RESUMO

Tank bromeliads are unique canopy microhabitats that offer freshwater and organic nutrient-rich substrates in the Neotropics. In them it is possible to thoroughly characterize environmental factors and species composition of terrestrial and aquatic biota. Therefore, these plants have been used as natural models to study how communities are distributed and assembled. Here we used amplicon sequencing of the 16S rRNA gene and their functional annotations to study the diversity and metabolic potential of prokaryotic communities in tank bromeliads in five different forests along an elevation gradient in tropical Mexico. Furthermore, we analyzed the effects of vegetation type and environmental factors inside the tanks on prokaryotic composition. We found a high prokaryotic diversity in tank bromeliads along the elevation gradient. Prokaryotes commonly observed in acidic environments rich in organic carbon, and the potential pathogen Pasteurella multocida, were present in all samples, but few amplicon sequence variants were shared between forests. The prokaryotic composition was affected by forest type, and comparisons against null models suggest that it was shaped by non-neutral processes. Furthermore, prokaryotic community changes significantly covaried with tank water temperature, pH, and inorganic carbon. We found a high diversity of putative metabolic groups dominated by chemoheterotrophs and fermenters, but taxonomic groups involved in nitrogen and sulfur cycling were also present in all samples. These results suggest that tank bromeliads promote taxonomic and metabolic diversity of the prokaryotic community at a local and regional scale and play an important role in the biogeochemistry of forest canopies in the Neotropics.

RESUMO

Blueberry production is affected by fungal postharvest pathogens, including Botrytis cinerea and Alternaria alternata, the causative agents of gray mold disease and Alternaria rot, respectively. Biocontrol agents adapted to blueberries and local environments are not known to date. Here, we report on the search for and the identification of cultivable blueberry epiphytic bacteria with the potential to combat the aforementioned fungi. Native, blueberry-borne bacterial strains were isolated from a plantation in Tucumán, Argentina and classified based on 16S rRNA gene sequences. Antagonistic activities directed at B. cinerea and A. alternata were studied in vitro and in vivo. The 22 bacterial strains obtained could be attributed to eleven different genera: Rosenbergiella, Fictibacillus, Bacillus, Pseudomonas, Microbacterium, Asaia, Acinetobacter, Curtobacterium, Serratia, Sphingomonas and Xylophilus. Three strains displaying antagonistic impacts on the fungal pathogens were identified as Bacillus velezensis (BA3 and BA4) and Asaia spathodeae (BMEF1). These strains are candidates for biological control agents of local blueberry production and might provide a basis for the development of eco-friendly, sustainable alternatives to synthetic pesticides.

RESUMO

Diamante Lake located at 4589 m.a.s.l. in the Andean Puna constitutes an extreme environment. It is exposed to multiple extreme conditions such as an unusually high concentration of arsenic (over 300 mg L-1) and low oxygen pressure. Microorganisms thriving in the lake display specific genotypes that facilitate survival, which include at least a multitude of plasmid-encoded resistance traits. Hence, the genetic information provided by the plasmids essentially contributes to understand adaptation to different stressors. Though plasmids from cultivable organisms have already been analyzed to the sequence level, the impact of the entire plasmid-borne genetic information on such microbial ecosystem is not known. This study aims at assessing the plasmidome from Diamante Lake, which facilitates the identification of potential hosts and prediction of gene functions as well as the ecological impact of mobile genetic elements. The deep-sequencing analysis revealed a large fraction of previously unknown DNA sequences of which the majority encoded putative proteins of unknown function. Remarkably, functions related to the oxidative stress response, DNA repair, as well as arsenic- and antibiotic resistances were annotated. Additionally, all necessary capacities related to plasmid replication, mobilization and maintenance were detected. Sequences characteristic for megaplasmids and other already known plasmid-associated genes were identified as well. The study highlights the potential of the deep-sequencing approach specifically targeting plasmid populations as it allows to evaluate the ecological impact of plasmids from (cultivable and non-cultivable) microorganisms, thereby contributing to the understanding of the distribution of resistance factors within an extremophilic microbial community.

Assuntos

Bactérias/genética , DNA Bacteriano/análise , Extremófilos/genética , Lagos/microbiologia , Microbiota , Plasmídeos/análise , Bactérias/classificação , Bactérias/crescimento & desenvolvimento , Bactérias/isolamento & purificação , DNA Bacteriano/genética , Farmacorresistência Bacteriana , Extremófilos/crescimento & desenvolvimento , Extremófilos/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala , Filogenia , Plasmídeos/genética , Plasmídeos/isolamento & purificação , Esgotos/microbiologia

RESUMO

Klebsiella pneumoniae carbapenemase (KPC) actively hydrolyzes carbapenems, antibiotics often used a last-line treatment for multidrug-resistant bacteria. KPC clinical relevance resides in its widespread dissemination. In this work, we report the genomic context of KPC coding genes blaKPC-2, blaKPC-3 and blaKPC-30 in multidrug-resistant Klebsiellapneumoniae isolates from Brazil. Plasmids harboring blaKPC-3 and blaKPC-30 were identified. Fifteen additional carbapenem-resistant K. pneumoniae isolates were selected from the same tertiary hospital, collected over a period of 8 years. Their genomes were sequenced in order to evaluate the prevalence and dissemination of blaKPC-harboring plasmids. We found that blaKPC genes were mostly carried by one of two isoforms of transposon Tn4401 (Tn4401a or Tn4401b) that were predominantly located on plasmids highly similar to the previously described plasmid pKPC_FCF3SP (IncN). The identified pKPC_FCF3SP-like plasmids carried either blaKPC-2 or blaKPC-30. Two K. pneumoniae isolates harbored pKpQIL-like (IncFII) plasmids, only recently identified in Brazil; one of them harbored blaKPC-3 in a Tn4401a transposon. Underlining the risk of horizontal spread of KPC coding genes, this study reports the prevalence of blaKPC-2 and the recent spread of blaKPC-3, and blaKPC-30, in association with different isoforms of Tn4401, together with high synteny of plasmid backbones among isolates studied here and in comparison with previous reports.

RESUMO

Worldwide, the green rot caused by Penicillium digitatum is one of the most aggressive postharvest diseases of lemons. Searching for sustainable alternatives to chemical fungicides, epiphytic yeasts as potential biocontrol agents were isolated from citrus fruits using a tailor-made selective medium. For disclosing their antagonistic potential against P. digitatum, obtained isolates were subjected to direct screening methods, both in vitro and in vivo. In the course of the primary in vitro screening that comprised dual culture assays, 43 yeast strains displaying antagonistic activities against the pathogen were selected. Subsequently, such strains were subjected to an in vivo screening that consisted of a microscale test, allowing the selection of six yeast strains for further analysis. In the final screening using macroscale in vivo tests, three strains (AcL2, AgL21, and AgL2) displaying the highest efficiencies to control P. digitatum were identified. The protection efficiencies in lemons were 80 (AcL2), 76.7 (AgL21), and 75% (AgL2). Based on sequence analysis of the PCR amplified D1/D2 domains of the 26S rRNA genes, they were identified as representatives of the species Clavispora lusitaniae. Interestingly, the strains exhibited a broad action spectrum among citrus fruits as they were also able to combat the green mold disease in grapefruit and two orange varieties. The direct screening methods applied in this study favored the recovery of efficient candidates for application as biological control agents to combat fungal infestations of citrus fruits.

RESUMO

Mobile genetic elements, including plasmids, drive the evolution of prokaryotic genomes through the horizontal transfer of genes allowing genetic exchange between bacteria. Moreover, plasmids carry accessory genes, which encode functions that may offer an advantage to the host. Thus, it is expected that in a certain ecological niche, plasmids are enriched in accessory functions, which are important for their hosts to proliferate in that niche. Puquio de Campo Naranja is a high-altitude lake from the Andean Puna exposed to multiple extreme conditions, including high UV radiation, alkalinity, high concentrations of arsenic, heavy metals, dissolved salts, high thermal amplitude and low O2 pressure. Microorganisms living in this lake need to develop efficient mechanisms and strategies to cope under these conditions. The aim of this study was to characterize the plasmidome of microbialites from Puquio de Campo Naranja, and identify potential hosts and encoded functions using a deep-sequencing approach. The potential ecological impact of the plasmidome, including plasmids from cultivable and non-cultivable microorganisms, is described for the first time in a lake representing an extreme environment of the Puna. This study showed that the recovered genetic information for the plasmidome was novel in comparison to the metagenome derived from the same environment. The study of the total plasmid population allowed the identification of genetic features typically encoded by plasmids, such as resistance and virulence factors. The resistance genes comprised resistances to heavy metals, antibiotics and stress factors. These results highlight the key role of plasmids for their hosts and impact of extrachromosomal elements to thrive in a certain ecological niche.

RESUMO

Pseudomonas aeruginosa is an important opportunistic pathogen in hospitals, responsible for various infections that are difficult to treat due to intrinsic and acquired antibiotic resistance. Here, 20 epidemiologically unrelated strains isolated from patients in a general hospital over a time period of two decades were analyzed using whole genome sequencing. The genomes were compared in order to assess the presence of a predominant clone or sequence type (ST). No clonal structure was identified, but core genome-based single nucleotide polymorphism (SNP) analysis distinguished two major, previously identified phylogenetic groups. Interestingly, most of the older strains isolated between 1994 and 1998 harbored exoU, encoding a cytotoxic phospholipase. In contrast, most strains isolated between 2011 and 2016 were exoU-negative and phylogenetically very distinct from the older strains, suggesting a population shift of nosocomial P. aeruginosa over time. Three out of 20 strains were ST235 strains, a global high-risk clonal lineage; these carried several additional resistance determinants including aac(6')Ib-cr encoding an aminoglycoside N-acetyltransferase that confers resistance to fluoroquinolones. Core genome comparison with ST235 strains from other parts of the world showed that the three strains clustered together with other Brazilian/Argentinean isolates. Despite this regional relatedness, the individuality of each of the three ST235 strains was revealed by core genome-based SNPs and the presence of genomic islands in the accessory genome. Similarly, strain-specific characteristics were detected for the remaining strains, indicative of individual evolutionary histories and elevated genome plasticity.

Assuntos

Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/classificação , Pseudomonas aeruginosa/genética , Brasil , Genoma Bacteriano , Humanos , Filogenia , Infecções por Pseudomonas/patologia , Pseudomonas aeruginosa/isolamento & purificação , Análise de Sequência de DNA

RESUMO

The two linear plasmids pLMA1 (109,112 bp) and pLMA7 (82,075 bp) from Micrococcus strains were isolated from a high-altitude lake in the Argentinean Puna, sequenced, and annotated. These extrachromosomal elements are probably conjugative and harbor genes potentially involved in coping with the harsh conditions in such extreme environments.