Revisiting fecal metatranscriptomics analyses of macaques with idiopathic chronic diarrhoea with a focus on trichomonad parasites.

Trichomonads, anaerobic microbial eukaryotes members of the phylum Parabasalia, are common obligate extracellular symbionts that can lead to pathological or asymptomatic colonization of various mucosal surfaces in a wide range of animal hosts. Results from previous in vitro studies have suggested a number of intriguing mucosal colonization strategies by Trichomonads, notably highlighting the importance of interactions with bacteria. However, in vivo validation is currently lacking. A previous metatranscriptomics study into the cause of idiopathic chronic diarrhoea in macaques reported the presence of an unidentified protozoan parasite related to Trichomonas vaginalis. In this work, we performed a reanalysis of the published data in order to identify the parasite species present in the macaque gut. We also leveraged the information-rich metatranscriptomics data to investigate the parasite behaviour in vivo. Our results indicated the presence of at least 3 genera of Trichomonad parasite; Tetratrichomonas, Pentatrichomonas and Trichomitus, 2 of which had not been previously reported in the macaque gut. In addition, we identified common in vivo expression profiles shared amongst the Trichomonads. In agreement with previous findings for other Trichomonads, our results highlighted a relationship between Trichomonads and mucosal bacterial diversity which could be influential in health and disease.

Cytokine array analysis of mediators produced by human macrophages stimulated with Trichomonastenax.

The incidence of oral colonization by the protozoan Trichomonas tenax correlates with gingival inflammation and periodontitis in humans. To determine whether T. tenax might contribute to inflammation by eliciting cytokines from human cells, differentiated THP-1 (dTHP-1) macrophages were cultured with live or sonicated T. tenax trophozoites, and the conditioned media were assayed for 36 different mediators by a membrane-based cytokine array. Scanning densitometry of the membranes revealed that live T. tenax trophozoites stimulated secretion of interleukin-8 (IL-8), macrophage migration inhibitory factor (MIF), IL-1ß, intercellular adhesion molecule-1 (ICAM-1), and IL-1 receptor antagonist (IL-1ra) from dTHP-1 macrophages. T. tenax lysates stimulated release of IL-8, MIF, and IL-1ra. Despite often being classified as a commensal organism, T. tenax elicited a wider variety of cytokines than the human urogenital pathogen, T. vaginalis, which elicited only IL-8 and MIF production from dTHP-1 cells.

Prevalence of trichomonads in the cloaca of wild wetland birds in the Netherlands.

Severe granulomatosis in productive layer chickens due to Tetratrichomonas gallinarum strain 13/16632 infection occurred in 2013 and 2017 on farms situated in a wetland area in the Netherlands. We hypothesized that wetland birds could be the source of the infection. Therefore, a prevalence study on trichomonads was performed by analysing cloaca swabs of 526 birds belonging to 13 species of wetland birds. The number of birds sampled ranged from 1 to 275 per species. Birds were sampled at 15 locations in the Netherlands. DNA extracted from the cloaca swabs was subjected to nested PCR using trichomonad-specific primers targeting the internal transcribed spacer 1 (ITS1)-5.8S rRNA-ITS2 region followed by cloning and sequencing. In nine bird species, trichomonads were detected; the overall prevalence was 9% (47/526), while the prevalence in the five species for which a substantial number of birds were examined (at least 39 per species) ranged from 4% to 24%. Three trichomonad species were found: T. gallinarum, Trichomonas tenax and Simplicimonas sp. of which T. gallinarum dominated. The virulent T. gallinarum strain 13/16632 was not detected, but closely related strains were. Phylogenetic analysis revealed that all T. gallinarum isolates belonged to two clusters within lineage 15 of Tetratrichomonas lineages. All T. tenax isolates were identical and clustered with reference strain H95, while Simplicimonas sp. isolates showed large genetic diversity. Some isolates may represent a new species of the genus Simplicimonas. We conclude that trichomonads are widespread amongst wetland birds, raising the question, amongst others, of their relevance for commercial poultry.RESEARCH HIGHLIGHTSTrichomonads occur among wild wetland birds in the Netherlands.T. gallinarum is the dominant trichomonad species in the cloaca of wetland birds.Some T. gallinarum isolates are closely related to a strain causing granulomas in layer chickens.Some isolates may represent a new species of the genus Simplicimonas.

Sympatric western lowland gorillas, central chimpanzees and humans are infected with different trichomonads.

We investigated intestinal trichomonads in western lowland gorillas, central chimpanzees and humans cohabiting the forest ecosystem of Dzanga-Sangha Protected Area in Central African Republic, using the internal transcribed spacer (ITS) region and SSU rRNA gene sequences. Trichomonads belonging to the genus Tetratrichomonas were detected in 23% of the faecal samples and in all host species. Different hosts were infected with different genotypes of Tetratrichomonas. In chimpanzees, we detected tetratrichomonads from 'novel lineage 2', which was previously reported mostly in captive and wild chimpanzees. In gorillas, we found two different genotypes of Tetratrichomonas. The ITS region sequences of the more frequent genotype were identical to the sequence found in a faecal sample of a wild western lowland gorilla from Cameroon. Sequences of the second genotype from gorillas were almost identical to sequences previously obtained from an anorexic French woman. We provide the first report of the presence of intestinal tetratrichomonads in asymptomatic, apparently healthy humans. Human tetratrichomonads belonged to the lineage 7, which was previously reported in domestic and wild pigs and a domestic horse. Our findings suggest that the ecology and spatial overlap among hominids in the tropical forest ecosystem has not resulted in exchange of intestinal trichomonads among these hosts.

Presence of trichomonads in subgingival biofilm of patients with periodontitis: preliminary results.

This study aims to evaluate the prevalence of trichomonads in the subgingival biofilm of patients with periodontitis. Secondarily, the trichomonad presence was related to patient characteristics and periodontal clinical parameters, in order to highlight the factor favoring the development of these protozoans. Subgingival biofilm samples were collected from at least two diseased and one healthy site in 50 patients suffering from periodontitis. Trichomonads were identified using phase contrast microscopy. All patient characteristics and periodontal clinical parameter data were then statistically analyzed. From the 50 patients examined, 195 sites were sampled, including 145 diseased ones. Trichomonads were only observed on 16 of the 145 diseased sites (11%) and none in the other 50 healthy sites. Based on these results, 20% (n = 10) of patients were positive for the presence of trichomonads from at least one of the diseased sites collected. Tooth mobility, substantial supra-gingival dental deposits, and severe clinical attachment loss were statistically associated with trichomonad presence. If the subgingival biofilm of male patients over the age of 50 seemed to be more frequently contaminated with trichomonads, this data was not statistically supported. This preliminary study indicates for the first time that in periodontitis-involved patients, trichomonads are observed in the subgingival biofilm collected from diseased sites with severe bone loss, but not from healthy teeth. Further investigations are needed to fully explore the role of this microorganism in the etiology of periodontal disease.

Prevalence of intestinal parasites in companion dogs with diarrhea in Beijing, China, and genetic characteristics of Giardia and Cryptosporidium species.

Companion animals including dogs are one of the important components in One Health. Parasites may cause not only diseases in pet animals but also many zoonotic diseases infecting humans. In this study, we performed a survey of intestinal parasites in fecal specimens (n = 485) collected from outpatient pet dogs with diarrhea in Beijing, China, for the entire year of 2015 by microscopic examination (all parasites) and SSU rRNA-based nested PCR detection (Giardia and Cryptosporidium). We observed a total of 124 (25.6%) parasite-positive specimens that contained one or more parasites, including Giardia duodenalis (12.8%), Cryptosporidium spp. (4.9%), Cystoisospora spp. (4.3%), trichomonads (4.3%), Toxocara canis (3.5%), Trichuris vulpis (0.6%), and Dipylidium caninum (0.2%). Among the 55 dog breeds, infection rates were significantly higher in border collies and bulldogs, but lower in poodles (p < 0.05). Risk factor analysis suggested that age was negatively correlated with the infection rate (p < 0.00001), while vaccination and deworming in the past 12 months could significantly reduce the parasite infections (p < 0.01). Among the 62 Giardia-positive specimens, 21 were successfully assigned into assemblages using glutamate dehydrogenase (gdh) and/or beta-giardin (bg) genes, including assemblage D (n = 15), C (n = 5), and F (n = 1). Among the 24 Cryptosporidium-positive specimens by SSU rRNA PCR, 20 PCR amplicons could be sequenced and identified as Cryptosporidium canis (n = 20). Collectively, this study indicates that parasites are a significant group of pathogens in companion dogs in Beijing, and companion dogs may potentially transmit certain zoonotic parasites to humans, particularly those with weak or weakened immunity.

Cytokine response of human THP-1 macrophages to Trichomonas tenax.

Trichomonas tenax is a protozoan that inhabits the oral cavity of humans, most often those with poor oral hygiene. Although T. tenax is widely considered a commensal, recent studies have suggested a pathogenic role for the protozoan in persons with periodontitis. Here we investigated the capacity of T. tenax to induce pro-inflammatory cytokine secretion in human macrophages, with the idea that elicitation of inflammation may be one mechanism by which T. tenax contributes to oral pathology. Human THP-1 cells differentiated to the macrophage phenotype (dTHP-1) were incubated with live or sonicated T. tenax at trophozoite:dTHP-1 ratios of 1:5, 1:10, and 1:20. Culture media removed from the wells after 4, 8, and 16 h of stimulation were assayed by ELISA for tumor necrosis factor alpha, interleukin-1 beta, interleukin-8, and the immunoregulatory cytokine interleukin-10. Live T. tenax trophozoites failed to induce production of any of the cytokines tested, regardless of trophozoite:dTHP-1 cell ratio or length of co-incubation. T. tenax lysates stimulated interleukin-8 synthesis, but only after 16 h of incubation at the 1:5 trophozoite:dTHP-1 cell ratio. These results suggest that pro-inflammatory cytokine synthesis by human macrophages in direct response to T. tenax contributes little to oral pathology.

Observation of trichomonads infection in a child with periodontitis by direct microscopy at the dental office.

The pathogenicity of Trichomonas species is well documented. Although their exact involvement in gum disease is not fully understood, recent studies suggest a correlation between these protozoa and periodontitis. This case report details the first chair-side observation in Europe of an oral trichomonad infection in a child with periodontitis, by direct microscopy. The dramatic recovery of the patient, observed following administration of an anti-parasitic treatment, confirms the necessity of further investigation in this field.

Host origin determines pH tolerance of Tritrichomonas foetus isolates from the feline gastrointestinal and bovine urogenital tracts.

The ability for protozoan parasites to tolerate pH fluctuations within their niche is critical for the establishment of infection and require the parasite to be capable of adapting to a distinct pH range. We used two host adapted Tritrichomonas foetus isolates, capable of infecting either the digestive tract (pH 5.3-6.6) of feline hosts or the reproductive tract (pH 7.4-7.8) of bovine hosts to address their adaptability to changing pH. Using flow cytometry, we investigated the pH tolerance of the bovine and feline T. foetus isolates over a range of physiologically relevant pH in vitro. Following exposure to mild acid stress (pH 6), the bovine T. foetus isolates showed a significant decrease in cell viability and increased cytoplasmic granularity (p-value < 0.003, p-value < 0.0002) compared to pH 7 and 8 (p-value > 0.7). In contrast, the feline genotype displayed an enhanced capacity to maintain cell morphology and viability (p-value > 0.05). Microscopic assessment revealed that following exposure to a weak acidic stress (pH 6), the bovine T. foetus transformed into rounded parasites with extended cell volumes and displays a decrease in viability. The higher tolerance for acidic extracellular environment of the feline isolate compared to the bovine isolate suggests that pH could be a critical factor in regulating T. foetus infections and host-specificity.

Persistence of Two Isolates of Trichomonas gallinae in Simulated Bird Baths With and Without Organic Material.

Trichomonas gallinae, a well-documented protozoan parasite of avian hosts, has been implicated in major passerine mortality events recently and historically throughout the literature. It has been suggested that bird baths and artificial water sources could serve as a source of infection for naive birds; however, trichomonad persistence in water is not well understood. We measured the persistence of T. gallinae isolates from two avian hosts in distilled water and distilled water with the addition of organic material. We inoculated plastic containers in a laboratory setting with 1 × 10(6) trichomonads and then sampled 500 µl from each container at various time points postinoculation (0-20 hr). The 500-µl aliquots were inoculated into flasks with 5 ml of modified Diamond media at each time point. Flasks were incubated at 37 C and examined by light microscopy for five consecutive days for the characteristic movements of live trichomonads. The maximum persistence was 16 hr with a Cooper's hawk (Accipiter cooperii) isolate in the organic material treatment, far longer than the 1 hr persistence previously reported. We show that T. gallinae isolates are capable of persisting for long periods of time in water, illustrating that bird baths may be validated as a potential source of transmission in epidemics.

Prevalence of intestinal trichomonads in captive non-human primates in China.

Trichomonads are protozoan symbionts with the capacity to infect vertebrates including humans and non-human primates (NHPs), sometimes with pathogenic effects. However, their diversity and prevalence in NHPs in China are poorly understood. A total of 533 fecal samples were collected from captive NHPs in Yunnan Province, China, of which 461 samples from Macaca fascicularis and 72 from Macaca mulatta. Trichomonadidae species were identified using PCR amplification of the ITS-1/5.8S/ITS-2 sequences. The overall prevalence of trichomonads in NHPs was determined to be 11.4% (61/533), with gender, diarrhea, and region identified as potential risk factors for the infections. Sequence alignment and phylogenetic analysis identified three species of trichomonads, i.e., Trichomitopsis minor (n = 45), Pentatrichomonas hominis (n = 11), and Tetratrichomonas sp. (n = 5). To the best of our knowledge, this is the first study to report Trichomitopsis minor infection in NHPs in China. Of note, Pentatrichomonas hominis is generally recognized as a parasitic organism affecting humans. Collectively, our results suggest that NHPs are potential sources of zoonotic trichomonad infections, highlighting the importance of surveillance and control measures to protect human and animal populations.

Title: Prévalence des Trichomonadidae intestinaux chez les primates non humains captifs en Chine. Abstract: Les Trichomonadidae sont des symbiotes protozoaires capables d'infecter les vertébrés, notamment les humains et les primates non humains (PNH), parfois avec des effets pathogènes. Cependant, leur diversité et leur prévalence chez les PNH en Chine sont mal comprises. Au total, 533 échantillons fécaux ont été collectés sur des PNH captifs dans la province du Yunnan, en Chine, dont 461 échantillons de Macaca fascicularis et 72 de Macaca mulatta. Les espèces de Trichomonadidae ont été identifiées par amplification PCR des séquences ITS-1/5.8S/ITS-2. La prévalence globale des Trichomonadidae dans les PNH a été déterminée à 11,4 % (61 / 533) et le sexe, la diarrhée et la région ont été identifiés comme facteurs de risque potentiels d'infection. L'alignement des séquences et l'analyse phylogénétique ont identifié trois espèces de Trichomonadidae, à savoir Trichomitopsis minor (n = 45), Pentatrichomonas hominis (n = 11) et Tetratrichomonas sp. (n = 5). À notre connaissance, il s'agit de la première étude à signaler une infection par Trichomitopsis minor chez les PNH en Chine. Il convient de noter que Pentatrichomonas hominis est généralement reconnu comme un organisme parasitaire affectant les humains. Collectivement, nos résultats suggèrent que les PNH sont des sources potentielles d'infections zoonotiques à Trichomonadidae, soulignant l'importance des mesures de surveillance et de contrôle pour protéger les populations humaines et animales.

Molecular Identification and Survey of Trichomonad Species in Pigs in Shanxi Province, North China.

Several trichomonad species have already been identified in pigs, and their pathogenic potential may not be ruled out. To date, however, no information is available regarding the prevalence of trichomonads in pigs in Shanxi Province, North China. In the present study, a total of 362 fecal samples collected from pigs in three representative counties (Qi, Jishan, and Shanyin) in this province were examined for Tetratrichomonas buttreyi, Tritrichomonas foetus, and Pentatrichomonas hominis using a nested polymerase chain reaction (PCR) with primers targeting the small subunit ribosomal RNA (SSU rRNA) gene. The overall prevalence of T. buttreyi was 49.72%, and region and age were found to be significantly associated with T. buttreyi infection, respectively. Only one pig fecal sample from Qi County was found to be positive for T. foetus, and all samples were negative for P. hominis. Molecular evolutionary analysis revealed that some T. buttreyi isolates showed complete genetic identity with those reported previously, and some T. buttreyi isolates and one T. foetus isolate showed minor allelic variations compared with those reported previously. This is the report of the molecular epidemiology of T. foetus and T. buttreyi in pigs in Shanxi Province, North China. These findings not only enrich the knowledge on the distribution of these trichomonad species in pigs in China but also provide baseline information for planning future research and control strategies.

Detection of trichomonads in patients with lung cancer and transcription analysis on the response of human pulmonary epithelial cells to Trichomonas tenax invasion.

INTRODUCTION: Lung cancer is one of the most prevalent malignancies worldwide. Substantial research has illuminated the intricate interplay between microorganisms and human health, revealing their role in disease regulation. Trichomonads is a flagellated protozoan in the human cavity and have been previously identified as a pathogen associated with pneumonia, contributing to tissue chronic inflammation and carcinogenesis. METHODS: Nested polymerase chain reaction methods were employed to scrutinize the prevalence of trichomonads in the bronchovesicular fluid of patients diagnosed with lung cancer. Subsequently, the influence of Trichomonas tenax invasion on lung cancer cells was elucidated through proliferation assays, migration assays, and transcription analysis. RESULTS: Bronchoalveolar fluid samples from lung cancer patients yielded positive nested PCR results for eight out of twenty-seven samples. Seven of these samples were identified as Trichomonas tenax, while one was identified as Tetratrichomonas spp. Our findings revealed a significant upregulation of pathways associated with carcinogenesis, including cellular proliferation, migration, and drug resistance, in response to T. tenax invasion. CONCLUSIONS: This study underscores the importance of recognizing the presence of trichomonads and the influence of T. tenax invasion on host responses to respiratory diseases. The identified pathways implicated in cancer development may pave the way for developing targeted treatment strategies for pulmonary diseases. These findings hold promise for informing and improving the precision of therapeutic interventions in the context of pulmonary ailments.

The parabasal filaments of Trichomonas vaginalis: A new filament and observations using 0.8 nm-resolution scanning electron microscopy.

Trichomonas vaginalis is the etiologic agent of trichomoniasis, the most common nonviral sexually transmitted infection worldwide, with an estimated 260 million new cases annually. T. vaginalis contains organelles common to all eukaryotic cells, uncommon cell structures such as hydrogenosomes, and a complex and elaborate cytoskeleton constituting the mastigont system. The mastigont system is mainly formed by several proteinaceous structures associated with basal bodies, the pelta-axostylar complex made of microtubules, and striated filaments named the costa and the parabasal filaments (PFs). Although the structural organization of trichomonad cytoskeletons has been analyzed using several techniques, observation using a new generation of scanning electron microscopes with a resolution exceeding 1 nm has allowed more detailed visualization of the three-dimensional organization of the mastigont system. In this study, we have investigated the cytoskeleton of T. vaginalis using a diverse range of scanning probe microscopy techniques, which were complemented by electron tomography and Fast-Fourier methods. This multi-modal approach has allowed us to characterize an unknown parabasal filament and reveal the ultrastructure of other striated fibers that have not been published before. Here, we show the differences in origin, striation pattern, size, localization, and additional details of the PFs, thus improving the knowledge of the cell biology of this parasite.

Clinical implications of trichomonads detected in bronchoalveolar fluid by metagenomic next-generation sequencing: a multicenter retrospective study.

Background: Pulmonary trichomoniasis is considered a neglected disease due to failures in recognizing it, stemming from insensitive microbial methods and a lack of specific clinical features. This study aims to analyze the clinical implications of trichomonads detected in bronchoalveolar lavage fluid (BALF) by metagenomic next-generation sequencing (mNGS). Methods: This multicenter retrospective study included patients diagnosed with pneumonia, admitted to three tertiary hospitals in China from July 2018 to September 2022, with trichomonads detected in BALF through mNGS. The analysis covered demographics, comorbidities, symptoms, laboratory findings, mNGS results, clinical treatment, and outcomes of these patients. Results: A total of 17 patients were enrolled, comprising 14 males and 3 females. Trichomonas tenax and Trichomonas vaginalis were detected by mNGS in BALF samples of 15 and 2 patients, respectively. Patients were categorized into two groups based on the presence of risk factors for trichomonad infection, including immunocompromised conditions, uncontrolled diabetes mellitus, oral/periodontal diseases, and aspiration. Among 11 patients with risk factors (Case 1-11), 4 received nitromidazoles as part of comprehensive treatment, achieving a 100% treatment success rate. The remaining 7 patients, who did not receive nitromidazoles, had only one achieving relief after broad-spectrum antimicrobial therapy, resulting in a 14.3% treatment success rate. For the 6 patients without any risk factors for trichomonad infection (Case 12-17), none received nitromidazoles during hospitalization. However, 4 out of these 6 patients (66.7%) eventually recovered. Conclusion: mNGS proves to be an efficient tool for detecting trichomonads in BALF samples. Comprehensive analysis of clinical features and laboratory indicators is essential to distinguish between infection and colonization of trichomonads. Pulmonary trichomoniasis should not be overlooked when trichomonads are detected in BALF from patients with risk factors.

Induction of apoptosis by essential oil of Dracocephalum kotschyi on Trichomonas vaginalis.

BACKGROUND: Trichomonas vaginalis is a protist parasite that causes trichomoniasis, a sexually transmitted disease. Metronidazole is the current treatment for trichomoniasis. However, this drug can provoke severe side effects, and some strains present resistance, making the development of alternative treatments for trichomoniasis urgent. OBJECTIVES: We investigate the use of essential oil obtained from Dracocephalum kotschyi on T. vaginalis. D. kotschyi has antispasmodic and analgesic properties and is well known in Iran. METHODS: The essential oil was obtained by hydrodistillation from 1000 g of the powdered plant. Gas chromatography-mass spectrometry analysis was used for the chemical composition of the essential oil, and 11 substances were identified, corresponding to 91.5% of the oil. Copaene (22.15%), Methyl geranate (16.31%), Geranial (13.78%) and Carvone (11.34%) were the main substances. A cell viability test was used to determine the percentage of growth inhibition (GI%) and the half-maximal inhibitory concentration (IC50) on T. vaginalis after incubation with the prepared essential oil. RESULTS: The oil induced an IC50 of 84.07 µg/ml after 24 h contact with trophozoites. Cytotoxicity was determined by MTT assay on the J774.A1 haematopoietic cell line. In addition, the initial stage of apoptosis was assayed using the fluorescein isothiocyanate Annexin V Apoptosis Detection Kit. Evaluation of the in vitro anti-trichomonal properties of D. kotschyi essential oils showed that it effectively induces apoptosis on T. vaginalis between 100 and 700 µg/ml after 48 h without toxicity on haematopoietic cells, suggesting that D. kotschyi essential oil can induce programmed death in T. vaginalis. CONCLUSIONS: The anti-trichomonal properties of D. kotschyi essential oil indicate that they could be suitable for new pharmacologic studies after new tests with human vaginal epithelial cells.

Drug susceptibility testing for oxygen-dependent and oxygen-independent resistance phenotypes in trichomonads.

Trichomonas vaginalis is the most prevalent, non-viral sexually transmitted human infection, causing 170 million cases of trichomoniasis annually. Since the 1950s, treatment has relied on 5-nitroimidazoles (5NIs), leading to increasing drug resistance. A similar drug resistance problem is present in the veterinary pathogen, Tritrichomonas foetus. There are currently no agreed standards for defining 5NI resistance, due in part to two distinct oxygen-dependent ("aerobic") and oxygen-independent ("anaerobic") resistance phenotypes. Diagnostic tools to detect 5NI resistance are lacking, and current assays used to phenotypically assess 5NI resistance in vitro are complicated by these two resistance phenotypes. We demonstrate that microaerophilic conditions support sufficient parasite growth to interrogate oxygen-dependent resistance of 5NIs against known resistant and susceptible isolates of T. vaginalis and T. foetus. We further demonstrate that microaerophilic conditions allow sufficient growth for compatibility with existing growth assays, including our TriTOX assay. Adopting microaerophilic conditions eliminates traditional 'by-eye' estimates of minimum inhibitory concentrations and opens up options for increased throughput and automation, scalable to higher-throughput analyses of 5NI resistance. This would further allow the development of quantitative phenotypic standards to benchmark oxygen-dependent or oxygen-independent trichomonad 5NI resistance towards standardised surveillance programs to combat drug resistance.

High-throughput sequencing of faeces provides evidence for dispersal of parasites and pathogens by migratory waterbirds.

Examination of faecal material has demonstrated how a broad range of organisms are distributed by bird movements. Such research has largely focused on dispersal of plant seeds by frugivores and of freshwater organisms by waterbirds. However, with few exceptions (e.g. avian influenza, Ebola virus), there is a dearth of evidence for transport of parasites and pathogens. High-throughput sequencing methods now provide a powerful means of addressing this knowledge gap by elucidating faecal contents in unprecedented detail. We collected faeces excreted by a range of migratory waterbirds in south-west Spain and pooled faecal DNA to create libraries reflective of feeding behavior. We created sets of libraries using high-throughput metagenomic and amplicon sequencing. For the latter we employed two sets of primers to broadly target the V4 region of the 18S rRNA gene (one set amplifying the region across all eukaryotes, the other excluding amplification of metazoans). Libraries revealed a wide diversity of eukaryotes, including parasites of the faecal producers themselves, parasites of food items, or those incidentally ingested. We also detected novel microbial eukaryotic taxa and found that parasite assemblage profiles were relatively distinct. Comparing the performance of the methods used supports their joint use for future studies of diversity and abundance. Because viable stages of many parasites are likely to be present in faeces, our results suggest significant levels of bird-mediated dispersal of parasites (both from avian and other hosts). Our methods revealed much hidden biodiversity, and allowed identification of the individuals who produced the faecal samples to species level, facilitating the study of interaction networks.

Isolation and Characterization of Novel Trichomonas gallinae Ribotypes Infecting Domestic and Wild Birds in Riyadh, Saudi Arabia.

Trichomonas gallinae, a single-celled protozoan parasite, is a causative agent of the disease trichomonosis, which is distributed worldwide and has recently been highlighted as a pandemic threat to several wild bird species. The aim of this study was to determine the prevalence and genotypic diversity of T. gallinae in Riyadh, Saudi Arabia. For this purpose, 273 oral swab samples from different bird species (feral pigeon Columba livia, common mynah Acridotheres tristis, chicken Gallus gallus domesticus, turkey Meleagris gallopavo, and ducks Anatidae) were collected and tested for T. gallinae infection with InPouch™ TV culture kits. The results showed that the overall prevalence of T. gallinae in these samples was 26.4% (n = 72). The PCRs were used to detect the internal transcribed spacer (ITS) region of T. gallinae, and the results of the sequence analysis indicated genetic variation. Among 48 sequences, we found 15 different ribotypes, of which 12 were novel. Three had been previously described as ribotypes A, C, and II. To our knowledge, this study demonstrated the presence of T. gallinae strain diversity in Saudi Arabian birds for the first time and revealed that ribotypes A and C are predominant among Riyadh birds.

Aislamiento y caracterización de nuevos ribotipos de Trichomonas gallinae que infectan aves domésticas y silvestres en Riad, Arabia Saudita. Trichomonas gallinae, un parásito protozoario unicelular, es un agente causal de la enfermedad tricomonasis, que se distribuye en todo el mundo y recientemente se ha destacado como una amenaza pandémica para varias especies de aves silvestres. El objetivo de este estudio fue determinar la prevalencia y la diversidad genotípica de T. gallinae en Riad, Arabia Saudita. Para este propósito, se recolectaron y analizaron 273 muestras de hisopos orales de diferentes especies de aves (paloma silvestre Columba livia, miná común Acridotheres tristis, pollos Gallus gallus domesticus, pavos Meleagris gallopavo y patos Anatidae) para detectar la infección por T. gallinae mediante cultivo con el estuche InPouch ™ TV. Los resultados mostraron que la prevalencia general de T. gallinae en estas muestras fue del 26.4% (n = 72). Se utilizó la técnica de PCR para detectar la región espaciadora transcrita interna (ITS) de T. gallinae, y los resultados del análisis de secuencias indicaron variación genética. Entre 48 secuencias, se encontraron 15 ribotipos diferentes, de los cuales 12 eran nuevos. Tres habían sido descritos previamente como ribotipos A, C y II. Hasta donde se conoce, este estudio demostró la presencia de diversidad entre cepas de T. gallinae en aves de Arabia Saudita por primera vez y reveló que los ribotipos A y C son los predominantes entre las aves de Riad.

Potential role of autophagy in proteolysis in Trichomonas vaginalis.

BACKGROUND: Autophagy has been shown to be involved in the pathogenesis of several protists, offering prospects for the developments of new drugs targeting autophagy. However, there is no evidence illustrating functional autophagy in the deep-branching trichomonads. The human parasitic protist Trichomonas vaginalis has been predicted to possess reduced autophagic machinery, with only autophagy-related protein 8 (Atg8) conjugation system required for autophagosome formation. METHODS: The recombinant protein of TvAtg8 (rTvAtg8) and the polyclonal antibody against rTvAtg8 were generated. The expression and localization of TvAtg8 was monitored upon autophagy induction by glucose restriction (GR) compared with glucose-rich cultivation. The role of TvAtg8 in proteolysis was clarified. RESULTS: Here, we report that T. vaginalis Atg8 (TvAtg8) is upregulated and conjugated to autophagosome-like vesicles upon autophagy induction by GR. Moreover, we investigate, for the first time, the role of autophagy in T. vaginalis. Proteasome inhibition (PI)-induced autophagy compensates for the removal of polyubiquitinated proteins under glucose-rich condition. GR-induced autophagy is a major proteolytic system in T. vaginalis. These results suggest that autophagy is vital for proteolysis in T. vaginalis with an impaired ubiquitin-proteasome system or under glucose-limited environment. CONCLUSION: Our findings unveiled previously unidentified functions of autophagy in proteostasis in trichomonads, advancing our understanding of this highly conserved process in the ancient eukaryote.