Experimental infection by Neospora caninum in gerbil reduces activity of enzymes involved in energy metabolism.

Neospora caninum is a protozoan that has tropism for the central nervous system. The aim of this study was to determine whether experimental infection of gerbils would interfere with activity of enzymes associated with energy metabolism. We randomized 20 gerbils into two groups (ten animals per group): the control group (healthy animals; uninfected) and the infected group (experimentally infected with dose 7.8 × 102 tachyzoites of N. caninum per gerbil). On day six and twelve post-infection (PI), brain and spleen tissues were collected for biochemical and histopathological analyses. No histopathological lesions were observed in the brains of infected animals; however, inflammatory infiltrates were found in the spleen. Significantly greater levels of reactive oxygen species (ROS) were observed in the brain and spleen of infected gerbils than in the control group at 12 days PI. Cytosolic creatine kinase (CK-CYT), mitochondrial creatine kinase (CK-MIT), and pyruvate kinase (PK) activities were lower in the brains of infected gerbils than in those of the control group on day 12 PI. There was significantly less CK-CYT activity in the spleens of infected gerbils on day 6 and 12 PI. Finally, there was significantly less sodium-potassium ion pump (Na+/K+ ATPase) activity in the brains and spleens of infected gerbils on day 12 PI. These data suggest that experimental infection with N. caninum interfered with energy metabolism associated with ATP homeostasis in the brain and spleen, directly or indirectly, apparently mediated by ROS overproduction, contributing to inhibition of Na+/K+ ATPase activity.

Vaccination of pigs with the S48 strain of Toxoplasma gondii--safer meat for human consumption.

As clinical toxoplasmosis is not considered a problem in pigs, the main reason to implement a control strategy against Toxoplasma gondii (T. gondii) in this species is to reduce the establishment of T. gondii tissue cysts in pork, consequently reducing the risk of the parasite entering the human food chain. Consumption of T. gondii tissue cysts from raw or undercooked meat is one of the main sources of human infection, with infected pork being considered a high risk. This study incorporates a mouse bioassay with molecular detection of T. gondii DNA to study the effectiveness of vaccination (incomplete S48 strain) in its ability to reduce tissue cyst burden in pigs, following oocyst (M4 strain) challenge. Results from the mouse bioassay show that 100% of mice which had received porcine tissues from vaccinated and challenged pigs survived compared with 51.1% of mice which received tissues from non-vaccinated and challenged pigs. The presence (or absence) of T. gondii DNA from individual mouse brains also confirmed these results. This indicates a reduction in viable T. gondii tissue cysts within tissues from pigs which have been previously vaccinated with the S48 strain. In addition, the study demonstrated that the main predilection sites for the parasite were found to be brain and highly vascular muscles (such as tongue, diaphragm, heart and masseter) of pigs, while meat cuts used as human food such as chop, loin, left tricep and left semitendinosus, had a lower burden of T. gondii tissue cysts. These promising results highlight the potential of S48 strain tachyzoites for reducing the number of T. gondii tissues cysts in pork and thus improving food safety.

Prevalence of Anti-Toxoplasma gondii and Anti-Brucella Spp. Antibodies in Pregnant Women From Mogadishu, Somalia.

Toxoplasmosis and brucellosis are zoonotic diseases of worldwide distribution. They both cause abortion and infertility in human and animals. Limited data are available about these pathogens in Somali people and their animals. Hence, this study has evaluated the prevalence of anti-Toxoplasma gondii and anti-Brucella spp. antibodies in pregnant women in Mogadishu, Somalia. Serum samples from 307 pregnant women from Mogadishu, Somalia were tested for anti-T. gondii antibodies by Latex Agglutination Test (LAT) and anti-Brucella spp. antibodies by Rose Bengal Plate Test (RBPT) and a commercial competitive-ELISA (cELISA). A total of 119/307 (38.76%) pregnant women had a prior history of abortion. A total of 159/307 (51.79%; 95% CI: 46.2-57.35%) pregnant women were seroreactive for T. gondii by LAT at different stages of pregnancy. For Brucella spp., when RBPT and cELISA were combined 4/307 (1.30%; 95% CI: 0.36-3.30%) pregnant women were seroreactive to anti-Brucella spp. antibodies, being 2/307 (0.65%; 95% CI: 0.18-2.34%) by RBPT and 3/307 (0.98%; 95% CI: 0.33-2.83%) by cELISA. Two women were seroreactive for both agents. A high seropositivity to T. gondii and low seropositivity to Brucella spp. have been found in pregnant women from Mogadishu, Somalia. Considering the high number of abortions in the country associated to the fact that awareness on other zoonotic abortifacient pathogens in Somalis is very low, further studies should be conducted to evaluate the potential causes of abortions.

A one health approach to vaccines against Toxoplasma gondii.

Toxoplasmosis is a serious disease with global impact, now recognised as one of the most important food borne diseases worldwide and a major cause of production loss in livestock. A one health approach to develop a vaccination programme to tackle toxoplasmosis is an attractive and realistic prospect. Knowledge of disease epidemiology, parasite transmission routes and main risk groups has helped to target key host species and outcomes for a vaccine programme and these would be to prevent/reduce congenital disease in women and sheep; prevent/reduce T. gondii tissue cysts in food animal species and to prevent/reduce T. gondii oocyst shedding in cats. Most animals, including humans, develop good protective immunity following infection, involving cell mediated immune responses, which may explain why live vaccines are generally more effective to protect against T. gondii. Recent advances in our knowledge of parasite genetics and gene manipulation, strain variation, key antigenic epitopes, delivery systems and induction of immune responses are all contributing to the prospects of developing new vaccines which may be more widely applicable. A key area in progressing vaccine development is to devise standard vaccine efficacy models in relevant animal hosts and this is where a one health approach bringing together researchers across different disciplines can be of major benefit. The tools and technologies are in place to make a real impact in tackling toxoplasmosis using vaccination and it just requires a collective will to make it happen.

Cell mediated and innate immune responses in pigs following vaccination and challenge with Toxoplasma parasites.

Toxoplasma gondii has a worldwide distribution and can infect almost all warm blooded animals including pigs and humans. This study aims to examine the immune responses induced in pigs following vaccination (live S48 tachyzoites) and/or challenge with T. gondii oocysts, through the examination of changes in levels of transcription in CD4, CD8α, IFN-γ, IL-12p35, CXCR3, MyD88. The experiment involved four groups of animals; pigs in group 1 (Challenged) (Chal) were challenged orally with (1 × 103 oocysts) on day 28 of the experiment. Pigs in group 2 (Vaccinated /Challenged) (Vac/Chal) were vaccinated (S48 isolate tachyzoites) on day 0, then challenged on day 28. The group 3 (Vaccinated) (Vac) animals were vaccinated (S48 isolate tachyzoites) on day 0 of the experiment. Finally the group 4 (control) pigs remained non-vaccinated and non-challenged. All animals were culled 6 weeks post challenge. At post mortem samples of retropharyngeal lymph node (RLN), mesenteric LN (MLN) and spleen were collected, RNA was extracted and cDNA synthesised. The results showed significant increases in IFN-γ expression in samples from groups 1 (Chal) and 2 (Vac/Chal) (RLN) and groups 1, 2 and 3 (Vac) (spleen) and in MyD88 expression (RLN) in samples from groups 1, 2 and 3 compared to the group 4 (control) animals. Significant increases were also observed in CD8α expression in group 1 (Chal) (RLN) and groups 1 and 2 (Vac/Chal) (RLN and MLN) compared against group 4 (control) and group 3 (Vac) respectively. Conversely, significant down regulation of CD4 and/or IL-12p35 transcription was found in at least one sample from groups 1 (Chal), 2 (Vac/Chal) and 3 (Vac) compared to group 4 (control) pigs. This study demonstrates that cell mediated and innate immune responses are generated in pigs following exposure to T. gondii parasites (oocysts or tachyzoites), key amongst them appear to be IFN-γ, MyD88 and CD8α.

Canine morbillivirus (canine distemper virus) with concomitant canine adenovirus, canine parvovirus-2, and Neospora caninum in puppies: a retrospective immunohistochemical study.

A retrospective immunohistochemical study was designed to investigate the frequency of concomitant traditional infectious disease pathogens in puppies that died suddenly and review the aspects of associated pathogenesis. Fifteen puppies were evaluated; the pathology reports and histopathologic slides of these animals were reviewed to determine the pattern of histopathologic lesions. The intralesional identification of antigens of canine (distemper) morbillivirus (CDV), canine adenovirus-1 and -2 (CAdV-1 and -2), canine parvovirus-2 (CPV-2), Toxoplasma gondii, and Neospora caninum was evaluated by IHC within the histopathologic patterns observed. All puppies contained CDV nucleic acid by molecular testing. The most frequent histopathologic patterns were intestinal crypt necrosis (n = 8), white matter cerebellar demyelination (n = 7), necrohaemorrhagic hepatitis (n = 7), interstitial pneumonia (n = 7), and gallbladder oedema (n = 5). All puppies contained intralesional antigens of CDV in multiple tissues resulting in singular (n = 3), and concomitant dual (n = 3), triple (n = 5) and quadruple (n = 4) infections by CAdV-1, and -2, CPV-2, and N. caninum; T. gondii was not identified. Concomitant infections by CDV was observed with N. caninum (100%; 1/1), CPV-2 (100%; 8/8), CAdV-1 (100%; 8/8), and CAdV-2 (100%; 8/8). Intralesional antigens of CDV and not CAdV-1 were identified in cases of gallbladder oedema. The "blue eye" phenomenon was histologically characterized by corneal oedema and degenerative lesions to the corneal epithelium, without inflammatory reactions.

Hemotropic mycoplasmas infection in water buffaloes (Bubalus bubalis) from northeastern Brazil.

Two species of hemotropic mycoplasmas (HM) are known to infect large domestic ruminants, Mycoplasma wenyonii and 'Candidatus Mycoplasma haemobos'. Although HM has been described in cattle worldwide, data in water buffaloes (Bubalus bubalis) remain scarce. Accordingly, the aim was to determine the occurrence of HM in water buffaloes from northeastern Brazil. A total of 101/290 (34.83%) buffaloes were positive for HM (16 M. wenyonii alone, 6 'Ca. M. haemobos' alone and 79 both). This was the first report of M. wenyonii infection in ruminants from Brazil. Clinical signs of hemoplasmosis in buffaloes remain unknown.

Mycoplasma ovis infection in goat farms from northeastern Brazil.

Although Mycoplasma ovis (formerly Eperythrozoon ovis) has been described in small ruminants worldwide, data on M. ovis in goats remain scarce. Accordingly, the aims of the present study were to i) determine the prevalence of hemoplasmas in goats, ii) identify the tick species parasitizing the animals, and iii) determine factors associated with infection in five dairy and three beef goat farms from the Paraíba State, northeastern Brazil. Blood samples were obtained from 402 goats. Samples were screened for hemoplasmas using a pan-hemoplasma PCR. The positive samples were confirmed by sequencing. An epidemiological questionnaire was given to each farm owner addressing age, gender, and presence of ticks. A total of 158/402 (39.3%) goats were positive for M. ovis by PCR. Sequencing of PCR positive samples has shown ≥99% identity with multiple M. ovis 16S rDNA sequences deposited in GenBank, including M. ovis isolates from humans. Dairy (OR=2.15; 95% CI: 1.40-3.32%; P=0.0004) and anemic goats (OR=2.33; 95% CI: 1.51-3.71%; P=0.0001) were more likely to be infected than beef and non-anemic animals, respectively. Amblyomma parvum (49/52, 94.23%) and Rhipicephalus microplus (3/52, 5.77%) were the tick species found parasitizing the animals, with no significant association between the presence of ticks and infection by M. ovis (P=0.1164). This is the first reportedly molecular detection of M. ovis infection in goats from South America. In conclusion, M. ovis is highly prevalent in goats from northeastern Brazil, mainly in dairy animals.

First Detection of Toxoplasma gondii DNA in the Fresh Leafs of Vegetables in South America.

Despite the high dispersion of Toxoplasma gondii oocysts in the environment, there are few studies investigating their presence in vegetables consumed by the general population. This has led us to investigate its occurrence in raw vegetables. A total of 238 samples of vegetables were collected, including crisp lettuce, regular lettuce, chicory, rocket, and parsley, both organic and nonorganic, locally in northwestern Parana, Southern Brazil. Each sample (50 g) was washed and filtered separately. A PCR was performed to detect the parasite DNA from the sediment of each sample, using B1 (B22-23) and Toxo4-5 primers. We found contamination in 3.8% of the samples, 0.8% with the primer Toxo4-5 and 2.9% with B22-B23. The results were positive in 0.6% (1/62) of the samples of smooth lettuce, 3.7% (4/106) of crisp head lettuce, 5.0% (2/40) of chicory, 14.3% (1/7) of rocket, and 20% (1/5) of parsley. These data show the contamination by T. gondii in raw vegetables directly from production sites and stores, in both organic and nonorganic samples.

CCp5A Protein from Toxoplasma gondii as a Serological Marker of Oocyst-driven Infections in Humans and Domestic Animals.

Considering that the current immunoassays are not able to distinguish the infective forms that cause Toxoplasma gondii infection, the present study was carried out to evaluate the reactivity of two recombinant proteins (CCp5A and OWP1) from oocyst/sporozoite, in order to differentiate infections occurring by ingestion of oocysts or tissue cysts. The reactivity of the recombinant proteins was assessed against panels of serum samples from animals (chickens, pigs, and mice) that were naturally or experimentally infected by different infective stages of the parasite. Also, we tested sera from humans who have been infected by oocysts during a well-characterized toxoplasmosis outbreak, as well as sera from pregnant women tested IgM(+)/IgG(+) for T. gondii, which source of infection was unknown. Only the sporozoite-specific CCp5A protein was able to differentiate the parasite stage that infected chickens, pigs and mice, with specific reactivity for oocyst-infected animals. Furthermore, the CCp5A showed preferential reactivity for recent infection by oocyst/sporozoite in pigs and mice. In humans, CCp5A showed higher reactivity with serum samples from the outbreak, compared with serum from pregnant women. Altogether, these findings demonstrate the usefulness of the CCp5A protein as a new tool to identify the parasite stage of T. gondii infection, allowing its application for diagnosis and epidemiological investigations in animals and humans. The identification of parasite infective stage can help to design effective strategies to minimize severe complications in immunocompromised people and, particularly, in pregnant women to prevent congenital infection.

Toxoplasma gondii genotypes isolated from pregnant women with follow-up of infected children in southern Brazil.

BACKGROUND: Toxoplasma gondii populations that perpetuate in South America's natural ecosystems display broad genetic diversity, but the impact of this diversity on humans is generally unknown. In this short communication, we depict the genotypic traits of four isolates related to congenital parasitism as it emerges in Southern Brazil. METHODS: Using the PCR-restriction fragment length polymorphism markers SAG1, 5'3'SAG2, alt. SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, Apico, and CS3. Three of the four strains (TgCTBral, TgCTBrv, and TgCTBrac) were identified as ToxoDB genotype #166. RESULTS: Three children displayed normal cognitive/psychomotor development, and one child displayed Sabin's tetrad (TgCTBral). CONCLUSION: Vertical transmission of the two genotypes was observed and contributes to knowledge of T. gondii strains isolated from humans in Brazil.

Estudo epidemiológico e avaliação de fatores de risco da infecção por Toxoplasma gondii e achados clinico-patológicos da infecção aguda em cães admitidos em um Hospital Escola Veterinário / Epidemiological study and evaluation of risk factors for infection with Toxoplasma gondii and clinical and pathological findings of acute infection in dogs admitted at a Veterinary School Hospital

We aimed to verify the prevalence and risk factors for Toxoplasma gondii infection in dogs addimitted at the hospital. We also assessed the occurrence rates and the clinical and pathological repercussion of the acute infection by T. gondii in these animals. Antibodies were detected in 7% (26/386) of a population of 386 dogs of both genders and different breeds and ages. Only variables, eating offal, rural origin and contact with cattle have significant values of p 0.05. Dogs from rural areas showed higher risk (OD=7.00) of infection than those of urban origin. In 6.5% (25/386) contact titles were detected (between 16 and 256); these titles do not necessarily mean acute infection, but only prior exposure. The recognition of prior infection by T. gondii is vital in those hospital patients. Depending on the cause of admission, although not being toxoplasmosis the responsible, the patient should receive prophylactic anti-parasite treatment or be monitored for further treatment in case of further acute occurrence of the disease by recrudescence of encysted bradyzoites. Only one dog (3:44%, 1/386) was admitted with high titer, what may be suggestive of acute infection (title of 4096). Although the dog with acute infection has shown neurological signs, caution is required not to extrapolate a false interpretation that toxoplasmosis is the main responsible for neurological signs, since numerous other cases included in this study had neurological signs without title of acute infection, even title of prior contact. Acute toxoplasmosis was not a significant clinic disease in this hospital; however differential diagnosis should be made in ill patients, especially those from rural areas, and definitive diagnosis must be reached for the correct therapeutic approach.

Esse trabalho teve como objetivo estudar a prevalência e respectivos fatores de risco para infecção do Toxoplasma gondii em cães provenientes de uma população hospitalar. Além disso, avaliou-se as taxas de ocorrência e as repercussões clínico-patológicas da infecção aguda pelo T. gondii nesses animais. Anticorpos foram detectados em 7% (26/386) da população estudada, composta de 386 cães de ambos os sexos e diferentes raças e idades. Somente as variáveis, ingestão de vísceras, origem rural e contato com bovinos apresentaram valores significativos com p 0.05. Adicionalmente os cães de origem rural apresentaram maiores risco (OD=7.00) de infecção do que aqueles de origem urbana. Em 6,5% (25/386) foram detectados títulos de contato (entre 16 e 256); esses títulos não significam necessariamente infecção aguda e sim apenas exposição prévia. É de fundamental importância o reconhecimento da infecção prévia por T. gondii nesses pacientes hospitalares. Dependendo da causa da admissão, mesmo não sendo a toxoplasmose a responsável, o paciente deve receber o tratamento anti-protozoário profilaticamente ou ser monitorado para posterior tratamento em caso de reagudização da enfermidade por recrudescência dos bradizoítos encistados. Apenas um animal (3.44%, 1/386) foi admitido com titulação elevada, o qual pode ser sugestivo de infecção aguda (titulo de 4096). Embora o animal com infecção aguda tenha sido apresentado com sinais neurológicos, cautela é necessária para não extrapolar uma falsa interpretação que a toxoplasmose é a grande responsável por quadros neurológico, uma vez que inúmeros outros casos incluídos nesse estudo tinham manifestações neurológicas e não tinham títulos de infecção aguda, nem mesmo título de contato prévio. A toxoplasmose aguda não foi uma afecção clínica expressiva nessa ambiência hospitalar, no entanto diagnóstico diferencial [...]

Estudo epidemiológico e avaliação de fatores de risco da infecção por Toxoplasma gondii e achados clinico-patológicos da infecção aguda em cães admitidos em um Hospital Escola Veterinário / Epidemiological study and evaluation of risk factors for infection with Toxoplasma gondii and clinical and pathological findings of acute infection in dogs admitted at a Veterinary School Hospital

Esse trabalho teve como objetivo estudar a prevalência e respectivos fatores de risco para infecção do Toxoplasma gondii em cães provenientes de uma população hospitalar. Além disso, avaliou-se as taxas de ocorrência e as repercussões clínico-patológicas da infecção aguda pelo T. gondii nesses animais. Anticorpos foram detectados em 7% (26/386) da população estudada, composta de 386 cães de ambos os sexos e diferentes raças e idades. Somente as variáveis, ingestão de vísceras, origem rural e contato com bovinos apresentaram valores significativos com p<0.05. Adicionalmente os cães de origem rural apresentaram maiores risco (OD=7.00) de infecção do que aqueles de origem urbana. Em 6,5% (25/386) foram detectados títulos de contato (entre 16 e 256); esses títulos não significam necessariamente infecção aguda e sim apenas exposição prévia. É de fundamental importância o reconhecimento da infecção prévia por T. gondii nesses pacientes hospitalares. Dependendo da causa da admissão, mesmo não sendo a toxoplasmose a responsável, o paciente deve receber o tratamento anti-protozoário profilaticamente ou ser monitorado para posterior tratamento em caso de reagudização da enfermidade por recrudescência dos bradizoítos encistados. Apenas um animal (3.44%, 1/386) foi admitido com titulação elevada, o qual pode ser sugestivo de infecção aguda (titulo de 4096). Embora o animal com infecção aguda tenha sido apresentado com sinais neurológicos, cautela é necessária para não extrapolar uma falsa interpretação que a toxoplasmose é a grande responsável por quadros neurológico, uma vez que inúmeros outros casos incluídos nesse estudo tinham manifestações neurológicas e não tinham títulos de infecção aguda, nem mesmo título de contato prévio. A toxoplasmose aguda não foi uma afecção clínica expressiva nessa ambiência hospitalar, no entanto diagnóstico diferencial deve ser feito nos pacientes enfermos, principalmente os da área rural, e diagnostico definitivo deve ser alcançado para a correta conduta terapêutica.(AU)

We aimed to verify the prevalence and risk factors for Toxoplasma gondii infection in dogs addimitted at the hospital. We also assessed the occurrence rates and the clinical and pathological repercussion of the acute infection by T. gondii in these animals. Antibodies were detected in 7% (26/386) of a population of 386 dogs of both genders and different breeds and ages. Only variables, eating offal, rural origin and contact with cattle have significant values of p<0.05. Dogs from rural areas showed higher risk (OD=7.00) of infection than those of urban origin. In 6.5% (25/386) contact titles were detected (between 16 and 256); these titles do not necessarily mean acute infection, but only prior exposure. The recognition of prior infection by T. gondii is vital in those hospital patients. Depending on the cause of admission, although not being toxoplasmosis the responsible, the patient should receive prophylactic anti-parasite treatment or be monitored for further treatment in case of further acute occurrence of the disease by recrudescence of encysted bradyzoites. Only one dog (3:44%, 1/386) was admitted with high titer, what may be suggestive of acute infection (title of 4096). Although the dog with acute infection has shown neurological signs, caution is required not to extrapolate a false interpretation that toxoplasmosis is the main responsible for neurological signs, since numerous other cases included in this study had neurological signs without title of acute infection, even title of prior contact. Acute toxoplasmosis was not a significant clinic disease in this hospital; however differential diagnosis should be made in ill patients, especially those from rural areas, and definitive diagnosis must be reached for the correct therapeutic approach.(AU)

A prospective study of Toxoplasma-positive pregnant women in southern Brazil: a health alert.

We evaluated anti-Toxoplasma gondii IgM-reactive pregnant women seen at a high-risk pregnancy outpatient clinic. From March 2005 to January 2008 in Paraná, Brazil, pregnant women seen by the Brazilian Public Health System, in any gestational period, who were anti-T. gondii IgM-positive, were followed. Clinical symptoms were noted, and tests performed including IgA, IgG avidity, ultrasonogram, and amniocentesis (PCR/inoculation in mice). Of 75 patients, 8 showed low, 3 intermediate and 31 high IgG avidity. Of those who underwent the avidity test, 31 (70.5%) were in the second trimester of pregnancy. Thirty-two (42.7%) pregnant women received specific treatment. Six received triple combination treatment; in three, tachyzoites were isolated, although only one was PCR-positive, showing changes in the cerebral sonogram, borderline IgA, and the Sabin tetrad. One fetus died, and one non-reactive IgM pregnant woman showed ocular recurrence. The municipality of residence, contact with cats during adulthood, and ingestion of unpasteurized milk were shown to be important risk factors. Congenital toxoplasmosis was observed in a pregnancy referred late for treatment. Follow-up of children born to mothers with diagnosed or suspected acute toxoplasmosis is crucial in the management of the changes that toxoplasmosis may cause.