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Objective: To determine the presence of Tritrichomonas foetus in the accessory sex glands, epididymis, and testicular tissue of T foetus-positive bulls and to assess the impact of post mortem tissue acidity on quantitative reverse-transcription PCR results., Methods: 8 bulls were euthanized from June 12 through June 15, 2023, and samples from their accessory sex glands, testicles, and epididymis were collected for T foetus testing. Preputial smegma was obtained before and after euthanasia and tested for T foetus by quantitative reverse-transcription PCR. The pH of the penile and preputial epithelial surfaces was measured., Results: Tritrichomonas foetus was not detected in samples from the testicle, epididymis, or epididymal semen. A post mortem decrease in the pH of the penis and prepuce correlated with a rise in cycle threshold values., Conclusions: Tritrichomonas foetus was not present in samples taken, suggesting its absence in these specific reproductive tissues. A decrease in the pH of the penis and prepuce over time post mortem correlated with an increase in cycle threshold values in the samples. This relationship indicates that changes in pH may impact the detectability or stability of genetic material in these tissues, possibly affecting diagnostic outcomes as time from death progresses., Clinical Relevance: This research enhances the understanding of T foetus pathology in bulls and suggests that epididymal semen recovery could be a viable method for preserving genetic material from valuable bulls that test positive for T foetus. It also emphasizes the need for timely post mortem sampling to ensure accurate detection and management of T foetus in the cattle industry.

Tritrichomonas foetus (T. foetus) , the causative agent of bovine trichomoniasis, is an obligate protozoan parasite of the bovine reproductive tract and can be found on the penis, prepuce, and distal urethra of the bull and from the cranial vagina to the oviduct in the infected cow. To date, the microbiome of bulls infected with T. foetus has not been described. The objectives of this study were to (1) describe the preputial and penile microbiome of bulls chronically infected by T. foetus, (2) describe the seminal microbiome of T. foetus -infected bulls, and (3) evaluate different collection devices that could be used for sampling. Eleven bulls naturally infected with T. foetus were utilized for the collection of samples. Samples were obtained during the process of a routine breeding soundness exam utilizing either a dacron swab, pizzle stick, double-guarded swab, or semen collection. The preputial and seminal microbiome of T. foetus -infected bulls was dominated by bacterial members of the phyla Fusobacteriota, Firmicutes , Bacteroidota, Actinobacteria, and Campylobacterota. Semen collection yielded the most microbial diversity; however, there was no significant difference between the four methods ( p ≥ 0.05). This study characterizes both the preputial and seminal microbial communities of bulls chronically infected by T. foetus.

Background: In recent years, the trichomonosis in raccoon dogs in China had occurred frequently. Pentatrichomonas hominis had been described in raccoon dogs in China in some previous studies. PURPOSE TO REVEAL: whether raccoon dogs can be infected by other trichomonad species besides P. hominis, and clarify the prevalence and species distribution of trichomonad in raccoon dogs., Methods: Herein, the 389 fecal samples were collected from farm-raised raccoon dogs in Hebei Province, all the samples were detected using the microscopic examination and several fecal samples containing trichomonad-like organisms were processed, cultured, stained, and photographed. Meanwhile, all the samples were screened by the species-specific nested PCR based on the small subunit rRNA (SSU rRNA) gene of P. hominis,Tritrichomonas foetus and Tetratrichomonas buttreyi, respectively, and all positive secondary PCR amplications obtained in this study were sequenced, aligned and analysed., Results: 62 fecal samples (15.9%,62/389) were trichomonad-positive under light microscopy, and the trichomonad-like cells were clearly observed in the culture contents. The PCR results showed that 100 samples were trichomonad-positive, including 45 P. hominis-positive samples (11.6%,45/389), 32 T. foetus-positive samples (8.2%,32/389), and 33 T. buttreyi-positive samples (8.5%,33/389), respectively. Double mixed infections were observed in 10 samples. The prevalence of T. foetus and P. hominis were both significantly higher in raccoon dogs with diarrhea (13.9%, and 25.0%) than that in raccoon dogs without diarrhea (7.6%, and 9.3%) (p < 0.05).All samples confirmed as trichomonad-positive under microscopy were also found to be trichomonad-positive by PCR analysis. The sequencing and phylogenetic analysis demonstrated the sequences obtained in this study belonged to P. hominis, T. foetus and T. buttreyi SSU rRNA, respectively. Among them, the T. buttreyi SSU rRNA sequences obtained in this study harbored the new sequence polymorphisms. Based on preliminary morphological and molecular analyses, raccoon dogs are considered as the new host of T. foetus and T. buttreyi., Conclusion: This is the first report about the identifcation and prevalence of T. foetus and T. buttreyi in raccoon dogs in China, and the results increase our knowledge about the host range and prevalence of trichomonad species., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Objective: To provide a video tutorial on how to perform preputial scraping for the testing of Tritrichomonas foetus in bulls., Animals: Postpubertal bulls., Methods: A preputial scraping device is inserted in the prepuce, and back-and-forth scraping movements are made to collect smegma and T foetus organisms. The sample is placed in an appropriate media and prepared for shipment to the diagnostic laboratory., Results: Preputial sampling is an effective method for diagnosing T foetus in infected bulls., Clinical Relevance: Bulls are the primary carriers of T foetus, a protozoan parasite responsible for bovine trichomoniasis. By sampling and testing bulls, veterinarians can identify infected animals and implement control measures to prevent the spread of the disease within herds. Trichomoniasis can lead to significant reproductive problems in cattle, including infertility, embryonic death, and abortions. Sampling bulls allows for the detection of infection, enabling prompt intervention to protect the reproductive health of the entire herd. Trichomoniasis outbreaks can result in substantial economic losses for cattle producers due to reduced conception rates, increased calving intervals, and decreased calf crops. Sampling bulls helps to mitigate these losses by identifying and removing infected animals from the breeding pool, thereby minimizing the spread of the disease and its associated reproductive inefficiencies. In many regions, T foetus testing in bulls is a regulatory requirement for cattle movement and trade. Sampling bulls and obtaining negative test results are often necessary for obtaining health certificates and complying with interstate or international movement regulations.

The incorporation of different molecules by eukaryotic cells occurs through endocytosis, which is critical to the cell's survival and ability to reproduce. Although this process has been studied in greater detail in mammalian and yeast cells, several groups working with pathogenic protists have made relevant contributions. This review analysed the most relevant data on the endocytic process in anaerobic protists (Entamoeba histolytica, Giardia intestinalis, Trichomonas vaginalis, and Tritrichomonas foetus). Many protozoa can exert endocytic activity across their entire surface and do so with great intensity, as with E. histolytica. The available data on the endocytic pathway and the participation of PI-3 kinase, Rab, and Rho molecular complexes is reviewed from a historical perspective.

Background: Trichomonosis is a common infection in small animals, mostly manifesting in gastrointestinal symptoms such as diarrhea. Although oral trichomonads are also known, the species found colonizing the large intestine are more frequently detected protozoa., Methods: In the present study, four wildcats, 94 domestic cats, and 25 dogs, originating from 18 different locations in Hungary, were investigated for the presence of oral and large intestinal trichomonads based on the 18S rRNA gene and ITS2., Results: All oral swabs were negative by polymerase chain reaction (PCR). However, Tritrichomonas foetus was detected in a high proportion among tested domestic cats (13.8%) and dogs (16%), and Pentatrichomonas hominis only in two domestic cats. In addition, a novel Tritrichomonas genotype was identified in one cat, probably representing a new species that was shown to be phylogenetically most closely related to Tritrichomonas casperi described recently from mice. All positive dogs and half of the positive cats showed symptoms, and among cats, the most frequent breed was the Ragdoll., Conclusions: With molecular methods, this study evaluated the prevalence of oral and intestinal trichomonads in clinical samples of dogs and cats from Hungary, providing the first evidence of T. foetus in dogs of this region. In contrast to literature data, P. hominis was more prevalent in cats than in dogs. Finally, a hitherto unknown large intestinal Tritrichomonas species (closely related to T. casperi) was shown to be present in a cat, raising two possibilities. First, this novel genotype might have been a rodent-associated pseudoparasite in the relevant cat. Otherwise, the cat was actually infected, thus suggesting the role of a predator-prey link in the evolution of this trichomonad., (© 2024. The Author(s).)

Tritrichomonas foetus (T. foetus) is a protozoal pathogen that infects cats and constitutes a significant cause of chronic colitis and diarrhea. Perturbations in the gut microbiota (GM) are affected by Trichomonas infection. Furthermore, dysregulation of the host GM enhances Trichomonas pathogenicity. However, it remains unclear whether the occurrence of diarrhea is associated with a dysregulation in GM following T. foetus infection in cats. Hence, the primary objective of this investigation was to explore the correlation between T. foetus infection and dysregulation in GM by analyzing fecal samples obtained from pet cats in Henan Province, central China. We randomly collected 898 fecal samples from pet cats living in 11 prefectural cities within Henan Province, and T. foetus was screened with polymerase chain reaction (PCR) amplification based on the 18 S rRNA gene. Subsequently, six T. foetus-positive and six T. foetus-negative samples underwent analysis through 16 S rRNA gene sequencing to evaluate the gut microbiota's composition. The overall prevalence of T. foetus infection among the collected samples was found to be 6.01% (54/898). Notably, a higher prevalence of infection was observed in young, undewormed, unimmunized, and diarrheic pet cats. T. foetus infection was found to significantly alter the composition of the pet cat fecal microbiota, leading to dysfunctions. Moreover, it resulted in a substantial increase in the abundance of Bacteroidetes, Proteobacteria, and Phascolarctobacterium spp., while decreasing the ratio of Firmicutes to Bacteroidetes (F/B) and the abundance of Actinobacteria, Clostridiaceae&#95;Clostridium spp., Phascolarctobacterium spp., SMB53 spp., and Blautia spp. We constructed ROC curves to assess the diagnostic value of specific bacterial taxa in discriminating T. foetus infection. The analysis revealed that Proteobacteria and Clostridiaceae&#95;Clostridium spp. were the most reliable single predictors for T. foetus infection. This finding suggests that alterations in the GM may be strongly associated with T. foetus infections., Competing Interests: Declaration of Competing Interest None., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The protozoan parasite Tritrichomonas foetus (T. foetus) is the causative organism of bovine trichomonosis (also referred to as trichomoniasis), a sexually-transmitted infection that reduces fertility in cattle. Efforts to control trichomonosis on cattle farms are hindered by the discouragement of antibiotic use in agriculture, and the incomplete, short-lived protection conferred by the current vaccines. A more complete mechanistic understanding of what effective immunity to T. foetus entails could enable the development of more robust infection control strategies. While neutrophils, the primary responders to infection, are present in infected tissues and have been shown to kill the parasite in vitro, the mechanism they use for parasite killing has not been established. Here, we show that primary bovine neutrophils isolated from peripheral blood rapidly kill T. foetus in vitro in a dose-dependent manner, and that optimal parasite killing is reduced by inhibitors of trogocytosis. We also use imaging to show that bovine neutrophils surround T. foetus and trogocytose its membrane. These findings are consistent with killing via trogocytosis, a recently described novel neutrophil antimicrobial mechanism., (© 2023. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Bovine genital campylobacteriosis (BGC) and bovine trichomonosis (BT) are sexually transmitted diseases (STDs) that affect bovine breeding herds, decreasing their reproductive efficiency. The objective of this work was to estimate the prevalence of these diseases and their temporal-spatial distribution in the province of Formosa, Argentina. The cross-sectional study conducted between 2018 and 2021 included a total of 15,571 bulls, inter-herd prevalence being 29.62% and 17.23% for BGC and BT, respectively. The prevalence of positive animals was 2.05% for BGC and 0.43% for BT. The temporal-spatial analysis of BGC showed two distinct spatial groupings, one group had a low risk of contracting the disease (RR = 0.13; p < 0.001; 2018-2021) while the other group had a high risk (RR = 2.84; p < 0.001; 2020-2021). BT had a high-risk group for the disease (RR = 35.24; p < 0.001; 2019). This study shows that STDs are endemic in the region, providing updated and valuable information as a tool for the health management of these diseases., (Copyright © 2023. Publicado por Elsevier España, S.L.U.)

Bovine genital campylobacteriosis, caused by the gram-negative bacteria Campylobacter fetus venerealis, and bovine trichomonosis, caused by the parasite protozoan Tritrichomonas foetus, are venereal diseases that occur with long intercalving periods and abortion. The control of both diseases relies on microbiological testing and culling infected bulls. Vaccination and antibiotic treatment may help in controlling campylobacteriosis but are not recommended for trichomonosis control. Several regions of the world have active control programs for trichomonosis, not campylobacteriosis. In Argentina, the state of La Pampa aims to eradicate trichomonosis and campylobacteriosis by imposing annual diagnostic testing of every bull and slaughtering positive animals. Prior studies indicated a declining trend in the prevalence of campylobacteriosis and trichomonosis in La Pampa. It was also proposed that the prevalence of one disease could be estimated from the prevalence of the other. The purpose of this retrospective analysis of data gathered from 2008 to 2021 was to determine the La Pampa program's efficacy. Descriptive statistics were employed to determine the reason behind the correlation between tricomonosis and campylobacteriosis diagnostic results. The outcomes refute the notion that this program of venereal eradication was a success. Furthermore, an excess of false positives in both diagnoses may have contributed to the correlation between the prevalences of campylobactriosis and trichomonosis. The practice of killing animals without verifying positive results hinders the determination of disease prevalence and results in the death of numerous healthy animals., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

There are no recommended drugs to treat cattle infected with the protozoan Tritrichomonas foetus (TF). Ivermectin, widely used in the treatment of intestinal parasites, was found effective against some protozoa growing in vitro. Here, its effectiveness against a TF line was investigated. Trophozoites were incubated in media with increasing concentrations of ivermectin and mortality was determined after 24 h. Ivermectin killed cells with a mean maximum effective concentration (EC50) of 2.47 μg mL-1. The effective concentration of ivermectin was rather high for a formulation suitable for systemic treatment. However, topical treatment of animals against TF could still be considered and tested.

Tritrichomonas foetus is a protozoan parasite that causes a venereal disease in cattle limiting reproduction by abortions and sterility. The immune response against this parasite is poorly understood. Since the iron and calcium ions are important regulators of the microenvironment of the urogenital tract in cattle, we decided to evaluate the role of these divalent cations on the antigenicity of membrane proteins of T. foetus on macrophage activation as one of the first inflammatory responses towards this pathogen. Colorimetric methods and ELISA were used to detect the nitric oxide and oxygen peroxide production and expression of cytokines in culture supernatant from macrophage incubated with membrane proteins from T. foetus cultured in iron- and calcium-rich conditions. qRT-PCR assays were used to evaluate the transcript expression of genes involved in the inflammatory response on the macrophages. The membrane proteins used for in vitro stimulation caused the up-regulation of the iNOS and NOX-2 genes as well as the generation of NO and H 2 O 2 in murine macrophages on a dependent way of the metal concentrations. Additionally, after stimulation, macrophages showed a considerable rise in pro-inflammatory cytokines and a downregulation of anti-inflammatory cytokines, as well as up-regulation in the transcription of the TLR4 and MyD88 genes. These data suggest that membrane proteins of T. foetus induced by iron and calcium can activate an inflammatory specific macrophage response via TLR4/MyD88 signalling pathway., (© 2024 John Wiley & Sons Ltd.)

Tritrichomonas foetus is a flagellated parasite that primarily infects the reproductive tissues of livestock, causing bovine trichomoniasis. The cytoplasmic membrane of T. foetus contains various compounds that contribute to adherence, colonization, and pathogenicity. Metronidazole (MTZ) is the main treatment for trichomoniasis, but the emergence of drug-resistant strains is a concern due to improper use and dosing. T. foetus infection induces inflammation, and macrophages are key players in the immune response. However, our understanding of the host's immune response to T. foetus is limited, and the specific mechanisms underlying these responses are not well understood. This study aimed to investigate the impact of T. foetus surface proteins from trophozoites cultured under different sublethal MTZ conditions (MTZ-treated T. foetus MPs) on macrophage activation. By analyzing cytokine levels and gene expression in murine macrophages, we demonstrated that MTZ-treated T. foetus MPs induce a specific proinflammatory response. MTZ-treated T. foetus MPs-exposed macrophages exhibited a higher NO and H 2 O 2 production and overexpression of iNOS and NOX-2 genes in comparison to untreated T. foetus. Additionally, MTZ-treated T. foetus MPs triggered a significant induction of the proinflammatory cytokines IL-1β, IL-6, TNF-α, and IFN-γ, as well as the overexpression of the TLR4, MyD88, and NF-κB genes on murine macrophages. The study aimed to unravel the immunological response and potential proinflammatory pathways involved in T. foetus infection and MTZ stress. Understanding the immune responses and mechanisms through which T. foetus surface proteins activate macrophages can contribute to the development of new therapeutic strategies for controlling bovine trichomoniasis., (© 2023 International Society of Protistologists.)

Importance: Nowadays, the routine herd diagnosis is usually performed exclusively on bulls, as they remain permanently infected, and prevention and control of Tritrichomonas foetus transmission are based on identifying infected animals and culling practices. The existence of other forms of transmission and the possible role of pseudocysts or cyst-like structures as resistant forms requires rethinking the current management and control of this parasitic disease in the future in some livestock regions of the world., Competing Interests: The authors declare no conflict of interest.

Light microscopy has significantly advanced in recent decades, especially concerning the increased resolution obtained in fluorescence images. Here we present the Expansion Microscopy (ExM) technique in two parasites, Trichomonas vaginalis and Tritrichomonas foetus, which significantly improved the localization of distinct proteins closely associated with cytoskeleton by immunofluorescence microscopy. The ExM techniques have been used in various cell types, tissues and other protist parasites. It requires the embedment of the samples in a swellable gel that is highly hydrophilic. As a result, cells are expanded 4.5 times in an isotropic manner, offering a spatial resolution of ∼70 nm. We used this new methodology not only to observe the structural organization of protozoa in more detail but also to increase the resolution by immunofluorescence microscopy of two major proteins such as tubulin, found in structures formed by microtubules, and costain 1, the only protein identified until now in the T. foetus's costa, a unique rod-shaped like structure. The individualized microtubules of the axostyle were seen for the first time in fluorescence microscopy and several other details are presented after this technique., Competing Interests: Declaration of competing interest We do not have., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Bovine Trichomonosis (BT), a sexually transmitted disease endemic in countries with extensive cattle farming and natural service, is one of the most common causes of reproductive failure. 5-nitroimidazoles and their derivatives are used for its treatment, mainly metronidazole. The emergence of drug resistance mechanisms and treatment failures raise the need to investigate the effectiveness of new active compounds that contribute to parasite control. In this regard, extracts of Lantana camara (Verbenacea) have shown high biocidal potential against isolates of Trypanosoma cruzi and Leishmania braziliensis in vitro assays, although their effect on Tritrichomonas foetus has not been demonstrated yet. The available information on in vitro susceptibility of trichomonicidal drugs comes from the use of a diversity of methodologies and criteria, especially the observation of parasite motility under the optical microscope to assess their viability. Recently, in our laboratory, the use of flow cytometry has been described for the first time as a rapid and efficient method to evaluate the viability of T. foetus against metronidazole. The present study aimed to evaluate the cytostatic effect of L. camara extracts against T. foetus isolates by flow cytometry. Under aerobic conditions, IC 50 values of 22.60 µg/mL were obtained on average. Under anaerobic conditions, the IC 50 oscilated around 29.04 µg/mL. The results obtained allowed describing the susceptibility exhibited by these protozoa, being a valuable information for the development of potential BT treatments., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Tritrichomonas foetus is the causative agent of bovine trichomonosis, a venereal disease that can lead to fetal loss. T. foetus proliferates in the vagina and cervix and invades the uterus and fetal cavities. It is not clear how T. foetus grows in the host or how its infection rarely demonstrable after 4 months, is controlled. Cervical vaginal mucus (CVM) is a protective barrier against potentially harmful microorganisms. Here, we demonstrate that bovine CVM is a medium in which this protozoan parasite can grow in vitro. T. foetus multiplied at different rates depending on the time of the estrous period from which the CVM was obtained. Growth rates were higher in CVM obtained 7-10 days after estrus., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Bovine trichomonosis (BT), a disease of the bovine urogenital tract, is caused by the protozoan Tritrichomonas foetus (Tf). Tf causes endometritis, infertility, and premature death of the embryo, which generates considerable economic losses. The proteins released can mediate fundamental interactions between the pathogen and the host, triggering factors associated with the symptomatology, immune evasion and pathogenesis characteristic of the species. However, little is known about the profile of the proteins released by Tf. In order to contribute to their knowledge, we performed an isolation protocol and a proteomic profiling of the supernatant (SN) content of six Tf isolates. A total of 662 proteins present in the SN of Tf were detected, out of which 121 were shared by the six isolates, while the remaining 541 were found in at least one of the isolates studied. The comparative analyses using the databases of Tf strain genome K revealed 32.9% of uncharacterized proteins. The bioinformatic analyses showed that the main molecular functions predicted were binding (47.9%) and catalytic activity (38.2%). Additionally, we performed immunodetection assays to evidence the antigenic potential of SN proteins. Interestingly, we observed great ability to detect SN proteins from all six isolates using serum from immunized mice and infected bulls. A complementary mass spectrometry assay allowed us to determine that the proteins that showed the strongest signal intensity in the immunoassays were Grp78 (A0A1J4IZS3) and Ap65 (A0A1J4JSR1). This work represents the first proteomic characterization of Tf SN proteins and their antigenic potential, which might be interesting for the future design of new diagnosis and treatment methods for BT., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Tritrichomonas foetus (TF) is a significant reproductive pathogen of cattle, and sample collection, handling, transport, and testing are significant hurdles to surveillance programs. Recent methods have been developed that allow for the direct detection of TF using a reverse transcription real-time PCR (direct RT-qPCR) approach. To evaluate these methods, a comparative analysis was conducted to assess the technical performance of this assay with a commercially available real-time PCR (qPCR) assay. In addition, the evaluation of two types of collection media (PBS and TF transport tube) was conducted that evaluated sample stability from 0 to 3 days when stored at 4°C or 25°C. Extended incubation times for PBS media were also evaluated (5, 7, and 14 days) at both refrigeration and frozen temperatures to evaluate the effect of extended transport time on samples. Limits of detection (LODs), dynamic range, and RNA stability were assessed using lab-cultured TF spiked into samples of normal bovine smegma collected in PBS or TF transport media, and performance was assessed on field samples collected in parallel. 100% agreement was found between direct RT-qPCR and qPCR at 10 parasites/extraction and a LOD of 1 parasite/extraction. Differences in detection were not observed in either collection media when incubated at either temperatures for up to 3 days of incubation. In addition, the extended incubation experiments indicate that samples containing 10 parasites/extraction can be detected at 4°C for 5 days with a mean Cq 26.34 (95% CI: 23.11-29.58) and detected at -20°C for 7 or 14 days, with a mean Cq 29.55 (95% CI: 27.73-31.37). A significant decrease in detectable RNA was observed in samples containing <10 parasites/extraction at -20°C for 14 days, which should be considered for long-term storage. In summary, direct RT-qPCR was found to be equivalent or superior to qPCR and PBS was not significantly different from TF transport media. The findings of the current study allows for more flexibility during sample collection and transport and ultimately enhancement of TF surveillance programs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Loy, Spuri Gomes, Dutta, Brodersen and Loy.)

Background and Aim: Bovine trichomoniasis, caused by Tritrichomonas foetus , is a venereal disease that is distributed in many countries, including Iraq. Compared with other abortive infectious diseases, prevalence of T. foetus is expected to be relatively low in the field by veterinarians. This study aimed to estimate the prevalence of T. foetus in aborted cows by conventional polymerase chain reaction (PCR) and phylogenetic analysis of local T. foetus isolates was documented in the National Center for Biotechnology Information as the first sequenced isolates from Iraq., Materials and Methods: Vaginal fluids were collected from 62 aborted cows and examined by PCR. Data were reported for the following parameters: Vital signs (body temperature and respiratory and pulse rates), age (<4, 4-8, and >8 years), reproductive health status (premature calving, embryonic death, pyometra, and healthy newborn), breed (pure or crossbred), type of breeding (natural or artificial), bull-to-cow ratio (1:<10, 1:10-20, and 1:>20), contact of cow with bull(s) from other farmers (yes or no), and contact with stray animals (dogs and cats)., Results: A total of 20.97% of aborted cows were positive for T. foetus . Phylogenetic analysis for 10 positive local T. foetus isolates demonstrated high identity with the Thai (MN560972.2) and Chinese (MH115435.1) isolates, with an identity range of 98.8%-99.5% and 98.6%-99.3%, respectively. Clinical data showed that the vital signs differed insignificantly between cows positive and negative for T. foetus . Prevalence and risk of infection increased significantly in <4-year-old, early calving, embryonic death, crossbred, and naturally inseminated cows that had direct contact with bulls from other farmers, and contact with stray animals. Fetal pneumonia and death of premature calves were significant among positive aborted fetuses., Conclusion: Tritrichomonas foetus is highly prevalent in aborted cows in Iraq and phylogenetic analysis demonstrated an identity between the local and global isolates, that is, Thai and Chinese, of cats., Competing Interests: The author declares that he has no competing interests., (Copyright: © Gharban.)

Trichomonosis is a venereal disease of cattle caused by the protozoan Tritrichomonas foetus. T. foetus infection in cattle herds can be economically costly for cattle producers; therefore, testing is important for detection of the agent. Given that bulls are considered to be subclinical carriers of T. foetus , it is important to detect T. foetus infection prior to movement and/or breeding season. We have described previously the development of an updated set of PCR primers and probes that offer increased sensitivity of T. foetus detection in preputial washings collected in PBS by utilizing reverse-transcription real-time PCR (RT-rtPCR) that targets the 5.8S ribosomal RNA of the T. foetus organism. Here, we report improvements in the updated RT-rtPCR reagents as well as the evaluation of testing of pooled preputial washings. We found that up to 5 preputial washings can be pooled, similar to routine testing practices (InPouch culture), without reducing the sensitivity of detection of T. foetus.

Tritrichomonas foetus and Trichomonas vaginalis are extracellular flagellated parasites that inhabit animals and humans, respectively. Cell division is a crucial process in most living organisms that leads to the formation of 2 daughter cells from a single mother cell. It has been assumed that T. vaginalis and T. foetus modes of reproduction are exclusively by binary fission. However, here, we showed that multinuclearity is a phenomenon regularly observed in different T. foetus and T. vaginalis strains in standard culture conditions. Additionally, we revealed that nutritional depletion or nutritional deprivation led to different dormant phenotypes. Although multinucleated T. foetus are mostly observed during nutritional depletion, numerous cells with 1 larger nucleus have been observed under nutritional deprivation conditions. In both cases, when the standard culture media conditions are restored, the cytoplasm of these multinucleated cells separates, and numerous parasites are generated in a short period of time by the fission multiple. We also revealed that DNA endoreplication occurs both in large and multiple nuclei of parasites under nutritional deprivation and depletion conditions, suggesting an important function in stress nutritional situations. These results provide valuable data about the cell division process of these extracellular parasites. IMPORTANCE Nowadays, it's known that T. foetus and T. vaginalis generate daughter cells by binary fission. Here, we report that both parasites are also capable of dividing by multiple fission under stress conditions. We also demonstrated, for the first time, that T. foetus can increase its DNA content per parasite without concluding the cytokinesis process (endoreplication) under stress conditions, which represents an efficient strategy for subsequent fast multiplication when the context becomes favorable. Additionally, we revealed the existence of novel dormant forms of resistance (multinucleated or mononucleated polyploid parasites), different than the previously described pseudocysts, that are formed under stress conditions. Thus, it is necessary to evaluate the role of these structures in the parasites' transmission in the future.

Bovine tritrichomonosis, a sexually transmitted disease caused by the protozoan Tritrichomonas foetus, is characterized by producing reproductive alterations in cattle. Carbohydrates on the surface of the uterine epithelium are involved in the process of adhesion and colonization of the protozoan. The murine model has proved to be an inexpensive, practical and representative alternative to study the lesions produced in the natural host. For this work, during the first stage, 6-8 week old female BALB/c mice were inoculated with 24 different T. foetus isolates in order to classify them according to their pathogenicity. Then, seven isolates were selected and processed with lectin histochemistry to determine if the differences in pathogenicity corresponded to the changes found in the uterine carbohydrate expression pattern. In this work, we demonstrate the differences in the expression of the carbohydrate pattern between infected and uninfected mice. In addition, within the group of infected mice, differences were found in the degree of pathogenicity of the isolates, thus evidencing their biological variability., Competing Interests: Declaration of Competing Interest All authors of this manuscript declare that they have no financial competing interests., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The aim of this study was to identify Neosopora caninum, Toxoplasma gondii and Tritricomonas foetus in all cattle aborted fetus samples and N. caninum and T. gondii in sheep and goat aborted fetuses sent to Elazig Veterinary Control Institute during two years. Total genomic DNAs were obtained using a commercial kit. Real-time PCR analysis was performed separately for each agent. Conventional PCR was set up for confirmation of positive samples. Then, fetal brain, heart, lung and liver samples were analysed by hematoxylin-eosin (HE) and Avidin-Biotin Complex (ABC) Immunohistochemistry (IHC) methods. Totally, we tested 55 aborted fetus samples. Of these samples, seven (12.7 %) was belonged to goats, 18 (32.7 %) to sheep and 30 (54.5 %) to cattle. T. gondii was detected in six (10.90 %) samples, and four (7.27 %) of them were positive with Real-time PCR, while only one of these four samples was positive for both classical PCR and IHC. N. caninum was determined by at least one of the three tests in 14 (25.45 %) of the samples studied, while 8 (14.54 %) of the positive samples were detected by Real-time PCR, only two of them were also positive with conventional PCR, eight (14.54 %) samples was determined as positive by IHC. Considering T. foetus in the samples, positivity was determined in two (3.63 %) of 55 aborted fetus (both of which were aborted cattle fetus) by Real-time PCR, while only one of them was positive with conventional PCR, while no positivity was detected with the IHC., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The parasite T. foetus causes trichomonosis in cattle but is generally asymptomatic in males. Thus, many bulls carrying the disease go unnoticed, making the detection of T. foetus in bulls an important aspect for its control. Due to drawbacks posed by its cultivation, PCR is a preferred option for diagnostic laboratories. Most published PCR protocols target the genomic region compring the 18S, 5.8S, and 28S rRNA genes and internal transcribed spacers 1 and 2 (rRNA-ITS region), homologous to that of other Tritrichomonas species. There is minimal information on alternative genetic targets and no comparative studies have been published. We compared a protocol based on the microsatellite TfRE (called H94) and five protocols based on the rRNA-ITS region (called M06, M15, G02, G05, and N02). We also designed and evaluated a novel PCR-based assay on the EF1-alpha-Tf1 gene (called V21). The analytical sensitivity and specificity assays for the PCR protocols were performed according to the World Organisation for Animal Health (OIE) directives and the comparative study was performed with a widely used PCR (M06) on clinical samples from 466 breeding bulls. V21 showed a high degree of agreement with our reference M06 (kappa = 0.967), as well as M15 (kappa = 0.958), G05 (kappa = 0.948), and H94 (kappa = 0.986). Protocols H94 and V21 appear to be good approaches for confirming clinical cases in preputial bull samples when genomic regions alternative to rRNA-ITS are required. By contrast, N02 gave false negatives and G02 false positives., (© 2022. The Author(s).)

Feline trichomonosis occurs worldwide, with gastrointestinal symptoms such as chronic large-bowel diarrhea and abdominal pain. The inclusion of molecular methods in diagnostic and epidemiological studies has necessitated an effective method for extracting DNA from feces. We tested four extraction commercial kits: ZR Fecal DNA MiniPrep (50 preps) (Zymo Research, Irvine, CA, USA), QIAamp ® DNA Stool Mini Kit (Qiagen Inc., Valencia, CA, USA), UltraClean Fecal DNA Kit (50 preps) (MO BIO, San Diego, CA, USA), and Sherlock AX/100 isolations (A&A Biotechnology, Gdynia, Poland). We assessed the sensitivity of detection of Tritrichomonas foetus in spiked fecal samples for the four kits combined with two molecular assays: PCR and LAMP. The extraction efficacy was quantified using defined aliquots of fecal samples spiked with 5 μL of suspensions containing serial dilutions of trophozoites (0.1; 1; 10; 100; 1000; 10,000), with six replicates for each concentration. In our study, we proved that the ZR Fecal DNA MiniPrep (50 preps) kit combined with LAMP and PCR had the highest efficiency among all the compared methods for the detection of feline T. foetus from fecal samples.

Bovine trichomoniasis, caused by the protozoal parasite Tritrichomonas foetus, is a highly contagious venereal disease characterised by early pregnancy loss, abortion and pyometra. Persistently infected bulls and cows are the primary reservoirs of infection in infected herds. This research investigated the prevalence of T. foetus infection in bulls from properties located across northern Australia and New South Wales. Preputial samples were collected from 606 bulls at slaughter and tested for T. foetus using the VetMAX-Gold Trich Detection Kit (Thermo Fisher Scientific). The apparent prevalence of T. foetus infection varied between regions, with northern regions in the Northern Territory, Queensland and Western Australia showing a prevalence of 15.4%, 13.8% and 11.4%, respectively. There was some evidence of an association between infection and postcode (P = 0.06) and increasing bull age (P = 0.054). This study confirms that T. foetus infection is likely to be present in many beef breeding herds and contributing to lower than expected reproductive performance, particularly across northern Australia., (© 2022 Thermo Fisher Scientific Australia Pty Ltd. Australian Veterinary Journal published by John Wiley & Sons Australia, Ltd on behalf of Australian Veterinary Association.)

A 2-year-old, 4.2 kg, spayed female, Maine coon cat was referred to the veterinary hospital for evaluation of hyporexia, slow growth, and chronic, intermittent, mucoid, bloody, voluminous, and fetid diarrhea. The diarrhea had been observed since the cat was acquired from a cattery at 4 months of age; with acute worsening in the 5 d before presentation. Abdominal palpation revealed moderate pain. Ultrasonographic examination showed thickening of the jejunal wall and ileal loops, increased echogenicity of the jejunal mucosa, and enlargement of the jejunal and ileocolic lymph nodes. Histopathology of full-thickness intestinal biopsies showed moderate, diffuse, lymphoplasmacytic, erosive enteritis with hemorrhage and edema. Diffuse, lymphoplasmacytic, erosive colitis with mild, interstitial fibrosis and hemorrhage was also noted. The ileocecal lymph node biopsy showed eosinophilic lymphadenitis. Based on the immunohistochemical evaluation of intestinal samples with CD3 and CD79a antibodies, a diagnosis of lymphoma was ruled out. Fecal polymerase chain reaction testing was positive for Tritrichomonas foetus . Based on these results, inflammatory bowel disease and trichomonosis were diagnosed. Treatment for the cat included a hypoallergenic diet and an oral omega-3 fatty acid supplement, in conjunction with prednisolone, to manage the inflammatory bowel disease. Ronidazole was administered to target the Tritrichomonas foetus . The cat was clinically normal during a follow-up examination after 6 months of treatment., (Copyright and/or publishing rights held by the Canadian Veterinary Medical Association.)

Protists members of the Trichomonadidae and Tritrichomonadidae families include agents of trichomoniasis that constitute important parasitic diseases in humans and in animals of veterinary interest. One of the characteristic features of these eukaryotic microorganisms is that they contain a fibrous structure known as the costa as an important cytoskeleton structure, that differs in several aspects from other cytoskeleton structures found in eukaryotic cells. Previous proteomic analysis of an enriched costa fraction revealed the presence of several hypothetical proteins. Here we describe the localization of one of the most prevalent protein found in this previously made proteomic assay to confirm its presence in the costa of Tritrichomonas foetus. A peptide sequence of the hypothetical protein ARM19800.1 was selected for the production of specific polyclonal antibodies and its specificity was confirmed by Western Blotting using an enriched costa fraction. Next, the specific localization of the selected protein was evaluated by immunofluorescence and electron microscopy immunocytochemistry. Our observations clearly showed that the ARM 19800.1 protein is indeed localized in the costa and displays an almost periodic labeling pattern. Since this is the first protein identified in the costa, it was designated as costain 1. A better understanding of a structure as peculiar as the costa is of great biological and evolutionary importance due to the fact that it contains unique proteins, it may represent a possible chemotherapy target and it may correspond to antigens of interest in immunodiagnosis and/or vaccine development., (Copyright © 2021. Published by Elsevier Inc.)

Tritrichomonas foetus causes chronic large-bowel diarrhoea in cats, complicated not only by its chronicity but for its difficulty to treat, with few registered or effective products available, subsequently resulting in ronidazole frequently used off-label. The extent of T. foetus resistance to ronidazole is unknown. To demonstrate in vitro resistance of feline T. foetus to ronidazole and the self-resolution of the infection, we isolated novel T. foetus "feline" genotype strains from two cats in 2017 from the same household and monitored these cats over a three-year period. Axenic cultures of T. foetus from the cats were tested for in vitro resistance to metronidazole and ronidazole using anaerobic and aerobic culture assays. The minimum lethal concentration for the novel strains after incubation in aerobic conditions for 48 h were 50-100 μg/ml and 6.25-12.50 μg/ml for metronidazole and ronidazole, respectively. For susceptible T. foetus strains, the minimum lethal concentrations ranged from 1.56-12.50 μg/ml for metronidazole, and 0.39-3.13 μg/ml for ronidazole. The self-resolution of the infection was assessed using repeat diagnostic qPCR and culture. Whilst positive in 2018, the cat samples are no longer returning positive qPCR results in 2020 and 2021, indicating self-resolution over this period. This study demonstrated resistance to ronidazole for the first time in Australian cats and infection self-resolution without antimicrobial intervention is demonstrated. This study provides clinicians with the evidence to reduce the use of off-label ronidazole, and advice on the resolution timeframe for cats in single households., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Trichomonas vaginalis and Tritrichomonas foetus are extracellular flagellated parasites that inhabit humans and other mammals, respectively. In addition to motility, flagella act in a variety of biological processes in different cell types, and extra-axonemal structures (EASs) have been described as fibrillar structures that provide mechanical support and act as metabolic, homeostatic, and sensory platforms in many organisms. It has been assumed that T. vaginalis and T. foetus do not have EASs. However, here, we used complementary electron microscopy techniques to reveal the ultrastructure of EASs in both parasites. Such EASs are thin filaments (3-5 nm diameter) running longitudinally along the axonemes and surrounded by the flagellar membrane, forming prominent flagellar swellings. We observed that the formation of EAS increases after parasite adhesion on the host cells, fibronectin, and precationized surfaces. A high number of rosettes, clusters of intramembrane particles that have been proposed as sensorial structures, and microvesicles protruding from the membrane were observed in the EASs. Our observations demonstrate that T. vaginalis and T. foetus can connect to themselves by EASs present in flagella. The protein VPS32, a member of the ESCRT-III complex crucial for diverse membrane remodeling events, the pinching off and release of microvesicles, was found in the surface as well as in microvesicles protruding from EASs. Moreover, we demonstrated that the formation of EAS also increases in parasites overexpressing VPS32 and that T. vaginalis -VPS32 parasites showed greater motility in semisolid agar. These results provide valuable data about the role of the flagellar EASs in the cell-to-cell communication and pathogenesis of these extracellular parasites., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Coceres, Iriarte, Miranda-Magalhães, Santos de Andrade, de Miguel and Pereira-Neves.)

Bovine trichomonosis is caused by Tritrichomonas foetus. Thirty-three US states have state rules on this disease and render it reportable due to potential huge economic losses to cattle industry. The various rules of different states generally mandate testing and culling T. foetus-positive bulls as well as prohibiting import of T. foetus-positive animals. Wyoming has enforced these rules for over 20 year beginning in 2000. From 2017 to 2019, 3 years in a row, not even one T. foetus-positive bull has been detected throughout the entire state among over ten thousand bulls tested annually. Wyoming is the first US state to achieve total control and eradication of bovine trichomonosis by testing and culling T. foetus-positive bulls., (© 2021. The Author(s).)

Background: We previously reported that the tomato glycoalkaloid tomatine inhibited the growth of Trichomonas vaginalis strain G3, Tritrichomonas foetus strain D1, and Tritrichomonas foetus-like strain C1 that cause disease in humans and farm and domesticated animals. The increasing prevalence of antibiotic resistance requires development of new tools to enhance or replace medicinal antibiotics., Methods: Wild tomato plants were harvested and divided into leaves, stems, and fruit of different colors: green, yellow, and red. Samples were freeze dried and ground with a handheld mill. The resulting powders were evaluated for their potential anti-microbial effects on protozoan parasites, bacteria, and fungi. A concentration of 0.02% (w/v) was used for the inhibition of protozoan parasites. A high concentration of 10% (w/v) solution was tested for bacteria and fungi as an initial screen to evaluate potential anti-microbial activity and results using this high concentration limits its clinical relevance., Results: Natural powders derived from various parts of tomato plants were all effective in inhibiting the growth of the three trichomonads to varying degrees. Test samples from leaves, stems, and immature 'green' tomato peels and fruit, all containing tomatine, were more effective as an inhibitor of the D1 strain than those prepared from yellow and red tomato peels which lack tomatine. Chlorogenic acid and quercetin glycosides were present in all parts of the plant and fruit, while caffeic acid was only found in the fruit peels. Any correlation between plant components and inhibition of the G3 and C1 strains was not apparent, although all the powders were variably effective. Tomato leaf was the most effective powder in all strains, and was also the highest in tomatine. S. enterica showed a minor susceptibility while B. cereus and C. albicans fungi both showed a significant growth inhibition with some of the test powders. The powders inhibited growth of the pathogens without affecting beneficial lactobacilli found in the normal flora of the vagina., Conclusions: The results suggest that powders prepared from tomato leaves, stems, and green tomato peels and to a lesser extent from peels from yellow and red tomatoes offer potential multiple health benefits against infections caused by pathogenic protozoa, bacteria, and fungi, without affecting beneficial lactobacilli that also reside in the normal flora of the vagina., (© 2021. The Author(s).)

Trichomonads are an order of parasitic protists which infect a wide range of hosts. The human parasite Trichomonas vaginalis and the bovine parasite Tritrichomonas foetus which also infects cats and swine are of considerable medical and veterinary importance, respectively. Since trichomonads are microaerophiles/anaerobes they are susceptible to 5-nitroimidazoles such as metronidazole. 5-nitroimidazoles are exclusively toxic to microaerophilic/anaerobic organisms because reduction, i.e. activation, of the drug can only occur in a highly reductive environment. 5-nitroimidazoles have remained a reliable treatment option throughout the last decades but drug resistance can be a problem. Clinical resistance to 5-nitroimidazoles has been studied in more detail in T. vaginalis and has been ascribed to defective oxygen scavenging mechanisms which lead to higher intracellular oxygen concentrations and, consequently, to less drug being reduced. Two enzymes, flavin reductase (FR) and NADH oxidase have been suggested to be the major oxygen scavenging enzymes in T. vaginalis. The loss, or at least an impairment of FR which reduces oxygen to hydrogen peroxide, has been proposed as the central mechanism that enables the emergence of 5-nitroimidazole resistance. In this study we explored if T. foetus also encodes a homolog of FR and if it is, likewise, involved in resistance. T. foetus was indeed found to express a FR but it was only weakly active as compared to the T. vaginalis homolog. Further, activity of FR in T. foetus was unchanged in metronidazole-resistant cell lines, ruling out that it has a role in metronidazole resistance. Finally, we measured oxygen scavenging rates in metronidazole-sensitive and -resistant cell lines and found that NADH oxidase and FR are not the major oxygen scavenging enzymes in trichomonads and that oxygen scavenging is possibly a consequence, rather than a cause of metronidazole resistance., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The prevalence of bovine Tritrichomonas foetus infection has decreased almost to zero in most European countries, such as Poland, which has been Tritrichomonas foetus -free since 1997. However, trichomonosis is a notifiable disease and there is a duty to examine samples from cattle. In this study, we present an unexpected cross-reaction with Honigbergiella -like DNA in a specimen from a bull. The bovine sample was submitted to the Department of Parasitology National Veterinary Research Institute in Pulawy (NVRI) for confirmatory testing after having been examined at the Regional Veterinary Laboratory, during a routine T. foetus diagnosis. Positive results from microscopic observation and cultures were confirmed. Noteworthily, parasites grew on Diamond's medium only after seven days of incubation, while optimal growth of trichomonads is generally observed after two to four days for this medium. Moreover, by using PCR we obtained positive results for the presence of T. foetus . However, sequencing of the amplification product revealed 99.62% identity with Honigbergiella sp. Our data suggest that false-positive results may occur in commonly used PCR tests. Thus, unexpected results should be carefully interpreted.

Trichomonas vaginalis is a neglected urogenital parasitic protist that causes 170 million cases of trichomoniasis annually, making it the most prevalent non-viral, sexually transmitted disease. Trichomoniasis treatment relies on nitroheterocyclics, such as metronidazole. However, with increasing drug-resistance, there is an urgent need for novel anti-trichomonals. Little progress has been made to translate anti-trichomonal research into commercialised therapeutics, and the absence of a standardised compound-screening platform is the immediate stumbling block for drug-discovery. Herein, we describe a simple, cost-effective growth assay for T. vaginalis and the related Tritrichomonas foetus. Tracking changes in pH were a valid indicator of trichomonad growth (T. vaginalis and T. foetus), allowing development of a miniaturised, chromogenic growth assay based on the phenol red indicator in 96- and 384-well microtiter plate formats. The outputs of this assay can be quantitatively and qualitatively assessed, with consistent dynamic ranges based on Z' values of 0.741 and 0.870 across medium- and high-throughput formats, respectively. We applied this high-throughput format within the largest pure-compound microbial metabolite screen (812 compounds) for T. vaginalis and identified 43 hit compounds. We compared these identified compounds to mammalian cell lines, and highlighted extensive overlaps between anti-trichomonal and anti-tumour activity. Lastly, observing nanomolar inhibition of T. vaginalis by fumagillin, and noting this compound has reported activity in other protists, we performed in silico analyses of the interaction of fumagillin with its molecular target methionine aminopeptidase 2 for T. vaginalis, Giardia lamblia and Entamoeba histolytica, highlighting potential for fumagillin as a broad-spectrum anti-protistal against microaerophilic protists. Together, this new platform will accelerate drug-discovery efforts, underpin drug-resistance screening in trichomonads, and contributing to a growing body of evidence highlighting the potential of microbial natural products as novel anti-protistals., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Tritrichomonas foetus (Trichomonadida, Tritrichomonadidae) is a parasitic protozoan which causes a diarrheic disease in domestic cats worldwide. The epizootiological and pathogenic impact of this parasite in many geographic locations is still to be unraveled; thus, the present study has investigated the occurrence of T. foetus in cats from central Italy, along with the analysis of risk factors and clinical signs. One hundred and five cats referred to the University Veterinary Teaching Hospital of the University of Teramo were tested with copromicroscopic techniques for common intestinal and extra-intestinal parasites and a specific PCR for T. foetus. A statistical analysis evaluating associations between T. foetus positivity and possible risk factors was performed. One animal scored positive for Giardia (0.95%), 2 for Toxocara cati (1.90%), 1 for Eucoleus aerophilus (syn. Capillaria aerophila) (0.95%), 9 for Aelurostrongylus abstrusus (8.57%) and 1 for Dipylididum caninum (0.95%), while T. foetus was molecularly recorded in 18/105 (17.14%) of the study cats. In two cats, T. foetus was found in association respectively with Giardia and A. abstrusus. Animals with chronic gastrointestinal signs were more significantly infected by T. foetus (40.90% vs 10.84%; p < 0.01; 95% CI = 1.90-17.04; OR = 5.69) in comparison to cats with no gastrointestinal distresses. No other statistically significant associations were found. This study confirms that T. foetus is enzootic in cats in Italy and that it should be included in the differential diagnosis of feline chronic gastrointestinal diseases., (Copyright © 2021 Elsevier B.V. All rights reserved.)

An 8-year-old, spayed female Rottweiler dog, under immunosuppressant treatment for protein-losing enteropathy (PLE) and intestinal lymphangiectasia, was presented for anorexia, poor general conditions and episodes of diarrhea. A subcutaneous mass between the caudal abdominal mammary glands was found. A fine-needle aspiration cytology was performed and revealed the presence of inflammatory cells mixed with pear- or round-shaped microorganisms with cytomorphological features of flagellated protozoan trophozoites, belonging to Trichomonadida order. The final diagnosis of Tritrichomonas foetus infection was achieved with molecular analysis (Real-Time PCR) and sequencing. T. foetus has often been reported as a pathogenic agent in cattle and cats, and rarely reported in dogs, where its pathogenetic role is poorly understood. To the author's knowledge, this is the first case of T. foetus infection described in a location different from the gastrointestinal tract in a dog., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Human trichomoniasis, caused by the pathogenic parasitic protozoan Trichomonas vaginalis, is the most common non-viral sexually transmitted disease that contributes to reproductive morbidity in affected women and possibly to prostate cancer in men. Tritrichomonas foetus strains cause the disease trichomoniasis in farm animals (cattle, bulls, pigs) and diarrhea in domestic animals (cats and dogs). Because some T. vaginalis strains have become resistant to the widely used drug metronidazole, there is a need to develop alternative treatments, based on safe natural products that have the potential to replace and/or enhance the activity of lower doses of metronidazole. To help meet this need, this overview collates and interprets worldwide reported studies on the efficacy of structurally different classes of food, marine, and medicinal plant extracts and some of their bioactive pure compounds against T. vaginalis and T. foetus in vitro and in infected mice and women. Active food extracts include potato peels and their glycoalkaloids α-chaconine and α-solanine, caffeic and chlorogenic acids, and quercetin; the tomato glycoalkaloid α-tomatine; theaflavin-rich black tea extracts and bioactive theaflavins; plant essential oils and their compounds (+)-α-bisabolol and eugenol; the grape skin compound resveratrol; the kidney bean lectin, marine extracts from algae, seaweeds, and fungi and compounds that are derived from fungi; medicinal extracts and about 30 isolated pure compounds. Also covered are the inactivation of drug-resistant T. vaginalis and T. foetus strains by sensitized light; anti-trichomonad effects in mice and women; beneficial effects of probiotics in women; and mechanisms that govern cell death. The summarized findings will hopefully stimulate additional research, including molecular-mechanism-guided inactivations and human clinical studies, that will help ameliorate adverse effects of pathogenic protozoa.

Plant-derived anthraquinones were evaluated in cell assays for their inhibitory activities against the parasitic protozoa Trichomonas vaginalis human strain G3 that causes the sexually transmitted disease trichomoniasis in women, Tritrichomonas foetus bovine strain D1 that causes sexually transmitted diseases in farm animals (bulls, cows, and pigs), Tritrichomonas foetus -like strain C1 that causes diarrhea in domestic animals (cats and dogs), and bacteria and fungi. The anthraquinones assessed for their inhibitory activity were anthraquinone, aloe-emodin (1,8-dihydroxy-3-hydroxymethylanthraquinone), anthrarufin (1,5-dihydroxyanthraquinone), chrysazin (1,8-dihydroxyanthraquinone), emodin (1,3,8-trihydroxy-6-methylanthraquinone), purpurin (1,2,4-trihydroxyanthraquinone), and rhein (1,8-dihydroxy-3-carboxyanthraquinone). Their activities were determined in terms of IC 50 values, defined as the concentration that inhibits 50% of the cells under the test conditions and calculated from linear dose response plots for the parasitic protozoa, and zone of inhibition for bacteria and fungi, respectively. The results show that the different substituents on the anthraquinone ring seem to influence the relative potency. Analysis of the structure-activity relationships in protozoa indicates that the aloe-emodin and chrysazin with the highest biological activities merit further study for their potential to help treat the diseases in women and domestic and farm animals. Emodin also exhibited antifungal activity against Candida albicans . The suggested mechanism of action and the additional reported beneficial biological properties of anthraquinones suggest that they have the potential to ameliorate a broad spectrum of human diseases.

Tritrichomonas foetus is a causative agent of feline trichomonosis, resulting in large-bowel diarrhea in cats. Feline trichomonosis has been reported in the USA, Europe and some Asian countries but there is limited information for Thailand. This study investigated the prevalence of T. foetus infection in cats in the Bangkok metropolitan area and evaluated the in vitro efficacy of metronidazole (MDZ) and ronidazole (RDZ) against T. foetus Thai isolates. In total, 215 fecal samples were collected from 121 owned cats and 94 stray cats. All fecal samples were cultivated in InPouch™ TF-feline medium. Afterward, polymerase chain reaction (PCR) assays targeting the 5.8S rRNA gene, the ITS regions and DNA sequencing were used for the confirmation of T. foetus. The overall prevalence of T. foetus infection was 4.18% (9/215) based on cultivation and PCR. The sequencing results showed 100% homology to T. foetus sequences from GenBank. The average minimal lethal concentrations (MLCs) of MDZ were 333.33 and 66.67 μg/ml at 24 and 48 h, respectively. The average MLCs of RDZ were 29.16 and 12.5 μg/ml at 24 and 48 h, respectively. The MLC of the MDZ results revealed that T. foetus Thai isolates had a tendency to be MDZ-resistant. To the best of the authors' knowledge, this study was the first using in vitro cultivation and molecular techniques to report and confirm the presence of T. foetus in cats living in the Bangkok metropolitan area. Further studies are needed to determine the genuine infection rate of T. foetus in a greater population sample and the infection status in cats with signs of diarrhea in Thailand., Competing Interests: Declaration of Competing Interest The authors have no competing interest to declare., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Tritrichomonas foetus is described as a pathogen of cattle and cats and also exhibits commensalism with pigs. In order to estimate the prevalence and determine the risk factors for parasite infection, specimens from animal hosts (cat, pigs, and cattle) from Poland were investigated. To our best knowledge, this is the first such study to examine samples from wild boars ( Sus scrofa ) for the presence of T. foetus. Data were collected from 117 cats, 172 pigs, 236 wild boars, and 180 cattle. The sensitivity of T. foetus identification was increased by using two molecular assays: PCR and loop-mediated isothermal amplification (LAMP). The prevalence of feline tritrichomonosis was 20.51%, and statistically significant differences were obtained between groups of animals regarding age, breed, number of cats, diarrhea, and place of living. Positive PCR and LAMP results for T. foetus were estimated for 16.28% of pigs, and the obtained data were significantly correlated with age. Conversely, no significant differences were observed concerning the farm size factor. In our survey, no cases of bovine tritrichomonosis were found, which is consistent with the data from the other countries of the European Union. Similarly, all wild boar samples were also T. foetus -negative according to LAMP and PCR.

Tritrichomonas foetus is a protozoan parasite that colonizes the reproductive tract of cattle as well as the gastrointestinal tract of cats. Bovine tritrichomonosis is a sexually transmitted disease whereas feline tritrichomonosis is thought to be transmitted by the fecal-oral route. Furthermore, T. foetus is known as an essentially apathogenic commensal located in the nasal cavity of pigs. Transmission of T. foetus between the different hosts has to be considered a realistic scenario that may have important implications for the epidemiology of infections and disease. In our study, we generated whole genome sequencing (WGS) data from bovine, feline and porcine T. foetus strains to investigate the genetic (dis)similarities among these diverse strains. As a reference, we used a previously released draft assembly from a bovine T. foetus strain K isolated from an infected bull in Brazil. In particular, we identified single nucleotide polymorphisms (SNPs) and the insertion-deletion (indel) variations within the genomes of the different strains. Interestingly, only a low degree of polymorphism (68 SNPs and indels) was found between the bovine and the porcine strains in terms of variants with a predicted impact of moderate or high and where one species is homozygous for one allele and the other homozygous for the other allele. Conversely, however, a 964 times higher number of such differences was detected by comparing the feline with either the bovine (65,569) or the porcine (65,615) strain. These data clearly indicated a close phylogenetic relationship between bovine and porcine T. foetus but a remarkable genetic distinctness of these two strains from the feline strain. The latter observation was confirmed by PCR-based sequencing of 20 in silico-selected indel markers and five in silico-selected SNP markers that uniformly demonstrated a relatively distant phylogenetic relationship of three independent feline T. foetus isolates in comparison to the bovine and porcine strains investigated. In summary, our comparative genome sequencing approach provided further insights into the genetic diversity of T. foetus in relation to the different host origins of the parasite. Furthermore, our study identified a large number of SNP- and indel-containing sequences that may be useful molecular markers for future epidemiological studies aimed at the elucidation of the transmission patterns of T. foetus within different host species., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Several Tritrichomonas species have been found in mammalian hosts. Among these trichomonads T. foetus is often found in the urogenital tract of cattle and the gastrointestinal tract of the domestic cat, resulting in sexually transmitted bovine trichomonosis and fecal-orally transmitted feline trichomonosis, respectively. The aims of the current study were to molecularly characterize clinical isolates of T. foetus in cattle populations in Wyoming, South Dakota, and Montana of the United States of America and to phylogenetically analyze Tritrichomonas species of mammalian hosts., Results: DNA sequencing of rRNA genes showed over 99% identity of the newly described isolates to other bovine isolates. Further, T. foetus isolates of various mammalian hosts originated in different geographic regions worldwide were clustered into two well-defined clades by phylogenetic analysis of rRNA and cysteine protease 2 genes. Clade I consisted of isolates originated from cattle, pig, and human whereas clade II contained isolates of cat and dog., Conclusion: It is concluded that all mammalian Tritrichomonas spp. apparently belong to T. foetus. Analysis of more sequences is warranted to support this conclusion.

Tritrichomonas foetus is a protozoan parasite that has been traditionally identified as a cause of reproductive tract disease in cattle and gastrointestinal tract infection in cats. Moreover, T. foetus is also well known as a commensal of the nasal cavity, intestines, and stomach in swine. In this review we describe T. foetus as a pathogen dangerous to more than one animal host, diagnostic and taxonomic aspects of this infection, and the extent to which isolates from different hosts share genetic identity., Competing Interests: Conflict of Interest Conflict of Interests Statement: The authors declare that there is no conflict of interests regarding the publication of this article., (© 2019 Dąbrowska et al. published by Sciendo.)

Tritrichomonas foetus isolates from feline and bovine origin has been previously shown to carry a certain degree of genetic heterogeneity. Here, novel candidate molecular markers were developed by means of multilocus sequence typing of the gap2 gene (encoding for T. foetus glyceraldehyde-3-phosphate dehydrogenase), ITS region, the TR7/TR8 variable-length repeat and microsatellite genotyping. These markers were used to characterize T. foetus field isolates from bulls and domestic cats and to compare phylogenetically with the following ATCC isolates: T. foetus isolated from cattle and pig (syn. Tritrichomonas suis), Tritrichomonas mobilensis, Tetratrichomonas gallinarum and Pentatrichomonas hominis. Among them, TFMS10 and TFMS7 were found to be the most polymorphic markers. Moreover, an 809 bp fragment of the gap2 gene was successfully amplified from all the trichomonads included in this study and the sequence analysis revealed differences between T. foetus porcine and feline genotypes and T. mobilensis in comparison to the bovine T. foetus ATCC isolate. The TR7/TR8 repeat pattern was not reproducible, being only consistent the fragments of approximately 110 and 217 bp. Sequence analysis of the latter revealed the existence of 3 SNPs resulting in 98.6 % homology between bovine and feline isolates. A search for similar sequences was carried out to develop a Restriction Length Fragment Polymorphism analysis. A 503 bp region, named TF1, revealed the existence of two BbvI restriction enzyme sites that were able to generate different length fragments for T. foetus feline and bovine isolates. Finally, the neighbour-joining analyses showed that T. foetus porcine genotype clusters together with bovine genotype, whereas T. mobilensis and the feline genotype form a separate cluster., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Tritrichomonas foetus is a common cause of large bowel diarrhea in cats. Probiotics have been suggested to be effective for many intestinal pathogens; however, there are a lack of studies evaluating the effect of probiotics in T. foetus infection. In vitro studies were performed to evaluate the effect of a probiotic containing Enterococcus faecium (Efm) SF68 and a novel probiotic, Enterococcus hirae, on the inhibition of T. foetus growth, adhesion to, and cytotoxicity towards the intestinal epithelium. The effect of enterococci on T. foetus proliferation during co-culture was evaluated throughout log phase T. foetus growth. The previously validated in vitro co-culture model system using porcine intestinal epithelial cells (IPEC-J2) was used to evaluate the effect of enterococci on T. foetus adhesion and cytotoxicity towards intestinal epithelial cells. Cytotoxicity was assessed using fluorescent microscopy and spectrophotometry. Interactions of T. foetus, enterococci, and intestinal epithelial cells were assessed using scanning electron microscopy and immunofluorescence assays (IFA). Enterococcus-induced inhibition of T. foetus growth was demonstrated at concentrations as low as 10 4 enterococci colony forming units (CFU)/mL and was dependent, in part, on environmental pH and the presence of viable enterococci organisms. T. foetus adhesion, including with a ronidazole-resistant strain, was reduced with pretreatment of intestinal epithelial cells with enterococci but was not significantly affected when enterococci were introduced simultaneously or following T. foetus infection. Compared to Efm, E. hirae more effectively decreased T. foetus adhesion, suggesting its superior potential as a novel probiotic for T. foetus infection. There was no effect of enterococci treatment on T. foetus-induced intestinal epithelial cell cytotoxicity. Our results support further study into the investigation of a possible benefit of enterococci-containing probiotic treatment for prevention of T. foetus infection in at-risk uninfected cats., (Copyright © 2019 Elsevier B.V. All rights reserved.)