Search Results (27)

Background/Objectives: Non-syndromic cleft lip with or without palate (NSCL/P) is a common, multifactorial congenital anomaly. As genetic associations can be population-specific, this study aimed to investigate single-nucleotide polymorphisms (SNPs) in the VAX1, MAFB, and WNT3 genes for association with NSCL/P in a Japanese cohort. Methods: A case–control study was conducted with 310 Japanese patients with NSCL/P and 308 ethnically matched healthy controls from Aichi Gakuin Dental Hospital. We genotyped SNPs rs7078160 (VAX1), rs13041247 (MAFB), and rs3809857 (WNT3) using TaqMan assays. Associations were assessed using chi-squared tests, with results stratified by sex and corrected for multiple comparisons using the Bonferroni method. Results: The VAX1 rs7078160 A allele was significantly associated with an increased risk for NSCL/P (OR = 1.67, p < 0.00001). The association was particularly strong in females (OR = 1.93, p < 0.00001) but not significant in males after correction. The MAFB rs13041247 variant showed a nominal protective association with the NSCLO subtype that was not significant after Bonferroni correction. No significant association was found for WNT3. A notable gene–gene interaction was observed, where carrying risk alleles for both VAX1 and MAFB significantly increased overall NSCL/P risk (OR = 2.65, p = 0.00008). Conclusions: VAX1 rs7078160 is a significant risk factor for NSCL/P in the Japanese population, with a pronounced female-specific effect. A synergistic interaction between VAX1 and MAFB elevates disease risk, whereas WNT3 was not implicated in this cohort. These findings underscore the population-specific genetic architecture of NSCL/P. Full article

Background: Non-syndromic orofacial clefts (NSOFCs) are one of the common congenital malformations in Vietnam, with 1.4 per 1000 live births, with notable sex differences in occurrence. This case–control study aims to investigate potential sex-specific interactions of WNT3 and NOG polymorphisms across NSOFC subtypes in a Vietnamese population. Methods: A total of 720 participants were separated into 4 groups with a male/female ratio of 1:1 (160 individuals with cleft lip and palate (NSCLP), 160 with cleft lip only (NSCLO), 160 with cleft palate only (NSCPO), 240 healthy controls). Two single-nucleotide polymorphisms (SNPs), rs3809857 and rs227731, were genotyped by using the StepOnePlus Real-Time PCR System. Results: The most significant findings were found in the male NSCLO group under a recessive model of WNT3 rs3809857 after applying Bonferroni correction, as a five-fold protective factor with OR = 0.18 (95% confidence interval: 0.05–0.64, p = 0.0033). Additionally, the weak or moderate protective association between rs3809857 and male NSCLP was found with p < 0.05 under the dominant model. However, there were no significant findings in the female NSOFC subtypes associated with WNT3. Conversely, NOG rs227731 results showed a weak increased risk in female NSCLO and NSCPO with p < 0.05. Conclusion: this study identified the critical role of WNT3 rs3809857 in reducing NSCLO risk in males. These findings support the potential influence of sex as a modifying factor in the genetic susceptibility to non-syndromic orofacial clefts. Full article

Trypanosomes are parasitic protozoa that cause severe diseases in humans and animals. The most important species of Trypanosmes include Trypanosoma evansi and Trypanosoma brucei gambiense. The most well-known human diseases are sleeping sickness in Africa and Chagas disease in South America. The most identified animal diseases include Nagana in the African tsetse fly belt and Surra in South Asia, North Africa, and the Middle East. Surra is caused by Trypanosoma evansi. Diminazene resistance is an emerging threat caused by T. evansi infecting animals. The underlying mechanism of diminazene resistance is poorly understood. Trypanosoma brucei gambiense causes African sleeping sickness. The development of diminazene resistance in Trypanosoma brucei gambiense is associated with the alterations in the corresponding P2 adenosine transporter-1 (AT-1) gene. In the present study, by extrapolating the findings from Trypanosoma brucei gambiense, we analyzed genetic diversity in the P2 adenosine transporter-1 gene (AT-1) from T. evansi to explore a potential link between the presence of mutations in this locus and diminazene treatment in ruminants. We examined T. evansi-infected blood samples collected from goats, sheep, camels, buffalo, and cattle in seven known endemic regions of the Punjab province of Pakistan. Heterozygosity (H_e) indices indicated a high level of genetic diversity between seven T. evansi field isolates that had resistance-type mutations at codons 178E/S, 239Y/A/E, and 286S/H/I/D/T of the P2 adenosine transporter-1 (AT-1) locus. A low level of genetic diversity was observed in 19 T. evansi field isolates with susceptible-type mutations at codons A178, G181, D239, and N286 of the P2 adenosine transporter-1 (AT-1) locus. Our results on T. evansi warrant further functional studies to explore the relationship between diminazene resistance and the mutations in AT-1. Full article

Animal trypanosomiasis, such as nagana, surra, and dourine, represent a significant challenge to animal health and economic development, especially in tropical and subtropical regions where livestock production is an essential component of a country’s economy. Despite advances in the control of human trypanosomiasis, animal diseases caused by several species of trypanosomes remain neglected. The lack of funding for the development of new treatments and vaccines contributes to sustaining the severe economic impacts these diseases have on the farming industry, especially in low-income rural areas. Recent advances in the understanding of the immune processes involved during infection have been essential for the development of new approaches towards disease control including vaccines. These new approaches must be part of integrated control programs, which must also include vector management and the awareness of good veterinary practices. Addressing the challenges posed by the control of animal trypanosomiasis requires collaborative and continuous efforts shared among scientists, governments, and the farming industry, if significant progress is to be made to mitigate the impact of these diseases. In this literature review, we discuss the main challenges for the development of vaccines for animal trypanosomiasis and the research underway, including the prospects for employing new vaccine platforms, such as an mRNA vaccine, vector-based vaccine, and CRISPR-attenuated parasite vaccine. Full article

Metabolomics has been used extensively to capture the exposome. We investigated whether prospectively measured metabolites provided predictive power beyond well-established risk factors among 758 women with adjudicated cancers [n = 577 breast (BC) and n = 181 colorectal (CRC)] and n = 758 controls with available specimens (collected mean 7.2 years prior to diagnosis) in the Women’s Health Initiative Bone Mineral Density subcohort. Fasting samples were analyzed by LC-MS/MS and lipidomics in serum, plus GC-MS and NMR in 24 h urine. For feature selection, we applied LASSO regression and Super Learner algorithms. Prediction models were subsequently derived using logistic regression and Super Learner procedures, with performance assessed using cross-validation (CV). For BC, metabolites did not increase predictive performance over established risk factors (CV-AUCs~0.57). For CRC, prediction increased with the addition of metabolites (median CV-AUC across platforms increased from ~0.54 to ~0.60). Metabolites related to energy metabolism: adenosine, 2-hydroxyglutarate, N-acetyl-glycine, taurine, threonine, LPC (FA20:3), acetate, and glycerate; protein metabolism: histidine, leucic acid, isoleucine, N-acetyl-glutamate, allantoin, N-acetyl-neuraminate, hydroxyproline, and uracil; and dietary/microbial metabolites: myo-inositol, trimethylamine-N-oxide, and 7-methylguanine, consistently contributed to CRC prediction. Energy metabolism may play a key role in the development of CRC and may be evident prior to disease development. Full article

The parasites Trypanosoma brucei (Tb) and Leishmania major (Lm) cause the tropical diseases sleeping sickness, nagana, and cutaneous leishmaniasis. Every year, millions of humans, as well as animals, living in tropical to subtropical climates fall victim to these illnesses’ health threats. The parasites’ frequent drug resistance and widely spread natural reservoirs heavily impede disease prevention and treatment. Due to pteridine auxotrophy, trypanosomatid parasites have developed a peculiar enzyme system consisting of dihydrofolate reductase-thymidylate synthase (DHFR-TS) and pteridine reductase 1 (PTR1) to support cell survival. Extending our previous studies, we conducted a comparative study of the T. brucei (TbDHFR, TbPTR1) and L. major (LmDHFR, LmPTR1) enzymes to identify lead structures with a dual inhibitory effect. A pharmacophore-based in silico screening of three natural product databases (approximately 4880 compounds) was performed to preselect possible inhibitors. Building on the in silico results, the inhibitory potential of promising compounds was verified in vitro against the recombinant DHFR and PTR1 of both parasites using spectrophotometric enzyme assays. Twelve compounds were identified as dual inhibitors against the Tb enzymes (0.2 μM < IC₅₀ < 85.1 μM) and ten against the respective Lm enzymes (0.6 μM < IC₅₀ < 84.5 μM). These highly promising results may represent the starting point for the future development of new leads and drugs utilizing the trypanosomatid pteridine metabolism as a target. Full article

This study aims to identify potential variants in the TP63–IRF6 pathway and GREM1 for the etiology of non-syndromic orofacial cleft (NSOFC) among the Vietnamese population. By collecting 527 case–parent trios and 527 control samples, we conducted a stratified analysis based on different NSOFC phenotypes, using allelic, dominant, recessive and over-dominant models for case–control analyses, and family-based association tests for case–parent trios. Haplotype and linkage disequilibrium analyses were also conducted. IRF6 rs2235375 showed a significant association with an increased risk for non-syndromic cleft lip and palate (NSCLP) and cleft lip with or without cleft palate (NSCL/P) in the G allele, with p_allele values of 0.0018 and 0.0003, respectively. Due to the recessive model (p = 0.0011) for the NSCL/P group, the reduced frequency of the GG genotype of rs2235375 was associated with a protective effect against NSCL/P. Additionally, offspring who inherited the G allele at rs2235375 had a 1.34-fold increased risk of NSCL/P compared to the C allele holders. IRF6 rs846810 and a G-G haplotype at rs2235375–rs846810 of IRF6 impacted NSCL/P, with p-values of 0.0015 and 0.0003, respectively. In conclusion, our study provided additional evidence for the association of IRF6 rs2235375 with NSCLP and NSCL/P. We also identified IRF6 rs846810 as a novel marker associated with NSCL/P, and haplotypes G-G and C-A at rs2235375–rs846810 of IRF6 associated with NSOFC. Full article

Demographic and clinical factors influence the metabolome. The discovery and validation of disease biomarkers are often challenged by potential confounding effects from such factors. To address this challenge, we investigated the magnitude of the correlation between serum and urine metabolites and demographic and clinical parameters in a well-characterized observational cohort of 444 post-menopausal women participating in the Women’s Health Initiative (WHI). Using LC-MS and lipidomics, we measured 157 aqueous metabolites and 756 lipid species across 13 lipid classes in serum, along with 195 metabolites detected by GC-MS and NMR in urine and evaluated their correlations with 29 potential disease risk factors, including demographic, dietary and lifestyle factors, and medication use. After controlling for multiple testing (FDR < 0.01), we found that log-transformed metabolites were mainly associated with age, BMI, alcohol intake, race, sample storage time (urine only), and dietary supplement use. Statistically significant correlations were in the absolute range of 0.2–0.6, with the majority falling below 0.4. Incorporation of important potential confounding factors in metabolite and disease association analyses may lead to improved statistical power as well as reduced false discovery rates in a variety of data analysis settings. Full article

African Animal Trypanosomiasis (AAT), caused predominantly by Trypanosoma brucei brucei, T. vivax and T. congolense, is a fatal livestock disease throughout Sub-Saharan Africa. Treatment options are very limited and threatened by resistance. Tubercidin (7-deazaadenosine) analogs have shown activity against individual parasites but viable chemotherapy must be active against all three species. Divergence in sensitivity to nucleoside antimetabolites could be caused by differences in nucleoside transporters. Having previously characterized the T. brucei nucleoside carriers, we here report the functional expression and characterization of the main adenosine transporters of T. vivax (TvxNT3) and T. congolense (TcoAT1/NT10), in a Leishmania mexicana cell line (‘SUPKO’) lacking adenosine uptake. Both carriers were similar to the T. brucei P1-type transporters and bind adenosine mostly through interactions with N3, N7 and 3′-OH. Expression of TvxNT3 and TcoAT1 sensitized SUPKO cells to various 7-substituted tubercidins and other nucleoside analogs although tubercidin itself is a poor substrate for P1-type transporters. Individual nucleoside EC₅₀s were similar for T. b. brucei, T. congolense, T. evansi and T. equiperdum but correlated less well with T. vivax. However, multiple nucleosides including 7-halogentubercidines displayed pEC50>7 for all species and, based on transporter and anti-parasite SAR analyses, we conclude that nucleoside chemotherapy for AAT is viable. Full article

Trypanosoma brucei is a species of kinetoplastid causing sleeping sickness in humans and nagana in cows and horses. One of the peculiarities of this species of parasites is represented by their redox metabolism. One of the proteins involved in this redox machinery is the monothiol glutaredoxin 1 (1CGrx1) which is characterized by a unique disordered N-terminal extension exclusively conserved in trypanosomatids and other organisms. This region modulates the binding profile of the glutathione/trypanothione binding site, one of the functional regions of 1CGrx1. No endogenous ligands are known to bind this protein which does not present well-shaped binding sites, making it target particularly challenging to target. With the aim of targeting this peculiar system, we carried out two different screenings: (i) a fragment-based lead discovery campaign directed to the N-terminal as well as to the canonical binding site of 1CGrx1; (ii) a structure-based virtual screening directed to the 1CGrx1 canonical binding site. Here we report a small molecule that binds at the glutathione binding site in which the binding mode of the molecule was deeply investigated by Nuclear Magnetic Resonance (NMR). This compound represents an important step in the attempt to develop a novel strategy to interfere with the peculiar Trypanosoma Brucei redox system, making it possible to shed light on the perturbation of this biochemical machinery and eventually to novel therapeutic possibilities. Full article

Nicotinamide adenine dinucleotide (NAD⁺) is an essential cofactor required for proper functioning of all cells and its decline is correlated with advancing age and disease. This randomized, triple-blind, placebo-controlled, crossover pilot study assessed the efficacy and safety of a combination of nicotinamide with D-ribose (RiaGev) for NAD metabolome enhancement and related benefits in healthy middle-aged adults. Supplementing with 1520 mg RiaGev twice daily for 7 days significantly increased the NAD⁺ metabolome in blood, especially NADP⁺ by 27% compared to the placebo group (p = 0.033) and over the baseline (p = 0.007). Increases in glutathione and high energy phosphates were also observed in the blood. Seven-day supplementation with RiaGev significantly (p = 0.013) reduced overall blood glucose without significant changes in insulin secretion (p = 0.796), suggesting an improved insulin sensitivity and glucose tolerance. The waking salivary cortisol of the subjects steadily and significantly decreased (p = 0.026) in the RiaGev group in contrast to the placebo. Subjects in the RiaGev group showed less fatigue, improved mental concentration and motivation over the baseline (p = 0.015, 0.018, and 0.012, respectively) as observed through the Checklist Individual Strength (CIS) questionnaire. There were no clinically relevant adverse events, or alterations in hematology, electrolytes, liver, and kidney markers pre- and post-supplementation. RiaGev appears to be safe and efficacious in increasing NAD⁺ metabolome in healthy middle-aged adults, as shown by this study. Full article

Inorganic polyphosphate (polyP) is an ancient biopolymer that is well preserved throughout evolution and present in all studied organisms. In mammals, it shows a high co-localization with mitochondria, and it has been demonstrated to be involved in the homeostasis of key processes within the organelle, including mitochondrial bioenergetics. However, the exact extent of the effects of polyP on the regulation of cellular bioenergetics, as well as the mechanisms explaining these effects, still remain poorly understood. Here, using HEK293 mammalian cells under Wild-type (Wt) and MitoPPX (cells enzymatically depleted of mitochondrial polyP) conditions, we show that depletion of polyP within mitochondria increased oxidative stress conditions. This is characterized by enhanced mitochondrial O₂⁻ and intracellular H₂O₂ levels, which may be a consequence of the dysregulation of oxidative phosphorylation (OXPHOS) that we have demonstrated in MitoPPX cells in our previous work. These findings were associated with an increase in basal peroxiredoxin-1 (Prx1), superoxide dismutase-2 (SOD2), and thioredoxin (Trx) antioxidant protein levels. Using ¹³C-NMR and immunoblotting, we assayed the status of glycolysis and the pentose phosphate pathway (PPP) in Wt and MitoPPX cells. Our results show that MitoPPX cells display a significant increase in the activity of the PPP and an increase in the protein levels of transaldolase (TAL), which is a crucial component of the non-oxidative phase of the PPP and is involved in the regulation of oxidative stress. In addition, we observed a trend towards increased glycolysis in MitoPPX cells, which corroborates our prior work. Here, for the first time, we show the crucial role played by mitochondrial polyP in the regulation of mammalian redox homeostasis. Moreover, we demonstrate a significant effect of mitochondrial polyP on the regulation of global cellular bioenergetics in these cells. Full article

The parasite Trypanosoma brucei (T. brucei) is responsible for human African trypanosomiasis (HAT) and the cattle disease “Nagana” which to this day cause severe medical and socio-economic issues for the affected areas in Africa. So far, most of the available treatment options are accompanied by harmful side effects and are constantly challenged by newly emerging drug resistances. Since trypanosomatids are auxotrophic for folate, their pteridine metabolism provides a promising target for an innovative chemotherapeutic treatment. They are equipped with a unique corresponding enzyme system consisting of the bifunctional dihydrofolate reductase-thymidylate synthase (TbDHFR-TS) and the pteridine reductase 1 (TbPTR1). Previously, gene knockout experiments with PTR1 null mutants have underlined the importance of these enzymes for parasite survival. In a search for new chemical entities with a dual inhibitory activity against the TbPTR1 and TbDHFR, a multi-step in silico procedure was employed to pre-select promising candidates against the targeted enzymes from a natural product database. Among others, the sesquiterpene lactones (STLs) cynaropicrin and cnicin were identified as in silico hits. Consequently, an in-house database of 118 STLs was submitted to an in silico screening yielding 29 further virtual hits. Ten STLs were subsequently tested against the target enzymes in vitro in a spectrophotometric inhibition assay. Five compounds displayed an inhibition over 50% against TbPTR1 as well as three compounds against TbDHFR. Cynaropicrin turned out to be the most interesting hit since it inhibited both TbPTR1 and TbDHFR, reaching IC₅₀ values of 12.4 µM and 7.1 µM, respectively. Full article

Tsetse flies cause major health and economic problems as they transmit trypanosomes causing sleeping sickness in humans (Human African Trypanosomosis, HAT) and nagana in animals (African Animal Trypanosomosis, AAT). A solution to control the spread of these flies and their associated diseases is the implementation of the Sterile Insect Technique (SIT). For successful application of SIT, it is important to establish and maintain healthy insect colonies and produce flies with competitive fitness. However, mass production of tsetse is threatened by covert virus infections, such as the Glossina pallidipes salivary gland hypertrophy virus (GpSGHV). This virus infection can switch from a covert asymptomatic to an overt symptomatic state and cause the collapse of an entire fly colony. Although the effects of GpSGHV infections can be mitigated, the presence of other covert viruses threaten tsetse mass production. Here we demonstrated the presence of two single-stranded RNA viruses isolated from Glossina morsitans morsitans originating from a colony at the Seibersdorf rearing facility. The genome organization and the phylogenetic analysis based on the RNA-dependent RNA polymerase (RdRp) revealed that the two viruses belong to the genera Iflavirus and Negevirus, respectively. The names proposed for the two viruses are Glossina morsitans morsitans iflavirus (GmmIV) and Glossina morsitans morsitans negevirus (GmmNegeV). The GmmIV genome is 9685 nucleotides long with a poly(A) tail and encodes a single polyprotein processed into structural and non-structural viral proteins. The GmmNegeV genome consists of 8140 nucleotides and contains two major overlapping open reading frames (ORF1 and ORF2). ORF1 encodes the largest protein which includes a methyltransferase domain, a ribosomal RNA methyltransferase domain, a helicase domain and a RdRp domain. In this study, a selective RT-qPCR assay to detect the presence of the negative RNA strand for both GmmIV and GmmNegeV viruses proved that both viruses replicate in G. m. morsitans. We analyzed the tissue tropism of these viruses in G. m. morsitans by RNA-FISH to decipher their mode of transmission. Our results demonstrate that both viruses can be found not only in the host’s brain and fat bodies but also in their reproductive organs, and in milk and salivary glands. These findings suggest a potential horizontal viral transmission during feeding and/or a vertically viral transmission from parent to offspring. Although the impact of GmmIV and GmmNegeV in tsetse rearing facilities is still unknown, none of the currently infected tsetse species show any signs of disease from these viruses. Full article

Trypanosomes are endemic and retard cattle health in Shimba Hills, Kenya. Wildlife in the area act as reservoirs of the parasites. However, wild animal species that harbor and expose cattle to tsetse-borne trypanosomes are not well known in Shimba Hills. Using xeno-monitoring surveillance to investigate wild animal reservoirs and sources of trypanosomes in Shimba Hills, we screened 696 trypanosome-infected and uninfected tsetse flies for vertebrate DNA using multiple-gene PCR-High Resolution Melting analysis and amplicon sequencing. Results revealed that tsetse flies fed on 13 mammalian species, preferentially Phacochoerus africanus (warthogs) (17.39%, 95% CI: 14.56–20.21) and Bos taurus (cattle) (11.35%, 95% CI: 8.99–13.71). Some tsetse flies showed positive cases of bloodmeals from multiple hosts (3.45%, 95% CI: 2.09–4.81), including warthog and cattle (0.57%, 95% CI: 0.01–1.14). Importantly, tsetse flies that took bloodmeals from warthog had significant risk of infections with Trypanosoma vivax (5.79%, 95% CI: 1.57–10.00), T. congolense (7.44%, 95% CI: 2.70–12.18), and T. brucei sl (2.48%, 95% CI: −0.33–5.29). These findings implicate warthogs as important reservoirs of tsetse-borne trypanosomes affecting cattle in Shimba Hills and provide valuable epidemiological insights to underpin the parasites targeted management in Nagana vector control programs in the area. Full article