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Coccidiosis is one of the most prevalent diseases seen in the poultry industry leading to excessive economic losses. The aim of this study was to investigate the effect of butyric acid glycerol esters (BE) on the ileal and cecal microbiota in birds challenged with Eimeria maxima(EM). Ross 708 male broilers were fed a diet supplemented with 0 (control) or 0.25% BE from day 1. On day 21, half of the birds were infected with 103 EM oocysts. For determing microbiota, ileal and cecal contents and epithelial scrapings were collected at 7 and 10 D postinfection (PI). Alpha diversity of bacterial communities was mostly affected (P< 0.05) by time PI and EM infection. The richness of luminal bacterial populations in the ileum and ceca was affected (P< 0.05) by addition of BE and by time PI × EM × BE interaction, respectively. In the ileal and cecal luminal and mucosal bacterial communities, permutational multivariate analysis of variance (PERMANOVA, unweighted UniFrac) showed significant (P < 0.05) differences because of time PI and interaction between time PI, EM, and BE. Significant (P< 0.05) differences in taxonomic composition at the family level were observed in microbiota of luminal and mucosal populations of the ileum and ceca owing to time PI, EM, BE, and their interactions. The bacterial community present in the cecal lumen was characterized by the lowest number of differential bacteria, whereas the cecal mucosal community was characterized by the highest number of differentially abundant bacteria. In conclusion, our results show that EM infection and time PI has the biggest impact on microbial diversity in the chicken gut. The presence of BE in the diet had a limited effect on gut microbiota.

Vaccination of chickens against avian coccidiosis in chickens often involves storing Eimeriaoocysts for months after oocyst propagation and sporulation. The purpose of this study was to determine how long E. acervulina, E. maxima, and E. tenellaoocysts remained viable when stored at refrigeration (4°C) or egg room (20°C) temperatures. Separate tubes containing E. acervulina, E. maxima, or E. tenellaoocysts were stored at these temperatures and a sample removed every 3 mo for inoculating chickens for evidence of a patent infection. Also, an aliquot of each Eimeriaspecies at each time–temperature combination was subjected to in vitro excystation to quantify the relative number of released sporozoites to intact (nonexcysted) sporocysts. Eimeria tenellaappeared to be most susceptible to storage in that no oocyst production was observed at 9 mo at either temperature. Although E. maximaoocysts were viable at 9 mo, no oocyst production was observed at 12 mo storage at these 2 temperatures. Quite unexpected was that E. acervulinawas much more stable than E. tenella and E. maximaremaining viable up to and including 27 mo at 4°C and up to and including 12 mo at 20°C. No consistent correlation was observed between in vivo oocyst production and in vitro excystation arising from these 2 respective temperatures (E. acervulinar = 0.58, r = 0.54; E. maximar = 0.90, r = 0.54; E. tenellar = 0.38, r = 0.90). These data indicate that attention must be paid to time and temperature of Eimeriaoocyst storage, and that sporozoite excystation may not be a good indicator of oocyst viability, particularly at later timepoints in incubation.

The prevalence, size, genome, and life cycle of Eimeria acervulinamake this organism a good surrogate for Cyclospora cayetanensis, a protozoan that causes gastroenteritis in humans, including recent outbreaks in the United States and Canada associated with contaminated raspberries and basil. Laboratory studies of C. cayetanensisare difficult because of the lack of readily available oocysts and of infection models and assays. UV radiation and high-hydrostatic-pressure processing (HPP) are both safe technologies with potential for use on fresh produce. Raspberries and basil were inoculated with sporulated E. acervulinaoocysts at high (106oocysts) and low (104oocysts) levels, and inoculated and control produce were treated with UV (up to 261 mW/cm2) or HPP (550 MPa at 40°C for 2 min). Oocysts recovered from produce were fed to 3-week-old broiler chickens, which were scored for weight gain, oocyst shedding, and lesions at 6 days postinoculation. Oocysts exhibited enhanced excystation on raspberries but not on basil. Birds fed oocysts from UV-treated raspberries had reduced infection rates, which varied with oocyst inoculum level and UV intensity. Birds fed oocysts from UV-treated raspberries (104oocysts) were asymptomatic but shed oocysts, and birds fed oocysts from UV-treated basil (104oocysts) were asymptomatic and did not shed oocysts. Birds fed oocysts from HPP-treated raspberries and basil were asymptomatic and did not shed oocysts. These results suggest that UV radiation and HPP may be used to reduce the risk for cyclosporiasis infection associated with produce. Both treatments yielded healthy animals; however, HPP was more effective, as indicated by results for produce with higher contamination levels.

Coccidiosis is a ubiquitous disease caused by intestinal protozoan parasites belonging to several distinct species of the genus EIMERIA: Cell-mediated immunity (CMI) is critically important for protection against Eimeria; thus, our approach utilizes the bacterial type III secretion system (TTSS) to deliver an antigen directly into the cell cytoplasm of the immunized host and into the major histocompatibility complex class I antigen-processing pathway for induction of CMI and antigen-specific cytotoxic T-lymphocyte responses in particular. To accomplish this goal, Eimeria genes encoding the sporozoite antigen EASZ240 and the merozoite antigen EAMZ250 were fused to the Salmonella enterica serovar Typhimurium effector protein gene sptP in the parental pYA3653 vector, yielding pYA3657 and pYA3658, respectively. SptP protein is secreted by the TTSS of Salmonella and translocated into the cytoplasm of immunized host cells. The host strain chromosomal copy of the sptP gene was deleted and replaced by a reporter gene, xylE. The newly constructed vectors pYA3657 and pYA3658 were introduced into host strain χ8879 (ΔphoP233 ΔsptP1033::xylEΔ asdA16). This strain is an attenuated derivative of the highly virulent strain UK-1. When strain χ8879(pYA3653) as the vector control and strain χ8879 harboring pYA3657 or pYA3658 were used to orally immunize day-of-hatch chicks, colonization of the bursa, spleen, and liver was observed, with peak titers 6 to 9 days postimmunization. In vitro experiments show that the EASZ240 antigen is secreted into the culture supernatant via the TTSS and that it is delivered into the cytoplasm of Int-407 cells by the TTSS. In vivo experiments indicate that both humoral and cell-mediated immune responses are induced in chickens vaccinated with a recombinant attenuated Salmonella serovar Typhimurium vaccine, which leads to significant protection against Eimeria challenge.

ABSTRACTCoccidiosis is a ubiquitous disease caused by intestinal protozoan parasites belonging to several distinct species of the genus Eimeria. Cell-mediated immunity (CMI) is critically important for protection against Eimeria; thus, our approach utilizes the bacterial type III secretion system (TTSS) to deliver an antigen directly into the cell cytoplasm of the immunized host and into the major histocompatibility complex class I antigen-processing pathway for induction of CMI and antigen-specific cytotoxic T-lymphocyte responses in particular. To accomplish this goal, Eimeriagenes encoding the sporozoite antigen EASZ240 and the merozoite antigen EAMZ250 were fused to the Salmonella entericaserovar Typhimurium effector protein gene sptPin the parental pYA3653 vector, yielding pYA3657 and pYA3658, respectively. SptP protein is secreted by the TTSS of Salmonellaand translocated into the cytoplasm of immunized host cells. The host strain chromosomal copy of the sptPgene was deleted and replaced by a reporter gene, xylE. The newly constructed vectors pYA3657 and pYA3658 were introduced into host strain χ8879 (ΔphoP233ΔsptP1033::xylEΔ asdA16). This strain is an attenuated derivative of the highly virulent strain UK-1. When strain χ8879(pYA3653) as the vector control and strain χ8879 harboring pYA3657 or pYA3658 were used to orally immunize day-of-hatch chicks, colonization of the bursa, spleen, and liver was observed, with peak titers 6 to 9 days postimmunization. In vitro experiments show that the EASZ240 antigen is secreted into the culture supernatant via the TTSS and that it is delivered into the cytoplasm of Int-407 cells by the TTSS. In vivo experiments indicate that both humoral and cell-mediated immune responses are induced in chickens vaccinated with a recombinant attenuated Salmonellaserovar Typhimurium vaccine, which leads to significant protection against Eimeriachallenge.

Butyrate is a feed additive that has been shown to have antibacterial properties and improve gut health in broilers. Here, we examined the performance and gene expression changes in the ileum of tributyrin–supplemented broilers infected with coccidia. Ninety-six, Ross 708 broilers were fed either a control corn–soybean–based diet (−BE) or a diet supplemented with 0.25% (w/w) tributyrin (+BE). Birds were further divided into groups that were inoculated with Eimeria maximaoocysts (EM) or sham-inoculated (C) on day 21 posthatch. At 7 d postinfection (7 d PI), the peak of pathology in E. maximainfection, tributyrin-supplemented birds had significantly improved feed conversion ratios (FCR, P< 0.05) and body weight gain (BWG, P< 0.05) compared with -BE-infected birds, despite both groups having similar feed intake (FI, P> 0.05). However, at 10 d post-infection (10 d PI) no significant effects of feed type or infection were observed. Gene expression in the ileum was examined for insights into possible effects of infection and tributyrin supplementation on genes encoding proteins related to immunity, digestion, and gut barrier integrity. Among immune-related genes examined, IL-1Band LEAP2were only significantly affected at 7 d PI. Transcription of genes related to digestion (APN, MCT1, FABP2, and MUC2) were primarily influenced by infection at 7 d PI and tributyrin supplementation (FABP2and MUC2) at 10 d PI. With exception of ZO1,tight junction genes were affected by either infection or feed type at 7 d PI. At 10 d PI, only CLDN1was not affected by either infection or feed type. Overall tributyrin shows promise as a supplement to improve performance during coccidiosis in broiler chickens; however, its effect on gene expression and mode of action requires further research.

Although water treatment processes to remove Cryptosporidium are improving and detection methods for identifying the parasite in water are becoming more sensitive, outbreaks of cryptosporidiosis continue in the human population. Animals, especially dairy calves, often become infected as C. parvum oocysts are present in high numbers, remain viable for long periods of time after excretion and no disinfectants, prophylactic or therapeutic reagents exist. Vaccination against C. parvum is being attempted in ruminants for the purpose of generating hyperimmune colostrum containing antibodies that may be effective in passive immunotherapy against cryptosporidiosis in the young. A number of recombinant C. parvum surface or internal antigens have been expressed by DNA-cloning technology. Immune colostrum specific for several recombinant C. parvum proteins have demonstrated efficacy in murine and ruminant models against cryptosporidiosis. The P23 and CP15 antigens appear to be the most promising candidates for vaccine development. Recent studies have demonstrated efficacy of the drug nitazoxanide against C. parvum infection in humans. In the near future, control of this parasitic disease in humans and animals will rely on a combination of passive immunotherapy and selective drug treatment.