RESUMO
Toxocara spp. are responsible for causing toxocariasis, a zoonotic disease of global significance. In some countries of South America, toxocariasis is considered the most prevalent human helminthic infection. The objective of this study was to evaluate LIVE/DEAD® Viability/Cytotoxicity kit as an alternative method to analyze the viability of Toxacara cati larvae. Two control groups were used to confirm the usage of this methodology: 100 untreated T. cati larvae as a negative control (G1) and 100 T. cati larvae killed by thermal shock as a positive control (G2). Subsequently, the viability of T. cati larvae was assessed by the exclusion of the trypan blue dye and by LIVE/DEAD® Viability/Cytotoxicity kit, as well as observation of motility and morphology. In order to confirm the larvicidal effect, T. cati larvae G1 and G2 were inoculated in mice to evaluate their progression in vivo. As expected, G1 showed negative staining by Trypan blue and was stained green by LIVE/DEAD® Viability/Cytotoxicity kit in all the exposure periods. Moreover, G1 presented 100% of relative motility (RM) (score of 5). G2 group was stained blue by Trypan blue and red by LIVE/DEAD® Viability/Cytotoxicity kit, and had 0% RM (score zero) in 24 h of incubation period. In mice, G2 was not viable and, therefore, was not able to infect the animals. In mice inoculated with G1, however, larvae were recovered from all the evaluated organs, except eyes. These results demonstrate that the viability of T. cati larvae was accurately obtained by the LIVE/DEAD® Viability/Cytotoxicity kit, making it an alternative method for viability evaluation.
Assuntos
Toxocara/crescimento & desenvolvimento , Análise de Variância , Animais , Membrana Celular/fisiologia , Sobrevivência Celular , Cães , Feminino , Larva/citologia , Camundongos , Camundongos Endogâmicos BALB C , Coloração e Rotulagem , Toxocara/citologia , Toxocara/fisiologia , Toxocaríase/parasitologia , Azul TripanoRESUMO
Toxocara spp. are responsible for causing toxocariasis, a zoonotic disease of global importance, which is difficult to treat as the available drugs have moderate efficacy in the clinical resolution of the disease. A promising alternative to the existing drugs is Propolis, which is known for having biological and pharmacological properties such as antiparasitic, antioxidant, and antitumor activities. In this study, we report the in vitro anthelmintic activity of essential oil from Brazilian Red Propolis (EOP) against larvae of Toxocara cati. Approximately 100 larvae per well were cultivated in microplates containing RPMI-1640 medium and incubated in the presence of EOP (18.75, 37.5, 75, 150, 300 and 600⯵g/mL) to determine the Minimum Inhibitory Concentration (MIC) and IC50 (concentration required to inhibit 50% of the population) values. Then, T. cati larvae treated with the MIC of EOP were inoculated in mice to evaluate their progression in vivo. A concentration of 600⯵g/mL of EOP showed 100% larvicidal activity after exposure for 48â¯h, while 300⯵g/mL represented the IC50 and CC50. The anthelmintic activity of EOP was confirmed by the inability of the treated T. cati larvae to infect the mice. Our findings demonstrate the potential of EOP as an anthelmintic.
Assuntos
Anti-Helmínticos/farmacologia , Óleos Voláteis/farmacologia , Própole/química , Toxocara/efeitos dos fármacos , Animais , Anti-Helmínticos/isolamento & purificação , Anti-Helmínticos/toxicidade , Células CHO , Corantes , Cricetinae , Cricetulus , Feminino , Concentração Inibidora 50 , Cinética , Larva/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Movimento/efeitos dos fármacos , Óleos Voláteis/isolamento & purificação , Óleos Voláteis/toxicidade , Toxocara/fisiologia , Azul TripanoRESUMO
Serodiagnosis of human toxocariasis is difficult in tropical areas where other helminthiasis are endemic. Many studies have shown that glycans from helminths may be the responsible for cross-reactions in the immunoassays. In this study, we have evaluated the deglycosylation of the Toxocara canis excretory-secretory (TES) antigens for the detection of IgG antibodies using a panel of 228 serum samples (58 patients with toxocariasis, 75 patients with other helminth infections and 95 healthy individuals) by ELISA and Western blot assays. Our results showed that the deglycosylation of TES antigens resulted in a single fraction of 26 kDa (dTES) and was able to detect IgG antibodies with a sensitivity and specificity of 100% in both above-mentioned assays. The rate of cross-reactions, observed in ELISA with TES (13·3%), was significantly reduced (5·3%) when the dTES antigens were used. Likewise, the cross-reactivity observed with the fractions of 32, 55 and 70 kDa of the TES antigens was totally eliminated when the dTES were used in the Western blot. All these results showed that the deglycosylation of the TES antigens really improves the specificity of the serodiagnosis of human toxocariasis in endemic areas for helminth infections.
Assuntos
Antígenos de Helmintos/sangue , Ensaio de Imunoadsorção Enzimática/métodos , Proteínas de Helminto/sangue , Toxocara/fisiologia , Toxocaríase/diagnóstico , Animais , Western Blotting , Reações Cruzadas , Feminino , Glicosilação , Humanos , Imunoglobulina G/sangue , Masculino , Sensibilidade e Especificidade , Toxocaríase/imunologiaRESUMO
Toxocara canis and Toxocara cati are nematode parasites in dogs and cats, respectively, transmitted by ingestion of embryonated eggs, transmammary and transplacental (T. canis) routes and paratenic host predation. Many parasites use mechanisms that change the behaviour of their hosts to ensure continued transmission. Several researchers have demonstrated behavioural changes in mouse models as paratenic hosts for T. canis. However, there have been no studies on behavioural changes in laboratory rats (Rattus norvegicus) experimentally infected with T. cati. This study investigated behavioural changes and muscle strength in male and female rats experimentally infected with T. cati or T. canis in acute and chronic phases of infection. Regardless of sex, rats infected with T. cati showed a greater decrease in muscle strength 42 days post infection compared to rats infected with T. canis. However, behavioural changes were only observed in female rats infected with T. canis.
Assuntos
Comportamento Animal , Força Muscular , Toxocara/fisiologia , Toxocaríase/patologia , Animais , Feminino , Masculino , RatosRESUMO
Experimental inoculations of approximately 100,000 infective Toxocara cati larval eggs were done in twelve pigs. The T. cati eggs used for inoculation were collected from cat's feces. Another group of three pigs served as an uninfected control. Groups of infected pigs were euthanized at seven, 14, 21, and 28 days post-inoculation (dpi). Tissue samples were taken for digestion and histopathology changes in early phase. The number of larvae recovered from the lungs peaked at seven and 14 dpi and were also present at 21, and 28 dpi. Larvae of T. cati were present in the lymph nodes of the small and large intestine at seven, 14, and 28 dpi and at seven, 14, 21, and 28 dpi respectively. In other studied tissues, no larvae or less than one larva per gram was detected. The pathological response observed in the liver and lungs at seven and 14 dpi, showed white spots on the liver surface and areas of consolidation were observed in the lungs. The lungs showed an inflammatory reaction with larvae in center at 28 dpi. In the liver we observed periportal and perilobular hepatitis. The lymph nodes of the intestines displayed eosinophil lymphadenitis with reactive centers containing parasitic forms in some of them. The granulomatous reaction was not observed in any tissues. The role of the other examined tissues had less significance. The relevance of this parasite as an etiological agent that leads to disease in paratenic hosts is evident.
Assuntos
Toxocara/crescimento & desenvolvimento , Toxocaríase/parasitologia , Animais , Gatos , Modelos Animais de Doenças , Feminino , Masculino , Suínos , Fatores de Tempo , Toxocara/fisiologia , Toxocaríase/patologiaRESUMO
Experimental inoculations of approximately 100,000 infective Toxocara cati larval eggs were done in twelve pigs. The T. cati eggs used for inoculation were collected from cat's feces. Another group of three pigs served as an uninfected control. Groups of infected pigs were euthanized at seven, 14, 21, and 28 days post-inoculation (dpi). Tissue samples were taken for digestion and histopathology changes in early phase. The number of larvae recovered from the lungs peaked at seven and 14 dpi and were also present at 21, and 28 dpi. Larvae of T. cati were present in the lymph nodes of the small and large intestine at seven, 14, and 28 dpi and at seven, 14, 21, and 28 dpi respectively. In other studied tissues, no larvae or less than one larva per gram was detected. The pathological response observed in the liver and lungs at seven and 14 dpi, showed white spots on the liver surface and areas of consolidation were observed in the lungs. The lungs showed an inflammatory reaction with larvae in center at 28 dpi. In the liver we observed periportal and perilobular hepatitis. The lymph nodes of the intestines displayed eosinophil lymphadenitis with reactive centers containing parasitic forms in some of them. The granulomatous reaction was not observed in any tissues. The role of the other examined tissues had less significance. The relevance of this parasite as an etiological agent that leads to disease in paratenic hosts is evident.
Se realizó la infección experimental de doce cerdos con aproximadamente 100.000 huevos infectivos de Toxocara cati. Los huevos de T. cati utilizados en la inoculación fueron recolectados de heces felinas. Otro grupo de tres cerdos no infectados se utilizó como control. Grupos de cerdos infectados se eutanaciaron a los 7,14,21 y 28 días posinoculación (pi). Se tomaron muestras de tejidos para digestión y evaluación de cambios histopatológicos en la etapa temprana de la infección. El número de larvas recuperadas de los pulmones se incrementó en los días 7 y 14 pi, recuperándose también los días 21 y 28 pi. Se encontraron larvas de T. cati en los linfonódulos del intestino delgado y grueso los días 7,14 y 28 pi y los días 7,14,21 y 28 pi respectivamente. En los restantes tejidos estudiados o no se recuperaron larvas o los valores fueron menores a una larva por gramo de tejido. La respuesta patológica observada en el hígado y los pulmones a los 7 y 14 días posinoculación, mostró en la superficie del hígado manchas blancas y en los pulmones áreas de consolidación. Los pulmones presentaron una reacción inflamatoria con presencia de larva en el centro en el día 28 pi. En el hígado se observó una hepatitis periportal y perilobular. Los linfonódulos del intestino presentaron una linfoadenitis eosinofílica con un centro reactivo conteniendo formas parasitarias en algunos de ello. En ninguno de los tejidos se observó la típica reacción granulomatosa. El rol de los restantes tejidos examinados fue de menor significancia. Queda evidenciada la importancia de éste parasito como un agente etiológico que desarrolla la enfermedad en hospederos paraténicos.
Assuntos
Animais , Gatos , Feminino , Masculino , Toxocara/crescimento & desenvolvimento , Toxocaríase/parasitologia , Modelos Animais de Doenças , Suínos , Fatores de Tempo , Toxocara/fisiologia , Toxocaríase/patologiaRESUMO
Toxocara cati is a common feline parasite transmitted by the ingestion of embryonated eggs, by the transmammary route or by predation of paratenic hosts harbouring third-stage larvae in their bodies. In the present study, the larval distribution of T. cati in tissues and organs of Rattus norvegicus experimentally infected with 300 embryonated eggs was analysed. Third-stage larvae were recovered from livers, lungs, kidneys, eyes, brains and carcasses of infected rats, following tissue digestion with HCl 0.5% for 24 h at 37 degrees C. Some differences from the known larval distribution of Toxocara canisin the same rodent species were found.
Assuntos
Reservatórios de Doenças/parasitologia , Toxocara/fisiologia , Toxocaríase/parasitologia , Animais , Gatos , Larva/crescimento & desenvolvimento , Masculino , Ratos , Ratos Wistar , Toxocara/classificaçãoRESUMO
Toxocara cati is a common feline parasite transmitted by the ingestion of embryonated eggs, by the transmammary route or by predation of paratenic hosts harbouring third-stage larvae in their bodies. In the present study, the larval distribution of T. cati in tissues and organs of Rattus norvegicus experimentally infected with 300 embryonated eggs was analysed. Third-stage larvae were recovered from livers, lungs, kidneys, eyes, brains and carcasses of infected rats, following tissue digestion with HCl 0.5 percent for 24 h at 37°C. Some differences from the known larval distribution of Toxocara canisin the same rodent species were found.
Assuntos
Animais , Gatos , Masculino , Ratos , Reservatórios de Doenças/parasitologia , Toxocara/fisiologia , Toxocaríase/parasitologia , Larva/crescimento & desenvolvimento , Ratos Wistar , Toxocara/classificaçãoRESUMO
To understand the development of the inflammatory responses in the wall of the gut, during the process of expulsion of the parasites from the host, samples of tissues were removed from the small intestines from four groups of naturally infected buffalo calves with Toxocara vitulorum during the beginning of the infection, at the peak of egg output, during the period of expulsion and post-expulsion of the worms, as well as from uninfected calves. Cells (mast cells, eosinophils, intraepithelial lymphocytes - IEL and goblet cells) present in the epithelial layer (intraepithelial) of the small intestine were counted. In the duodenum, jejunum and ileum, the population of mast cells, eosinophils and lymphocytes increased significantly during the peak of the infection. Goblet cell numbers increased also during the beginning and at the peak of the infection. The decline of the number of these cells occurred during the periods of expulsion of the worms reaching to uninfected control counts at the post-expulsion period indicating a role of these cells in the process of expulsion of T. vitulorum by the buffalo calves. The layers of the intestinal wall (villus, crypt, submucosa and muscular) were also measured. Morphological examinations showed a significant vilar atrophy, particularly in the duodenum during the beginning, peak and during the period of expulsion of the worms, but smooth muscle hypertrophy or other alteration was not observed in any period of the infection.
Assuntos
Búfalos/parasitologia , Células Epiteliais/patologia , Células Epiteliais/parasitologia , Mucosa Intestinal/patologia , Mucosa Intestinal/parasitologia , Intestino Delgado/patologia , Intestino Delgado/parasitologia , Toxocara/fisiologia , Toxocaríase/parasitologia , Animais , Feminino , Hiperplasia , Toxocaríase/patologiaRESUMO
Toxocara vitulorum is a pathogenic nematode from the small intestine of very young buffalo calves. To understand the development of the inflammatory responses in the wall of the gut, samples of tissues were removed from the duodenum, jejunum and ileum of buffalo calves naturally infected with T. vitulorum during the beginning of the infection, at the peak of egg output, as well as during the periods of rejection of the worms and post-rejection. Two additional control groups of uninfected calves (by anti-helminthic therapy of their mothers and after the birth) were also necropsied on days 30 and 50 after birth. Blood samples were fortnightly collected from birth to 174 days post-birth. Blood smears were prepared and stained with Giemsa for eosinophils. The parasitological status of buffalo calves was evaluated through weekly fecal egg counts (EPG) from 1 to 106 days after birth, which revealed that T. vitulorum egg shedding started on day 11, reached the peak of the infection on day 49 and finally expelled the parasites between days 50 and 85 after birth. In the infected buffalo calves, the mast cell population increased significantly, by two-fold in the mucosa (villus-crypt unit (VCU)) of the duodenum and four-fold in the proximal jejunum; but these increases were statistically significant only at the peak of the infection. Although mast cell numbers increased in the mucosa of the ileum as well as in both the submucosal and muscle tissues of the duodenum, proximal jejunum and ileum, the data was not significantly different from the controls. Eosinophil numbers increased in the mucosa of the duodenum (two-five times higher than the control) and proximal jejunum (three-five-fold) during the period of the infection (beginning, peak and rejection). The relative numbers of eosinophils increased in the blood stream from the second to the seventh week. In conclusion, T. vitulorum infection elicited mastocytosis and tissue eosinophilia in the duodenum and proximal jejunum, as well as eosinophilia in the blood stream, during the beginning, at the peak and during the rejection of the worm. After the rejection of the worms, the numbers of these cells returned to normal levels suggesting that these cells may have a role in the process of rejection of T. vitulorum by the host.