Rapid detection and quantification of Cryptosporidium baileyi oocysts in feces and organs of chickens using a microscopic slide flotation method

Abstract

A simple semiquantitative microscopic slide flotation (MSF) method using modified Sheather's sugar solution (MSSS) is presented for the rapid detection and quantification of Cryptosporidium baileyi oocysts in the feces and mucosal and/or organ scrapings of chickens. Oocyst shedding was evaluated by examination of the surface of coverslips, and the average quantitative score (0-5) recorded for 10 microscopic fields (magnification x250) is reported. The equivalence between these scores and the actual number of oocysts counted per gram of feces was assessed (rs = 0.89; P < 0.001). The applicability of this method was tested by comparison of the kinetics of oocyst shedding in feces of inoculated chickens with those reported by other authors working under similar conditions. In organs the MSF method was compared to histology. Fewer false-negative results were obtained using MSF versus the histology method. The MSF method was particularly more efficient in tracheae with low levels of infection and in the lungs, regardless of the level of infection. The MSF method was also very efficient in detecting oocysts in air sacs from chickens with aerosacculitis. It provides a specific and sufficiently sensitive, simple, rapid, reliable, and low-cost means of diagnosing C. baileyi in the feces and organs of chickens. This method can be used in the routine diagnosis of cryptosporidia in chickens, and it could be extended to other avian species and used in epidemiology studies to evaluate the prevalence of cryptosporidiosis in fowl.

Effect of Cryptosporidium baileyi in specific pathogen free chickens vaccinated (CVI988/Rispens) and challenged with HPRS-16 strain of Marek's

“…The strain of C. baileyi was initially isolated from the bursa of Fabricius (BF) of ducks and then maintained for several years by serial passage in SPF chickens in controlled conditions (Naciri et al, 1994;Abbassi et al, 2000). The C. baileyi oocysts collected from the bursa were used for chicken inoculation in our present study and those from the faeces were used to produce the soluble antigen used in serology after concentration and purification.…”

“…The C. baileyi oocysts collected from the bursa were used for chicken inoculation in our present study and those from the faeces were used to produce the soluble antigen used in serology after concentration and purification. The isolation and storage of oocysts and preparation of inoculum were performed as described previously (Abbassi et al, 2000). The oocysts obtained after concentration procedures were purified by discontinuous sucrose gradient similar to that previously described (Arrowood & Sterling, 1987;Ortega-Mora et al, 1992).…”

“…In addition to these organs, samples of larynx, intestinal portions (duodenum, jejunum, ileum and caecum) and, sometimes, air sacs, oral cavity, oesophagus, crop and proventriculus were removed and placed separately in Petri dishes. These samples, which were used for the evaluation of C. baileyi infection by the microscopic slide flotation (MSF) method (Abbassi et al, 2000), were preserved at 4°C until use after necropsy (within 48 h). Samples of the sciatic nerve and brachial plexus were removed from each chicken that died or was killed for histological diagnosis of MD.…”

“…), of the parasite in C4, R0C4M15 and C4R8M15 groups, and once a week in the other groups. Individual oocyst shedding was measured by the MSF method as previously described (Abbassi et al, 2000). At days 42 and 62, blood samples were collected from surviving chickens in all groups and corresponding sera were stored at -20°C until use for C. baileyi serology.…”

“…The kinetics of oocyst 628 H. Abbassi et al (Abbassi et al, 2000 shedding of C. baileyi-inoculated groups are presented in Figure 3, with the number of shedding chickens expressed as a percentage of survivors. All three groups had the same prepatent period (from 4 to 7 days p.i.).…”