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Abstract Tick-borne diseases in animals are increasing rapidly worldwide, but there is insufficient information about tick-borne diseases infecting dogs in southern Egypt. Thus, in the current study, we detected the presence of Anaplasma marginale (A. marginale) and Babesia canis vogeli (B. canis vogeli) in the blood of dogs. The results revealed that 4/100 (4%) were positive, and a higher infection rate was found in males (75%), than females (25%). The phylogenetic analysis for the major surface protein 4 (msp4) gene in this study was compared with amplicons separate from other reported isolates with alignment by identity 100% with cattle and camels from Egypt, and the phylogenetic analysis for the B. canis vogeli small subunit ribosomal RNA (SSU rRNA) gene in this study identified identity by 99.89% with dogs from Egypt. This report is considered the first report in southern Egypt about A. marginale in dogs based on the sequence analysis of the msp4 gene, providing new data for the classification and identification of A. marginale in dogs compared to A. marginale isolated from other animals in southern Egypt.

Bovine neosporosis is an infection caused by the protozoan parasite Neospora caninum and has substantial veterinary hazards. Neosporosis cannot be controlled by vaccination or chemotherapy. Thus, accurate diagnosis followed by isolation and culling of infected animals is regarded as the most efficient method of control. In vivo diagnosis often relies on serologic testing of the animals, and milk represents a non-invasive and easy-to-collect sample matrix. However, indirect enzyme-linked immunosorbent assay (ELISA) specifically designed for antibody detection in milk are sometimes not easily available and it is tempting to use ELISA kits that are originally designed for use in serum in milk samples instead. Herein, we evaluated a widely used commercial ELISA (ID Screen® Neospora caninum competition Multispecies ELISA (ID. Vet, Grabels, France)), developed for detection of N. caninum antibodies in serum samples, for its performance on milk samples. Milk samples from dairy ruminants (cows, buffaloes, sheep, and goats; n = 149) were tested in parallel with the serum ELISA and a commercial milk ELISA as a standard test (Neospora caninum Milk Competitive ELISA, ID. Vet, Grabels, France). The detected prevalence values were 28.2% (42/149), 17.4% (26/149), and 17.4% (26/149) using milk ELISA, serum ELISA, and both ELISAs, respectively. Sensitivity, specificity, positive predictive value, and negative predictive value for the serum ELISA used with milk samples were 61.9%, 100%, 100%, and 87%, respectively. The agreement and kappa value between the two ELISAs were 89.3% and 0.70, respectively, suggesting substantial agreement. High values of Pearson correlation coefficient (0.904, p ≥ 0.0001) and area under the receiver operating characteristic (ROC) curve (0.789, p ≥ 0.0001) demonstrated the high diagnostic performance of the serum ELISA in milk samples. Also, a Bland–Altman Plot and histogram describing the frequency of distribution of ELISA optical densities confirmed the high agreement of both serum and milk ELISAs. The current results revealed the high specificity but moderate sensitivity of the serum ELISA used for milk samples compared with the milk ELISA. However, the excellent positive predictive value of the serum ELISA makes it an alternative option in case of the unavailability of milk ELISAs. With this study, we provided additional evidence that a widely used serum ELISA test kit may also be used for the detection of N. caninum antibodies in milk samples.

Toxoplasma gondii is an intracellular protozoan parasite of veterinary and public health importance. Infection may lead to abortion in susceptible pregnant animals and women, and potentially fatal health complications in immunocompromised individuals. In this study, we aimed to provide an update on the seroprevalence of, and risk factors for, T. gondii antibodies in cattle from Qena, southern Egypt. Additionally, we investigated if raw milk and serum samples from the same animals reacted similarly in a commercial ELISA, thus potentially reducing the invasiveness of future serosurveillance studies. Cattle serum samples (n = 362) from three locations in the Qena governorate (Qena, Qus, and Al Waqf cities), of both sexes and different ages were collected. From most dairy cows, a corresponding milk sample (n = 154) was additionally obtained. We found that the overall seroprevalence in serum samples was 9.1% (33/362). Increasing age was the sole risk factor identified in our study among all tested parameters (location, age, sex, lactating yes or no). Thus, older cattle (more than 3 years old) exhibited significantly higher rates of T. gondii antibodies (11.7%; p = 0.033, odd ratio = 4.3) in comparison to animals younger than 1 year (2.9%). In the corresponding serum and milk samples, the prevalence was 12.3% (19/154) in serum samples, and 9.7% (15/154) in milk samples, respectively. A high correlation was observed between the two sample types with a concordance of 97.4%, a kappa value of 0.87, and a Pearson r correlation coefficient of 0.85. When the serum ELISA was taken as the gold standard, the milk ELISA had the following characteristics: sensitivity (78.9%), specificity (100%), positive predictive value (100%), negative predictive value (97.1%), and area under the receiver operating characteristic (ROC) curve (0.6, p = 0.0011). In this study, we confirmed the frequent occurrence of T. gondii antibodies in cattle in southern Egypt and demonstrated that non-invasive milk samples may be used instead of serum samples for seroprevalence studies in dairy cows.

Published data on tick-borne pathogens (TBPs) in camels worldwide have been collected to provide an overview of the global prevalence and species diversity of camelid TBPs. Several TBPs have been detected in dromedary camels, raising concerns regarding their role as natural or maintenance hosts for tick-borne pathogens. Insubstantial evidence exists regarding the natural infection of camels with Babesia spp., Theileria spp., Anaplasma spp., and Ehrlichia spp., particularly because most of the camels were considered healthy at the time of sampling. Based on polymerase chain reaction (PCR) testing, a pooled prevalence of 35.3% (95% CI: 22.6–48.1%) was estimated for Anaplasma, which was the most frequently tested TBP in dromedaries, and DNA of Anaplasma marginale, Anaplasma centrale, Anaplasma ovis, Anaplasma platys, and A. platys-like were isolated, of which ruminants and dogs are reservoirs. Similarly, the estimated pooled prevalence for the two piroplasmid genera; Babesia and Theileria was approximately equal (10–12%) regardless of the detection method (microscopy or PCR testing). Nevertheless, Babesia caballi, Theileria equi, and Theileria annulata DNA have frequently been detected in camels but they have not yet been proven to be natural hosts. Scarce data detected Babesia microti, Anaplasma phagocytophilum, and Borrelia burgdorferi sensu lato (s.l.) DNA in blood of dromedaries, although ticks of the genus Ixodes are distributed in limited areas where dromedaries are raised. Interestingly, a pooled seroprevalence of 47.7% (26.3–69.2%) was estimated for Crimean–Congo hemorrhagic fever virus, and viral RNA was detected in dromedary blood; however, their contribution to maintain the viral transmission cycles requires further experimental investigation. The substantially low incidence and scarcity of data on Rickettsia and Ehrlichia species could imply that camels were accidentally infected. In contrast, camels may play a role in the spread of Coxiella burnetii, which is primarily transmitted through the inhalation of aerosols emitted by diseased animals and contaminated environments. Bactrian camels showed no symptoms due to the examined TBPs, meanwhile, clinical disease was seen in alpacas infected with A. phagocytophilum. Similar to dromedaries, accidental tick bites may be the cause of TBP DNA found in the blood of Bactrian camels.

Brucellosis and coxiellosis/Q fever are bacterial infections caused by Brucella species and Coxiella burnetii, respectively; camels are highly susceptible to both pathogens. Trichinellosis is a parasitic infection caused by various Trichinella nematode species. Reportedly, camels are susceptible to experimental infection with Trichinella spp., but information on this potential host species is scarce. All three infections are of zoonotic nature and thus of great public health concern. The current study aimed to determine antibodies against the three pathogens in recently imported camels (n = 491) from Sudan at the two main ports for the entrance of camels into southern Egypt using commercial indirect ELISAs. Samples were collected in two sampling periods. The seropositivity rates of Brucella spp., C. burnetii, and Trichinella spp. were 3.5%, 4.3%, and 2.4%, respectively. Mixed seropositivity was found in 1% for Brucella spp. and C. burnetii. Marked differences were found between the two study sites and the two sampling periods for Brucella. A higher rate of seropositivity was recorded in the Red Sea/older samples that were collected between 2015 and 2016 (4.3%, 17/391; odds ratio = 9.4; p < 0.030) than in those collected in Aswan/recent samples that were collected between 2018 and 2021 (0/100). Concerning C. burnetii, samples collected during November and December 2015 had a significantly higher positivity rate than the other samples (13%, 13/100; OD = 4.8; p < 0.016). The same effect was observed for antibodies to Trichinella spp., with samples collected during November and December 2015 showing a higher positivity rate than the other samples (7%, 7/100; OD = 10.9; p < 0.001). This study provides valuable information on the seroprevalence of Brucella spp. and additional novel information on C. burnetii and Trichinella spp. in recently imported camels kept in quarantine before delivery to other Egyptian regions. This knowledge can be utilized to reduce health hazards and financial burdens attributable to brucellosis, Q fever, and trichinellosis in animals and humans in Egypt.

Pork production is a niche economy in Egypt, and pigs are typically raised as backyard animals with no sanitary control, potentially exposing them to various pathogens. Commercially available ELISAs were used to detect specific antibodies to the food-borne zoonotic parasites Toxoplasma gondii and Trichinella spp., as well as to Neospora caninum, in serum samples of pigs slaughtered at Egypt’s only licensed pig abattoir, the El-Bassatin abattoir in Cairo. Among the tested sera (n = 332), seroreactivity for T. gondii was 45.8% (95% confidence interval: 40.4–51.3), N. caninum was 28.0% (95% CI: 23.3–33.2), and Trichinella spp. was 1.2% (95% CI: 0.4–3.3). Mixed infection was only detected for T. gondii and N. caninum (18.7%; 95% CI: 14.7–23.4). The seroprevalence of T. gondii was significantly higher (p = 0.0003) in animals collected from southern Cairo (15 May city slum) than in eastern Cairo (Ezbet El Nakhl slum). Seroprevalence for N. caninum was higher in western (Manshiyat Naser slum; p = 0.0003) and southern Cairo (15 May city slum; p = 0.0003) than in that of eastern Cairo (Ezbet El Nakhl slum; p = 0.0003). Moreover, female pigs exhibited a higher rate of N. caninum antibodies than male ones (p < 0.0001). This study provides the first seroprevalence data for N. caninum in pigs in Egypt, and updates the prevalence of the zoonotic parasites Trichinella spp. and T. gondii.

IntroductionNeospora caninum and Toxoplasma gondii are closely related obligate intracellular protozoan parasites. They are considered to be the major causes of infectious abortions and congenital abnormalities in livestock worldwide resulting in huge economic losses. Currently, there are no reports on the prevalence of neosporosis or toxoplasmosis in cattle in Beheira, Egypt's most important region for cattle industry.MethodsThe current study investigated the presence of anti-N. caninum and anti-T. gondii antibodies in apparent healthy cattle from eight localities representing the whole area of Beheira. A total of 358 plasma samples were randomly collected from 6 dairy and 10 beef farms and analyzed by commercially available ELISAs. Production type (dairy versus beef), sex (female vs male), age (< 3 years, 3–5, and > 5 years old), breed (mixed vs Holstein vs Colombian Zebu), and location (various locations) were assessed as risk factors for N. caninum and T. gondii infections.Results and discussionOf the samples, 88 (24.6%) and 19 (5.3%) were positive for anti-N. caninum and anti-T. gondii antibodies, respectively, and mixed infection was detected in 7. Of the 16 herds, 6 dairy and 7 beef herds were positive for antibodies to N. caninum. Antibodies to T. gondii were detected in 4, and 5 of dairy and beef herds, respectively. Production type (dairy) and, therewith, sex (female), age (aged over 5 years), and location were considered as risk factors for N. caninum infection. No factors statistically associated with T. gondii infection were identified. Overall, this study provided the first serological detection of N. caninum and T. gondii infections in cattle from Beheira, demonstrating the endemicity of both parasites in the main cattle rearing region of Egypt. This study also confirmed earlier reports of N. caninum being more present in dairy cattle than in beef cattle. Routine monitoring of N. caninum and T. gondii infections and the implementation of control strategies are urgently needed.

IntroductionFascioliasis is a parasitic foodborne disease caused by the liver flukes, Fasciola hepatica and F. gigantica. Such parasites cause serious illness in numerous domestic animals and also in humans. Following infection, the parasite secretes a variety of molecules that immediately interact with the host immunity to establish successful infection. These molecules include cathepsin L peptidase 1 (CatL1); the highly investigated diagnostic and vaccine antigens using various animal models. However, a few studies have analyzed the potentials of FhCatL1 as a diagnostic or vaccine antigen using bioinformatic tools and much less for FgCatL1. The present study provides inclusive and exclusive information on the physico-chemical, antigenic and immunogenic properties of F. hepatica cathepsin L1 (FhCatL1) protein using multiple bioinformatic analysis tools and several online web servers. Also, the validation of our employed available online servers was conducted against a huge collection of previously published studies focusing on the properties of FhCatL1as a diagnostic and vaccine antigen.MethodsFor this purpose, the secondary, tertiary, and quaternary structure of FhCatL1 protein were also predicted and analyzed using the SWISS-MODEL server. Validation of the modeled structures was performed by Ramachandran plots. The antigenic epitopes of the protein were predicted by IEDB server.Results and discussionOur findings revealed the low similarity of FhCatL1 with mammalian CatL1, lacking signal peptides or transmembrane domain, and the presence of 33 phosphorylation sites. Also, the containment of FhCatL1 for many topological, physico-chemical, immunological properties that favored its function of solubility and interaction with the immune components were reported. In addition, the earlier worldwide reports documented the high efficacy of FhCatL1 as a diagnostic and vaccine antigen in different animals. Altogether, FhCatL1 is considered an excellent candidate for using in commercialized diagnostic assays or vaccine products against fascioliasis in different animal species. Our assessment also included FgCatL1 and reported very similar findings and outputs to those of FhCatL1.

IntroductionToxoplasma gondii and Neospora caninum are closely related intracellular protozoan parasites of medical and veterinary concern by causing abortions and systemic illness. Limited or ambiguous data on the prevalence of T. gondii and N. caninum in camels triggered us to conduct this study.MethodsCamels (n = 460) recently imported from Sudan and destined mainly for human consumption, were tested for specific antibodies against these protozoans using commercially available ELISAs. From the two only quarantine stations for camels from Sudan, 368 camels were sampled between November 2015 and March 2016 in Shalateen, Red Sea governorate, and 92 samples were collected between September 2018 and March 2021 from Abu Simbel, Aswan governorate.Results & DiscussionOverall, seropositive rates in camels were 25.7%, 3.9% and 0.8% for T. gondii, N. caninum and mixed infection, respectively. However, marked differences were found between the two study sites and/or the two sampling periods: For T. gondii, a higher rate of infection was recorded in the Red Sea samples (31.5%, 116/368; odds ratio 20.7, 5.0-85.6; P

Domestic cats are a potential source of Toxoplasma gondii infection for humans. This study was conducted to determine the seroprevalence and risk factors for T. gondii infection in domestic cats. Cat sera (n = 182) were tested for T. gondii IgG antibodies using the latex agglutination test (LAT) and the GRA7 of T. gondii (TgGRA7)-indirect enzyme-linked immunosorbent assay (iELISA). Univariable and multivariable logistic regression analyses were used to identify the factors associated with T. gondii infection. The overall prevalence rates were 18.1% (33/182) according to LAT, 19.2% (35/182) according to the iELISA and 17.0% (31/182) according to LAT and iELISA. Univariable analyses identified, outdoor access (P = 0.006), being a former stray cat (P = 0.001) and successful hunting behaviors (P = 0.04) as risk factors for T. gondii infection. Outdoor access (OR 2.63, 95% confidence interval (CI) 1.03–6.72) and is a former stray cat (OR 3.69, 95% CI 1.52–8.96) remained significant risk factors in multivariable analyses. This study indicated a relatively high seroprevalence of T. gondii among domestic cats. Cat owners can reduce the risk for T. gondii infection by not allowing their cats to roam free. Furthermore, education about the transmission of the parasite should be provided to prevent infection to the owners.

The Barki sheep industry is becoming increasingly important in Egypt because of the high quality of their meat and wool. This sheep breed is also commonly known for its resistance to arid and harsh environmental conditions. Such characteristics can be exploited in solving the problematic situation of inadequate animal protein for human consumption, particularly under climatic changes. However, very few studies have investigated aspects of breeding, nutrition, and susceptibility to infectious or non-infectious diseases in Barki sheep. Herein, we propose to unravel the differences in the clinical and biochemical profiles among Barki sheep of different growth rates. We measured clinical and biochemical parameters in stunted (n = 10; test group) and in good body condition (n = 9; control group) Barki sheep. Animals subjected to this experiment were of the same sex (female), age (12 months old), and housed in the same farm with similar conditions of feeding, management practice, and vaccination and deworming regimens. Regarding clinical examination, stunted/tested sheep showed a significantly higher pulse and respiratory rate compared to sheep with a good body condition/control group. The appetite, body temperature, and digestion processes were the same in both groups. In biochemical investigations, nutritional biomarkers were reduced markedly in stunted sheep compared with the control sheep, including total protein (p = 0.0445), albumin (p = 0.0087), cholesterol (p = 0.0007), and triglycerides (p = 0.0059). In addition, the Barki sheep test group suffered from higher levels of urea and blood urea nitrogen than the control group. Consistently, growth and thyroid hormone levels were lower in stunted sheep than the control sheep, although the differences were not statistically significant (p > 0.05). No significant differences were detected in both groups for serum levels of calcium, phosphorus, magnesium, iron, and zinc (p > 0.05). To detect the reasons for emaciation, certain debilitating infections were tested. All tested sheep showed negative coprological tests for gastrointestinal parasites, and had no obvious seropositivity to brucellosis, toxoplasmosis, neosporosis, or Q fever. This study demonstrates the useful biochemical markers for monitoring growth performance in Egyptian Barki sheep and unravels the usefulness of this breed in nationwide breeding and farming.

Toxoplasma gondii and Neospora caninum are apicomplexan protozoan parasites that have been associated with reproductive problems in ruminants. Despite the high seroprevalence of T. gondii and N. caninum reported in goats worldwide, little information about the seroprevalence of these parasites in goats in Thailand. In this study, we investigated the seroprevalence of T. gondii and N. caninum in 389 goat samples from five districts of Kanchanaburi province, Thailand. An indirect enzyme-linked immunosorbent assay (iELISA) using recombinant TgGRA7 and NcSAG1 antigens was used for the detection of anti-T.gondii and anti-N.caninum antibodies, respectively. Risk factors for the occurrence of these protozoan parasites on goat farms were also assessed. Specific IgG against T. gondii and N. caninum was detected in 28.5% and 16.7% samples, respectively. Co-infections were observed in 11.8% of samples. The risk factors significantly associated with T. gondii seroprevalence was the presence of cats (odds ratio [OR]= 2.55, 95% confidential interval [CI]=1.67- 3.89). The presence of other domestic animals on farm contributed to the risk of T. gondii and N. caninum infection on goat farms (OR=1.692, 95% CI= 1.08- 2.63). The present study provides data on the seroprevalence and risk of T. gondii and N. caninum infections among goats in Kanchanaburi province, Thailand. Integrated and efficient measures are required to prevent and control infections on goat farms.

Background: Toxoplasma gondii and Neospora caninum are major protozoan parasites of worldwide distribution and significance in veterinary medicine and, for T. gondii, in public health. Cats and dogs, as final hosts for T. gondii and N. caninum, respectively, have a key function in environmental contamination with oocysts and, thus, in parasite transmission. Very little is known about the prevalence of T. gondii infections in dogs and cats in Egypt, and even less about the prevalence of N. caninum in the same hosts. Methods: In the current study, 223 serum samples of both dogs (n = 172) and cats (n = 51) were investigated for specific antibodies to T. gondii and N. caninum using commercially available ELISAs. A risk factor analysis was conducted to identify factors associated with seropositivity. Results & discussion: Exposure to T. gondii was reported in 23.3% of the dogs and in 9.8% of the cats, respectively. In addition, N. caninum-specific antibodies were recorded in 5.8% of dogs and in 3.4% of cats. A mixed infection was found in two dogs (1.2%) and in one cat (2%). Antibodies to T. gondii in dogs were significantly more frequent in dogs aged 3 years or more and in male German Shepherds. As this breed is often used as watchdogs and was the most sampled breed in Alexandria governorate, the purpose “watchdog” (compared to “stray” or “companion”), the male sex, and the governorate “Alexandria” also had a significantly higher seroprevalence for T. gondii. No factors associated with antibodies to N. caninum could be identified in dogs, and no significant factors were determined in cats for either T. gondii or N. caninum infection. Our study substantially adds to the knowledge of T. gondii infection in dogs and cats and presents data on N. caninum infection in cats for the first and in dogs in Egypt for the second time.

Toxoplasmosis and neosporosis are major protozoan diseases of global distribution. Toxoplasma gondii is the cause of toxoplasmosis, which affects almost all warm-blooded animals, including humans, while Neospora caninum induces neosporosis in many animal species, especially cattle. The current defective situation with control measures is hindering all efforts to overcome the health hazards and economic losses of toxoplasmosis and neosporosis. Adequate understanding of host-parasite interactions and host strategies to combat such infections can be exploited in establishing potent control measures, including vaccine development. Macrophages are the first defense line of innate immunity, which is responsible for the successful elimination of T.gondii or N. caninum. This action is exerted via the immunoregulatory interleukin-12 (IL-12), which orchestrates the production of interferon gamma (IFN-γ) from various immune cells. Cellular immune response and IFN-γ production is the hallmark for successful vaccine candidates against both T. gondii and N. caninum. However, the discovery of potential vaccine candidates is a highly laborious, time-consuming and expensive procedure. In this review, we will try to exploit previous knowledge and our research experience to establish an efficient immunological approach for exploring potential vaccine candidates against T. gondii and N. caninum. Our previous studies on vaccine development against both T. gondii and N. caninum revealed a strong association between the successful and potential vaccine antigens and their ability to promote the macrophage secretion of IL-12 using a murine model. This phenomenon was emphasized using different recombinant antigens, parasites, and experimental approaches. Upon these data and research trials, IL-12 production from murine macrophages can be used as an initial predictor for judgment of vaccine efficacy before further evaluation in time-consuming and laborious in vivo experiments. However, more studies and research are required to conceptualize this immunological approach.

Toxoplasmosis, neosporosis, and brucellosis are devastating diseases causing infectious abortion and, therefore, substantial economic losses in farm animals. Toxoplasmosis and neosporosis are caused by the intracellular protozoan parasites Toxoplasma gondii (T. gondii) and Neospora caninum (N. caninum), respectively. Brucellosis is a bacterial disease caused by numerous Brucella species in multiple hosts. Toxoplasmosis and brucellosis are also considered foodborne zoonotic diseases. In the current study, specific antibodies to T. gondii and N. caninum, in addition to those to Brucella spp., were detected to gain a better understanding of the epidemiological situation for these three pathogens. Sheep and goat sera from Egypt (n = 360) of animals with and without a history of abortion were tested using commercial ELISAs. Seropositivity rates of 46.1%, 11.9%, and 8.6% for T. gondii, N. caninum, and Brucella spp., respectively, were revealed. Mixed infections with T. gondii and Brucella spp. (4.4%), T. gondii and N. caninum (4.2%), N. caninum and Brucella spp. (1.4%), and even some triple infections (0.6%) have been observed. Animals with a history of abortion had a significantly higher seroprevalence for Brucella spp. infection than those without abortion (12.6%; 28/222 vs. 2.2%; 3/138) (p = 0.0005; Odds ratio = 1.9–21.8), while none of the other pathogens showed a similar effect. This result suggests brucellosis as a possible cause of abortion in the study population. However, the high seroprevalence for T. gondii and N. caninum revealed in our study warrants further investigations.

The prevalence of Neospora caninum and Toxoplasma gondii antibodies in raw milk samples was estimated in different ruminants and Egyptian governorates. Of 13 bulk milk samples tested by ELISA, five (38.5%) were positive for antibodies to N. caninum, and two samples were additionally positive for antibodies to T. gondii, resulting in a seroprevalence of 15.4% for both T. gondii and co-infection. In individual milk samples (n = 171) from the same bulks, antibodies to N. caninum were detected in 25.7%, to T. gondii in 14%, and 3.5% had antibodies to both parasites. A strong correlation between the OD values of the bulk samples and of the relevant individual milk samples was found for T. gondii (Pearson r = 0.9759) and moderately strong for N. caninum (Pearson r = 0.5801). Risk factor assessment for individual milk samples revealed that antibodies to T. gondii were significantly influenced by animal species, while no risk factors were detected for N. caninum antibodies. Additionally, DNA of N. caninum was detected in a bulk milk sample of cattle for the first time in Egypt, and DNA of T. gondii was found in bulk milk samples of cattle, sheep and goats. This is the first study in Egypt in which bulk milk samples of different ruminants were tested for the presence of N. caninum and T. gondii antibodies and DNA. Both individual and bulk milk samples are useful tools for monitoring antibody response to N. caninum and T. gondii infections in different ruminants in Egypt.

Toxoplasma gondii infects almost all warm-blooded animals, including humans, leading to both cellular and humoral immune responses in the host. The virulence of T. gondii is strain specific and is defined by secreted effector proteins that disturb host immunity. Here, we focus on nuclear factor-kappa B (NFκB) signaling, which regulates the induction of T-helper type 1 immunity. A luciferase assay for screening effector proteins, including ROPs and GRAs that have biological activity against an NFκB-dependent reporter plasmid, found that overexpression of GRA7, 14, and 15 of a type II strain resulted in a strong activity. Thus, our study was aimed at understanding the involvement of NFκB in the pathogenesis of toxoplasmosis through a comparative analysis of these three molecules. We found that GRA7 and GRA14 were partially involved in the activation of NFκB, whereas GRA15 was essential for NFκB activation. The deletion of GRA7, GRA14, and GRA15 in the type II Prugniaud (Pru) strain resulted in a defect in the nuclear translocation of RelA. Cells infected with the PruΔgra15 parasite showed reduced phosphorylation of inhibitor-κBα. GRA7, GRA14, and GRA15 deficiency decreased the levels of interleukin-6 in RAW246.7 cells, and RNA-seq analysis revealed that GRA7, GRA14, and GRA15 deficiency predominantly resulted in downregulation of gene expression mediated by NFκB. The virulence of all mutant strains increased, but PruΔgra14 only showed a slight increase in virulence. However, the intra-footpad injection of the highly-virulent type I RHΔgra14 parasites in mice resulted in increased virulence. This study shows that GRA7, 14, and 15-induced host immunity via NFκB limits parasite expansion.

Toxoplasma gondii is the causative agent of toxoplasmosis in humans and various animal species worldwide. In Thailand, seroprevalence studies on T. gondii have focused on domestic animals, and information on infections in Asian elephants (Elephas maximus indicus) is scarce. This study was conducted to determine the seroprevalence of T. gondii infection in archival sera collected from 268 elephants living in Thailand. The serum samples were analyzed for anti-T. gondii immunoglobulin G antibodies using the latex agglutination test (LAT) and indirect enzyme-linked immunosorbent assay (iELISA) based on T. gondii lysate antigen (TLA-iELISA) and recombinant T. gondii dense granular antigen 8 protein (TgGRA8-iELISA). The prevalence of antibodies against T. gondii was 45.1% (121/268), 40.7% (109/268), and 44.4% (119/268) using LAT, TLA-iELISA, and TgGRA8-iELISA, respectively. Young elephants had a higher seropositivity rate than elephants aged >40 years (odds ratio = 6.6; p < 0.001; 95% confidence interval: 2.9–15.4). When LAT was used as the reference, TLA-iELISA and TgGRA8-iELISA showed a substantial (κ = 0.69) and moderate (κ = 0.42) agreement, respectively. Although our findings suggest the widespread exposure of Asian elephants to T. gondii in Thailand, the source of infection was not investigated. Therefore, investigation of the predisposing factors associated with toxoplasmosis is necessary to identify the potential risk factors for infection.

The obligate intracellular parasite Toxoplasma gondii infects warm-blooded animals, including humans. We previously revealed through a whole-brain transcriptome analysis that infection with T. gondii in mice causes immune response-associated genes to be upregulated, for instance, chemokines and chemokine receptors such as CXC chemokine receptor 3 (CXCR3) and its ligand CXC chemokine ligand 10 (CXCL10). Here, we describe the effect of CXCR3 on responses against T. gondii infection in the mouse brain. In vivo assays using CXCR3-deficient mice showed that the absence of CXCR3 delayed the normal recovery of body weight and increased the brain parasite burden, suggesting that CXCR3 plays a role in the control of pathology in the brain, the site where chronic infection occurs. Therefore, to further analyze the function of CXCR3 in the brain, we profiled the gene expression patterns of primary astrocytes and microglia by RNA sequencing and subsequent analyses. CXCR3 deficiency impaired the normal upregulation of immune-related genes during T. gondii infection, in astrocytes and microglia alike. Collectively, our results suggest that the immune-related genes upregulated by CXCR3 perform a particular role in controlling pathology when the host is chronically infected with T. gondii in the brain.

Neospora caninum is an intracellular protozoan parasite responsible for numerous abortion outbreaks and neonatal abnormalities in cattle. Rapid and accurate diagnosis is critical for N. caninum control owing to the lack of vaccine or drug-based control strategies. Herein, we evaluated the performance of four frequently used antigens in the diagnosis of N. caninum infection using immunochromatographic tests (ICTs) as a rapid, affordable, and field applicable tool. These antigens included recombinant proteins of N. caninum surface antigen 1 (NcSAG1), dense granule proteins 7 (NcGRA7) and 6 (NcGRA6), in addition to native Neospora lysate antigen (NLA). Our study revealed the utility of all antigen-based ICTs for detection of specific antibodies to N. caninum. However, the NcSAG1-based ICT was the best for detection of all control N. caninum-infected mouse or cattle sera, while NcGRA7 and NcGRA6-based ICTs exhibited specific ability to detect samples from acute and sub-acute infection in mice and cattle, respectively. Analyses of the NcSAG1-based ICT against enzyme-linked immunosorbent assays (ELISAs) of the same antigen revealed its efficiency in detection of field cattle samples as observed in high sensitivity (84.2%), specificity (93.5%), agreement (90%), and kappa value (0.78). The current knowledge provides an efficient platform for N. caninum control through on-site diagnosis of infected cattle.

Vaccination is the mainstay of preventative measures for numerous infectious diseases. Neospora caninum infection induces storms of abortion in pregnant cows and ewes, resulting in drastic economic losses because of fetal losses and culling of the dams. Herein, we evaluated the potential of recombinant protein of N. caninum dense granule protein 6 fused with glutathione-S-transferase (NcGRA6+GST) as a vaccine candidate against neosporosis in a pregnant mouse model. The protective efficacy was investigated by subcutaneous inoculation of BALB/c mice with recombinant NcGRA6+GST (25 pmol), and GST alone (25 pmol) or phosphate-buffered saline (PBS) as the controls. This study revealed the partial ability of NcGRA6+GST to protect the dams and offspring from N. caninum infection during the critical period of pregnancy. This ability was revealed by higher survival rate and lower parasite burden in brains of offspring of the NcGRA6+GST-immunized group in comparison with the control groups. In addition, mouse dams from NcGRA6+GST-immunized groups exhibited lower clinical score and minimum alteration in body weight in comparison with PBS or GST groups after challenge with N. caninum tachyzoites. Taken together, our results suggest the efficacy of recombinant NcGRA6 for interrupting the vertical transmission of N. caninum in mice by reducing the severity of infections in dams and offspring.

Neospora caninum is an intracellular protozoan parasite affecting numerous animal species. It induces significant economic losses because of abortion and neonatal abnormalities in cattle. In case of infection, the parasite secretes numerous arsenals to establish a successful infection in the host cell. In the same context but for a different purpose, the host resorts to different strategies to eliminate the invading parasite. During this battle, numerous key factors from both parasite and host sides are produced and interact for the maintaining and vanishing of the infection, respectively. Although several reviews have highlighted the role of different compartments of the immune system against N. caninum infection, each one of them has mostly targeted specific points related to the immune component and animal host. Thus, in the current review, we will focus on effector molecules derived from the host cell or the parasite using a comprehensive survey method from previous reports. According to our knowledge, this is the first review that highlights and discusses immune response at the host cell–parasite molecular interface against N. caninum infection in different susceptible hosts.