Sporulated Oocysts Research Articles

Anticoccidial activities of Piper betle L essential oil on Eimeria tenella oocysts

Coccidiosis poses a significant threat to the poultry industry, with synthetic antibiotics and disinfectants being the primary tools for control. This study investigated the potential of Piper betle L essential oil (PBEO) as a natural alternative against Eimeria tenella, one of the most pathogenic Eimeria species affecting poultry. Our findings revealed that PBEO exhibits significant anticoccidial effects through two primary mechanisms: (i) oocysticidal activity by disintegrating oocyst walls and (ii) inhibition of the sporulation process. PBEO demonstrated oocysticidal activities ranging from 8.67 to 95.33% across concentrations from 0.04 to 40%. Notably, at 72 h post-incubation, a 0.04% PBEO concentration significantly reduced the number of sporulated oocysts (P ≤ 0.05) to 71.67%, showing effects comparable to those of formalin. PBEO reduced 50% of oocyst sporulation (IC50) in the concentration of 1.31% at 72 h. Gas chromatography-mass spectrometry (GC-MS) identified the primary constituents of PBEO, including eugenol, beta-caryophyllene, and other key compounds, collectively constituting 96% of the oil. This research underscores the potential of PBEO as a natural anticoccidial agent and lays the groundwork for further studies aimed at identifying, isolating, and developing active compounds that may specifically target the sporogony process in coccidian parasites.

Viability time of sporulated oocysts of bovine Eimeria spp. in water and efficacy of disinfectants in a region with tropical climate.

The water can be a significant risk factor for the occurrence of cattle eimeriosis on farms located in tropical climate regions. This study aimed to assess the viability period of sporulated oocysts of bovine Eimeria spp. in water containing organic matter and to evaluate the efficacy of disinfectants against sporulated oocysts of this protozoa. Two experiments were conducted, each comprising in vitro and in vivo evaluations. In Experiment 1, the viability period of oocysts sporulated in buckets containing a solution composed of a mixture of feces with oocysts, water, and potassium dichromate was assessed. Quantifications and identification of Eimeria spp. oocysts were performed on days + 30, + 60, + 90, and every 30days thereafter until no more oocysts were detected. Naïve calves were inoculated with this solution kept in the bucket for 6 and 12months. In Experiment 2, the efficacy of various disinfectants (acetic acid, sodium hypochlorite, ethanol + acetic acid, and ammonia quaternary) against a new solution of sporulated oocysts and pH changes over time (10', 30', and 24h) were evaluated in the laboratory. The most effective disinfectant was used to treat the water solution and inoculate the calves, and its efficacy was calculated. In Experiment 1, Eimeria spp. oocysts remained viable in the solution for up to 12months. E. zuernii persisted until the end of the study period. Calves inoculated with the solution kept in the buckets for 6 and 12months excreted at least four Eimeria species (E. zuernii, E. bovis, E. ellipsoidalis, and E. cylindrica). In Experiment 2, among the tested disinfectants, 3% acetic acid demonstrated the highest efficacy (10' = 80.9%; 30' = 87.9%; 24h = 91.7%). The pH values of the solutions containing acetic acid ranged from 2.4 to 3.5. Calves that received the inoculum treated with 3% acetic acid did not excrete Eimeria spp. oocysts in feces during the study period (efficacy = 100%). In contrast, all animals in the control group excreted Eimeria spp. oocysts in feces. Future studies should be carried out to better understand the possible effects of cattle drinking water with 3% acetic acid.

The Anticoccidial In Vitro Effects and Antioxidant Properties of Several Plants Traditionally Used for Coccidiosis in Togo.

Coccidiosis is a parasitic disease that often affects livestock. Identifying plants with inhibitory effects on the development of the parasite could help in finding new natural treatments. This study aimed to evaluate the anticoccidial potentials of extracts from Azadirachta indica leaves (AILs), Combretum micranthum leaves (CMLs), Carica papaya seeds (CPSs), Sarcocephalus latifolius roots (SLRs), and Vernonia amygdalina leaves (VALs). The in vitro anticoccidial efficacy of the extracts was evaluated through oocyst sporulation inhibition and sporozoite viability inhibition assays of Eimeria oocysts. The setup was examined for 72 h (every 24 h) of incubation. The DPPH radical scavenging activity and ferric reducing antioxidant power were used to evaluate the antioxidant potential of the extracts. Among the tested extracts, the SLR, CPS, and AIL extracts exhibited the maximum oocyst sporulation inhibition (75.85 ± 1.21%, 74.53 ± 1.65%, and 71.58 ± 0.24%, respectively) at a concentration of 75 mg/mL of plant extracts against the Eimeria species. The Sarcocephalus latifolius root extract showed the highest radical scavenging capacity (76.25 ± 0.53) and reducing power (86.21 ± 4.28). The biochemical screening of the selected plant extracts revealed the presence of antioxidant compounds such as phenols, flavonoids, alkaloids, saponins, and carbohydrates. The SLR extract contained the highest amounts of phenols (56.11 ± 0.33 µg/mL) and flavonoids (36.65 ± 1.85 µg/mL). In conclusion, the selected hydro-ethanolic extracts from these plants possess excellent anticoccidial and antioxidant activities, which can be attributed to the presence of medicinally important phytochemicals. Further research is needed to identify and isolate the active anticoccidial compounds from these plants, which could be utilized in the development of drugs against coccidiosis.

Trifolium pratense as a novel phytogenic supplement, is an anticoccidial agent in chickens

Coccidiosis, caused by a protozoan parasite of the genus Eimeria, is one of the most severe contagious parasite diseases affecting the poultry industry worldwide. Using phytogenics to prevent chicken coccidiosis is a strategy aimed at combating the increasing issue of drug-resistant strains of Eimeria spp. This study demonstrates the anticoccidial activities of a medicinal herb, Trifolium pratense (TP) powder, and its ethanolic extract (designated TPE) against Eimeria spp. TPE exhibited significant suppressive activity against E. maxima oocyst sporulation and E. tenella sporozoite invasion and reproduction in Madin-Darby bovine kidney cells. Furthermore, administration of basal chicken diets containing TP powder or TPE to Eimeria-infected chickens significantly reduced the output of oocysts and severity of intestinal lesions. Dietary supplementation with TP significantly improved relative weight gain in E. tenella- and E. acervulina-infected chickens, while there was no significant improvement in E. maxima-infected chickens. The anticoccidial activities of TP and TPE on E. acervulina, E. tenella and E. maxima were further supported by anticoccidial index scores, which showed greater efficacy than those of amprolium, a commercial coccidiostat used in poultry. TP supplementation positively impacted the primary metabolism of chickens challenged with E. tenella or E. acervulina. The chemical fingerprints of TPE were established using liquid column chromatography; TPE contained 4 major compounds: ononin, sissotrin, formononetin, and biochanin A. In addition, various spectrometric methods were used to ensure the batch-to-batch consistency of TP/TPE. In conclusion, T. pratense is demonstrated to be a novel phytogenic supplement that can be used to control Eimeria-induced coccidiosis in chickens.

Efeito do gás amônia em cama aviária contaminada com Eimeria spp.

Avian coccidiosis is a common disease that affects poultry worldwide. The practices used to control this pathology in poultry are based mainly on the use of antimicrobials, which have generated increased resistance of Eimeria and other microorganisms. Research is therefore needed to devise new control models and strategies. The objective of our research was to evaluate the effect of different concentrations of ammonia gas on poultry litter artificially contaminated with unsporulated Eimeria spp. oocysts. Samples of sterilized poultry litter were placed in containers artificially contaminated with unsporulated Eimeria spp. oocysts. Samples were placed in plastic bags and ammonia gas was injected in concentrations of 0.2%, 0.4%, 0.8%, 1% and 1.2%. The tested dosages of ammonia gas inhibited the sporulation of oocysts in poultry litter by 100%. Our study indicates that ammonia gas is efficient in eliminating Eimeria spp. in poultry litter at concentrations equal to or higher than 0.2% after one hour of exposure. These results are promising because, through the use of the bedding disinfection method (shallow fermentation with ammonia injection) it will be possible to eliminate field oocysts, improving the result of coccidiosis vaccines.

Genetic diversity and occurrence of Eimeria species causing cattle coccidiosis in Kashmir, India

This study focuses on the occurrence, identification, and molecular characterization of Eimeria species causing coccidiosis in cattle in the Kashmir Valley, India. Coccidiosis, caused by apicomplexan parasites of the genus Eimeria, poses a significant threat to global cattle farming. Conventional techniques for identification, which rely on the morphology of sporulated oocysts, have drawbacks, leading to the adoption of molecular techniques to accurately delimit species. A total of 190 cattle were sampled in nine farms and parasitological examination revealed an occurrence of 45.7% for Eimeria spp. Molecular analysis using PCR and sequencing identified three predominant species: E. zuernii, E. alabamensis, and E. bovis. The study highlights the widespread occurrence of these species globally, as supported by previous research conducted in Bangladesh, Austria, Egypt, and Brazil. The phylogenetic analysis based on internal transcribed spacer (ITS-1) gene sequences revealed distinct clusters for E. zuernii and E. bovis, while E. alabamensis formed a separate clade. The genetic diversity and phylogenetic connections provide insights into the evolutionary relationships among these Eimeria species. This study contributes valuable information for understanding the epidemiology and genetic diversity of cattle coccidiosis in the Kashmir Valley, emphasizing the importance of molecular characterization for accurate species identification.

Molecular characterization and functional analysis of Eimeria tenella ankyrin repeat-containing protein

Chicken coccidiosis causes disastrous losses to the poultry industry all over the world. Eimeria tenella is the most prevalent of these disease-causing species. Our former RNA-seq indicated that E. tenella ankyrin repeat-containing protein (EtANK) was expressed differently between drug-sensitive (DS) and drug-resistant strains. In this study, we cloned EtANK and analyzed its translational and transcriptional levels using quantitative real-time PCR (qPCR) and western blotting. The data showed that EtANK was significantly upregulated in diclazuril-resistant (DZR) strain and maduramicin-resistant (MRR) strain compared with the drug-sensitive (DS) strain. In addition, the transcription levels in the DZR strains isolated from the field were higher than in the DS strain. The translation levels of EtANK were higher in unsporulated oocysts (UO) than in sporozoites (SZ), sporulated oocysts (SO), or second-generation merozoites (SM), and the protein levels in SM were significantly higher than in UO, SO, and SZ. The results of the indirect immunofluorescence localization showed that the protein was distributed mainly at the anterior region of SZ and on the surface and in the cytoplasm of SM. The fluorescence intensity increased further with its development in vitro. An anti-rEtANK polyclonal antibody inhibited the invasive ability of E. tenella in DF-1 cells. These results showed that EtANK may be related to host cell invasion, required for the parasite’s growth in the host, and may be involved in the development of E. tenella resistance to some drugs.

Evaluation of the anticoccidial effect of jataí bee (Tetragonisca angustula) propolis extract against Eimeria spp. oocysts in broiler

The use of anticoccidial compounds is the primary strategy for controlling avian coccidiosis. The emergence of drug resistance, regulatory restrictions, and societal demands for meat production without the use of antimicrobials have increased the need for natural alternatives to control this disease. Propolis is known for its antimicrobial effects and may be an alternative. In this context, this study aimed to evaluate the anticoccidial activity of jataí bee (Tetragonisca angustula) propolis against the sporulation of Eimeria spp. oocysts in broiler chicken feces. Propolis ethanolic extracts were tested at 10, 15, and 20% against Eimeria oocyst sporulation. Direct use of the extract on oocysts completely inhibited sporulation. When applied to feces, the extract significantly reduced the number of sporulated oocysts. Thus, the jatai bee propolis extract is a promising alternative to commercial anticoccidial agents.

Comparative molecular analyses of Eimeria Schneider (Apicomplexa: Eimeriidae) species from rock ptarmigan in Iceland, Svalbard-Norway, and Japan.

The rock ptarmigan (Lagopus muta) has a Holarctic breeding distribution and is found in arctic and sub-arctic regions. Isolated populations and glacial relicts occur in alpine areas south of the main range, like the Pyrenees in Europe, the Pamir mountains in Central Asia, and the Japanese Alps. In recent decades considerable effort has been made to clarify parasite infections in the rock ptarmigan. Seven Eimeria spp. have been reported parasitizing rock ptarmigan. Two of those species, E. uekii and E. raichoi parasitizing rock ptarmigan (L. m. japonica) in Japan, have been identified genetically. Here we compare partial sequences of nuclear (18S rRNA) and mitochondrial (COI) genes and we detail the morphology of sporulated oocysts of E. uekii and E. raichoi from Japan, E. muta and E. rjupa, from the rock ptarmigan (L. m. islandorum) in Iceland, and two undescribed eimerian morphotypes, Eimeria sp. A, and Eimeria sp. B, from rock ptarmigan (L. m. hyperborea) in Norway (Svalbard in the Norwegian Archipelago). Two morphotypes, ellipsoidal and spheroidal, are recognized for each of the three host subspecies. Our phylogenetic analysis suggests that the ellipsoidal oocyst types, E. uekii, E. muta, and Eimeria sp. A (Svalbard-Norway) are identical and infects rock ptarmigan in Japan, Iceland, and Svalbard-Norway, respectively. Eimeria uekii was first described in Japan in 1981 so that E. muta, described in Iceland in 2007, and Eimeria sp. A in Svalbard-Norway are junior synonyms of E. uekii. Also, phylogenetic analysis shows that the spheroidal oocyst types, E. rjupa and Eimeria sp. B (Svalbard-Norway), are identical, indicating that rock ptarmigan in Iceland and Svalbard-Norway are infected by the same Eimeria species and differ from E. raichoi in Japan.

Comparative proteomic analysis across the developmental stages of the Eimeria tenella

Eimeria tenella is the main pathogen responsible for coccidiosis in chickens. The life cycle of E. tenella is, arguably, the least complex of all Coccidia, with only one host. However, it presents different developmental stages, either in the environment or in the host and either intracellular or extracellular. Its signaling and metabolic pathways change with its different developmental stages. Until now, little is known about the developmental regulation and transformation mechanisms of its life cycle. In this study, protein profiles from the five developmental stages, including unsporulated oocysts (USO), partially sporulated (7 h) oocysts (SO7h), sporulated oocysts (SO), sporozoites (S) and second-generation merozoites (M2), were harvested using the label-free quantitative proteomics approach. Then the differentially expressed proteins (DEPs) for these stages were identified. A total of 314, 432, 689, and 665 DEPs were identified from the comparison of SO7h vs USO, SO vs SO7h, S vs SO, and M2 vs S, respectively. By conducting weighted gene coexpression network analysis (WGCNA), six modules were dissected. Proteins in blue and brown modules were calculated to be significantly positively correlated with the E. tenella developmental stages of sporozoites (S) and second-generation merozoites (M2), respectively. In addition, hub proteins with high intra-module degree were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Gene and Genomes (KEGG) pathway enrichment analyses revealed that hub proteins in blue modules were involved in electron transport chain and oxidative phosphorylation. Hub proteins in the brown module were involved in RNA splicing. These findings provide new clues and ideas to enhance our fundamental understanding of the molecular mechanisms underlying parasite development.

Evaluation of anthelminthic and antiprotozoal activity of myrrh (Commiphora myrrha) methanolic extract.

To treat and control parasitic infections, traditional medical remedies using plant products are utilized as antiparasitic agents rather than standard synthetic chemicals due to drug resistance. Myrrh, a resinous exudate of Commiphora myrrha (Burseraceae), is a powerful antioxidant with a variety of medicinal uses. This study aimed to investigate the effect of the myrrh methanolic extract (MyE) of three concentrations (100, 50, and 25 mg/ml) on the sporulation of oocysts and as an anthelminthic effector via in vitro study. Characterization of the plant was done by Fourier-transform infrared spectroscopy (FT-IR). The earthworm, Eisenia fetida, is used as a model worm to evaluate the anthelminthic activity of MyE. Eimeria labbeana-like oocysts are used as a model protozoan parasite in anticoccidial assays. The sporulation and inhibition (%) of E. labbeana-like were assessed by MyE compared to other chemical substances. FT-IR revealed the presence of twelve active compounds. Our results showed that paralysis and death of earthworms at MyE (100 mg/ml) were 7.88 ± 0.37 and 9.24 ± 0.60 min, respectively, which is more potency when compared to mebendazole (reference drug). In all treated worms, microscopic examinations revealed obvious surface architecture abnormality. This study shows that MyE affects oocysts sporulation in a dose-dependent manner. At 24 and 36 hr, a high concentration of MyE (100 mg/ml) inhibits sporulation by 90.95 and 87.17 %. At 36 hr, other concentrations of MyE (50 and 25 mg/ml), as well as amprolium, DettolTM, and phenol inhibits oocyst sporulation by 40.17 %, 29.34 %, 45.09 %, 85.11 %, and 61.58 %, respectively. According to our research, the MyE extract had powerful anthelmintic and anticoccidial properties.

Metaphylactic strategies using toltrazuril against coccidiosis in goat kids

Goat coccidiosis compromises animal welfare, reduces productivity and may cause mortality and delayed growth rates in goat kids around the weaning period worldwide. This field study was conducted to evaluate the efficacy of metaphylactic treatments with two doses of toltrazuril (20 or 40 mg/kg body weight – BW, p. o.), at different timing, in kids naturally infected with Eimeria spp. A total of 97 healthy goat kids (Majorera milk aptitude breed) were divided into five groups, depending on the age of treatment (2 or 7 weeks). One group remained untreated as a negative control until the end of the study. Faecal oocyst shedding, faecal consistency, and body weight of the animals were monitored at day 0 and at weekly intervals. Counts of oocysts per gram of faeces (OPG) were determined by a modified McMaster technique. Morphometric identification of Eimeria species was carried out on individual faecal samples from each experimental group after oocyst sporulation. Goat kids treated at two weeks of age maintained OPG values close to zero during the 5 weeks post-treatment and, overall, had lower faecal oocyst counts than untreated control animals. No significant differences were observed between the two doses of toltrazuril used in two-week-old treated animals. By contrast, when treatment was carried out at seven weeks of age, the dose of 40 mg/kg BW of toltrazuril reduced oocyst levels for longer and to a greater extent than the 20 mg/kg dose. Irrespectively of the treatment and dose, toltrazuril delayed the appearance of pathogenic Eimeria species, i. e. Eimeria ninakohlyakimovae and Eimeria arloingi. As a whole, Eimeria christenseni, with a rather moderate pathogenicity, was highly predominant throughout the study period, including the untreated control group, which was probably the reason why clinical signs of coccidiosis were barely observed throughout the experiment. Under these circumstances, the positive effect of toltrazuril on body weight condition observed in some treated groups was difficult to correlate to the timing and doses. Metaphylactic treatments with 20 mg/kg BW toltrazuril given at two weeks of age are sufficient to control oocyst excretion in goat kids; whereas if administered later in 7-week-old animals, thereby coinciding with the frequently observed peak of oocyst elimination in goat kids under field conditions, a higher dose might be advisable to prevent environmental contamination with infectious oocysts.

Fighting eimeriosis by using the anti-eimerial and anti-apoptotic properties of rhatany root extract.

Over the last decade, extensive use of coccidiostats to treat and control Eimeria infection has developed drug resistance, prompting the search for new alternative therapies. Rhatany is proven to have various pharmacological properties. The present study aimed to in vitro and in vivo evaluate the effect of Rhatany roots extract (RRE) as an anti-eimerial and anti-apoptotic agent against murine eimeriosis induced by Eimeria papillata. Phytochemical screening by gas chromatography-mass spectrometry analysis (GC-MS) was used to detect active compounds in RRE. In vitro anti-eimerial activity of RRE (200, 100, 50 mg/ml), amprolium, phenol, Dettol™, and formalin were studied after incubation with non-sporulated Eimeria oocysts. For the in vivo study, twenty-five male C57BL/6 mice were randomly allocated into five groups. Animals in the first group were just given distilled H2O, while those in the second group were given 200 mg/kg RRE for 5 days. The Eimeria parasite's oocysts were infected into the third, fourth, and fifth groups. For treatment, RRE (200 mg/kg) and amprolium (120 mg/kg) were orally given to the 4th and 5th groups for five days, respectively. All mice were euthanized, on day 5 post-infection, to collect the jejunal tissues under study. Investigations were undertaken into the oocyst output in feces and goblet cells in mice jejuna. Assays for glutathione peroxidase (GPx), hydrogen peroxide (H2O2), and myeloperoxidase (MPO) were also performed. In jejunal tissue, cysteine aspartic acid protease-3 (Caspase-3) was counted using immunohistochemistry, while BCL2-associated X protein (Bax) and B-cell lymphoma-2 (BCL-2) were assayed using ELISA. In addition, mRNA expression of the goblet cell response gene (MUC2) was detected using real-time PCR. Phytochemical screening by GC-MS demonstrated the presence of 22 compounds in the RRE. The in vitro study revealed that RRE significantly inhabited the oocyst sporulation in a dose-dependent manner. By day 5 after infection with the Eimeria parasite, the number of oocysts in mice feces was significantly reduced after RRE treatment (1.308 × 106 ± 1.36 × 105 oocysts/g feces) compared to the infected group (5.387 × 106 ± 4.29 × 105 oocysts/g feces). Moreover, the Eimeria infection reduced the number of goblet cells of mice jejuna and its specific gene, MUC2. The treatment with RRE increased the number of goblet cells/villus from 3.45 ± 0.17 to 6.04 ± 0.23, associated with upregulation for MUC2 from 0.26 to 2.39-fold. Also, the Eimeria experimental infection lowered the activity of the antioxidant enzyme represented by GPx (23.99 ± 3.68 mg/g tissue), while increasing the stress parameters of hydrogen peroxide (0.07 ± 0.01 mM/g) as well as the activity of MPO (66.30 ± 3.74 U/mg). The production of apoptotic markers including Caspase-3 (68.89 ± 2.67 U/g) and Bax (159.05 ± 6.50 pg/ml) was significantly elevated while decreasing the anti-apoptotic marker of BCL2 (0.42 ± 0.07 pg/ml). Our study proved that RRE significantly reduced oxidative stress, and apoptotic markers as well as the inflammatory activity of MPO. Also, antioxidant enzyme and anti-apoptotic activity in the jejunum of E. papillata-infected mice were enhanced after RRE treatment. Our study highlights the potential of RRE as a natural solution for coccidiosis management by modulating apoptosis in E. papillata host cells. However, further research is needed to fully understand the underlying mechanisms and enhance our understanding of its therapeutic efficacy.

A questionnaire-based survey in Spain provides relevant information to improve the control of ovine coccidiosis.

Ovine coccidiosis is a widespread intestinal parasitic disease caused by Eimeria spp. Lambs are infected by the ingestion of sporulated oocysts, experiencing diarrhea and low growth rates. Control should be based on measures to reduce infection pressure and stress on the animals as well as on appropriate diagnosis and strategic treatment. To obtain information on how control measures are implemented in the ovine sector in Spain, a questionnaire-based survey was completed in 2022 by 154 veterinarians and 173 farmers working in this sector. Coccidiosis was highlighted as a relevant disease by 34% of the respondents. The period of greatest risk seemed to differ between production systems, being mainly early after weaning (7-15 days after weaning) in meat flocks and feedlots and later (1-2 months after weaning) in dairy flocks. The absence of cleaning and disinfection measures was identified as a risk factor by 51% of the veterinarians, with 22% mentioning overcrowding of animals and 22% indicating that coccidiosis has more incidence in flocks with large number of animals. The use of laboratory diagnosis methods (fecal oocyst count) was unusual in 70 and 84% of the veterinarians and farmers, respectively. Regarding control, dairy flocks usually housed a larger number of animals under intensive conditions, and they implemented more frequently control measures for coccidiosis than meat flocks. Anticoccidial drugs were used in 79% of the flocks, and in 74-82% of them, they were applied based on clinical criteria. Comparing protocols for anticoccidial treatment among different production systems, in meat flocks, anticoccidial drugs were applied more frequently when clinical signs were observed, and coccidiostats were used for less than 28 days compared to dairy flocks. These results highlight the need for improvement in the use of anticoccidial treatments adjusted to the new regulatory framework in the EU, which in turn will rationalize the use of antimicrobial compounds and may help to mitigate the impact of coccidiosis in flocks.

Proteomic Analysis of Fractionated Eimeria tenella Sporulated Oocysts Reveals Involvement in Oocyst Wall Formation.

Eimeria tenella is the most pathogenic intracellular protozoan parasite of the Eimeria species. Eimeria oocyst wall biogenesis appears to play a central role in oocyst transmission. Proteome profiling offers insights into the mechanisms governing the molecular basis of oocyst wall formation and identifies targets for blocking parasite transmission. Tandem mass tags (TMT)-labeled quantitative proteomics was used to analyze the oocyst wall and sporocysts of E. tenella. A combined total of 2865 E. tenella proteins were identified in the oocyst wall and sporocyst fractions; among these, 401 DEPs were identified, of which 211 were upregulated and 190 were downregulated. The 211 up-regulated DEPs were involved in various biological processes, including DNA replication, fatty acid metabolism and biosynthesis, glutathione metabolism, and propanoate metabolism. Among these proteins, several are of interest for their likely role in oocyst wall formation, including two tyrosine-rich gametocyte proteins (EtGAM56, EtSWP1) and two cysteine-rich proteins (EtOWP2, EtOWP6). Concurrently, 96 uncharacterized proteins may also participate in oocyst wall formation. The present study significantly expands our knowledge of the proteome of the oocyst wall of E. tenella, thereby providing a theoretical basis for further understanding of the biosynthesis and resilience of the E. tenella oocyst wall.

A Comparison in Terms of Prevalence, Morphological and Histopathological Changes between Eimeria spp. that Infect Local Rabbits (Oryctolagus cuniculus) in Some Regions of Saudi Arabia

Background: The epidemiology of parasitic infections is common in rabbit flocks and poses a significant threat to public health. And the most famous and influential of these causes are coccidia parasites, Coccidiosis in rabbits is an infection caused by intracellular protozoan parasites belonging to Eimeria, which is considered a major cause of significant morbidity and mortality. Methods: A total of 210 fresh feces samples of adult domestic rabbits (Oryctolagus cuniculus) were collected from Riyadh City, Al Kharj and ADilam in Saudi Arabia to examine the incidence of the Eimeria infection of microscopically. Feces concentrated by the floatation technique. After that, freshly collected non-sporulated oocysts were suspended in 2.5% (W/V) potassium dichromate, placed in Petri dishes in the air, and allowed to sporulate at room temperature (25±3oC). The morphometric and specific characteristics of sporulated oocysts were studied by an Olympus microscope (BX51ggTF, OLYMPUS, Tokyo, Japan) using the oil immersion lens. Specimens were taken from all parts intestine and liver, and the tissues were kept in 10% formalin immediately and make tissue sections and that to study the histopathological change under light microscopic. Result: An investigation was conducted in three cities in the Riyadh area (Riyadh, Al Kharj and Adilam) to determine the incidence of Eimeria species that parasitize local rabbits (Oryctolagus cuniculus). The overall frequency of these coccidial illnesses in rabbits in Riyadh City was 45.7%, in Adilam City it was 44.2% and in Al Kharj City it was 37.1%, respectively. There were five different species of Eimeria: E. magna, E. flavescens, E. exigua, E. striedae and E. coecicola, respectively. More than one species of Eimeria oocyst was obtained in the same examined sample. single-infection 13/90 (14.4%), dual-infection 39/90 (43.3%) and triple-infection 38/90 (42.2%). E. magna; E. flavescens; E. exigua; E. stiedae and E. coecicola, respectively. Significant differences (p less than 0.05) in prevalence were observed between the rabbits. Eimeria infection that affects the liver was confirmed by histopathological changes that were noted through the development of parasitic stages in the jejunum, ileum, cecum, end cecum and liver. It is possible to conclude that the current research offers pertinent information that aids in determining potential infection and future preventative measures against rabbit coccidiosis to lessen financial losses in the rabbit industry.

Methods of coprological diagnostics of animal parasitoses

The purpose of the research is to review the classical methods of coprological diagnostics, taking into account modern recommendations and proposals. Materials and methods. Literature sources on the issues of coprological methods of diagnosing parasitosis have been studied. Results and discussion. Diagnosis during life is an important element of practical veterinary activity, the success of which largely depends on the correct choice and competent use of suitable laboratory methods. For the diagnosis of parasitic diseases, the study of feces is most often used. Coprological methods are highly informative due to the fact that they can detect not only parasites of the digestive tract, liver, pancreas, but also a number of helminths of the respiratory system and even some tissue parasites that excrete eggs or larvae with feces. Unlike existing indirect immunological and molecular genetic methods, classical research methods are based on the detection and identification of the pathogens themselves at different stages of development. At the same time, the researcher has the opportunity to study morphological features and conduct morphometric analysis. The widespread use of classical coprological methods is also associated with their availability, they are quite simple to perform and relatively inexpensive. A limitation of coprological methods is the inability to diagnose pathogens that are at a non-propagative stage. The effectiveness of diagnostics is influenced by the method of collecting samples, the terms and conditions of transportation, and the choice of an adequate method for examining samples. The article presents the rules for sampling and options for the use of fixatives, a classification of coprological methods is given, and the procedure for performing research is described. Special attention is paid to the choice of flotation solutions, the method of cultivation of strongylid larvae and the method of sporulation of coccidia oocysts. Attention is drawn to the importance of micrometric measurements of detected objects. To ensure the quality of the conducted coprological studies, it is necessary to control the stages of the studies, the technical condition of the equipment and train the personnel. The material presented in the article is intended for both students and practitioners.

Atividade antiparasitária in vitro do extrato etanólico das folhas de Anethum graveolens

ABSTRACT Natural products are safe environmentally friendly agents and have no negative impact on the environment, they can be used to combat parasitic diseases. Helminthiasis and coccidiosis are parasitic diseases that harm both health and the economy. This research aimed to see how Anethum graveolens leaves extract (AGLE) worked as an anti-parasitic modulator during oocyst sporulation of an Eimeria papillata infection. FT-IR phytochemical analysis revealed the presence of eight compounds. The time required to induce paralysis and death in worms at the highest concentration (200 mg/mL) was 4.57±0.26 and 5.22±0.10 min, respectively. In an in vitro study, AGLE (300 mg/ml) inhibited sporulation by approximately 100% after 72 and 96 hr. AGLE (200, 100, and 50 mg/ml), amprolium, DettolTM, and phenol induced variable inhibition levels at 96 hr of 5.54%, 1.01%, 37.33%, 81.33%, and 89.33%, respectively. Our findings suggest that AGLE has potent anthelmintic and anticoccidial properties that could be further developed into a novel therapeutic agent.

Relationship between Eimeria oocyst infectivity for chickens and in vitro excystation of E. acervulina, E. maxima, and E. tenella oocyst during long-term storage

Vaccination of chickens against avian coccidiosis in chickens often involves storing Eimeria oocysts for months after oocyst propagation and sporulation. The purpose of this study was to determine how long E. acervulina, E. maxima, and E. tenella oocysts remained viable when stored at refrigeration (4°C) or egg room (20°C) temperatures. Separate tubes containing E. acervulina, E. maxima, or E. tenella oocysts 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 Eimeria species at each time-temperature combination was subjected to in vitro excystation to quantify the relative number of released sporozoites to intact (nonexcysted) sporocysts. Eimeria tenella appeared to be most susceptible to storage in that no oocyst production was observed at 9 mo at either temperature. Although E. maxima oocysts were viable at 9 mo, no oocyst production was observed at 12 mo storage at these 2 temperatures. Quite unexpected was that E. acervulina was much more stable than E. tenella and E. maxima remaining 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. acervulina r=0.58, r=0.54; E. maxima r=0.90, r=0.54; E. tenella r=0.38, r=0.90). These data indicate that attention must be paid to time and temperature of Eimeria oocyst storage, and that sporozoite excystation may not be a good indicator of oocyst viability, particularly at later timepoints in incubation.

Administration of Ethanolic Extract of Spinacia oleracea Rich in Omega-3 Improves Oxidative Stress and Goblet Cells in Broiler Chickens Infected with Eimeria tenella.

This study investigated the anticoccidial activity of spinach (Spinacia oleracea) whole-plant extract against Eimeria tenella, both in vitro and in vivo. For this purpose, one hundred 8-day-old broiler chicks of both sexes were divided into four groups (n = 25 in each group). Chicks in the first group served as the negative control (non-treated-non-infected). Chicks in the second group were challenged at 18 days old with 5 × 104E. tenella sporulated oocysts. The third group was challenged with 5 × 104 sporulated E. tenella oocysts at 18 days old after receiving spinach extract at a dose of 50 mg/kg at 8 days old. The fourth group received 0.2 mg/kg diclazuril (Coxiril® 0.2%) in their diet two days before being orally infected with 5 × 104 sporulated E. tenella oocysts and this continued till day 10 post-infection (PI). The growth performance, clinical symptoms, oocyst shedding, histological findings, and biochemical parameters were used to evaluate the efficacy on day 8 PI when the infection was at its peak. A gas chromatography examination revealed that omega-3 fatty acids were the main constituents of the spinach extract, followed by oleic acid, palmitic acid, and phytol, with amounts of 23.37%, 17.53%, 11.26%, and 7.97%, respectively. The in vitro investigation revealed that the spinach extract at concentrations of 10% and 5% inhibited the oocyst sporulation by 52.1% and 45.1%, respectively. The 5% concentration was selected for the in vivo trial based on the results of the in vitro study. The infected-untreated group showed high levels of OPG; lower body weight; a greater number of parasite stages; few goblet cells; decreased SOD, CAT, and GPX levels; and increased MDA and NO levels. The spinach-treated group, on the other hand, showed a significant decrease in oocyst output per gram of feces (OPG), increased body weight, decreased parasitic stages, and a nearly normal number of goblet cells. Additionally, it reduced malondialdehyde (MDA) and nitric oxide (NO), while increasing superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX). In conclusion, spinach produced significant antioxidant effects, increased body weight, reduced the number of oocysts and parasite stages in the caecum, and restored the number of goblet cells relative to those of an uninfected control. Furthermore, spinach extract inhibits the sporulation percentage of E. tenella oocysts. The ethanolic extract of S. oleracea (whole plant) contained high concentrations of fatty acids, palmitic acid, Phytol, betulin, and ursolic aldehyde, all of which are known to regulate the antioxidant pathway and modulate inflammatory processes and may be the main reason for its anticoccidial activity.