Pedro Soto

Tritrichomonas foetus : Experimental infection in pregnant BALB/c mice

Bovine genital tritrichomonosis is a venereal disease produced by the flagellate Tritrichomonas foetus. The disease is characterized by the repetition of oestrus and infertility due to embryonic or foetal death. Numerous experimental rodent models have been developed, but none of them has been applied in pregnant females. In this work, we reproduced genital tritrichomonosis in pregnant BALB/c mice. The results were analysed considering the following pregnancy phases: early, middle and final. In the infected group, embryonic loss was significantly higher and occurred in the early and middle phases, in accordance with the time of embryo death in infected bovines. In infected animals at the early phase of pregnancy there was evidence of embryonic death without inflammatory changes in the uterus, suggesting a pathogenic mechanism that does not involve direct tissue damage. In the later days, conceptus loss was associated with endometritis and changes in the decidua.

Experimentally induced intravaginal Tritrichomonas foetus infection in a mouse model

The interest to develop research on the host-parasite relationship in bovine tritrichomonosis has accomplished the use of experimental models alternative to cattle. The BALB/c mouse became the most appropriate species susceptible to vaginal Tritrichomonas foetus infection requiring previous estrogenization. For the need of an experimental model without persistent estrogenization and with normal estrous cycles, the establishment and persistence of vaginal infection on BALB/c mouse with different concentrations of T. foetus in two experimental groups was evaluated. Group A was treated with 5mg of b-estradiol 3-benzoate to synchronize the estrous, 48 hours before the T. foetus vaginal inoculation, and Group B was inoculated in natural estrus. At 5-7 days after treatment, estrogenic effect decreased allowing all animals to cycle regularly during the experiment. From the first week post-infection, samples of vaginal mucus were taken from all animals during 34 weeks, in order to evaluate the course of infection and the stage of the estrus cycle. Group A showed 93.6% of infected animals, and Group B showed 38%. Different doses of T. foetus were assayed to establish the vaginal infection, with a persistence of 34 weeks. Although different behavior was observed in each subgroup belonging to either Group A or Group B, there were no significant differences among the infecting doses used. The b-estradiol 3-benzoate treatment had a favorable effect on the establishment of the infection (P<0.0001), but it did not influence its persistence (P=0.1097). According to the results, an experimental mouse model is presented, appropriate for further studies on mechanisms of pathogenicity, immune response, protective evaluation of immunogen and therapeutic effect of drugs.

Variation in the saccharide lectin binding pattern from different isolates of Tritrichomonas foetus.

Tritrichomonas foetus (T. foetus) is the causal agent of bovine tritrichomonosis (BT), a venereal disease that causes significant economic losses in the bovine livestock industry. The structural organization of T. foetus presents a cell membrane, an undulating membrane which extends along the parasite, three anterior flagella and a recurrent posterior flagellum. The interaction between the superficial glycoconjugates of the parasite and the host cell is one of the most relevant pathogenic mechanisms. In the present study, we analyzed the saccharide pattern through lectincytochemistry of the cell membrane, undulating membrane, cytoplasm and flagella of 28 isolates of T. foetus. Lectins that labeled most of the isolates were WGA, Con-A, RCA-I, LCA, GS-II and PHA-E showing the presence of D-mannose, D-glucose, N-acetylglucosamine and sialic acid. On the other hand, no labeling was observed in any of the structures with VVA, STA, LEA, Jacalin, GS-I, SJA, PHA-L, DSA, and weak labeling was observed with DBA, PNA, SBA and UEA I, showing therefore a low expression of N-acetylgalactosamine, L-fucose and galactose. In addition, GS II labeled in a granular pattern when lectincytochemistry was positive, whereas LCA strongly labeled the membranes and weakly the cytoplasms. The labeling variations observed among the isolates analyzed in the present work, could be related to differences in the pathogenic behavior of the isolates.

Análise quantitativa da adesão de Campylobacter fetus venerealis em culturas de células do aparelho reprodutor bovino

Campylobacter fetus e o agente etiologico da campilobacteriose genital bovina, uma doenca sexualmente transmissivel que esta associada com perdas reprodutivas em bovinos. Campylobacter coloniza a vagina e o utero e entao infecta as celulas epiteliais do endometrio. O objetivo deste trabalho foi desenvolver um modelo ex vivo para quantificar a adesao de Campylobacter as celulas-alvo naturais especificas; este e um passo fundamental para o estabelecimento da infeccao e estudos acerca da adesao e citotoxicidade sobre as celulas do hospedeiro natural nao estao disponiveis. Os ensaios foram realizados a traves da semeadura de Campylobacter fetus venerealis em culturas celulares epiteliais vaginais e uterinas.Celulas HeLa foram utilizadas como controle.A aderencia bacteriana foi confirmada por microscopia optica e a determinacao da porcentagem de bacterias aderidas foi realizada em lâminas tingidas imunoquimicamente. Os resultados sao apresentados como porcentagem de celulas com Campylobacter aderente e como o numero de bacterias por celulas. Em comparacao com as celulas HeLa controle, a analise estatistica revelou que as culturas primarias mostram uma maior porcentagem de celulas infectadas e uma menor variacao dos parâmetros avaliados. Este modelo de cultura primaria pode ser util para estudos sobre citopatogenicidade e adesao de diferentes cepas de campo de Campylobacter fetus.

Caracterización de la Adhesión de Campylobacter fetus subsp. venerealis a Espermatozoides Bovinos

Campylobacter fetus i extracellular bacteria of the genital tract of cattle. They cause infertility and abortion, but there is no documented information on the susceptibility of bovine sperm cells to this bacteria. The aim of this present work w as to study the effects provoked by Campylobacter fetus ubsp. venerealis when in interaction with bovine sperm cells. The bovine spermatozoa were obtained frozen bovine semen pooled from uninfected bulls, and were exposed to living campylobacter over different periods of time. Light microscopy and scanning electron microscopy first revealed a tropism, then a close proximity followed by tight adhe sion between these two different cells. A decrease in the spermatozoa motility was observed. Motile bacteria were observed during th next 3 h, this process began with a tight membrane–membrane adhesion. The adhesion between Campylobacter fetus to the sperm cell occurred either by the flagella or by sperm head. Results from this study demonstrated with light microscopy scanning electron microscop y allowed us to characterize some aspects of the interaction of Campylobacter fetus ubsp. venerealis and bovine sperm while preserving the cellular and bacterial structure. This ex vivo model might be useful for studies on adhesion and cytopathogenicity of different field strains of Campylobacter fetus .

Apoptosis and cell proliferation in the mouse model of embryonic death induced by Tritrichomonas foetus infection.

Bovine tritrichomonosis is a sexually transmitted disease caused by the protozoon Tritrichomonas foetus and characterised by embryonic-death and abortion. During pregnancy, the processes of cell proliferation and death play a crucial role for blastocyst implantation and the subsequent maintenance of early pregnancy, and their misbalance may lead to the abortion. In this study, we aimed to investigate whether cell proliferation and death may be altered during tritrichomonosis. For this purpose, we used pregnant BALB/c mice as an alternative experimental animal model that has successfully reproduced the infection. We analysed the immunohistochemical expression of active caspase-3 and proliferating cell nuclear (PCNA) antigens in the endometrium of infected mice. We found an increase in the number of caspase-3 positive cells in infected mice that were not pregnant at the necropsy. Besides, the number of positive proliferating cells increased in the uterine luminal epithelium of infected animals killed at 5-7 days post coitum (dpc). Pregnant infected mice killed at 8-11 dpc showed higher proliferation than control animals. We suggest that the cytopathic effect induced by T. foetus in the uteri of infected mice may induce the apoptosis of the epithelial cells and, as a result, promote a compensatory proliferative response. The information described here will be helpful to further study the pathogenesis of the bovine tritrichomonosis.

Campylobacter fetus subsp. venerealis adhesion to MDBK cells.

Fil: Chiapparrone, Maria Laura. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Tandil. Centro de Investigacion Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigacion Veterinaria de Tandil. Provincia de Buenos Aires. Gobernacion. Comision de Investigaciones Cientificas. Centro de Investigacion Veterinaria de Tandil; Argentina