Differential Drug Susceptibility across Trichomonasvirus Species Allows for Generation of Varied Isogenic Clones of Trichomonas vaginalis.

Trichomonas vaginalis (Tvag) is a sexually transmitted human pathogen that is commonly infected with strains of one or more of five known species of Trichomonas vaginalis viruses (TVVs), members of genus Trichomonasvirus. TVVs are thought not to have an extracellular phase to their lifecycle and instead to be transmitted vertically from mother to daughter cells. As a result, generation of isogenic virus-positive and virus-negative sets of Tvag clones has been a major barrier to studying interactions between TVVs and their host. Nucleoside analog 2'-C-methylcytidine (2CMC) has been recently reported to clear trichomonads of infections with TVV1, TVV2, and TVV3. We used 2CMC to treat a panel of Tvag isolates that collectively harbor at least one representative strain of each TVV species and thereby provided evidence that infections with TVV4 and TVV5 can also be cleared by 2CMC. Furthermore, our results suggest a newly identified difference in drug susceptibility between TVV species. We took advantage of these susceptibility difference to generate isogenic sets of Tvag clones harboring different combinations of the five TVV species. These results provide both new insight into differences between these species and new avenues for generating tools to study the potential roles of TVVs in Tvag biology.

Trichomonas vaginalis: comparison of primers for implementation as an in-house PCR in rural Vellore, South India.

BACKGROUND: Trichomonas vaginalis (TV) accounts for the highest burden of curable, non-viral sexually transmitted infections worldwide. Prevalence in India ranges from 0.4 to 27.4% in women and 0.0-5.6% in men. In 2015, the prevalence of TV among pregnant women of rural Vellore was 3.11% using Sekisui OSOM® Trichomonas test and culture methods. Molecular methods are the most sensitive, rapid diagnostic tool for Sexually Transmitted Infection's (STI) albeit cost hinders implementation of commercial platforms. To determine a sensitive, sustainable molecular method, we compared three targets (Adhesin AP65, cytoskeleton Beta-tubulin BTUB 9/2 and TVK 3/7) with the highest published diagnostic accuracy against microscopy, culture and Real Time PCR (RT- PCR). MATERIALS & METHODS: Six-hundred adult, sexually active women attending the Obstetrics-Gynaecology rural out-patient clinic the Rural Unit for Health and Social Affairs (RUHSA) from July 2020 - February 2021 were enrolled. A vaginal lateral and posterior fornix specimen was inoculated, onsite, into Biomed InPouch® TV culture and smeared onto a slide for fluorescence microscopy using Acridine orange. A flocked nylon swab specimen for PCR was used to determine the sensitivities of the Adhesin AP65, cytoskeleton Beta-tubulin BTUB 9/2 and TVK 3/7 gene targets. Seegene Allplex™ STI Essential Assay, S.Korea was used to confirm TV positives. RESULTS: Nine specimens (9/600, 1.5%) were positive for TV. There was a 100% correlation between Biomed InPouch TV® culture, PCR with TVK 3/7 and RT-PCR while a correlation of 66.6% with BTUB 9/2 and AP65 gene targets. Clinically, 77.7% (n = 7) presented with white-greenish discharge per vagina, 11% (n = 1) with infertility, 22.2% (n = 2) were asymptomatic. Eight of nine patients (88.9%) had co-infections with other bacterial STIs. Prevalence of TV coinfection with Neisseria gonorrhoea was 1.1%. CONCLUSION: Current hospital-based prevalence of TV in rural Vellore was 1.5%. Repetitive DNA target TVK 3/7 was more sensitive than AP65 and BTUB 9/2 primers.

Structures of Native Doublet Microtubules from Trichomonas vaginalis Reveal Parasite-Specific Proteins as Potential Drug Targets.

Doublet microtubules (DMTs) are flagellar components required for the protist Trichomonas vaginalis (Tv) to swim through the human genitourinary tract to cause trichomoniasis, the most common non-viral sexually transmitted disease. Lack of DMT structures has prevented structure-guided drug design to manage Tv infection. Here, we determined the cryo-EM structure of native Tv-DMTs, identifying 29 unique proteins, including 18 microtubule inner proteins and 9 microtubule outer proteins. While the A-tubule is simplistic compared to DMTs of other organisms, the B-tubule features specialized, parasite-specific proteins, such as TvFAP40 and TvFAP35 that form filaments near the inner and outer junctions, respectively, to stabilize DMTs and enable Tv locomotion. Notably, a small molecule, assigned as IP6, is coordinated within a pocket of TvFAP40 and has characteristics of a drug molecule. This first atomic model of the Tv-DMT highlights the diversity of eukaryotic motility machinery and provides a structural framework to inform rational design of therapeutics.

An uncommon cause of persistent infection by Trichomonas vaginalis.

Trichomonas vaginalis infection typically exhibits a favorable response to treatment. Nonetheless, there are instances where complete eradication proves challenging, necessitating multiple treatment cycles. Understanding patient history and conducting thorough examinations are crucial in identifying the reasons behind therapeutic failures. We present a case study involving a patient with persistent trichomoniasis despite multiple treatment cycles, attributed to the presence of an intrauterine device inserted several years prior. This case underscores the intricacies involved in managing recurrent Trichomonas vaginalis infections and the importance of a comprehensive evaluation.

Dynamin 2-mediated endocytosis of BLT1 is required for IL-8 production in HMC-1 cells induced by Trichomonas vaginalis-derived secretory products.

We previously reported that leukotriene B4 (LTB4) contained in Trichomonas vaginalis-derived secretory products (TvSP) play an essential role in interleukin-8 (IL-8) production in human mast cell line (HMC-1 cells) via LTB4 receptor (BLT)-mediated Nuclear Factor-kappa B (NF-кB) activation. Dynamin, a GTPase, has been known to be involved in endocytosis of receptors for signaling of production of cytokine or chemokines. In the present study, we investigated the role of dynamin-mediated BLT1 endocytosis in TvSP-induced IL-8 production. When HMC-1 cells were transfected with BLT1 or BLT2 siRNA, TvSP-induced IL-8 production was significantly inhibited compared with that in cells transfected with control siRNA. In addition, pretreatment of HMC-1 cells with a dynamin inhibitor (Dynasore) reduced IL-8 production induced by TvSP or LTB4. TvSP- or LTB4- induced phosphorylation of NF-кB was also attenuated by pretreatment with Dynasore. After exposing HMC-1 cells to TvSP or LTB4, BLT1 was translocated from the intracellular compartments to the plasma membrane within 30 min. At 60 min after stimulation with TvSP or LTB4, BLT1 remigrated from the cell surface to intracellular areas. Pretreatment of HMC-1 cells with dynamin-2 siRNA blocked internalization of BLT1 induced by TvSP or LTB4. Co-immunoprecipitation experiments revealed that dynamin-2 strongly interacted with BLT1 60 min after stimulation with TvSP or LTB4. These results suggest that T. vaginalis-secreted LTB4 induces IL-8 production in HMC-1 cells via dynamin 2-mediated endocytosis of BLT1 and phosphorylation of NF-кB.

CysLT receptor-mediated NOX2 activation is required for IL-8 production in HMC-1 cells induced by Trichomonas vaginalis-derived secretory products.

Trichomoniasis is caused by a sexually transmitted flagellate protozoan parasite Trichomonas vaginalis. T. vaginalis-derived secretory products (TvSP) contain lipid mediators such as leukotriene B4 (LTB4) and various cysteinyl leukotrienes (CysLTs) which included LTC4, LTD4, and LTE4. However, the signaling mechanisms by which T. vaginalis-induced CysLTs stimulate interleukin (IL)-8 production in human mast cells remain unclear. In this study, we investigated these mechanisms in human mast cells (HMC-1). Stimulation with TvSP resulted in increased intracellular reactive oxygen species (ROS) generation and NADPH oxidase 2 (NOX2) activation compared to unstimulated cells. Pre-treatment with NOX2 inhibitors such as diphenyleneiodonium chloride (DPI) or apocynin significantly reduced ROS production in TvSP-stimulated HMC-1 cells. Additionally, TvSP stimulation increased NOX2 protein expression and the translocation of p47phox from the cytosol to the membrane. Pretreatment of HMC-1 cells with PI3K or PKC inhibitors reduced TvSP-induced p47phox translocation and ROS generation. Furthermore, NOX2 inhibitors or NOX2 siRNA prevented CREB phosphorylation and IL-8 gene expression or protein secretion induced by TvSP. Pretreatment with a CysLTR antagonist significantly inhibited TvSP-induced ROS production, CREB phosphorylation, and IL-8 production. These results indicate that CysLT-mediated activation of NOX2 plays a crucial role in ROS-dependent IL-8 production in human mast cells stimulated by T. vaginalis-secreted CysLTs. These findings enhance our understanding of the inflammatory response in trichomoniasis and may inform the development of targeted therapies to mitigate this response.

Laboratory study: Synthesis and optimization of nano nisomes containing Bunium persicum essential oil and investigating its toxicity on Trichomonas vaginalis parasite and HFF cell line.

The use of nanotechnology can reduce the challenges facing the use of herbal compounds in the fight against infectious agents. The aim of the present research is to produce nano niosomes containing Bunium persicum essential oil with high efficiency in the temperature and acidity of the living environment of Trichomonas vaginalis parasite and to investigate its toxicity on this parasite. First, Essential oil compounds were identified using GC-Mass. Then six niosomal formulations were made using Tween 40, 60, and 80 and cholesterol by thin film method. Three formulations that have more suitable particle size, zeta potential, and essential oil release and encapsulation efficiency were selected by MTT method to investigate the toxicity on HFF (Human foreskin fibroblasts) cell line. The formulation with lower toxicity was optimized using DSPE-mPEG(2000) polymer. Encapsulation efficiency, particle size, zeta potential, release of essential oil (in temperature and acidity similar to Trichomonas vaginalis living environment), particle morphology and toxicity of optimal formulation (on HFF and Trichomonas vaginalis) were investigated. At the end, the stability of the optimized nanoparticles was studied for 120 days. 12 chemical compounds including γ-Terpinene, Cuminic aldehyde and Para-cymene were identified Bunium persicum essential oil. The optimized formulation has a particle size of 159.73 nm, a zeta potential of -25.1 mV and an encapsulation efficiency of 63.11 %, which has a slow and continuous release at the similar temperature and acidity as Trichomonas vaginalis. Niosomal nanoparticles have a spherical shape and a smooth surface and have little toxicity on the HFF cell line. Also, the toxicity of nano niosomes containing essential oil on Trichomonas vaginalis is higher than free essential oil in all concentrations. The optimized niosomal nanoparticles have good stability because their physicochemical properties have changed very little during 120 days. In conclusion optimized Niosomal formulation containing Bunium persicum essential oil compared to free essential oil can have a higher efficiency to deal with Trichomonas parasite in laboratory conditions.

Sandwich enzyme-linked aptamer-based assay for the detection of Trichomonas vaginalis.

Trichomoniasis is the most prevalent curable, non-viral sexually transmitted infection (STI), with an estimated 156 million new infections in 2020. It can potentially result in adverse birth outcomes as well as infertility in men, whilst it also increases the risk of acquiring HIV and contracting other vaginal infections. It is mostly prevalent among women in low-income countries and especially in Africa and the Americas. This STI is caused by Trichomonas vaginalis (TV) and a robust, cost-effective, sensitive, specific and rapid diagnostic test is urgently required. We report the screening of 6 full-length and 4 truncated aptamers previously selected in our group for use in a microplate-based sandwich assay. The combination of dual aptamers comprising a short 14-mer truncated capture aptamer (termed A1_14mer) and a full-length non-truncated reporter aptamer (A6) was elucidated to be the optimum pair for a sensitive sandwich enzyme-linked aptamer assay (ELAA) for the detection of TV achieving a detection limit of 3.02 × 104 TV cells/mL. The results obtained with the A1_14mer-A6 ELAA correlate excellently with wet-mount microscopy for the detection of TV in clinical specimens, cervicovaginal lavages and vaginal swabs, highlighting the potential clinical application of this assay for cost-effective population screening and subsequent prevention of the onset of complications associated with undiagnosed and untreated TV.

Vaginitis and risk of sexually transmitted infections: results of a multi-center U.S. clinical study using STI nucleic acid amplification testing.

Significant increases in rates of sexually transmitted infections (STIs) caused by Trichomonas vaginalis (TV), Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and Mycoplasma genitalium (MG) are occurring in the United States. We present results of a U.S. study examining the intersection of STIs and vaginitis. Among 1,051 women with diagnoses for the presence or absence of bacterial vaginosis (BV) and/or symptomatic vulvovaginal candidiasis (VVC), 195 (18.5%) had one or more STIs, including 101 (9.6%) with TV, 24 (2.3%) with CT, 9 (0.8%) with NG, and 93 (8.8%) with MG. STI prevalence in BV-positive women was 26.3% (136/518), significantly higher than STI prevalence of 12.5% (59/474) in BV-negative women (P < 0.0002). Unlike infections with CT or NG, solo infections of MG or TV were each significantly associated with a diagnosis of BV-positive/VVC-negative (OR 3.0751; 95% CI 1.5797-5.9858, P = 0.0113, and OR 2.873; 95% CI 1.5687-5.2619, P = 0.0017, respectively) and with mixed infections containing MG and TV (OR 3.4886; 95% CI 1.8901-6.439, P = 0.0042, and OR 3.1858; 95% CI 1.809-5.6103, P = 0.0014, respectively). TV and MG infection rates were higher in all Nugent score (NS) categories than CT and NG infection rates; however, both STIs had similar comparative prevalence ratios to CT in NS 6-10 vs NS 0-5 (CT: 3.06% vs 1.4%, 2.2-fold; MG: 10.7% vs 6.1%, 1.8-fold; TV: 14.5% vs 7.0%, 2.1-fold). NG prevalence was relatively invariant by the NS category. These results highlight the complexity of associations of STIs with two major causes of vaginitis and underscore the importance of STI testing in women seeking care for abnormal vaginal discharge and inflammation. IMPORTANCE: This study reports high rates for sexually transmitted infections (STIs) in women seeking care for symptoms of vaginitis and bacterial vaginosis, revealing highly complex associations of STIs with two of the major causes of vaginal dysbiosis. These results underscore the importance of STI testing in women seeking care for abnormal vaginal discharge and inflammation.

Bacterial vaginosis (BV) and Trichomonas vaginalis (TV) co-infection, and bacterial antibiogram profile of pregnant women studied in Lagos, Nigeria.

AIM: This study was undertaken to determine the prevalence of Bacterial Vaginosis (BV), Trichomonas Vaginalis (TV) co-infection, and the antibacterial sensitivity profile of bacterial isolates. METHODS: The study was a cross-sectional study of 232 pregnant women on a routine antenatal visit between April 2019 and Sept. 2020, at Amukoko clinic in Lagos, Nigeria. The gynaecologist conducted the clinical examination on each patient looking for vaginal discharge and its consistency/homogeneity, colour and odour. Two High Vaginal Swab (HVS) samples were taken from every patient and a semi-structured questionnaire was used to gather the socio-demographic, practices/attitudes, and clinical information of each participant. One sample was employed for wet preparation to identify the TV and BV diagnosis using Amsel's criteria and Whiff's test. The second sample was used for bacterial culture and antibiogram was conducted using the disc diffusion technique. The Clinical Laboratory Standard Institutes' (CLSI) interpretative criteria were used to categorise the results. RESULTS: The mean age of the clients was 28.11 ± 7.08 years of age. The majority (88%) were aged 15-35 years. Only 81 (34.9%) had microbial organisms isolated or seen from their specimens and 19 (8.2%) of such were classified as having BV (Bacteriods or Gardnerella isolated). Of the 81 infected, 33 (40.8%) had only bacterial infection, 36 (44.4%) had TV alone and 12 (14.8%) had bacteria co-infected with TV. From the clinical records, the population that was classified as having UTI or vaginitis was only 46 (20.7%) The study observed age (15-35 years) related association between vaginosis/ TV co-infection (X2 = 7.9; P = 0.005). Participants with symptoms of vaginitis or UTI (mainly E. coli & pseudomonas spp. isolated), BV/co-infection with TV significantly associated with female traders (X2 = 8.5; P = 0.003) and were more associated with those from polygamous relationships (X2 = 18.79, P = 0.0001). Women in their 3rd and 2nd. trimester were more significantly associated with vaginal infection (X2 = 9.47, P = 0.002; X2 = 4.79, P = 0.029) respectively. The Pseudomonas showed susceptibility to ciprofloxacin (CIP) and cefuroxime (CXM). While, E. coli isolates were susceptible to cefepime, ciprofloxacin, and imipenem. CONCLUSION: There is a relatively low prevalence of BV and flagellate co-infection in the community studied. RECOMMENDATION: We recommend screening of antenatal women with underlying symptoms for BV and flagellates co-infection to avoid its progression to vaginitis.

Epidemiology of Trichomonas vaginalis infection in the Middle East and North Africa: systematic review, meta-analyses, and meta-regressions.

BACKGROUND: Trichomoniasis, caused by the parasite Trichomonas vaginalis (TV), remains an underappreciated sexually transmitted infection (STI), primarily due to inadequate understanding of its epidemiology and public health implications. This study aimed to characterize TV epidemiology in the Middle East and North Africa (MENA). METHODS: Systematic review and analysis of evidence sourced from international, regional, and national databases were conducted. Findings were reported following PRISMA guidelines. Random-effects meta-analyses and meta-regressions were performed to determine pooled mean prevalence, investigate associations with prevalence, and identify sources of between-study heterogeneity. FINDINGS: The review identified 263 relevant publications, encompassing 462 TV prevalence measures. The pooled mean TV prevalence was estimated as follows: 4.7% (95% CI: 3.9-5.6%) in the general population of women, 17.2% (95% CI: 5.4-33.6%) among intermediate-risk populations, 10.3% (95% CI: 6.2-15.3%) among female sex workers, 13.9% (95% CI: 12.3-15.6%) among symptomatic women, 7.4% (95% CI: 1.9-15.5%) among infertility clinic attendees, 2.3% (95% CI: 0.1-6.3%) among women with miscarriages or ectopic pregnancies, and 1.6% (95% CI: 0.8-2.7%) among STI clinic attendees. Limited data were found for men. Multivariable meta-regressions explained >40% of the prevalence variation, unveiling a hierarchical prevalence pattern by population type, an inverse correlation with national income, and a prevalence decline at a rate of 1% per calendar year. INTERPRETATION: Despite conservative sexual norms, MENA has a substantial TV prevalence, comparable to the global TV prevalence. The unexpectedly high prevalence of this curable infection may, in part, be attributed to limited access to and underutilization of STI screening and treatment services. FUNDING: This work was supported by the Qatar Research, Development, and Innovation Council [ARG01-0522-230273] and by the Biomedical Research Program at Weill Cornell Medicine-Qatar.

The Prevalence of Sexually Transmitted Infections among Male Patients at a Dermato-Venereology Outpatient Clinic in Gdansk, Poland: Findings from a Single-Center Study.

Background: Sexually transmitted infections (STIs) are a significant public health concern worldwide, yet data on their prevalence and epidemiology, particularly in Central and Eastern Europe, remain scarce. This study aimed to assess the prevalence, anatomical localization, symptomatic/asymptomatic course, and co-infection patterns of STIs among men. Methods: This retrospective study analyzed data collected between May 2021 and July 2023, including sociodemographic, sexual behavior, and clinical data from 139 male participants. Molecular polymerase chain reaction (PCR) tests were conducted for Neisseria gonorrhoeae (NG), Chlamydia trachomatis (CT), Mycoplasma genitalium, and Trichomonas vaginalis. Results: Of the participants, 36% tested positive for at least one STI, with the urethra being the most common site of infection. NG and CT were the most prevalent infections. The majority of infections were asymptomatic, highlighting the importance of comprehensive screening, especially in high-risk populations like men who have sex with men (MSM). Conclusions: This study emphasizes the need for targeted screening strategies, particularly for extragenital STIs, and underscores the role of MSM in STI epidemiology. The findings highlight the importance of routine screening, even for asymptomatic individuals, to effectively control STI spread. Future research should validate and expand upon these findings to enhance STI prevention and management efforts.

A hybrid TIM complex mediates protein import into hydrogenosomes of Trichomonas vaginalis.

BACKGROUND: Hydrogenosomes are a specific type of mitochondria that have adapted for life under anaerobiosis. Limited availability of oxygen has resulted in the loss of the membrane-associated respiratory chain, and consequently in the generation of minimal inner membrane potential (Δψ), and inefficient ATP synthesis via substrate-level phosphorylation. The changes in energy metabolism are directly linked with the organelle biogenesis. In mitochondria, proteins are imported across the outer membrane via the Translocase of the Outer Membrane (TOM complex), while two Translocases of the Inner Membrane, TIM22, and TIM23, facilitate import to the inner membrane and matrix. TIM23-mediated steps are entirely dependent on Δψ and ATP hydrolysis, while TIM22 requires only Δψ. The character of the hydrogenosomal inner membrane translocase and the mechanism of translocation is currently unknown. RESULTS: We report unprecedented modification of TIM in hydrogenosomes of the human parasite Trichomonas vaginalis (TvTIM). We show that the import of the presequence-containing protein into the hydrogenosomal matrix is mediated by the hybrid TIM22-TIM23 complex that includes three highly divergent core components, TvTim22, TvTim23, and TvTim17-like proteins. The hybrid character of the TvTIM is underlined by the presence of both TvTim22 and TvTim17/23, association with small Tim chaperones (Tim9-10), which in mitochondria are known to facilitate the transfer of substrates to the TIM22 complex, and the coupling with TIM23-specific ATP-dependent presequence translocase-associated motor (PAM). Interactome reconstruction based on co-immunoprecipitation (coIP) and mass spectrometry revealed that hybrid TvTIM is formed with the compositional variations of paralogs. Single-particle electron microscopy for the 132-kDa purified TvTIM revealed the presence of a single ring of small Tims complex, while mitochondrial TIM22 complex bears twin small Tims hexamer. TvTIM is currently the only TIM visualized outside of Opisthokonta, which raised the question of which form is prevailing across eukaryotes. The tight association of the hybrid TvTIM with ADP/ATP carriers (AAC) suggests that AAC may directly supply ATP for the protein import since ATP synthesis is limited in hydrogenosomes. CONCLUSIONS: The hybrid TvTIM in hydrogenosomes represents an original structural solution that evolved for protein import when Δψ is negligible and remarkable example of evolutionary adaptation to an anaerobic lifestyle.

Trichomonas vaginalis extracellular vesicles up-regulate and directly transfer adherence factors promoting host cell colonization.

Trichomonas vaginalis, a common sexually transmitted parasite that colonizes the human urogenital tract, secretes extracellular vesicles (TvEVs) that are taken up by human cells and are speculated to be taken up by parasites as well. While the crosstalk between TvEVs and human cells has led to insight into host:parasite interactions, roles for TvEVs in infection have largely been one-sided, with little known about the effect of TvEV uptake by T. vaginalis. Approximately 11% of infections are found to be coinfections of multiple T. vaginalis strains. Clinical isolates often differ in their adherence to and cytolysis of host cells, underscoring the importance of understanding the effects of TvEV uptake within the parasite population. To address this question, our lab tested the ability of a less adherent strain of T. vaginalis, G3, to take up fluorescently labeled TvEVs derived from both itself (G3-EVs) and TvEVs from a more adherent strain of the parasite (B7RC2-EVs). Here, we showed that TvEVs generated from the more adherent strain are internalized more efficiently compared to the less adherent strain. Additionally, preincubation of G3 parasites with B7RC2-EVs increases parasite aggregation and adherence to host cells. Transcriptomics revealed that TvEVs up-regulate expression of predicted parasite membrane proteins and identified an adherence factor, heteropolysaccharide binding protein (HPB2). Finally, using comparative proteomics and superresolution microscopy, we demonstrated direct transfer of an adherence factor, cadherin-like protein, from TvEVs to the recipient parasite's surface. This work identifies TvEVs as a mediator of parasite:parasite communication that may impact pathogenesis during mixed infections.

Structural characterization of the (deoxy)hypusination in Trichomonas vaginalis questions the bifunctionality of deoxyhypusine synthase.

Trichomonas vaginalis, the causative agent of trichomoniasis, is a prevalent anaerobic protozoan parasite responsible for the most common nonviral sexually transmitted infection globally. While metronidazole and its derivatives are approved drugs for this infection, rising resistance necessitates the exploration of new antiparasitic therapies. Protein posttranslational modifications (PTMs) play crucial roles in cellular processes, and among them, hypusination, involving eukaryotic translation factor 5A (eIF5A), has profound implications. Despite extensive studies in various organisms, the role of hypusination in T. vaginalis and its potential impact on parasite biology and pathogenicity remain poorly understood. This study aims to unravel the structural basis of the hypusination pathway in T. vaginalis using X-ray crystallography and cryo-electron microscopy. The results reveal high structural homology between T. vaginalis and human orthologs, providing insights into the molecular architecture of eIF5A and deoxyhypusine synthase (DHS) and their interaction. Contrary to previous suggestions of bifunctionality, our analyses indicate that the putative hydroxylation site in tvDHS is nonfunctional, and biochemical assays demonstrate exclusive deoxyhypusination capability. These findings challenge the notion of tvDHS functioning as both deoxyhypusine synthase and hydroxylase. The study enhances understanding of the hypusination pathway in T. vaginalis, shedding light on its functional relevance and potential as a drug target, and contributing to the development of novel therapeutic strategies against trichomoniasis.

Prevalence of Trichomonas Infection in Relation to Human Papillomavirus (HPV) in Pap Smear Samples of Female Patients Referred to Shahid Sadoughi Hospital, Yazd (Iran).

Objectives Human papillomavirus (HPV) and Trichomonas vaginalis (TV) infections have been proposed as risk factors for cervical cancer. This study has been conducted with the aim of investigating the prevalence of TV and its relationship with HPV in women who underwent Pap smear testing as part of cancer screenings. Materials and methods The sampling of liquid-based cervical tissue was conducted among 500 women referred to the women's clinic of Shahid Sadoughi Hospital in Yazd, Iran. The studied samples were examined for Pap smear tests and microscopic identification of TV, as well as HPV-DNA detection and the determination of high-risk and low-risk HPV types by the polymerase chain reaction (PCR) method. The results were analyzed using the IBM SPSS Statistics for Windows, Version 24 (Released 2016; IBM Corp., Armonk, New York) software. Results The individuals included in the study were 16-72 years old. The prevalence rate of TV infection in this population was found to be 29.2%, and the frequency rate of HPV was reported to be approximately 19.4%, with high-risk HPV, including HPV-56, having the highest frequency. The Pap smear test results were reported as abnormal in 20.2%, and a significant correlation was observed between HPV infection and an abnormal Pap smear test (P < 0.05). In addition, a notable correlation was detected between TV infection and high-risk and low-risk HPV (P < 0.05). Conclusion According to the significant relationship found between the two pathogens, TV and HPV, in the abnormal Pap smear test results, TV infection can be considered a risk factor for HPV infection, as well as uterine lesions and cancer.

Development of a DNA-Based Lateral Flow Strip Membrane Assay for Rapid Screening and Genotyping of Six High-Incidence STD Pathogens.

Sexually transmitted diseases (STDs) are a global concern because approximately 1 million new cases emerge daily. Most STDs are curable, but if left untreated, they can cause severe long-term health implications, including infertility and even death. Therefore, a test enabling rapid and accurate screening and genotyping of STD pathogens is highly awaited. Herein, we present the development of the DNA-based 6STD Genotyping 9G Membrane test, a lateral flow strip membrane assay, for the detection and genotyping of six STD pathogens, including Trichomonas vaginalis, Ureaplasma urealyticum, Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma hominis, and Mycoplasma genitalium. Here, we developed a multiplex PCR primer set that allows PCR amplification of genomic materials for these six STD pathogens. We also developed the six ssDNA probes that allow highly efficient detection of the six STD pathogens. The 6STD Genotyping 9G Membrane test lets us obtain the final detection and genotyping results in less than 30 m after PCR at 25 °C. The accuracy of the 6STD Genotyping 9G membrane test in STD genotyping was confirmed by its 100% concordance with the sequencing results of 120 clinical samples. Therefore, the 6STD Genotyping 9G Membrane test emerges as a promising diagnostic tool for precise STD genotyping, facilitating informed decision-making in clinical practice.

The presence of Trichomonas vaginalis in urogenital samples can affect the sensitivity of Mycoplasma hominis identification techniques, leading to an underestimation of bacterial infections.

Trichomonas vaginalis and Mycoplasma hominis, two microorganisms causing infections of the urogenital tract, are closely associated in that they establish an endosymbiosis relationship, the only case among human pathogens. As a result, the presence of one microorganism may be considered a sign that the other is present as well. Identification of the two pathogens in clinical samples is based on cultivation techniques on specific media, even though in recent years, new sensitive and rapid molecular techniques have become. Here, we demonstrate that the concomitant presence of T.vaginalis in urogenital swabs may lead to a delay in the identification of M.hominis, and thus to an underestimation of bacterial infections when cultural techniques are used.

A novel fluoro colorimetric Loop mediated isothermal amplification (LAMP) assay for detection of Trichomonasvaginalis.

BACKGROUND: Trichomoniasis is a curable, non-viral, sexually transmitted infection. Early diagnosis and treatment of cases can prevent complications and further spread of infection. Rapid diagnostics tests, which can be performed on-site, will help in early diagnosis. The study aims to develop a rapid diagnostic test based on the principle of fluoro-colorimetric LAMP for detecting Trichomonas vaginalis (TV). MATERIALS AND METHODS: T. vaginalis was grown in a modified CPLM medium, and DNA was extracted. Three pairs of LAMP primers targeting the actin gene were designed using the primerexplorer V.5 online tool. The LAMP assay was standardized for temperature and time. To determine the LAMP assay's detection limit, diluted TV DNA and spiked urine samples were used. Conventional PCR was done using previously published primers and compared with LAMP results. The sensitivity and specificity to detect TV from clinical specimens were assessed. RESULTS: The optimum performance of the LAMP assay was determined to be 63 °C for 60 min and terminated at 80 °C for 5 min. The LAMP assay could detect 60 fg/µl of diluted TV DNA and up to 1 parasite/ml of spiked samples. The assay was 1000 times more sensitive than PCR. The LAMP assay was 100% sensitive and specific with crude extract, and reactions were visually discernible. INTERPRETATION & CONCLUSIONS: The LAMP assay developed in the study is easy to perform and interpret, affordable, rapid, and highly sensitive to detect T. vaginalis. It is ideally suited for the point-of-care test, as it fulfills WHO's recommended ASSURED characteristics.

Characterization of Actin-based Genotypes and Mycoplasma Endosymbionts of Trichomonas vaginalis Isolated in Sapporo, Japan.

PURPOSE: Trichomonas vaginalis is a causative agent of common non-viral sexually transmitted infections worldwide. However, the biological features, such as genotypes and endosymbionts, of T. vaginalis isolated in Japan remain unclear. The aim of this study was to characterize the actin-based genotypes and the endosymbionts of T. vaginalis isolated in Sapporo, Japan. METHODS: Three T. vaginalis clinical strains were isolated in Sapporo, Japan between 2019 and 2022. Actin-based genotyping was conducted by sequencing and phylogenetic analyses. The endosymbionts, such as Mycoplasma sp. and Trichomonasvirus, were detected using PCR and RT-PCR, respectively. Furthermore, the detected Mycoplasma spp. were identified using 16S rRNA gene sequencing. RESULTS: Of the three T. vaginalis strains, two belonged to genotype E, whereas one was genotype G as determined by actin-based genotyping. Two of the T. vaginalis strains harbored Mycoplasma spp. Using nearly full-length 16S rRNA gene sequencing, both were identified as Candidatus Mycoplasma girerdii. In contrast, the Trichomonasvirus was not found in the T. vaginalis strains. CONCLUSION: To our knowledge, this is the first report on the characterization of actin-based genotypes and the presence of endosymbiotic Ca. M. girerdii in T. vaginalis strains in Japan. Thus, this study will provide an important impetus for future research.