ABSTRACT
Antimicrobial resistance presents a substantial threat to global public health, demanding urgent attention and action. This study focuses on lanthipeptides, ribosomally encoded peptides that display significant structural diversity and hold promising potential as antibiotics. Genome mining was employed to locate biosynthetic gene clusters (BGCs) containing class II lanthipeptide synthetases encoded by lanM genes. A phylogenetic study analyzing homologous sequences of functional LanM sequences revealed a unique evolutionary clade of 17 LanM proteins associated with 12 Clostridium bacterial genomes. In silico exploration identified nine complete BGCs, including one super-cluster containing two co-localized operons from Clostridium cellulovorans 743B, that encode for two new peptides named clostrisin and cellulosin. Each operon was heterologously expressed in Escherichia coli. Molecular weights associated with the expected post-translational modifications of the purified lanthipeptide were confirmed by MS-MS/MS analysis for cellulosin, while clostrisin was not post-translationally modified. Both peptides demonstrated antimicrobial activity against multidrug-resistant bacteria, such as a clinical strain of Staphylococcus epidermidis MIQ43 and Pseudomonas aeruginosa PA14. This is the first report of lanthipeptides from the Clostridium genus produced with its native biosynthetic machinery, as well as chemically and biologically characterized. This study showcases the immense potential of genome mining in identifying new RiPP synthetases and associated bioactive peptides.
ABSTRACT
It is estimated that 6-7 million people worldwide are infected with Trypanosoma cruzi, the parasite that causes Chagas disease. In Venezuela, Chagas disease remains a public health problem. In this work, T. cruzi isolates from six species of triatomines and mammals of the orders Didelphimorphia and Xenarthra, captured in rural communities of Monagas, underwent parasitological and molecular characterization. A total of 471 triatomines and 17 mammals were captured, with a natural infection rate of 41.4% and 70.6%, respectively. In the male NMRI mouse model used for parasitological characterization (prepatent period, parasitemia curve, mouse mortality, and tissular parasitism), T. cruzi isolates exhibited high lethality due to their pronounced virulence, irrespective of the parasite load in each mouse, resulting in a mortality rate of 75%. Among the vector isolates, in the mouse model, only 2 out of 6 remained alive, while the rest perished during the evaluation. Conversely, the isolates from mammals proved fatal for all the inoculated mice. All isolates were identified as belonging to DTU TcI, based on the molecular markers such as the intergenic region of the miniexon, D7 divergent domain of the 24Sα rDNA, size-variable domain of the 18S rDNA, and hsp60-PCR-RFLP-EcoRV. This study demonstrates the presence of vectors and mammalian reservoirs naturally infected with T. cruzi in communities of Monagas, the 9th largest and 9th most populous state in Venezuela. This situation represents a neglected epidemiological problem demanding urgent attention and imperative health care intervention.
Subject(s)
Chagas Disease , Marsupialia , Trypanosoma cruzi , Animals , Male , Humans , Mice , Venezuela/epidemiology , Chagas Disease/epidemiology , Chagas Disease/parasitology , Mammals/parasitology , DNA, RibosomalABSTRACT
Methicillin-resistant Staphylococcus aureus (MRSA) increases its antibiotic resistance by forming biofilms. Natural products (NP) or specialized metabolites have demonstrated their ability to decrease the virulence and pathogenesis of MRSA infections by inhibiting biofilm formation. The present study evaluated the antimicrobial and antibiofilm potential against MRSA of a small library of fungal NP isolated from Mexican biodiversity. The most potent antibacterial activity was observed for myrotecisin B, epiequisetin, equisetin, stachybotrolide acetate, monorden A, zearalenone, fuscin, and fusarubin. On the other hand, epifiscalin C, fiscalin C, dimethylglyotoxin, aspernolide B, and butyrolactones I and IV inhibited the biofilm formation without decreasing bacterial growth. To determine the putative mechanism of action of these compounds, docking analyses were performed against SarA and AgrA proteins, targets known to regulate biofilm production in MRSA. Overall, the results demonstrate that fungal NP may act as potential antibiofilm agents for treating MRSA infections.
Subject(s)
Methicillin-Resistant Staphylococcus aureus , Biofilms , Anti-Bacterial Agents/pharmacology , Virulence , Microbial Sensitivity TestsABSTRACT
The bean fruit pericarp accumulates a significant amount of starch, which starts to be degraded 20 days after anthesis (DAA) when seed growth becomes exponential. This period is also characterized by the progressive senescence of the fruit pericarp. However, the chloroplasts maintained their integrity, indicating that starch degradation is a compartmentalized process. The process coincided with a transient increase in maltose and sucrose levels, suggesting that ß-amylase is responsible for starch degradation. Starch degradation in the bean fruit pericarp is also characterized by a large increase in starch phosphorylation, as well as in the activities of cytosolic disproportionating enzyme 2 (DPE2, EC 2.4.1.25) and glucan phosphorylase (PHO2, EC 2.4.1.1). This suggests that the rate of starch degradation in the bean fruit pericarp 20 DAA is dependent on the transformation of starch to a better substrate for ß-amylase and the increase in the rate of cytosolic metabolism of maltose.
Subject(s)
Arabidopsis , beta-Amylase , Maltose/metabolism , Fruit/metabolism , beta-Amylase/metabolism , Arabidopsis/metabolism , Starch/metabolismABSTRACT
We evaluated the microbial diversity and metabolome profile of an uncommon hypersaline elastic microbial mat from Cuatro Ciénegas Basin (CCB) in the Chihuahuan Desert of Coahuila, México. We collected ten samples on a small scale transect (1.5-m) and described its microbial diversity through NGS-based ITS and 16S rDNA gene sequencing. A very low number of taxa comprised a considerable proportion of the mat and were shared across all sampling points, whereas the rare biosphere was more phylogenetically diverse (Faith's Phylogenetic Diversity (FPD) index) and phylogenetically disperse (using a null model distribution of Phylogenetic Species Clustering (nmdPSC)) than the abundant (high read count) taxa for both analyzed libraries. We also found a distinctive metabolome profile for each sample and were able to tentatively annotate several classes of compounds with relevant biological properties.
Subject(s)
Environment , Phylogeny , DNA, Ribosomal , MexicoABSTRACT
Cenotes are habitats with unique physical, chemical, and biological features. Unexplored microorganisms from these sinkholes represent a potential source of bioactive molecules. Thus, a series of cultivable fungi (Aspergillus spp. NCA257, NCA264, and NCA276, Stachybotrys sp. NCA252, and Cladosporium sp. NCA273) isolated from the cenote Tza Itzá were subjected to chemical, coculture, and metabolomic analyses. Nineteen compounds were obtained and tested for their antimicrobial potential against ESKAPE pathogens, Mycobacterium tuberculosis, and nontuberculous mycobacteria. In particular, phenylspirodrimanes from Stachybotrys sp. NCA252 showed significant activity against MRSA, MSSA, and mycobacterial strains. On the other hand, the absolute configuration of the new compound 17-deoxy-aspergillin PZ (1) isolated from Aspergillus sp. NCA276 was established via single-crystal X-ray crystallography. Also, the chemical analysis of the cocultures between Aspergillus and Cladosporium strains revealed the production of metabolites that were not present or were barely detected in the monocultures. Finally, molecular networking analysis of the LC-MS-MS/MS data for each fungus was used as a tool for the annotation of additional compounds, increasing the chemical knowledge on the corresponding fungal strains. Overall, this is the first systematic chemical study on fungi isolated from a sinkhole in Mexico.
ABSTRACT
Chemical investigation of Punctularia atropurpurascens strain HM1 (Punctulariaceae), a corticioid isolated from a decorticated piece of Quercus bark collected in Bosque de Tlalpan, Mexico City, led to the isolation of a new drimane, 1-α-hydroxy-isodrimenine (1: ) and a new tetrahydroxy kauranol, 16-hydroxy-phlebia-nor-kauranol (2: ), together with the known N-phenylacetamide (3: ). Structures of all compounds were elucidated by spectroscopic and spectrometric methods, and the absolute configuration of 1: and 2: was confirmed via single-crystal X-ray crystallography. The isolated compounds showed modest antimycobacterial activity.
Subject(s)
Basidiomycota , Terpenes , Anti-Bacterial Agents/pharmacology , Crystallography, X-Ray , Fungi , Molecular Structure , Terpenes/pharmacologyABSTRACT
Barley malting depends on hydrolytic enzymes that degrade storage macromolecules. Identifying barley cultivars with proteolytic activity that guarantees appropriate foaming, flavor, and aroma in the beer is of great importance. In this work, the proteolytic activity and profiles of brewing malt from Mexican barley cultivars were analyzed. Data showed that Cys- (at 50°C) and Ser-proteases (at 70°C) are the major contributors to proteolytic activity during mashing. Essential amino acids, necessary for fermentation and production of good flavor and aroma in beer, were detected at the end of mashing. According to our results, Mexican cultivar HV2005-19 exhibits similar proteolytic activities as those from cultivar Metcalfe, which is one of the most utilized for the brewing industry. Moreover, we propose Cys- and Ser-proteases as biochemical markers during mashing at 50 and 70°C, respectively, to select barley cultivars for beer production. PRACTICAL APPLICATIONS: Proteolytic activity, which depends on activation and de novo synthesis of proteases in the aleurone layer of barley seeds, is crucial in beer production. Identifying new barley varieties that have optimal proteolytic activities is of great interest for genetic improvement programs. In this study, we propose the variety HV2005-19 as a genotype with Cys- and Ser-proteases activity similar to that from Metcalfe, which is a top variety in the brewing industry.
Subject(s)
Hordeum , Beer/analysis , Fermentation , Hordeum/chemistry , Hordeum/genetics , Peptide Hydrolases/genetics , Seeds/chemistryABSTRACT
A collection of 29 cultivable fungal strains isolated from deep-sea sediments of the Gulf of Mexico were cultivated under the "one strain, many compounds" approach to explore their chemical diversity and antimicrobial potential. From the 87 extracts tested, over 50% showed antimicrobial activity, and the most active ones were those from cultures grown at 4 °C in darkness for 60 days (resembling deep-sea temperature). PCA analysis of the LC-MS data of all the extracts confirmed that culture temperature is the primary factor in the variation of the 4462 metabolite features, accounting for 21.3% of the variation. The bioactivity-guided and conventional chemical studies of selected fungal strains allowed the identification of several active and specialized metabolites. Finally, metabolomics analysis by GNPS molecular networking and manual dereplication revealed the biosynthetic potential of these species to produce interesting chemistry. This work uncovers the chemical and biological study of marine-derived fungal strains from deep-sea sediments of the Gulf of Mexico.
Subject(s)
Anti-Infective Agents/chemistry , Fungi/chemistry , Anti-Infective Agents/metabolism , Anti-Infective Agents/pharmacology , Biological Products/chemistry , Biological Products/metabolism , Biological Products/pharmacology , Fungi/metabolism , Geologic Sediments/microbiology , Gulf of Mexico , MetabolomeABSTRACT
Mangrove sediment ecosystems in the coastal areas of the Yucatan peninsula are unique environments, influenced by their karstic origin and connection with the world's largest underground river. The microbial communities residing in these sediments are influenced by the presence of mangrove roots and the trading chemistry for communication between sediment bacteria and plant roots can be targeted for secondary metabolite research. To explore the secondary metabolite production potential of microbial community members in mangrove sediments at the "El Palmar" natural reserve in Sisal, Yucatan, a combined meta-omics approach was applied. The effects of a cultivation medium reported to select for actinomycetes within mangrove sediments' microbial communities was also analyzed. The metabolome of the microbial communities was analyzed by high-resolution liquid chromatography-tandem mass spectrometry, and molecular networking analysis was used to investigate if known natural products and their variants were present. Metagenomic results suggest that the sediments from "El Palmar" harbor a stable bacterial community independently of their distance from mangrove tree roots. An unexpected decrease in the observed abundance of actinomycetes present in the communities occurred when an antibiotic-amended medium considered to be actinomycete-selective was applied for a 30-day period. However, the use of this antibiotic-amended medium also enhanced production of secondary metabolites within the microbial community present relative to the water control, suggesting the treatment selected for antibiotic-resistant bacteria capable of producing a higher number of secondary metabolites. Secondary metabolite mining of "El Palmar" microbial community metagenomes identified polyketide synthase and non-ribosomal peptide synthetases' biosynthetic genes in all analyzed metagenomes. The presence of these genes correlated with the annotation of several secondary metabolites from the Global Natural Product Social Molecular Networking database. These results highlight the biotechnological potential of the microbial communities from "El Palmar", and show the impact selective media had on the composition of communities of actinobacteria.
Subject(s)
Actinobacteria/isolation & purification , Geologic Sediments/microbiology , Microbiota , Actinobacteria/genetics , Actinobacteria/metabolism , Metabolome , Metabolomics , Metagenome , MetagenomicsABSTRACT
Epidemiological studies related to androgens and prostate cancer (PC) have focused on serum determination of testosterone, androstenedione (A4), and DHEA, with inconsistent results. Herein, we hypothesized that differences in androgen biosynthetic and metabolic pathways, rather than differences in specific androgen concentrations, are associated with prostatic carcinogenesis. Therefore, spot urine samples from 111 incident PC cases with Gleason score at diagnosis and 227 healthy population controls, were analyzed. Urinary androgen concentrations (nanograms/milligrams of creatinine) were determined by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS). Using a factor analysis, we identified three androgen urinary excretion patterns. In a subsample, we evaluated a modification effect of the androgen receptor (AR) CAG polymorphism. Pattern I, characterized by A4 and testosterone hydroxylated metabolites (11ß-OHT; 2ß-OHT; 15ß-OHT; 2α-OHT; 6ß-OHT), was associated with high PC odds among carriers of AR gene (CAG)>19 repeats (OR: 3.67 95% CI: 1.23-11.0; P for interaction= 0.009). Conversely, higher testosterone excretion (pattern III), was marginally associated with lower (OR: 0.35 95% CI: 0.12-1.00, P for trend= 0.08) poorly differentiated PC (Gleason ≥8). No clear association was observed with pattern II (DHEA; 16α and 16ß-OHT). Our results were consistent with the previous evidence which suggests that the C11-oxy backdoor pathway is important for prostatic carcinogenesis. Androgen urine excretion analysis could be useful for PC diagnosis, treatment, and prognosis; however, further studies with a larger number of samples and the urinary determination of 11-ketoandrogens are necessary.
Subject(s)
Androgens , Prostatic Neoplasms , Androgens/metabolism , Carcinogenesis , Chromatography, Liquid , Dehydroepiandrosterone , Humans , Male , Prostatic Neoplasms/metabolism , Receptors, Androgen/genetics , Tandem Mass Spectrometry , Testosterone/metabolismABSTRACT
The marine-facultative Aspergillus sp. MEXU 27854, isolated from the Caleta Bay in Acapulco, Guerrero, Mexico, has provided an interesting diversity of secondary metabolites, including a series of rare dioxomorpholines, peptides, and butyrolactones. Here, we report on the genomic data, which consists of 11 contigs (N50~3.95 Mb) with a ~30.75 Mb total length of assembly. Genome annotation resulted in the prediction of 10,822 putative genes. Functional annotation was accomplished by BLAST searching protein sequences with different public databases. Of the predicted genes, 75% were assigned gene ontology terms. From the 67 BGCs identified, ~60% belong to the NRPS and NRPS-like classes. Putative BGCs for the dioxomorpholines and other metabolites were predicted by extensive genome mining. In addition, metabolomic molecular networking analysis allowed the annotation of all isolated compounds and revealed the biosynthetic potential of this fungus. This work represents the first report of whole-genome sequencing and annotation from a marine-facultative fungal strain isolated from Mexico.
Subject(s)
Aspergillus/metabolism , Metabolomics , Morpholines/metabolism , Peptides, Cyclic/metabolism , Aspergillus/genetics , Aspergillus/isolation & purification , Mexico , Molecular Structure , Morpholines/chemistry , Peptides, Cyclic/chemistry , Peptides, Cyclic/geneticsABSTRACT
Temephos (Tem) is the larvicide of choice to control mosquito transmission of dengue, Zika, and chikungunya. The toxicokinetic and toxicological information of temephos is very limited. The aim of this work was to determine the toxicokinetics and dosimetry of temephos and its metabolites. Male Wistar rats were orally administered temephos (300 mg/kg) emulsified with saline solution and sacrificed over time after dosing. Temephos and its metabolites were analyzed in blood and tissues by high performance liquid chromatography-diode array detector. At least eleven metabolites were detected, including temephos-sulfoxide (Tem-SO), temephos-oxon (Tem-oxon), temephos-oxon-sulfoxide (Tem-oxon-SO), temephos-oxon-SO-monohydrolyzed (Tem-oxon-SO-OH), 4,4´-thiodiphenol, 4,4´-sulfinyldiphenol, and 4,4´-sulfonyldiphenol or bisphenol S (BPS). The mean blood concentrations of temephos were fitted to a one-compartment model for kinetic analysis. At 2 h, the peak was reached (t1/2 abs = 0.38 h), and only trace levels were detected at 36 h (t1/2 elim = 8.6 h). Temephos was detected in all tissues and preferentially accumulated in fat. Temephos-sulfone-monohydrolyzed (Tem-SO2-OH) blood levels remained constant until 36 h and gradually accumulated in the kidney. Tem-oxon was detected in the brain, liver, kidney, and fat. Clearance from the liver and kidney were 7.59 and 5.52 ml/min, respectively. These results indicate that temephos is well absorbed, extensively metabolized, widely distributed and preferentially stored in adipose tissue. It is biotransformed into reactive metabolites such as Tem-oxons, Tem-dioxons, and BPS. Tem-SO2-OH, the most abundant metabolite of temephos, could be used as an exposure biomarker for toxicokinetic modeling. These results could provide critical insight into the dosimetry and toxicity of temephos and its metabolites.
Subject(s)
Biomarkers/metabolism , Insecticides/administration & dosage , Models, Biological , Temefos/administration & dosage , Administration, Oral , Animals , Chromatography, High Pressure Liquid , Insecticides/pharmacokinetics , Insecticides/toxicity , Male , Rats , Rats, Wistar , Temefos/pharmacokinetics , Temefos/toxicity , Time Factors , Tissue Array Analysis , ToxicokineticsABSTRACT
Testosterone regulates the male reproductive system and acts directly or indirectly on nearly all systems during fetal, pubertal and adult life. Testosterone homeostasis depends on its synthesis and degradation. The major biotransformation reactions are hydroxylation by different cytochrome P450 (CYP) isoforms. There are no described methods to determine the profile of testosterone-hydroxylated metabolites in human urine. The aim of this study was to develop an analytical method to determine testosterone-hydroxylated metabolites in human urine using UPLC-MS. Seven testosterone-hydroxylated metabolites, androstenedione, and testosterone, were identified by comparison of their tret and positive electrospray ionization (ESI+) data, with those of analytical standards. The method developed is sensitive, specific, repeatable, and precise. Limits of detection and quantitation for all compounds ranged from 1.360 to 13.054 ng/ml and 4.234-39.679 ng/ml, respectively. The percentages of recovery were between 81.2 and 128.8%. The applicability of the analytical method was confirmed by analysis of urine samples obtained from two groups of healthy men (25-30 and 50-75 years old). All analytes were identified with slightly different metabolites profiles in both groups. In conclusion, the UPLC-MS method developed here was validated for the analysis of testosterone-hydroxylated metabolites in human urine.
Subject(s)
Testosterone/urine , Adult , Aged , Calibration , Chromatography, High Pressure Liquid , Healthy Volunteers , Humans , Hydroxylation , Male , Mass Spectrometry , Middle Aged , Testosterone/metabolismABSTRACT
Temephos (Tem) is an organophosphorus pesticide widely used to kill and prevent the growth of the main vectors for the transmission of dengue, zika, and chikungunya viruses. In chlorinated water, Tem is oxidized to its dioxon-sulfoxide (Tem-dox-SO), dioxon-sulfone (Tem-dox-SO2), and sulfoxide (Tem-SO) derivatives; however, these compounds are not commercially available to be used as standards and in toxicological studies. In the present study, we synthesized and characterized the Tem-oxidation products and the compound 4,4'-sulfinyldiphenol. These compounds were obtained by a simple reaction between Tem or 4,4'-thiodiphenol with sodium hypochlorite or potassium periodate, and were characterized by IR, NMR, and UPLC-HRESIMS. The in vitro evaluation of inhibitory potency of Tem-oxidized products on human red blood cell acetylcholinesterase (RBC AChE) showed that Tem-dox-SO2 was the most potent inhibitor of human RBC AChE, and its effect was more pronounced than that observed for ethyl-paraoxon, a potent typical inhibitor of AChE. An HPLC-DAD method for the analysis of metabolic products of Tem was developed, which may be useful for monitoring in biological and environmental samples. The ability of Tem-oxidized metabolites to inhibit human RBC AChE suggests that the addition of Tem to chlorinated drinking water could result in an increase in the risk of RBC AChE inhibition after exposure.
Subject(s)
Cholinesterase Inhibitors/adverse effects , Erythrocytes/drug effects , Pesticides/adverse effects , Temefos/adverse effects , Acetylcholinesterase/metabolism , Cholinesterase Inhibitors/chemistry , Erythrocytes/enzymology , Humans , Oxidation-Reduction , Pesticides/chemistry , Temefos/analogs & derivativesABSTRACT
Herbal medicines are an integral element of alternative medical care in Mexico, and the best testimony to their efficacy and cultural value is their persistence in contemporary Mexican marketplaces where the highest percentages of medicinal and aromatic plants are sold. This chapter summarizes current trends in research on medicinal plants in Mexico, with emphasis on work carried out at the authors' laboratories. The most relevant phytochemical and pharmacological profiles of a selected group of plants used widely for treating major national health problems are described.From this contribution, it is evident that in the last five decades a significant amount of research on medicinal plants has been performed by Mexican scientists. Such efforts have led to the publication of many research papers in noted peer-reviewed journals and technical books. The isolation and structural characterization of hundreds of bioactive secondary metabolites have been accomplished, and most importantly, these studies have tended to support the ethnomedical uses of many different species. A multidisciplinary approach for investigating these plants has led to an increased emphasis on areas such as phytopharmacology, phytotoxicology, quality control, regulation, and conservation issues for these valuable resources. The medicinal plants analyzed so far have shown a very broad chemical diversity of their constituents, which have a high potential for exhibiting novel mechanistic effects biologically. The chapter shows also that there is need to conduct additional clinical studies on herbal drugs, in particular because the longstanding traditional evidence for their safety is not always sufficient to assure their rational use. There is also need to move to "omics" approaches for investigating the holistic effect and the influence of groups of phytochemicals on the whole organism. Mexican scientists may be expected to have bright prospects in this regard, which will imbue medicinal plant research with a new dynamism in the future.
Subject(s)
Phytochemicals/pharmacology , Plants, Medicinal/chemistry , Medicine, Traditional , Mexico , Phytotherapy , Plant Extracts/pharmacologySubject(s)
Biological Products , Plants, Medicinal , History, 20th Century , History, 21st Century , Mexico , VenezuelaABSTRACT
During our ongoing research on fungal strains from unexplored sources, the reinvestigation of the CHCl3-MeOH extract of the marine-facultative Aspergillus sp. MEXU 27854 yielded a new N-methyl cyclic pentapeptide (1) along with known butyrolactone II and PF1233 A. In addition, from the marine-facultative Gymnoascus hyalinosporus MEXU 29901, a new alternariol glucoside, 10-O-[ß-D-(4-methoxyl-glucopyranosyl)]-4-O-methylalternariol (2) and known alternariol 4-O-methyl ether, alternariol and beauvericin, were isolated. The structures of 1 and 2 were established by detailed spectroscopic data, and their absolute configuration was ascertained by Marfey's analysis and HRESIMS-MS/MS data for 1, and by chemical degradation and optical rotation analysis for 2.