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Tritrichomonas foetus is a protozoan parasite that causes bovine trichomonosis (also referred to as trichomoniasis) resulting in substantial economic loss in extensive grazing systems. The parasite colonises the reproductive tracts of both male and female cattle, being asymptomatic in males but causing early reproductive failure in infected females. This systematic review aimed to examine research manuscripts describing the development of T. foetus vaccines, the strategies employed, and the immune response associated with T. foetus infection. A systematic review was conducted using indexed sources on Scopus, PubMed, Web of Science, and Embase to search for published vaccine development studies in English utilising either experimental or commercial T. foetus vaccines for bovine trichomonosis. Search terms (cattle, bull, heifer, vaccine, immune response, and Tritrichomonas foetus) were entered into the indexed sources and with no limit was set for the year of publication. Among 374 studies imported for screening, 96 were duplicates, and 255 were excluded for irrelevancy as these studies did not meet the inclusion criteria (report original data, vaccine tests in cattle, and used either commercial or experimental vaccines that incorporated antigens from T. foetus). A further five studies were excluded after full-text review as either the publication described a different target pathogen or the full text was not in English. Eighteen studies met the inclusion criteria and were subjected to data extraction using the Covidence platform. Studies included in the systematic review reported three vaccine strategies, including subunit, cell fraction, and whole-cell killed vaccines. This review considers the design, rationale, and results of each of the 18 studies to provide a comprehensive overview of the current knowledge and to inform future research agendas. While whole-cell killed vaccines are most prevalent, they have shown superior efficacy compared to subunit vaccines studied to date. One study using a purified fractions of T. foetus cell membranes as vaccine demonstrated higher efficacy and higher calving rates compared to the whole-cell killed vaccine. All of these methods require cell culture growth of T. foetus which can be challenging compared to the production of recombinant proteins. At this stage, no T. foetus recombinant antigens have been reported. Advances in understanding the parasite's genome, pathogenesis, host-parasite interactions, and host immune responses to T. foetus will provide opportunities for the development of novel vaccine strategies for bovine trichomonosis., (© 2025 The Author(s). Parasite Immunology published by John Wiley & Sons Ltd.)

The utility of therapeutic vaccination of bulls against Tritrichomonas foetus has been advocated in previous studies, but anecdotal reports suggest this practice does not clear infections and may additionally confound diagnostic testing by reducing parasite burdens below detectable limits. The objective of this study was to characterize the systemic humoral immune response to therapeutic vaccination in T. foetus-infected bulls over a period of four months using an indirect ELISA and to compare the dynamics of this response to culture and PCR results to establish the existence of a relationship (or lack thereof) between immunization and infection status. A study population of 4- to 6-year-old T. foetus-infected beef bulls (n = 20) was divided equally into a treatment group and a control group. The treatment group received two doses of commercially prepared whole cell killed vaccine 2 weeks apart while the control group received injections of vaccine diluent. Blood samples were collected at each injection and at 4 subsequent dates every 4 weeks thereafter (i.e. 0, 2, 6, 10, 14, and 18 wks) to measure IgG 1 and IgG 2 antibody subisotype response via an indirect ELISA. Preputial smegma samples were collected at the four monthly intervals following vaccination for diagnosis of infection via InPouch™ culture, Modified Diamond's Medium (MDM) culture, and PCR. Humoral response for both IgG isotypes from week 2 through week 18 were significantly increased in vaccinates compared to controls. No significant decrease in infection prevalence was detected in the treatment group for any of the diagnostic methods used. The apparent lack of pathogen clearance during a stimulated immune response suggests that therapeutic vaccination may not be a useful T. foetus management practice., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Vaccines against Tritrichomonas foetus have been shown to reduce the time of infection after natural or experimental exposure. The object of this study was to assess the protection against T. foetus infection conferred by a single vaginal instillation of formaldehyde fixed T. foetus cells. Aberdeen Angus virgin heifers were randomly allocated to 3 groups of 12 individuals to receive placebo or formaldehyde fixed T. foetus cells prepared following one of two procedures (formalin or freshly prepared solution) and six weeks later they were challenged with 10 6 T. foetus trophozoites. The median time for clearance among control heifers was 93.75 days while in animals immunized with formaldehyde fixed T. foetus it was 45 days. A single vaginal dose of cells fixed with fresh formaldehyde solution gave a rate of decay of infection per unit of time of 2.54 (CI 95% = 1.07;6.01)., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Tritrichomonas foetus (T. foetus) is a flagellated protozoa that infects the distal ileum and proximal colon of domestic cats, as well as the urogenital tract of cattle. Feline trichomonosis is recognized as a prevalent cause of chronic diarrhea in cats worldwide. The suspected route of transmission is fecal-oral, with cats in densely crowded environments at highest risk for infection. Thus, the recommended strategy for minimizing spread of infection is to identify and isolate T. foetus-positive cats from the general population. Rapid identification of infected cats can be challenging due to the inability to accurately and quickly detect the organism in samples at point of care facilities. Thus, identification of targets for use in development of a novel diagnostic test, as well as a vaccine or therapy for T. foetus infection is a significant area of research. Despite a difference in organ tropism between T. foetus genotypes, evidence exists for conserved virulence factors between feline and bovine T. foetus. The bovine T. foetus surface antigen, TF1.17, is an adhesin that is conserved across isolates. Vaccination with the purified antigen results in amelioration of cytopathogenicity and more rapid clearance of infection in cattle. We previously showed that three feline isolates of T. foetus were positive for TF1.17 antigen so we further hypothesized that TF1.17 is conserved across feline T. foetus isolates and that this antigen would represent an attractive target for development of a novel diagnostic test or therapy for feline trichomonosis. In these studies, we used monoclonal antibodies previously generated against 1.15 and 1.17 epitopes of the bovine T. foetus TF1.17 antigen, to evaluate for the presence and role of TF1.17 in the cytopathogenicity of feline T. foetus. A previously validated in vitro co-culture approach was used to model feline T. foetus infection. Immunoblotting, immunofluorescence assays, and flow cytometric analysis confirmed the presence and surface localization of antigen TF1.17 across all feline T. foetus isolates tested. Antigen TF1.17 was notably absent in the presumably nonpathogenic intestinal trichomonad, Pentatrichomonas hominis, a parasite that can be confused microscopically with T. foetus. Similar to bovine trichomoniasis, TF1.17 was found to promote T. foetus adhesion to the intestinal epithelium. These results support further characterization and development of the TF1.17 antigen as a possible target for the diagnosis and prevention of feline T. foetus infection., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Tritrichomonas foetus is a sexually transmitted reproductive pathogen of cattle that causes transient infertility, early embryonic death, metritis, pyometra, and sporadic abortions. The objective of this research was to assess the impact on reproductive health of vaccinating naïve heifers with a killed T. foetus vaccine (TrichGuard) before experimental exposure followed by breeding. A total of 40 beef heifers were randomly assigned into two treatment groups. Heifers where then vaccinated with two doses of TrichGuard or sham vaccinated with 0.9% sterile saline according to their respective groups. Sixty days following vaccination or sham vaccination, heifers were intravaginally inoculated with 2 × 10 6 organisms of a cloned isolate of T. foetus of bovine origin (CDTf-4) during synchronized estrus. Three days following inoculation of T. foetus, bulls free of T. foetus were introduced for natural breeding. Three bulls were maintained with the 40 heifers (20 vaccinated; 20 sham vaccinated) for a 49-day breeding season. Cervical mucous samples were obtained from each heifer at Day 0 and at 29 additional time points throughout the study for T. foetus culture. Pregnancy assessments were performed routinely by using transrectal palpation and ultrasonography. Pregnancies were detected in 19/20 (95%) vaccinated heifers and 14/20 (70%) sham-vaccinated heifers (P = 0.046). Only 4/20 (20%) of the sham-vaccinated heifers gave birth to a live calf compared with 10/20 (50%) of the vaccinated heifers (P = 0.048). Thus, embryonic or fetal loss was detected in 9/19 (47%) vaccinated heifers and 10/14 (71%) sham-vaccinated heifers (P = 0.153). The interval of time between inoculations with T. foetus and conceptions of pregnancies that were maintained until birth did not differ significantly between groups (vaccinated = 18.7 days; sham-vaccinated = 17.3 days; P = 0.716). The infectious challenge in this study proved to be very rigorous as a positive culture was detected from all heifers. The culture-positive results on the last culture day did not differ significantly (P = 0.115) between vaccinated heifers (63.9 days) and sham-vaccinated heifers (79.2 days). All uterine culture samples collected from the 26 nonpregnant heifers on Day 207 postinoculation did not result in the detection of T. foetus. These findings indicate that the killed, whole cell vaccine used in this study (TrichGuard) was effective in improving reproductive health evidenced by significantly reducing losses associated with T. foetus infections., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Tritrichomonas foetus is a flagellated protozoan that causes a sexually transmitted disease in cattle. Trichomonosis is characterized by early abortions, subfertility and a significant decrease in productivity. Vaccine preparations containing whole T. foetus can reduce the time of residence of the pathogen in the host cervix after experimental infection. Here, T. foetus vaccines prepared with different adjuvants were tested, in parallel with a commercial vaccine, for their efficacy to clear the infection. The median time for clearance of infection was 69days in non-immunized animals, 55days in animals treated with aluminum hydroxide, 41days with oil-in-water or saponin based vaccines or with a commercial vaccine and 27days in animals treated with saponin plus aluminum hydroxide. A slight increase in the risk of T. foetus clearance from the genital tract was found with the saponin based vaccine (hazard ratio, 2.52; 95% confidence interval, 1.03-6.17) or the commercial vaccine (hazard ratio, 2.61; 95% confidence interval, 1.07-6.38). A significant increase in the risk of T. foetus clearance was found with the combination of saponin plus aluminum hydroxide based vaccine (hazard ratio, 5.12; 95% confidence interval, 2.04-12.83)., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The objective of this study was to determine the level and duration of IgG antibodies induced against killed whole Tritrichomonas foetus and T foetus-purified surface antigen (TF1.17) in serum, vaginal, and uterine secretions after systemic immunization of beef cows with a vaccine containing killed whole T foetus. Twenty nonpregnant beef cows were randomly assigned to vaccine or control groups as follows: Vaccine (n = 10): cows received 2 mL of a commercial vaccine containing killed whole T foetus subcutaneously and a 2-mL booster 2 weeks later. Control (n = 10): cows received 2 mL of sterile saline on the same schedule. Vaginal secretions and blood samples were collected on Days 0, 8, 15, 22, 29, 36, 43, 50, 60, 75, 89, 110, 146, and 182 relative to day of primary vaccination. Uterine flush fluid was collected on Days 0, 15, 29, and 43 after the day of primary vaccination. Samples were assayed for IgG antibodies to the killed whole T foetus and surface antigen TF1.17 using enzyme-linked immunosorbent assay. Serum whole T foetus-specific IgG levels were significantly increased (between Days 15 and 182) following vaccination with T foetus or with saline. No differences between vaccinates and controls in uterine responses to whole-cell antigen were detected. Serum anti-TF1.17 IgG responses to vaccination were significantly higher than Day 0 throughout the immunization period (P < 0.001) and were higher than responses in control animals on each day post immunization through Day 146 (P < 0.001). A significant rise in TF1.17-specific IgG levels was observed in vaginal and uterine fluids from Day 15 post vaccination compared to the Day 0 levels. These levels remained significantly elevated in vaginal and uterine fluids through Days 75 (P < 0.05) and 43 (P < 0.001) after primary vaccination, respectively. Antibody levels in serum, vaginal, and uterine secretions against TF1.17 remained low in the control group throughout the study. In conclusion, vaccination of beef cows with a commercial vaccine containing T foetus induced significant increase in the levels of IgG to the T foetus TF1.17 surface antigen in serum, vaginal secretions, and uterine fluid, which remained elevated through Days 43, 75, and 182 in uterine fluids, vaginal secretions, and serum, respectively. Since purified TF1.17 antigen has been shown to protect against experimental T foetus infection in heifers, the vaccine-induced TF1.17-specific IgG response is likely to be important in the prevention of trichomoniasis in beef cattle., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Tritrichomonas foetus is a protist that causes bovine trichomoniasis and presents a well-developed Golgi. There are very few studies concerning the Golgi in trichomonads. In this work, monoclonal antibodies were raised against Golgi of T. foetus and used as a tool on morphologic and biochemical studies of this organelle. Among the antibodies produced, one was named mAb anti-Golgi 20.3, which recognized specifically the Golgi complex by fluorescence and electron microscopy. By immunoblotting this antibody recognized two proteins with 60 and 66 kDa that were identified as putative beta-tubulin and adenosine triphosphatase, respectively. The mAb 20.3 also recognized the Golgi complex of the Trichomonas vaginalis, a human parasite. In addition, the nucleotide coding sequences of these proteins were identified and included in the T. foetus database, and the 3D structure of the proteins was predicted. In conclusion, this study indicated: (1) adenosine triphosphatase is present in the Golgi, (2) ATPase is conserved between T. foetus and T. vaginalis, (3) there is new information concerning the nucleic acid sequences and protein structures of adenosine triphosphatase and beta-tubulin from T. foetus and (4) the mAb anti-Golgi 20.3 is a good Golgi marker and can be used in future studies.

Trichomonas vaginalis and Tritrichomonas foetus are parasitic protists of the human and bovine urogenital tracts, respectively. Several studies have described the cytotoxic effects of trichomonads on urogenital tract epithelial cells. However, little is known about the host cell response against trichomonads. The aim of this study was to determine whether T. foetus and T. vaginalis stimulated the release of the cytokine interleukin (IL)-10 from cultured bovine epithelial cells. To characterise the inflammatory response induced by these parasites, primary cultures of bovine oviduct epithelial cells were exposed to either T. vaginalis or T. foetus. Within 12 h after parasite challenge, supernatants were collected and cytokine production was analysed. Large amounts of IL-10 were detected in the supernatants of cultures that had been stimulated with T. foetus. Interestingly, T. vaginalis induced only a small increase in the release of IL-10 upon exposure to the same bovine cells. Thus, the inflammatory response of the host cell is species-specific. Only T. foetus and not T. vaginalis induced the release of IL-10 by bovine oviduct epithelial cells.

Tritrichomonas foetus has been identified as the causative agent of feline intestinal trichomonosis, characterized by clinical signs of chronic large bowel diarrhoea. This disease has been reported in cats from the USA, Europe and Australia. However, its epidemiology is still unclear. The aim of the present study was to describe T. foetus infection in a Persian cattery in Spain. T. foetus infection was sequentially diagnosed in 20 cats by direct faecal smear examined under the microscope, specific culture (In Pouch TF medium) and PCR. A standard coprological sedimentation method was also performed in order to screen for other intestinal parasites in all the cats included. In addition, sera were tested for IgG antibodies against Leishmania infantum, Toxoplasma gondii, and for the detection of feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV). Five out of 20 cats were positive for T. foetus (25%), two of them by microscopy, culture and PCR and three by culture and PCR. No association was found between T. foetus infection and age or sex. L. infantum and T. gondii seroprevalence rates were 15% and 10%, respectively. The prevalence of FeLV p27 antigen and of FIV antibodies in the study population was zero. Cystoisospora spp. oocysts were detected in one cat. These preliminary results show that the transmission of T. foetus infection in cluster conditions may occur between asymptomatic cats and young or immunocompromised animals.

The mucosa of the bovine prepuce has unique immunological characteristics critical to defense against sexually transmitted diseases. Tritrichomonas foetus and Campylobacter fetus subspecies venerealis persistently colonize the lower genital tract of bulls but usually do not cause either major clinical signs or inflammation. These microbes may be sexually transmitted to female cattle to cause reproductive failure. Although the male genital immune responses to T. foetus and C. fetus subspecies venerealis are inefficient in clearing infection, systemic immunization with T. foetus and C. fetus subspecies venerealis antigens does prevent or eliminate these infections with induction of IgG antibodies in genital secretions and serum., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Systemic and genital immune responses in bulls were determined after infection with Tritrichomonas foetus and systemic vaccination with whole cell antigens. Vaccinated bulls resisted infection, developed IgG1 and IgG2 antibodies against T. foetus in preputial secretions and serum, and had increased MHC II(+) and CD205(+) cells (probably dendritic cells), CD3(+) and CD8(+) T cells, and B cells including IgG1 and IgA plasma cells in the prepuce. Non-vaccinated bulls challenged with T. foetus were persistently infected and had no detectable antibodies to T. foetus in either preputial secretions or serum for 6 weeks post challenge. We conclude that genital and serum IgG antibodies to T. foetus accounts for resistance of vaccinated bulls to T. foetus infection and that the lack of an antibody response in infected bulls accounts for persistent infection.

The potential pathogenicity of non-Tritrichomonas foetus trichomonads (NTfTs) recently isolated from the prepuce of virgin bulls is not known. The purpose of this study was to determine the ability of these NTfTs to cause disease in the female reproductive tract relative to T. foetus. Forty-four virgin heifers were experimentally infected intravaginally with either one of two NTfTs (Pentatrichomonas hominis or Tetratrichomonas spp.), T. foetus, or sterile media and cultured weekly from 0 time until slaughter at 8 weeks. Serum and vaginal antibody responses during infection were assessed, and the reproductive tracts were histologically examined, scored, and compared based on numbers of neutrophils, eosinophils, lymphocytes, and plasma cells as well as the qualitative appearance of the reproductive tract. The NTfTs did not persist in the reproductive tract, while T. foetus persisted for at least 6-8 weeks. Further, no vaginal IgA response to infection was found in NTfT-infected and control heifers, but a vaginal IgA response was present in the T. foetus-infected group. Heifers infected with NTfT or controls showed little mucosal inflammatory response compared to T. foetus-infected heifers. Among the trichomonads studied, persistent infection by T. foetus alone seems responsible for uterine inflammatory lesions usually associated with pregnancy loss. The NTfTs studied in this work only transiently infected the vagina and were associated with strictly mild inflammatory changes, which probably do not cause significant disease, i.e., pregnancy loss.

Tritrichomonas foetus is the cause of trichomoniasis in cattle. Severe infection is often associated with heavy neutrophil and macrophage accumulation, although it is not known how this response protects during early parasite colonization. The goal of this study was to examine the effects of an early host response upon initial T. foetus colonization within the murine reproductive tract. Mice depleted of neutrophils before T. foetus infection had a significantly higher parasite burden within the reproductive tract compared with mock-depleted control mice. Additionally, gp91(phox-/-)/ iNOS(-/-), and iNOS(-/-) mice had substantially larger parasite burdens than C57BL/6 control mice, whereas gp91l(Phox-/-) mice had similar parasite burden to C57BL/6 control mice. Interestingly, phorbol 12-myristate 13-acetate-stimulated neutrophils and macrophages isolated from all groups of mice were unable to kill T. foetus in vitro. However, macrophages isolated from gp91l(phox-/-) and C57BL/6 mice stimulated with interferon-gamma and lipopolysaccharide were able to kill T. foetus in vitro, whereas macrophages isolated from gp91(phox(-/-)/ iNOS(-/-) and iNOS(-/-) mice were unable to kill T. foetus, suggesting the ability of macrophages to produce reactive nitrogen species but not reactive oxygen species (ROS) is critical for parasite killing during early infection in vivo and in vitro. Additionally, neutrophils seem to control early dissemination of T. foetus throughout the reproductive tract, although production of ROS is not critical for this process.

Environmental stress and endocrine control can affect pathogenesis of sexually transmitted diseases such as trichomoniasis. Acute Tritrichomonas foetus infection was compared in female BALB/c mice to infections in mice treated with high doses of estradiol or housed in constant bright illumination (stressed). In untreated mice, T. foetus readily colonized the reproductive tract, causing minimal epithelial damage and inflammation. Several fold increases of IFN-gamma, TNF-alpha, MCP-1, and IL-6 cytokines were detected after estradiol-treatment of mice, resulting in greatly enhanced inflammation and tissue damage throughout the reproductive tract. Interestingly, estradiol-treatment of mice resulted in reduced T. foetus colonization compared to untreated mice. Infection in stressed mice resulted in increased tissue damage, inflammation, and inflammatory cytokine expression, although parasite colonization within the reproductive tract was similar to that in untreated mice. These results indicate that either estradiol-treatment or stress result in pathogenesis often observed during severe disease. Alternatively, infection in non-treated mice results in chronic colonization, with little inflammation or pathology.

Tritrichomonas foetus, the agent of bovine trichomoniasis, is a flagellate protozoan responsible for substantial economic losses to the dairy and calf industries worldwide. As yet, there is no approved treatment nor is there a sensitive diagnostic method. All these problems suggest that immunization is the best control strategy. In view of this, we isolated and partially purified flagella of the parasite by vortex homogenization followed by low-speed differential centrifugation. The resulting enriched flagellar preparation termed "crude flagellar prep" was purified further by sucrose and percoll gradients. Microscopic analysis showed that the flagellar membrane was intact. Analysis by sodium dodecyl-sulfate polyacrylamide gel electrophoresis revealed three prominent protein bands of 42, 49, and >250 kDa, and several minor bands. Immunoblotting of flagellar and whole-cell extracts revealed many flagellar antigens.

We showed earlier that Tritrichomonas foetus-specific bovine immunoglobulin (Ig)G1 and IgA antibodies in uterine and vaginal secretions are correlated with clearance of this sexually transmitted infection. Eosinophils have been noted in previous studies of bovine trichomoniasis but the role of mast cells and IgE responses have not been reported. The hypothesis that IgE and mast cell degranulation play a role in clearance was tested in 25 virgin heifers inseminated experimentally and infected intravaginally with T. foetus strain D1 at estrus and cultured weekly. Groups were euthanatized at 3, 6, 9, or 12 weeks, when tissues were fixed and secretions were collected for culture and antibody analysis. Immunohistochemistry using a monoclonal antibody to a soluble lipophosphoglycan (LPG)-containing surface antigen (TF1.17) demonstrated antigen uptake by uterine epithelial cells. Lymphoid nodules were detected below antigen-positive epithelium. Little IgG2 antibody was detected but IgG1, IgA, IgM, and IgE T. foetus-specific antibodies increased in uterine secretions at weeks 6 and 9 after infection. This was inversely proportional to subepithelial mast cells numbers and most animals cleared the infection by the sampling time after the lowest mast cell count. Furthermore, soluble antigen was found in uterine epithelium above inductive sites (lymphoid nodules). Cross-linking of IgE on mast cells by antigen and perhaps LPG triggering appears to have resulted in degranulation. Released cytokines may account for production of predominantly Th2 (IgG1 and IgE) and IgA antibody responses, which are related to clearance of the infection.

Vaccines against both bovine venereal campylobacteriosis and trichomonosis were tested. Heifers were assigned to three groups. Groups 1 (n = 21 heifers) and group 2 (n = 20) received a commercial or experimental vaccine, respectively, containing both Campylobacter fetus and Tritrichomonas foetus antigens. Group 3 (n = 21) received adjuvant alone. Preparations were injected SQ in groups 1 and 3 at days -60 and -30 (day 0 was considered the first day of a 90-day breeding period), and in group 2 SQ at days -30 and +11 and into the vaginal submucosa at day -9. Heifers were exposed to two pathogen-infected bulls for 90 days (from day 0 to day +90); furthermore, half of the heifers in each group were challenged at day +39 by an intravaginal instillation of C. fetus venerealis and T. foetus. Pregnancy diagnosis, vaginal culture, and determination of systemic IgG for both organisms were performed. Compared to controls, vaccinated heifers resisted or quickly cleared both pathogens, had a higher pregnancy rate and a higher systemic immune response during and after the breeding period. Overall, the experimental vaccine was superior to the commercial vaccine (groups 2 and 1, respectively). In conclusion, an experimental vaccine containing both C. fetus and T. foetus antigens, given both SQ and intravaginal immediately before breeding and early in the breeding season, yielded superior protection for heifers exposed to bulls harboring C. fetus and T. foetus.

The performance of a whole-cell vaccine and the other vaccine with cellular membranes of Tritrichomonas foetus applied to heifers naturally challenged by mating with an infected bull was determined. Forty heifers were divided into three groups: a control group (n=16) without immunizing, another group (n=12) immunized with whole cells (10(8)/dose) and a third group (n=12) immunized with cellular membranes (300 micro g of membranes/dose protein). The females were subcutaneously vaccinated at 3-week on two occasions and received a third intravaginal booster dose. After 3 weeks of the last vaccinal doses, the heifers were served by a T. foetus infected bull over 90-day period. The mean duration of infection for membrane-vaccinated heifers was 60 days +/-25, compared with 63 days +/-35.8 of infection for whole-cell-vaccinated heifers and 79 days +/-41.3 for control heifers. Calving rates were 6/12 for membrane-vaccinated heifers, 3/12 for whole-cell-vaccinated animals, and 2/16 for control animals. Fetal mortality rates were 3/12 for membrane-vaccinated animals, 4/12 for those vaccinated with whole cells and 10/16 for control animals. These reproductive parameters were significantly different (P<0.05) between heifers vaccinated with membranes and control heifers. The hemolytic test and enzyme-linked immunoabsorbent assay (ELISA) with T. foetus antigen showed that serum immunoglobulins peaked before and during the breeding period. The heifers vaccinated with membranes developed an important response during the critical period of fetal loss, second and third month of the breeding time, and another month after the same period. The ELISA method was more sensitive and more reliable than the hemolytic test for the evaluation of the systemic immune response in females infected and/or vaccinated with T. foetus.

The antibody response patterns of cattle after subcutaneous and intranasal immunizations with adhesin Tf190 of Tritrichomonas foetus were investigated. Reactions of antibody from cattle parenterally immunized with Tf190 revealed antigen specificity and Tf190 sensitization in the majority of the animals, as determined by Western blotting. The results also demonstrated strong preimmune immunoglobulin G2 (IgG2) binding to T. foetus antigens not seen in IgG1 profiles. Subcutaneous injections of Tf190 resulted in significant (P < 0.05) increases in serum IgG1 and IgG2 titers over time, as determined by parasite specific enzyme-linked immunosorbent assay. Immune sera also significantly inhibited parasite adhesion to mammalian cell lines compared to the level of inhibition obtained with preimmune sera (P < 0.05). Intranasal immunization with Tf190 failed to produce measurable parasite-specific antibody in serum; however, this immunization route did result in significant (P < 0.05) increases in parasite-specific IgA titers in cervical mucus secretions from immunized animals that were more resistant to intravaginal challenge with T. foetus than controls. These results suggest that systemic immunization with Tf190 results in serum antibody production and antiparasitic adhesin antibodies. Additionally, the results of challenge experiments with intranasally immunized animals suggests that Tf190 primes protective immune responses that lead to lower rates of infection among these animals.

The cellular immune responses of cattle immunized with antigens of Tritrichomonas foetus were investigated. Subcutaneous injections of antigen preparations primed bovine peripheral blood mononuclear cells (PBMC) by 30 days of immunization as demonstrated by antigen-specific proliferation and by cytokine production upon antigen challenge of PBMC. Antigen-specific T-cells derived from PBMC responded by production of interferon (IFN)-gamma message detected by reverse transcriptase polymerase chain reaction, secreted IFN-gamma detected by enzyme-linked immunosorbent assay, and intracellular IFN-gamma detected by flow cytometry. Phenotypic analysis of PBMC responding in vitro to parasite antigen demonstrated a shift from a mixed CD4+, CD8+, gammadelta+, to predominantly CD4+, CD8-, gammadelta- phenotype in the Tf190-primed PBMC. In conclusion, systemic immunization of cattle with parasite antigen results in priming of bovine T-cells that are antigen specific and can produce an anamnestic IFN-gamma response to subsequent stimulation with antigens of T. foetus.

Immunoaffinity-purified TF1.17 adhesin antigen was compared biochemically and antigenically to Tritrichomonas foetus (TF) lipophosphoglycan (LPG) and a soluble glycosylated antigen (SGA) released from T. foetus and implicated in pathogenesis and immunity. The monoclonal antibodies (Mabs TF1.15 and TF1.17) specific for a glycosylated TF1.17 antigen were previously shown to prevent adhesion of the T. foetus parasites to bovine vaginal epithelial cells and to mediate killing by bovine complement. SGA was isolated from T. foetus-conditioned buffer and purified by octyl-Sepharose hydrophobic column chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of SGA showed a major SGA1 component (approximately 190 kDa) and a minor SGA2 component (50-70 kDa), which migrated close to TF-LPG and TF1.17. The carbohydrate and lipid compositional analyses of affinity-purified TF1.17 and SGA2 by high-performance liquid chromatography (HPLC) and gas-liquid chromatography revealed the presence of monosaccharides and fatty acids as found in TF-LPG. All antigens contained terminal fucose as determined by alpha-fucosidase digestion followed by HPLC. ELISA and western blots were used to further characterize these glycosylated antigens and to analyze their relationships. The Mabs TF1.15 and TF1.17 reacted very strongly to TF-LPG and SGA2. as well as TF1.17 antigen, indicating that these molecules share common epitopes. These Mabs did not react with the SGA1 component either in ELISA and western blot analyses. Also, the monosaccharide composition of SGA1 was very different from the other three antigen, suggesting SGA1 was different from LPG, SGA2 and TF1.17. Although LPG reacted with Mabs to native TF1.17 antigen, LPG did not induce an immune response in cattle with the same route and adjuvant used to produce strong antibody responses to the native antigen. The latter response suggests that the tightly bound peptide present in the immunoaffinity-purified antigen is necessary for induction of a response to (an) epitope(s) in TF-LPG and TF1.17. Furthermore, vaginal fluid from T. foetus-infected heifers and serum from a cow with a T. foetus-associated pyometra recognized both TF1.17 and TF-LPG in western blots. These results suggest that T. foetus LPG and SGA2 are related to TF1.17 antigen, which was previously shown to play an important role in the pathogenesis and host response in bovine trichomoniasis.

Portions of penis and prepuce were collected from 24 bulls with current or recent Tritrichomonas foetus infection. Epididymides were collected from seven of the bulls, and seminal vesicles and prostate were collected from four. Following immunohistochemical staining with two monoclonal antibodies (34.7C4.4 and TF1.15) prepared against T. foetus surface antigens, trichomonads were identified in sections from 15 of the bulls. Organisms were most often located in penile crypts in the midshaft and caudal regions and less often in preputial crypts. Trichomonads were not observed in sections from other genitalia or in subepithelial tissue. T. foetus antigen, however, was present in the cytoplasm of some epithelial cells and the cytoplasm of some mononuclear cells in subepithelial lymphoid aggregates and follicles. Preputial smegma was collected from 16 T. foetus-infected bulls and from 16 control bulls with negative T. foetus cultures. Preputial antibody levels to TF1.17, a surface antigen of T. foetus, were determined by an enzyme-linked immunosorbent assay. Preputial secretions from infected bulls contained specific antibody of each isotype and subisotype tested. IgG1 responses were the greatest, IgM and IgA responses were approximately equal, and IgG2 responses were low. Each isotype and subisotype response in infected bulls was significantly greater than that in the controls. These results confirm previous speculation concerning anatomical sites of infection and suggest that parasite antigen can be taken up and processed locally, resulting in deposition of specific IgG1, IgG2, IgA, and IgM antibodies in the preputial cavity.

Tritrichomonas foetus is a parasite of particular veterinary importance causing bovine tritrichomonosis, a sexually transmitted disease leading to infertility and abortion. The present review summarizes the current knowledge on potential mechanisms of pathogenicity of T. foetus, the immunology of host-parasite interaction in bovine tritrichomonosis, and the experimental model systems of this parasitic disease.

Tritrichomonas foetus is an obligate parasite of the bovine urogenital tract and is recognized as 1 of the more common infectious agents causing decreased reproductive efficiency in beef cattle. Infections result in reproductive failure and produce considerable economic loss. Vaccination of heifers with vaccines containing T. foetus induces elevated serological responses to many T. foetus antigens, decreases the rate and/or length of infection with T. foetus, and decreases fetal loss caused by infection. Because T. foetus infections are usually limited to lumen and mucosal surfaces of the reproductive tract, it has been assumed that protection from infection and abortion is partially mediated by immunoglobulins in the uterus and vagina. The objective of this study was to identify and characterize specific antigens of T. foetus that show promise for use in a recombinant vaccine that will generate a protective mucosal immune response in cattle. Surface proteins were identified by using polyclonal rabbit anti-trichomonal sera eluted from paraformaldehyde-fixed cells. Analyses of these proteins, utilizing mucosal antibodies from vaccinated and convalescent cows, have identified proteins involved in generating a local immune response. Western immunoblot analysis indicates that these proteins are well conserved and are excellent candidates for incorporation into a recombinant vaccine.

Tritrichomonas foetus is an obligate parasite of the bovine urogenital tract producing infection associated with inflammatory changes, abortion, and infertility, Tritrichomonas mobilensis was isolated from squirrel monkey colon, and symptoms involve diarrheal complications. Both tritrichomonads produced hemagglutinins with the properties of sialic acid-specific lectins. Assays on the adherence of these protozoans to Chinese hamster ovary (CHO) cells and to bovine cervical and monkey colon mucus were performed to assess the function of the lectins in adhesion. Sialic acid at concentration as low as 2 mM inhibited the adhesion to CHO cells, less effectively to the mucus. Predigestion with Clostridium perfringens sialidase prevented the adhesion to both epithelial cells and the mucus. Inhibition of endogenous sialidases with 2,3-dehydro-2-deoxy-NeuAc increased the adhesion of T. mobilensis to CHO cells. Specific anti-T. foetus lectin (TFL) and anti-T. mobilensis lectin (TML) antibodies inhibited adhesion of the trichomonads to the epithelial cells and to the mucus. TFL histochemistry disclosed the presence of lectin ligands on keratinized vaginal epithelia, cervical mucosa, and mucin and on endometrial glands and their secretions. TML histochemistry showed reactivity with the luminal membranes of colonic glandular epithelium and less with the colonic mucin. Both lectins bound to the surface membrane of CHO cells. Anti-lectin antibodies showed granular cytoplasmic and strong membrane localization of the lectins in both tritrichomonads. Although the 2 tritrichomonads have different habitats, the results indicate that both these protozoa use lectins with sialic acid specificity for adhesion to mucosal surfaces.

A reliable laboratory animal model would be useful for the study of immune responses to trichomoniasis, a sexually transmitted disease of human beings and cattle. Murine models are available, but pretreatment with estrogen is used, which may influence immune responses. To evaluate whether vaginal trichomoniasis could be established in nonestrogenized mice and to define the immune responses associated with the infection, CD1 and BALB/c mice were studied with or without estrogen treatment prior to inoculation with Tritrichomonas foetus. Tritrichomonas Foetus was cultured from the vagina and uterus of both estrogen-treated and untreated control mice for up to 26 wk. The infection was sustained better in BALB/c than in CD1 mice, suggesting that the former strain was most susceptible. In CD1 mice, infection was sustained less well in estrogen-treated than in untreated control mice, but there was no difference between treatment groups of BALB/c mice. IgA and IgG antibodies in vaginal secretions, uterine secretions, and serum specific for a surface antigen of T. foetus (TF1.17) were measured by enzyme-linked immunosorbent assay. In infected CD1 mice, vaginal IgA and IgG antibodies were detected by 8 wk postinoculation (PI). In infected BALB/c mice, vaginal IgA and IgG antibodies were detected by 12 wk PI. Uterine IgG responses predominated over IgA in estrogen-treated and untreated CD1 and BALB/c mice. There were high levels of IgG, but relatively no IgA in the sera of CD1 and BALB/c mice. Overall, the highest IgA response was in the vaginal secretions of infected CD1 mice, and some animals of this strain cleared the infection. These results show that a chronic trichomonad infection was established in mice without prior treatment with estrogen. The infection was associated with antibody responses in reproductive secretions and serum. This animal model will be useful in studying immunization to protect against trichomoniasis in mice not immunocompromised by estrogen.

Tritrichomonas foetus is the causative agent of bovine tritrichomonosis, a sexually transmitted disease leading to infertility and abortion. Diagnosis is hampered by putative contamination of samples with intestinal or coprophilic trichomonadid protozoa which might be mistaken for T. foetus. Therefore, we developed a PCR test optimized for applicability in routine diagnosis. Amplification is based upon primers TFR3 and TFR4 directed to the rRNA gene units of T. foetus. In order to avoid potential carryover contamination by products of previous amplification reactions, conditions were adapted to the use of the uracil DNA glycosylase system. Furthermore, documentation and interpretation of results were facilitated by including a DNA enzyme immunoassay for the detection of amplification products. Specificity was confirmed with genomic material from different related trichomonadid protozoa. The high sensitivity of the test allowed the detection of a single T. foetus organism in diagnostic culture medium or about 50 parasites per ml of preputial washing fluid. The present methods are thus proposed as (i) confirmatory tests for microscopic diagnosis following diagnostic in vitro cultivation and (ii) a direct T. foetus screening test with diagnostic samples.

Histopathologic changes and local antibody responses were studied in immunized and control heifers after intravaginal challenge with 10(6) Tritrichomonas foetus. Animals were given 3 intramuscular inoculations of immunoaffinity-purified superficial antigen, TF 1.17, in 2 different adjuvant combinations (incomplete Freund's adjuvant or dextran sulfate plus IFA-8 animals each) or adjuvant alone at 3-wk intervals and were challenged with T. foetus 2 wk later. Histologically, a nonsuppurative endometritis with nodular lymphoid aggregates in the stratum spongiosum was present in 9 of 24 heifers. Twice as many control heifers as immunized had moderate to severe endometritis at 10 wk and the rate of clearance of the organism was significantly faster in immunized than in control heifers. Furthermore, time of clearance was statistically correlated with severity of endometritis at 10 wk postinfection, when necropsies were done (P < 0.02). Because 9-10 wk postinfection is thought to be the critical period for determining fetal loss associated with endometritis, this correlation with early clearance is important to protection against disease. In heifers with moderate to severe infiltration of mononuclear cells in the endometrium, lymphoid nodules and some secondary follicles were detected. In the subgroup of 12 animals from which uterine secretions were collected. IgA antibody responses to antigen were detected by 6 wk in infected animals with increases in mean responses at 8 and 10 wk, but not in uninfected animals. A rationale is presented for consideration of the lymphoid nodules as a possible inductive site for this local antibody response to T. foetus.

Tritrichomonas foetus is a protozoan parasite of cattle that can be cultured axenically. Three monoclonal antibodies specific for surface antigens of T. foetus were found to be rapidly internalized and degraded by these cells after binding. Degradation was not due to secreted or artificially liberated proteases but depended on targeting to internal degradative compartments. Radiolabeled catabolites of the antibodies were subsequently incorporated into the parasite's own proteins. Antibody degradation could be inhibited by certain protease inhibitors or lowered temperatures; a sharp reduction in degradation between 20 C and 15 C was similar to a well documented block in endocytic transport to degradative compartments of mammalian cells. Growth and proliferation of T.foetus in the continuous presence of the antibodies appeared unhindered, but there was a general shift toward expression of both more and less of each epitope among cells within each population. Subclones of these populations always exhibited striking variability in epitope expression levels, with patterns similar to the parent cultures. These findings may lead to a better understanding of how T. foetus resists host immune responses.

We developed a serological assay for detection of (l) an erythrocyte-adhering molecule(s) shed by the bovine venereal pathogen Tritrichomonas foetus and (II) serum antibodies to this antigen(s) in exposed cattle. Sera from exposed and unexposed cattle were tested for their ability to induce complement-mediated lysis of bovine erythrocytes that had been previously incubated overnight at room temperature in pH-adjusted supernatants of T. foetus culture media. Eight of 180 serum specimens from six groups of presumably unexposed cows or heifers showed a positive (> or = 1:2) hemolytic titer (specificity = 95.6%). Thirteen of 14 females in two experimentally infected groups showed a positive hemolytic titer following infection (sensitivity = 94%). In experimentally infected heifers, there was little correlation (r2 = 0.33) between serum hemolytic titers with respect to shed antigen and titers obtained in serum enzyme-linked immunosorbent assays in which whole T. foetus served as the antigen. Serum hemolytic titers rose 3 to 4 weeks sooner than did previously described vaginal mucus immunoglobulin G1 or immunoglobulin A titers with respect to whole-cell antigen or TF1.17 subunit antigen, respectively. Among 14 chronically infected bulls, only 6 (43%) showed a positive hemolytic titer. This study is the first, to our knowledge, to show a specific serological response in the host to an in vitro-shed antigen(s) of T. foetus and suggests a useful diagnostic test for potentially exposed herds.

Ninety-seven BALB/c mice were inoculated intravaginally with 8.0 x 10(5) Tritrichomonas foetus organisms, using either isolate ATCC 30003 or field isolate MU Y22 2 days after estrogenization with 15 micrograms 17 beta-estradiol. Reproductive tracts were examined at several time points post-inoculation to determine gross and histologic responses to trichomonad infection as compared to estrogenized, uninfected control animals. The two isolates varied greatly in ability to maintain chronic infection; no ATCC 30003-inoculated animals remained culture-positive beyond 7 weeks post-inoculation, whereas MU Y22-inoculated animals were infected for greater than 26 weeks. Lesions were seen in 40-60% of animals prior to 10 weeks post-inoculation and included moderate uterine dilation and glandular atrophy, uterine gland abscesses, pyometra, intramural perivascular lymphoid infiltrates, and ovarian bursitis. The severity of lesions was independent of the T. foetus isolate. Lesions became more severe at 10 weeks post-inoculation, and at 10 and 26 weeks post-inoculation, lesions were seen in 60% and 75% of animals, respectively. In addition to lesions described above, epithelial changes were marked at these late necropsies, including ulceration, flattening, hypertrophy, and squamous metaplasia. The lesions seen in these mice closely resemble those described in natural bovine infection, suggesting that the estrogenized BALB/c mouse is an excellent model for study of bovine trichomoniasis.

Bovine trichomoniasis is a prevalent sexually transmitted disease of cattle caused by the protozoan Tritrichomonas foetus. Currently, diagnosis is most often made by culture. In order to provide a faster immunodiagnostic approach, a specific enzyme-linked immunosorbent assay (ELISA) was investigated. A protective surface antigen (TF1.17 antigen) of T. foetus was immunoaffinity purified and used in an ELISA to detect antibodies in vaginal mucus from heifers inoculated with T. foetus. In preliminary studies, antibodies of the immunoglobulin A (IgA) isotype were detected in mucus from all experimentally infected heifers which were tested at 6 weeks postinoculation, whereas IgG1 and IgG2 were not. In addition, IgA responses detected in postinoculation samples were all greater than those detected in preinoculation samples, unlike those detected by a whole-cell antigen ELISA. For these two reasons, IgA antibodies appeared to be useful diagnostically. Further investigation of IgA antibodies used vaginal mucus collected weekly from heifers inoculated intravaginally with 10(2), 10(4), or 10(6) T. foetus organisms. Heifers with positive cultures for T. foetus had similar IgA responses to TF1.17 antigen over the 10 weeks of infection regardless of the initial inoculum dose. This indicates that if the dose is sufficient to establish infection, the magnitude and duration of the immune response are no longer dependent on dose. All of the infected animals receiving all dosages responded with high absorbance values in the IgA anti-TF1.17 antigen ELISA by 6 weeks postinoculation, and all absorbance values remained high at 10 weeks. To determine the duration of the IgA response, four other heifers inoculated with 7 x 10(6) T. foetus organisms were studied through 24 weeks postinoculation. IgA antibody responses to TF1.17 antigen were still high at 24 weeks, even though some heifers cleared the infection several weeks earlier. These results indicate that experimentally infected heifers produced detectable long-lasting IgA responses to TF1.17 antigen in vaginal mucus, which may be useful in a herd test for the diagnosis of bovine trichomoniasis.

Thirty-five heifers were allotted to 3 groups. Group 1 (control) consisted of 10 heifers that were not vaccinated and were challenge exposed by breeding to infected bulls. Group 2 (natural challenge exposure) consisted of 10 heifers that were vaccinated and challenge exposed by breeding to infected bulls. Group 3 (experimental challenge exposure) consisted of 15 heifers that were vaccinated and challenge exposed by breeding to infected bulls and by intravaginal inoculation with 10(7) Tritrichomonas foetus. Total immunoglobulin concentrations and specific trichomonal antibodies were determined in serum and vaginal secretions of heifers, using radial immunodiffusion and ELISA procedures. Control heifers remained infected for a mean of 10.6 weeks (range, 0 to 18 weeks), and heifers of the natural and experimental challenge-exposure groups remained infected for 3.2 and 5.0 weeks, respectively (range, 0 to 12 weeks). Total serum and cervicovaginal mucus concentrations of IgM, IgA, IgG1, and IgG2 did not change significantly after vaccination or challenge exposure. However, ELISA titers of total trichomonal antibodies increased up to 1:10,000 (range, 1:400 to 1:10,000) in serum after vaccination, and increased approximately tenfold above background in cervicovaginal mucus. In serum, the predominant trichomonal antibody isotype was IgG1, although trichomonal IgA and IgM antibodies also increased. The predominant trichomonal antibody detected in cervicovaginal mucus was IgA. Antibody titers in serum and cervicovaginal mucus of vaccinated heifers were not increased by infection. However, in control heifers, the total local trichomonal antibody response increased three- to fivefold after infection.(ABSTRACT TRUNCATED AT 250 WORDS)

The evolution of hydrogenosomes, energy-generating organelles of rumen ciliate protozoa and the flagellate trichomonads has been the subject of much speculation. Polypeptides of the hydrogenosome-enriched fractions from the rumen ciliates, Dasytricha ruminantium, Isostricha spp., Polyplastron multivesiculatum and Eudiplodinium maggii were separated by SDS-PAGE and compared to analogous polypeptide preparations from Tritrichomonas foetus. Immunoblotting with antisera specific to the hydrogenosomes of T. foetus identified common immunoreactive polypeptides present at estimated molecular masses of 28, 35, 38, 44, 48, 58, 100 and 120 kDa. That at 120 kDa corresponds to a single subunit of the purified pyruvate:ferredoxin oxidoreductase from the hydrogenosome of Trichomonas vaginalis.

Bovine trichomoniasis is a sexually transmitted disease caused by the flagellated protozoan Tritrichomonas foetus. A protective surface antigen was previously identified and immunoaffinity purified from T. foetus isolate D1 with cross-reactive monoclonal antibodies (MAbs) TF1.15 and TF1.17 (BonDurant, R. H., R. R. Corbeil, and L. B. Corbeil, Infect. Immun. 61:1385-1394, 1993). This antigen elicited antibody responses in the serum and cervicovaginal mucus of heifers. Thus, it may be useful as an immunodiagnostic reagent as well as a subunit vaccine. Conservation of the antigen in all strains would be crucial for either application. We investigated the conservation of this antigen among 36 isolates of T. foetus from Argentina, Costa Rica, and the United States using MAbs TF1.15 and TF1.17 in an enzyme-linked immunosorbent assay. MAb TF1.17 reacted with 32 of the 36 isolates, whereas MAb TF1.15 reacted with all of the isolates tested. One of the isolates which did not react with MAb TF1.17 (i.e., D1#3) was investigated further by Western blotting (immunoblotting) to determine the reason for the lack of reactivity with one of the two cross-reactive MAbs. The antigenic band that was reactive with MAb TF1.15 had a molecular mass slightly lower than that of the corresponding band from isolate D1, which reacted with both MAbs TF1.15 and TF1.17. Thus, at least a major portion of the antigen appeared to be conserved. This was confirmed in a study of heifers infected with isolate D1#3. The vaginal immunoglobulin A antibodies of these infected heifers reacted with the antigen of isolate D1 that was immunoaffinity purified with MAb TF1.17. Therefore, even though the epitope recognized by MAb TF1.17 was missing in the challenge isolate (D1#3), the heifers developed an immune response to the rest of the molecule. These results indicate that the major portion of the previously described protective antigen is conserved in different isolates of T. foetus. This portion contains the epitope that reacts with MAb TF1.15. Most isolates express the whole antigen, which possesses both TFl.15 and TF1.17 epitopes, but the few isolates that are missing the portion containing the TF1.17 epitope may still elicit an immune reponse to the conserved portion. Thus, the protective surface antigen is promising for use in immunodiagnosis or vaccination against bovine trichomoniasis.

The fine structure of normal and antibody-coated Tritrichomonas foetus cells and their interaction with rat peritoneal neutrophils was studied. Peritoneal neutrophils were obtained by glycogen stimulation. The neutrophils readily associated with and killed the parasites, which were subsequently ingested. The process involved activation of the respiratory burst, as demonstrated by the use of cytochemical methods. Images were obtained indicating that binding of parasites to the neutrophil surface triggers an exocytic response with release of oxygen-derived products. Cytochemical localization of acid phosphatase and peroxidase activities showed that leukocyte granules fused with the parasite-containing phagocytic vacuoles. We also showed the cytochemical localization of alkaline phosphatase in the parasite-neutrophil interaction.

The role of bovine antibody and complement in bovine neutrophil-mediated killing of Tritrichomonas foetus was investigated. No neutrophil-mediated trichomonacidal activity was detected when Hanks' balanced salt solution, a widely utilized and weakly buffered medium, was used. This lack of neutrophil activity was evident even in the presence of specific bovine antibody and bovine complement. Moreover, the pH of the weakly buffered Hanks' balanced salt solution was observed to fall from pH 7.0 to 5.8 in 4 h at 37 degrees C in the presence of T. foetus. The pH of 5.8 inhibited the bactericidal activity of bovine neutrophils for Staphylococcus epidermidis by 53.2% and may have contributed to the lack of neutrophil-mediated trichomonacidal activity in the weakly buffered salt solution. However, T. foetus was susceptible to bovine neutrophil-mediated destruction when a HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid)-buffered Hanks' balanced salt solution was used (21.8% killing by neutrophils alone). Neither specific bovine immune serum nor purified immune bovine immunoglobulin G2 alone enhanced bovine neutrophil-mediated killing. When complement-sensitized trichomonads were incubated with bovine neutrophils, killing of T. foetus was observed, a result which represented the additive effects of each treatment. Significant (P < 0.05) killing of trichomonads was observed when antibody- and complement-opsonized trichomonads were exposed to bovine neutrophils (> 70% parasite destruction), an effect which reflected the additive nature of each treatment.

Protection by surface antigen TF1.17 of Tritrichomonas foetus was investigated because it reacted with a monoclonal antibody which immobilized and mediated complement killing of the organism and prevented adherence to vaginal epithelial cells. This monoclonal antibody was used to demonstrate conservation of the antigen in most strains and to immunoaffinity purify the 50- to 70-kDa glycoprotein antigen. In preparation for immunization studies, the appropriate challenge dose of parasites was determined by intravaginal inoculation of 23 virgin cows (heifers) with 10(2), 10(4), or 10(6) live organisms at the time of estrus. More animals became infected and vaginal infection was maintained at a higher rate (P < 0.005) over 10 weeks for the group that received 10(6) organisms than in the other two groups. Therefore, this dose was used for challenge of immunized animals. Animals immunized with immunoaffinity-purified TF1.17 antigen in incomplete Freund's adjuvant or incomplete Freund's adjuvant plus dextran sulfate cleared the infection more quickly than adjuvant controls (P < 0.005). Isotype-specific enzyme-linked immunosorbent assay with T. foetus antigen showed that serum immunoglobulin G1 (IgG1) and IgG2 antibody responses as well as cervicovaginal mucus IgG1 and IgA antibodies peaked at about the time of clearance of infection in vaccinated animals. Controls developed later cervicovaginal mucus IgA antibody responses as would be expected in a primary local immune response to infection. These results indicate that vaccination with this immunoaffinity-purified surface antigen of T. foetus enhances antibody responses as well as clearance of the parasite from the female reproductive tract.

A photometrically-measured indirect fluorescent antibody (IFA) test was developed to measure antibody levels in three heifers vaccinated intramuscularly with two injections (two-week interval) of a vaccine containing formalin-killed Tritrichomonas foetus in oil adjuvant; a separate animal served as a nonvaccinated control. Sera were collected weekly and tested for specific IgG against T. foetus. Antibodies were detectable within two weeks of the initial injection and reached reciprocal titers as high as 7700 as estimated from previously tested reference sera. Titers peaked at six to eight weeks and remained at relatively high levels for the eleven-week study period. The fluorometric assay was easily developed and economical to perform; in addition, it more accurately estimated IgG levels than standard slide IFA tests.

To test the efficacy of a polyvalent Tritrichomonas foetus vaccine, 130 nulliparous heifers were randomly assigned to either receive the test T foetus vaccine or to serve as nonvaccinated controls. The polyvalent test vaccine consisted of a Campylobacter fetus/Leptospira canicola-grippotyphosa-hardjo-icterohaemorrhagiae-pamona bacterine containing 5 x 10(7) killed T foetus/dose. The polyvalent control vaccine consisted of the aforementioned formulation without T foetus. Heifers were administered 2 doses of control or experimental vaccine at 3-week intervals. Heifers were bred to T foetus-infected bulls and their conception and pregnancy rates were determined throughout gestation. In addition, serum samples were analyzed to determine induced concentrations of antitrichomonal antibodies and vaginal secretions were sampled to determine T foetus infection rates in control and vaccinated animals. One week after each of the 15-day breeding periods, 60% (6 of 10) of tested vaccinates and 80% (8 of 10) of tested control animals were T foetus culture-positive. The mean duration of infection of vaccinates was 3.8 weeks (+/- 7.5 days), compared with 5.4 weeks (+/- 7.5 days) of infection for control heifers. All vaccinates developed increased immunofluorescence and serum neutralizing antibody titers following the first immunization, and had additional increases of at least fourfold in response to the second injection. In contrast, no consistent increase in immunofluorescence or serum neutralizing antibodies was observed in control animals. Conception rates were 89.2% for vaccinates and 85.9% for control animals 30 days after breeding and 80 to 90% of these remained pregnant 60 days after breeding.(ABSTRACT TRUNCATED AT 250 WORDS)

A better method for diagnosis of bovine trichomoniasis is needed because culture is slow and somewhat lacking in sensitivity. Immunodiagnosis of Tritrichomonas foetus infection usually involves detection of antigen-antibody reactions with an anti-immunoglobulin conjugate. However, nonspecific immunoglobulin (Ig), bound to the surface of T. foetus, would also be detected by an anti-Ig conjugate and thus would interfere with the specificity of the immunoassay. The goals of this study were to define the binding of bovine immunoglobulins to T. foetus. To determine whether nonimmune binding of Ig to T. foetus occurs, we immunized rabbits with organisms that had been grown in medium containing normal bovine serum and vigorously washed three times with phosphate-buffered saline. The rabbit antiserum had similar titers to T. foetus and to normal bovine serum by enzyme-linked immunosorbent assay (ELISA). Furthermore, two bovine serum proteins were immunoprecipitated by the rabbit antiserum in an immunoelectrophoretogram. One of the serum proteins had a distribution characteristic of IgG2. The rabbit antiserum was then shown to react with purified bovine IgG and IgM by ELISA. Reactivity to IgG was greater. To identify the IgG subisotypes bound and to confirm nonimmune binding of Ig, we grew T. foetus in agammaglobulinemic fetal calf serum and reacted it with IgG1, IgG2, and IgM specific for dinitrophenol (DNP) in ELISA. The binding of IgG2 was greatest, that of IgG1 was next, and that of IgM was least. Little competitive inhibition by DNP was detected, indicating that binding of DNP-specific antibodies was predominantly nonimmune rather than antigen-specific Ig binding. We also demonstrated that T. foetus grew well in medium containing agammaglobulinemic fetal calf serum or serum made agammaglobulinemic by ammonium sulfate precipitation of Igs. This may overcome the problem of low specificity in diagnostic assays as a result of antigen with Ig bound by nonimmune mechanisms.