Tritrichomonas foetus causes bovine trichomonosis, a venereal disease that reduces productivity in naturally mated cattle. Its high prevalence in Northern Australian herds underscores the need for a locally made strain-specific vaccine. This study developed and tested a whole-cell killed T. foetus vaccine using the Queensland isolate TfOz5 (vaccine strain) and TfOz-N36 (Northern Territory isolate) as the challenge strain. The heat-inactivated vaccine, adjuvanted with Montanide ISA 61 VG, was administered subcutaneously in 2 doses (5 × 10⁷ cells/dose) at a 1-month interval to mature bulls (n = 6) (4–7 years old), while controls (n = 6) (4–8 years old) received adjuvant with PBS. Bulls were experimentally challenged intrapreputially with live cultures of T. foetus at 2- and 6-months post first vaccination. A therapeutic trial with T. foetus-positive, persistently infected mature bulls (n = 10) (4–7 years old) used the same vaccine regime without the subsequent T. foetus challenges. The vaccine was found to be safe, causing only mild local reactions. The vaccine challenge experiment demonstrated similar duration of T. foetus positivity, confirmed by quantitative polymerase chain reaction (qPCR), compared to controls (94 vs. 106 days, P = 0.73). In the therapeutic experiment, 2/10 treated bulls tested negative for T. foetus at the end of the trial, while the remaining eight remained positive. Vaccinated bulls in both experiments showed significantly elevated serum anti-T. foetus IgG antibody levels, confirming the vaccine’s immunogenicity. These findings demonstrate that the experimental vaccine is safe and capable of eliciting a specific immune response in mature bulls.

Bovine trichomonosis caused by Tritrichomonas foetus is a significant reproductive disease of cattle. Preputial samples were collected using sheath washing technique in bulls in Namibia. Thirty-six trichomonad cultures were characterized using the TaqMan-probe commercial real-time polymerase chain reaction (PCR) diagnostic assay (VetMAX™-Gold Trich Detection Kit) and CYBR real-time PCR assay based on TFR3/4 primers. Diagnostic real-time PCRs and DNA sequencing of the internal transcribed region confirmed presence of T. foetus in 35 out of 36 samples. Multilocus genotyping using cysteine proteases (CP1, CP2, CP4, CP5, CP6, CP7, CP8, CP9) and malate dehydrogenase (MDH1) gene sequences demonstrate that the T. foetus in Namibia are genetically distinct from those characterized elsewhere. We report the discovery of a novel genotype of T. foetus in Namibian cattle, distinct from other T. foetus genotypes in Europe, South and North America and Australia. We suggest recognition of a ‘Southern African’ genotype of T. foetus. Identification of the new genotype of T. foetus demonstrates the need for wider global sampling to fully understand the diversity and origin of T. foetus causing disease in cattle or cats.

Iron is an essential element for the survival of trichomonads during host–parasite interaction. The availability of this metal modulates several metabolic pathways of the parasites and regulates the expression of virulence factors such as adhesins and proteolytic enzymes. In this study, we investigated the effect of iron depletion on the morphology and life cycle of Tritrichomonas foetus. Scanning and transmission electron microscopy analyses revealed that depletion of iron from the culture medium (named TYM-DIP inducer medium) induces morphological transformation of typical pear-shaped trophozoites into spherical and non-motile pseudocysts. Remarkably, inoculation of pseudocysts into an iron-rich medium (standard TYM medium), or addition of FeSO4 to a TYM-DIP inducer medium reverted the morphological transformation process and typical trophozoites were recovered. These results show that pseudocysts are viable forms of the parasite and highlight the role of iron as a modulator of the parasite phenotype. Although iron is required for the survival of T. foetus, iron depletion does not cause a cellular collapse of pseudocysts, but instead induces phenotypic alterations, probably in order to allow the parasite to survive conditions of nutritional stress. Together, these findings support previous studies that suggest pseudocysts are a resistance form in the life cycle of T. foetus and enable new approaches to understanding the multifactorial role of iron in the cell biology of this protozoan parasite.

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.

Malate dehydrogenase (decarboxylating) from Tritrichomonas foetus hydrogenosomes was purified close to homogeneity by a combination of differential centrifugation, zwitterionic detergent solubilization, Red-Sepharose chromatography and anion-exchange chromatography. The enzyme with apparent subunit size of 59 kDa and native molecular mass of 308 kDa utilized NAD+ preferentially to NADP+ as a cofactor and required Mn2+ or Mg2+ for its activity. Affinity curves for malate and coenzymes were hyperbolic. Km for malate was 100 μM and 458 μM in the presence of NAD+ and NADP+, respectively. Km for NAD+ and for NADP+ in the presence of malate was 18 μM and 207 μM, respectively. The enzyme is proposed to be a tetramer with a possible physiological role in the maintenance of an appropriate NAD+/NADH ratio in hydrogenosomes.

Lysis of Tritrichomonas foetus with a solution of the non-ionic detergent Triton X-114 at 0 °C, followed by low-speed centrifugation, resulted in a detergent-insoluble pellet and a detergent-soluble supernatant. The supernatant was further fractionated by phase separation at 30 °C into a detergent-rich phase and an aqueous phase. Neuraminidase activity was mostly located in the detergent-insoluble pellet. When the parasites were incubated with bacterial phosphatidylinositol phospholipase C (PI–PLC) prior to detergent solubilization and phase separation neuraminidase activity was predominantly recovered in aqueous phase, rather than in the pellet and detergent phase. The molecular mass determined by gel permeation in high performance liquid chromatography (HPLC) and SDS–PAGE was 80000 Da. Indirect immunofluorescence microscopy using polyclonal antibodies raised in rabbits against the purified neuraminidase, indicated that the enzyme is exposed on the cell surface. Previous treatment of the cells with PI–PLC significantly reduced antibody binding. Incubation of cryo-sections with the antibodies followed by detection using gold-labelled anti-rabbit IgG confirmed the presence of neuraminidase in the plasma membrane enclosing the cell body and flagella and in the membrane of vesicles preferentially located at the peripheral region of the protozoan.