Assessment of selected molecular factors and 17-ß estradiol dosage in response to Toxoplasma gondii infection in swine.

Background and Aim: Toxoplasma gondii is a global zoonotic parasite infecting virtually all warm-blooded species, although a species-specific variability is evident referring to symptoms frame. Both the success of T. gondii and the outcome of infection depend on a delicate balance between host cellular pathways and the evasion or modulation strategies elicited by the parasite. The hormonal and molecular mechanisms involved in this delicate host-parasite balance are still unclear, especially when considering intermediate host species other than mouse. This study aimed to assess any correlation between T. gondii infection and selected molecular and hormonal factors involved in responses to infection in susceptible species such as swine. Moreover, blood counts and hematochemical assays (glucose, total cholesterol, and triglycerides dosage) were performed to evaluate the overall health condition of animals. Materials and Methods: Toxoplasmosis was diagnosed by enzyme-linked immunosorbent assay for antibodies determination and real-time polymerase chain reaction (RT-PCR) for T. gondii DNA detection. Target genes coding for key factors of cell responses to T. gondii infection were selected, and their transcription was assessed in various tissues by quantitative RT-PCR. 17-ß estradiol concentrations were assessed by fluorimetric enzyme-linked immunoassay and the AIA-360 automated immunoassay analyzer. Blood count and hematochemical analyses were performed by a blood cell counter and a spectrophotometer, respectively. Results: The present research highlighted significant differences among infected and uninfected swine (control group) for both transcription profiles of some of the molecular factors considered and 17-ß estradiol concentrations. Referring to the assessed hematological and biochemical parameters, no statistically significant differences were observed in infected swine compared to the control group. Conclusion: Our results contribute to the enrichment of data available about the subject and could be useful for a deeper knowledge of the interaction between this parasite and its hosts. However, more aspects are still unclear, such as the effective response of downstream molecules from the same pathways to the variation of factors observed in this study either assessing how the same factors respond to Toxoplasma gondii infection in other host speciesand further analyses should be performed on other host species.

Edible Insects and Toxoplasma gondii: Is It Something We Need To Be Concerned About?

ABSTRACT: Novel foods, such as edible insects and food products on the basis of insects, could play an important role in both human and animal nutrition in the future. The identification of dangers associated with insect consumption is fundamental to guarantee consumer safety and adequate regulatory guidelines for operators of the food sector. Although former studies have focused on the microbiological contamination of fresh or processed edible insects, so far little information is available about the occurrence of foodborne parasites, such as Toxoplasma gondii, whose life cycles make them candidates for potential insect breeding substrate contamination. Hence, we investigated the presence of contaminating T. gondii in farmed edible insects to rule out this further hazard for consumers. Four species of insects most commonly used as food for human consumption were analyzed: mealworm; African migratory locust, house cricket, and silkworm. Samples included live specimens but also minimally (dehydrated) and highly processed edible insects. Traces of T. gondii DNA were detected in samples of dehydrated mealworm. These results highlight the need for implementing good farming and processing practices with particular care paid to safe storage and handling of feed and substrates used for edible insects to reduce the chance of T. gondii entering the human food chain.

Toxoplasma gondii in edible fishes captured in the Mediterranean basin.

The issue of whether market fish can be involved in the transmission of Toxoplasma gondii in the marine environment is highly debated since toxoplasmosis has been diagnosed frequently in cetaceans stranded along the Mediterranean coastlines in recent times. To support the hypothesis that fishes can harbour and effectively transmit the parasite to top-of-the-food-chain marine organisms and to human consumers of fishery products, a total of 1,293 fishes from 17 species obtained from wholesale and local fish markets were examined for T. gondii DNA. Real-time PCR was performed in samples obtained by separately pooling intestines, gills and skin/muscles collected from each fish species. Thirty-two out of 147 pooled samples from 12 different fish species were found contaminated with T. gondii DNA that was detected in 16 samples of skin/muscle and in 11 samples of both intestine and gills. Quantitative analysis of amplified DNA performed by both real-time PCR and digital PCR (dPCR) confirmed that positive fish samples were contaminated with Toxoplasma genomic DNA to an extent of 6.10 × 10-2 to 2.77 × 104  copies/ml (quantitative PCR) and of 1 to 5.7 × 104  copies/ml (dPCR). Fishes are not considered competent biological hosts for T. gondii; nonetheless, they can be contaminated with T. gondii oocysts flowing via freshwater run-offs (untreated sewage discharges, soil flooding) into the marine environment, thus acting as mechanical carriers. Although the detection of viable and infective T. gondii oocysts was not the objective of this investigation, the results here reported suggest that fish species sold for human consumption can be accidentally involved in the transmission route of the parasite in the marine environment and that the risk of foodborne transmission of toxoplasmosis to fish consumers should be further investigated.

Development and validation of a real-time PCR assay for the detection of Toxoplasma gondii DNA in animal and meat samples.

We report a rapid and reliable method for the detection of Toxoplasma gondii in meat and animal tissues based on real-time polymerase chain reaction (PCR). Samples were collected from cattle, small ruminants, horses, and pigs raised or imported into Sicily, Italy. All DNA preparations were assayed by real-time PCR tests targeted to a 98-bp long fragment in the AF 529-bp repeat element and to the B1 gene using specific primers. Diagnostic sensitivity (100%), diagnostic specificity (100%), limit of detection (0.01 pg), efficiency (92-109%), and precision (mean coefficient of variation = 0.60%), repeatability (100%), reproducibility (100%), and robustness were evaluated using 240 DNA extracted samples (120 positives and 120 negative as per the OIE nested PCR method) from different matrices. Positive results were confirmed by the repetition of both real-time and nested PCR assays. Our study demonstrates the viability of a reliable, rapid, and specific real-time PCR on a large scale to monitor contamination with Toxoplasma cysts in meat and animal specimens. This validated method can be used for postmortem detection in domestic and wild animals and for food safety purposes.