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R Fayer

Publications and source records attributed to R Fayer.

At least 37 records · Page 2Linked to original sources
PubMed2001

Cryptosporidium canis n. sp. from domestic dogs.

Oocysts of Cryptosporidium, from the feces of a naturally infected dog and from an HIV-infected human, were identified as the previously reported canine genotype of Cryptosporidium parvum, hereafter referred to as Cryptosporidium canis n. sp. Also among the oocysts from the dog, a trace amount of C. parvum bovine genotype was detected. Cryptosporidium canis oocysts from both the dog and human were infectious for calves. Oocysts excreted by calf 1 (dog source) were approximately 90% C. canis and 10% C. parvum, whereas those excreted by calf 3 (human source) were 100% C. canis. Oocysts from calf 1 infected calf 2 resulting in excretion by calf 2 of oocysts approximately 90% C. parvum and 10% C. canis. Oocysts of C. canis were not infectious for BALB/c neonatal mice or immunosuppressed C57 juvenile mice, although all control mice became infected with the C. parvum Beltsville isolate. Oocysts of C. canis from calf 1 and the human were structurally indistinguishable from oocysts of the C. parvum Beltsville isolate (bovine). However, C. canis oocysts differed markedly at the molecular level from all known species of Cryptosporidium based on sequence data for the 18S rDNA and the HSP 70 gene. The differences in genetics and host specificity clearly differentiate C. canis as a new species.

Animals↗
PubMed2000

Estimating viability of Cryptosporidium parvum oocysts using reverse transcriptase-polymerase chain reaction (RT-PCR) directed at mRNA encoding amyloglucosidase.

The purpose of the present study was to determine if reverse transcriptase-polymerase chain reaction (RT-PCR) directed at mRNA encoding the enzyme amyloglucosidase (CPAG) could serve as a indicator for C. parvum oocyst viability. Oocysts were stored for 1-11 months in the refrigerator and at monthly intervals extracted for total RNA for RT-PCR analysis. An aliquot of these C. parvum oocysts was inoculated into neonatal mice which were necropsied 4 days later for ileal tissue that was analyzed by semi-quantitative PCR to determine the level of parasite replication. The CPAG RT-PCR assay detected RNA from as few as 10(3) C. parvum oocysts. An effect of storage time on both RT-PCR signal and mouse infectivity was observed. RNA from oocysts stored for 1-7 months, unlike oocysts stored for 9 or 11 months, contained CPAG mRNA that was detectable by RT-PCR. A gradual decrease in the RT-PCR signal intensity was observed between 5 and 7 months storage. The intensity of RT-PCR product from oocysts and the signal from semi-quantitative PCR of ileal tissue DNA from mice infected with these same aged oocysts were comparable. The RT-PCR assay of CPAG mRNA in cultured cells infected with viable C. parvum oocysts first detected expression at 12 h with highest expression levels observed at 48 h post-infection. These results indicate that CPAG RT-PCR may be useful for differentiating viable from non-viable C. parvum oocysts and for studying the expression of the gene for amyloglucosidase in vitro.

Animals↗
PubMed2000

Prevalence of Cryptosporidium, Giardia and Eimeria infections in post-weaned and adult cattle on three Maryland farms.

The prevalence of Cryptosporidium, Giardia and Eimeria, in healthy, asymptomatic, post-weaned and mature cattle was investigated on three Maryland farms. One farm, a dairy research facility, had 150 multiparous Holstein milking cows; 24 were examined and Cryptosporidium andersoni was detected in three (12.5%) but neither Giardia nor Eimeria was detected. The second farm, a commercial dairy, had 57 multiparous Holstein milking cows and an equal number of heifers. Of 19 cows examined, C. parvum, Giardia duodenalis, and Eimeria bovis and/or E. ellipsoidalis were detected in two (10.5%), two (10.5%) and one (5.26%) cow, respectively. Of 23 heifers examined, C. parvum, Giardia, and E. bovis and E. ellipsoidalis, was detected in two (8.7%), four (17.4%), and five (21.7%), heifers, respectively. The third farm, a beef cattle breeding and genetics research facility, had 180 7- to 9-month old purebred black Angus. Of 118 examined for C. parvum and Giardia, 34 (28.8%) and 44 (37.3%) were positive, respectively, of 97 examined for E. bovis and/or E. ellipsoidalis 32 (33.0%) were positive. These findings, based on a method with a minimum detection level of 100 oocysts of C. parvum/g of feces, which underestimates the number of infected cattle, clearly demonstrate the presence of low level, asymptomatic infections in post-weaned and adult cattle in the United States and indicate the potential role of such cattle as reservoirs of infectious parasites.

Animals↗
PubMed2000

Epidemiology of Cryptosporidium: transmission, detection and identification.

There are 10 valid species of Cryptosporidium and perhaps other cryptic species hidden under the umbrella of Cryptosporidium parvum. The oocyst stage is of primary importance for the dispersal, survival, and infectivity of the parasite and is of major importance for detection and identification. Because most oocysts measure 4-6 microm, appear nearly spherical, and have obscure internal structures, there are few or no morphometric features to differentiate species and in vitro cultivation does not provide differential data as for bacteria. Consequently, we rely on a combination of data from three tools: morphometrics, molecular techniques, and host specificity. Of 152 species of mammals reported to be infected with C. parvum or an indistinguishable organism, very few oocysts have ever been examined using more than one of these tools. This paper reviews the valid species of Cryptosporidium, their hosts and morphometrics; the reported hosts for the human pathogen, C. parvum; the mechanisms of transmission; the drinking water, recreational water, and food-borne outbreaks resulting from infection with C. parvum; and the microscopic, immunological, and molecular methods used to detect and identify species and genotypes.

Animals↗
PubMed2000

Cryptosporidium systematics and implications for public health.

There is controversy in the taxonomy of Cryptosporidium parasites and the public health significance of Cryptosporidium isolates from various animals. Recent advances in molecular characterization of Cryptosporidium parasites have allowed the re-examination of species structure of the genus Cryptosporidium. Non-parvum Cryptosporidium spp and new C. parvum genotypes in immunocompromised humans can now be clearly detected. In this article, Lihua Xiao and colleagues summarize the current biological and molecular evidence for different Cryptosporidium spp, and the public health importance of these species and new C. parvum genotypes.

Animals↗
PubMed2000

Cryptosporidium meleagridis in an Indian ring-necked parrot (Psittacula krameri).

OBJECTIVE: To perform a morphological and genetic characterisation of a Cryptosporidium infection in an Indian ring-necked parrot (Psittacula krameri) and to compare this with C meleagridis from a turkey. DESIGN: Tissue and intestinal sections from an Indian ring-necked parrot were examined microscopically for Cryptosporidium. The organism was also purified from the crop and intestine, the DNA extracted and a portion of the 18S rDNA gene amplified, sequenced and compared with sequence and biological information obtained for C meleagridis from a turkey as well as sequence information for other species of Cryptosporidium. RESULTS: Morphological examination of tissue sections from an Indian ring-necked parrot revealed large numbers of Cryptosporidium oocysts attached to the apical border of enterocytes lining the intestinal tract. Purified Cryptosporidium oocysts measured about 5.1 x 4.5 microns, which conformed morphologically to C meleagridis. The sequence obtained from this isolate was identical to sequence information obtained from a C meleagridis isolate from a turkey. CONCLUSION: Cryptosporidium meleagridis was detected in an Indian ring-necked parrot using morphological and molecular methods. This is the first time that this species of Cryptosporidium has been reported in a non-galliform host and extends the known host range of C meleagridis.

Animals↗
PubMed2000

Cryptosporidium spp. in domestic dogs: the "dog" genotype.

Genetic and phylogenetic characterization of Cryptosporidium isolates at two loci (18S rRNA gene and heat shock gene) from both Australian and United States dogs demonstrated that dog-derived Cryptosporidium isolates had a distinct genotype which is conserved across geographic areas. Phylogenetic analysis provided support for the idea that the "dog" genotype is, in fact, a valid species.

Animals↗
PubMed2000

Transforming growth factor beta1 ameliorates intestinal epithelial barrier disruption by Cryptosporidium parvum in vitro in the absence of mucosal T lymphocytes.

Exposure to oocysts of the protozoan Cryptosporidium parvum causes intestinal epithelial cell dysfunction in vivo and in vitro, but effective means by which mucosal injury might be prevented remain unclear. We examined the ability of transforming growth factor beta1 (TGF-beta1)-a cytokine synthesized and released by cells in the intestine-to preserve the barrier function of human colonic epithelia when challenged with C. parvum oocysts and then studied the mechanisms involved. Epithelial barrier function was monitored electrophysiologically, receptors for TGF-beta1 were localized by confocal microscopy, and TGF-beta1-induced protein kinase C activation was detected intracellularly by translocation of its alpha isozyme. TGF-beta1 alone enhanced intestinal epithelial barrier function, while exposure to C. parvum oocysts (> or =10(5)/monolayer) markedly reduced barrier function to < or =40% of that of the control. When epithelial monolayers were pretreated with TGF-beta1 at 5.0 ng/ml, the barrier-disrupting effect of C. parvum oocysts was almost completely abrogated for 96 h. Further investigation showed that (i) the RI and RII receptors for TGF-beta1 were present on 55 and 65% of human epithelial cell line cells, respectively, over a 1-log-unit range of receptor protein expression, as shown by flow cytometry and confirmed by confocal microscopy; (ii) only basolateral and not apical TGF-beta1 exposure of the polarized epithelial monolayer resulted in a protective effect; and (iii) TGF-beta1 had no direct effect on the organism in reducing its tissue-disruptive effects. In exploring mechanisms to account for the barrier-preserving effects of TGF-beta1 on epithelium, we found that the protein kinase C pathway was activated, as shown by translocation of its 80-kDa alpha isozyme within 30 s of epithelial exposure to TGF-beta1; the permeability of epithelial monolayers to passage of macromolecules was reduced by 42% with TGF-beta1, even in the face of active protozoal infection; and epithelial cell necrosis monitored by lactate dehydrogenase release was decreased by 50% 70 h after oocyst exposure. Changes in epithelial function, initiated through an established set of surface receptors, likely accounts for the remarkable barrier-sparing effect of nanogram-per-milliliter concentrations of TGF-beta1 when human colonic epithelium is exposed to an important human pathogen, C. parvum.

Animals↗
PubMed1999

Hyperimmune bovine colostrum specific for recombinant Cryptosporidium parvum antigen confers partial protection against cryptosporidiosis in immunosuppressed adult mice.

Preparturient cows were immunized three times over a six-week period with recombinant plasmid DNA encoding the Cryptosporidium parvum CP15/60 antigen by injecting the DNA in the mammary gland. Serum was collected at each immunization and first colostrum was collected after parturition; all were assayed for Cryptosporidium-specific antibodies (Ab). A serological response to C. parvum sporozoite and oocyst antigen was detected in cows immunized with pCP15/60 plasmid DNA. Colostrum from these cows, unlike colostrum from normal controls, contained Ab specific for C. parvum sporozoites and oocysts as indicated by immunofluorescence Ab (IFA) staining. Colostrum was also tested for conferring passive immunity against C. parvum infection by oral administration to immunosuppressed adult inbred mice. Immune colostrum and control colostrum were administered to separate groups of dexamethasone (DEX)-treated adult C57BL/6NCr mice beginning 12 h before and at 12 h intervals for 3 days after oral C. parvum oocyst infection. Cryptosporidium development was assayed in ilea of immune- and control-colostrum-treated mice 96 h postinfection by semiquantitative PCR. Mice receiving immune colostrum showed partial protection (about 50% reduction) against intestinal C. parvum development compared to mice receiving control colostrum. This protection was evident at a challenge dose of 10(3) C. parvum oocysts per mouse; no differences were noted in parasite development between groups receiving immune or control colostrum and infected with 10(4) oocysts. This study showed that serum and colostrum Ab response to C. parvum can be elicited in preparturient cows by direct injection of recombinant pCP15/60 plasmid DNA and that passive protection against cryptosporidiosis can be obtained by treating immunosuppressed mice with immune colostrum before and after C. parvum infection.

Animals↗
PubMed1999

Evaluation of the recovery of waterborne Giardia cysts by freshwater clams and cyst detection in clam tissue.

The Asian freshwater clam, Corbicula fluminea, inhabits environments recognized to be contaminated with waterborne Giardia cysts. Sixty-four tissue samples of Giardia-free clams were spiked with various numbers of Giardia duodenalis cysts within the range of 50-700 cysts. Regression analysis showed that paired numbers of spiked (x) versus recovered (y) cysts regressed significantly (P < 0.01) according to the equation y = 42.57 +/- 1.81x (+/- 64.3). The cyst detection threshold was 43 cysts/clam, the coefficient of determination was 77%, and the overall sensitivity of cyst detection was 42.9%. All 20 values of cyst numbers in clam tissue samples that were processed blind were located within the 95% prediction limits of the linear regression equation. The cyst retention rate of 160 clams kept in an aquarium with 38 l of water spiked with 1.00 x 10(5) G. duodenalis cysts was approximately 1.3 x 10(3) cysts/clam. No waterborne cysts were detected by the membrane filtration method 90 min after spiking the aquarium water. G. duodenalis cysts were detected in clam tissue up to 3 weeks post-exposure. Filtration of water by clams substantially depleted the aquarium water of its particulate matter. The sampling program demonstrated that the population of 160 clams examined during the study could be accurately assessed for exposure to waterborne Giardia cysts by random sampling of 86 (54%) clams. The results indicate that C. fluminea clams can be used for biological monitoring of contamination with Giardia.

Analysis of Variance↗
PubMed1999

Cryptosporidium oocysts in Bent mussels (Ischadium recurvum) in the Chesapeake Bay.

Filter-feeding molluscan shellfish can concentrate environmentally derived waterborne pathogens of humans, which can be utilized in the sanitary assessment of water quality. In the present study, oocysts of Cryptosporidium were detected in Bent mussels (Ischadium recurvum) at two Chesapeake Bay sites from which C. parvum-contaminated oysters had previously been collected. Spiking of Cryptosporidium-free blue mussel (Mytilus edulis) tissue with C. parvum oocysts showed a 51.1% recovery rate of oocysts, giving an oocyst detection limit of 19 oocysts/0.7 ml of mussel tissue homogenate. The results indicate that Bent mussels, which are common throughout the Chesapeake Bay region, may prove to be useful as biological indicators of water contamination with Cryptosporidium oocysts.

Animals↗
PubMed1999

Variation in Cryptosporidium: towards a taxonomic revision of the genus.

Cryptosporidium is an important cause of enteric disease in humans and other animals. Limitations associated with conventional diagnostic methods for cryptosporidiosis based on morphological features, coupled with the difficulty of characterising parasites isolated in the laboratory, have restricted our ability to clearly identify species. The application of sensitive molecular approaches has obviated the necessity for laboratory amplification. Such studies have found considerable evidence of genetic heterogeneity among isolates of Cryptosporidium from different species of vertebrate, and there is now mounting evidence suggesting that a series of host-adapted genotypes/strains/species of the parasite exist. In this article, studies on the molecular characterisation of Cryptosporidium during the last 5 years are reviewed and put into perspective with the past and present taxonomy of the genus. The predictive value of achieving a sound taxonomy for the genus Cryptosporidium with respect to understanding its epidemiology and transmission and controlling outbreaks of the disease is also discussed.

Animals↗
PubMed1999

Phylogenetic analysis of Cryptosporidium parasites based on the small-subunit rRNA gene locus.

Biological data support the hypothesis that there are multiple species in the genus Cryptosporidium, but a recent analysis of the available genetic data suggested that there is insufficient evidence for species differentiation. In order to resolve the controversy in the taxonomy of this parasite genus, we characterized the small-subunit rRNA genes of Cryptosporidium parvum, Cryptosporidium baileyi, Cryptosporidium muris, and Cryptosporidium serpentis and performed a phylogenetic analysis of the genus Cryptosporidium. Our study revealed that the genus Cryptosporidium contains the phylogenetically distinct species C. parvum, C. muris, C. baileyi, and C. serpentis, which is consistent with the biological characteristics and host specificity data. The Cryptosporidium species formed two clades, with C. parvum and C. baileyi belonging to one clade and C. muris and C. serpentis belonging to the other clade. Within C. parvum, human genotype isolates and guinea pig isolates (known as Cryptosporidium wrairi) each differed from bovine genotype isolates by the nucleotide sequence in four regions. A C. muris isolate from cattle was also different from parasites isolated from a rock hyrax and a Bactrian camel. Minor differences were also detected between C. serpentis isolates from snakes and lizards. Based on the genetic information, a species- and strain-specific PCR-restriction fragment length polymorphism diagnostic tool was developed.

Animals↗