Evaluation of two commercial disinfectants on the viability and infectivity of Cryptosporidium parvum oocysts

Cryptosporidiosis is mainly a problem in neonatal ruminants. Not only do Cryptosporidium spp. spread ubiquitously in our environment, but the protozoa are highly resistant to harsh environmental conditions and disinfectants, and a control measure is urgently required. This study investigated the potential biocidal activity on Cryptosporidium parvum oocysts of two commercial disinfectants developed originally to be used in farms and food-processing industries. The products, containing formaldehyde and hydrogen peroxide respectively, both had some anticryptosporidial effects. The viability and infectivity of purified C. parvum oocysts exposed to both disinfectants at different concentrations and exposure times were evaluated by inclusion or exclusion of vital dye (propidium iodide), use of an excystation technique and infection of suckling mice. Viability assays showed a decrease in oocyst viability associated with an increase in exposure time for each of the concentrations used. The intensity of infection in neonatal mice was significantly lower (P<0.05) than in the control litters.     

Study of the combined influence of environmental factors on viability of cryptosporidium parvum oocysts in water evaluated by fluorogenic vital dyes and excystation techniques

Using a factorial experimental design, the combined effect of salinity, temperature and storage time on the viability of Cryptosporidium parvum oocysts in water was evaluated by fluorogenic vital dyes (4',6-diamidino-2-phenylindole and propidium iodide) and an excystation technique. Salinity, storage time and their interaction seemed to be the most influential factors, whereas temperature was not a significant factor. Under unfavourable conditions (salinity 35 per thousand, storage time 40 days), even more than 20% of oocysts remain viable, indicating a high risk of infection for immunocompromised individuals.     

In vitro and in vivo efficacy of alpha-cyclodextrin for treatment of experimental cryptosporidiosis

The efficacy of alpha-cyclodextrin against infection by Cryptosporidium parvum was evaluated using in vitro and in vivo models. Cyclodextrins are water-soluble cyclic hexamers of glucose units with hydrophobic cavities capable of solubilizing lipophiles and are widely used as drug excipients in the pharmaceutical industry. The viability of purified C. parvum oocysts, exposed for 30, 60, 90, 120 min and 24h to different concentrations of alpha-cyclodextrin (2.5, 5, 7.5, 10, 12.5 and 15%), was evaluated by inclusion or exclusion of two fluorogenic vital dyes and by an excystation technique. Preventive and curative efficacies against cryptosporidial infections, at different doses (2.5 and 5%) and regimes of administration of alpha-cyclodextrin, were determined in an experimental neonatal mice model. Results of the viability assay showed a decrease in oocyst viability that was associated with an increase in exposure time, for each of the concentrations used. Moreover, a high proportion of nonviable oocysts (81%) was observed when C. parvum oocysts were exposed to alpha-cyclodextrin (2.5%) for 24h. The intensity of infection, determined 7 days post-inoculation by examination of intestinal homogenates, was significantly lower (P<0.05) than in the control litters, for all the assays carried out with alpha-cyclodextrin. Only 38.8% of the animals became infected when the alpha-cyclodextrin solution (5%) was administered 2h before inoculated oocysts, and every 24h at 1 and 2 days post-inoculation.     

用核酸染色和小鼠感染力方法评估脱囊和热处理后的隐孢子虫卵囊的活力

温度是影响隐孢子虫活力的重要环境因素之一.对于疾病暴发的风险评估需要有效的方法来准确分析卵囊的活力.本试验应用核酸染色和小鼠感染力2种方法研究了脱囊和热处理后完整卵囊和不完整卵囊的活力,并与新鲜卵囊进行了对比.结果表明,脱囊和中性温度热处理后的完整卵囊保持活力.然而,高温处理后的完整卵囊以及脱囊和热处理后的不完整卵囊完全丧失活力.对于新鲜卵囊,灭活卵囊,以及在脱囊后或在40℃和70℃处理后的完整和非完整卵囊,核酸染色法和小鼠感染力法的结果相对应;但是对于50℃和60℃处理后的完整卵囊这2种方法不对应.     

In vitro and in vivo efficacy of lasalocid for treatment of experimental cryptosporidiosis

In vitro viability of purified Cryptosporidium parvum oocysts, exposed for 30, 60, 90 and 120 min to 0.27 mg/ml lasalocid suspension was evaluated by inclusion or exclusion of two fluorogenic vital dyes and an excystation technique. Continuously, preventive and curative efficacies at different doses (9, 6.75, 5.625 and 4.5 mg/kg body weight) and regimens of lasalocid against cryptosporidial infection were evaluated on an experimental neonatal mice model. In vitro assays demonstrated a decrease in the oocyst viability related to an increase in exposure time for exposure to the lasalocid suspension. The infection was eradicated when the suspension was administered with a dose of ≥6.75 mg/kg body weight. No apparent toxic effects were observed.     

Comparison of viability and infectivity of Cryptosporidium parvum oocysts stored in potassium dichromate solution and chlorinated tap water

The present study was undertaken to compare the viability and infectivity of Cryptosporidium parvum oocysts that had been stored for 1, 4, 7, 10, 13, 16, 20, 25 and 30 months at 4 degrees C in 2.5% potassium dichromate (Cr) or chlorinated tap water, respectively. An excystation protocol was performed in vitro to evaluate viability. One hundred and eighty female BABL/c mice were used to evaluate the infectivity of oocysts by investigating the prepatent period of C. parvum infection, the quantity of oocysts excreted, and the number of parasites that colonized the villi of the ileum. The results showed that C. parvum oocysts preserved in Cr for 1-16 months or in water for 1-13 months were capable of excystation in vitro and infection of mice. The excystation rates of oocysts and the prepatent periods in mice infected by oocysts stored in Cr and water were not significantly different (p>0.05), and there was a strong correlation between prepatent period and duration of oocyst storage (Cr: R2=0.92; water: R2=0.98). There were no significant differences in oocyst shedding from feces or parasitism of the terminal ilea of mice by Cryptosporidia between the two storage media (p>0.05). In conclusion, C. parvum oocysts may be stored at 4 degrees C in water instead of Cr for the purposes of laboratory research. However, the presence of viable C. parvum oocysts in water is a severe challenge to the drinking water treatment industry.     

Viability and infectivity of Cryptosporidium parvum oocysts detected in river water in Hokkaido,Japan

The viability and infectivity of Cryptosporidium parvum (C. parvum) oocysts, detected in water samples collected from river water in Hokkaido, were investigated using Severe Combined Immunodeficient (SCID) mice. The water samples collected from September 27 through October 10, 2001 by filtration using Cuno cartridge filters were purified and concentrated by the discontinuous centrifugal flotation method. From 1.2 x 10 (5) liters of the raw river water, approximately 2 x 10(4) oocysts were obtained and designated as Hokkaido river water 1 isolate (HRW-1). Oocyst identification was carried out using microscopic and immunological methods. Six 8-week-old female SCID mice were each inoculated orally with 1 x 10 (3) oocysts. Infection was successfully induced, resulting in fecal oocyst shedding. Oocysts were then maintained by sub-inoculation into SCID mice every 3 months. Infectivity was evaluated by making comparisons with two known C. parvum stocks, HNJ-1 and TK-1, which were bovine genotypes detected in fecal samples from a cryptosporidiosis patient and young cattle raised in Tokachi, Hokkaido respectively. The oocyst genotypes were determined from a small subunit ribosomal RNA (SSU-rRNA) gene by polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) analysis. No significant differences were observed in the average number of oocysts per gram of feces (OPG) in any of the isolates. Our data indicates that the C. parvum oocysts detected in the sampled river water were of C. parvum genotype 2. Moreover, our data on the continued isolation, detection and identification of the C. parvum isolates is consistent with the available epidemiological data for the Tokachi area.     

保存在自来水中的微小隐孢子虫卵囊活性及感染性研究

微小隐孢子虫卵囊(CPO)保存在4℃自来水中1~30个月,通过体外脱囊技术检测CPO的脱囊率评价其活性,通过检测CPO感染免疫抑制BALB/c小鼠的潜伏期、排卵囊数量和末端回肠绒毛中的隐孢子虫数量来评价其感染性。结果表明,保存在自来水中1~13个月的CPO出现脱囊;小鼠在感染保存1~13个月的CPO后3~8 d开始排出大量的CPO,在末端回肠绒毛中寄生有大量的隐孢子虫;CPO的保存时间与潜伏期之间存在强烈的相关性(r2=0.98)。因此,CPO在自来水中能保持活性和感染性至少13个月,水是保存CPO的良好介质,水中活性CPO的长期存在对饮用水工业是一个严重的挑战。     

Activity of decoquinate against Cryptosporidium parvum in cell cultures and neonatal mice

Cryptosporidium parvum is an apicomplexan parasite that is an important cause of diarrhea in neonatal calves and humans. No treatment is currently available for neonatal calves. We have recently learned from colleagues in the pharmaceutical industry that dairy practitioners are sometimes using decoquinate for the treatment of neonatal bovine cryptosporidiosis. Therefore, the present study was undertaken to determine whether the clinical observations in calves can be substantiated by laboratory investigation. Oocysts of the KSU-1 isolate of C. parvum were used to infect human ileocecal epithelial cells in vitro to measure the efficacy of treatment using an ELISA based assay. No activity was observed at 10 or 50microM decoquinate, but at 100microM an 8% inhibition of development was seen. Oocysts of the AUCp-1 isolate of C. parvum were then used to infect suckling mice. The numbers of oocysts observed in suckling mice treated with 2.5 or 5.0mg/kg decoquinate were not significantly different from untreated control suckling mice (p0.05). The results of our study suggest that decoquinate should have little efficacy for treatment of neonatal bovine cryptosporidiosis if administered once per day and that any clinical improvement observed in treated calves may be due to factors unrelated to decoquinate's effect on C. parvum.     

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

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

Efficacy of amine-based disinfectant KENO™COX on the infectivity of Cryptosporidium parvum oocysts

Cryptosporidium parvum is a zoonotic protozoan parasite that may cause severe neonatal diarrhoea or even mortality in newborn ruminants: its oocysts are extremely resistant to normal environmental conditions and to most common disinfectants. KENO?COX, a patent pending amine-based formula, was tested for its ability to inactivate C. parvum oocysts. The Daugschies assay (2002), a standardized assay for chemical disinfection initially described for Eimeria spp., was adapted for C. parvum oocysts. KENO?COX diluted in water at 2% and 3% concentration and incubated with oocyst suspensions for 2h, allowed a significant reduction in viability, lysing 89% and 91% of oocysts respectively. Infectivity of the remaining C. parvum oocysts was assessed by inoculation to C57 Bl/6 neonatal mice. Each mouse received 2.5 μl of a suspension initially containing 500,000 oocysts before contact with KENO?COX. Six days post inoculation, the intestinal parasite load was significantly reduced by 97.5% with KENO?COX 2% compared to that of the mice inoculated with untreated parasites. KENO?COX 3% completely eliminated infectivity of oocysts. The number of oocysts remaining infectious in the inoculum treated with KENO?COX 2% was calculated from an inoculated dose-response curve: it was estimated at about 48.6 oocysts among the 500,000 oocysts initially treated corresponding to 99.99% of inhibition. These results demonstrate the high efficacy of KENO?COX against C. parvum oocysts. Combined with an appropriate method of cleaning, the application of KENO?COX may be a useful tool to reduce cryptosporidial infectious load on farm level.     

Comparison of viability assays for Cryptosporidium parvum oocysts after disinfection

In order to test various viability assays for Cryptosporidium parvum oocysts were used to infect HCT-8 cells in vitro or baby mice. Infected cells were either stained with fluorescent anti-Cryptosporidium-antibody or lysed and subjected to C. parvum-specific PCR after 48 h. Titrations with infective oocysts were performed and compared to oocysts disinfected with Neopredisan for 2 h at varying concentrations. Caecal smears and histological sections from infected animals were examined in parallel. The number of foci of parasite development in vitro after immunofluorescent staining correlated well with the infection dose. PCR was less quantifiable and the results were not always reproducible, especially when low infection doses were used. Disinfection resulted in a dose-dependent reduction of oocyst infectivity when compared to the controls in all three assays. The infection of cells cultured in vitro with oocysts of C. parvum provides a suitable tool for the estimation of viability after treatment with chemical disinfectants. Immunofluorescence is easy to perform and gives quantitative results, while PCR-based detection of parasite DNA, although possible, requires the use of more sophisticated tools for quantification.     

The effect of heating against Cryptosporidium oocysts

The effect of heat treatment was examined against oocysts of Cryptosporidium parvum, Cryptosporidium muris and chicken Cryptosporidium sp. isolated in Japan. The oocysts of these species were exposed at 50, 55, 60 and 70 degrees C for 5, 15, 30 and 60 sec in water bath, respectively. To determine the infectivity of heated oocysts, the nice and chickens were inoculated with the treated oocysts and the oocyst output in the feces after inoculation was examined. In C. parvum and chicken Cryptosporidium sp., the oocysts were not detected from mice or chickens which were received oocysts heated at 55 degrees C for 30 sec, 60 degrees C for 15 sec and 70 degrees C for 5 sec. In C. muris, the oocysts were not detected from mice which were received oocysts heated at 55 degrees C for 15 sec, 60 degrees C for 15 sec and 70 degrees C for 5 sec. Consequently, it was clarified that the infectivity of Cryptosporidium oocysts to mice and chickens was lost by heating at 55 degrees C for 30 sec, 60 degrees C for 15 sec and 70 degrees C for 5 sec.     

Infectivity to experimental rodents of Cryptosporidium parvum oocysts from Siberian chipmunks (Tamias sibiricus) originated in the People's Republic of China

We isolated Cryptosporidium parvum-type oocysts from naturally infected siberian chipmunks which originated in the People's Republic of China and examined the infectivity to rodents as experimental animals. The naturally infected chipmunks did not show any clinical symptoms. The oocysts were 4.8 x 4.2 microm on average in size. They were ovoid and morphologically similar to the C. parvum oocysts isolated from human and cattle. Experimental rodents were inoculated with 1.6 x 10(6) original oocysts each. SCID mice began to shed oocysts on day 7 and the OPG value was 10(5) from 50 days. The oocysts were found from ICR mice on days 13 and 16 by only sugar flotation method, however, any oocysts were not detected from the rats, guinea pigs and rabbits until 30 days. Two infected SCID mice were necropsied on days 100 and 102 and examined for coccidian organisms. Merozoites and oocysts were found in the low part of jejunum and ileum, however, no parasites were detected in the stomach. Consequently, it was considered that the present species was C. parvum and was probably genotype 2 from result of infectivity to rodents.     

Comparative pathogenicity of selected bovine viral diarrhea virus isolates in gnotobiotic lambs

The infectivity and pathogenicity of selected bovine viral diarrhea virus (BVDV) isolates were determined in gnotobiotic, colostrum-deprived neonatal lambs. Five-day-old cesarean-derived gnotobiotic lambs were exposed to 1 of 10 BVDV isolates via aerosol suspension. These isolates were from tissues or secretions of calves or lambs affected with respiratory tract disease, weak neonatal calves, aborted bovine fetuses, or reference Singer or Draper BVDV. The pathogenicity of each isolate, relative to the others, was evaluated in lambs by measurement of the neutralizing antibody response, virus isolation from nasal secretions or tissues, and postmortem lesions. The BVDV isolates varied in their infectivity and pathogenicity. Singer, the cytopathic reference strain, was the most lymphotrophic isolate and stimulated the greatest neutralizing antibody response. Encephalitis was the most consistent lesion observed and was used as the final determinant of relative pathogenicity of the viruses. The most neuropathogenic isolates were the 2 viruses originating from lambs affected with respiratory tract disease, the 2 weak neonatal calf isolates, and 1 isolate from an aborted bovine fetus. The least pathogenic isolates were the 2 reference isolates, Draper and Singer; the 2 mucosal disease isolates; and 1 isolate originating from an aborted bovine fetus.     

Viability of Sarcocystis neurona sporocysts after long-term storage

The effect of long-term storage on the viability and infectivity of Sarcocystis neurona sporocysts was investigated. S. neurona sporocysts were harvested from the small intestine of Virginia opossums from 1996 to 2002 and stored at 4 degrees C. Viability of sporocysts was assessed by propidium iodide (PI) exclusion assay, in vitro excystation and development in tissue cultures, and bioassay in gamma-interferon gene knockout (gamma-IFN-KO) mice. The rate of excystation was apparently unaffected by long-term storage; sporocysts retained their ability to excyst after 7 years of storage at 4 degrees C. However, the ability of sporocysts to exclude PI stain, to invade and proliferate in cells in vitro, and to cause disease and lesions in gamma-IFN-KO mice appeared to decline as sporocysts age. The results demonstrated that sporocysts of S. neurona were able to survive and maintain moderate to high viability for up to 7 years when stored in phosphate buffered saline and Hank's balanced salt solution containing antibiotic-antimycotic mixture at 4 degrees C.     

Role of Cryptosporidium parvum as a pathogen in neonatal diarrhoea complex in suckling and dairy calves in France.

This study was carried out to find the importance of Cryptosporidium parvum in diarrhoea of neonatal calves in two types of breeding - suckling and dairy calves - in France. Different agents causing neonatal diarrhoea, E. coli, rotavirus, coronavirus, Salmonella and Cryptosporidium were systematically researched in faeces. 1. Suckling calves: In 40 livestock farms selected for diarrhoea, 311 calves 4 to 10 days old which had diarrhoea for less than 24h or no diarrhoea, were included in the study. A prophylaxis of neonatal diarrhoea had been carried out in 21 of the 40 livestock farms. On D0 (inclusion day), the mean age was 6 days, 82% presented a good initial general condition and 76.2% had a good appetite; 48.6% were diarrhoeic but 91.3% presented no sign of dehydration. Only 6.1% were infected by E. coli K99, 14.3% by rotavirus, 6.8% by coronavirus, 0.3% by Salmonella but 50% excreted C. parvum oocysts. This later percentage increases up to 84% and 86% by D3 and D7, respectively . We note that 16% of the 4-day-old calves on D0 are excreting oocysts and this percentage increases as a function of the age of the calf on D0 to reach 90% to 95% by the age of 8 days. 10 out of 12 dead calves excreted C. parvum oocysts. From D0 to D14 the other pathogen agents show a relative or a decreasing stability. 2. Dairy calves: 382 calves which had diarrhoea for less than 24 h or no diarrhoea, aged 8 to 15 days coming from six industrial livestock farms were included in the study. On D0, 99% of the calves presented a good initial general condition, 99.7% had a good appetite and no calf was dehydrated. At this date (D0), 16.8% of the calves excreted cryptosporidia. This percentage increases up to 23% and 51.8% on D3 and D8, respectively, then decreases to 31.9% on D14. The pressure of the other pathogenicagents remains relatively stable, excepted for rotavirus on D7 (from 9.9% on D0 to 27.2% on D7, then 12.6% on D14) which does not explain the concomitantpeak in diarrhoea because the infection by rotavirus on D7 is more frequent in non-diarrhoeic calves than in diarrhoeic calves. Our results show that Cryptosporidium prevalence is higher in suckling than in dairy calves and C. parvum constitutes actually in both cases the major aetiological agent of neonatal diarrhoea.     

Calf-level risk factors for neonatal diarrhea and shedding of Cryptosporidium parvum in Ontario dairy calves

This work was conducted to investigate calf-level factors that influence the risk of neonatal diarrhea and shedding of Cryptosporidium parvum oocysts in calves, on dairy farms in Ontario with histories of calf diarrhea or cryptosporidiosis. Fecal samples were collected weekly for 4 weeks from each of 1045 calves under 30 days of age on 11 dairy farms in south-western Ontario during the summer of 2003 and the winter of 2004. A questionnaire designed to gather information on calf-level management factors was administered on farm for each calf in the study. Samples were examined for C. parvum oocysts by microscopy, and a subset of specimens was also tested for enterotoxigenic Escherichia coli, Salmonella, bovine rotavirus and bovine coronavirus. The consistency of each sample was scored and recorded at the time of collection in order to assess the presence or absence of diarrhea. In addition, a blood sample was taken from each calf upon enrollment in the study, for assessment of maternal antibody transfer and for polymerase chain reaction testing for persistent bovine viral diarrhea virus infection. Using the GLLAMM function in Stata 9.0, multilevel regression techniques were employed to investigate associations between management practices and the risk of C. parvum shedding or diarrhea. C. parvum oocysts were detected in the feces of 78% of the 919 calves from which all four fecal samples had been collected. Furthermore, 73% of the 846 calves for which all four fecal consistency scores had been recorded were diarrheic at the time of collection of at least one sample. Significant predictors of the calf-level risk of C. parvum shedding included the use of calf diarrhea prophylaxis in pregnant cows, and the type of maternity facilities in which the calves were born. Factors associated with an increased risk of diarrhea were leaving the calf with the dam for more than an hour after birth, and the birth of a calf in the summer as opposed to winter. Calves shedding C. parvum oocysts had 5.3 (95% CI 4.4, 6.4) times the odds of diarrhea than non-shedding calves, controlling for other factors included in the final multivariable model. Furthermore, infected calves shedding more than 2.2 x 10(5) oocysts per gram of feces were more likely to scour than infected calves shedding lower numbers of oocysts (OR= 6.1, 95% CI 4.8, 7.8). The odds of diarrhea in calves shedding oocysts that had been allowed to remain with their dams for more than an hour were higher than the odds of diarrhea in shedding calves that had been separated from their dams within an hour after birth.     

Genetic and biologic characteristics of Toxoplasma gondii isolates in free-range chickens from Colombia, South America

The prevalence of Toxoplasma gondii in free-ranging chickens is a good indicator of the prevalence of T. gondii oocysts in the soil because chickens feed from the ground. The prevalence of T. gondii in 77 free-range chickens (Gallus domesticus) from Colombia, South America was determined. Antibodies to T. gondii were assayed by the modified agglutination test (MAT), and found in 32 (44.4%) of 72 chickens with titers of 1:5 in 4, 1:10 in 3, 1:20 in 1, 1:40 in 1, 1:80 in 8, 1:160 in 8, 1:320 in 3, and 1:640 or higher in 4. Hearts and brains of 31 seropositive chickens were pooled and bioassayed in mice. Tissues from 32 (16+16) seronegative chickens were pooled and fed to two, T. gondii-free cats, and tissues from nine chickens without matching sera were fed to one T. gondii-free cat. Feces of cats were examined for oocysts. T. gondii oocysts were excreted by a cat that was fed tissues of 16 seronegative chickens. T. gondii was isolated by bioassay in mice from 23 chickens with MAT titers of 1:20 or higher. All infected mice from 16 of the 23 isolates died of toxoplasmosis. Overall, 82 (81.1%) of 101 mice that became infected after inoculation with chicken tissues died of toxoplasmosis. Genotyping of these 24 isolates using polymorphisms at the SAG2 locus indicated that seven T. gondii isolates were Type I, 17 were Type III, and none was Type II. Phenotypically, T. gondii isolates from chickens from Colombia were similar to isolates from Brazil but different from the isolates from North America; most isolates from chickens from Brazil and Colombia were lethal for mice whereas isolates from North America did not kill inoculated mice. Genetically, none of the T. gondii isolates from Colombia and Brazil was SAG2 Type II, whereas most isolates from chickens from North America were Type II. This is the first report of genetic characterization of T. gondii isolates from Colombia, South America.     

Comparison of staining and concentration techniques for detection of Cryptosporidium oocysts in cat faecal specimens.

Modified Ziehl-Neelsen (MZN), auramine-phenol (A-P) and fluorescein isothiocyanate-labelled (FITC-labelled) monoclonal antibody (MAb) techniques were compared for detection of Cryptosporidium parvum oocysts in cat faecal specimens inoculated with known numbers of C. parvum oocysts. Of the three techniques, the FITC-labelled MAb technique detected more oocysts than the MZN and A-P techniques (P < 0.05), but A-P was more efficient than MZN (P < 0.05). Comparison of sucrose flotation, zinc sulphate (ZnSO4) flotation and formol-ether (F-E) sedimentation techniques revealed that F-E was the most efficient of the three (P < 0.05) for concentration of C. parvum oocysts from cat faecal specimens. On average, the F-E technique recovered 37% of oocysts from the original sample, whereas the sucrose and ZnSO4 flotation techniques recovered 33% and 11%, respectively. The findings of this study suggest that MZN and A-P staining are both useful for screening C. parvum oocysts in cat faecal materials containing 10(6) oocysts or more, but FITC-labelled MAb should be used when the number of oocysts is low. Also, the F-E sedimentation technique is recommended for concentrating oocysts in cat faecal specimens.