Distribution and molecular evolution of the anti-CRISPR family AcrIF7.

Anti-clustered regularly interspaced short palindromic repeats (CRISPRs) are proteins capable of blocking CRISPR-Cas systems and typically their genes are located on mobile genetic elements. Since their discovery, numerous anti-CRISPR families have been identified. However, little is known about the distribution and sequence diversity of members within a family, nor how these traits influence the anti-CRISPR's function and evolution. Here, we use AcrIF7 to explore the dissemination and molecular evolution of an anti-CRISPR family. We uncovered 5 subclusters and prevalent anti-CRISPR variants within the group. Remarkably, AcrIF7 homologs display high similarity despite their broad geographical, ecological, and temporal distribution. Although mainly associated with Pseudomonas aeruginosa, AcrIF7 was identified in distinct genetic backgrounds indicating horizontal dissemination, primarily by phages. Using mutagenesis, we recreated variation observed in databases but also extended the sequence diversity of the group. Characterisation of the variants identified residues key for the anti-CRISPR function and other contributing to its mutational tolerance. Moreover, molecular docking revealed that variants with affected function lose key interactions with its CRISPR-Cas target. Analysis of publicly available data and the generated variants suggests that the dominant AcrIF7 variant corresponds to the minimal and optimal anti-CRISPR selected in the family. Our study provides a blueprint to investigate the molecular evolution of anti-CRISPR families.

Phage-plasmids: missed links between mobile genetic elements.

Phages and plasmids are discrete mobile genetic elements (MGEs) with critical roles in gene dissemination across bacteria but limited scope for exchanging DNA between them. By investigating recent gene-sharing events, Pfeifer and Rocha describe how the hybrid elements phage-plasmids (P-Ps) promote gene flow between MGE types and evolve into new ones.

Impact of transient acquired hypermutability on the inter- and intra-species competitiveness of Pseudomonas aeruginosa.

Once acquired, hypermutation is unrelenting, and in the long-term, leads to impaired fitness due to its cumulative impact on the genome. This raises the question of why hypermutators arise so frequently in microbial ecosystems. In this work, we explore this problem by examining how the transient acquisition of hypermutability affects inter- and intra-species competitiveness, and the response to environmental insults such as antibiotic challenge. We do this by engineering Pseudomonas aeruginosa to allow the expression of an important mismatch repair gene, mutS, to be experimentally controlled over a wide dynamic range. We show that high levels of mutS expression induce genomic stasis (hypomutation), whereas lower levels of induction lead to progressively higher rates of mutation. Whole-genome sequence analyses confirmed that the mutational spectrum of the inducible hypermutator is similar to the distinctive profile associated with mutS mutants obtained from the airways of people with cystic fibrosis (CF). The acquisition of hypermutability conferred a distinct temporal fitness advantage over the wild-type P. aeruginosa progenitor strain, in both the presence and the absence of an antibiotic selection pressure. However, over a similar time-scale, acquisition of hypermutability had little impact on the population dynamics of P. aeruginosa when grown in the presence of a competing species (Staphylococcus aureus). These data indicate that in the short term, acquired hypermutability primarily confers a competitive intra-species fitness advantage.

Decreased efficacy of antimicrobial agents in a polymicrobial environment.

The airways of people with cystic fibrosis (CF) often harbour diverse polymicrobial communities. These airway infections can be impossible to resolve through antibiotic intervention, even though isolates of the individual species present are susceptible to the treatment when tested in vitro. In this work, we investigate how polymicrobial cultures comprised of key CF-associated pathogens respond to challenge with species-specific antimicrobial agents; colistin (targets Pseudomonas aeruginosa), fusidic acid (targets Staphylococcus aureus), and fluconazole (targets Candida albicans). We found that growth in a polymicrobial environment protects the target microorganism (sometimes by several orders of magnitude) from the effect(s) of the antimicrobial agent. This decreased antimicrobial efficacy was found to have both non-heritable (physiological) and heritable (genetic) components. Whole-genome sequencing of the colistin-resistant P. aeruginosa isolates revealed single nucleotide polymorphisms and indels in genes encoding lipopolysaccharide (LPS) biosynthesis and/or pilus biogenesis, indicating that a previously undescribed colistin resistance mechanism was in operation. This was subsequently confirmed through further genetic analyses. Our findings indicate that the polymicrobial nature of the CF airways is likely to have a significant impact on the clinical response to antimicrobial therapy.

A Novel Group of Promiscuous Podophages Infecting Diverse Gammaproteobacteria from River Communities Exhibits Dynamic Intergenus Host Adaptation.

Phages are generally described as species specific or even strain specific, implying an inherent limitation for some to be maintained and spread in diverse bacterial communities. Moreover, phage isolation and host range determination rarely consider the phage ecological context, likely biasing our notion on phage specificity. Here we isolated and characterized a novel group of six promiscuous phages, named Atoyac, existing in rivers and sewage by using a diverse collection of over 600 bacteria retrieved from the same environments as potential hosts. These podophages isolated from different regions in Mexico display a remarkably broad host range, infecting bacteria from six genera: Aeromonas, Pseudomonas, Yersinia, Hafnia, Escherichia, and Serratia Atoyac phage genomes are ∼42 kb long and highly similar to each other, but not to those currently available in genome and metagenome public databases. Detailed comparison of the phages' efficiency of plating (EOP) revealed variation among bacterial genera, implying a cost associated with infection of distant hosts, and between phages, despite their sequence similarity. We show, through experimental evolution in single or alternate hosts of different genera, that efficiency of plaque production is highly dynamic and tends toward optimization in hosts rendering low plaque formation. However, adaptation to distinct hosts differed between similar phages; whereas one phage optimized its EOP in all tested hosts, the other reduced plaque production in one host, suggesting that propagation in multiple bacteria may be key to maintain promiscuity in some viruses. Our study expands our knowledge of the virosphere and uncovers bacterium-phage interactions overlooked in natural systems.IMPORTANCE In natural environments, phages coexist and interact with a broad variety of bacteria, posing a conundrum for narrow-host-range phage maintenance in diverse communities. This context is rarely considered in the study of host-phage interactions, typically focused on narrow-host-range viruses and their infectivity in target bacteria isolated from sources distinct to where the phages were retrieved from. By studying phage-host interactions in bacteria and viruses isolated from river microbial communities, we show that novel phages with promiscuous host range encompassing multiple bacterial genera can be found in the environment. Assessment of hundreds of interactions in diverse hosts revealed that similar phages exhibit different infection efficiency and adaptation patterns. Understanding host range is fundamental in our knowledge of bacterium-phage interactions and their impact on microbial communities. The dynamic nature of phage promiscuity revealed in our study has implications in different aspects of phage research such as horizontal gene transfer or phage therapy.

Virulence and antimicrobial resistance genes are enriched in the plasmidome of clinical Escherichia coli isolates compared with wastewater isolates from western Kenya.

Many low-middle income countries in Africa have poorly-developed infectious disease monitoring systems. Here, we employed whole genome sequencing (WGS) to investigate the presence/absence of antimicrobial resistance (AMR) and virulence-associated (VA) genes in a collection of clinical and municipal wastewater Escherichia coli isolates from Kakamega, west Kenya. We were particularly interested to see whether, given the association between infection and water quality, the isolates from these geographically-linked environments might display similar genomic signatures. Phylogenetic analysis based on the core genes common to all of the isolates revealed two broad divisions, corresponding to the commensal/enterotoxigenic E. coli on the one hand, and uropathogenic E. coli on the other. Although the clinical and wastewater isolates each contained a very similar mean number of antibiotic resistance-encoding genes, the clinical isolates were enriched in genes required for in-host survival. Furthermore, and although the chromosomally encoded repertoire of these genes was similar in all sequenced isolates, the genetic composition of the plasmids from clinical and wastewater E. coli was more habitat-specific, with the clinical isolate plasmidome enriched in AMR and VA genes. Intriguingly, the plasmid-borne VA genes were often duplicates of genes already present on the chromosome, whereas the plasmid-borne AMR determinants were more specific. This reinforces the notion that plasmids are a primary means by which infection-related AMR and VA-associated genes are acquired and disseminated among these strains.

Selection of Functional Quorum Sensing Systems by Lysogenic Bacteriophages in Pseudomonas aeruginosa.

Quorum sensing (QS) in Pseudomonas aeruginosa coordinates the expression of virulence factors, some of which are used as public goods. Since their production is a cooperative behavior, it is susceptible to social cheating in which non-cooperative QS deficient mutants use the resources without investing in their production. Nevertheless, functional QS systems are abundant; hence, mechanisms regulating the amount of cheating should exist. Evidence that demonstrates a tight relationship between QS and the susceptibility of bacteria against the attack of lytic phages is increasing; nevertheless, the relationship between temperate phages and QS has been much less explored. Therefore, in this work, we studied the effects of having a functional QS system on the susceptibility to temperate bacteriophages and how this affects the bacterial and phage dynamics. We find that both experimentally and using mathematical models, that the lysogenic bacteriophages D3112 and JBD30 select QS-proficient P. aeruginosa phenotypes as compared to the QS-deficient mutants during competition experiments with mixed strain populations in vitro and in vivo in Galleria mellonella, in spite of the fact that both phages replicate better in the wild-type background. We show that this phenomenon restricts social cheating, and we propose that temperate phages may constitute an important selective pressure toward the conservation of bacterial QS.

Perspectivas y garantía de atención de salud mental en Ecuador / Mental Health Care Perspectives and Guarantees in Ecuador

Introducción: La salud mental constituye uno de los principales objetivos de la Organización Mundial de la Salud. Objetivo: Identificar la perspectiva individual sobre la salud mental y las garantías de atención de la población. Métodos: Se realizó una investigación básica, con un enfoque mixto y alcance descriptivo consistente en la aplicación del cuestionario denominado Kognitive: Estudio de Pertinencia. El universo estuvo constituido por 381 personas que asisten a servicios de salud mental privados del Distrito Metropolitano de Quito. La muestra quedó constituida por 340 personas. Resultados: Promedio de edad de 53,21 años, predominio de pacientes entre 40 y 49 años (55 por ciento), del sexo femenino (57,94 por ciento), solteros (56,18 por ciento) y mestizos (94,12 por ciento). El 55,59 por ciento no contaba con trabajo estable y 44,41 por ciento no contaba con seguro de salud. El 69,41 por ciento de las personas desconocían sobre la presencia de una red de servicios de salud mental (78,24 por ciento), 95,88 por ciento consideraba importante poseer cobertura de salud mental y el 93,24 por ciento mostraron interés por estos servicios. La falta de recursos económicos (56,18 por ciento) y la deficiente información (36,47 por ciento) fueron referidas como las causas más frecuentes de no asistencia a servicios de salud mental. Conclusiones: Las condiciones socioestructurales actuales no garantizan el adecuado estado de salud mental de la población; existe la necesidad de formar profesionales de salud capacitados y aumentar las redes de servicios y el acceso de la población a este tipo de servicios(AU)

Introduction: Mental health is one of the main goals of the World Health Organization. Objective: To identify individual perspective on mental health and the guarantees of population care. Methods: A basic investigation was carried out, with a mixed approach and a descriptive scope consisting in the application of the questionnaire called Kognitive: Pertinence Study. The universe was made up of 381 people who attend private mental health services in the Metropolitan District of Quito. The sample consisted of 340 people. Results: The average age of 53.21 years; there was a predominance of patients between 40 and 49 years (55 percent), of female sex (57.94 percent), single (56.18 percent) and of mixed race (94.12 percent). 55.59 percent did not have a stable job and 44.41 percent did not have health insurance. 69.41 percent of the people were unaware of the presence of a mental health services network (78.24 percent), 95.88 percent considered it important to have mental health coverage, and 93.24 percent showed interest in these services. The lack of financial resources (56.18 percent) and deficient information (36.47 percent) were mentioned as the most frequent causes for not attending mental health services. Conclusions: The current socio-structural conditions do not guarantee an adequate mental health state of the population. It is necessary to train health professionals in this field as well as increase service networks and population's access to these types of services(AU)