Interplay between Intestinal Bacterial Communities and Unicellular Parasites in a Morbidly Obese Population: A Neglected Trinomial.

Obesity is an epidemic causing a metabolic health crisis. Herein, the interactions between the gut prokaryotic and eukaryotic communities, metabolic comorbidities and diet were studied. Stool samples from 56 subjects, 47 with type III obesity and 9 with type II obesity and cardiovascular risk or metabolic disease, were assessed for the richness, diversity and ecology of the bacterial gut community through metagenomics, together with the study of the presence of common unicellular eukaryote parasites (Blastocystis sp., Dientamoeba fragilis and Giardia intestinalis) by qPCR. Clinical information regarding metabolic comorbidities and non-alcoholic hepatic fatty liver disease was gathered. To assess the quality of the patients' diet, each participant filled in three dietary questionnaires. The most prevalent parasite Blastocystis sp. (46.4%), together with D. fragilis (8.9%), was found to be associated with higher mean diversity indexes regarding non-colonized subjects; the opposite of that which was observed in those with G. intestinalis (16.1%). In terms of phyla relative abundance, with Blastocystis sp. and D. fragilis, very slight differences were observed; on the contrary, G. intestinalis was related to an increase in Bacteroidetes and Proteobacteria, and a decrease in Firmicutes and Actinobacteria, presenting the lowest Firmicutes/Bacteroidetes ratio. At genus level, Blastocystis sp. and/or D. fragilis was accompanied with an increase in Lactobacillus spp., and a decrease in Akkermansia spp., Bifidobacterium spp. and Escherichia spp., while G. intestinalis was associated with an increase in Bacteroides spp., and a decrease in Faecalibacterium spp., Prevotella spp. and Lactobacillus spp., and the highest Bacteroides spp./Prevotella spp. ratio. Participants with non-alcoholic hepatic fatty liver presented a higher Firmicutes/Bacteroidetes ratio, and those with type 2 diabetes displayed a significantly lower Faecalibacterium spp./Escherichia spp. ratio, due to an overrepresentation of the genus Escherichia spp. The presence of parasites was associated with variations in the richness, diversity and distribution of taxa in bacterial communities, confirming a gain in diversity associated with Blastocystis sp. and providing different functioning of the microbiota with a potential positive effect on comorbidities such as type 2 diabetes, insulin resistance and metabolic syndrome. Future basic and clinical studies should assess the beneficial or pathogenic effect of these eukaryotes on obese subjects and focus on deciphering whether they may imply a healthier metabolic profile.

Antimicrobial Activity, in silico Molecular Docking, ADMET and DFT Analysis of Secondary Metabolites from Roots of Three Ethiopian Medicinal Plants.

BACKGROUND: Uvaria scheffleri (Annonaceae), Clematis burgensis (Ranunculaceae), and Euphorbia schimperiana (Euphorbiaceae) are medicinal plants traditionally used to treat cough, tuberculosis, asthma, sore throat and skin infections. METHODS: Silica gel column chromatographic separation was used to isolate compounds. Crude extract and isolated compounds were evaluated for antimicrobial activity against Staphylococcus aureus, Escherichia coli, and Candida albicans via the broth dilution method. Docking studies were performed with E. coli DNA-Gyrase B and human DNA topoisomerase IIα by using AutoDock Vina. ADMET were predicted by SwissADME, PreADMET, and OSIRIS Property predictions. The optimized structures and molecular electrostatic potential surface of the isolated compounds were predicted by DFT analysis using B3LYP/6-31G basis levels. RESULTS: Silica gel column chromatographic separation afforded five compounds 1-5 of which N-methyl-2,3-bis(2-hydroxybenzyl)-1Н-indol (1) is reported herein for the first time, along with known C-benzylated dihydrochalcone uvaretin (2), bis(2-ethylheptyl) phthalate (3), lupeol (4) and suberosin derivative (5). Dichloromethane roots extract of U. scheffleri showed potent antibacterial activity against S. aureus (MIC = 6.25 µg/mL) compared to gentamicin (MIC=5 µg/mL). In silico, molecular docking analysis of compounds (1and 3-5) showed strong interaction with E. coli DNA gyrase B with a binding energy value ranging from -6.9 to -6.0 kcal/mol compared to ciprofloxacin -7.2 kcal/mol, whereas analysis against human topoisomerase IIα showed binding energy value ranging from -5.9 to -5.3 kcal/mol compared to vosaroxin (-6.2 kcal/mol). CONCLUSION: The results obtained suggest that N-methyl-2,3-bis(2-hydroxybenzyl)-1Н-indol (1) and coumarin (5) are potential topoisomerase II α inhibitors and might be used as anticancer agents. The ADMET studies showed the highest drug-likeness properties for studied compounds other than bis(2-ethylheptyl) phthalate (3). DFT calculations suggested that studied compounds showed the lowest gap energy and were chemically reactive, and isolated compounds may serve as potential drug candidates that corroborate with the traditional uses of studied plants.

Ciprofloxacin self-dissolvable Soluplus based polymeric films: a novel proposal to improve the management of eye infections.

Infections of the eye are among the leading causes of vision impairment and vision loss worldwide. The ability of a drug to access the anterior parts of the eye is negligible after systemic administration. Effective drug delivery to the eye is a major challenge due to the presence of protective mechanisms and physiological barriers that result in low ocular availability after topical application. The main purpose of this work was the improvement of the corneal and conjunctival permeation of the antibiotic Ciprofloxacin, a wide spectrum antibiotic used for the most common eye infection, using a self-dissolving polymeric film. Films were prepared by the solvent casting technique, using polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft co-polymer (Soluplus), polyvynyl alcohol, and propylene glycol. Films were homogeneous in drug content and thickness, as demonstrated by adapting the Swiss Roll technique followed by microscopy observation. These films proved in vitro to control the release of the Ciprofloxacin. Ex vivo permeability studies using Franz diffusion cells and porcine cornea and sclera showed an effective permeability of the drug without inducing irritation of the tissues. Films swelled in contact with artificial tears forming an in situ gel over 20 min, which will improve drug contact and reduce the need of multiple dosing. The antibiotic activity was also tested in vitro in five types of bacterial cultures, assuring the pharmacological efficacy of the films. The developed films are a promising drug delivery system to topically treat or prevent ocular infections.

Implementation of Antibiotic Discovery by Student Crowdsourcing in the Valencian Community Through a Service Learning Strategy.

Antibiotic misuse is a public health problem due to the appearance of resistant strains in almost all human pathogens, making infectious diseases more difficult to treat. The search for solutions requires the development of new antimicrobials as well as novel strategies, including increasing social awareness of the problem. The Small World Initiative (SWI) and the Tiny Earth (TE) network are citizen science programs pursuing the discovery of new antibiotics from soil samples and the promotion of scientific culture. Both programs aim to bring scientific culture and microbiological research closer to pre-university students through a crowdsourcing strategy and a Service Learning (SL) educational approach, with a 2-fold objective: to encourage students to pursue careers in science and to involve them in the discovery of soil microorganisms producing new antimicrobials. SWI and TE projects were put into practice in Spain under the common name MicroMundo. MicroMundo@Valencia was implemented at the Universitat de València (UV) during the academic years 2017-2018 and 2018-2019. It trained 140 university students to disseminate this initiative into 23 high/secondary schools, and one primary school, involving about 900 people (teachers and students) as researchers. A total of 7,002 bacterial isolates were obtained from 366 soil samples and tested for antibiosis at UV and high/secondary school centers. About 1 or 7% of them produced inhibition halos for the Escherichia coli or Bacillus cereus target strains, respectively. Geolocation of sampling sites by an application developed ad hoc and Kriging analysis also allowed detection of soil foci of antibiotic-producing bacteria. Evaluation of the project by university, high/secondary, and primary school students revealed their strong positive perception and their increased interest in science, as a consequence of acquiring new scientific and pedagogical concepts and skills that they were able to pass on to other classmates, younger students, or relatives. To further expand the dissemination of the project in the Valencian Community, diverse extramural activities deemed to include a gender perspective and aimed at different age groups, were also carried out, obtaining very satisfactory results, increasing sensitivity and awareness to the global antibiotic crisis.

Enterobacter and Klebsiella species isolated from fresh vegetables marketed in Valencia (Spain) and their clinically relevant resistances to chemotherapeutic agents.

Occurrence of antibiotic-resistant pathogenic or commensal enterobacteria in marketed agricultural foodstuffs may contribute to their incorporation into the food chain and constitutes an additional food safety concern. In this work, we have determined the clinically relevant resistances to 11 common chemotherapeutic agents in Enterobacter and Klebsiella isolates from fresh vegetables from various sources (supermarkets and greengrocers' shops in Valencia, Spain). A total of 96 isolates were obtained from 160 vegetables analyzed (50% positive samples): 68 Enterobacter isolates (59 E. cloacae, two E. aerogenes, two E. cancerogenus, one E. gergoviae, and four E. sakazakii, currently Cronobacter spp.), and 28 Klebsiella isolates (19 K. oxytoca and 9 K. pneumoniae). Only seven isolates were susceptible to all agents tested, and no resistances to ceftazidime, ciprofloxacin, gentamicin, and chloramphenicol were detected. Most isolates were resistant to amoxicillin/clavulanic acid (74 [58 Enterobacter and 16 Klebsiella]) or to ampicillin (80 [55/25]). Other resistances were less frequent: nitrofurantoin (13 isolates [12/1]), tetracycline (6 [5/1]), co-trimoxazole (3 [3/0]), cefotaxime (1 [1/0]), and streptomycin (2 [1/1]). Multiresistant isolates to two (56 [41/15]), three (10 E. cloacae isolates), four (one E. cloacae and one K. pneumoniae isolate), and five (two E. cloacae isolates) chemotherapeutic agents were also detected. The presence of potential pathogens points to marketed fresh produce, which often is eaten raw, as a risk factor for consumer health. In addition, these results support the usefulness of these bacterial species as indicators of the spreading of antibiotic resistances into the environment, particularly in the food chain, and suggest their role as carriers of resistance determinants from farms to consumers, which may constitute an additional "silent" food safety concern. Therefore, there is a need to improve the hygienic quality of marketed fresh vegetables, from better methods to prevent contamination in the farms to the use of sanitizing practices at home.

A role for the MAP kinase gene MKC1 in cell wall construction and morphological transitions in Candida albicans.

The Candida albicans MKC1 gene encodes a mitogen-activated protein (MAP) kinase, which has been cloned by complementation of the lytic phenotype associated with Saccharomyces cerevisiae slt2 (mpk1) mutants. In this work, the physiological role of this MAP kinase in the pathogenic fungus C. albicans was characterized and a role for MKC1 in the biogenesis of the cell wall suggested based on the following criteria. First, C. albicans mkc1 delta/mkc1 delta strains displayed alterations in their cell surfaces under specific conditions as evidenced by scanning electron microscopy. Second, an increase in specific cell wall epitopes (O-glycosylated mannoprotein) was shown by confocal microscopy in mkc1 delta/mkc1 delta mutants. Third, the sensitivity to antifungals which inhibit (1,3)-beta-glucan and chitin synthesis was increased in these mutants. In addition, evidence for a role for the MKC1 gene in morphological transitions in C. albicans is presented based on the impairment of pseudohyphal formation of mkc1 delta/mkc1 delta strains on Spider medium and on the effect of its overexpression on Sacch. cerevisiae colony morphology on SLADH medium. Using the two-hybrid system, it was also demonstrated that MKC1 is able to interact specifically with Sacch. cerevisiae Mkk1p and Mkk2p, the MAP-kinase kinases of the PKC1-mediated route of Sacch. cerevisiae, and to activate transcription in Sacch. cerevisiae when bound to a DNA-binding element. These results suggest a role for this MAP kinase in the construction of the cell wall of C. albicans and indicate its potential relevance for the development of novel antifungals.