can avoid false-negative results and are useful for monitoring of the C. parvum infection in cattle farms.

Curcumin: A promising treatment for Cryptosporidium parvum infection in immunosuppressed BALB/c mice

Members of the genus Cryptosporidium are frequent protozoan pathogens in humans and a wide range of animals. There is no consistently effective treatment against cryptosporidiosis, especially in immunodeficient patients. The present study was carried out to study the therapeutic effects of curcumin against cryptosporidiosis in immunosuppressed BALB/c mice. Mice were divided into five groups and immunosuppressed by dexamethasone. Three groups were inoculated with C. parvum oocysts, administered with curcumin, paromomycin, and without treatment. The reminders were regarded as controls. The oocysts in the fecal smear were counted daily. At days 0, 3, 7, and 11 post-treatment, the mice were sacrificed, and the efficacy of drugs was evaluated by comparing the histopathological alterations in jejunum and ileum, measuring the total antioxidant capacity, and malondialdehyde in the affected tissues. The infection was completely eliminated in the curcumin-treated group, and oocyst shedding stopped with no recurrence after drug withdrawal. On the contrary, paromomycin was unable to eliminate C. parvum infection completely, and oocyst shedding continued even 10 days after the drug withdrawal. Based on these findings, curcumin can be a trustworthy compound for the elimination of infection in immunosuppressed hosts. Further evaluation to find its accurate mechanism of action should be considered.

Oxidative stress and clinico-pathological alterations induced by Cryptosporidium parvum infection in a rat model

Cryptosporidium parvum is an apicomplexan zoonotic enteric protozoa that is principally responsible for severe necrotizing enteritis in immunocompromised hosts and has been associated with diarrhoea in a wide variety of animal species. The work was carried out on 48 rats which were equally divided into four groups: GI, GII, GIII and GIV. Rats in GI worked as healthy control. Rats in GII were administered C. parvum oocysts orally, GIII received dexamethasone, whereas GIV were given dexamethasone along with C. parvum oocysts. The maximum severity of experimental infection and oocyst shedding was observed on 8th DPI in Group IV rats who exhibited depression, inappetance, poor fur condition, decreased weight gain, bloody diarrhoea and highest mortality rate. Significant increase in LPO and decrease in SOD, CAT and GSH concentrations were seen in intestine in GIV rats at 8th DPI indicating the involvement of free radical induced oxidative stress in the pathogenesis of C. parvum infection. Severe necrotizing and haemorrhagic enteritis was seen in Group IV rats, whereas Group II animals had mild enteritis. Pathological examination of intestines in Group IV animals revealed extensive necrosis, blunting, fusion and sloughing of intestinal villi. Severe lymphocytic infiltration in lamina propria and submucosa was also observed. The diarrhoea and oocyst shedding gradually subsided after 8th DPI and completely resolved in GII whereas mild enteritis was still seen in GIV animals at 12thDPI. It was inferred from the present study that C. parvum strains prevalent in Jammu were capable of causing severe enteric infection in immunosuppressed rats.