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This review of literature analyses the effectiveness of three commercially available filters, namely the Pall Envirochek filters, IDEXX FiltaMax, and Whatman flatbed membrane, in the process of concentrating Cryptosporidium oocysts and Giardia cysts from water samples. Moreover, several research studies investigated the possibility of using ultrafiltration filters as an affordable substitute for pathogen concentration, and their efficacy for protozoa is reviewed here and compared against the existing commercial filter options. Because of the differences in sampling procedures, no one method for concentrating these protozoa consistently outperforms others. This comparative article summarises our current understanding of the recent water sampling techniques to concentrate waterborne protozoans and highlights the remaining knowledge gaps. While ultrafiltration is a financially practical alternative for the concentration of pathogens, this paper highlights that further investigation is required to optimise their efficacy, particularly for low numbers of oocysts and cysts. These insights play a substantial role in providing information for advancing enhanced water sampling and treatment systems, reducing the dangers connected with waterborne protozoans, and ensuring the protection of public health. [ABSTRACT FROM AUTHOR]

In this article, a novel model for the dynamics of toxoplasmosis in human and cat populations with vertical transmission and contribution of oocysts to the environment from the mildly infected cats is constructed. The non-negative properties of the model’s solutions are proved. We demonstrate that a secondary quantity that affects the overall dynamics of T. gondii in human and cat populations is the reproductive ratio R◦. The impact of the contribution of oocysts from the mildly infected cats as well as the impact of vertical transmission and the impact of effective contact between cat and cat and cat and humans on the reproductive ratio are shown. The model’s endemic and disease-free equilibria are derived, and their local and global stabilities are proved. The bifurcation and sensitivity of the model’s parameters to T. gondii dynamics are studied. Finally, simulations are performed with the aid of the computer-in-built Runge-Kutta package implemented in the software Maple to illustrate the behavior of the model graphically. The results indicate that vertical transmission, contact with the infected cats and contribution of T. gondii from the mildly infected cats have a significant impact on the dynamics of toxoplasmosis. [ABSTRACT FROM AUTHOR]

Background: Coccidia are a group of intracellular protozoal parasites within the phylum Apicomplexa. Eimeria tenella, one of the species that cause intestinal coccidiosis in poultry, can cause significant mortality and morbidity. Diploid oocysts of Eimeria species are shed in the feces of an infected host and must sporulate to achieve infectivity. This process results in eight haploid infectious units, called sporozoites, held within a single oocyst. Each Eimeria spp. parasite possesses a single apicoplast and a single mitochondrion, both of which carry multiple copies of their respective organellar genomes. Reports of copy numbers of organellar genomes have varied widely. Methods: We report the application of quantitative polymerase chain reaction (qPCR), supported by next-generation sequencing, for the quantification of the extranuclear genomes relative to the nuclear genome over the course of sporulation and following its completion. Results: At 64 elapsed hours, 93.0% of oocysts were fully sporulated; no increase in percent sporulation was observed after this time. Apicoplast relative genome copy number showed several significant shifts up to 72 elapsed hours, after which no significant shifts were observed. Oocysts were shed with approximately 60% the amount of apicoplast DNA present at 72 h, after which point no significant shifts in apicoplast genome relative abundance occurred. Mitogenome relative copy number showed only two significant shifts, from 16 to 24 elapsed hours and from 24 to 32 elapsed hours. Oocysts were shed with approximately 28% the amount of mitochondrial DNA that was present at the time sporulation was deemed morphologically complete, at 64 elapsed hours. Conclusions: The characterization of the dynamics of genome abundance in exogenous stages sheds new light on the basic biology of Eimeria spp. and supports the use of extranuclear targets for molecular modes of parasite quantification and identification with improved sensitivity and accuracy. [ABSTRACT FROM AUTHOR]

Background: Thioester-containing protein 1 (TEP1) is a crucial component of mosquitoes' natural resistance to parasites. To effectively combat malaria, there is a need to better understand how TEP1 polymorphism affects phenotypic traits during infections. Therefore, the purpose of this study was to determine the Tep1 genotype frequency in malaria vector populations from south-western Ethiopia and investigate its effect on Plasmodium oocyst development in Anopheles arabiensis populations. Methods: Using standard dippers, Anopheles mosquito larvae were collected from aquatic habitats in Asendabo, Arjo Dedessa, and Gambella in 2019 and 2020. Collected larvae were reared to adults and identified morphologically. Female An. gambiae s.l. were allowed to feed on infected blood containing the same number of gametocytes obtained from P. falciparum and P. vivax gametocyte-positive individuals using indirect membrane feeding methods. Polymerase Chain Reaction (PCR) was used to identify An. gambiae s.l. sibling species. Three hundred thirty An. gambiae s.l. were genotyped using Restricted Fragment Length Polymorphism (RFLP) PCR and sub samples were sequenced to validate the TEP1 genotyping. Results: Among the 330 samples genotyped, two TEP1 alleles, TEP1*S1 (82% frequency) and TEP1*R1 (18% frequency), were identified. Three equivalent genotypes, TEP1*S1/S1, TEP1*R1/R1, and TEP1*S1/R1, had mean frequencies of 65.15%, 2.12%, and 32.73%, respectively. The nucleotide diversity was ranging from 0.36554 to 0. 46751 while haplotype diversity ranged from 0.48871 to 0.63161, across all loci. All sample sites had positive Tajima's D and Fu's Fs values. There was a significant difference in the TEP1 allele frequency and genotype frequency among mosquito populations (p < 0.05), except populations of Anopheles arabiensis from Asendabo and Gambella (p > 0.05). In addition, mosquitoes with the TEP1 *RR genotype were susceptible and produced fewer Plasmodium oocysts than mosquitoes with the TEP1 *SR and TEP1 *SS genotypes. Conclusion: The alleles identified in populations of An. arabiensis were TEP1*R1 and TEP1*S1. There was no significant variation in TEP1*R1 allele frequency between the high and low transmission areas. Furthermore, An. arabiensis carrying the TEP1*R1 allele was susceptible to Plasmodium infection. Further studies on vector-parasite interactions, particularly on the TEP1 gene, are required for vector control techniques. [ABSTRACT FROM AUTHOR]

Background: Study of malaria parasite cell biology is challenged by their small size, which can make visualisation of individual organelles difficult or impossible using conventional light microscopy. In recent years, the field has attempted to overcome this challenge through the application of ultrastructure expansion microscopy (U-ExM), which physically expands a biological sample approximately 4.5-fold. To date, U-ExM has mostly been used to visualise blood-stage parasites and used exclusively on parasites in vitro. Methods: Here we develop Mosquito Tissue U-ExM (MoTissU-ExM), a method for preparing dissected mosquito salivary glands and midguts by U-ExM. MoTissU-ExM preserves both host and parasite ultrastructure, enabling visualisation of oocysts and sporozoites in situ. We also provide a point-by-point protocol for how to perform MoTissU-ExM. Results: We validate that MoTissU-ExM samples expand as expected, provide a direct comparison of the same dissected tissues before and after MoTissU-ExM, and highlight some of the key host and parasite structures that can be visualised following MoTissU-ExM. Discussion: We discuss potential use cases for MoTissU-ExM for study of malaria parasite biology, and more broadly. We detail drawbacks or challenges MoTissU-ExM and imaging these expanded tissues, along with information troubleshooting this technique. Finally, we discuss how MoTissU-ExM could be applied and adapted in future to increase its utility. [ABSTRACT FROM AUTHOR]

Background: Rabbits are the most important protein sources of animals for humans all over the world. Its production has increased significantly in recent years and it has become one of the most important animal sources in the Kingdom of Saudi Arabia. Coccidiosis is one of the most dangerous diseases affecting rabbits and other animals causing global economic losses. Plant extracts have been used as potential alternatives to chemotherapy because they do not have the negative consequences of tissue deposits and drug resistance. Methods: The current study examined the antioxidant activity of Vitis vinifera leaf extract (VVLE) in New Zealand rabbits infected with Eimeria stiedae. The extract was subjected to phytochemical analysis by FT-IR. A total of 35 male New Zealand rabbits were divided into seven groups. Group 1 served as the uninfected, untreated (negative control) group. Group 2 uninfected and treated at 400 mg/kg of extract to test for toxicity. Group 3 represented the infected, non-treated (positive control) group. While, groups 4, 5 and 6 comprised infected treated with at 100 mg/kg, 200 mg/kg and 400 mg/kg of extracts, respectively. Finally, group 7 consisted of infected rabbits treated with Toltrazuril at a dosage of 5 mg/kg body weight. Result: The analysis of phytochemicals by FT-IR for Vitis vinifera leaf extract revealed the existence of 15 biologically active compounds. VVLE was able to decrease the induced weight loss due to infection. Moreover, the oxidative status due to E. stiedae infection had been changed after treatment with VVLE where the level of glutathione, malondialdehyde and nitric oxide was improved indicating the antioxidant activity of the VVLE. Our findings suggested that VVLE could boost the induced oxidative stress in the liver of infected New Zealand rabbits, but studies are needed to determine the mechanism of VVLE. [ABSTRACT FROM AUTHOR]

[Display omitted] • Toxoplasma gondii DNA was detected in bivalves from the Kerguelen and Galapagos archipelagos. • Its occurrence differed between sampling sites in Kerguelen but not in Galapagos. • In both archipelagos, quantitative PCR-positive bivalves were found on islands without cats. • Our finding demonstrated the waterborne transport of oocysts over at least 20 km. • Exposure to currents and giant kelp density affect T. gondii detection in bivalves. Oocysts of the protozoan Toxoplasma gondii are found in felid feces and can be washed into coastal waters, where they persist for months, attaching to algae and accumulating in invertebrates. We used wild bivalves to assess contamination of coastal waters of the Kerguelen and Galapagos archipelagos by this zoonotic parasite. Additionally, we leveraged the contrasting situations of these archipelagos to identify some potential drivers of contamination. In the Galapagos, with a cat density reaching 142 per km2, 15.38% of the sampled oysters (Saccostrea palmula) tested positive for T. gondii by quantitative real-time PCR (qPCR) (n = 260), and positive samples were found in all eight sampling sites. In Kerguelen, with 1-3 cats per km2, 40.83% of 120 tested mussels (Mytilus edulis platensis) were positive, and positive samples were found in four out of the five sampling sites. These findings provide evidence of T. gondii contamination in the coastal waters of these archipelagos. Furthermore, T. gondii -positive bivalves were found on islands located 20 km away (Galapagos) and 5 km away (Kerguelen) from the nearest cat population, indicating that T. gondii oocysts can disperse through waterborne mechanisms over several kilometers from their initial deposition site. In the Galapagos, where runoff is infrequent and all sites are exposed to currents, the prevalence of qPCR-positive bivalves did not show significant variations between sites (p = 0.107). In Kerguelen where runoff is frequent and site exposure variable, the prevalence varied significantly (p < 0.001). The detection of T. gondii in Kerguelen mussels was significantly correlated with the site exposure to currents (odds ratio (OR) 60.2, p < 0.001) and the on-site density of giant kelp forests (OR 2.624, p < 0.001). This suggests that bivalves can be contaminated not only by oocysts transported by currents but also by consuming marine aggregates containing oocysts that tend to form in kelp forests. [ABSTRACT FROM AUTHOR]

The extraction of DNA from Cryptosporidium oocysts is challenging due to the robust oocyst wall. Nanoparticles have been applied to disinfect Cryptosporidium oocysts; here, we demonstrate the utilisation of nanoparticles to disrupt the oocyst wall to enable sporozoite lysis and detection via PCR. Both silver and zinc oxide nanoparticles are investigated under different conditions and compared to existing techniques. Zinc oxide nanoparticles are shown to be as effective as freeze–thaw methods, suggesting that a nanoparticle lysis approach offers a viable alternative to existing methods. [ABSTRACT FROM AUTHOR]

Simple Summary: Natural products offer significant potential in the fight against parasitic diseases. Coccidiosis is a disease caused by the Eimeria genus of parasites that infect many animals all over the world. The development of resistance to anticoccidial medications has refocused attention on finding new compounds for alternative therapeutic approaches. The purpose of this study was to determine the efficacy of Holothuria polii extract (HpE) in treating eimeriosis in mice. The results indicated that animals treated with 200 mg/kg of HpE showed a significant decrease in oocyst count and higher expression of the MUC2 gene. In conclusion, HpE functions as an anticoccidial substance. Eimeria is a parasite that causes coccidiosis in a variety of animals, leading to nutritional imbalances, issues with food digestion and absorption, low weight, and intestinal inflammation of varying degrees in its hosts. Anticoccidial medication resistance has become a significant obstacle to disease control efforts, leading to a renewed focus on discovering novel chemicals for alternative therapeutic approaches. The purpose of this study was to determine the efficacy of Holothuria polii extract (HpE) in treating eimeriosis in mice. Five groups of mice were studied, with E. papillata sporulated oocysts (1 × 103) used to infect three groups experimentally. The first group was treated with 200 mg/kg of HpE, the second group with Amprolium (120 mg/kg), and the third group was left untreated as a control. The fourth and fifth groups were uninfected, with one of them treated with 200 mg/kg of HpE and the other used as a negative control. The results revealed that HpE has 10 phytochemical compounds according to GC-MS analysis. Treatment with HpE resulted in a substantial decrease in oocyst output in feces. Also, the parasitic stages showed morphometric alterations, with reductions in the measurements compared with the infected, untreated mice. Moreover, expression of the mucin gene (MUC2) was higher in treated mice compared to infected ones, with significant increases in goblet cell numbers, which provided additional evidence for the activity of HpE as an anticoccidial product. In conclusion, there are a wide variety of natural products and many have been employed in folk medicine for treating a range of parasitic diseases. [ABSTRACT FROM AUTHOR]

Eimeria stiedae is one of the few eukaryotic pathogens that exclusively infect the liver and serves as a good model to study the host–pathogen interactions in this vital organ. In this study, we show that rabbits infected with E. stiedae develop severe but self‐healing cholangitis. RNA‐seq analysis of the liver gene expression landscapes over the long course of E. stiedae infection identified 912 differentially expressed genes (DEGs) in the prepatent period (794 up‐ and 118 downregulated genes), 2889 DEGs in the early oocyst shedding period (1870 up‐ and 1019 downregulated genes), 2859 DEGs in the peak oocyst shedding period (1923 up‐ and 936 downregulated genes), and 327 DEGs in the recovery period (164 up‐ and 163 downregulated genes). Combined with pathological observations, we identified dynamic changes in host–parasite interactions involving multiple pathways. They showed that E. stiedae infection induced full‐blown inflammatory, Th1 and Th17 immune responses at all time points. This was associated with the strong innate immune responses during the prepatent period, including increased Toll‐like and NOD‐like receptor signaling. Despite mounting several damage control and repair responses, such as PI3K‐Akt signaling, Ras signaling, and extracellular matrix‐receptor interactions, the liver underwent severe metabolic dysfunction, oxidative damage, and coagulopathy after patency and at peak infection, possibly as a result of suppressed peroxisome activities and downregulated PPAR signaling. These responses largely disappeared during late infection, suggesting that the liver self‐heals after severe cholangitis. These data provide new insights into host–pathogen interactions during Eimeria infection and improve our understanding of the pathogenesis of parasitic cholangitis. [ABSTRACT FROM AUTHOR]

Background: Transmission-blocking vaccines (TBVs) can effectively prevent the community's spread of malaria by targeting the antigens of mosquito sexual stage parasites. At present, only a few candidate antigens have demonstrated transmission-blocking activity (TBA) potential in P. vivax. Quiescin-sulfhydryl oxidase (QSOX) is a sexual stage protein in the rodent malaria parasite Plasmodium berghei and is associated with a critical role in protein folding by introducing disulfides into unfolded reduced proteins. Here, we reported the immunogenicity and transmission-blocking potency of the PvQSOX in P. vivax. Methods and findings: The full-length recombinant PvQSOX protein (rPvQSOX) was expressed in the Escherichia coli expression system. The anti-rPvQSOX antibodies were generated following immunization with the rPvQSOX in rabbits. A parasite integration of the pvqsox gene into the P. berghei pbqsox gene knockout genome was developed to express full-length PvQSOX protein in P. berghei (Pv-Tr-PbQSOX). In western blot, the anti-rPvQSOX antibodies recognized the native PvQSOX protein expressed in transgenic P. berghei gametocyte and ookinete. In indirect immunofluorescence assays, the fluorescence signal was detected in the sexual stages, including gametocyte, gamete, zygote, and ookinete. Anti-rPvQSOX IgGs obviously inhibited the ookinetes and oocysts development both in vivo and in vitro using transgenic parasites. Direct membrane feeding assays of anti-rPvQSOX antibodies were conducted using four field P. vivax isolates (named isolates #1-4) in Thailand. Oocyst density in mosquitoes was significantly reduced by 32.00, 85.96, 43.52, and 66.03% with rabbit anti-rPvQSOX antibodies, respectively. The antirPvQSOX antibodies also showed a modest reduction of infection prevalence by 15, 15, 20, and 22.22%, respectively, as compared to the control, while the effect was insignificant. The variation in the DMFA results may be unrelated to the genetic polymorphisms. Compared to the P.vivax Salvador (Sal) I strain sequences, the pvqsox in isolate #1 showed no amino acid substitution, whereas isolates #2, #3, and #4 all had the M361I substitution. Conclusions: Our results suggest that PvQSOX could serve as a potential P. vivax TBVs candidate, which warrants further evaluation and optimization. [ABSTRACT FROM AUTHOR]

Twenty-four treated municipal wastewater samples were collected during one-year study period and examined for the presence of helminthes eggs, Cryptosporidium oocysts, and Giardia cysts to compare between their occurrences. Modified Bailenger method was used in this study to detect and enumerate nematode eggs in the treated wastewater samples, while polymerase chain reaction (PCR) technique was used to detect Cryptosporidium and Giardia in the same treated wastewater samples. Results revealed that Ascaris eggs were detected in only 8.33% out of 24 treated wastewater samples, while Cryptosporidium oocysts and Giardia cysts were detected in 20.83 and 12.5% out of the same 24 treated wastewater samples, respectively. The public's health is at risk since treated wastewater used for agriculture still contains Giardia cysts and Cryptosporidium oocysts, even if it satisfies the nematode criteria. [ABSTRACT FROM AUTHOR]

Infections by gastrointestinal parasites are found in a variety of animals worldwide. For the diagnosis of such infections, the flotation method is commonly used to detect parasitic microorganisms, such as oocysts or eggs, in feces. Instead of adding a flotation solution after the final centrifugation step and using a cover slip to collect the parasites, the method using a wire loop for the recovery of the organisms has been reported as one of alternative methods. However, the recovery rates of microorganisms from the flotation method have not been analysed. In the present study, the utility of a flotation method with the use of a wire loop of 8 mm in diameter (the loop method) was evaluated using different numbers of E. tenella oocysts and Heterakis gallinarum eggs, and chicken fecal samples collected at the farms. Consequently, we found that the oocysts and eggs in tubes could be collected at a ratio of 2.00 to 3.08. Thus, our results indicate that the loop method is a simple and time saving method, implicating the application for the estimated OPG/ EPG (Oocysts/Eggs per gram) of the samples. [ABSTRACT FROM AUTHOR]

Objective: This study was to determine the number of chickens in intensively managed poultry farms and to assess the relationship between the number of chickens and risk factors in specific poultry farm. Methods: A cross-sectional study was conducted in Adele town between December 2014 and November 2015. The presence of oocysts in the fecal samples was examine by floatation methods and post-mortem examinations looked for intestinal lesions. Results: Out of the 262 chicken sample sources examined, 29 (11.01%) were positive for coccidian oocysts, and out of the 84 total Environmental sample source examined, 12 (14.4%) were positive for coccidian parasites. There were no statistically significant differences in prevalence across the sample type and source (p>0.05). There were no changes in the prevalence of coccidiosis between the sexes of birds. P<0.05 indicates a statistically significant difference between the age groups. Furthermore, a strong correlation was found between the sample times. The prevalence of coccidia was considerably higher in chicks than in adults (P<0.05). The environment had a greater prevalence of coccidiosis than faeces did. Of the environmental samples, the majority of coccidial infections are linked to management issues, and the prevalence of water and feed is higher than that of litter material. Conclusions: This study showed that coccidiosis is a significant disease affecting poultry in the studied area and that effective control measures must be developed to reduce the illness's impact. [ABSTRACT FROM AUTHOR]

Cryptosporidiosis is a neglected tropical zoonotic disease, commonly associated with a diarrhoea, caused by a protozoan parasite of the genus Cryptosporidium. The aim of this study was to determine the occurrence of Cryptosporidium oocysts in faeces, milk, and water sources, and its associated risk factors in sedentary Fulani herds. Seven hundred and fifty faecal samples (240, 180, 240 and 90 from cattle, sheep, goats and dogs, respectively), and 120 samples each, of milk and water were collected for this study. Faecal samples were concentrated by formol ether concentration technique, then stained using Modified Ziehl Neelsen staining method, to detect Cryptosporidium oocysts. Cryptosporidium oocysts in milk and water samples were also concentrated and stained with same stain. The prevalence of Cryptosporidium oocysts in (cattle, dog, sheep and goat faeces were 18.3 %, 15.6 %, 13.9 % and 4.2 %, respectively.), Significantly higher prevalence was observed in calves ≤ 1 year (P = 0.002), female cattle (P = 0.007) and animals with poor body condition (P < 0.001) than in the other groups. Goats with poor body condition scores had significantly higher prevalence (P = 0.008) than other groups while dogs ≤ 6 months of age also had significantly higher prevalence (P = 0.03) than older dogs. The prevalence of Cryptosporidium oocysts in bulk cow milk was 11.7 % while a prevalence of 13.3 % was observed in the water bodies sampled. Hence the findings of this study are of great public health significance, therefore, inhabitants of these LGAs should be informed and educated on the need for improvement of sanitary measures during the course of milking these animals, and the need for adequate pasteurization, of milk before consumption. [ABSTRACT FROM AUTHOR]

Purpose: Nothing is known about coccidians (Apicomplexa: Eimeriidae) from the Pacific blue-tailed skink, Emoia caeruleocauda. Here, we report mensural and morphometric data on a new species of Isospora from E. caeruleocauda from Guam, US Territory. Methods: Feces from four E. caeruleocauda collected by hand in November 2023 were placed in individual vials containing 2.5% potassium dichromate. They were examined for sporulated oocysts after flotation in Sheather's sugar solution, measured, and photographed. Results: A single (25%) E. caeruleocauda was found to be passing oocysts representing a new species of Isospora. Oocysts of Isospora guamensis n. sp. are ellipsoidal to ovoidal with a bi-layered wall, measure (L × W) 16.5 × 11.8 μm, and have a length/width (L/W) ratio of 1.4; a micropyle and an oocyst residuum were absent but a polar granule was present. Sporocysts are ovoidal and measure 9.4 × 6.5 μm, L/W 1.4; Stieda and sub-Stieda bodies were present but a para-Stieda body was absent. The sporocyst residuum is composed various-sized granules in a compact rounded or irregular mass, sometimes dispersed between the sporozoites. The new species can be differentiated from all other isosporans from skinks by possessing the smallest oocysts known from this host family. Conclusion: This is the first time an isosporan coccidian has been reported from E. caeruleocauda as well as the first report of a coccidian from a Guam-inhabiting skink. [ABSTRACT FROM AUTHOR]

Toxoplasma gondii is a zoonotic parasite that can cause severe morbidity and mortality in warm-blooded animals, including marine mammals such as sea otters. Free-ranging cats can shed environmentally resistant T. gondii oocysts in their feces, which are transported through rain-driven runoff from land to sea. Despite their large population sizes and ability to contribute to environmental oocyst contamination, there are limited studies on T. gondii oocyst shedding by free-ranging cats. We aimed to determine the frequency and genotypes of T. gondii oocysts shed by free-ranging domestic cats in central coastal California and evaluate whether genotypes present in feces are similar to those identified in sea otters that died from fatal toxoplasmosis. We utilized a longitudinal field study of four free-ranging cat colonies to assess oocyst shedding prevalence using microscopy and molecular testing with polymerase chain reaction (PCR). T. gondii DNA was confirmed with primers targeting the ITS1 locus and positive samples were genotyped at the B1 locus. While oocysts were not visualized using microscopy (0/404), we detected T. gondii DNA in 25.9% (94/362) of fecal samples. We genotyped 27 samples at the B1 locus and characterized 13 of these samples at one to three additional loci using multi locus sequence typing (MLST). Parasite DNA detection was significantly higher during the wet season (16.3%, 59/362) compared to the dry season (9.7%; 35/362), suggesting seasonal variation in T. gondii DNA presence in feces. High diversity of T. gondii strains was characterized at the B1 locus, including non-archetypal strains previously associated with sea otter mortalities. Free-ranging cats may thus play an important role in the transmission of virulent T. gondii genotypes that cause morbidity and mortality in marine wildlife. Management of free-ranging cat colonies could reduce environmental contamination with oocysts and subsequent T. gondii infection in endangered marine mammals and people.

Abstract Background Coccidia are a group of intracellular protozoal parasites within the phylum Apicomplexa. Eimeria tenella, one of the species that cause intestinal coccidiosis in poultry, can cause significant mortality and morbidity. Diploid oocysts of Eimeria species are shed in the feces of an infected host and must sporulate to achieve infectivity. This process results in eight haploid infectious units, called sporozoites, held within a single oocyst. Each Eimeria spp. parasite possesses a single apicoplast and a single mitochondrion, both of which carry multiple copies of their respective organellar genomes. Reports of copy numbers of organellar genomes have varied widely. Methods We report the application of quantitative polymerase chain reaction (qPCR), supported by next-generation sequencing, for the quantification of the extranuclear genomes relative to the nuclear genome over the course of sporulation and following its completion. Results At 64 elapsed hours, 93.0% of oocysts were fully sporulated; no increase in percent sporulation was observed after this time. Apicoplast relative genome copy number showed several significant shifts up to 72 elapsed hours, after which no significant shifts were observed. Oocysts were shed with approximately 60% the amount of apicoplast DNA present at 72 h, after which point no significant shifts in apicoplast genome relative abundance occurred. Mitogenome relative copy number showed only two significant shifts, from 16 to 24 elapsed hours and from 24 to 32 elapsed hours. Oocysts were shed with approximately 28% the amount of mitochondrial DNA that was present at the time sporulation was deemed morphologically complete, at 64 elapsed hours. Conclusions The characterization of the dynamics of genome abundance in exogenous stages sheds new light on the basic biology of Eimeria spp. and supports the use of extranuclear targets for molecular modes of parasite quantification and identification with improved sensitivity and accuracy. Graphical Abstract

The most common disease of young poultry in commercial farms of the Russian Caspian region is eimeriosis. In most cases, after convalescence from coccidiosis caused by one of Eimeria species poultry remains susceptible to other species. This parasite has a very short life cycle and immense reproductive capacity that is why it can cause large-scale outbreaks of the disease in commercial poultry houses. To control avian eimeriosis, various coccidiostats are used in combination with probiotics and vitamins. Frequent and long-term use of the same drugs against this infection can potentially result in the emergence of resistant Eimeria populations. This suggests that this coccidiosis control requires rotation of eimeriocidal drugs. Studies on eimeriosis prevalence were performed in the laboratory of the Caspian Regional Research Veterinary Institute and in different poultry farms of the Republic of Dagestan. Swabs of the floor, litter, equipment, droppings, feedstuffs, cecum smears from dead poultry were used for testing. High infection rate with eimerias was established in floor-housed poultry in the plain and piedmont zones of the Republic (Khasavyurtovsky and Karabudakhkentsky raions), where the infection rates were 81.6 and 82.4%, respectively. In batter-cage system poultry farms of the mountain and mountain valley zones (Khunzakhsky and Gergebilsky raions) the infection rates were significantly lower – 61.2 and 58.0%, respectively. The comparative efficacy study of two eimeriocidal drugs showed that “Robenidine”, used daily from the first day of life during the entire rearing period at a dose of 33 g per 1 ton of feedstuffs controls coccidiosis in poultry. At the same time, the survival rate of the experimental young poultry during the observation period was 98.0% compared with “Sarucoxum 12%” group (96.7%).

Background & objectives: Anopheles stephensi is responsible for the transmission of malaria in urban areas. Vector competence of An. stephensi from a non-malarious (Coimbatore) and highly malarious (Chennai) urban area were investigated to find out the reason for the non-transmission of malaria in Coimbatore. Methods: Vector competence (Susceptibility/refractoriness) of An. stephensi mosquitoes from Chennai (Malarious) and Coimbatore (Non-malarious), Tamil Nadu, India to Plasmodium vivax (Chennai) were investigated. Bioassays were carried out concurrently in both these strains by artificial membrane feeding technique using the same malaria infected blood. An. stephensi were dissected to observe infection in the midgut and salivary glands. The parasite infection, oocyst and sporozoite positivity rate, the oocyst load, correlation between male-female gametocyte ratio and infection, and Survival Analysis of parasitic stages during sporogony were analyzed and compared. Results: The overall infection rate was 45.8 and 41.2 per cent in Chennai and Coimbatore. Oocyst count ranged from 1-80 and 1-208 respectively and not statistically significant. Oocyst positivity was high from Day 8-21in both strains. The Mean Survival Day (MSD) for oocyst was Day 14 in both strains. Sporozoite was observed in four experiments in each of the strains and the MSD for sporozoites was Day 20 and Day 17 in Chennai and Coimbatore. Interpretation & conclusion: An. stephensi of Chennai and Coimbatore are equally susceptible to P. vivax infection and the non-transmission of malaria in Coimbatore can be attributed to external factors such as the presence of preferential breeding habitat, vector density, vector survival, and weather. The only difference observed was the comparatively shortened oocyst maturation time in the Coimbatore strain which requires further investigation.

Abstract Background Plasmodium falciparum oocysts undergo growth and maturation in a unique setting within the mosquito midgut, firmly situated between the epithelium and the basal lamina. This location exposes them to specific nutrient exchange and metabolic processes while in direct contact with the mosquito haemolymph. The limited availability of in vitro culture systems for growth of the various P. falciparum mosquito stages hampers study of their biology and impedes progress in combatting malaria. Methods An artificial in vitro environment was established to mimic this distinctive setting, transitioning from a 2D culture system to a 3D model capable of generating fully mature oocysts that give rise to in vitro sporozoites. Results A two-dimensional (2D) chamber slide was employed along with an extracellular matrix composed of type IV collagen, entactin, and gamma laminin. This matrix facilitated development of the optimal medium composition for cultivating mature P. falciparum oocysts in vitro. However, the limitations of this 2D culture system in replicating the in vivo oocyst environment prompted a refinement of the approach by optimizing a three-dimensional (3D) alginate matrix culture system. This new system offered improved attachment, structural support, and nutrient exchange for the developing oocysts, leading to their maturation and the generation of sporozoites. Conclusions This technique enables the in vitro growth of P. falciparum oocysts and sporozoites.

Abstract The availability of nutrients from mosquito blood meals accelerates the development of Plasmodium falciparum laboratory strains in artificially infected Anopheles gambiae mosquitoes. The impact of multiple blood meals on the number of P. falciparum genotypes developing from polyclonal natural human malaria infections (field-isolates) remains unexplored. Here, we experimentally infect An. gambiae with P. falciparum field-isolates and measure the impact of an additional non-infectious blood meal on parasite development. We also assess parasite genetic diversity at the blood stage level of the parasite in the human host and of the sporozoites in the mosquito. Additional blood meals increase the sporozoite infection prevalence and intensity, but do not substantially affect the genetic diversity of sporozoites in the mosquito. The most abundant parasite genotypes in the human blood were transmitted to mosquitoes, suggesting that there was no preferential selection of specific genotypes. This study underlines the importance of additional mosquito blood meals for the development of parasite field-isolates in the mosquito host.

Background and Aim: Cryptosporidium spp. members of the phylum Apicomplexa are obligate protozoan parasites capable of infecting various vertebrate hosts, including rodents and chickens. Infection caused by these parasites may lead to zoonotic diseases in humans. The aim of this study was to estimate the prevalence of Cryptosporidium spp. in rodents and domestic chickens sampled in Franceville, Gabon. Materials and Methods: Two hundred and eighty-five samples were collected, of which 185 samples were from rodents and 100 from domestic chickens. Microscopy after modified Ziehl-Neelsen staining and nested polymerase chain reaction targeting the small subunit (SSU) rRNA gene were used to examine Cryptosporidium spp. Results: The overall prevalence of Cryptosporidium oocysts was 55.8%, with a prevalence of 72.4% in rodents and 25.0% in domestic chickens. Molecular analysis showed that Cryptosporidium spp. were present in 4.0% of the samples. No significant correlation was observed between Cryptosporidium spp. carriage and sex or location in this study. These results indicate that Cryptosporidium spp. persist and circulate in the studied animal species in Franceville, Gabon. Conclusion: Infection with Cryptosporidium is very common in rodents and chickens in Franceville. The potential risk of human contamination cannot be ruled out. More research should be conducted to characterize Cryptosporidium species circulating in rodents and chickens in Gabon. Such studies are essential to better understand the epidemiology of this protozoan and its potential impact on public health.

Background: Plasmodium falciparum oocysts undergo growth and maturation in a unique setting within the mosquito midgut, firmly situated between the epithelium and the basal lamina. This location exposes them to specific nutrient exchange and metabolic processes while in direct contact with the mosquito haemolymph. The limited availability of in vitro culture systems for growth of the various P. falciparum mosquito stages hampers study of their biology and impedes progress in combatting malaria. Methods: An artificial in vitro environment was established to mimic this distinctive setting, transitioning from a 2D culture system to a 3D model capable of generating fully mature oocysts that give rise to in vitro sporozoites. Results: A two-dimensional (2D) chamber slide was employed along with an extracellular matrix composed of type IV collagen, entactin, and gamma laminin. This matrix facilitated development of the optimal medium composition for cultivating mature P. falciparum oocysts in vitro. However, the limitations of this 2D culture system in replicating the in vivo oocyst environment prompted a refinement of the approach by optimizing a three-dimensional (3D) alginate matrix culture system. This new system offered improved attachment, structural support, and nutrient exchange for the developing oocysts, leading to their maturation and the generation of sporozoites. Conclusions: This technique enables the in vitro growth of P. falciparum oocysts and sporozoites. [ABSTRACT FROM AUTHOR]

The study was conducted to investigate the effect of dietary encapsulated organic acids (EOAs) and anticoccidials on the age-dependent development trend of intestinal Lactobacillus, E. coli, coliforms, and Eimeria in Eimeria spp.-infected broiler chickens from reused litter. In total, 525 mixed-sex 1-day-old broiler chickens were used in an uninfected/un-supplemented control plus a 2 (no EOA or 0.1% EOA) × 3 (no anticoccidial, 0.05% maduramicin, and 0.02% diclazuril) factorial arrangement of treatments as a completely randomized design with five replicates of 15 chickens. Results indicated that the cubic model is the best model for explaining the development trends of the intestinal microbial population in uninfected and infected chickens (affected by the EOAs and anticoccidials). Based on the cubic models, the microbial populations had development trends with a decreasing slope from 1-day-old until the early or middle finisher period. EOAs and anticoccidials, especially their simultaneous usage, improved (P < 0.05) the linear and cubic models' slope (affected negatively by Eimeria infection). A polynomial model (order = 6) was determined as the best model for explaining the EOAs and anticoccidial effects on the trend of intestinal Eimeria oocysts in infected chickens. The infection peak (which happened at 25 days) was reduced by EOAs and anticoccidials, especially their simultaneous usage. In conclusion, cubic and polynomial (order = 6) regressions are the best models fitted for explaining the microbiota and Eimeria oocysts trends, respectively. EOAs and anticoccidials, especially their simultaneous usage, had beneficial effects on the microbiota and Eimeria development trends and gastrointestinal health in coccidia-infected broiler chickens.‏ RESEARCH HIGHLIGHTS Cubic regression is the best model for explaining intestinal microbiota development. Polynomial regression is the best model for intestinal Eimeria oocysts development. Age-development trends are affected by dietary encapsulated organic acids and anticoccidials. [ABSTRACT FROM AUTHOR]

This study determines the effects of probiotics - Saccharomyces cerevisiae on oocysts shedding, haematology and serum proteins of broilers infected with mixed Eimeria oocysts. 120-day-old broiler chicks were randomly assigned to four groups (Groups A - D) of 30 birds each. Group A - was fed a plain diet and not infected, Group B - was fed a supplemented diet and infected, Group C - was fed a plain diet and infected, and Group D - was fed a supplemented diet and not infected. S. cerevisiae was supplemented at 0.1 %/kg of feed. At week 3, Groups B and C were infected with sporulated mixed Eimeria oocysts orally, followed by a daily faecal examination of all groups. The rate of oocyst shedding, packed cell volume (PCV), haemoglobin concentration (Hb), red blood cell (RBC) count, total white blood cell (WBC) count, differential WBC count and serum levels of total protein, albumin and globulin were determined at five days' intervals. The data collected were analyzed using a one-way analysis of variance at a 95% confidence interval (p<0.05). Group C had the significantly higher (p<0.05) number of oocysts in faeces, total WBC count and globulin levels, with the lowest PCV, Hb, RBC count, total protein and albumin levels. Group D had significantly higher (p<0.05) heterophil, eosinophil and monocyte counts. In conclusion, dietary supplementation with S. cerevisiae (1 g/kg) ameliorated the effect of Eimeria infection in broilers, by reducing the rate of oocyst shedding, anaemia and hypoproteinaemia due to the activity of the parasite. [ABSTRACT FROM AUTHOR]

The availability of nutrients from mosquito blood meals accelerates the development of Plasmodium falciparum laboratory strains in artificially infected Anopheles gambiae mosquitoes. The impact of multiple blood meals on the number of P. falciparum genotypes developing from polyclonal natural human malaria infections (field-isolates) remains unexplored. Here, we experimentally infect An. gambiae with P. falciparum field-isolates and measure the impact of an additional non-infectious blood meal on parasite development. We also assess parasite genetic diversity at the blood stage level of the parasite in the human host and of the sporozoites in the mosquito. Additional blood meals increase the sporozoite infection prevalence and intensity, but do not substantially affect the genetic diversity of sporozoites in the mosquito. The most abundant parasite genotypes in the human blood were transmitted to mosquitoes, suggesting that there was no preferential selection of specific genotypes. This study underlines the importance of additional mosquito blood meals for the development of parasite field-isolates in the mosquito host. [ABSTRACT FROM AUTHOR]

Intestinal epithelial cells possess the requisite molecular machinery to initiate cell-intrinsic defensive responses against intracellular pathogens, including intracellular parasites. Interferons(IFNs) have been identified as cornerstones of epithelial cell-intrinsic defense against such pathogens in the gastrointestinal tract. Long non-coding RNAs (lncRNAs) are RNA transcripts (>200 nt) not translated into protein and represent a critical regulatory component of mucosal defense. We report here that lncRNA Nostrill facilitates IFN-γ-stimulated intestinal epithelial cell-intrinsic defense against infection by Cryptosporidium, an important opportunistic pathogen in AIDS patients and a common cause of diarrhea in young children. Nostrill promotes transcription of a panel of genes controlled by IFN-γ through facilitating Stat1 chromatin recruitment and thus, enhances expression of several genes associated with cell-intrinsic defense in intestinal epithelial cells in response to IFN-γ stimulation, including Igtp, iNos, and Gadd45g. Induction of Nostrill enhances IFN-γ-stimulated intestinal epithelial defense against Cryptosporidium infection, which is associated with an enhanced autophagy in intestinal epithelial cells. Our findings reveal that Nostrill enhances the transcription of a set of genes regulated by IFN-γ in intestinal epithelial cells. Moreover, induction of Nostrill facilitates the IFN-γ-mediated epithelial cell-intrinsic defense against cryptosporidial infections. [ABSTRACT FROM AUTHOR]

The community's awareness of the prevalence and impact of zoonotic diseases has been significantly underestimated, leading to insufficient implementation of control measures. This study was carried out in Northern Tanzania between 2019 and 2023 to investigate zoonotic parasites and the risks associated with human activities that contribute to zoonotic diseases. Cross‐sectional surveys were conducted in 12 villages, including nine in Loliondo Division and three in Babati District. Focus Group Discussions and Key Informant Interviews were conducted to assess the community's knowledge and practices regarding the risks associated with zoonotic diseases in the surveyed areas. A total of 255 samples were collected from various sources, including latrines, households, livestock enclosures, domestic dogs, and chickens. Out of these samples, 152 tested positive for identifiable parasite eggs and oocysts. These parasites included hookworms (21.7%), Trichuris sp. (14.5%), Strongyloides sp. (13.8%), Eimeria sp. (19.7%), Taeniids (5.9%), Hymenolepis sp. (3.3%), Spirometra sp. (2.6%), and Dipylidium sp. (0.7%). Taeniids and Spirometra species were predominantly found in villages near protected areas such as Arash Sokoni, Oloipiri, Sukenya, Wasso, Orkuyiene, Haytemba, and Loliondo. Hookworms were most commonly detected in Arash Sokoni, Loliondo, Isuguro, and Hyatemba, while Strongyloides sp. was prevalent in Wasso, Sukenya, and Olobo villages. The quantitative analysis reveals significant associations between hygiene practices, proximity to livestock enclosures, ecological factors, and human–animal interaction, highlighting their pivotal roles in determining soil‐transmitted helminth (STH) prevalence across different villages. This study reveals that there was a generally low level of awareness regarding zoonotic diseases and STHs. The detection of STH samples, combined with the limited understanding of zoonotic diseases, emphasizes the importance of taking proactive measures to reduce transmission risks. Prioritizing education and promoting awareness along with implementing comprehensive strategies are essential steps to effectively tackle the problems linked to STH infections and substantially lessen the public health burden caused by zoonotic diseases. [ABSTRACT FROM AUTHOR]

Two local isolates of Eimeria tenella gametocytes against coccidiosis were immunologically evaluated in chickens. Cell mediated immune response was detected by modified splenic cell migration inhibition assay (MSCMIA) and data were expressed in terms of per cent migration index. No significant difference in per cent migration index was detected for the chickens immunized either with Vaccine-I (local isolate-I) or with Vaccine-II (local isolate-II); however per cent migration index was comparatively lower in chickens immunized with Vaccine-II as compared to Vaccine-I; indicating a somewhat higher cell mediated immune (CMI) response. Humoral immune response was monitored by ELISA in vaccinated and control chickens. Significantly elevated (P<0.05) antibody titer (IgG) in chickens immunized with Vaccine-II as compared to Vaccine-I was detected. Significantly higher protection (67%) in chickens immunized with Vaccine-II followed by vaccine-I (49%) was recorded. Further, oocyst count was significantly lower (P<0.05) in chickens immunized with Vaccine-II as compared to those immunized with Vaccine-I. It was concluded that vaccinal strain (Vaccine-II) contained additional protein of high molecular weight (49.23 kDa) in its gametocytes provided cross protection and can be used to prepare commercial vaccine against coccidiosis in poultry. [ABSTRACT FROM AUTHOR]

Background and Aim: Cryptosporidium spp. members of the phylum Apicomplexa are obligate protozoan parasites capable of infecting various vertebrate hosts, including rodents and chickens. Infection caused by these parasites may lead to zoonotic diseases in humans. The aim of this study was to estimate the prevalence of Cryptosporidium spp. in rodents and domestic chickens sampled in Franceville, Gabon. Materials and Methods: Two hundred and eighty-five samples were collected, of which 185 samples were from rodents and 100 from domestic chickens. Microscopy after modified Ziehl-Neelsen staining and nested polymerase chain reaction targeting the small subunit (SSU) rRNA gene were used to examine Cryptosporidium spp. Results: The overall prevalence of Cryptosporidium oocysts was 55.8%, with a prevalence of 72.4% in rodents and 25.0% in domestic chickens. Molecular analysis showed that Cryptosporidium spp. were present in 4.0% of the samples. No significant correlation was observed between Cryptosporidium spp. carriage and sex or location in this study. These results indicate that Cryptosporidium spp. persist and circulate in the studied animal species in Franceville, Gabon. Conclusion: Infection with Cryptosporidium is very common in rodents and chickens in Franceville. The potential risk of human contamination cannot be ruled out. More research should be conducted to characterize Cryptosporidium species circulating in rodents and chickens in Gabon. Such studies are essential to better understand the epidemiology of this protozoan and its potential impact on public health. [ABSTRACT FROM AUTHOR]

Malaria drug research and development efforts have resurged in the last decade following the decelerating rate of mortality and malaria cases in endemic regions. The inefficiency of malaria interventions is largely driven by the spreading resistance of the Plasmodium falciparum parasite to current drug regimens and that of the malaria vector, the Anopheles mosquito, to insecticides. In response to the new eradication agenda, drugs that act by breaking the malaria transmission cycle (transmission-blocking drugs), which has been recognized as an important and additional target for intervention, are being developed. These drugs take advantage of the susceptibility of Plasmodium during population bottlenecks before transmission (gametocytes) and in the mosquito vector (gametes, zygotes, ookinetes, oocysts, sporozoites). To date, compounds targeting stage V gametocytes predominate in the chemical library of transmission-blocking drugs, and some of them have entered clinical trials. The targeting of Plasmodium mosquito stages has recently renewed interest in the development of innovative malaria control tools, which hold promise for the application of compounds effective at these stages. In this review, we highlight the major achievements and provide an update on the research of transmission-blocking drugs, with a particular focus on their chemical scaffolds, antiplasmodial activity, and transmission-blocking potential. [ABSTRACT FROM AUTHOR]

Coccidiosis, a parasitic disease caused by single or multiple Eimeria species, leads to significant economic losses in the poultry industry. The Eimeria life cycle includes schizogony, gametogony, and sporogony. To investigate the dynamics of gene expression and regulatory networks during the development of Eimeria acervulina, we employed time-course transcriptomics to rigorously compare the gene expression patterns between a precocious line (PL) and the wild type (WT) of E. acervulina. The results revealed that the PL enters into gametogony 12 h earlier than the WT, and both the PL and WT exhibited distinct clustering patterns during the development phase. A weighted gene co-expression network analysis (WGCNA) identified genes specifically expressed at four distinct developmental stages, schizogony, gametogony, sporulated oocysts, and unsporulated oocysts, clarifying the key biological processes at each stage. This study used global transcriptome profiling to elucidate molecular variations throughout the E. acervulina life cycle, providing critical insights into molecular characterization and valuable resources for investigating other apicomplexan parasites of public health importance. [ABSTRACT FROM AUTHOR]

Simple Summary: Coccidiosis is an infectious disease that mainly affects cattle, chickens, goats, and sheep. Its source is the genus Eimeria, which produces coccidian protozoans that cause intestinal damage to their hosts. The oral intake of eimerian oocysts, which release infectious sporozoites within the colon, is the first step in the infection process. Eventually, oocysts are released with feces after they successively enter intestinal epithelial cells, where asexual multiplication occurs. Due to the widespread drug resistance brought on by the widespread use of currently available anticoccidial medicines, research into novel therapeutic approaches is being conducted. Various substances obtained from plants have shown promising and optimal anticoccidial as well as extra-therapeutic effects, in addition to other options. In Saudi Arabia, natural sources have been examined as potential controls for murine coccidiosis. Among these sources is Teucrium polium. Due to the aforementioned characteristics, the objective of this study was to examine T. polium leaf extract's anticoccidial and antioxidant effects in Eimeria papillata-infected mice. Eimeria spp. are responsible for the economic loss of both domestic and wild animals due to coccidiosis, the most common parasitic disease. The resistance to currently available drugs used to treat coccidiosis has been proven. Medicinal plants that contain physiologically active phytochemicals have been widely used in traditional medicine. Teucrium polium leaf extract (TPLE) has been shown to exhibit pharmacological, antioxidant, and anticoccidial properties in different experiments. Here, our investigation focused on how T. polium leaf extract affected the way that Eimeria papillate caused intestinal injury in mice. Thirty-five male Swiss albino mice were divided into seven groups, as follows: group I: untreated and uninfected (negative control); group II: uninfected, treated group with TPLE (150 mg/kg b.w); and group III: infected untreated (positive control). Groups III–VII were orally administered 103 sporulated E. papillata oocysts. A total of 60 min after infection, groups IV–VI were treated for five successive days with 50, 150, and 250 mg/kg b.w TPLE, respectively, while group VII was treated with amprolium (120 mg/kg b.w.). The mice had been euthanized on the fifth day post-infection, and the jejunum tissues were prepared for histology and oxidative stress studies. A total of 150 mg/kg of TPLE was the most effective dosage, significantly decreasing oocyst output by about 80.5%, accompanied by a significant reduction in the number of developmental parasitic phases in jejunal sections. In addition, the decrease in the number of goblet cells in the jejuna of mice raised after treatment. Also, TPLE greatly diminished the body weight loss of infected mice. Moreover, our research proved that TPLE reduced oxidative damage due to E. papillata infection via decreasing intestinal malondialdehyde (MDA) and nitric oxide (NO) levels and increasing reduced superoxide dismutase (SOD) and glutathione (GSH) levels. These results demonstrated that TPLE had potent anticoccidial properties. TPE's efficacy as a natural antioxidant has also been demonstrated in reducing oxidative stress and enhancing antioxidant systems to mitigate biochemical and histological changes in the jejunum caused by E. papillata. [ABSTRACT FROM AUTHOR]

The World Health Organization (WHO) has recommended two vaccines for malaria prevention, but neither vaccine meets the desired efficacy and durability for an optimal malaria vaccine. However, there are multiple next-generation vaccines in clinical testing that aim to provide higher efficacy, decrease vaccine delivery demands, and improve cost effectiveness. Computational biology plays a crucial role in informing the design of these vaccines, with techniques such as mRNA-lipid nanoparticle technology offering advantages such as quick manufacturing and safety for infants and pregnant women. The article also discusses the strategy of combining existing vaccines with additional antigens to improve efficacy and the potential of transmission-blocking vaccines. Overall, next-generation vaccines are seen as crucial in achieving malaria elimination goals and improving child health in endemic areas. [Extracted from the article]

Very little is known about the process of meiosis in the apicomplexan parasite Cryptosporidium despite the essentiality of sex in its life cycle. Most cell lines only support asexual growth of Cryptosporidium parvum (C. parvum), but stem cell derived intestinal epithelial cells grown under air-liquid interface (ALI) conditions support the sexual cycle. To examine chromosomal dynamics during meiosis in C. parvum, we generated two transgenic lines of parasites that were fluorescently tagged with mCherry or GFP on chromosomes 1 or 5, respectively. Infection of ALI cultures or Ifngr1-/- mice with mCherry and GFP parasites resulted in cross-fertilization and the formation of "yellow" oocysts, which contain 4 haploid sporozoites that are the product of meiosis. Recombinant oocysts from the F1 generation were purified and used to infect HCT-8 cultures, and phenotypes of the progeny were observed by microscopy. All possible phenotypes predicted by independent segregation were represented equally (~25%) in the population, indicating that C. parvum chromosomes exhibit a Mendelian inheritance pattern. The most common pattern observed from the outgrowth of single oocysts included all possible parental and recombinant phenotypes derived from a single meiotic event, suggesting a high rate of crossover. To estimate the frequency of crossover, additional loci on chromosomes 1 and 5 were tagged and used to monitor intrachromosomal crosses in Ifngr1−/− mice. Both chromosomes showed a high frequency of crossover compared to other apicomplexans with map distances (i.e., 1% recombination) of 3–12 kb. Overall, a high recombination rate may explain many unique characteristics observed in Cryptosporidium spp. such as high rates of speciation, wide variation in host range, and rapid evolution of host-specific virulence factors. Author summary: Although sex is essential for the transmission and maintenance of infection of Cryptosporidium, it has been historically challenging to study the process of meiosis in this medically relevant protist. We utilize recent methodological advances such as a specialized in vitro culture system, cell sorting, and the generation of transgenic parasites to cross identical strains of parasites in the absence of selection pressure to identify intrinsic chromosome behavior during meiosis. By specifically examining the phenotypes from the first generation of parasites, we reveal that cross-fertilization frequently occurs in parasite populations, chromosomes segregate in a Mendelian manner, and the rate of crossover is high on Chromosomes 1 and 5. Understanding these baseline meiotic mechanisms is essential for planning and interpreting future genetic studies of Cryptosporidium seeking to identify genes associated with phenotypes of interest. [ABSTRACT FROM AUTHOR]

Avian coccidiosis, a parasitic disease prevalent in poultry, is caused by Eimeria species and leads to significant economic losses. The use of attenuated live oocyst vaccines has been adopted as an alternative to the use of anticoccidial drugs. However, the accurate detection and differentiation of vaccine strains from virulent ones remain challenging. Therefore, this study presents a novel TaqMan polymerase chain reaction (PCR) detection method that offers enhanced sensitivity, specificity, and reproducibility compared with traditional PCR techniques. Through whole-genome resequencing and bioinformatics analysis, we identified a molecular marker gene, Em_marker6, with a unique 21-base pair deletion specific to the Eimeria maxima attenuated vaccine strain. Optimized primers and probes targeting this marker enabled rapid quantification cycle value achievement and high fluorescence intensity. The standard curve's slope of -3.540 and correlation coefficient of 0.9971 confirmed precise quantification capabilities. The TaqMan PCR method detected as few as 30 plasmid DNA copies and 50 oocysts per reaction, outperforming traditional PCR techniques by an order of magnitude. No cross-reactivity was observed with other E. maxima wide-type strains or common intestinal pathogens, ensuring the exclusive detection of the E. maxima EMPY vaccine strain. Weekly testing over 3 weeks demonstrated minimal variability, indicating robust consistency in the method's application. Testing on 61 clinical samples revealed a 57.38% positivity rate for E. maxima species and 13.11% for the vaccine strain. The Em_marker6 gene exhibited genetic stability across multiple generations, confirming the detection method's robust stability for the attenuated E. maxima vaccine strain. This study significantly advances the field of avian coccidiosis research and control by providing a valuable tool for monitoring vaccine purity and preventing inadvertent infections in vaccinated flocks, aligning with global efforts to curb antibiotic use in animal feed. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the effect of Calotropis procera (Asclepiadaceae) extract on Eimeria piriformis (E. piriformis) oocysts and sporozoites (coccidiosis) isolated from commercial rabbits. Methods: Calotropis procera extract was obtained from dried, pulverized leaves, and macerated in 80 % methanol for 30 h. The extract was analyzed using infrared spectroscopy (IR). Twelve-well plates of 3 mL containing 1 x 104 non-sporulated oocyst were tested in 6 groups: Negative (untreated) control received 2.5 % potassium dichromate solution. Other groups received four concentrations of C. procera leaf extract (12.5, 25, 50, and 100 mg/mL) for oocysts vitality test. Toltrazuril (25 µl/mL) was used as positive control. Moreover, 250, 500, 750, and 1000 µg/mL of C. procera extract were tested for sporozoites vitality test. The mixture was examined daily for 4 days for oocysts vitality and after 12 and 24 h for sporozoite vitality. Results: Findings of IR showed that extracts contained 5 biologically active chemical components, indicating the presence of alkaloids, flavonoids, saponins, and phenols. The extract showed a significant inhibitory effect on E. piriformis oocysts at 100 mg/mL with suppression rates of approximately 89 % after 96 h, nearly similar to toltrazuril (p ≤ 0.01). Furthermore, C. procera extract showed inhibition at the highest repression (p ≤ 0.01) of 89 % of E. piriformis sporozoites viability at 1000 µg/mL after 24 h. The inhibitory rate increased proportionately with an extended incubation duration and high concentration. Conclusion: C. procera extract is as effective as the positive control in treating coccidiosis in rabbits. However, further studies are necessary to determine the active constituents of C. procera and their mode(s) of action. [ABSTRACT FROM AUTHOR]

Abstract This article reports on a golden-billed saltator Saltator aurantiirostris Vieillot, 1817, kept in captivity outside its natural distribution area, in the proximities of the Itatiaia National Park, as a new host for Isospora similisi Coelho, Berto, Neves, Oliveira, Flausino & Lopes, 2013. Additionally, a supplementary molecular identification is provided through the sequencing of three non-overlapping loci of mitochondrial DNA and one locus of the 18S small subunit ribosomal RNA (18S) gene. All the taxonomic features of the I. similisi oocysts shed by S. aurantiirostris were equivalent to those originally described from Saltator similis d’Orbigny & Lafresnaye, 1837. The new sequenced loci were identical, or showed 99.9% similarity, to the samples of I. similisi from S. similis and S. aurantiirostris, confirming the same species from both hosts. Lastly, I. similisi is estimated as a junior synonym of Isospora formarum McQuistion & Capparella, 1992, due to the morphological similarities and wide distribution of its hosts in the Neotropical region. Therefore, this study encourages future taxonomic inquiries into I. similisi collected from other Saltator spp. in order to establish this synonymization of I. formarum with I. similisi, and hence, its wide distribution and dispersion in the Neotropical region, including across the Andes mountains.

Toxoplasma gondii oocysts, which are shed in large quantities in the feces from infected felines, are very stable in the environment, resistant to most inactivation procedures, and highly infectious. The oocyst wall provides an important physical barrier for sporozoites contained inside oocysts, protecting them from many chemical and physical stressors, including most inactivation procedures. Furthermore, sporozoites can withstand large temperature changes, even freeze-thawing, as well as desiccation, high salinity, and other environmental insults; however, the genetic basis for this environmental resistance is unknown. Here, we show that a cluster of four genes encoding Late Embryogenesis Abundant (LEA)-related proteins are required to provide Toxoplasma sporozoites resistance to environmental stresses. Toxoplasma LEA-like genes (TgLEAs) exhibit the characteristic features of intrinsically disordered proteins, explaining some of their properties. Our in vitro biochemical experiments using recombinant TgLEA proteins show that they have cryoprotective effects on the oocyst-resident lactate dehydrogenase enzyme and that induced expression in E. coli of two of them leads to better survival after cold stress. Oocysts from a strain in which the four LEA genes were knocked out en bloc were significantly more susceptible to high salinity, freezing, and desiccation compared to wild-type oocysts. We discuss the evolutionary acquisition of LEA-like genes in Toxoplasma and other oocyst-producing apicomplexan parasites of the Sarcocystidae family and discuss how this has likely contributed to the ability of sporozoites within oocysts to survive outside the host for extended periods. Collectively, our data provide a first molecular detailed view on a mechanism that contributes to the remarkable resilience of oocysts against environmental stresses. IMPORTANCE Toxoplasma gondii oocysts are highly infectious and may survive in the environment for years. Their resistance against disinfectants and irradiation has been attributed to the oocyst and sporocyst walls by acting as physical and permeability barriers. However, the genetic basis for their resistance against stressors like changes in temperature, salinity, or humidity, is unknown. We show that a cluster of four genes encoding Toxoplasma Late Embryogenesis Abundant (TgLEA)-related proteins are important for this resistance to environmental stresses. TgLEAs have features of intrinsically disordered proteins, explaining some of their properties. Recombinant TgLEA proteins show cryoprotective effects on the parasite's lactate dehydrogenase, an abundant enzyme in oocysts, and expression in E. coli of two TgLEAs has a beneficial effect on growth after cold stress. Moreover, oocysts from a strain lacking all four TgLEA genes were more susceptible to high salinity, freezing, and desiccation compared to wild-type oocysts, highlighting the importance of the four TgLEAs for oocyst resilience.

AbstractRaising of rabbits for wealth creation and to improve protein intake among resource-poor communities is a potentially impactful venture. However, common parasitic infestations are among the major challenges to the sustenance of such ventures. Parasites such as Eimeria spp. which cause coccidiosis affect rabbit growth rate and may result in high mortalities. This study was conducted in response to increasing kit mortalities in three municipalities in the Greater Accra Region of Ghana. Two hundred and ninety-six samples of fresh faecal droppings were collected from the faecal trays of rabbits and individually examined for Eimeria oocysts using light microscopy. Prevalence of Eimeria ranged from 60% (Ga South) to 78.4% (Ga Central), with overall prevalence of 67.6%. Eight species of Eimeria were identified namely, E. stiedae, E. intestinalis, E. magna, E. coecicola, E. piriformis, E. irresidua, E. media, and E. perforans. The hepatic species, E. stiedae was dominant in two municipalities and was the overall second most dominant. Rabbits in parts of the Greater Accra region are infested with Eimeria spp. To the best of our knowledge, this is the first research-backed report of rabbit Eimeria spp. in the Greater Accra region. More research is required on the impact of coccidiosis in rabbits in Ghana.

Toxoplasma gondii is a ubiquitous zoonotic parasite that can infect warm-blooded vertebrates, including humans. Felids, the definitive hosts, drive T. gondii infections by shedding the environmentally resistant stage of the parasite (oocysts) in their feces. Few studies characterize the role of climate and anthropogenic factors in oocyst shedding among free-ranging felids, which are responsible for the majority of environmental contamination. We determined how climate and anthropogenic factors influence oocyst shedding in free-ranging domestic cats and wild felids using generalized linear mixed models. T. gondii oocyst shedding data from 47 studies were systematically reviewed and compiled for domestic cats and six wild felid species, encompassing 256 positives out of 9,635 total fecal samples. Shedding prevalence in domestic cats and wild felids was positively associated with human population density at the sampling location. Larger mean diurnal temperature range was associated with more shedding among domestic cats and warmer temperature in the driest quarter was associated with lower oocyst shedding in wild felids. Increasing human population density and temperature fluctuation can exacerbate environmental contamination with the protozoan parasite T. gondii. Management of free-ranging domestic cats could lower the burden of environmental oocysts due to their large population sizes and affinity with human settlements.

Background: Coccidiosis outbreaks in susceptible laying hens can significantly decrease egg production and cause substantial economic loss to the egg industry. The supplementation of poultry diets with chemotherapeutic agents is limited due to antimicrobial resistance and residue in poultry meat or processed products. Therefore, alternative strategies to control coccidiosis are needed, and Artemisia annua (AA) might have the potential to be a phytogenic feed additive, an alternative to anticoccidial agents. This study aimed to investigate the effect of the dietary supplementation of powdered AA on the performance and gut health of laying hens infected with coccidiosis by Eimeria spp. Methods: A total of 225 Hy-Line W-36 laying hens at 23 weeks of age were allocated into 5 treatment groups: 1) control (NC), 2) pair-fed (PF) control, 3) challenged control (CC), 4) CC with dietary inclusion of 0.5% AA (0.5AA), and v) CC with dietary inclusion of 1% AA (1AA). The hens in the CC, 0.5AA, and 1AA groups were orally inoculated with sporulated oocysts of Eimeria maxima (12,500), Eimeria tenella (12,500), and Eimeria acervulina (62,500) at week 25. The PF hens received the same amount of feed consumed by the CC hens from 0--14 days post-inoculation (dpi) of Eimeria spp. The performance of the laying hens, including body weight (BW), hen--day egg production (HDEP), feed intake (FI), and feed conversion ratio (FCR), was measured weekly. Additionally, markers of intestinal health, including gut permeability, lesion score, intestinal morphometry, and immune responses, were evaluated at 6, 14, and 21 dpi. Results: At 6 and 14 dpi, laying hens challenged with Eimeria spp. had a lower BW than PF and NC hens (p < 0.0001). Supplementation of 1% AA improved the HDEP by 8.1% compared to CC hens; however, it was still 15.4% lower than that of PF hens (p < 0.0001). The inclusion of 1% AA did not have any beneficial effect on FI; however, the FCR was improved by 0.61 (2.46) than that of CC hens (3.07; p < 0.0001). The inclusion of 1% AA reduced the severity of the intestinal lesions and increased the recovery of intestinal villi (p < 0.05). Additionally, gut permeability was significantly different between the challenged and non-challenged hens; however, among the challenged hens, the inclusion of AA reduced the gut permeability by 29% compared to CC hens (p < 0.0001). Furthermore, the inclusion of 0.5% AA reduced the inflammatory responses in the infected hens. Conclusion: Dietary inclusion of AA partially restored the performance and gut health of the laying hens and modulated their inflammatory immune response following Eimeria infection; however, further studies are needed to better understand the mode of action and effective dosages to improve the gut health without negative impacts on the performance. [ABSTRACT FROM AUTHOR]

The biological control of gastrointestinal (GI) parasites using predatory fungi has been recently proposed as an accurate and sustainable approach in birds. The current study aimed to assess for the first time the efficacy of using the native ovicidal fungus Mucor circinelloides (FMV-FR1) in reducing coccidia parasitism in peacocks. For this purpose, an in vivo trial was designed in the resident peacock collection (n = 58 birds) of the São Jorge Castle, at Lisbon, Portugal. These animals presented an initial severe infection by coccidia of the genus Eimeria (20106 ± 8034 oocysts per gram of feces, OPG), and thus received commercial feed enriched with a M. circinelloides suspension (1.01 × 108 spores/kg feed), thrice-weekly. Fresh feces were collected every 15 days to calculate the coccidia shedding, using the Mini-FLOTAC technique. The same bird flock served simultaneously as control (t0 days) and test groups (t15–t90 days). The average Eimeria sp. shedding in peacocks decreased up to 92% following fungal administrations, with significant reduction efficacies of 78% (p = 0.004) and 92% (p = 0.012) after 45 and 60 days, respectively. Results from this study suggest that the administration of M. circinelloides spores to birds is an accurate solution to reduce their coccidia parasitism. [ABSTRACT FROM AUTHOR]

Background: Toxoplasmosis affects a quarter of the world's population. Toxoplasma gondii (T.gondii) is an intracellular parasitic protozoa. Macrophages are necessary for proliferation and spread of T.gondii by regulating immunity and metabolism. Family with sequence similarity 96A (Fam96a; formally named Ciao2a) is an evolutionarily conserved protein that is highly expressed in macrophages, but whether it play a role in control of T. gondii infection is unknown. Methodology/principal findings: In this study, we utilized myeloid cell-specific knockout mice to test its role in anti-T. gondii immunity. The results showed that myeloid cell-specific deletion of Fam96a led to exacerbate both acute and chronic toxoplasmosis after exposure to T. gondii. This was related to a defectively reprogrammed polarization in Fam96a-deficient macrophages inhibited the induction of immune effector molecules, including iNOS, by suppressing interferon/STAT1 signaling. Fam96aregulated macrophage polarization process was in part dependent on its ability to fine-tuning intracellular iron (Fe) homeostasis in response to inflammatory stimuli. In addition, Fam96a regulated the mitochondrial oxidative phosphorylation or related events that involved in control of T. gondii. Conclusions/significance: All these findings suggest that Fam96a ablation in macrophages disrupts iron homeostasis and inhibits immune effector molecules, which may aggravate both acute and chronic toxoplasmosis. It highlights that Fam96a may autonomously act as a critical gatekeeper of T. gondii control in macrophages. Author summary: An important feature of macrophages is their ability to adopt many different phenotypes and functions in response to extracellular or intracellular cues. Functionally well-adapted macrophages are required for the defense against T. gondii. Here, we found that Fam96a expressed in macrophages played a role in control of T. gondii infection. Mechanically speaking, Fam96a regulated intracellular iron homeostasis, which affected immune effector molecules. Our study reveals that Fam96a may autonomously act as a critical gatekeeper of intracellular parasitic control by coupling Fe metabolism to adaptative remodeling of macrophage function. It provides a novel target for prevention and treatment of toxoplasmosis and other intracellular pathogens in the future. [ABSTRACT FROM AUTHOR]

Coccidiosis is an intestinal protozoan disease that affects the poultry industry worldwide. The severity of this disease varies depending on the identity of the infectious agents. Therefore, this study was carried out to identify the Eimeria species that affect broiler chickens, Gallus gallus domesticus, through morphological and molecular phylogenetic analyses. Twenty‐five faecal samples were collected from the broiler chickens in a commercial poultry farm in Riyadh (Saudi Arabia). Using the floatation technique, faeces were examined microscopically for the Eimeria occurrence. Identification of Eimeria species was performed based on morphological criteria and molecular tools (DNA amplification for the partial small subunit ribosomal RNA (18S rRNA), internal transcribed spacer (ITS)‐1, and mitochondrial cytochrome c oxidase I (COI) genes. In this study, 32% (8 out of 25) of collected samples were found to be positive for coccidiosis. After sporulation in potassium dichromate (K2Cr2O7), the sporulated oocysts were observed as ovoid and measured 18.37–23.19 µm (19.87) long and 15.07–18.67 µm (16.46) wide, with the anterior location of a polar granule and absence of micropyle. These Eimeria oocysts were assumed to size and shape characteristics of Eimeria acervulina. Molecular analysis was conducted on the sequences of the polymerase chain reaction products from the three genes studied (18S rRNA, ITS‐1, and COI). At the three genes, results showed that the resultant sequences clustered with E. acervulina from different regions confirming morphological description. This study highlighted the importance of molecular techniques to detect avian Eimeria species more than the traditional morphology‐based tool to optimise the appropriate anticoccidial strategies for long‐term control in the studied area. [ABSTRACT FROM AUTHOR]

It is currently unknown whether all Plasmodium falciparum-infected mosquitoes are equally infectious. We assessed sporogonic development using cultured gametocytes in the Netherlands and naturally circulating strains in Burkina Faso. We quantified the number of sporozoites expelled into artificial skin in relation to intact oocysts, ruptured oocysts, and residual salivary gland sporozoites. In laboratory conditions, higher total sporozoite burden was associated with shorter duration of sporogony (p<0.001). Overall, 53% (116/216) of infected Anopheles stephensi mosquitoes expelled sporozoites into artificial skin with a median of 136 expelled sporozoites (interquartile range [IQR], 34-501). There was a strong positive correlation between ruptured oocyst number and salivary gland sporozoite load (ρ = 0.8; p<0.0001) and a weaker positive correlation between salivary gland sporozoite load and number of sporozoites expelled (ρ = 0.35; p=0.0002). In Burkina Faso, Anopheles coluzzii mosquitoes were infected by natural gametocyte carriers. Among salivary gland sporozoite positive mosquitoes, 89% (33/37) expelled sporozoites with a median of 1035 expelled sporozoites (IQR, 171-2969). Again, we observed a strong correlation between ruptured oocyst number and salivary gland sporozoite load (ρ = 0.9; p<0.0001) and a positive correlation between salivary gland sporozoite load and the number of sporozoites expelled (ρ = 0.7; p<0.0001). Several mosquitoes expelled multiple parasite clones during probing. Whilst sporozoite expelling was regularly observed from mosquitoes with low infection burdens, our findings indicate that mosquito infection burden is positively associated with the number of expelled sporozoites. Future work is required to determine the direct implications of these findings for transmission potential. [ABSTRACT FROM AUTHOR]

Background: In 2022, fluralaner was launched on the market for use in the control of the cattle tick Rhipicephalus microplus after showing 100% efficacy in registration trials against the causative agents of cattle tick fever (TFAs). The aim of the present study was to determine whether a strategic control regimen against R. microplus using fluralaner (FLU) in Holstein calves grazing in a tropical region would alter the enzootic stability status of cattle tick fever, triggering outbreaks in these animals up to 22 months age. Methods: In this study, a group of calves treated with FLU was compared with a control group treated with the regimen currently being used on the farm, which consisted of the fipronil + fluazuron formulation (FIFLUA). In the first experiment, the efficacy of the FIFLUA pour-on formulation was evaluated in a field study. In the second experiment, which lasted 550 days, two experimental groups (n = 30/group) of Holstein calves naturally infested with R. microplus were analyzed. Calves aged 4 to 10 months received either a specific treatment regimen with FLU (experimental group) or FIFLUA (control group). During this period, tick counts, animal weight measurement, feces collection (to determine eggs and oocysts per gram of feces), tick fever monitoring, blood smears (to ascertain enzootic stability of the herd), PCR testing for TFAs and serology (indirect enzyme-linked immunosorbent assay [iELISA]) were performed. All calves were evaluated for signs of tick fever between ages 11 and 22 months. Results: FIFLUA showed an acaricidal efficacy of > 90% from post-treatment days 14 to 35. Regarding treatments against the TFAs, the average number of treatments was similar between groups, but animals treated with FLU had a smaller reduction in packed cell volume on some of the evaluation dates of the second and third treatment against TFAs. In calves aged 10 months in the FLU group, B. bovis was not detected by PCR (0/15 samples), 40% of the samples had antibody titers and 33% (10/30) of the samples had positive blood smears. Regarding B. bigemina, > 86% of the samples in both groups tested positive for B. bigemina DNA and antibodies; there was no difference in the antibody titers between the groups. There were no clinical cases of cattle tick fever in calves aged 11 to 22 months. Conclusions: In comparison with the control treatment, the strategic control regimen against R. microplus with FLU that was implemented in the present study did not negatively affect the enzootic stability status of A. marginale and B. bigemina in the herd up to 22 months of age. The enzootic stability status of B. bovis was not reached by either group. These results likely represent a characteristic of the local tick population, so further studies should be performed. [ABSTRACT FROM AUTHOR]

Plasmodium sporozoite development in and egress from oocysts in the Anopheles mosquito remains largely enigmatic. In a previously performed high‐throughput knockout screen, the putative subunit 5 of the prefoldin complex (PbPCS5, PBANKA_0920100) was identified as essential for parasite development during mosquito and liver stage development. Here we generated and analyzed a PbPCS5 knockout parasite line during its development in the mosquito. Interestingly, PbPCS5 deletion does not significantly affect oocyst formation but leads to a growth defect resulting in aberrantly shaped sporozoites. Sporozoites produced in the absence of PbPCS5 were thinner, markedly elongated, and did, in most cases, not contain a nucleus. Sporozoites contained fewer subpellicular microtubules, which reached deep into the sporoblast during sporogony where they contacted and indented nuclei. These aberrantly shaped sporozoites did not reach the salivary glands, and we, therefore, conclude that PbPCS5 is essential for sporogony and the life cycle progression of the parasite during its mosquito stage. [ABSTRACT FROM AUTHOR]