Heligmosomoides bakeri and Toxoplasma gondii co-infection leads to increased mortality associated with changes in immune resistance in the lymphoid compartment and disease pathology.

Co-infections are a common reality but understanding how the immune system responds in this context is complex and can be unpredictable. Heligmosomoides bakeri (parasitic roundworm, previously Heligmosomoides polygyrus) and Toxoplasma gondii (protozoan parasite) are well studied organisms that stimulate a characteristic Th2 and Th1 response, respectively. Several studies have demonstrated reduced inflammatory cytokine responses in animals co-infected with such organisms. However, while general cytokine signatures have been examined, the impact of the different cytokine producing lymphocytes on parasite control/clearance is not fully understood. We investigated five different lymphocyte populations (NK, NKT, γδ T, CD4+ T and CD8+ T cells), five organs (small intestine, Peyer's patches, mesenteric lymph nodes, spleen and liver), and 4 cytokines (IFN©, IL-4, IL-10 and IL-13) at two different time points (days 5 and 10 post T. gondii infection). We found that co-infected animals had significantly higher mortality than either single infection. This was accompanied by transient and local changes in parasite loads and cytokine profiles. Despite the early changes in lymphocyte and cytokine profiles, severe intestinal pathology in co-infected mice likely contributed to early mortality due to significant damage by both parasites in the small intestine. Our work demonstrates the importance of taking a broad view during infection research, studying multiple cell types, organs/tissues and time points to link and/or uncouple immunological from pathological findings. Our results provide insights into how co-infection with parasites stimulating different arms of the immune system can lead to drastic changes in infection dynamics.

Molecular Models of Toxoplasma gondii in Humans and Animals.

Toxoplasma gondii (T. gondii) is an obligate intracellular, zoonotic protozoan parasite of interest to physicians and veterinarians with its highly complex structure. It is known to infect about one-third of the world's population. Since it is a zoonotic disease, it is necessary to keep the animal population under control in order to prevent human exposure. Many studies have been conducted on the detection of T. gondii and it has been determined that there are three clonal groups consisting of types 1, 2, 3. Developments in molecular studies have led to changes in the taxonomy and new developments in parasitic diseases. It has helped in diagnosis, treatment, development of antiparasitic drugs and research on resistance. They also provided research on vaccine studies, genetic typing and phylogenetics of parasitic diseases. Conventional polymerase chain reaction (PCR), real-time PCR and genotyping studies conducted today increase our knowledge about T. gondii. Methods such as B1, SAG1, SAG2, GRA1, 529-bp repeat element, OWP genes and 18S rRNAs are mostly used in PCR, and methods such as MS, MLST, PCR-RFLP, RAPD-PCR and HRM are used in genotyping. Toxoplasmosis is a disease that is within the framework of the concept of one health and must attract attention, has not yet been eradicated in the world and needs joint studies for humans, animals and ecosystems to be eradicated. This can only be possible by establishing interdisciplinary groups, conducting surveys and training.

[Preliminary observation on the development and dynamic changes of chronic toxoplasmosis in mice].

OBJECTIVE: To investigate the development and dynamic changes of cysts in the brain of mice following infection with different forms of Toxoplasma gondii, so as to provide insights into for toxoplasmosis prevention and control. METHODS: ICR mice at ages of 6 to 8 weeks, each weighing 20 to 25 g, were intraperitoneally injected with tachyzoites of the T. gondii PRU strain at a dose of 1 × 105 tachyzoites per mouse, orally administered with cysts at a dose of 20 oocysts per mouse or oocysts at a dose of 200 oocysts per mouse for modeling chronic T. gondii infection in mice, and the clinical symptoms and survival of mice were observed post-infection. Mice were orally infected with T. gondii cysts at doses of 10 (low-dose group), 20 (medium-dose group), 40 cysts per mouse (high-dose group), and the effect of different doses of T. gondii infections on the number of cysts was examined in the mouse brain. Mice were orally administered with T. gondii cysts at a dose of 20 cysts per mouse, and grouped according to gender (female and male) and time points of infections (20, 30, 60, 90, 120, 150, 180 days post-infection), and the effects of gender and time points of infections on the number of cysts was examined in the mouse brain. In addition, mice were divided into the tachyzoite group (Group T), the first-generation cyst group (Group C1), the second-generation cyst group (Group C2), the third-generation cyst (Group C3) and the fourth-generation cyst group (Group C4). Mice in the Group T were intraperitoneally injected with T. gondii tachyzoites at a dose of 1 × 105 tachyzoites per mouse, and the cysts were collected from the mouse brain tissues 30 days post-infection, while mice in the Group C1 were orally infected with the collected cysts at a dose of 30 cysts per mouse. Continuous passage was performed by oral administration with cysts produced by the previous generation in mice, and the effect of continuous passage on the number of cysts was examined in the mouse brain. RESULTS: Following infection with T. gondii tachyzoites, cysts and oocysts in mice, obvious clinical symptoms were observed on days 6 to 13 and mice frequently died on days 7 to 12. The survival rates of mice were 67.0%, 87.0% and 53.0%, and the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0) and (581.0 ± 183.1) in the mouse brain (F = 11.94, P < 0.01) on day 30 post-infection with T. gondii tachyzoites, cysts and oocysts, respectively, and the numbers of cysts in the brain tissues were significantly lower in mice infected with T. gondii tachyzoites and oocysts than in those infected with cysts (all P values < 0.01). The survival rates of mice were 87.0%, 87.0% and 60.0%, and the mean numbers of cysts were (953.0 ± 355.5), (1 084.0 ± 474.3) and (1 113.0 ± 546.0) in the mouse brain in the low-, medium- and high-dose groups on day 30 post-infection, respectively (F = 0.42, P > 0.05). The survival rates of male and female mice were 73.0% and 80.0%, and the mean numbers of cysts were (946.4 ± 411.4) and (932.1 ± 322.4) in the brain tissues of male and female mice, respectively (F = 1.63, P > 0.05). Following continuous passage, the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0), (896.8 ± 332.3), (782.5 ± 423.9) and (829.2 ± 306.0) in the brain tissues of mice in the T, C1, C2, C3 and C4 groups, respectively (F = 4.82, P < 0.01), and the number of cysts was higher in the mouse brain in Group 1 than in Group T (P < 0.01). Following oral administration of 20 T. gondii cysts in mice, cysts were found in the moues brain for the first time on day 20 post-infection, and the number of cysts gradually increased over time, peaked on days 30 and 90 post-infection and then gradually decreased; however, the cysts were still found in the mouse brain on day 180 post-infection. CONCLUSIONS: There is a higher possibility of developing chronic T. gondii infection in mice following infection with cysts than with oocysts or tachyzoites and the most severe chronic infection is seen following infection with cysts. The number of cysts does not correlate with the severity of chronic T. gondii infection, and the number of cysts peaks in the mouse brain on days 30 and 90 post-infection.

Detection of Toxoplasma gondii infection in buffaloes (Bubalus bubalis) and cattle (Bos taurus) at the Tabriz abattoir, Iran.

BACKGROUND: Toxoplasma gondii is a widely prevalent zoonotic protozoan parasite in humans and warm-blooded animals worldwide. Infection of humans by this parasite can result in severe clinical symptoms, particularly in individuals with congenital toxoplasmosis or immunocompromised patients. Contamination mainly occurs through foodborne routes, especially the consumption of raw or undercooked meat from animals. OBJECTIVES: The aim of this study was to use PCR to detect T. gondii in tissues and organs of buffaloes and cattle slaughtered at Tabriz slaughterhouse, in Iran. METHODS: Fifty grams of heart, thigh, diaphragm and tongue from 50 buffaloes and 100 cattle slaughtered at the Tabriz industrial slaughterhouse were selected for sampling using a combination of convenience sampling. The samples were tested using a previously published PCR method. RESULTS: Out of the 150 animal samples, T. gondii was detected in 10 (6.7%, 95%CI: 3.2-11.9), including one buffalo (2%, 95%CI: 0.1-10.6) and nine cattle (9%, 95%CI: 4.2-16.4). There was no statistically significant difference in the rate of T. gondii infection among cattle based on age and sex (p > 0.05). CONCLUSIONS: The results indicated a potential risk of T. gondii transmission to humans through the consumption of infected meat. Therefore, appropriate and effective preventive measures should be taken to limit the transmission of this parasite to humans, and the consumption of raw and undercooked meat should be discouraged.

Early immune response to Toxoplasma gondii lineage III isolates of different virulence phenotype.

Introduction: Toxoplasma gondii is an intracellular parasite of importance to human and veterinary health. The structure and diversity of the genotype population of T. gondii varies considerably with respect to geography, but three lineages, type I, II and III, are distributed globally. Lineage III genotypes are the least well characterized in terms of biology, host immunity and virulence. Once a host is infected with T.gondii, innate immune mechanisms are engaged to reduce the parasite burden in tissues and create a pro-inflammatory environment in which the TH1 response develops to ensure survival. This study investigated the early cellular immune response of Swiss-Webster mice post intraperitoneal infection with 10 tachyzoites of four distinct non-clonal genotypes of lineage III and a local isolate of ToxoDB#1. The virulence phenotype, cumulative mortality (CM) and allele profiles of ROP5, ROP16, ROP18 and GRA15 were published previously. Methods: Parasite dissemination in different tissues was analyzed by real-time PCR and relative expression levels of IFNγ, IL12-p40, IL-10 and TBX21 in the cervical lymph nodes (CLN), brain and spleen were calculated using the ΔΔCt method. Stage conversion was determined by detection of the BAG1 transcript in the brain. Results: Tissue dissemination depends on the virulence phenotype but not CM, while the TBX21 and cytokine levels and kinetics correlate better with CM than virulence phenotype. The earliest detection of BAG1 was seven days post infection. Only infection with the genotype of high CM (69.4%) was associated with high T-bet levels in the CLN 24 h and high systemic IFNγ expression which was sustained over the first week, while infection with genotypes of lower CM (38.8%, 10.7% and 6.8%) is characterized by down-regulation and/or low systemic levels of IFNγ. The response intensity, as assessed by cytokine levels, to the genotype of high CM wanes over time, while it increases gradually to genotypes of lower CM. Discussion: The results point to the conclusion that the immune response is not correlated with the virulence phenotype and/or allele profile, but an early onset, intense pro-inflammatory response is characteristic of genotypes with high CM. Additionally, high IFNγ level in the brain may hamper stage conversion.

Altered landscape of total RNA, tRNA and sncRNA modifications in the liver and spleen of mice infected by Toxoplasma gondii.

BACKGROUND: Pathogens can impact host RNA modification machinery to establish a favorable cellular environment for their replication. In the present study, we investigated the effect of Toxoplasma gondii infection on host RNA modification profiles and explored how these modifications may influence the host-parasite interaction. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed the modification levels of ∼ 80 nt tRNA and 17-50 nt sncRNAs in mouse liver, spleen, and serum using liquid chromatography and tandem mass spectrometry analysis. The results revealed alterations in RNA modification profiles, particularly during acute infection. The liver exhibited more differentially abundant RNA modifications than the spleen. RNA modification levels in serum were mostly downregulated during acute infection compared to control mice. Correlations were detected between different RNA modifications in the liver and spleen during infection and between several RNA modifications and many cytokines. Alterations in RNA modifications affected tRNA stability and protein translation. CONCLUSIONS/SIGNIFICANCE: These findings provide new insight into the role of RNA modifications in mediating the murine host response to T. gondii infection.

Molecular detection of Toxoplasma gondii in domestic and wild guinea pigs (Cavia spp.) from the Marangani district in Cuzco, Peru.

Toxoplasmosis is a foodborne disease caused by the protozoan Toxoplasma gondii, and transmitted to humans by eating raw or undercooked meat, mainly. Poultry, beef, and pork are the main meats consumed in Peru; despite this, guinea pig meat is also widely consumed. For this reason, the objective of this study was to molecularly detect T. gondii in domestic and wild guinea pigs from the Marangani district in Cuzco, Peru, and identify some risk factors associated with this pathogen. DNA was extracted from the brain tissue samples of guinea pigs (30 domestic and 30 wild), and PCR protocols were used to amplify the internal transcribed spacer (ITS-1) region and a 529 bp fragment from the T. gondii genome. T. gondii DNA was detected in 14 (23.3%) guinea pigs. T. gondii frequency was 33.3% in domestic guinea pigs and 13.3% in wild guinea pigs. Our results demonstrated that guinea pigs represent an important source for T. gondii infection in human populations in this locality.

Differential transcriptome study on the damage of testicular tissues caused by chronic infection of T. gondii in mice.

BACKGROUND: Toxoplasma gondii is an intracellular protozoan parasite that is widely distributed in humans and warm-blooded animals. T. gondii chronic infections can cause toxoplasmic encephalopathy, adverse pregnancy, and male reproductive disorders. In male reproduction, the main function of the testis is to provide a stable place for spermatogenesis and immunological protection. The disorders affecting testis tissue encompass abnormalities in the germ cell cycle, spermatogenic retardation, or complete cessation of sperm development. However, the mechanisms of interaction between T. gondii and the reproductive system is unclear. The aims were to study the expression levels of genes related to spermatogenesis, following T. gondii infection, in mouse testicular tissue. METHODS: RNA-seq sequencing was carried out on mouse testicular tissues from mice infected or uninfected with the T. gondii type II Prugniaud (PRU) strain and validated in combination with real-time quantitative PCR and immunofluorescence assays. RESULTS: The results showed that there were 250 significant differentially expressed genes (DEGs) (P < 0.05, |log2fold change| â‰§ 1). Bioinformatics analysis showed that 101 DEGs were annotated to the 1696 gene ontology (GO) term. While there was a higher number of DEGs in the biological process classification as a whole, the GO enrichment revealed a significant presence of DEGs in the cellular component classification. The Arhgap18 and Syne1 genes undergo regulatory changes following T. gondii infection, and both were involved in shaping the cytoskeleton of the blood-testis barrier (BTB). The number of DEGs enriched in the MAPK signaling pathway, the ERK1/2 signaling pathway, and the JNK signaling pathway were significant. The PTGDS gene is located in the Arachidonic acid metabolism pathway, which plays an important role in the formation and maintenance of BTB in the testis. The expression of PTGDS is downregulated subsequent to T. gondii infection, potentially exerting deleterious effects on the integrity of the BTB and the spermatogenic microenvironment within the testes. CONCLUSIONS: Overall, our research provides in-depth insights into how chronic T. gondii infection might affect testicular tissue and potentially impact male fertility. These findings offer a new perspective on the impact of T. gondii infection on the male reproductive system.

The Toxoplasma oxygen-sensing protein, TgPhyA, is required for resistance to interferon gamma-mediated nutritional immunity in mice.

As Toxoplasma gondii disseminates through its host, the parasite must sense and adapt to its environment and scavenge nutrients. Oxygen (O2) is one such environmental factor and cytoplasmic prolyl 4-hydroxylases (PHDs) are evolutionarily conserved O2 cellular sensing proteins that regulate responses to changes in O2 availability. Toxoplasma expresses 2 PHDs. One of them, TgPHYa hydroxylates SKP1, a subunit of the SCF-E3 ubiquitin ligase complex. In vitro, TgPHYa is important for growth at low O2 levels. However, studies have yet to examine the role that TgPHYa or any other pathogen-encoded PHD plays in virulence and disease. Using a type II ME49 Toxoplasma TgPHYa knockout, we report that TgPHYa is important for Toxoplasma virulence and brain cyst formation in mice. We further find that while TgPHYa mutant parasites can establish an infection in the gut, they are unable to efficiently disseminate to peripheral tissues because the mutant parasites are unable to survive within recruited immune cells. Since this phenotype was abrogated in IFNγ knockout mice, we studied how TgPHYa mediates survival in IFNγ-treated cells. We find that TgPHYa is not required for release of parasite-encoded effectors into host cells that neutralize anti-parasitic processes induced by IFNγ. In contrast, we find that TgPHYa is required for the parasite to scavenge tryptophan, which is an amino acid whose levels are decreased after IFNγ up-regulates the tryptophan-catabolizing enzyme, indoleamine dioxygenase (IDO). We further find, relative to wild-type mice, that IDO knockout mice display increased morbidity when infected with TgPHYa knockout parasites. Together, these data identify the first parasite mechanism for evading IFNγ-induced nutritional immunity and highlight a novel role that oxygen-sensing proteins play in pathogen growth and virulence.

RPA-CRISPR/Cas12a-LFA combined with a digital visualization instrument to detect Toxoplasma gondii in stray dogs and cats in Zhejiang province, China.

Toxoplasma gondii, which causes toxoplasmosis, is prevalent in warm-blooded animals, such as cats, dogs, and humans. T. gondii causes economic losses to livestock production and represents a potential risk to public health. Dogs and cats are common hosts in the epidemiology of toxoplasmosis. The current molecular diagnostic tools for T. gondii infection require high technical skills, a laboratory environment, and complex instruments. Herein, we developed a recombinase polymerase amplification (RPA)-clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 12a (Cas12a) assay to detect T. gondii. The lowest limit of detection of the assay was 31 copies/µL for the T. gondii B1 gene. In addition, we established a visual RPA-CRISPR/Cas12a lateral flow band assay (RPA-CRISPR/Cas12a-LFA) combined with a digital visualization instrument, which minimized the problem of false-negative results for weakly positive samples and avoided misinterpretation of the results by the naked eye, making the LFA assay results more accurate. The assay established in this study could identify T. gondii within 55 min with high accuracy and sensitivity, without cross-reaction with other tested parasites. The developed assay was validated by establishing a mouse model of toxoplasmosis. Finally, the developed assay was used to investigate the prevalence of T. gondii in stray cats and dogs in Zhejiang province, Eastern China. The positive rates of T. gondii infection in stray cats and dogs were 8.0% and 4.0%, respectively. In conclusion, the RPA-CRISPR/Cas12a-LFA is rapid, sensitive, and accurate for the early diagnosis of T. gondii, showing promise for on-site surveillance. IMPORTANCE: Toxoplasma gondii is a virulent pathogen that puts millions of infected people at risk of chronic disease reactivation. Hosts of T. gondii are distributed worldwide, and cats and dogs are common hosts of T. gondii. Therefore, rapid diagnosis of early T. gondii infection and investigation of its prevalence in stray dogs and cats are essential. Here, we established a visual recombinase polymerase amplification-clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 12a-assay combined with a lateral flow band assay and a digital visualization instrument. Detailed analyses found that the assay could be used for the early diagnosis of T. gondii without false-negative results. Moreover, we detected the prevalence of T. gondii in stray cats and dogs in Zhejiang province, China. Our developed assay provides technical support for the early diagnosis of T. gondii and could be applied in prevalence surveys of T. gondii in stray dogs and cats.

Toxoplasma gondii infection in European brown hares (Lepus europaeus) in Brazil.

Toxoplasmosis is a worldwide zoonosis that affects warm-blooded animals, including humans. Wild animals can act as intermediate hosts of this pathogen; thus, this study aims to detect Toxoplasma gondii infection in invasive European brown hares in Brazil. For this, 72 wild European brown hares were captured from July 2020 to June 2022 in three Brazilian states: São Paulo, Paraná, and Rio Grande do Sul. The diagnostic of Toxoplasma gondii infection was performed by bioassay in mouse, histopathology in Hematoxylin-Eosin-stained tissue sections (brain, liver, lungs, kidneys, and small intestine), serology by IFAT, and molecular techniques by conventional PCR and qPCR. The combined prevalence of the different diagnostic methods was 51.4% (37/72, CI= 40.1 - 62.6 %), and there was no statistical difference between sexes, age range, or geographical region of the hosts. Mouse bioassay was the technique that detected more positive hares. To our knowledge, this is the first confirmation of Toxoplasma gondii infection in invasive European brown hares in Brazil. These animals act as reservoirs and potential infection source for carnivores and other wild and domestic animals, including humans, thus contributing to perpetuate the disease cycle in São Paulo, Paraná, and Rio Grande do Sul States. Research such as the present study is necessary to raise awareness about the role of animals in the disease cycle.

Seroprevalence and risk factors of Toxoplasma gondii infection among goats in Algeria.

Toxoplasmosis is one of the most common zoonotic parasitic diseases worldwide and is caused by Toxoplasma gondii. It is implicated in reproductive disorders in small ruminants. This study aims to determine, for the first time in Algeria, the seroprevalence and associated factors of T. gondii infection in goats. The study was conducted in four regions, Ghardaia, Laghouat and Djelfa, southern Algeria, and Jijel region, northern Algeria. A total of 92 blood samples were collected including 74 females and 18 males. All sera were tested using an enzyme-linked immunosorbent assay (ELISA) to detect the T. gondii antibodies. The presence of anti-T. gondii antibodies was detected in 35 out of 92 goats (38.04%) (95% CI: 31.64%-44.44%) and in all flocks (100%). Risk factors that have a significant influence on the seroprevalence of T. gondii infection are breed, regions, production system, presence of cats, clinics and abortion history. However, variables such as age and gender were note significantly associated with toxoplasma infection in goats. The highest seroprevalences of infection was observed in saanen (52.94%) (p<0.001) and cross-breed race (44%) (p<0.01) in comparison with other breeds. Regarding regions, Jijel and Laghouat were most infected with seroprevalences of 50% (p<0.001) and 40.91% (p<0.01), respectively. Animals in intensive production systems were most infected, showing a seroprevalence of 51.85%, in comparison with extensive (28.13%) and semi-intensive systems (36.36%) (p<0.001). The presence of cats in farms was significantly associated with high seroprevalence (44.64%) (p<0.001). The infection was more prevalent in previously aborted females (50%) than females that had never aborted (3.35%) (p<0.001)and animals that have diarrhoea or poor health (41.67%) were significantly more infected than healthy animals (37.50%) (p<0.01). Seroprevalence in males (38.89%) was very close to those in females (37.84%) (p>0.05). Age-related seroprevalence did not vary significantly (ranged from 36.37% to 40%) between the three age classes. These results indicate that goat toxoplasmosis is widespread in Algeria, and goats may represent a high risk of contamination for humans. This requires more attention during consumption of goat meat.

Construction of Toxoplasma gondii SRS29C nucleic acid vaccine and comparative immunoprotective study of an SRS29C and SAG1 combination.

Toxoplasma gondii is an intracellular protozoan parasite that infects all nucleated cells except the red blood cells. Currently, nucleic acid vaccines are being widely investigated in Toxoplasma gondii control, and several nucleic acid vaccine candidate antigens have shown good protection in various studies. The aim of this study was to construct a nucleic acid vaccine with Toxoplasma gondii SRS29C as the target gene. We explored the nucleic acid vaccine with Toxoplasma surface protein SRS29C and the combined gene of SRS29C and SAG1 and evaluated its immunoprotective effect against Toxoplasma gondii. To amplify the gene fragment and clone it to the expression vector, the recombinant plasmid pEGFP-SRS29C was constructed by PCR. Eukaryotic cells were transfected with the plasmid, and the expression of the target protein was assessed using the Western blot method. The level of serum IgG was determined via ELISA, and the splenic lymphocyte proliferation ability was detected using the CCK-8 method. The percentages of CD4+ and CD8+ T cells were measured by flow cytometry. Mice were immunised three times with single-gene nucleic acid vaccine and combination vaccine. Splenic lymphocytokine expression was determined using ELISA kits. The mice's survival time was monitored and recorded during an in vivo insect assault experiment, and the vaccine's protective power was assessed. The outcomes showed that PCR-amplification of an SRS29C gene fragment was successful. The 4,733-bp vector fragment and the 1,119-bp target segment were both recognised by double digestion. Additionally, after transfection of the recombinant plasmid pEGFP-SRS29C, Western blot examination of the extracted protein revealed the presence of a target protein strip at 66 kDa. The test results demonstrated that the IgG content in the serum of the pEGFP-SRS29C group and the co-immunization group was significantly higher than that of the PBS group and the empty vector group. The IgG potency induced by the co-immunization group was higher than that of the pEGFP-SRS29C group and the pEGFP-SAG1 group, the number of splenic lymphocyte proliferation number was higher than that of the PBS group and the empty vector group. The CD4+/CD8+ T ratio was higher than that of the PBS group and the empty vector group. The expression of IFN-γ and TNF-α in the splenocytes of the pEGFP-SRS29C group and the combined immunisation group was significantly higher following antigen stimulation. In the worm attack experiments, mice in the PBS and empty vector groups perished within 9 days of the worm attack, whereas mice in the pEGFP-SRS29C group survived for 18 days, mice in the pEGFP-SAG1 group survived for 21 days, and mice in the co-immunization group survived for 24 days. This demonstrates that the constructed Toxoplasma gondii nucleic acid vaccine pEGFP-SRS29C and the combined gene vaccine can induce mice to develop certain humoral and cellular immune responses, and enhance their ability to resist Toxoplasma gondii infection.

Toxoplasma gondii exposure in wildlife in Spain: Is there any predictable threat for humans and domestic animals?

Toxoplasmosis is a parasitic zoonosis of key importance in veterinary and public health. This article summarizes the available data (from 2000 to 2023) of exposition to Toxoplasma gondii in wildlife species in Spain based on a systematic bibliographic search, as well as further analysis of its potential relationship with environmental variables, biodiversity, anthropogenic impact on the habitat, and the reported human cases of toxoplasmosis. The overall seroprevalence of T. gondii in carnivorous mammals, birds, ungulate and lagomorph species in Spain was estimated at 69.3 %, 36.4 %, 18.4 %, and 16.2 %, respectively. Among the studies considered, great heterogeneity was observed both between and within taxonomic groups [Cohen's d > 0.8; X2 = 1039.10, df = 4 (p < 0.01) I2 = 97 %, r2 = 1.88, (p < 0.001)] and between and within bioregions [Cohen's d > 0.5; X2 = 368.59, df = 4 (p < 0.01)]. The results of a generalized linear model explaining T. gondii seroprevalence in wild animals suggest the influence of abiotic variables [wetland (p < 0.001), unvegetated (p < 0.001), isothermality (p < 0.001), and mean temperature during wettest quarter (p < 0.05)] and number of intermediate host species as positively associated with increased exposure of wildlife to T. gondii (p < 0.01). Toxoplasma gondii DNA was detected in both wild birds and wild mammals (range: 0.0-51.2 %) mainly from north-centre, northeast, and central-west of Spain. Regarding hospitalisation rates due to toxoplasmosis in humans, some abiotic variables [permanent crops (p < 0.05) and mean temperature during wettest quarter (p < 0.05)] showed a positive association. Despite certain limitations, this research evidences a substantial gap of knowledge on the implication of wildlife in the life cycle of T. gondii in Spain. This lack of knowledge is particularly evident in areas where the human-livestock-wildlife interface overlaps, preventing us from accurately determining its true distribution in different habitats, as well as its potential direct or indirect implications on public and veterinary health.

A combination of GRA3, GRA6 and GRA7 peptides offer a useful tool for serotyping type II and III Toxoplasma gondii infections in sheep and pigs.

The clinical consequences of toxoplasmosis are greatly dependent on the Toxoplasma gondii strain causing the infection. To better understand its epidemiology and design appropriate control strategies, it is important to determine the strain present in infected animals. Serotyping methods are based on the detection of antibodies that react against segments of antigenic proteins presenting strain-specific polymorphic variations, offering a cost-effective, sensitive, and non-invasive alternative to genotyping techniques. Herein, we evaluated the applicability of a panel of peptides previously characterized in mice and humans to serotype sheep and pigs. To this end, we used 51 serum samples from experimentally infected ewes (32 type II and 19 type III), 20 sheep samples from naturally infected sheep where the causative strain was genotyped (18 type II and 2 type III), and 40 serum samples from experimentally infected pigs (22 type II and 18 type III). Our ELISA test results showed that a combination of GRA peptide homologous pairs can discriminate infections caused by type II and III strains of T. gondii in sheep and pigs. Namely, the GRA3-I/III-43 vs. GRA3-II-43, GRA6-I/III-213 vs. GRA6-II-214 and GRA6-III-44 vs. GRA6-II-44 ratios showed a statistically significant predominance of the respective strain-type peptide in sheep, while in pigs, in addition to these three peptide pairs, GRA7-II-224 vs. GRA7-III-224 also showed promising results. Notably, the GRA6-44 pair, which was previously deemed inefficient in mice and humans, showed a high prediction capacity, especially in sheep. By contrast, GRA5-38 peptides failed to correctly predict the strain type in most sheep and pig samples, underpinning the notion that individual standardization is needed for each animal species. Finally, we recommend analyzing for each animal at least 2 samples taken at different time points to confirm the obtained results.

Geoepidemiology, seroprevalence and factors associated with Toxoplasma gondii infection in domicilied cats from Paraíba (Brazil).

Toxoplasma gondii is a parasite responsible for toxoplasmosis, an emerging and often neglected zoonosis in South America, particularly Brazil. Felines, the only definitive hosts, excrete oocysts in their feces, potentially infecting all homeotherms. Domestic cats are primarily responsible for contaminating human environments with these oocysts. Monitoring their populations is therefore essential to ensure proper toxoplasmosis prophylaxis. The aim of this study was to estimate the prevalence of T. gondii and exposure factors in a population of owner cats in the city of João Pessoa, Paraíba, Brazil. A total of 267 blood samples were collected from domestic cats aged between 1 and 15 years and tested with an immunofluorescence antibody test. The seroprevalence of antibodies against T. gondii was only 17.22% (46/267 individuals). This result therefore suggests a low contribution of domestic cats to T. gondii contamination of the urban environment. The cats' age and living environment were identified as risk factors for cat exposure to T. gondii.

Title: Géoépidémiologie, séroprévalence et facteurs associés à l'infection à Toxoplasma gondii chez les chats domiciliés à Paraíba (Brésil). Abstract: Toxoplasma gondii est le parasite responsable de la toxoplasmose, une zoonose émergente et souvent négligée en Amérique du Sud, notamment au Brésil. Les félins, seuls hôtes définitifs, excrètent des oocystes dans leurs selles, infectant potentiellement tous les homéothermes. Les chats domestiques sont les premiers responsables de la contamination des environnements humains avec ces oocystes. La surveillance de leurs populations est donc essentielle pour garantir une prophylaxie adéquate contre la toxoplasmose. Le but de cette étude était d'estimer la prévalence de T. gondii et les facteurs d'exposition dans une population de chats domestiques de la ville de João Pessoa, Paraíba, Brésil. Au total, 267 échantillons de sang ont été prélevés sur des chats domestiques âgés de 1 à 15 ans et testés avec un test d'immunofluorescence des anticorps. La séroprévalence des anticorps contre T. gondii n'était que de 17,22 % (46/267 individus). Ce résultat suggère donc une faible contribution des chats domestiques à la contamination du milieu urbain par T. gondii. L'âge et le milieu de vie des chats ont été identifiés comme facteurs de risque d'exposition du chat à T. gondii.

First report on prevalence, molecular characterization and phylogenetic study of Toxoplasma gondii infecting sheep of the Malakand Division of Pakistan.

BACKGROUND: Toxoplasma gondii is an apicomplexan protozoan parasite that infects one-third of the population of the world, including humans, animals, birds, and other vertebrates. The present investigation is the first molecular attempt in the Malakand Division of Pakistan to determine the epidemiology and phylogenetic study of Toxoplasma gondii infecting small ruminants. METHODOLOGY: A total of (N = 450) blood samples of sheep were randomly collected during the study period (December 2020 to November 2021), and DNA detection was done using PCR by amplifying ITS-1 genes. SPSS.20 and MEGA-11 software were used for statistical significance and phylogenetic analysis. RESULTS: The overall prevalence of T. gondii infection among sheep was 14.44 % (65/450). A high infection rate was found in more than five-year-olds at 18.33 % (11/60). Sequencing and BLAST analysis of PCR-positive samples confirmed the presence of T. gondii. Randomly, three isolates were sequenced and submitted to GenBank under accession numbers (PP028089-PP028091), respectively. The BLAST analysis of the obtained sequences based on the ITS-1 gene showed 99 % similarities with reported genotypes found in goats of Malakand, Pakistan (PP028089) and dogs of Brazil (MF766454). The study concludes that T. gondii is notably prevalent among the sheep population in the region, emphasizing the significant role of risk factors in disease transmission across animals and potentially to humans. Further research, zoonotic potential analysis, and targeted control measures are warranted to address and manage this parasitic infection effectively.

An investigation into Toxoplasma gondii at the human-livestock-wildlife interface, South Africa.

Interface areas shared by humans, domestic and wild animals may serve as high transmission contexts for Toxoplasma gondii. However, knowledge about the epidemiology of T. gondii in such areas is currently limited. The present study assessed the seroprevalence of T. gondii in different hosts from Mpumalanga, South Africa. Furthermore, we investigated the local knowledge and related practices about T. gondii by conducting a questionnaire study in the community. Blood samples were obtained and analysed for T. gondii antibodies using a commercial multispecies latex agglutination kit. The seroprevalence detected in humans (n = 160; patients showing signs of acute febrile illness), cats (n = 9), chickens (n = 336) and goats (n = 358) was 8.8%, 0.0%, 4.2% and 11.2%, respectively. Seroprevalence in impalas (n = 97), kudus (n = 55), wild dogs (n = 54), wildebeests (n = 43), warthogs (n = 97) and zebras (n = 68) was calculated at 5.2%, 7.3%, 100.0%, 20.9%, 13.4% and 9.1%, respectively. The questionnaire revealed that 63.0% of household owners were subsistence farmers, and 35.9% were pet owners. A high level of female participation was found (75.3%) when compared to male participation (24.7%). The results show a low circulation of T. gondii in the domestic cycle and suggest the presence of possible bridges between the wildlife cycle and the surrounding domestic cycle.Contribution: The study contributes to identifying transmission patterns and risk factors of T. gondii within human and animal populations. This topic fits within the scope of the journal presenting original research in veterinary science, with the focus on wild and domestic populations on the African continent on a topic of universal importance.

Seroprevalence and risk factors of Toxoplasma gondii in goats (Capra hircus) in the state of Espírito Santo, Southeast Brazil.

Toxoplasma gondii is described as a potential cause of abortion in goats and as a threat to public health. To estimate the prevalence of goats infected by T. gondii, in different cities in the Espírito Santo State, and to identify possible risk factors for infection a serological study was conducted. A total of 146 goat serum samples from the cities of Cariacica, Serra and Vila Velha were analyzed. The presence of IgG Class Immunoglobulins was serologically evaluated by Immunofluorescence antibody test (IFAT) and by Enzyme-linked Immunosorbent Assay (ELISA). The seroprevalence of anti-T. gondii was 46.6% (68/146) in both techniques and the same samples got the same results in both techniques. Among the analyzed sera, 70.6% (48/68) exhibited high-avidity IgG antibodies, and 29.4% (20/68) exhibited low-avidity IgG antibodies, suggesting that the infection was chronic in the infected animals. Female sex, age group over two years old, water from the public supply system, storage of food and supplies in an open and unprotected place, and the presence of a domestic cat on the property were identified as risk factors for T. gondii infection in goats. The state of Espirito Santo has a high frequency of infected goats, and this is the first research on caprine toxoplasmosis seroepidemiology in that region.

Molecular prevalence, phylogeny and hematological impact of Toxoplasma gondii and Plasmodium spp. in common quails from Punjab, Pakistan.

This study investigates the molecular prevalence and phylogenetic characteristics of two prominent blood-borne pathogens, Toxoplasma gondii (T. gondii) and Plasmodium spp., in common quails (Coturnix coturnix) sampled from both wild (N = 236) and farmed (N = 197) populations across four districts (Layyah, Dera Ghazi Khan, Lahore, and Multan) in Punjab, Pakistan, during the hunting seasons from 2021 to 2023. Additionally, the impact of these pathogens on the complete blood count (CBC) of the hosts is examined. Out of 433 quails tested, 25 (5.8%) exhibited amplification of the internal transcribed spacer (ITS-1) gene for T. gondii, while 15 (3.5%) showed amplification of the Cytochrome b gene for Plasmodium spp. A risk factor analysis indicated that the prevalence of both pathogens was not confined to specific sampling sites or bird sexes (P > 0.05). District-wise analysis highlighted that hens were more susceptible to both T. gondii and Plasmodium spp. infections than cocks. Wild quails exhibited a higher susceptibility to T. gondii compared to farmed birds. Significant CBC variations were recorded in infected birds as compared to uninfected ones. BLAST analysis of generated sequences has confirmed the identity of recovered PCR amplicons as T. gondii and Plasmodium relictum. Phylogenetic analysis revealed that Pakistani isolates clustered with those reported from various countries globally. This study provides the first documentation of T. gondii and Plasmodium sp. infections in Pakistani quails, underscoring the need for detailed investigations across different regions to enhance our understanding of infection rates and the zoonotic potential of these parasites.