The “self-cleaving” 2A sequence of picornavirus, which mediates ribosome-skipping events, enables the generation of two or more separate peptide products from one mRNA containing one or more “self-cleaving” 2A sequences. In this study, we introduced a single 2A sequence of porcine teschovirus-1 (P2A) linked to two fluorescent protein genes, the enhanced yellow fluorescent protein (EYFP) gene and the red fluorescent protein (RFP) gene, in a single cassette into transgenic Eimeria tenella (EtER). As expected, we obtained two separated protein molecules rather than a fused protein, although the two molecules were translated from the same mRNA carrying a single “self-cleaving” 2A sequence. Importantly, RFP led by a secretion signal was secreted into parasitophorous vacuoles, while EYFP localized mainly to the nucleus of EtER. Our results demonstrate that the “self-cleaving” 2A sequence actively mediated cleavage of polyproteins in the apicomplexan parasite E. tenella.

Coccidiosis is a challenging disease of wild and domestic rabbits both, caused by Eimeria and thereby leads enormous economic losses at rabbit farms. The present study carried out to survey the prevalence and intensity of coccidial infection among the rabbits in Sichuan Province, southwest China. A total of 110 faecal samples were collected from 11 farms situated in eight main rabbits rearing administrative regions. Oocysts in faecal samples were purified, sporulated and identified according to morphological features. The overall prevalence of infection was 56.4 % (62/110), with prevalence of 64 % (47/75) for local meat breeds of rabbit and 51.4 % (18/35) for Rex Rabbits (local fur rabbits). Weanling rabbits had the highest prevalence (74 %, 37/50), followed by young rabbits (45 %, 13/29) and the adult rabbits showed the lowest prevalence (42 %, 13/31). Concurrent infection with two to seven Eimeria species was found. In total, 9 species of Eimeria were identified from oocyst-positive samples. E. perforans was the most prevalent specie (42.73 %), followed in order by Eimiera media, E. irresidua, E. magna, and E. intestinalis with prevalences of 35.45, 34.55, 31.82, and 23.64 %, respectively. Results of the present investigation indicated that the prevalence of coccidial infection is high among the rabbit population in southwest China. This study also elucidate about the coccidial infection and emphasis to adopt control strategies in commercial rabbitories.

Parasitic diseases are major trouble in many parts of the world. We consider that if a chemical can break a DNA barcode sequence, it might be used to develop a species-specific anti-parasitic agent. To examine this hypothesis, we constructed sgRNAs that target both the control (5.8S rDNA) and a DNA barcode (ITS) sequence in Eimeria tenella. In vitro experiment showed that Cas9 mRNA combined with sgRNAs could reduce the sporulation percentage of oocysts and the survival rate of sporulated oocysts and sporozoites. Quantitative real-time PCR showed that the DNAs of parasites exposed to Cas9 mRNA and sgRNAs were significantly affected, regardless of whether they were exposed to a combination of two sgRNAs or just a single sgRNA. The DNA sequencing also indicated that the experimental group exposed to two sgRNAs mixed with Cas9-induced deletion of large parts and a single sgRNA mixed with Cas9-induced mutation at sgRNA targeted fragments. In vivo trial, the effect of sgRNA and Cas9 RNA on the pathogenicity of E. tenella in chicken showed less lesion score and oocysts score (P < 0.05) in experimental groups than control groups. The results and concepts presented in this research can lead to discovering novel nucleic acid therapeutic drugs for Eimeriasis and other parasitic infections, which provide insights into the development of species-specific anti-parasitic agents.

Eimeria species are obligate intracellular apicomplexan parasites, causing great economic losses in the poultry industry. Currently wild-and attenuated- type anticoccidial vaccines are used to control coccidiosis. However, their use in fast growing broilers is limited by vaccination side effects caused by medium and/or low immunogenic Eimeria spp. There is, therefore, a need for a vaccine with high immunogenicity for broilers. The avian yolk sac IgY Fc is the avian counterpart of the mammalian IgG Fc, which enhances immunogenicity of Fc-fusion proteins. Here, we developed a stable transgenic Eimeria mitis expressing IgY Fc (Emi.chFc) and investigated whether the avian IgY Fc fragment enhances the immunogenicity of E. mitis. Two-week-old broilers were immunized with either Emi.chFc or wild type Eimeria and challenged with wild type E. mitis to analyze the protective properties of transgenic Emi.chFc. Chickens immunized with Emi.chFc had significantly lower oocyst output, in comparison with PBS, mock control (transgenic E. mitis expressing HA1 from H9N2 avian influenza virus) and wildtype E. mitis immunized groups after challenge, indicating that IgY Fc enhanced the immunogenicity of E. mitis. Our findings suggest that IgY Fc-expressing Eimeria may be a better coccidiosis vaccine, and transgenic Eimeria expressing Fc-fused exogenous antigens may be used as a novel vaccine-delivery vehicle against a wide variety of pathogens.

Immune mapped protein-1 (IMP1) of E. maxima has been identified as a vaccine antigen candidate for E. maxima infection.In the current study, the N- and C-terminal derivative of EmIMP1 were expresse...

This study evaluated whether oyster crude polysaccharides (OPS) attenuates lipopolysaccharide (LPS)-induced immune stress in weanling piglets. Thirty healthy crossbred piglets (28 ± 1 days old) were randomly divided into five groups (6 piglets/group). Blank control and LPS groups were fed with the basal diet, while low, medium and high dose of OPS groups were fed with the basal diet supplemented with 0.5, 0.8 and 1.2 % OPS, respectively, for 30 days. LPS group, as well as low, medium and high dose of OPS groups were then injected intraperitoneally with LPS (100 μg/kg body weight), whereas the blank control group was given phosphate buffered saline. The concentrations of TNF-α, IL-1β and IL-6 in plasma were detected by ELISA. The mRNA levels of PPARγ in liver, spleen, adrenal gland and thymus were evaluated by quantitative real-time PCR. The results showed that compared with the blank control, LPS treatment significantly increased plasma IL-1β, IL-6 and TNF-α levels, which was significantly attenuated by supplementing 0.5, 0.8 or 1.2 % OPS in the diet. In addition, LPS significantly induced expression of PPARγ mRNA in liver, spleen, adrenal gland, and thymus, which was blocked by adding OPS regardless of the doses. These results indicate that dietary supplementation of OPS was able to alleviate the immune stress induced by LPS.