Genômica comparativa e filogenômica de Xylella fastidiosa

The gamma-proteobacterium Xylella fastidiosa is an insect-transmitted, xylem-inhabiting pathogen and the causal agent of several plant diseases, most notably Pierce\'s disease of grapes (PD), citrus variegated chlorosis (CVC) and olive quick decline syndrome (OQDS). The first two complete genomes of X. fastidiosa sequenced in the early 2000s were from CVC (9a5c) and PD (Temecula1) strains. Since then, genomes of various isolates of X. fastidiosa have been sequenced, which are very similar to each other regardless of the host and/or geographical region from which such strains were isolated. Despite the available genomes in public databases, potential determinants of host adaptation and the heterogenicity among the prophage regions are still unknown in a wide genomic level. In this study, the CDSs of 46 X. fastidiosa genomes were compared using the new computational tool GTACG that deals with phylogenetically close organisms. Also, in order to explore the content of Mobile Genetic Elements (MGE) we have analyzed predicted prophages, genomic islands and insertion sequences harbored in X. fastidiosa chromosome. A total of 4942 and 1518 orthologs were found in the pan- and core-genome of X. fastidiosa, respectively. The phylogenomic trees showed three main clades related with the subspecies pauca, fastidiosa and multiplex, and subclades most related with the predicted sequence type and geographic region of isolation while the plant host information had less relationship. Most of the virulence and pathogenicrelated orthologs were found in the core-genome of X. fastidiosa. In one case, the afimbrial adhesin orthologs XadA1 and XadA3 sequence diversities showed a relative congruence with the plant host. The MGE content per genome ranged from 12% to 28% in the 46 strains of X. fastidiosa analyzed. The mean of prophages and genomic island regions per genome were 8.6 and 8.9, respectively. Around half (56%) of predicted insertion sequences were located into prophage regions. In summary, the X. fastidiosa comparative genomics and phylogenomics analyses showed that the geographic region of isolation is strongly supported at the strains genomic level differently from strain-plant host adaptation. Moreover, a relevant and heterogeneous amount of MGE are harbored in the chromosome of all 46 X. fastidiosa strains analyzed.A gamaproteobacteria Xylella fastidiosa é transmitida por insetos, restrita ao xilema e agente causal de várias doenças de plantas, principalmente a doença de Pierce de videiras (PD), a clorose variegada dos citros (CVC) e a síndrome do rápido declínio de oliveiras (OQDS). Os dois primeiros genomas completos de X. fastidiosa sequenciados no início dos anos 2000 foram das cepas 9a5c e Temecula1, respectivamente causadoras de CVC e PD. Desde então, os genomas de vários isolados de X. fastidiosa foram sequenciados, os quais são muito semelhantes entre si, independentemente do hospedeiro e/ou da região geográfica de origem dessas cepas. Apesar da disponibilidade desses genomas em bancos de dados públicos, os potenciais determinantes da adaptação do hospedeiro e a heterogeneidade de regiões do prófagos ainda não foram analisados em nível genômico mais amplo. No presente estudo, as CDSs de 46 genomas de X. fastidiosa foram comparadas usando a nova ferramenta computacional GTACG dedicada à análise de genomas de organismos filogeneticamente próximos. Além disso, visando explorar o conteúdo de Elementos Genéticos Móveis (MGE), também foram analisadas as regiões preditas como prófagos, ilhas genômicas e sequências de inserção no cromossomo de X. fastidiosa. Foram encontrados 4942 e 1518 ortólogos no pan- e core-genoma de X. fastidiosa, respectivamente. As árvores filogenômicas mostraram os três clados principais relacionados às subespécies pauca, fastidiosa e multiplex, e os subclados foram mais relacionados ao sequence type predito e região geográfica de isolamento, enquanto as informações do hospedeiro vegetal tiveram menor relação. A maior parte dos ortólogos relacionados a virulência e patogenicidade foram encontrados no coregenoma de X. fastidiosa. Em um caso, a diversidade de sequência dos ortólogos das adesinas afimbriais XadA1 e XadA3 apresentou relativa congruência com o hospedeiro vegetal. O conteúdo de MGE por genoma variou de 12% a 28% nas 46 cepas de X. fastidiosa analisadas. A média de regiões de prófagos e ilhas genômicas por genoma foi de 8,6 e 8,9, respectivamente. Cerca de metade (56%) das sequências de inserção previstas foram localizadas em regiões de prófagos. Em resumo, as análises de genômica comparativa e de filogenômica de X. fastidiosa mostraram forte relação entre a região geográfica de isolamento e os distintos genomas, mas não com a adaptação cepa-hospedeiro vegetal. Além disso, verificamos que todos os 46 genomas de X. fastidiosa analisados contém um conteúdo relevante e heterogêneo de MGE

Actividad Bactericida de Superficies de Cobre sobre Bacterias Multirresistentes Asociados a Infecciones Intrahospitalarias

Objetivo. Evaluar la actividad bactericida de superficies de cobre sobre bacterias multirresistentes asociados a infecciones intrahospitalarias. Materiales y métodos: Estudio experimental, donde la población estuvo constituido por bacterias multirresistente y la muestra por nueve cepas bacterianas, 3 de Acinetobacter baumannii, 3 de Pseudomonas aeruginosa y 3 de 2 Staphylococcus aureus; los mismos que fueron inoculadas sobre láminas metálicas de 1cm de cobre y acero inoxidable e incubados a temperatura ambiente durante 0, 5, 10, 20, 40 y 60 minutos para luego determinar la sobrevivencia en Mueller Hinton a través del recuento de unidades formadoras de colonias. Resultados. Las cepas de A. baumannii fueron las más sensible al cobre mientras que S. aureus la más resistente. Sin embargo todas las cepas permaneciendo viables a superficies de acero inoxidable hasta los 60 minutos de exposición. Hubo diferencias estadistas significativas (P<0,05) entre los promedios de las variables metal, cepa y tiempo; existiendo diferencias en el efecto de ambos metales, crecimiento de unidades formadoras de colonias durante el tiempo de exposición y diferencias en la respuesta de cada tipo de bacteria al contacto con superficies de cobre. Conclusiones. Las superficies del cobre tiene efecto bactericida al disminuir las unidades formadoras de colonias de Acinetobacter baumannii, Pseudomonas aeruginosa y Staphylococcus aureus a medida que aumenta el tiempo de exposición

Unraveling the diversity of prokaryotic immune systems by in silico comparative analysis

&lt;p&gt;Bacteria and Archaea have evolved a variety of antiphage immune systems including the well-known Restriction-Modification (RM) and CRISPR-Cas, and several recently discovered systems. Nevertheless, which traits are shared between these systems and how their diversity is originated are underexplored questions. Thus, we sought to compare the immune system family diversity using a robust sequence similarity approach and a network to analysis. The immune systems were predicted using Defense-Finder followed by a robust sequence comparison of all-vs-all immune system sequences (instances) using HH-suite. A network was built, the properties of the network were determined, and the protein domains associated to the systems were identified. We obtained a network of 15,289 nodes (instances) distributed among 82 immune system families. The network highlighted several conserved traits shared between instances belonging to different families and one big cluster containing ~90% of the instances of 45 immune system families. These families comprise the most known family RM as well as the less widely distributed CBASS, BREX, Lamassu, Gabija, and Wadjet families. The network centrality analysis showed that the median of the instances belongs to RM type I, Hachiman and PsyrTA families being the most central in the network. In contrast, several of the newly reported families such as Thoeris, Druantia and Zorya among others are distributed in the network peripheral region. We found that more than 80% of the relationship revealed in the network reflect at least one known shared protein domain related to DNA metabolism such as helicases and nucleases. Our results show that behind the prokaryotic immune system diversity, there are several conserved traits. We also show the importance of RM type I to support the network of 45 immune systems, suggesting this type of RM could be key in the generation of the observed diversity.&lt;/p&gt

Transcriptome and Secretome Analyses of Endophyte Methylobacterium mesophilicum and Pathogen Xylella fastidiosa Interacting Show Nutrient Competition.

Xylella fastidiosa is the causal agent of several plant diseases affecting fruit and nut crops. Methylobacterium mesophilicum strain SR1.6/6 was isolated from Citrus sinensis and shown to promote plant growth by producing phytohormones, providing nutrients, inhibiting X. fastidiosa, and preventing Citrus Variegated Chlorosis. However, the molecular mechanisms involved in the interaction among these microbes are still unclear. The present work aimed to analyze physiological and molecular aspects of M. mesophilicum SR1.6/6 and X. fastidiosa 9a5c in co-culture. The transcriptome and secretome analyses indicated that X. fastidiosa down-regulates cell division and transport genes and up-regulates stress via induction of chaperones and pathogenicity-related genes including, the lipase-esterase LesA, a protease, as well as an oligopeptidase in response to M. mesophilicum competition. On the other hand, M. mesophilicum also down-regulated transport genes, except for iron uptake, which was up-regulated. Secretome analysis identified four proteins in M. mesophilicum exclusively produced in co-culture with X. fastidiosa, among these, three are related to phosphorous uptake. These results suggest that M. mesophilicum inhibits X. fastidiosa growth mainly due to nutrient competition for iron and phosphorous, thus promoting X. fastidiosa starvation, besides producing enzymes that degrade X. fastidiosa cell wall, mainly hydrolases. The understanding of these interactions provides a direction for control and management of the phytopathogen X. fastidiosa, and consequently, helps to improve citrus growth and productivity

Transcriptome and Secretome Analyses of Endophyte <i>Methylobacterium mesophilicum</i> and Pathogen <i>Xylella fastidiosa</i> Interacting Show Nutrient Competition

Xylella fastidiosa is the causal agent of several plant diseases affecting fruit and nut crops. Methylobacterium mesophilicum strain SR1.6/6 was isolated from Citrus sinensis and shown to promote plant growth by producing phytohormones, providing nutrients, inhibiting X. fastidiosa, and preventing Citrus Variegated Chlorosis. However, the molecular mechanisms involved in the interaction among these microbes are still unclear. The present work aimed to analyze physiological and molecular aspects of M. mesophilicum SR1.6/6 and X. fastidiosa 9a5c in co-culture. The transcriptome and secretome analyses indicated that X. fastidiosa down-regulates cell division and transport genes and up-regulates stress via induction of chaperones and pathogenicity-related genes including, the lipase-esterase LesA, a protease, as well as an oligopeptidase in response to M. mesophilicum competition. On the other hand, M. mesophilicum also down-regulated transport genes, except for iron uptake, which was up-regulated. Secretome analysis identified four proteins in M. mesophilicum exclusively produced in co-culture with X. fastidiosa, among these, three are related to phosphorous uptake. These results suggest that M. mesophilicum inhibits X. fastidiosa growth mainly due to nutrient competition for iron and phosphorous, thus promoting X. fastidiosa starvation, besides producing enzymes that degrade X. fastidiosa cell wall, mainly hydrolases. The understanding of these interactions provides a direction for control and management of the phytopathogen X. fastidiosa, and consequently, helps to improve citrus growth and productivity

The XadA trimeric autotransporter adhesins in Xylella fastidiosa differentially contribute to cell aggregation, biofilm formation, insect transmission and virulence to plants.

Surface adhesion strategies are widely employed by bacterial pathogens during establishment and systemic spread in their host. A variety of cell surface appendages such as pili, fimbriae and afimbrial adhesins are involved in these processes. The phytopathogen Xylella fastidiosa employs several of these structures for efficient colonization of its insect and plant hosts. Among the adhesins encoded in the X. fastidiosa genome, three afimbrial adhesins, XadA1, Hsf/XadA2, and XadA3, are predicted to be trimeric autotransporters with a C-terminal YadA-anchor membrane domain. We analyzed the individual contributions of XadA1, XadA2, and XadA3 to various cellular behaviors both in vitro and in vivo. Using isogenic X. fastidiosa mutants, we found that cell-cell aggregation and biofilm formation were severely impaired in the absence of XadA3. No significant reduction of cell-surface attachment was found with any mutant under flow conditions. Acquisition by insect vectors and transmission to grapevines were reduced in the XadA3 deletion mutant. While the XadA3 mutant was hypervirulent in grapevines, XadA1 or XadA2 deletion mutants conferred lower disease severity than the wild-type strain. This insight of the importance of these adhesive proteins and their individual contributions to different aspects of X. fastidiosa biology should guide new approaches to reduce pathogen transmission and disease development

Comparative Genomics of <i>Xylella fastidiosa</i> Explores Candidate Host-Specificity Determinants and Expands the Known Repertoire of Mobile Genetic Elements and Immunity Systems

Xylella fastidiosa causes diseases in many plant species. Originally confined to the Americas, infecting mainly grapevine, citrus, and coffee, X. fastidiosa has spread to several plant species in Europe causing devastating diseases. Many pathogenicity and virulence factors have been identified, which enable the various X. fastidiosa strains to successfully colonize the xylem tissue and cause disease in specific plant hosts, but the mechanisms by which this happens have not been fully elucidated. Here we present thorough comparative analyses of 94 whole-genome sequences of X. fastidiosa strains from diverse plant hosts and geographic regions. Core-genome phylogeny revealed clades with members sharing mostly a geographic region rather than a host plant of origin. Phylogenetic trees for 1605 orthologous CDSs were explored for potential candidates related to host specificity using a score of mapping metrics. However, no candidate host-specificity determinants were strongly supported using this approach. We also show that X. fastidiosa accessory genome is represented by an abundant and heterogeneous mobilome, including a diversity of prophage regions. Our findings provide a better understanding of the diversity of phylogenetically close genomes and expand the knowledge of X. fastidiosa mobile genetic elements and immunity systems

Molecular Basis of Histone H3K4me3 Recognition by ING4*S⃞

The inhibitors of growth (ING) family of tumor suppressors consists of five homologous proteins involved in chromatin remodeling. They form part of different acetylation and deacetylation complexes and are thought to direct them to specific regions of the chromatin, through the recognition of H3K4me3 (trimethylated K4 in the histone 3 tail) by their conserved plant homeodomain (PHD). We have determined the crystal structure of ING4-PHD bound to H3K4me3, which reveals a tight complex stabilized by numerous interactions. NMR shows that there is a reduction in the backbone mobility on the regions of the PHD that participate in the peptide binding, and binding affinities differ depending on histone tail lengths Thermodynamic analysis reveals that the discrimination in favor of methylated lysine is entropy-driven, contrary to what has been described for chromodomains. The molecular basis of H3K4me3 recognition by ING4 differs from that of ING2, which is consistent with their different affinities for methylated histone tails. These differences suggest a distinct role in transcriptional regulation for these two ING family members because of the antagonistic effect of the complexes that they recruit onto chromatin. Our results illustrate the versatility of PHD fingers as readers of the histone code