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The protozoan Toxoplasma gondii infects virtually all warm-blooded animals, including bats. Depending on the diet, bats are classified as frugivorous, insectivorous, omnivorous, hematophagous, nectarivorous and carnivorous. The seroprevalence of T. gondii was higher in insectivores bats than fructivores bats. Owls, eagles, and cats can be predators of bats. Bats may be important in the epidemiology of T. gondii because they can be sentinels and can spread infection. Viable T. gondii has been isolated from brain, heart and pectoral muscle of bats. ToxoDB genotypes #9, #10, #6, #19, #69, #162 were identified from bats tissues. Genotypes #9 is prevalent in Asia. Genotypes #6 is widely distributed across Africa and Brazil. This result was matched with genotypes distribution from other hosts. The present review summarizes worldwide information on the seroprevalence, molecular epidemiology, isolation, genotypes and clinical cases of T. gondii infection in bats. Further studies are needed to verify the validity of serological and molecular tests, and the transmission routes of T. gondii infection in bats., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The morbidity due to congenital toxoplasmosis in humans is very high. Most of these infected children are likely to develop symptoms of clinical toxoplasmosis. Sequelae in fetus resulting from Toxoplasma gondii infections in women who become infected with this parasite during pregnancy can be devastating and enormous efforts are directed in some countries to prevent these consequences. Here, an update on congenital toxoplasmosis in humans, especially the rate of congenital infections in humans worldwide, is provided. Although several countries have surveillance programmes, most information on the rate of congenital transmission is from France and Brazil. Because of compulsory national screening programme in France to detect and treat women with recently acquired T. gondii infection with anti-toxoplasma therapy, the rate of congenital transmission and the severity of disease in children are declining. Infections by this parasite are widely prevalent in Brazil. The severity of clinical toxoplasmosis in Brazilian children is very high and may be associated with the genetic characteristics of T. gondii isolates prevailing in animals and humans in Brazil. Virtually little or no information is available on this topic from China, India and other countries in Asia.

Background: Toxoplasma gondii infections are common in humans and animals worldwide. Among all intermediate hosts of T. gondii, captive marsupials from Australia and New Zealand are highly susceptible to clinical toxoplasmosis. However, most free-range marsupials establish chronic T. gondii infection. Infected marsupial meat may serve as a source of T. gondii infection for humans. Differences in mortality patterns in different species of kangaroos and other marsupials are not fully understood. Lifestyle, habitat, and the genotype of T. gondii are predicted to be risk factors. For example, koalas are rarely exposed to T. gondii because they live on treetops whereas wallabies on land are frequently exposed to infection., Methods: The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, and genetic diversity of T. gondii infecting Australasian marsupials in their native habitat and among exported animals over the past decade. The role of genetic types of T. gondii and clinical disease is discussed., Results: Fatal toxoplasmosis has been diagnosed in captive Australasian marsupials in Argentina, Chile, China, Germany, Hungary, Japan, Spain, Turkey, and the USA. Most deaths occurred because of disseminated toxoplasmosis. Genetic characterization of T. gondii strains isolated from fatal marsupial infections identified Type III as well as atypical, nonclonal genotypes. Fatal toxoplasmosis was also diagnosed in free-ranging wombats (Vombatus ursinus) in Australia. Genetic characterization of DNA amplified directly from host tissues of subclinical culled kangaroos at slaughter identified many mixed-strain infections with both atypical and recombinant genotypes of T. gondii., Conclusions: Most Australasian marsupials in their native land, Australia and New Zealand, have high prevalence of T. gondii, and kangaroo meat can be a source of infection for humans if consumed uncooked/undercooked. The genotypes prevalent in kangaroos in Australia and New Zealand were genetically distinct from those isolated or genotyped from most macropods in the USA and other countries. Thus, clinical toxoplasmosis in marsupials imported from Australia is most likely to occur from infections acquired after importation.

MicroRNAs are molecules belonging to an evolutionarily conserved family of small non-coding RNAs, which act on post-transcriptional gene regulation, causing messenger RNA (mRNA) degradation or inhibiting mRNA translation into proteins. These molecules represent potential biomarkers for diagnosis, non-invasive prognosis, and monitoring the development of the disease. Moreover, they may provide additional information on the pathophysiology of parasitic infections and guide strategies for treatment. The Apicomplexan parasite Toxoplasma gondii modifies the levels of microRNAs and mRNAs in infected host cells by modulating the innate and adaptive immune responses, facilitating its survival within the host. Some studies have shown that microRNAs are promising molecular markers for developing diagnostic tools for human toxoplasmosis. MicroRNAs can be detected in human specimens collected using non-invasive procedures. changes in the circulating host microRNAs have been associated with T. gondii infection in mice and ocular toxoplasmosis in humans. Besides, microRNAs can be amplified from samples using sensitive and molecular-specific approaches such as real-time PCR. This review presents recent findings of the role that microRNAs play during T. gondii infection and discuss their potential use of these small nuclei acid molecules to different approaches such as laboratory diagnosis, modulation of cell and tissue infected as other potential applications in human toxoplasmosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 de Faria Junior, Murata, Lorenzi, Castro, Assoni, Ayo, Brandão and de Mattos.)

Toxoplasma gondii infections are common in humans and animals worldwide. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, and genetic diversity of T. gondii infections in bears. Seroprevalence estimates of T. gondii in black bears (Ursus americanus) are one of the highest of all animals. In Pennsylvania, seroprevalence is around 80% and has remained stable for the past 4 decades. Approximately 3,500 bears are hunted yearly in Pennsylvania alone. The validity of different serological tests is discussed based on bioassay and serological comparisons. Seroprevalence in grizzly bears (Ursus arctos) is lower than that in black bears. Even polar bears (Ursus maritimus) are infected; infections in these animals are ecologically interesting because of the absence of felids in the Arctic. Clinical toxoplasmosis in bears is rare and not documented in adult animals. The few reports of fatal toxoplasmosis in young bears need confirmation. Viable T. gondii has been isolated from black bears and a grizzly bear. The genetic diversity of isolates based on DNA from viable T. gondii isolates is discussed. Genetic typing of a total of 26 T. gondii samples from bears using 10 PCR-RFLP markers revealed 8 PCR-RFLP ToxoDB genotypes: #1 (clonal type II) in 3 samples, #2 (clonal type III) in 8 samples, #4 (haplogroup 12) in 3 samples, #5 (haplogroup 12) in 3 samples, #74 in 5 samples, #90 in 1 sample, #147 in 1 sample, and #216 in 2 samples. These results suggest relatively high genetic diversity of T. gondii in bears. Overall, T. gondii isolates in bears range from those circulating in a domestic cycle (genotypes #1 and #2) to those mainly associated with wildlife (such as genotypes #4 and #5, together known as haplogroup 12). A patient who acquired clinical Trichinella spiralis infection after eating undercooked bear meat also acquired T. gondii infection. Freezing of infected meat kills T. gondii, including the strains isolated from bears., (© American Society of Parasitologists 2021.)

Toxoplasma gondii infections are common in humans and animals worldwide. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, diagnosis, and genetic diversity of T. gondii in non-human primates (NHP) for the past decade. Seroprevalence estimates of T. gondii worldwide were tabulated for each host. Risk factors associated with T. gondii infections are evaluated. New World NHP in captivity are highly susceptible to T. gondii infection with high mortality associated with disseminated toxoplasmosis. T. gondii can be transmitted to NHP in contact with symptomatic NHP. Therefore, precautions should be taken to prevent transmission of T. gondii to humans while handling symptomatic NHP. There were no reports of clinical toxoplasmosis in Old World NHP. Among the different genera of New World NHP, susceptibility to clinical toxoplasmosis varies a great deal; however, factors affecting this susceptibility are not fully understood. Genetic characteristics of T. gondii strains from monkeys is summarized., (Published by Elsevier Ltd.)

Toxoplasmosis is a zoonosis of global distribution, and Toxoplasma gondii infections are common in humans and animals worldwide. Hares and rabbits are important small game species, and their meat is consumed by humans in many countries. Demand for rabbit meat for human consumption is increasing; therefore, toxoplasmosis in rabbits and hares is of epidemiological significance. Viable T. gondii has been isolated from rabbits. The present review summarizes worldwide information on the seroprevalence, parasitological investigations, clinical cases, isolation, and genetic diversity of T. gondii in wild rabbits, free domestic rabbits, hares, and other rabbits from 2010 to 2020. Differences in prevalence, susceptibility, genetic variants, and clinical implications of T. gondii infection in rabbits and hares are discussed. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers. Additional studies are needed to increase our knowledge of genetic variants and the population structure of T. gondii in rabbits and hares and to understand the differences in susceptibility to T. gondii in hares in different areas.

Toxoplasma gondii infections are common in humans and animals worldwide. Rodents are one of the most important intermediate hosts for T. gondii because they are preyed on by cats, who in turn excrete the environmentally resistant oocysts in their feces and thus spread the infection. Information on T. gondii infections is spread in numerous reports and is not easily accessible to readers. Here, we review prevalence, persistence of infection, clinical disease, epidemiology, and genetic diversity of T. gondii infections in wild rodents worldwide. Data are tabulated by country, by each rodent species alphabetically, and chronologically. Recent genetic diversity of T. gondii strains in rodents is critically evaluated., (© American Society of Parasitologists 2021.)

Toxoplasma gondii infections are common in humans and animals worldwide. The ingestion of food or water contaminated with oocysts excreted by infected cats or ingestion of uncooked or undercooked meat containing tissue cysts of T. gondii are the 2 major modes of transmission of T. gondii. Deer are a popular game. Recently, outbreaks of clinical toxoplasmosis were reported in humans in North America linked to ingestion of undercooked venison. Here, we review prevalence, persistence of infection, clinical disease, epidemiology, and public health risks of T. gondii infections in deer and other cervids for the past decade. Estimates of worldwide serological prevalence are summarized individually for each species of deer, elk, moose, and caribou. Genetic diversity of 112 viable isolates of T. gondii from cervids is discussed, including its public health significance. Prevalence of T. gondii in deer is very high. Any part of a deer, including liver, spleen, and muscles, should be cooked thoroughly before human consumption., (© American Society of Parasitologists 2021.)

Toxoplasma gondii infections are common in humans and animals worldwide. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, diagnosis, and genetic diversity of T. gondii in wild canids and other carnivores for the past decade. Seroprevalence estimates of T. gondii worldwide were tabulated for each host. Seroprevalence in wild foxes was very high compared with farmed Arctic foxes. Economic and public health aspects of some of the carnivore species raised for fur and meat (raccoon dogs, mink) are discussed. Diagnostic efficacies of different serological methods and PCR methods are discussed. Clinical toxoplasmosis was observed mainly in carnivores concurrently infected with immunosuppressive Canine Distemper Virus infection. Abortion and blindness were noted in mink. Genetic diversity of isolates using DNA derived from 162 (89 viable T. gondii isolates and 73 DNA extracted from tissues) of wild carnivores from several countries is discussed. However, 69 of the 162 T. gondii isolates were strains from USA and these were genetically diverse with predominance of ToxoDB genotypes #4 and #5 (haplogroup 12). Only limited information is available concerning genotyping of T. gondii isolates from other countries; none of the 93 T. gondii isolates from other countries (Brazil, China, France, Grenada) were haplogroup 12., (Published by Elsevier B.V.)

Toxoplasma gondii infections are common in humans and animals worldwide. Wild and domestic avian species are important in the epidemiology of T. gondii infections because felids prey on them and excrete millions of oocysts in the environment, disseminating the infection. Herbivorous birds are also excellent sentinels of environmental contamination with T. gondii oocysts because they feed on the ground. Toxoplasma gondii infections in birds of prey reflect infections in intermediate hosts. Humans can become infected by consuming undercooked avian tissues. Here, the authors reviewed prevalence, persistence of infection, clinical disease, epidemiology and genetic diversity of T. gondii strains isolated from turkeys, geese, ducks, ratites and avian species (excluding chickens) worldwide 2009-2020. Genetic diversity of 102 T. gondii DNA samples isolated worldwide is discussed. The role of migratory birds in dissemination of T. gondii infection is discussed.

Toxoplasma gondii infections are common in humans and animals worldwide. Toxoplasma gondii infection in pigs continues to be of public health concern. Pigs are important for the economy of many countries, particularly, USA, China, and European countries. Among the many food animals, pigs are considered the most important for T. gondii transmission in USA and China because viable parasites have rarely been isolated from beef or indoor raised chickens. Besides public health issues, T. gondii causes outbreaks of clinical toxoplasmosis in pigs in China, associated with a unique genotype of T. gondii (ToxoDB genotype #9 or Chinese 1), rarely found in other countries. The safety of ready to eat pork products with respect to T. gondii infection is a matter of recent debate. Here, we review in detail seroprevalence, prevalence of viable and nonviable T. gondii, epidemiology, risk assessment, diagnosis, and curing of pork products containing T. gondii for the past decade. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers., (Published by Elsevier B.V.)

Toxoplasma gondii infections are common in humans and animals worldwide. T. gondii causes mortality in several species of marine mammals, including threatened Southern sea otters (Enhydra lutris) and endangered Hawaiian monk seals (Monachus schauinslandi). Marine mammals are now considered sentinels for environmental exposure to protozoan agents contaminating marine waters, including T. gondii oocysts. Marine mammals also serve as food for humans and can result in foodborne T. gondii infections in humans. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, and genetic diversity of T. gondii infecting marine mammals in the past decade. The role of genetic types of T. gondii and clinical disease is discussed., (Published by Elsevier B.V.)

Toxoplasma gondii infections are common in humans and animals worldwide. The ingestion of food or water contaminated with oocysts excreted by infected cats or ingesting uncooked or undercooked meat containing tissue cysts of T. gondii are the 2 major modes of transmission of T. gondii. Viable T. gondii is more prevalent in pork and lamb than in beef. In the past decade, there have been many articles on the high seroprevalence in cattle, particularly from China. There is a report of an outbreak of acute toxoplasmosis in humans suspected to be linked to the ingestion of Artisan fresh cheese from cow's milk. There are conflicting reports concerning the rate of congenital transmission of T. gondii in cattle, especially from Brazil. In a report from Brazil, viable T. gondii was isolated from the blood of 1 of 60 pregnant cows slaughtered at an abattoir and from 1 fetus. The role of beef in the epidemiology of T. gondii infections is still not clear. Here, we review prevalence, persistence of infection, clinical disease, epidemiology, and public health risks of T. gondii infections in cattle from beef and cow's milk worldwide for the past decade., (© American Society of Parasitologists 2020.)

Toxoplasma gondii infections are common in humans and animals worldwide. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, and genetic diversity of T. gondii in horses, donkeys and other equids in the past decade. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers., Competing Interests: Conflict of interest None., (Published by Elsevier Ltd.)

Toxoplasma gondii infections are common in humans and animals worldwide. Domestic free-range chickens (Gallus domesticus) are excellent sentinels of environmental contamination with T. gondii oocysts because they feed on the ground. Chickens can be easily infected with T. gondii; however, clinical toxoplasmosis is rare in these hosts. Chickens are comparatively inexpensive and thus are good sentinel animals for T. gondii infections on the farms. Here, the authors reviewed prevalence, the persistence of infection, clinical disease, epidemiology and genetic diversity of T. gondii strains isolated from chickens worldwide for the past decade. Data on phenotypic and molecular characteristics of 794 viable T. gondii strains from chickens are discussed, including new data on T. gondii isolates from chickens in Brazil. This paper will be of interest to biologists, epidemiologists, veterinarians and parasitologists.

Toxoplasma gondii infections are common in humans and animals worldwide. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, diagnosis, control, and genetic diversity of T. gondii in sheep in the past decade. There is debate and uncertainty concerning repeat congenital infection as evidenced by finding T. gondii DNA in progeny of chronically infected sheep. However, there is no concrete evidence that T. gondii is the cause of repeated abortions in sheep. Recent data concerning pathogenesis of abortion in acutely infected sheep are reviewed. PCR-RFLP typing of T. gondii DNA derived from viable T. gondii isolates or tissues of infected sheep revealed low genetic diversity in sheep in Europe, Africa, Asia and North America but high diversity in South America. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers., (Published by Elsevier B.V.)

Toxoplasma gondii infections are common in humans and animals worldwide. Toxoplasmosis in goats continues to be of public health and economic concern. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, diagnosis, treatment, and genetic diversity of T. gondii in goats in the past decade. There is debate and uncertainty regarding excretion of T. gondii in milk and the ingestion of raw milk as sources of T. gondii infection for humans. Toxoplasmosis can cause abortion and deaths even in adult goats. PCR-RFLP typing of T. gondii DNA derived from viable T. gondii isolates has revealed genetic diversity in goats in North and South America. The significance of T. gondii isolates in domestic goats usually associated with wildlife in USA is discussed. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers., Competing Interests: Declaration of Competing Interest None., (Published by Elsevier Ltd.)

Toxoplasma gondii infections are common in humans and animals worldwide. The present review summarizes worldwide information on the prevalence of clinical and subclinical infections, epidemiology, diagnosis, and genetic diversity of T. gondii in dogs (Canis familiaris) from 2009-2020. Seroprevalence estimates of T. gondii worldwide were tabulated. Reports of high seroprevalence in canine population and high congenital transmission of T. gondii in dogs in Brazil are reviewed. Most reports from China were published in Chinese, and these reports are now summarized here. Dogs have an additional importance in some countries such as China, Vietnam, and Nigeria; whereas in many cities dog meat is sold commercially for human consumption and given to felids, and transmission of T. gondii could occur if meat is not cooked properly. Dogs can ingest T. gondii-infected cat feces and these oocysts remain viable after passage through the digestive tract of the dog; T. gondii DNA was found in feces of dogs from New York City parks in USA. Most clinical canine cases of toxoplasmosis were in immunosuppressed dogs, and ulcerative dermatitis was one of the main presentations. Genetic diversity based on PCR-RFLP markers using DNA derived from 133 viable T. gondii isolates from dogs from several countries is discussed. T. gondii strains from Asia and Americas were more genetically diverse than those from Africa. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers., (Published by Elsevier B.V.)

Abstract: Toxoplasmosis has been recognized as a major public health problem worldwide. The consumption of uncooked or undercooked meat infected with Toxoplasma gondii tissue cysts is one of the main means of transmission of this parasite. Although sheep, goats, and pigs are commonly infected with T. gondii, little information is available on the distribution of T. gondii tissue cysts in naturally infected meat. In this study, we investigated the distribution of viable T. gondii tissue cysts in shoulder muscles of naturally infected lambs and goats. Hearts and shoulders of 46 lambs and 39 goats from a local grocery store were tested for T. gondii infection. Animals were evaluated for the presence of anti-T. gondii antibodies in heart blood and clots by the modified agglutination test. Fourteen of the 85 animals (seven lambs and seven goats) were seropositive. Six to 12 samples weighing 5, 10, and 50 g were obtained from shoulder muscles of each seropositive animal and used for bioassay in mice. The distribution of viable T. gondii differed according to the size of the sample analyzed, but in general larger sample sizes resulted in higher isolation rates (P < 0.05). Results of the study revealed an uneven distribution of T. gondii in muscle samples of lambs and goats and that T. gondii can be transmitted by consumption of very small servings (5 and 10 g) of meat when it is consumed raw or is undercooked., (Published 2020 by the International Association for Food Protection. Not subject to U.S. Copyright.)

Toxoplasma gondii infections are common in humans and animals worldwide. Toxoplasmosis continues to be of public health concern. Cats (domestic and wild felids) are the most important host in the epidemiology of toxoplasmosis because they are the only species that can excrete the environmentally resistant oocysts in feces. Cats can excrete millions of oocysts and a single cat can spread infection to many hosts. The present paper summarizes information on prevalence, persistence of infection, clinical signs, and diagnosis of T. gondii infections in domestic and wild cats for the past decade. Special emphasis is paid to genetic diversity of T. gondii isolates from cats. Review of literature indicates that a unique genotype (ToxoDB genotype #9 or Chinese 1) is widely prevalent in cats in China and it has been epidemiologically linked to outbreaks of clinical toxoplasmosis in pigs and deaths in humans in China; this genotype has rarely been detected in other countries. This review will be of interest to biologists, parasitologists, veterinarians, and public health workers., (Published by Elsevier B.V.)

To assess the role of white-tailed deer (Odocoileus virginianus, WTD) in the epidemiology of toxoplasmosis, we conducted a national survey of WTD across the USA for Toxoplasma gondii infection. To do this, we combined serology with parasite isolation to evaluate the prevalence and genetic diversity of T. gondii in this game species. From October 2012 to March 2019, serum and tissues were collected from 914 WTD across the USA. Serum samples were screened for antibodies to T. gondii, and then the tissues of seropositive WTD were bioassayed in mice. Antibodies were detected in 329 (36%) of 914 WTD tested by the modified agglutination test (positive reaction at 1:25 or higher). Viable T. gondii was isolated from the heart of 36 WTD from 11 states. Three of the 36 isolates were pathogenic but not highly virulent to outbred Swiss Webster mice and all 36 isolates could be propagated further in cell culture and were genotyped. For genotyping, DNA extracted from cell culture-derived tachyzoites was characterized by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using the genetic markers SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico. Genotyping revealed seven ToxoDB PCR-RFLP genotypes, including 24 isolates for genotype #5 (haplogroup 12), four isolates for #2 (type III, haplogroup 3), three isolates for genotypes #1 (type II, haplogroup 2), two isolates for genotypes #3 (type II, haplogroup 2) and one isolate each for #39, #221 and #224. Genotype #5 was the most frequently isolated, accounting for 66.6% (24 of 36) of the isolates. Combining the 36 isolates from this study with previously reported 69 isolates from WTD, 15 genotypes have been identified. Among these, 50.4% (53/105) isolates belong to genotype #5. Our results indicate moderate genetic diversity of T. gondii in WTD. The results also indicate that undercooked venison should not be consumed by humans or fed to cats.

Feral swine are known reservoirs of various pathogens, including Toxoplasma gondii. Here, we report the first national survey of viable T. gondii in feral swine in the USA. We paired serological surveys with parasite isolation and bioassay to evaluate the prevalence and genetic diversity of these parasites. From 2012-2017, sera and tissues from 1517 feral swine across the USA were collected for the isolation of viable T. gondii. Serum samples were initially screened for antibodies to T. gondii, and then the tissues of seropositive feral swine were bioassayed in mice. Antibodies were detected in 27.7% of feral swine tested by the modified agglutination test (1:25 or higher). Antibody positive rates increased significantly with age, with 10.1% of juveniles, 16.0% of sub-adults and 38.4% of adults testing seropositive. Myocardium (50 g) from 232 seropositive feral swine was digested in pepsin and bioassayed in mice. Viable T. gondii was isolated from 78 feral swine from 21 states. Twelve of the 78 isolates were pathogenic to outbred Swiss Webster mice and 76 of the 78 isolates could be propagated further in cell culture and were genotyped. For genotyping, deoxyribonucleic acid extracted from cell culture-derived tachyzoites was characterized by polymerase chain reaction restriction fragment length polymorphism using the genetic markers SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico. Genotyping revealed 15 ToxoDB genotypes, including 43 isolates for genotype #5 (haplogroup 12), 11 isolates for #24, four isolates for #2 (haplogroup 3), two isolates for each of genotypes #3 (haplogroup 2), #4 (haplogroup 12), #216, #221, #289 and #297 and one isolate for each of genotypes #1 (haplogroup 2), #39, #66, #260, #261 and #299. Genotype #5 was the most frequently isolated, accounted for 57% (43/76) of the isolates, followed by #24, accounted for 14% (11/76). Genotypes #260, #289, #297 and #299 are new types. Genotype #289 was highly virulent to mice and originated from feral swine collected in Louisiana on the same day at the same location. Genotype #216 was previously demonstrated to be highly virulent to mice. Our results indicate moderate genetic diversity of T. gondii in feral swine in the USA, with the genotype #5 (haplogroup 12) dominant in the continental USA, whereas genotype #24 (10/14) was dominant in Hawaii, suggesting different population structures of the parasites among the two distinct geographical locations.

In a national survey of fresh, unfrozen, American pasture-raised lamb and pork, the prevalence of viable Toxoplasma gondii was determined in 1500 samples selected by random multistage sampling (750 pork, 750 lamb) obtained from 250 retail meat stores from 10 major geographic areas in the USA. Each sample consisted of a minimum of 500g of meat purchased from the retail meat case. To detect viable T. gondii , 50g meat samples of each of 1500 samples were bioassayed in mice. Viable T. gondii was isolated from 2 of 750 lamb samples (unweighted: 0.19%, 0.00-0.46%; weighted: 0.04%, 0.00-0.11%) and 1 of 750 pork samples (unweighted: 0.12%, 0.00-0.37%; weighted: 0.18%, 0.00-0.53%) samples. Overall, the prevalence of viable T. gondii in these retail meats was very low. Nevertheless, consumers, especially pregnant women, should be aware that they can acquire T. gondii infection from ingestion of undercooked meat. Cooking meat to an internal temperature of 66°C kills T. gondii ., Competing Interests: Conflict of interest None.

Ocular toxoplasmosis is one of the most common complications caused by the infection with the parasite Toxoplasma gondii . The risk of developing eye lesions and impaired vision is considered higher in Brazil than other countries. The clinical diagnosis is difficult and the use of sensitive and specific laboratorial methods can aid to the correct diagnosis of this infection. We compared serological methods ELISA and ELFA, and molecular cPCR, Nested PCR and qPCR for the diagnosis of T. gondii infection in groups of patients clinically evaluated with ocular diseases non-toxoplasma related (G1 = 185) and with lesions caused by toxoplasmosis (G2 = 164) in an Ophthalmology clinic in Brazil. Results were compared by the Kappa index, and sensitivity (S), specificity (E), positive predictive value (PPV), and negative (NPV) were calculated. Serologic methods were in agreement with ELISA more sensitive and ELFA more specific to characterize the acute and chronic infections while molecular methods were discrepant where qPCR presented higher sensitivity, however, lower specificity when compared to cPCR and Nested PCR., (Copyright © 2020 Murata, Previato, Frederico, Barbosa, Nakashima, Faria, Silveira Carvalho, Meira Strejevitch, Pereira-Chioccola, Castiglioni, de Mattos, Siqueira and Brandão de Mattos.)

Background: Toxoplasma gondii infects millions of individuals worldwide. This protozoan is food and water-borne transmitted but blood transfusion and organ transplantation constitute alternative forms for transmission. However, the influence of IgG anti- T. gondii antibodies in molecular analysis carried out in peripheral blood still remain unclear. This study aimed to investigate the serum IgG anti- T. gondii antibody concentrations correlate Nested PCR results in blood donors. Methods: 750 blood donors were enrolled. IgM and IgG anti- T. gondii antibodies were assessed by ELISA (DiaSorin, Italy). Nested PCR was performed with primers JW62/JW63 (288 bp) and B22/B23 (115 bp) of the T. gondii B1 gene. The mean values of IgG concentration were compared for PCR positive and PCR Negative blood donors using the t -test or Mann-Whitney according to the normal distribution ( p -value ≤ 0.05). Results: 361 (48.1%) blood donors presented positive serology as follow: IgM + /IgG - : 5 (0.6%); IgM + /IgG + : 21 (2.8%); IgM - /IgG + : 335 (44.7%) and 389 (51.9%), negative serology. From 353 blood donors with positive serology tested, the Nested PCR was positive in 38 (10.8%) and negative in 315 (89.2%). There were no differences statistically significant between the mean values of serum IgG anti- T. gondii antibody concentrations and the Nested PCR results. Conclusions: In conclusion, our data show that variations in the serum IgG anti- T. gondii antibody concentrations do not correlate T. gondii parasitemia detected by Nested PCR in chronically infected healthy blood donors., (Copyright © 2020 Nakashima, Pardo, Miola, Murata, Paduan, Longo, Brandão de Mattos, Pereira-Chioccola, Ricci and de Mattos.)

Toxoplasma gondii infections are prevalent in most warm-blooded animals worldwide. During the 2018 November hunting season in Pennsylvania, fresh (unfixed, not frozen) samples obtained from 99 harvested elk ( Cervus canadensis ) were tested for T. gondii infection. Antibodies to T. gondii were detected in 69 of 99 (69.7%) elk tested by the modified agglutination test (MAT, 1:25 cut-off). Tongues and hearts from 16 elk with high MAT titers ( > 1:200) were bioassayed for T. gondii by inoculation in outbred Swiss Webster (SW) and interferon-gamma gene knockout (KO) mice. Viable T. gondii was isolated from tongues of 2 elk with MAT titers of 1:200 and 1:3,200. Toxoplasma gondii from both isolates were successfully propagated in cell culture. Genetic typing on DNA extracted from culture-derived tachyzoites using the PCR restriction fragment length polymorphism with 10 genetic markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico) revealed that both isolates belonged to ToxoDB PCR-RFLP genotype #5 that is widely prevalent in wildlife in the United States. Our results suggest that elk may clear T. gondii organisms from their tissues.

Cats are important in the epidemiology of Toxoplasma gondii infection because they are the only hosts that can excrete the environmentally resistant oocysts in the environment. Although exposure is common (approximately 30% of cats in the USA), clinical toxoplasmosis is relatively rare. Here, we report overwhelming disseminated toxoplasmosis in two litter mate 8-week-old kittens, thought to have acquired toxoplasmosis postnatally. Five domestic shorthair kittens, approximately 2-3 weeks of age, and the queen were found in upstate New York by a rescue group in spring of 2018. The kittens and queen were placed in a foster home for approximately 4-5 weeks and then transferred to a shelter. Two kittens died unexpectedly following a short illness. Postmortem examination of the two deceased kittens revealed overwhelming toxoplasmosis and the presence of entero-epithelial stages in small intestine, suggestive of recent ingestion of infected tissues. Antibodies to T. gondii were found in the deceased kittens and the queen but not in the three asymptomatic littermate kittens. No obvious cause of immunosuppression was demonstrated. Genetic typing of T. gondii from DNA extracted from liver and lungs of both kittens revealed Toxo DB #4 genotype, commonly found in wildlife. Owners and veterinarians should be aware of dangers of feeding raw meat to cats and contact with infected cat feces. Procedures to safely handle T. gondii infected feces in hospital setting are outlined., (Copyright © 2019. Published by Elsevier B.V.)

Neosporosis is a common cause of abortion in cattle worldwide but is rare in horses. Here, the first case of histologically, ultrastructurally, immunohistochemically, and molecularly confirmed equine abortion caused by neosporosis is reported. Samples of lung, heart, liver, skeletal muscle, tongue, brain, and the placenta from a female fetus aborted at 280 days of gestation were fixed in formalin and submitted for diagnosis. Histologically, there was disseminated neosporosis with severe lesions in lungs, liver and the heart. Protozoal tachyzoites in all tissues reacted with polyclonal anti-Neospora caninum rabbit antibodies. Transmission electron microscopic observation on lung tissue revealed tachyzoites consistent with Neospora, including many rhoptries. Polymerase-chain reaction (PCR) using primers designed to amplify the rRNA gene internal transcribed spacer 1 (ITS1) of the Sarcocystidae was performed on DNA extracted from fetal tissues. Comparison of the ITS1 amplified from the foal tissue to sequences available in GenBank revealed 100% sequence identity to the ITS1 from three isolates of Neospora hughesi., (Published by Elsevier B.V.)

Toxoplasmosis in wild turkeys ( Meleagris gallopavo ) is of epidemiological interest because turkeys feed from the ground, and detection of infection in turkeys indicates contamination by oocysts in the environment. During the 2018 spring hunting season in Pennsylvania, fresh (unfixed, not frozen) samples were obtained from 20 harvested wild turkeys and tested for Toxoplasma gondii infection. Hearts from all wild turkeys and skeletal muscle from 1 were bioassayed for T. gondii by inoculation in outbred Swiss Webster (SW) and interferon-gamma gene knockout (KO) mice. Antibodies to T. gondii were detected in 1:5 dilution of neat serum from 5 of 15 wild turkeys and in fluid from the heart of 1 of 4 wild turkeys with the modified agglutination test (MAT); neat serum was not available from 4 wild turkeys. Viable T. gondii was isolated from hearts of 5 wild turkeys, 1 with MAT of 1:10, 1 with MAT of 1:5, and 3 seronegative (MAT < 1:5). Toxoplasma gondii was isolated from both heart and skeletal muscle in the 1 wild turkey that had skeletal muscle submitted. The KO mice inoculated with tissue from all 5 infected wild turkeys died or were euthanatized when ill, 7-21 days post-inoculation (PI). Tachyzoites were detected in lungs of all KO mice, and the T. gondii strains were successfully propagated in cell culture. The SW mice inoculated with tissues of wild turkeys remained asymptomatic, and tissue cysts were seen in the brains of infected mice when euthanatized in good health at 46 days PI; 1 of the 2 SW mice inoculated with the heart of 1 turkey died on day 26, and tachyzoites were detected in its lung. Genetic typing on DNA extracted from culture-derived tachyzoites using the PCR restriction fragment length polymorphism with 10 genetic markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico) revealed that 4 isolates belonged to ToxoDB PCR-RFLP genotype #5 and 1 was genotype #216.

Among the meat sources of Toxoplasma gondii, pork is considered important in the epidemiology of toxoplasmosis in the USA. How soon after infection T. gondii forms tissue cysts in pork is unknown. In the present study, eight serologically negative ˜ 3 months old pigs were fed mouse tissues infected with VEG (Type III) strain of T. gondii and euthanized 7 (4 pigs) and 14 days (4 pigs) post-inoculation (p.i.). Meat from the right shoulder of each pig was bioassayed in mice for T. gondii tissue cysts by peptic digestion. From each pig, the shoulder muscle was cut at random spots into 5 g, 10 g and 50 g portions. Extreme care was taken to use different scalpels and forceps to minimize cross contamination among 17 samples (6 replicates of each 5 g and 10 g portions and 5 replicates of 50 g). From the four pigs euthanized at 7 days p.i., a composite of ˜ 200 g of leftover meat from each shoulder was bioassayed in cats and their feces were tested for oocyst excretion. All eight pigs developed T. gondii antibodies (modified agglutination test, MAT, 1: 80 or higher) and viable T. gondii was isolated from shoulder meat of each pig. All four cats fed pork from excreted T. gondii oocysts. The density of T. gondii, based on mouse infectivity, varied within 5-50 g samples each pig, and between pigs within the same group, day 7 versus day 14 p.i. There were no significant differences in mouse bioassay results obtained with day 7 versus day 14 infected pigs. Overall, the rate of isolation of T. gondii increased with sample size of meat bioassayed. Results demonstrate that tissue cysts are formed early in infection and they are unevenly distributed., (Published by Elsevier B.V.)

Here, we report confirmation of sarcocysts of Sarcocystis jamaicensis in an experimental intermediate host, IFN-γ gene knockout (KO) mice orally inoculated sporocysts from its natural definitive host, a red-tailed hawk ( Buteo jamaicensis) (RTH). A RTH submitted to the Carolina Raptor Center, Huntersville, North Carolina, was euthanized because it could not be rehabilitated and released. Fully sporulated sporocysts from intestinal scrapings of the RTH were orally fed to 2 laboratory-reared outbred Swiss Webster mice (SW; Mus musculus) and to 2 KO mice. The sporocysts were infective for KO mice but not to SW mice. Both SW mice remained asymptomatic, and neither schizonts nor sarcocysts were found in their tissues when euthanized on day 54 post-inoculation (PI). The KO mice developed neurological signs and were necropsied 38-54 days PI. Schizonts/merozoites were found in both KO mice euthanized and they were confined to the brain. The predominant lesion was meningoencephalitis. Microscopic sarcocysts were found in muscles of both KO mice. When viewed with light microscopy, the sarcocyst wall appeared thin (<1 μm thick) and smooth. Ultrastructural details of sarcocysts are described.

Sarcocystis sarcocysts are common in many species of domestic and wild animals. Here, we report sarcocystosis in muscles from 91 free range elk (Cervus elaphus) from Pennsylvania, USA, tested by histopathology, transmission electron microscopy (TEM), and DNA sequencing. Sarcocysts were detected in hematoxylin and eosin (HE)-stained sections from 83 of 91 (91.2%) elk, including 83/91 (91.2%) tongues and 15/17 (88.2%) hearts. With respect to age, sarcocysts were found in 0/5 calves, 8/9 (88.8%) yearlings, and 75/77 (97.4%) adults. Sarcocysts were identified in 62/69 (89.4%) females and 21/22 (91.2%) males. Associated lesions were mild and consisted of inflammatory foci around degenerate sarcocysts. There were two morphologically distinct sarcocysts based on wall thickness, thin (< 0.5 μm) and thick-walled (> 4.0 μm). Thin-walled sarcocysts had a TEM "type 2" and villar protrusions (vps), identical to Sarcocystis wapiti previously described from elk in western USA. This species was present both in tongue and heart samples and was detected in all infected elk. Thick-walled sarcocysts consisted of three morphologic variants, referred to herein as subkinds A, B, C. Subkind A sarcocysts were rare; only four sarcocysts were found in three elk. Histologically, they had a 5-8-μm thick wall with tufted vp. By TEM, the sarcocyst wall was "type 12" and appeared similar to Sarcocystis sybillensis, previously described from elk in USA. Subkind B, Sarcocystis sp.1 sarcocysts were also rare, found in only 1 elk. These sarcocysts had 6.7-7.3-μm-thick wall with TEM "type 15b" vp. Subkind C Sarcocystis sp.2 sarcocysts were more common (22/91). Morphologically, the sarcocyst wall was 6.1-6.8 μm thick and contained "type 10b" vp. Comparisons of ribosomal DNA loci with published sequences indicated all sarcocysts were similar to what has previously been isolated from cervid hosts across the northern hemisphere. Phylogenetic analysis placed the thin-walled S. wapiti within a strongly supported clade with S. linearis and S. taeniata, while the thick-walled cysts were very closely related to S. truncata, S. elongata, S. silva, and S. tarandi. Further sequencing is needed to produce molecular diagnostics to distinguish among these species. North American elk are hosts to multiple Sarcocystis species with diverse morphology, deriving from two separate evolutionary lineages.

Rodents are known to be reservoir hosts of Toxoplasma gondii infection for other animals, such as cats and pigs. From February to July 2017, 167 rats ( Rattus norvegicus) were trapped in Grenada, and serum, heart, skeletal muscle, and brain were examined for T. gondii infection by serological examination (modified agglutination test, 1:25) for T. gondii antibodies and for viable parasites by bioassay in mice. Samples of heart, skeletal muscle, and brain of all rats were bioassayed in Swiss Webster (SW) outbred albino mice and interferon gamma gene knockout (KO) mice. Toxoplasma gondii was isolated from heart and brain from 1 rat; this was the only seropositive rat. The T. gondii strain was avirulent for SW mice but killed KO mice. Tissue cysts were detected in the brains of SW mice, and tachyzoites were detected in the lungs of KO mice that died of acute toxoplasmosis. The strain was propagated in cell culture, and DNA derived from cell-cultured tachyzoites was genotyped using the 10 PCR restriction fragment length polymorphisms (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico). The strain was a clonal Type III (ToxoDB genotype no. 2) strain. Although the prevalence of T. gondii in humans and animals in Grenada is high, rats seem to have little importance in the transmission of T. gondii on this island.

CC chemokine receptor type 5 (CCR5) is a chemokine receptor that influences the immune response to infectious and parasitic diseases. This study aimed to determine whether the CCR5Δ32 and CCR5 59029 A/G polymorphisms are associated with the development of ocular toxoplasmosis in humans. Patients with positive serology for Toxoplasma gondii were analyzed and grouped as 'with ocular toxoplasmosis' (G1: n=160) or 'without ocular toxoplasmosis' (G2: n=160). A control group (G3) consisted of 160 individuals with negative serology. The characterization of the CCR5Δ32 and CCR5 59029 A/G polymorphisms was by PCR and by PCR-RFLP, respectively. The difference between the groups with respect to the mean age (G1: mean age: 47.3, SD±19.3, median: 46 [range: 18-95]; G2: mean age: 61.3, SD±13.7, median: 61 [range: 21-87]; G3: mean age: 38.8, SD±17.9, median: 34 [range: 18-80]) was statistically significant (G1 vs.G2: p-value <0.0001; t=7.21; DF=318; G1 vs.G3: p-value <0.0001; t=4.32; DF=318; G2 vs. G3: p-value <0.0001; t=9.62; DF=318). The Nagelkerke r 2 value was 0.040. There were statistically significant differences for the CCR5/CCR5 (p-value=0.008; OR=0.261), AA (p-value=0.007; OR=2.974) and AG genotypes (p-value=0.018; OR=2.447) between G1 and G2. Individuals with the CCR5/CCR5 genotype and simultaneously the CCR5-59029 AA or AG genotypes have a greater risk of developing ocular toxoplasmosis (4% greater), which may be associated with a strong and persistent inflammatory response in ocular tissue., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Bovine besnoitiosis, caused by Besnoitia besnoiti, is an economically important disease of cattle in many countries but its transmission remains a mystery. Wild felids are suspected to be its definitive hosts. The domestic cat (Felis catus) is known experimental definitive host for Besnoitia species of rodents. Here, we report for Besnoitia darlingi the first identification of a natural definitive host, the bobcat (Lynx rufus). Oocysts resembling Toxoplasma gondii (unsporulated; 10.9±0.8×12.1±0.2μm; n=5) were detected microscopically in the feces of two of 25 free ranging wild bobcats from Mississippi, USA. After detailed investigation, we identified these oocysts as B. darlingi and not T. gondii. The IFN-γ gene knockout (KO) mice fed oocysts from bobcats died of acute besnoitiosis and tachyzoites were found in their tissues. Oocysts were also mildly pathogenic to outbred Swiss Webster mice (SW) (Mus musculus). The SW mice fed oocysts became ill but generally survived and developed characteristic thick-walled Besnoitia tissue cysts in their tongue and heart muscles and brains. Two laboratory-raised domestic cats (Felis catus) excreted B. darlingi oocysts after ingesting murine tissues infected with bobcat-derived oocysts. The parasite was successfully cultivated in African green monkey kidney fibroblast cells (CV-1 cell line) seeded with infected murine tissue homogenate. The multilocus PCR-DNA sequencing (18S rDNA, 28S rDNA, and ITS-1) from culture-derived tachyzoites confirmed the parasite as B. darlingi. Our results suggest that bobcats may be an important link in the sylvatic cycle of Besnoitia species and bioassay or molecular tests are needed to differentiate Toxoplasma gondii-like oocysts in feces of felids, both domestic and wild cats., (Published by Elsevier B.V.)

Toxoplasmosis during pregnancy can have severe consequences. The use of sensitive and specific serological and molecular methods is extremely important for the correct diagnosis of the disease. We compared the ELISA and ELFA serological methods, conventional PCR (cPCR), Nested PCR and quantitative PCR (qPCR) in the diagnosis of Toxoplasma gondii infection in pregnant women without clinical suspicion of toxoplasmosis (G1=94) and with clinical suspicion of toxoplasmosis (G2=53). The results were compared using the Kappa index, and the sensitivity, specificity, positive predictive value and negative predictive value were calculated. The results of the serological methods showed concordance between the ELISA and ELFA methods even though ELFA identified more positive cases than ELISA. Molecular methods were discrepant with cPCR using B22/23 primers having greater sensitivity and lower specificity compared to the other molecular methods., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Toxoplasmosis is a worldwide zoonosis. The ingestion of uncooked/undercooked meat and consumption of water contaminated with Toxoplasma gondii oocysts excreted by felids are the main modes of transmission of this parasite. T. gondii has been reported in multiple cervid species; however, little is known of the parasite in North American elk (Cervus canadensis). In the present study, antibodies to T. gondii were detected in serum of wild elk from Pennsylvania collected during 2013-2016 by the modified agglutination test (MAT, cut-off 1:25); 221 of 317 (69.7%) had MAT titers of 1:25 in 19, 1:50 in 28, 1:100 in 34, and 1:200 or higher in 140. Thus most (44.1%) elk had relatively high titers. Seroprevalence was slightly higher in males (76.9%) than females (67.5%, not statistically significant, Chi-square tests, P<0.0001) and was higher in adults (76.5%) than yearlings (46.4%, Odds ratio 3.82; 95% CL 1.72-8.47; P=0.001) or calves (21.7%, Odds ratio 12.58; 95% CL 4.51-35.10; P<0.0001). Annual seroprevalence was relatively stable throughout the period tested and ranged from 66.6% to 72.2%. Of the 101 elk harvested in 2016, hearts were bioassayed from 20 elk and tongues were bioassayed from 56; all tongue samples were negative. Viable T. gondii was isolated from hearts of two female elk, one of these was a seronegative adult and the other was a calf with no serum available for testing. Both T. gondii isolates were cultivated in cell culture and DNA derived from tachyzoites was characterized using the PCR-RFLP markers including SAG1, SAG2 (5'- 3'SAG2 and altSAG2), SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico. One isolate belongs to ToxoDB PCR-RFLP genotype #2 and the other is genotype #5. Both genotypes are frequently identified in animals in North America., (Copyright © 2017. Published by Elsevier B.V.)