Coccidiosis in chicken causes great economic losses. Increasing resistance of Eimeria species to anticoccidials has forced the search for alternative methods of control. The present study evaluates the anticoccidial activity of some anti-Eimeria tenella antibody fragments expressed in pea plants. Both in vitro and in vivo infection assays including indirect immunofluorescence, in vivo evaluation of antibody neutralization and cell culture invasion-inhibition assays were used to study the inhibitory effect of these antibody fragments on E. tenella sporozoites. Seven of nine antibody fragments (Ab1, Ab4, Ab5, Ab6, Ab7, Ab8 and Ab9) showed binding to sporozoites of E. tenella in an indirect immunofluorescence test. Only two antibodies (Ab4 and Ab5) cross reacted with sporozoites of E. maxima, E. acervulina and E. brunetti. The localization of specific fluorescence differed between species. Ab binding with sporozoites was seen in the area of both anterior and posterior refractile bodies in case of E. tenella, E. brunetti, and E. maxima but was only observed in the posterior refractile body in case of E. acervulina. No antibody binding was observed on merozoites. The suitability of antibody fragments to alter the infectivity of E. tenella sporozoites to Madin Darby Bovine Kidney cells (MDBK) was examined in vitro and the invasion-inhibition rates were quantified by flow cytometry. To assess the inhibitory effect on parasite reproduction, the in vivo antibody neutralization assay was done by retrograde infection of chicken with sporozoites previously incubated with antibody fragments. In vitro invasion rates were reduced by incubation with antibody fragments by approximately 24 to 45 %, with Ab6 and Ab7 showing the most distinct effect. However, proliferation rates (PR) of the respective MDBK cultures were also clearly reduced by 15 to 26 %. PR of MDBK cells treated with 1:1000, 1:100, 1:10 and undiluted mixed antibody fragments were reduced by 1%, 10%, 16%, and 26% with a reduction of invasion rates by 0%, 9%, 15% and 18%, respectively. Immune sera reduced the invasion rates by 16% to 70% and increased PR of the host cells. It appeared that the preparations of the antibody fragments contained compounds cytotoxic to MDBK cells and thus invasion inhibition could not be unequivocally evaluated in vitro. However, after incubation with antibody fragments sporozoites displayed a reduced ability to reproduce after intracloacal application to chicken (especially Ab1, Ab3, Ab5 and Ab9). Other antibody fragments (Ab2, Ab4, Ab6, Ab7 and Ab8) were less capable to reduce sporozoite infectivity and reproduction. More investigations are still required to study the possible use of antibody fragments and their application to infected chicken exposed to coccidiosis.

<p dir="ltr">For this dissertation, two experiments were conducted to evaluate the effects of a multienzyme mix and partially defatted black soldier fly larvae meal on chicken coccidiosis, focusing on growth performance, intestinal health, and microbiota dynamics. Experiment 1 examined the growth performance, nutrient utilization, microbiota modulations, and other gut health-related indicators of broiler chickens under coccidia challenge, with dietary supplementation of multienzyme, including phytase, xylanase, β-glucanase, amylase, hemicellulase, and pectinase. Ross 308 broilers were assigned to 4 treatments in a 2×2 factorial arrangement comprising of 0 or 50 g·kg-1 multienzyme and oral challenge with PBS or mixed Eimeria spp. oocysts (250,000 E. acervulina, 50,000 E. maxima, and 50,000 E. tenella). Multienzyme reduced (P < 0.05) Eimeria-induced loss in feed efficiency and nutrient utilization, partially explained by reduced decrease of b0,+ amino acid transporter in jejunum. Multienzyme suppressed (P < 0.05) the overexpression of interleukin-8 in the duodenum and jejunum and ameliorated (P = 0.05) the decreased expression of antioxidant heme oxygenase 1 in ileum induced by Eimeria infections. Multienzyme facilitated (P < 0.01) the bloom of short-chain fatty acid-producing and fiber-degrading microbes. The study concluded that multienzyme supplementation partially alleviated the adverse effects of Eimeria infections through various mechanisms, including enhanced nutrient utilization, reduced local inflammations, and restoration of microbial homeostasis. Experiment 2 investigated the growth dynamics, nutrient assimilation, and gut health responses of broiler chickens under coccidia challenge, with dietary supplementation of partially defatted black soldier fly larvae meal (pBSFLM) with increasing concentrations of 0, 60, 120 g/kg. During the infection phase (from d 13 to 19), interactions between Eimeria and pBSFLM revealed significant associations with gain to feed ratio (G:F) (P < 0.05) and cecal interferon-γ (IFN-γ, P < 0.05), while showing tendencies for crypt depth (P = 0.088) and cecal acetate concentration (P = 0.06). The incremental inclusion of pBSFLM demonstrated a negative effect on the G:F and the generation of IFN-γ and acetate in the ceca during coccidia challenge. Conversely in the non-challenged birds, the impact of dietary pBSFLM varied from neutral (e.g. G:F) to potentially advantageous (e.g. acetate). Challenged broilers exhibited decreased (P < 0.01) BW, feed intake (FI), G:F, as well as the apparent ileal digestibility (AID) and total tract nutrient utilization (ATTU) of DM, gross energy (GE), and nitrogen (N). Eimeria challenge led to reduced (P < 0.01) serum carotenoid concentrations, increased (P < 0.01) ileal crypt depth (CD), and an increase in the generation of branched-chain fatty acids, specifically isobutyrate (P = 0.059) and isovalerate (P < 0.05) in the ceca. Dietary pBSFLM addition caused a linear reduction (P < 0.05) in BW, FI, G:F, and N utilization. Furthermore, a tendency (P < 0.06) was observed where pBSFLM linearly decreased the villi height: CD ratio and reduced goblet cell density in the villi. Results from this experiment reveal that higher levels of pBSFLM supplementation, especially at 12%, had detrimental effects on growth, ileal morphology, cecal acetate production, and downregulated the expression of key cytokines in response to coccidia infection. In summary, these studies shed light on the multifaceted effects of dietary interventions on Eimeria infections in broiler chickens, with a specific emphasis on growth, nutrient utilization, and indicators of gut health.</p>

Coccidiosis in chicken causes great economic losses. Increasing resistance of Eimeria species to anticoccidials has forced the search for alternative methods of control. The present study evaluates the anticoccidial activity of some anti-Eimeria tenella antibody fragments expressed in pea plants. Both in vitro and in vivo infection assays including indirect immunofluorescence, in vivo evaluation of antibody neutralization and cell culture invasion-inhibition assays were used to study the inhibitory effect of these antibody fragments on E. tenella sporozoites. Seven of nine antibody fragments (Ab1, Ab4, Ab5, Ab6, Ab7, Ab8 and Ab9) showed binding to sporozoites of E. tenella in an indirect immunofluorescence test. Only two antibodies (Ab4 and Ab5) cross reacted with sporozoites of E. maxima, E. acervulina and E. brunetti. The localization of specific fluorescence differed between species. Ab binding with sporozoites was seen in the area of both anterior and posterior refractile bodies in case of E. tenella, E. brunetti, and E. maxima but was only observed in the posterior refractile body in case of E. acervulina. No antibody binding was observed on merozoites. The suitability of antibody fragments to alter the infectivity of E. tenella sporozoites to Madin Darby Bovine Kidney cells (MDBK) was examined in vitro and the invasion-inhibition rates were quantified by flow cytometry. To assess the inhibitory effect on parasite reproduction, the in vivo antibody neutralization assay was done by retrograde infection of chicken with sporozoites previously incubated with antibody fragments. In vitro invasion rates were reduced by incubation with antibody fragments by approximately 24 to 45 %, with Ab6 and Ab7 showing the most distinct effect. However, proliferation rates (PR) of the respective MDBK cultures were also clearly reduced by 15 to 26 %. PR of MDBK cells treated with 1:1000, 1:100, 1:10 and undiluted mixed antibody fragments were reduced by 1%, 10%, 16%, and 26% with a reduction of invasion rates by 0%, 9%, 15% and 18%, respectively. Immune sera reduced the invasion rates by 16% to 70% and increased PR of the host cells. It appeared that the preparations of the antibody fragments contained compounds cytotoxic to MDBK cells and thus invasion inhibition could not be unequivocally evaluated in vitro. However, after incubation with antibody fragments sporozoites displayed a reduced ability to reproduce after intracloacal application to chicken (especially Ab1, Ab3, Ab5 and Ab9). Other antibody fragments (Ab2, Ab4, Ab6, Ab7 and Ab8) were less capable to reduce sporozoite infectivity and reproduction. More investigations are still required to study the possible use of antibody fragments and their application to infected chicken exposed to coccidiosis.