Occurrence of Toxoplasma gondii DNA in sheep naturally infected and slaughtered in abattoirs in Pernambuco, Brazil

The aim of the present study was to assess the occurrence of antibodies to Toxoplasma gondii and to detect genomic DNA of the parasite in the reproductive organs, fetuses and fetal membranes of sheep in slaughterhouses in the state of Pernambuco, Brazil. The Indirect Immunofluorescence technique (IFA) was used for screening. The Polymerase Chain Reaction (PCR) was used to detect DNA of T. gondii in the animals that were positive in the serology. In the serology, 13/50 samples were positive and genomic DNA of T. gondii was detected in one uterus, tube, ovary, placenta and fetus (heart, brain and umbilical cord) sample from a sheep that was positive in the serology. The present study provides evidence of the occurrence of T. gondii DNA in the organs of the reproductive system, placenta and fetus of a naturally infected sheep.

Glycosphingolipid antigens from Leishmania (L.) amazonensis amastigotes: Binding of anti-glycosphingolipid monoclonal antibodies in vitro and in vivo

Specific glycosphingolipid antigens of Leishmania (L.) amazonensis amastigotes reactive with the monoclonal antibodies (MoAbs) ST-3, ST-4 and ST-5 were isolated, and their structure was partially elucidated by negative ion fast atom bombardment mass spectrometry. The glycan moieties of five antigens presented linear sequences of hexoses and N-acetylhexosamines ranging from four to six sugar residues, and the ceramide moieties were found to be composed by a sphingosine d18:1 and fatty acids 24:1 or 16:0. Affinities of the three monoclonal antibodies to amastigote glycosphingolipid antigens were also analyzed by ELISA. MoAb ST-3 reacted equally well with all glycosphingolipid antigens tested, whereas ST-4 and ST-5 presented higher affinities to glycosphingolipids with longer carbohydrate chains, with five or more sugar units (slow migrating bands on HPTLC). Macrophages isolated from footpad lesions of BALB/c mice infected with Leishmania (L.) amazonensis were incubated with MoAb ST-3 and, by indirect immunofluorescence, labeling was only detected on the parasite, whereas no fluorescence was observed on the surface of the infected macrophages, indicating that these glycosphingolipid antigens are not acquired from the host cell but synthesized by the amastigote. Intravenous administration of 125I-labeled ST-3 antibody to infected BALB/c mice showed that MoAb ST-3 accumulated significantly in the footpad lesions in comparison to blood and other tissues

How are antibodies involved in the protective mechanism of susceptible mice infected with T. cruzi?

Host resistance to Trypanosoma cruzi is dependent on both natural and acquired immune responses. During the acute phase of the infection the presence of IFN-g, TNF-a, IL-12 and GM-CSF has been closely associated with resistance, whereas TGF-ß and IL-10 have been associated with susceptibility. Several investigators have demonstrated that antibodies are responsible for the survival of susceptible animals in the initial phase of infection and for the maintenance of low levels of parasitemia in the chronic phase. However, how this occurs is not yet understood. Our results and other data in the literature support the hypothesis that the protective role of antibodies in the acute phase of infection is dependent mostly on their ability to induce removal of bloodstream trypomastigotes from the circulation in addition to other concomitant cell-mediated events.

In vivo effect of deoxynivalenol (DON) naturally contaminated feed on porcine reproductive and respiratory syndrome virus (PRRSV) infection.

Deoxynivalenol (DON), also known as vomitoxin, is the most prevalent type B trichothecene mycotoxin worldwide. Pigs show a great sensitivity to DON, and because of the high proportion of grains in their diets, they are frequently exposed to this mycotoxin. The objective of this study was to determine the impact of DON naturally contaminated feed on porcine reproductive and respiratory syndrome virus (PRRSV) infection, the most important porcine viral pathogen in swine. Experimental infections were performed with 30 animals. Piglets were randomly divided into three groups of 10 animals based on DON content of diets (0, 2.5 and 3.5 mg/kg DON). All experimental groups were further divided into subgroups of 6 pigs and were inoculated with PRRSV. The remaining pigs (control) were sham-inoculated with PBS. Pigs were daily monitored for temperature, weight and clinical signs for 21 days. Blood samples were collected and tested for PRRSV RNA and for virus specific antibodies. Results of PRRSV infection showed that ingestion of diet highly contaminated with DON greatly increases the effect of PRRSV infection on weight gain, lung lesions and mortality, without increasing significantly viral replication, for which the tendency is rather directed toward a decrease of replication. These results suggest that PRRSV infection could exacerbate anorectic effect of DON, when ingested in large doses. Results also demonstrate a DON negative effect on PRRSV-specific humoral responses. This study demonstrate that high concentrations of DON naturally contaminated feed decreased the immune response against PRRSV and influenced the course of PRRSV infection in pigs.

Deoxynivalenol (DON) naturally contaminated feed impairs the immune response induced by porcine reproductive and respiratory syndrome virus (PRRSV) live attenuated vaccine

Cereal commodities are frequently contaminated with mycotoxins produced by the secondary metabolism of fungal infection. Among these contaminants, deoxynivalenol (DON), also known as vomitoxin, is the most prevalent type B trichothecene mycotoxin worldwide. Pigs are very sensitive to the toxic effects of DON and are frequently exposed to naturally contaminated feed. Recently, DON naturally contaminated feed has been shown to decrease porcine reproductive and respiratory syndrome virus (PRRSV) specific antibody responses following experimental infection. The objective of this study was to determine the impact of DON naturally contaminated feed on the immune response generated following vaccination with PRRSV live attenuated vaccine. Eighteen pigs were randomly divided into three experimental groups of 6 animals based on DON content of the diets (0, 2.5 and 3.5 mg DON/kg). They were fed these rations one week prior to the vaccination and for all the duration of the immune response evaluation. All pigs were vaccinated intra-muscularly with one dose of Ingelvac® PRRSV modified live vaccine (MLV). Blood samples were collected at day −1, 6, 13, 20, 27 and 35 post vaccination (pv) and tested for PRRSV RNA by RT-qPCR and for virus specific antibodies by ELISA. Results showed that ingestion of DON-contaminated diets significantly decreased PRRSV viremia. All pigs fed control diet were viremic while only 1 (17%) and 3 (50%) out of 6 pigs were viremic in the groups receiving 3.5 and 2.5 mg of DON/kg, respectively. Subsequently, all pigs fed control diet developed PRRSV specific antibodies while only viremic pigs that were fed contaminated diets have developed PRRSV specific antibodies. These results suggest that feeding pigs with DON-contaminated diet could inhibit vaccination efficiency of PRRSV MLV by severely impairing viral replication.

"A Sequence in Subdomain 2 of DBL1a of Plasmodium falciparum Erythrocyte Membrane Protein 1 Induces Strain Transcending Antibodies"

Immunity to severe malaria is the first level of immunity acquired to Plasmodium falciparum. Antibodies to the variant antigen PfEMP1 (P. falciparum erythrocyte membrane protein 1) present at the surface of the parasitized red blood cell (pRBC) confer protection by blocking microvascular sequestration. Here we have generated antibodies to peptide sequences of subdomain 2 of PfEMP1-DBL1a previously identified to be associated with severe or mild malaria. A set of sera generated to the amino acid sequence KLQTLTLHQVREYWWALNRKEVWKA, containing the motif ALNRKE, stained the live pRBC. 50% of parasites tested (7/14) were positive both in flow cytometry and immunofluorescence assays with live pRBCs including both laboratory strains and in vitro adapted clinical isolates. Antibodies that reacted selectively with the sequence REYWWALNRKEVWKA in a 15-mer peptide array of DBL1a-domains were also found to react with the pRBC surface. By utilizing a peptide array to map the binding properties of the elicited anti-DBL1a antibodies, the amino acids WxxNRx were found essential for antibody binding. Complementary experiments using 135 degenerate RDSM peptide sequences obtained from 93 Ugandan patient-isolates showed that antibody binding occurred when the amino acids WxLNRKE/D were present in the peptide. The data suggests that the ALNRKE sequence motif, associated with severe malaria, induces strain-transcending antibodies that react with the pRBC surface

A sequence in subdomain 2 of DBL1 alpha of plasmodium falciparum erythrocyte membrane protein 1 induces strain transcending antibodies

Immunity to severe malaria is the first level of immunity acquired to Plasmodium falciparum. Antibodies to the variant antigen PfEMP1 (P. falciparum erythrocyte membrane protein 1) present at the surface of the parasitized red blood cell (pRBC) confer protection by blocking microvascular sequestration. Here we have generated antibodies to peptide sequences of subdomain 2 of PfEMP1-DBL1 alpha previously identified to be associated with severe or mild malaria. A set of sera generated to the amino acid sequence KLQTLTLHQVREYWWALNRKEVWKA, containing the motif ALNRKE, stained the live pRBC. 50% of parasites tested (7/14) were positive both in flow cytometry and immunofluorescence assays with live pRBCs including both laboratory strains and in vitro adapted clinical isolates. Antibodies that reacted selectively with the sequence REYWWALNRKEVWKA in a 15-mer peptide array of DBL1 alpha-domains were also found to react with the pRBC surface. By utilizing a peptide array to map the binding properties of the elicited anti-DBL1 alpha antibodies, the amino acids WxxNRx were found essential for antibody binding. Complementary experiments using 135 degenerate RDSM peptide sequences obtained from 93 Ugandan patient-isolates showed that antibody binding occurred when the amino acids WxLNRKE/D were present in the peptide. The data suggests that the ALNRKE sequence motif, associated with severe malaria, induces strain-transcending antibodies that react with the pRBC surface.

Humoral immune response of water buffalo monitored with three different antigens of Toxocara vitulorum

Humoral immune response of water buffalo naturally infected with Toxocara vitulorum was monitored using three different antigens of this parasite in serum and colostrum of buffalo cows and calves. Soluble extract (Ex) and excretory/secretory (ES) larval antigens and perienteric fluid antigen (Pe) of adult T vitulorum were used to measure the antibody levels by an indirect ELISA. Serum of 7-12 buffalo cows for the first 365 days and colostrum of the same number of buffalo cows for the first 60 days of parturition, and serum of 8-10 buffalo calves for the first 365 days afterbirth were assayed. The ELISA detected antibodies against all three T vitulorum antigens in the colostrum and serum of 100% of buffalo cows and calves examined. The highest antibody levels against Ex, ES and Pe antigens were detected in the buffalo cow sera during the perinatal period and were maintained at high levels through 300 days after parturition. on the other hand, colostrum antibody concentrations of all three antigens were highest on the first day post-parturition, but decreased sharply during the first 15 days. Concomitantly to the monitoring of immune response, the parasitic status of the calves was also evaluated. In calves, antibodies passively acquired were at the highest concentrations 24 h after birth and remained at high levels until 45 days coincidentally with the peak of T vitulorum infection. The rejection of the worms by the calves occurred simultaneously with the decline of antibody levels, which reached their lowest levels between 76 and 150 days. Thereafter, probably because of the presence of adults/larvae stimulation, the calves acquired active immunity and the antibodies started to increase slightly in the serum and plateaued between the days 211 and 365. All three antigens were detected by the serum antibodies of buffalo calves; however, the concentration of anti-Pe antibody was higher than anti-EX and anti-ES, particularly after 90 days of age. By conclusion, the buffalo cows develop immunity and keep high levels of antibodies against T vitulorum-Ex, ES and Pe antigens and these antibodies are transferred to their calves through the colostrum. This passively acquired immunity does not protect the calves against the acquisition of the infection, but these antibodies, passively or actively acquired, may have an important role during worm rejection by the calves and prevention of intestinal reinfection. (C) 2004 Elsevier B.V. All rights reserved.

Antigenic characterization of Trypanosoma evansi using sera from experimentally and naturally infected bovines, equines, dogs, and coatis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Leukocyte entry into the CNS of Leishmania chagasi naturally infected dogs

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Resposta imune-humoral de búfalos (Bubalus bubalis) contra Anaplasma marginale (Theiler, 1910)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Taenia saginata: Production and characterization of monoclonal antibodies against Taenia saginata metacestode antigens

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

First identification of Sarcocystis tenella (Railliet, 1886) Moule, 1886 (Protozoa: Apicomplexa) by PCR in naturally infected sheep from Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Serological diagnosis of visceral leishmaniasis by an enzyme immunoassay using protein A in naturally infected dogs

Um ensaio de imunoadsorção enzimática para detecção de anticorpos contra Leishmania chagasi, utilizando antígeno total de formas promastigotas lisados foi desenvolvido. Cinqüenta cães com sintomas clínicos de leishmaniose visceral foram examinados. Esta técnica utilizou anti-IgG de cão conjugado a peroxidase ou proteína A conjugado a peroxidase. Foi verificado que nos animais positivos diagnosticados por exame parasitológico direto o ensaio ELISA utilizando proteína A conjugada a peroxidase (média da densidade óptica ± desvio padrão 2,078 ± 0,631) detecta mais anticorpos do que o sistema utilizando anti-IgG de cão conjugado a peroxidase (média da densidade óptica ± desvio padrão 1,008 ± 0,437), enquanto para os animais negativos o resultado obtido nos dois sistemas de detecção são similares. Esse resultado sugere que o sistema de ELISA utilizando proteína A conjugado a peroxidase pode ser útil na detecção de animais na fase aguda da infecção e desta forma auxiliar na identificação dos animais positivos e no controle desta importante zoonose.

Immune responses in sheep naturally infected with Oestrus ovis (Diptera: Oestridae) and gastrointestinal nematodes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)