Unearthing a Cryptic Biosynthetic Gene Cluster for the Piperazic Acid-Bearing Depsipeptide Diperamycin in the Ant-Dweller Streptomyces sp. CS113.

Piperazic acid is a cyclic nonproteinogenic amino acid that contains a hydrazine N-N bond formed by a piperazate synthase (KtzT-like). This amino acid, found in bioactive natural products synthesized by non-ribosomal peptide synthetases (NRPSs), confers conformational constraint to peptides, an important feature for their biological activities. Genome mining of Streptomyces strains has been revealed as a strategy to identify biosynthetic gene clusters (BGCs) for potentially active compounds. Moreover, the isolation of new strains from underexplored habitats or associated with other organisms has allowed to uncover new BGCs for unknown compounds. The in-house "Carlos Sialer (CS)" strain collection consists of seventy-one Streptomyces strains isolated from the cuticle of leaf-cutting ants of the tribe Attini. Genomes from twelve of these strains have been sequenced and mined using bioinformatics tools, highlighting their potential to encode secondary metabolites. In this work, we have screened in silico those genomes, using KtzT as a hook to identify BGCs encoding piperazic acid-containing compounds. This resulted in uncovering the new BGC dpn in Streptomyces sp. CS113, which encodes the biosynthesis of the hybrid polyketide-depsipeptide diperamycin. Analysis of the diperamycin polyketide synthase (PKS) and NRPS reveals their functional similarity to those from the aurantimycin A biosynthetic pathway. Experimental proof linking the dpn BGC to its encoded compound was achieved by determining the growth conditions for the expression of the cluster and by inactivating the NRPS encoding gene dpnS2 and the piperazate synthase gene dpnZ. The identity of diperamycin was confirmed by High-Resolution Mass Spectrometry (HRMS) and Nuclear Magnetic Resonance (NMR) and by analysis of the domain composition of modules from the DpnP PKS and DpnS NRPS. The identification of the dpn BGC expands the number of BGCs that have been confirmed to encode the relatively scarcely represented BGCs for depsipeptides of the azinothricin family of compounds and will facilitate the generation of new-to-nature analogues by combinatorial biosynthesis.

Uncovering the Cryptic Gene Cluster ahb for 3-amino-4-hydroxybenzoate Derived Ahbamycins, by Searching SARP Regulator Encoding Genes in the Streptomyces argillaceus Genome.

Genome mining using standard bioinformatics tools has allowed for the uncovering of hidden biosynthesis gene clusters for specialized metabolites in Streptomyces genomes. In this work, we have used an alternative approach consisting in seeking "Streptomyces Antibiotic Regulatory Proteins" (SARP) encoding genes and analyzing their surrounding DNA region to unearth cryptic gene clusters that cannot be identified using standard bioinformatics tools. This strategy has allowed the unveiling of the new ahb cluster in Streptomyces argillaceus, which had not been retrieved before using antiSMASH. The ahb cluster is highly preserved in other Streptomyces strains, which suggests a role for their encoding compounds in specific environmental conditions. By combining overexpression of three regulatory genes and generation of different mutants, we were able to activate the ahb cluster, and to identify and chemically characterize the encoded compounds that we have named ahbamycins (AHBs). These constitute a new family of metabolites derived from 3-amino-4-hydroxybenzoate (3,4-AHBA) known for having antibiotic and antitumor activity. Additionally, by overexpressing three genes of the cluster (ahbH, ahbI, and ahbL2) for the synthesis and activation of 3,4-AHBA, a new hybrid compound, AHB18, was identified which had been produced from a metabolic crosstalk between the AHB and the argimycin P pathways. The identification of this new BGC opens the possibility to generate new compounds by combinatorial biosynthesis.

Colibrimycins, Novel Halogenated Hybrid Polyketide Synthase-Nonribosomal Peptide Synthetase (PKS-NRPS) Compounds Produced by Streptomyces sp. Strain CS147.

The improvement of genome sequencing techniques has brought to light the biosynthetic potential of actinomycetes due to the large number of gene clusters they present compared to the number of known compounds. Genome mining is a recent strategy in the search for novel bioactive compounds, which involves the analysis of sequenced genomes to identify uncharacterized natural product biosynthetic gene clusters, many of which are cryptic or silent under laboratory conditions, and to develop experimental approaches to identify their products. Owing to the importance of halogenation in terms of structural diversity, bioavailability, and bioactivity, searching for new halogenated bioactive compounds has become an interesting issue in the field of natural product discovery. Following this purpose, a screening for halogenase coding genes was performed on 12 Streptomyces strains isolated from fungus-growing ants of the Attini tribe. Using the bioinformatics tools antiSMASH and BLAST, six halogenase coding genes were identified. Some of these genes were located within biosynthetic gene clusters (BGCs), which were studied by construction of several mutants for the identification of the putative halogenated compounds produced. The comparison of the metabolite production profile of wild-type strains and their corresponding mutants by ultrahigh-performance liquid chromatography-UV and high-performance liquid chromatography-mass spectrometry allowed us the identification of a novel family of halogenated compounds in Streptomyces sp. strain CS147, designated colibrimycins. IMPORTANCE Genome mining has proven its usefulness in the search for novel bioactive compounds produced by microorganisms, and halogenases comprise an interesting starting point. In this work, we have identified a new halogenase coding gene that led to the discovery of novel lipopetide nonribosomal peptide synthetase/polyketide synthase (NRPS/PKS)-derived natural products, the colibrimycins, produced by Streptomyces sp. strain CS147, isolated from the Attini ant niche. Some colibrimycins display an unusual α-ketoamide moiety in the peptide structure. Although its biosynthetic origin remains unknown, its presence might be related to a hypothetical inhibition of virus proteases, and, together with the presence of the halogenase, it represents a feature to be incorporated in the arsenal of structural modifications available for combinatorial biosynthesis.

Spontaneous verbal repetition in toddler-adult conversations: a longitudinal study with Spanish-speaking two- year-olds.

The role of children's verbal repetition of parents' utterances on vocabulary growth has been well documented (Masur, 1999). Nevertheless, few studies have analyzed adults' and children's spontaneous verbal repetition around the second birthday distinguishing between the types of repetition. We analyzed longitudinally Spanish-speaking parent-child dyads during spontaneous interaction at 21, 24 and 30 months. Linguistic level was measured using the Spanish version of the MacArthur CDI (López-Ornat et al., 2005). Children's and adults' repetitions are about 17% of the speech. Children repeated adults' utterances in a reduced manner whereas adults produced more extended repetitions. Adults' rate of repetition predicted children's linguistic level at 30 months. Children's rate of repetition did not predict linguistic level. These results suggest that parents adapt their speech to children's communicative abilities. Since children's rate of repetition did not predict linguistic level, we suggest that verbal imitation plays an indirect and complex role in communicative development.

A Spanish Sign Language (LSE) Adaptation of the Communicative Development Inventories.

This article presents the adaptation of the MacArthur Communicative Development Inventory (CDI; Fenson et al., 1993, Guide and technical manual for the MacArthur Communicative Development Inventories. San Diego, CA: Singular Press; Fenson et al. 1994, Variability in early communicative development. Monographs of the Society for Research in Child Development, 59, 1-173) to Spanish Sign Language (LSE). Data were collected from 55 participants (32 boys and 23 girls; 17 deaf signers, 38 hearing signers) who, evaluated by their caregivers every 4 months, presented a total of 170 records. The parents reported the signs that the children could understand or produce between 8 and 36 months. Results suggested that the CDI adapted to LSE is a valid and reliable instrument. Signing children could understand more signs than they produced at this early developmental stage. There were no significant differences between boys and girls, or between deaf and hearing children. The development of LSE is similar to other sign languages, although with a lower production of signs in the early stages, perhaps due to the bilingualism of most of the children of our study.

Cluster of Escherichia coli isolates producing a plasmid-mediated OXA-48 ß-lactamase in a Spanish hospital in 2012.

Three unrelated sequence type 131 (ST131), ST58, and ST83 Escherichia coli isolates with low-level resistance to imipenem and resistance to ertapenem were recovered in a Spanish hospital from July to October 2012. They were positive for blaOXA-48 carried by an IncL/M conjugative plasmid, which may have been acquired from Klebsiella pneumoniae.

Past, present, and future of microbiome-based therapies.

Technological advances in studying the human microbiome in depth have enabled the identification of microbial signatures associated with health and disease. This confirms the crucial role of microbiota in maintaining homeostasis and the host health status. Nowadays, there are several ways to modulate the microbiota composition to effectively improve host health; therefore, the development of therapeutic treatments based on the gut microbiota is experiencing rapid growth. In this review, we summarize the influence of the gut microbiota on the development of infectious disease and cancer, which are two of the main targets of microbiome-based therapies currently being developed. We analyze the two-way interaction between the gut microbiota and traditional drugs in order to emphasize the influence of gut microbial composition on drug effectivity and treatment response. We explore the different strategies currently available for modulating this ecosystem to our benefit, ranging from 1st generation intervention strategies to more complex 2nd generation microbiome-based therapies and their regulatory framework. Lastly, we finish with a quick overview of what we believe is the future of these strategies, that is 3rd generation microbiome-based therapies developed with the use of artificial intelligence (AI) algorithms.

Efficient mobilization of a resistance derivative of pSLT, the virulence plasmid specific of Salmonella enterica serovar Typhimurium, by an IncI1 plasmid.

pUO-StVR2 is a derivative of pSLT, the virulence plasmid specific of Salmonella enterica serovar Typhimurium, which confers multidrug resistance. This plasmid is widespread among closely related isolates of S. Typhimurium, and often coexists with other plasmids like pStR12. The latter belongs to incompatibility group IncI1, was assigned to ST48 by pMLST (plasmid multilocus sequence typing), and confers resistance to streptomycin/spectinomycin, chloramphenicol, trimethoprim and sulphonamides, with the responsible genes (aadA1/aadA2, cmlA1, dfrA12 and sul3) located on a sul3-class 1 integron. When using clinical isolates of S. Typhimurium containing one (pUO-StVR2) or both (pUO-StVR2 and pStR12) plasmids as donors and Escherichia coli K12 J53 resistant to rifampicin as recipient, the conjugation frequencies of pUO-StVR2 and pStR12 were 10⁻8 and 10⁻³-10⁻5 transconjugants/donor, respectively, while the transfer frequency of pUO-StVR2 increased 10² up to 105 times through mobilization by pStR12, depending on the donor strain and experimental conditions. Mobilization of pUO-StVR2, a plasmid which encodes virulence and resistance functions, by compatible plasmids which coexist in the same bacterium can facilitate the spread of these properties in S. Typhimurium, one of the most common serovars of S. enterica.

GutAlive® enables DNA-based microbiome analysis without disrupting the original composition and diversity.

Introduction: A precise fecal microbiome analysis requires normalized methods for microbiome sampling, transport and manipulation in order to obtain a representative snapshot of the microbial community. GutAlive® is the unique stool collection kit that generates an anaerobic atmosphere enabling oxygen sensitive bacteria to survive, maintaining the original microbiome composition and diversity. Methods: Five stool samples from different donors were collected using two different sampling devices, GutAlive® and Zymo DNA/RNA Shield®, and processed at four different time points. Shotgun metagenomics was used to evaluate the influence of the device and the processing timing on the microbial populations to unravel the potential fluctuations on the composition and diversity of the fecal microbiome and the metabolic pathways profiling. Additionally, RT-qPCR was used to quantify bacterial cell viability for downstream applications of microbiota samples beyond metagenomics. Results: Our results show that GutAlive® enables bacterial cell viability overtime preserving DNA integrity, obtaining high-quantity and high-quality DNA to perform microbiome analysis using shotgun metagenomics. Based on the taxonomic profiling, metabolic pathways analysis, phylogeny and metagenome-assembled genomes, GutAlive® displayed greater performance without significant variability over time, showcasing the stabilization of the microbiome preserving the original composition and diversity. Indeed, this DNA stabilization is enabled with the preservation of bacterial viability on an anaerobic environment inside of the sampling device, without the addition of any reagents that interact directly with sample. Conclusion: All the above makes GutAlive® an user-friendly kit for self-collection of biological samples, suitable for microbiome analysis, diagnostics, fecal microbiota transplant and bacterial isolation, maintaining the stability and bacterial viability over time, preserving the original composition and diversity of the microbiome.

Food deprivation induces chronic stress and affects thyroid hormone metabolism in Senegalese sole (Solea senegalensis) post-larvae.

In vertebrates, stress and thyroid systems interact closely, most likely because of the involvement of both systems in energy metabolism. However, studies on these interactions, especially during larval development, are scarce. Recently, cDNAs coding for corticotropin-releasing hormone (CRH) and CRH-binding protein (CRH-BP), two key players in the regulation of the neuroendocrine stress response, were characterized for the flatfish Senegalese sole (Solea senegalensis). To investigate the involvement of stress and thyroid systems in this species, the effects of food deprivation during early development of S. senegalensis were assessed. Growth was arrested in food-deprived post-larvae, which was also reflected by decreased carbon and nitrogen contents, indicating increased catabolism. Food deprivation induces chronic stress, as illustrated by enhanced whole-body cortisol levels, as well as an up regulation of crh and a decrease of crh-bp expression levels. Furthermore, whole-body total T3 concentrations of food-deprived post-larvae were reduced, although tshß subunit expression levels remained unaffected. Our results show that food deprivation is a chronic stressor that induces energy-releasing catabolic processes that compensate for the reduced energy intake, and inhibits anabolic processes via the peripheral thyroid system.

Oral Tradition as Context for Learning Music From 4E Cognition Compared With Literacy Cultures. Case Studies of Flamenco Guitar Apprenticeship.

The awareness of the last 20 years about embodied cognition is directing multidisciplinary attention to the musical domain and impacting psychological research approaches from the 4E (embodied, embedded, enactive, and extended) cognition. Based on previous research regarding musical teaching and learning conceptions of 30 young guitar apprentices of advanced level in three learning cultures: Western classical, jazz, and flamenco of oral tradition, two participants of flamenco with polarised profiles of learning (reproductive and transformative) were selected as instrumental cases for a prospective ex post facto design. Discourse and practice of the two flamenco guitarists were analysed in-depth to describe bodily issues and verbal discourse on the learning practice in their natural contexts. Qualitative analysis is performed on the posture, gestures, verbal discourse, and musical practice of the participants through the System for the Analysis of Music Teaching and Learning Practices (SAPIL). The results are organised attending: (a) the Embodied mind through differential postures and gestures of flamenco participants that showed a fusion among verbal, body language, and musical discourse with respect to the musical literacy cultures; (b) the Embedded mind and a detailed description of circumstances and relationships of the two flamenco participants, and how music is embedded in their way of life, family and social context, and therefore transcends musical activity itself; (c) the Enactive mind, regarding the active processes that make differences between the reproductive and the transformative flamenco apprentices, then tentative relationship are observed in the discourse of each apprentice and the way in which they practice; finally, (d) the Extended mind through the bodily, technical and symbolic tools they use during learning. Flamenco culture of oral tradition made use of listening, and temporary external representations instead of notational, but also the body played a central role in a holistic rhythm processing through multimodality, such as singing, playing, and dancing. Conclusions point out the embodied mind as a result of the culture of learning reflected through the body and the gesture in instrumental learning.

Adaptation and Validation of the Evasive Attitudes of Sexual Orientation Scale into Spanish.

This article presents an instrumental study to validate the adaptation of the Evasive Attitudes of Sexual Orientation Scale (EASOS) to Spanish. This instrument has been shown to be useful in detecting the potential lack of awareness about the situation of lesbian, gay, bisexual, and queer (LGBQ) people among psychology professionals and its possible relationship to contemporary homonegative attitudes. The 596 heterosexual psychology students who participated were given an adaptation into Spanish (back translation). A confirmatory factor analysis was performed to study the fit to the factorial structure of the original scale (aversive heterosexism, institutional heterosexism, and heterosexual privilege). The internal consistency of the subscales was adequate (.70-.83). The convergent validity showed positive correlations and significant predictive levels between the EASOS and various attitudinal scales and sociodemographic variables. The findings offer evidence that the EASOS is an adequate instrument to evaluate LGBQ-negativity, particularly in the field of psychosocial intervention.

On the Multimodal Path to Language: The Relationship Between Rhythmic Movements and Deictic Gestures at the End of the First Year.

The aim of this study is to analyze the relationship between rhythmic movements and deictic gestures at the end of the first year of life, and to focus on their unimodal or multimodal character. We hypothesize that multimodal rhythmic movement performed with an object in the hand can facilitate the transition to the first deictic gestures. Twenty-three children were observed at 9 and 12 months of age in a naturalistic play situation with their mother or father. Results showed that rhythmic movements with objects in the hand are a frequent behavior in children's repertoires. Rhythmic behaviors tend to decrease from 9 to 12 months, specifically when they are unimodal. Multimodal rhythmic behavior production at 9 months is positively related with proximal deictic gestures 3 months later. Multimodal rhythmic movements are not directly related to distal deictic gestures, but are indirectly related via proximal deictic gestures. These results highlight the relevance of multimodal behaviors in the transition to the use of early gestures, and can be considered as a transitional phenomenon between the instrumental action and early communicative gestures.

Multimodal Communication in Children with Autism Spectrum Disorder and Different Linguistic Development.

This study focuses on the multimodal communication of children with autism spectrum disorders (ASD) compared to typically developing (TD) children. Eleven children with ASD (aged from 28 to 79 months) and 11 TD children (from 12 to 30 months) were matched by their productive vocabulary. We observed their communicative production in a semi-structured play situation. Results showed no differences in the combinations of gestural and vocal elements between children with ASD and TD. By contrast, regarding the production of the three-element multimodal combinations, we found a different pattern between ASD and TD children depending on their lexical development. These results provide clues to understand some controversial findings regarding multimodal production of people with ASD described in the literature.

Correlate Attitudes Toward LGBT and Sexism in Spanish Psychology Students.

The present study evaluates the correlations between sexism, homonegativity, binegativity, pro-trans attitudes, political affiliation, contact with LGBT individuals and perceived stigma among psychology students. A study was conducted with 655 cis women (471 heterosexuals, 179 bisexuals and lesbians) and 174 cis men (120 heterosexuals, 54 bisexuals and gays). Descriptive, multivariate analysis of variance, bivariate correlations and multiple regression were used. In general, the groups of men and heterosexuals obtained higher negativity scores and lower acceptance scores, with significant correlations being more frequent in the heterosexual group. Predictive models confirmed the literature on social and ideological conservatism.

Intersections Around Ambivalent Sexism: Internalized Homonegativity, Resistance to Heteronormativity and Other Correlates.

This article explores the connections between the construct of sexism and other sociodemographic and attitudinal variables, such as internalized homonegativity and heteronormative resistances, among psychology students. Both unrefined and inferential analyses were used with a representative sample of 841 psychology students from public universities in Madrid. Results showed higher levels of sexism, internalized homonegativity and low resistances to heteronormativity among groups of men, heterosexuals and conservatives. Interactions were found that showed a higher degree of hostile sexism in: heterosexual people with respect to LGB and heterosexual men with respect to heterosexual women. Also, interactions were found to show a greater degree of heteronormative resistance in: LGB people with respect to heterosexuals and left-wing women with respect to right-wing women. Correlations with sexism varied according to gender identity and sexual orientation. In addition, heteronormative resistances correlated negatively with sexism, while some components of internalized homonegativity correlated positively. Political affiliation was the most frequent predictor of sexism. The results highlight the need for an intersectional approach to understanding the phenomenon of sexism.

A Plasmid-Encoded FetMP-Fls Iron Uptake System Confers Selective Advantages to Salmonella enterica Serovar Typhimurium in Growth under Iron-Restricted Conditions and for Infection of Mammalian Host Cells.

The resistance plasmid pUO-StVR2, derived from virulence plasmid pSLT, is widespread in clinical isolates of Salmonella enterica serovar Typhimurium recovered in Spain and other European countries. pUO-StVR2 carries several genes encoding a FetMP-Fls system, which could be involved in iron uptake. We therefore analyzed S. Typhimurium LSP 146/02, a clinical strain selected as representative of the isolates carrying the plasmid, and an otherwise isogenic mutant lacking four genes (fetMP-flsDA) of the fetMP-fls region. Growth curves and determination of the intracellular iron content under iron-restricted conditions demonstrated that deletion of these genes impairs iron acquisition. Thus, under these conditions, the mutant grew significantly worse than the wild-type strain, its iron content was significantly lower, and it was outcompeted by the wild-type strain in competition assays. Importantly, the strain lacking the fetMP-flsDA genes was less invasive in cultured epithelial HeLa cells and replicated poorly upon infection of RAW264.7 macrophages. The genes were introduced into S. Typhimurium ATCC 14028, which lacks the FetMP-Fls system, and this resulted in increased growth under iron limitation as well as an increased ability to multiply inside macrophages. These findings indicate that the FetMP-Fls iron acquisition system exceeds the benefits conferred by the other high-affinity iron uptake systems carried by ATCC 14028 and LSP 146/02. We proposed that effective iron acquisition by this system in conjunction with antimicrobial resistance encoded from the same plasmid have greatly contributed to the epidemic success of S. Typhimurium isolates harboring pUO-StVR2.

Neurokinin-1 receptors located in human retinoblastoma cell lines: antitumor action of its antagonist, L-732,138.

PURPOSE: The authors have recently demonstrated that substance P and L-733,060 induce cell proliferation and cell inhibition, respectively, in human retinoblastoma cell lines. However, the presence of neurokinin-1 receptors has not been demonstrated in such cell lines, nor is it known whether other neurokinin-1 receptor antagonists exert antitumoral action against retinoblastoma cell lines. The purpose of this study was to demonstrate the presence of neurokinin-1 receptors in the human retinoblastoma cell lines WERI-Rb-1 and Y-79 and to study the growth inhibitory capacity of the neurokinin-1 receptor antagonist L-732,138 against those cell lines. The authors also sought to demonstrate that the administration of L-732,138 or L-733,060 induces apoptosis in retinoblastoma cells and that neurokinin-1 receptors and substance P are present in primary retinoblastoma. METHODS: Immunoblot analysis was used to determine neurokinin-1 receptors, and a Coulter counter was used to determine viable cell numbers; this was followed by application of the tetrazolium compound WST-8, a colorimetric method, to evaluate cell viability. DAPI stain was applied to assess chromatin condensation, characteristic of apoptosis, and immunoperoxidase was used to demonstrate neurokinin-1 receptors and substance P in eyes with primary retinoblastoma. RESULTS: Neurokinin-1 receptors were present in both retinoblastoma cell lines studied. Three identical bands (isoforms of approximately 33, 58, and 75 kDa) were observed in both cell lines. Moreover, L-732,138 inhibited the growth of both cell lines studied, with and without previous administration of substance P. This inhibition occurred in a dose-dependent manner, with the IC50 values of 60.47 microM for WERI-Rb1 and 56.78 microM for Y-79. Moreover, apoptosis was observed in both cell lines after the administration of L-732,138 or L-733,060. In fixed eyes with primary retinoblastoma, a high density of neurokinin-1 receptors was observed in tumor cells, whereas a very low number of such cells contained substance P. CONCLUSIONS: This study showed that the same isoforms of the neurokinin-1 receptor are present in human retinoblastoma cell lines WERI-Rb-1 and Y-79. Both L-732,138 and L-733,060 can induce apoptosis in these cell lines and therefore can act as antitumoral agents. Primary retinoblastoma specimens display neurokinin-1 receptor immunolabeling. These results suggest that the neurokinin-1 receptor may be a promising new target for the treatment of retinoblastoma.

Incidence and Genetic Bases of Nitrofurantoin Resistance in Clinical Isolates of Two Successful Multidrug-Resistant Clones of Salmonella enterica Serovar Typhimurium: Pandemic "DT 104" and pUO-StVR2.

In this study, the incidence and genetic bases of nitrofurantoin resistance were established for clinical isolates of two successful clones of Salmonella enterica serovar Typhimurium, the pandemic "DT 104" and the pUO-StVR2 clone. A total of 61 "DT 104" and 40 pUO-StVR2 isolates recovered from clinical samples during 2008-2014 and assigned to different phage types, were tested for nitrofurantoin susceptibility. As previously shown for older isolates, all newly tested pUO-StVR2 isolates were highly resistant to nitrofurantoin (minimal inhibitory concentration [MIC] of 128 µg/ml), while 42.6%, 24.6%, and 32.8% of the "DT 104" isolates were susceptible, showed intermediate resistance or were highly resistant, with MICs of 8, 64, and 128 µg/ml, respectively. The genetic bases of nitrofurantoin resistance were established by PCR amplification and sequencing of the nfsA and nfsB genes encoding oxygen-insensitive nitroreductases. pUO-StVR2 isolates shared identical alterations in both nfsA (IS1 inserted into the coding region) and nfsB (in frame duplication of two codons). "DT 104" isolates with intermediate or high resistance had a missense mutation affecting the start codon of nfsA, while a single resistant isolate carried an additional frameshift mutation affecting nfsB. Complementation studies, performed with wild-type nfsA and nfsB, cloned independently and together into low and high copy-number vectors, confirmed NfsA and NfsB as responsible for nitrofurantoin toxicity. The same alterations persisted along time in isolates of each clone belonging to different phage types. Accordingly, changes leading to nitrofurantoin resistance have probably occurred before phage type diversification.

[Cardiovascular risk factor progression in young males at 15-year follow-up in the General Military Academy of Zaragoza (AGEMZA) Study]. / Evolución de los factores de riesgo cardiovascular en jóvenes varones tras 15 años de seguimiento en el estudio Academia General Militar de Zaragoza (AGEMZA).

INTRODUCTION AND OBJECTIVES: The AGEMZA cohort comprises military men whose risk factors were studied in 1985 when they were 20 years old. As these men reached the age of 35 years, we investigated the stability of or changes in anthropometric measures, lipid levels and arterial pressure, and looked for interrelationships between any changes. METHODS: In 2000, we collected new data (by cross-sectional study) on body mass index (BMI), cholesterol, cholesterol fractions, triglycerides and blood pressure, which could be compared with the original data. Persistence or tracking was evaluated using standardized regression coefficients and odds for persistence within the same quintile. Current data were modelled using multivariate regression models. RESULTS: In the 250 subjects studied, significant changes were observed in the following variables: weight +12.1 kg, BMI +3.9 kg/m(2), cholesterol +68.0 mg/dL, HDL cholesterol -5.2 mg/dL, LDL cholesterol +57.9 mg/dL, and triglycerides +76.3 mg/dL. The degree of persistence was high for all variables, except for diastolic blood pressure. Persistence was most pronounced for BMI, cholesterol, and LDL cholesterol. The changes observed indicate an increase in cardiovascular risk that adds to the effect of aging. The change in lipid profile was mainly influenced by the increase in BMI experienced, whereas blood pressure was mainly influenced by the final BMI attained. In addition, being a current smoker was associated with worse cholesterol fractions and triglyceride levels. CONCLUSIONS: Cardiovascular risk factors increase during the third decade of the life. Early evaluation (after adolescence) enables the identification of individuals who will later be at an increased risk. Modifiable risk factors were identified, such as weight increase and smoking. Preventive measures should be designed for these groups.