Genome sequencing analysis of Streptomyces coelicolor mutants that overcome the phosphate-depending vancomycin lethal effect

Abstract Background Glycopeptide antibiotics inhibit bacterial cell-wall synthesis, and are important for the treatment of infections caused by multi drug-resistant strains of enterococci, streptococci and staphylococci. The main mechanism by which bacteria resist the action of glycopeptides is by producing a modified cell-wall in which the dipeptide D-Alanine-D-Alanine is substituted by D-Alanine-D-Lactate or D-Alanine-D-Serine. Recently, it has been shown that inorganic phosphate (Pi) induces hypersensitivity to vancomycin in Streptomyces coelicolor (which is highly resistant to the antibiotic in low-Pi media). This finding was surprising because the bacterium possesses the entire set of genes responsible for vancomycin resistance (VR); including those coding for the histidine kinase/response regulator pair VanS/VanR that activates the system. Results This work shows that high Pi amounts in the medium hamper the activation of the van promoters and consequently inhibit VR in S. coelicolor; i.e. the repression effect being stronger when basic or acidic forms of the nutrient are used. In addition, this work shows that lysozyme resistance is also highly regulated by the Pi concentration in the medium. At least five different mutations contribute to the overcoming of this repression effect over VR (but not over lysozyme resistance). Therefore, the interconnection of VR and lysozyme resistance mechanisms might be inexistent or complex. In particular, two kinds of mutant in which Pi control of VR has been lost (one class expresses the van genes in a constitutive manner; the other retains inducibility by vancomycin) have been isolated and further characterized in this study. Sequencing revealed that the first class of mutation conferred a single amino acid substitution in the second transmembrane helix of the VanS protein; whereas the other class hampered the expression or activity of a putative homolog (SCO1213) to the staphylococcal GatD protein. Complementation, phenotypic and bioinformatics analyses identified SCO1213, and its upstream gene (i.e. murT), as relevant genetic determinants involved with VR in S. coelicolor. Conclusion The genomic approach of this study together with other genetic and phenotypic analyses has allowed the identification of the uncharacterized murT-gatD Streptomyces genes and the characterization of their involvement with the Pi control of VR in S. coelicolor

Mecanismo molecular del control por fosfato de la cascada de expresión de los genes de biosíntesis de metabolitos secundarios en Streptomyces coelicolor

216 p.Se analiza la estructura operadora de pitH2, glpQ1, glpQ2 y afsS al conocimiento de la caja PHO en Streptomyces coelicolor; La inducción y represión génica por la proteína PhoP; La respuesta primaria ejercida por PhoP ante la escasez de fosfato en el medio; La interacción de las regulaciones por carbono, nitrógeno y fósforo y el efecto de PhoP en el metabolismo secundari

Characterization of the keratinolytic activity of three streptomyces strains and impact of their co-cultivation on this activity

Producción CientíficaIn this study, we describe the characterization of three efficient chicken feather-degrading Streptomyces bacteria isolated from honeybee samples and assess the impact of their co-cultivation on this activity and antistaphylococcal activity. Streptomyces griseoaurantiacus AD2 was the strain showing the highest keratinolytic activity (4000 U × mL−1), followed by Streptomyces albidoflavus AN1 and Streptomyces drozdowiczii AD1, which both generated approximately 3000 U × mL−1. Moreover, a consortium constituted of these three strains was able to use chicken feathers as its sole nutrient source and growth in such conditions led to a significant increase in antibiotic production. S. griseoaurantiacus AD2 was the only strain that exhibited weak antimicrobial activity against Staphylococcus aureus. UPLC analyses revealed that a significant number of peaks detected in the extracts of co-cultures of the three strains were missing in the extracts of individual cultures. In addition, the production of specialized metabolites, such as undecylprodigiosin and manumycin A, was clearly enhanced in co-culture conditions, in agreement with the results of the antimicrobial bioassays against S. aureus. Our results revealed the benefits of co-cultivation of these bacterial species in terms of metabolic wealth and antibiotic production. Our work could thus contribute to the development of novel microbial-based strategies to valorize keratin waste.Fondo Europeo de Desarrollo Regional (FEDER), (grant TCUE 2021–2023) - (project 067/229111)Unión Europea, H2020-MSCA-IF-2016 - (grant UE-18-VANRESTREP-740080

Cross‐talk between two global regulators in Streptomyces: PhoP and AfsR interact in the control of afsS, pstS and phoRP transcription

[EN] The regulatory proteins AfsR and PhoP control expression of the biosynthesis of actinorhodin and undecylprodigiosin in Streptomyces coelicolor. Electrophoretic mobility shift assays showed that PhoPDBD does not bind directly to the actII-ORF4, redD and atrA promoters, but it binds to the afsS promoter, in a region overlapping with the AfsR operator. DNase I footprinting studies revealed a PhoP protected region of 26 nt (PHO box; two direct repeats of 11 nt) that overlaps with the AfsR binding sequence. Binding experiments indicated a competition between AfsR and PhoP; increasing concentrations of PhoPDBD resulted in the disappearance of the AfsR-DNA complex. Expression studies using the reporter luxAB gene coupled to afsS promoter showed that PhoP downregulates afsS expression probably by a competition with the AfsR activator. Interestingly, AfsR binds to other PhoP-regulated promoters including those of pstS (a component of the phosphate transport system) and phoRP (encoding the two component system itself). Analysis of the AfsR-protected sequences in each of these promoters allowed us to distinguish the AfsR binding sequence from the overlapping PHO box. The reciprocal regulation of the phoRP promoter by AfsR and of afsS by PhoP suggests a fine interplay of these regulators on the control of secondary metabolismSIThis project was supported by grants of the ‘Comisión Interministerial de Ciencia y Tecnología’ (BIO2003-01489, BIO2006-14853-C02-01); Ministry of Education, Science and Technology, Madrid (GEN2003-20245-C09-01); the ERA-NET SySMO Project (GEN2006-27745-E/SYS) and the European Union (ACTINOGEN LSHM-CT-2004-005224). F. Santos-Beneit received a fellowship of the FPI programme (Ministry of Education, Spain). We thank S. Horinouchi for the plasmids containing the afsR gene, M. Bibb for strain S. coelicolor M513 (ΔafsR) and B. Martín, J. Merino, A. Casenave and B. Aguado for excellent technical assistanc

Recent trends and advances in biogas upgrading and methanotrophs-based valorization

Producción CientíficaThe global quest for sustainability in industrial activities and waste management has recently boosted biogas production worldwide. However, the rapid decrease in the levelized cost of electricity of renewable energies will soon entail electricity prices from biogas much higher than those from solar or wind power. In this context, the upgrading of biogas into biomethane represents an alternative to on-site biogas combustion. Membrane separation technology is rapidly dominating the biogas upgrading market and displacing scrubbing and adsorption technologies driven by the recent breakthroughs in material science. Similarly, biogas biorefineries have recently emerged as an innovative platform for biogas valorization capable of biotransforming methane into added value products. The limited number of bioproducts naturally synthesized by methanotrophs can be boosted via metabolic engineering of methanotrophs, while novel bioreactor configurations capable of supporting a cost-effective methane mass transfer from the gas phase to the methanotrophic broth are currently under investigation to facilitate the full scale implementation of biogas biorefineries.Junta de Castilla y Leon - FEDER (program CLU 2017–09, CL-EI-2021–07 and UIC 315)European Commission-H2020-MSCA-IF-2019 project UP-GRAD (894515)Ministerio de Ciencia, Innovación y Universidades (project IJC2019–040495-I

Genome‐wide transcriptomic and proteomic analysis of the primary response to phosphate limitation in Streptomyces coelicolor M145 and in a ΔphoP mutant

[EN] Phosphate limitation in Streptomyces and in other bacteria triggers expression changes of a large number of genes. This response is mediated by the two-component PhoR-PhoP system. A Streptomyces coelicolor ΔphoP mutant (lacking phoP) has been obtained by gene replacement. A genome-wide analysis of the primary response to phosphate limitation using transcriptomic and proteomic studies has been made in the parental S. coelicolor M145 and in the ΔphoP mutant strains. Statistical analysis of the contrasts between the four sets of data generated (two strains under two phosphate conditions) allowed the classification of all genes into 12 types of profiles. The primary response to phosphate limitation involves upregulation of genes encoding scavenging enzymes needed to obtain phosphate from different phosphorylated organic compounds and overexpression of the high-affinity phosphate transport system pstSCAB. Clear interactions have been found between phosphate metabolism and expression of nitrogen-regulated genes and between phosphate and nitrate respiration genes. PhoP-dependent repressions of antibiotic biosynthesis and of the morphological differentiation genes correlated with the observed ΔphoP mutant phenotype. Bioinformatic analysis of the presence of PHO boxes (PhoP-binding sequences) in the upstream regions of PhoP-controlled genes were validated by binding of PhoP, as shown by electrophoretic mobility shift assaysSIThis work was supported by grants of the CICYT (Madrid,Spain) to J. F. M. (BIO2003-01489 and GEN2003-20245-C09-01) and of the European Union (Project ActinoGEN OJ 2003/C164). A. Rodríguez was supported by a Torres Quevedo contract (PTQ2002-0468), and F. Santos received a FPI fellowship from the Ministry of Education (Madrid, Spain). We thank J. Merino, B. Martín, A. Casenave, and B. Aguado for excellent technical assistance, and V. Mersinias, O. Domínguez, L. Lombardía, and J.C. Oliveros for helpful discussion of microarray procedure

Parental and self-reported dietary and physical activity habits in pre-school children and their socio-economic determinants.

OBJECTIVE: To assess the agreement between self-reported and parent-reported dietary and physical activity habits in children; and to evaluate the socio-economic determinants of healthier habits (Mediterranean diet and physical activity) among children. DESIGN: Cross-sectional analysis of children recruited to a cluster-randomized controlled trial (Program SI!). Information about children's and parents' dietary and physical activity habits was obtained through validated questionnaires (Program SI! questionnaires, Kidmed, Krece Plus and Predimed scores). SETTING: Twenty-four schools in Madrid, Spain. SUBJECTS: Children (n 2062) aged 3-5 years and their parents (n 1949). RESULTS: There was positive agreement between parental- and self-reporting for three of the six children's habits examined. Parents' dietary and physical activity patterns were associated with those of their children. The main determinants of higher scores in children were higher parental age, the mother's scores, Spanish origin and higher awareness of human health (P<0·005). Children from parents with a low educational level had lower odds for scoring positively on items such as using olive oil (OR=0·23; 95 % CI 0·13, 0·41) and not skipping breakfast (OR=0·36; 95 % CI 0·23, 0·55), but higher odds for meeting the recommendations for consuming pulses (OR=1·71; 95 % CI 1·14, 2·55). Other habits being influenced by parental socio-economic status included the consumption of vegetables, fish, nuts, avoidance of fast food, and consumption of bakery products for breakfast. CONCLUSIONS: Children's habits may be influenced by their parents' health awareness and other socio-economic characteristics. These findings suggest that intervention strategies, even in very young children, should also target parents in order to achieve maximum success

Phosphate control over nitrogen metabolism in Streptomyces coelicolor: direct and indirect negative control of glnR, glnA, glnII and amtB expression by the response regulator PhoP

[EN] Bacterial growth requires equilibrated concentration of C, N and P sources. This work shows a phosphate control over the nitrogen metabolism in the model actinomycete Streptomyces coelicolor. Phosphate control of metabolism in Streptomyces is exerted by the two component system PhoR-PhoP. The response regulator PhoP binds to well-known PHO boxescomposed of direct repeat units (DRus). PhoP binds to the glnR promoter, encoding the major nitrogen regulator as shown by EMSA studies, but not to the glnRII promoter under identical experimental conditions. PhoP also binds to the promoters of glnA and glnII encoding two glutamine synthetases, and to the promoter of the amtB - glnK - glnD operon, encoding an ammonium transporter and two putative nitrogen sensing/regulatory proteins. Footprinting analyses revealed that the PhoP-binding sequence overlaps the GlnR boxes in both glnA and glnII. 'Information theory' quantitative analyses of base conservation allowed us to establish the structure of the PhoP-binding regions in the glnR, glnA, glnII and amtB genes. Expression studies using luxAB as reporter showed that PhoP represses the above mentioned nitrogen metabolism genes. A mutant deleted in PhoP showed increased expression of the nitrogen metabolism genes. The possible conservation of phosphate control over nitrogen metabolism in other microorganisms is discussedSIFunding by Comisión Interministerial de Ciencia y Tecnología [BIO2003-01489, BIO2006-14853-C02-01]; the ‘Ministerio de Ciencia e Innovación’, Madrid [GEN2003-20245-C09-01]; the AECID (Agencia Española de Cooperación Internacional para el Desarrollo), ‘Ministerio de Asuntos Exteriores y de Cooperación’, Madrid [A/010257/07]; the ERA-NET SySMO Project [GEN2006-27745-E/SYS]; and the European Union (ACTINOGEN LSHM-CT-2004-005224). F.P.U. fellowship of the Ministerio de Ciencia e Innovación (Spain) (to K.A.); fellowship of the F.P.I. program (Ministerio de Ciencia e Innovación, Spain) (to F.S.B.). Funding for open access charge: Institute of Biotechnology of Leó

Body image and dietary habits in adolescents: a systematic review

Context: Adolescence is a critical developmental stage in which increasing concerns about body image (BI) coincide with the consolidation of dietary habits (DHs). Multiple studies have sought robust associations between BI and DHs to prevent unhealthy behaviors. Objective: The aim of this systematic review was to examine the available literature on the association between BI perception (BIP) and/or satisfaction (BIS) and DHs in adolescents. Data Sources: A search was carried out of 5 electronic databases (PubMed, SciELO, Cochrane, Embase, and PsycInfo) using a combination of keywords (and synonyms) related to adolescence, BI, and diet. Data Extraction: Data screening, extraction, and quality assessment were performed independently by 2 investigators using the PRISMA and AXIS guidelines. Data Analysis: Of 2496 articles screened, 30 articles, published in English or Spanish, that evaluated the relationship between BI and DHs in adolescents aged between 10 years and 18 years, were included. A relationship between accurate BI perception in adolescents and healthy DHs was reported in 5 articles (16.2%). A relationship between overestimation of body weight in adolescents and healthy DHs was reported in 4 articles (13.3%). A relationship between underestimation of body weight and unhealthy DHs was reported in 8 articles (26.7%). In addition, 4 articles (13.3%) reported a relationship between BIS and healthy DHs. The desire to gain weight was associated with unhealthy DHs in 3 (10%) of the articles, while the desire to lose weight was related to healthy DHs in 3 (10%) of the articles and to unhealthy DHs in 3 (10%) other articles. There were also gender differences in the relationship between BIP or BIS and DHs. Conclusion: Adolescents who underestimate their body weight tend to report less healthy DHs than body weight overestimators. Adolescents unsatisfied with their BI and with a drive for thinness frequently engage in DHs linked to losing weight. Systematic Review Registration: PROSPERO registration no. CRD42020184625