Non-lethal Inhibition of Gut Microbial Trimethylamine Production for the Treatment of Atherosclerosis.

Trimethylamine (TMA) N-oxide (TMAO), a gut-microbiota-dependent metabolite, both enhances atherosclerosis in animal models and is associated with cardiovascular risks in clinical studies. Here, we investigate the impact of targeted inhibition of the first step in TMAO generation, commensal microbial TMA production, on diet-induced atherosclerosis. A structural analog of choline, 3,3-dimethyl-1-butanol (DMB), is shown to non-lethally inhibit TMA formation from cultured microbes, to inhibit distinct microbial TMA lyases, and to both inhibit TMA production from physiologic polymicrobial cultures (e.g., intestinal contents, human feces) and reduce TMAO levels in mice fed a high-choline or L-carnitine diet. DMB inhibited choline diet-enhanced endogenous macrophage foam cell formation and atherosclerotic lesion development in apolipoprotein e(-/-) mice without alterations in circulating cholesterol levels. The present studies suggest that targeting gut microbial production of TMA specifically and non-lethal microbial inhibitors in general may serve as a potential therapeutic approach for the treatment of cardiometabolic diseases.

Systematic identification of genes encoding cell surface and secreted proteins that are essential for in vitro growth and infection in Leishmania donovani.

Leishmaniasis is an infectious disease caused by protozoan parasites belonging to the genus Leishmania for which there are no approved human vaccines. Infections localise to different tissues in a species-specific manner with the visceral form of the disease caused by Leishmania donovani and L. infantum being the most deadly in humans. Although Leishmania spp. parasites are predominantly intracellular, the visceral disease can be prevented in dogs by vaccinating with a complex mixture of secreted products from cultures of L. infantum promastigotes. With the logic that extracellular parasite proteins make good subunit vaccine candidates because they are directly accessible to vaccine-elicited host antibodies, here we attempt to discover proteins that are essential for in vitro growth and host infection with the goal of identifying subunit vaccine candidates. Using an in silico analysis of the Leishmania donovani genome, we identified 92 genes encoding proteins that are predicted to be secreted or externally anchored to the parasite membrane by a single transmembrane region or a GPI anchor. By selecting a transgenic L. donovani parasite that expresses both luciferase and the Cas9 nuclease, we systematically attempted to target all 92 genes by CRISPR genome editing and identified four that were required for in vitro growth. For fifty-five genes, we infected cohorts of mice with each mutant parasite and by longitudinally quantifying parasitaemia with bioluminescent imaging, showed that nine genes had evidence of an attenuated infection although all ultimately established an infection. Finally, we expressed two genes as full-length soluble recombinant proteins and tested them as subunit vaccine candidates in a murine preclinical infection model. Both proteins elicited significant levels of protection against the uncontrolled development of a splenic infection warranting further investigation as subunit vaccine candidates against this deadly infectious tropical disease.

Spatial and Temporal Detection of Ions Ejected from Coulomb Crystals.

Coulomb crystals have proven to be powerful and versatile tools for the study of ion-molecule reactions under cold and controlled conditions. Reactions in Coulomb crystals are typically monitored through a combination of in situ fluorescence imaging of the laser-cooled ions and destructive time-of-flight mass spectrometry measurements of the ejected ions. However, neither of these techniques is able to provide direct structural information on the positions of nonfluorescing "dark" ions within the crystal. In this work, structural information is obtained using a phosphor screen and a microchannel plate detector in conjunction with a Timepix3 camera. The Timepix3 camera simultaneously records the spatial and temporal distribution of all ions that strike the phosphor screen detector following crystal ejection at a selected reaction time. A direct comparison can be made between the observed Timepix3 ion distributions and the distributions established from SIMION simulations of the ion trajectories through the apparatus and onto the detector. Quantitative agreement is found between the measured Timepix3 signal and the properties of Coulomb crystals assigned using fluorescence imaging─independently confirming that the positions and numbers of nonfluorescing ions within Coulomb crystals can be accurately determined using molecular dynamics simulations. It is anticipated that the combination of high-resolution spatial and temporal data will facilitate new measurements of the ion properties within Coulomb crystals.

Fluoroquinolone resistance among fecal extended spectrum ßeta lactamases positive Enterobacterales isolates from children in Dar es Salaam, Tanzania.

BACKGROUND: Fluoroquinolones have been, and continue to be, routinely used for treatment of many bacterial infections. In recent years, most parts of the world have reported an increasing trend of fluoroquinolone resistant (FQR) Gram-negative bacteria. METHODS: A cross-sectional study was conducted between March 2017 and July 2018 among children admitted due to fever to referral hospitals in Dar es Salaam, Tanzania. Rectal swabs were used to screen for carriage of extended-spectrum ß-lactamase-producing Enterobacterales (ESBL-PE). ESBL-PE isolates were tested for quinolone resistance by disk diffusion method. Randomly selected fluroquinolone resistant isolates were characterized by using whole genome sequencing. RESULTS: A total of 142 ESBL-PE archived isolates were tested for fluoroquinolone resistance. Overall phenotypic resistance to ciprofloxacin, levofloxacin and moxifloxacin was found in 68% (97/142). The highest resistance rate was seen among Citrobacter spp. (100%, 5/5), followed by Klebsiella. pneumoniae (76.1%; 35/46), Escherichia coli (65.6%; 42/64) and Enterobacter spp. (31.9%; 15/47). Whole genome sequencing (WGS) was performed on 42 fluoroquinolone resistant-ESBL producing isolates and revealed that 38/42; or 90.5%, of the isolates carried one or more plasmid mediated quinolone resistance (PMQR) genes. The most frequent PMQR genes were aac(6')-lb-cr (74%; 31/42), followed by qnrB1 (40%; 17/42), oqx, qnrB6 and qnS1. Chromosomal mutations in gyrA, parC and parE were detected among 19/42 isolates, and all were in E. coli. Most of the E. coli isolates (17/20) had high MIC values of > 32 µg/ml for fluoroquinolones. In these strains, multiple chromosomal mutations were detected, and all except three strains had additional PMQR genes. Sequence types, ST131 and ST617 predominated among E. coli isolates, while ST607 was more common out of 12 sequence types detected among the K. pneumoniae. Fluoroquinolone resistance genes were mostly associated with the IncF plasmids. CONCLUSION: The ESBL-PE isolates showed high rates of phenotypic resistance towards fluoroquinolones likely mediated by both chromosomal mutations and PMQR genes. Chromosomal mutations with or without the presence of PMQR were associated with high MIC values in these bacteria strains. We also found a diversity of PMQR genes, sequence types, virulence genes, and plasmid located antimicrobial resistance (AMR) genes towards other antimicrobial agents.

Bilingual children's visual attention while reading digital picture books and story retelling.

This study examined Mandarin-English bilingual children's visual attention over repetitive readings of Mandarin enhanced digital books and static books as well as the effects visual attention has on story retelling. We assigned 89 4- and 5-year-old preschoolers in Singapore to one of three reading conditions: (a) digital books with visual and auditory enhancements, (b) digital books with only auditory enhancements, and (c) static digital books with neither visual nor auditory enhancements. We presented three stories to the children in four sessions over 2 weeks, traced their visual attention with an eye tracker, and examined their story retelling after the first and fourth readings. The results demonstrated that the digital books with visual and auditory enhancements maintained greater visual attention from children compared with that from children in the other two conditions across the four repetitive readings. Moreover, children's bilingual language proficiency significantly modulates the conditional effects of attention. Children with higher bilingual proficiency in the visual and auditory enhancements condition outperformed their peers in the other two conditions in terms of visual attention across most readings. However, for the children with lower bilingual proficiency, the digital books with auditory and visual enhancements only outperformed the static condition but not the auditory enhanced condition. Children with lower language proficiency maintained their attention at a relatively high level across the repetitive readings in the enhanced digital book conditions but demonstrated significantly decreased visual attention in the static digital book condition. Because children with better visual attention and higher bilingual proficiency retold the stories significantly better, the results indicate that influencing visual attention helps to improve story comprehension.

AWZ1066S, a highly specific anti-Wolbachia drug candidate for a short-course treatment of filariasis.

Onchocerciasis and lymphatic filariasis are two neglected tropical diseases that together affect ∼157 million people and inflict severe disability. Both diseases are caused by parasitic filarial nematodes with elimination efforts constrained by the lack of a safe drug that can kill the adult filaria (macrofilaricide). Previous proof-of-concept human trials have demonstrated that depleting >90% of the essential nematode endosymbiont bacterium, Wolbachia, using antibiotics, can lead to permanent sterilization of adult female parasites and a safe macrofilaricidal outcome. AWZ1066S is a highly specific anti-Wolbachia candidate selected through a lead optimization program focused on balancing efficacy, safety and drug metabolism/pharmacokinetic (DMPK) features of a thienopyrimidine/quinazoline scaffold derived from phenotypic screening. AWZ1066S shows superior efficacy to existing anti-Wolbachia therapies in validated preclinical models of infection and has DMPK characteristics that are compatible with a short therapeutic regimen of 7 days or less. This candidate molecule is well-positioned for onward development and has the potential to make a significant impact on communities affected by filariasis.

The Relationship Between Strength Measures and Task Performance in Specialist Tactical Police.

ABSTRACT: Orr, RM, Robinson, J, Hasanki, K, Talaber, KA, Schram, B, and Roberts, A. The relationship between strength measures and task performance in specialist tactical police. J Strength Cond Res 36(3): 757-762, 2022-Specialist tactical police officers (STPOs) carry heavier on-body loads than generalist police officers. Improvements in strength may mitigate the impacts of these heavier loads. The aim of this investigation was to determine the correlations between absolute and relative strength measures and occupational task performance in STPOs. Retrospective data were provided for 47 male specialist police officers from an elite Australian police unit. Data included body mass (mean = 89.0 ± 8.58 kg), strength measures (1 repetition maximum measures for a bench press, squat, deadlift, and pull-up), and task performance measures (85-kg victim drag wearing 15 kg of operational load and 5-km pack march wearing 40 kg of operational load). Pearson's correlations were conducted to determine relationships between measures and were plotted on a linear regressions model. Significant, moderate to strong correlations were found between all strength measures and victim drag performance and significant negative moderate correlations between relative bench press, absolute and relative squat, and absolute and relative pull-up and pack march times. The absolute deadlift had the strongest correlation to the victim drag (r = 0.747, p < 0.01) while the relative pull-up showed the strongest correlation with pack march performance (r = -0.466, p < 0.01). The requirement to lift a portion of the dummy off the ground during the victim drag may explain the increased importance of absolute strength while the requirement to transport load affixed to the body may explain the importance of relative strength requirements. Improvements in absolute and relative upper- and lower-body strength may improve task performance in this population.

Quasi-cyclical preferences in the ethics of Plato, Aristotle, and Kant.

Bermúdez describes the extensionality principle as being "almost unquestioned." This claim might come as a surprise to philosophers who work on agency and ethics. In Kantian deontological ethics and in Platonic or Aristotelian virtue ethics, our preferences for outcomes can be rationally affected by how those outcomes are framed in terms of maxims and character traits.

Bengali nasal vowels: lexical representation and listener perception.

This paper focuses on the question of the representation of nasality as well as speakers' awareness and perceptual use of phonetic nasalisation by examining surface nasalisation in two types of vowels in Bengali: underlying nasal vowels (CVC) and nasalised vowels before a nasal consonant (CVN). A series of three cross-modal forced-choice experiments was used to investigate the hypothesis that only unpredictable nasalisation is stored and that this sparse representation governs how listeners interpret vowel nasality. Visual full-word targets were preceded by auditory primes consisting of CV segments of CVC words with nasal vowels ([tʃɑ̃] for [tʃɑ̃d] 'moon'), oral vowels ([tʃɑ] for [tʃɑl] 'unboiled rice') or nasalised oral vowels ([tʃɑ̃(n)] for [tʃɑ̃n] 'bath') and reaction times and errors were measured. Some targets fully matched the prime while some matched surface or underlying representation only. Faster reaction times and fewer errors were observed after CVC primes compared to both CVC and CVN primes. Furthermore, any surface nasality was most frequently matched to a CVC target unless no such target was available. Both reaction times and error data indicate that nasal vowels are specified for nasality leading to faster recognition compared to underspecified oral vowels, which cannot be perfectly matched with incoming signals.

A novel hemA mutation is responsible for a small-colony-variant phenotype in Escherichia coli.

We identified a small colony variant (SCV) of an amoxicillin/clavulanic acid-resistant derivative of a clinical isolate of Escherichia coli from Malawi, which was selected for in vitro in a subinhibitory concentration of gentamicin. The SCV was auxotrophic for hemin and had impaired biofilm formation compared to the ancestral isolates. A single novel nucleotide polymorphism (SNP) in hemA, which encodes a glutamyl-tRNA reductase that catalyses the initial step of porphyrin biosynthesis leading to the production of haem, was responsible for the SCV phenotype. We showed the SNP in hemA resulted in a significant fitness cost to the isolate, which persisted even in the presence of hemin. However, the phenotype quickly reverted during sequential sub-culturing in liquid growth media. As hemA is not found in mammalian cells, and disruption of the gene results in a significant fitness cost, it represents a potential target for novel drug development specifically for the treatment of catheter-associated urinary tract infections caused by E. coli.

Detection of mobile genetic elements associated with antibiotic resistance in Salmonella enterica using a newly developed web tool: MobileElementFinder.

OBJECTIVES: Antimicrobial resistance (AMR) in clinically relevant bacteria is a growing threat to public health globally. In these bacteria, antimicrobial resistance genes are often associated with mobile genetic elements (MGEs), which promote their mobility, enabling them to rapidly spread throughout a bacterial community. METHODS: The tool MobileElementFinder was developed to enable rapid detection of MGEs and their genetic context in assembled sequence data. MGEs are detected based on sequence similarity to a database of 4452 known elements augmented with annotation of resistance genes, virulence factors and detection of plasmids. RESULTS: MobileElementFinder was applied to analyse the mobilome of 1725 sequenced Salmonella enterica isolates of animal origin from Denmark, Germany and the USA. We found that the MGEs were seemingly conserved according to multilocus ST and not restricted to either the host or the country of origin. Moreover, we identified putative translocatable units for specific aminoglycoside, sulphonamide and tetracycline genes. Several putative composite transposons were predicted that could mobilize, among others, AMR, metal resistance and phosphodiesterase genes associated with macrophage survivability. This is, to our knowledge, the first time the phosphodiesterase-like pdeL has been found to be potentially mobilized into S. enterica. CONCLUSIONS: MobileElementFinder is a powerful tool to study the epidemiology of MGEs in a large number of genome sequences and to determine the potential for genomic plasticity of bacteria. This web service provides a convenient method of detecting MGEs in assembled sequence data. MobileElementFinder can be accessed at https://cge.cbs.dtu.dk/services/MobileElementFinder/.

Targeted Inhibition of Gut Microbial Trimethylamine N-Oxide Production Reduces Renal Tubulointerstitial Fibrosis and Functional Impairment in a Murine Model of Chronic Kidney Disease.

OBJECTIVE: Gut microbial metabolism of dietary choline, a nutrient abundant in a Western diet, produces trimethylamine (TMA) and the atherothrombosis- and fibrosis-promoting metabolite TMA-N-oxide (TMAO). Recent clinical and animal studies reveal that elevated TMAO levels are associated with heightened risks for both cardiovascular disease and incident chronic kidney disease development. Despite this, studies focusing on therapeutically targeting gut microbiota-dependent TMAO production and its impact on preserving renal function are limited. Approach and Results: Herein we examined the impact of pharmacological inhibition of choline diet-induced gut microbiota-dependent production of TMA, and consequently TMAO, on renal tubulointerstitial fibrosis and functional impairment in a model of chronic kidney disease. Initial studies with a gut microbial choline TMA-lyase mechanism-based inhibitor, iodomethylcholine, confirmed both marked suppression of TMA generation, and consequently TMAO levels, and selective targeting of the gut microbial compartment (ie, both accumulation of the drug in intestinal microbes and limited systemic exposure in the host). Dietary supplementation of either choline or TMAO significantly augmented multiple indices of renal functional impairment and fibrosis associated with chronic subcutaneous infusion of isoproterenol. However, the presence of the gut microbiota-targeting inhibitor iodomethylcholine blocked choline diet-induced elevation in TMAO, and both significantly improved decline in renal function, and significantly attenuated multiple indices of tubulointerstitial fibrosis. Iodomethylcholine treatment also reversed many choline diet-induced changes in cecal microbial community composition associated with TMAO and renal functional impairment. CONCLUSIONS: Selective targeting of gut microbiota-dependent TMAO generation may prevent adverse renal structural and functional alterations in subjects at risk for chronic kidney disease.

Synthesis and biological evaluation of pentacyclic triterpenoid derivatives as potential novel antibacterial agents.

A series of ursolic acid (UA), oleanolic acid (OA) and 18ß-glycyrrhetinic acid (GA) derivatives were synthesized by introducing a range of substituted aromatic side-chains at the C-2 position after the hydroxyl group at C-3 position was oxidized. Their antibacterial activities were evaluated in vitro against a panel of four Staphylococcus spp. The results revealed that the introduction of aromatic side-chains at the C-2 position of GA led to the discovery of potent triterpenoid derivatives for inhibition of both drug sensitive and resistant S. aureus, while the other two series derivatives of UA and OA showed no significant antibacterial activity even at high concentrations. In particular, GA derivative 33 showed good potency against all four Staphylococcus spp. (MIC = 1.25-5 µmol/L) with acceptable pharmacokinetics properties and low cytotoxicity in vitro. Molecular docking was also performed using S. aureus DNA gyrase to rationalize the observed antibacterial activity. This series of GA derivatives has strong potential for the development of a new type of triterpenoid antibacterial agent.

The importance of air quality for underground spaces: An international survey of public attitudes.

Space is a resource that is constantly being depleted, especially in mega-cities. Underground workspaces (UGS) are increasingly being included in urban plans and have emerged as an essential component of vertical cities. While progress had been made on the engineering aspects associated with the development of high-quality UGS, public attitudes toward UGS as work environments (ie, the public's design concerns with UGS) are relatively unknown. Here, we present the first large-scale study examining preferences and attitudes toward UGS, surveying close to 2000 participants from four cities in three continents (Singapore, Shanghai, London, and Montreal). Contrary to previous beliefs, air quality (and not lack of windows) is the major concern of prospective occupants. Windows, temperature, and lighting emerged as additional important building performance aspects for UGS. Early adopters (ie, individuals more willing to accept UGS and thus more likely to be the first occupants) across all cities prioritized air quality. Present results suggest that (perceived) air quality is a key building performance aspect for UGS that needs to be communicated to prospective occupants as this will improve their attitudes and views toward UGS. This study highlights the importance of indoor air quality for the public.

A safety and pharmacodynamics study of temelimab, an antipathogenic human endogenous retrovirus type W envelope monoclonal antibody, in patients with type 1 diabetes.

AIM: To report the first study of temelimab, a monoclonal antibody neutralizing the pathogenic human endogenous retrovirus type W envelope, in patients with type 1 diabetes (T1D). MATERIALS AND METHODS: This double-blind, placebo-controlled, randomized clinical trial recruited adult patients with T1D within 4 years postdiagnosis and remaining C-peptide secretion. Sixty-four patients were randomized (2:1) to monthly temelimab 6 mg/kg or placebo during 24 weeks followed by a 24-week, open-label extension, during which all patients received temelimab. The primary objective was the safety and tolerability of temelimab. The secondary objective was to assess the pharmacodynamics response such as C-peptide levels, insulin use, HbA1c, hypoglycaemia and autoantibodies. RESULTS: Temelimab was well tolerated without any group difference in the frequency or severity of adverse events. Concerning exploratory endpoints, there was no difference in the levels of C-peptide, insulin use or HbA1c between treatment groups at weeks 24 and 48. The frequency of hypoglycaemia events was reduced with temelimab (P = 0.0004) at week 24 and the level of anti-insulin antibodies was lower with temelimab (P < 0.01); the other autoantibodies did not differ between groups. CONCLUSIONS: Temelimab appeared safe in patients with T1D. Pharmacodynamics signals (hypoglycaemia and anti-insulin antibodies) under temelimab were observed. Markers of ß-cell functions were not modified by treatment. These results need to be further explored in younger patients with T1D with earlier disease onset.

Associations of perceived indoor environmental quality with stress in the workplace.

Indoor environmental quality (IEQ) is a general indicator of the quality of conditions inside a building. We investigated associations of perceived IEQ including air quality, thermal comfort, noise, and light quality with stress at work and the extent to which workplace location modifies these associations. We recruited 464 full-time workers from four companies in Singapore. Data on socio-demographic characteristics, lifestyle/health-related factors, and workplace factors were collected through self-administered questionnaires. Perceived IEQ satisfaction scores of all four factors were collected using the validated OFFICAIR questionnaire. We fitted a logistic regression model to assess associations between each perceived IEQ score and stress at work, adjusting for potential confounders. The odds ratio for stress at work associated with a 1-unit increase in perceived air quality score was 0.88 (0.82-0.94), 0.89 (0.82-0.97) for thermal comfort, 0.93 (0.87-0.98) for noise, and 0.88 (0.82-0.94) for light quality. Significant associations were found in office and control rooms for all four perceived IEQ, except for thermal comfort in office rooms. Higher satisfaction levels of perceived air quality, thermal comfort, noise, and lighting, were significantly associated with a reduction in stress at work. Our findings could potentially provide a useful tool for environmental health impact assessment for buildings.

Employee experiences in underground workplaces: a qualitative investigation.

Underground workplaces (UWS) are increasingly being recognised as a sustainable solution to concerns regarding lack of space facing megacities. UWS can multiply available space by reducing urban sprawl thereby improving quality of life in cities. However, developing workplaces that respond to the needs and preferences of occupants will be a crucial factor in determining the success of this concept. We examine the experiences of existing full-time employees of a large underground facility in North America and analyse how these are related to company policies and other organisational factors. From in-depth interviews of 73 participants, three predominant themes emerged from the analyses: environmental effects (e.g. lack of sunlight), design (e.g. the unique working environment), and organisational factors and social dynamics (e.g. policies regarding breaks). We discuss these themes in relation with interventions that may improve employee comfort and retention and suggest that many factors are within organisational control. Practitioner Summary: Underground workplaces are a sustainable solution to lack of urban space. However, developing effective workplaces is crucial to their success. We examine the experiences of existing employees of an underground facility in North America. We suggest that most negative attitudes towards underground spaces can be mitigated through simple organisational interventions. Abbreviations: UWS: underground workplaces; IRB: institutional review board.

Pneumococcal Colonization in Healthy Adult Research Participants in the Conjugate Vaccine Era, United Kingdom, 2010-2017.

Pneumococcal colonization is rarely studied in adults, except as part of family surveys. We report the outcomes of colonization screening in healthy adults (all were nonsmokers without major comorbidities or contact with children aged <5 years) who had volunteered to take part in clinical research. Using nasal wash culture, we detected colonization in 6.5% of volunteers (52 of 795). Serotype 3 was the commonest serotype (10 of 52 isolates). The majority of the remaining serotypes (35 of 52 isolates) were nonvaccine serotypes, but we also identified persistent circulation of serotypes 19A and 19F. Resistance to at least 1 of 6 antibiotics tested was found in 8 of 52 isolates.

The shufflon of IncI1 plasmids is rearranged constantly during different growth conditions.

One of the factors that can affect conjugation of IncI1 plasmids, amongst others, is the genetic region known as the shufflon. This multiple inversion system modifies the pilus tip proteins used during conjugation, thus affecting the affinity for different recipient cells. Although recombination is known to occur in in vitro conditions, little is known about the regulation and the extent of recombination that occurs. To measure the recombination of the shufflon, we have amplified the entire shufflon region and sequenced the amplicons using nanopore long-read sequencing. This method was effective to determine the order of the segments of the shufflon and allow for the analysis of the shufflon variants that are present in a heterogeneous pool of templates. Analysis was performed over different growth phases and after addition of cefotaxime. Furthermore, analysis was performed in different E. coli host cells to determine if recombination is likely to be influenced. Recombination of the shufflon was constantly ongoing in all conditions that were measured, although no differences in the amount of different shufflon variants or the rate at which novel variants were formed could be found. As previously reported, some variants were abundant in the population while others were scarce. This leads to the conclusion that the shufflon is continuously recombining at a constant rate, or that the method used here was not sensitive enough to detect differences in this rate. For one of the plasmids, the host cell appeared to have an effect on the specific shufflon variants that were formed which were not predominant in another host, indicating that host factors may be involved. As previously reported, the pilV-A and pilV-A' ORFs are formed at higher frequencies than other pilV ORFs. These results demonstrate that the recombination that occurs within the shufflon is not random. While any regulation of the shufflon affected by these in vitro conditions could not be revealed, the method of amplifying large regions for long-read sequencing for the analysis of multiple inversion systems proved effective.

A role for trypanosomatid aldo-keto reductases in methylglyoxal, prostaglandin and isoprostane metabolism.

Trypanosomatid parasites are the infectious agents causing Chagas disease, visceral and cutaneous leishmaniasis and human African trypanosomiasis. Recent work of others has implicated an aldo-keto reductase (AKR) in the susceptibility and resistance of Trypanosoma cruzi to benznidazole, a drug used to treat Chagas disease. Here, we show that TcAKR and homologues in the related parasites Trypanosoma brucei and Leishmania donovani do not reductively activate monocyclic (benznidazole, nifurtimox and fexinidazole) or bicyclic nitro-drugs such as PA-824. Rather, these enzymes metabolise a variety of toxic ketoaldehydes, such as glyoxal and methylglyoxal, suggesting a role in cellular defence against chemical stress. UPLC-QToF/MS analysis of benznidazole bioactivation by T. cruzi cell lysates confirms previous reports identifying numerous drug metabolites, including a dihydro-dihydroxy intermediate that can dissociate to form N-benzyl-2-guanidinoacetamide and glyoxal, a toxic DNA-glycating and cross-linking agent. Thus, we propose that TcAKR contributes to benznidazole resistance by the removal of toxic glyoxal. In addition, three of the four enzymes studied here display activity as prostaglandin F2α synthases, despite the fact that there are no credible cyclooxygenases in these parasites to account for formation of the precursor PGH2 from arachidonic acid. Our studies suggest that arachidonic acid is first converted non-enzymatically in parasite lysates to (PGH2-like) regioisomers by free radical-mediated peroxidation and that AKRs convert these lipid peroxides into isoprostanes, including prostaglandin F2α and 8-iso-prostaglandin F2α.