The ciliate Tetrahymena paravorax strain S in a monoxenic culture was sterilized by treatment with antibiotics and then was cultured axenically in a proteose peptone-containing (PYG) medium. The optimal temperature of the cell growth was at 26 to 28℃. The initial cell density is not an important parameter for the early stage of the cell growth in the PYG medium because the cells started proliferation and grew at the same rate independent of the initial cell density. The culture medium in the logarithmic phase had activities to enhance the growth rate of the ciliate, while the culture medium in the stationary phase had growth-inhibitory activities. These results show release of factors from Tetrahymena to regulate its proliferation.
Trials were conducted in groups of 14 day old Ross Broiler and Lohmann Brown laying birds experimentally infected with 1,250, 5,000, 20,0000 80,000, 320,000, 1,280,000, or 5,120,000 oocysts of a field strain of Eimeria acervulina. The oocyst output, weight gain and performance, clinical signs and mortality were recorded for 14 days post infection. Birds were also necropsied for lesion scoring and histopathological examination. Performance was related to the level of challenge. There was good correlation between clinical signs and post mortem findings with severe lesions present, extending from the duodenum to the yolk-stalk, in birds in the more heavily infected groups. The histopathological observations related to both the level of parasite challenge and the interval post infection. The infected birds showed varying degrees of villous atrophy of the duodenum and upper small intestine with hyperaemia and inflammation of the mucosa resulting in villous thickening and “clubbing”. Parasitised cells were characteristically hypertrophied and, in the heavier infections (80,000 oocysts upwards) hyperplasia of the epithelial cells was evident in the crypts.
Seven different staining techniques, enzyme linked immunosorbent assay, immunofluorescence and two counting methods were compared for the detection of Cryptosporidium oocysts in faecal samples. Auramine phenol and modified Ziehl-Neelsen techniques were the most useful staining procedures with a detection level of 20,000 oocysts per gram. Both of these methods are suitable for the detection of C. parvum oocysts in samples from clinically affected hosts, where large numbers of oocysts are generally excreted. The former method was marginally clearer when lower numbers of oocysts were present. Immunofluorescence was more sensitive with a detection limit of 5,000 oocysts per gram, whereas enzyme linked immunosorbent assay had a detection limit of 50,000 oocysts per gram of faeces. The Fuchs Rosenthal counting method was relatively insensitive with a reproducible detection limit of 5×105 oocysts per gram, but very accurate with a percentage standard error over the range assessed of less than 5%. The “sensitive” counting method, though extremely laborious, had a detection limit of 50 oocysts per gram, but was less accurate at the lower end of the range with a percentage standard error of more than 20% when concentrations were of 500 oocysts per gram or less.
To generate transgenic mice carrying a protozoan gene, 3.1 kb DNA fragments carrying the CAG promoter ligated with a protozoan gene encoding a major surface antigen SAG-1 (p30) of Toxoplasma gondii, were microinjected into one of pronuclei of embryos of C57BL/6J and BALB/c mice. The embryos were transferred to the oviducts of ICR pseudopregnant recipients. The transgene was detected by polymerase chain reaction in DNA sequence purified from tissue biopsies. Out of 159 mice that developed from injected eggs two p30-founders (BALB/c and C57BL/6J) were obtained. The transgene was not detected in 52 pups of F1 progeny of C57BL/6J founder. However, 3(17.6%) out of 17 F1 progenies from BALB/c founder were found to have p30 gene and they inherited the transgene to 39(55.7%) of 70 F2 progenies. These results may afford the opportunity to study the role of SAG-1 (P30) gene in Toxoplasma gondii infection.