Cryptosporidium parvum causes life-threatening diarrhoea in immunocompromized, especially AIDS patients and the efficiency of proposed anti-cryptosporidial therapies is limited or doubtful. An immunosuppressed adult rat model of C. parvum infection was developed for screening molecules candidate for curative and preventive activity in human cryptosporidiosis. Among 31 drugs tested, lasalocid (2-10 mg/kg/24 h), and sinefungin (2-10 mg/kg/24 h), exhibited some activity against C. parvum infection. Oral sinefungin therapy resulted in a dose related suppression of oocysts shedding, which correlated with oocyst disappearance from ileum sections and was also efficient in preventing infection. Relapses were observed after discontinuation of curative sinefungin therapy, which suggests that the biliary tract, a major location and parasite reservoir which sustains persisting infection, was not cleared of parasites by the drug. Improved therapeutic procedures with sinefungin (or analogues) will result from current pharmacological studies.
An ICR outbred suckling mouse model of cryptosporidiosis was used to explain some of the variability associated with experimental Cryptosporidium parvum infections in neonate mice. Fourty four groups of 12 mice each, ranging in age from 4-12 days, each received 1.0 x 104 CsCl purified oocysts per os in 5 pm PBS. At 6 days post-inoculation (PI), mice were killed by C02 overdose and individually weighed. Intestines were then homogenized and oocysts were quantified by hemacytometer. Results revealed that both age and weight have pronounced effects on numbers of oocysts produced in vivo, with larger and older mice producing higher numbers of parasites. Mice 8-9 days of age at the time of inoculation displayed the least amount of weight dependent variability, produced the highest numbers of oocysts, and were judged to be superior over other ages for pharmaceutical screening. Significant reductions in numbers of oocysts occurred in mice inoculated at 10 days of age, and only a few oocysts were found in mice inoculated at 11-12 days of age. These studies suggest that at least some data on Cryptosporidium generated from suckling mouse studies to date are probably unreliable and should be viewed skeptically.
Cryptosporidium parvum, Tyzzer, 1912 is identified as a common cause of diarrhoea in immunocompetent individuals. In immunocompromised, especially HIV-infected subjects, cryptosporidiosis causes severe chronic diarrhoea. In this study, nitazoxanide (NTZ) was compared for curative activity with sinefungin (SNF) and paromomycin (PRM) in immunosuppressed rats, a screening model for anticryptosporidial agents. NTZ at either 50 mg/kg/day, 100 mg/kg/day or 200 mg/kg/day resulted in seven days in a dose-dependent inhibition of oocyst shedding similar to that obtained with SNF (10 mg/kg/day) and PRM (100 mg/kg/day). Further discontinuation of SNF or PRM 100 mg/kg/day therapy resulted in early relapse of oocyst shedding which reached the pre-treatment levels in 2-4 days. In contrast, seven days after discontinuation of therapy, shedding inhibition was unchanged in NTZ-treated rats. Data prompt further assessment of the activity of NTZ on sequestered C. parvum.
To investigate the transmission of species of Cryptosporidium Tyzzer, 1907 in Timis County, Romania, 48 isolates of Cryptosporidium coccidia from 11 children, 29 calves and eight pigs were characterised by molecular analysis of two loci (SSU rRNA and 60-kDa glycoprotein gene). Overall, 22 isolates were amplified and sequence analyses revealed that all isolates were Cryptosporidium parvum Tyzzer, 1912. Two subtype families were identified, IIa and IId. Subtype IIdA22G1 (n = 4) was the single C. parvum subtype found in children. Subtypes found in calves included IIdA27G1 (n = 8), a novel subtype, IIdA25G1 (n = 5), IIdA22G1 (n = 2), IIdA21G1a (n = 1), and IIaA16G1R1 (n = 1). Subtype IIdA26G1 was found in a pig. These results were significantly different from previous Romanian reports, as the five subtypes of family IId identified in this study were never identified previously in this country. Thus, cattle may be a source of Cryptosporidium infections for humans and the transmission dynamics of C. parvum in Romania is more complex than previously believed., Patrícia Manuela Vieira, Narcisa Mederle, Maria Luísa Lobo, Kálmán Imre, Ovidiu Mederle, Lihua Xiao, Gheorghe Darabus, Olga Matos., and Obsahuje bibliografii
Cryptosporidium parvum, the protozoan responsible for cryptosporidiosis, continues to defy eradication with existing therapies. A review of the anticryptosporidial activity of several drugs in the dexamethasone-immunosuppressed rat model illustrates the multitude of factors that may contribute to the difficulty of assessing a drug’s therapeutic efficacy against the protozoan and provides possible explanation for drug failure at the level of host-parasite interaction.
The objective of this study was to determine if primers, probes, and pHCl, a plasmid containing a 2.3 kilobasc insert of genomic DNA from Cryptosporidium parvum, would be useful for the detection of Cryptosporidium wrairi DNA using the polymerase chain reaction. C. wrairi DNA was prepared from oocysts recovered from guinea pigs and C. parvum DNA was prepared from the Iowa strain of C. parvum. Two 26-(bp) primers were used to amplify a 452-base pair bp target sequence within the cloned DNA. Similarly-sized PCR products were obtained with the pHCl plasmid DNA, C. parvum DNA, and C. wrairi DNA. However, a 20-base pair probe did not detect C. wrairi DNA. Sequencing of C. wrairi DNA homologous with the 452-bp segment of C. parvum DNA showed 18 bp changes including bp changes in the segment homologous with the probe. A new probe based on homologous sequences was useful for detection of both species of Cryptosporidia. The sequences of the homologous 452-bp segment from the Iowa strain of C. parvum and that segment of C. parvum DNA from the pHC I plasmid were very similar. Nine base pairs identical in the homologous bp segment of the Iowa strain of C. parvum, C. wrairi and the pHCl plasmid differed from those previously reported. Previously reported primers and a newly designed probe proved to be useful for detecting C. wrairi DNA.
A technique based on the analysis of banding patterns obtained by SDS-PAGE Western-blotting of an oocyst wall antigen obtained from faeces has been evaluated to subtype Cryptosporidium parvum Tyzzer, 1912. This technique appears to have sufficient stability to recognise multiple types of this parasite. A similar Western-blotting technique has also been used to assess antibody responses to cryptosporidial antigens in human sera. Two systems were developed: one against three antigens of apparent molecular weights 6, 14 and 17 kDa; the second against oocyst wall antigens of apparent molecular weights 57, 69, 75, 89, 128, 151 and 173 kDa. Antibodies to three antigens of apparent molecular weights 6, 14 and 17 kDa were most successful as diagnostic markers in that they were found in >88% of convalescent phase sera from confirmed cryptosporidiosis patients and were uncommon (>7%) in control subjects. Faecal samples from human and animal sporadic cases yielded a wide range of cryptosporidial antigen banding patterns. Samples from patients in a water-borne outbreak in South Devon (England) in 1995 also yielded a wide range of banding patterns including members within individual household family groups. These results are in contrast with those from samples collected from other defined geographical areas, including some from a second water-borne outbreak where much more homogeneous banding patterns were obtained. Sera collected for other purposes from apparently uninfected individuals 9 months after the South Devon 1995 outbreak were examined. Antibodies to the three antigens of molecular weights 6, 14 and 17 kDa were detected in 32-49% of individuals resident in the outbreak water supply area, and in 15-21%) of those resident in an adjacent water supply area. The significance of these findings is discussed in relation to data obtained from epidemiological field studies.
Water-borne transmission of the coccidium Cryptosporidium parvum Tyzzer, 1912 is frequently responsible for outbreaks of human cryptosporidiosis. One of the most important was reported in 1993 in Milwaukee in the United States, where 403,000 cases were recorded. The determination of the percentage of oocysts excystated is the first step in evaluating their viability, but it alone is not sufficient. This percentage depended on the conditions of storage and also the presence of oxidant or disinfectent agents in water. The percentage of excystation is not always related to viability. Therefore, determination of the viability of excysted sporozoites by determining their infectivity for enterocytic Caco2 cell lines in culture provides information essential for evaluating the risk of contaminated drinking water.