Expression, purification and characterization of the transcription termination factor Rho from Azospirillum brasilense.

The Transcription Termination factor Rho is a ring-shaped, homohexameric protein that causes transcript termination by interaction with specific sites on nascent mRNAs. The process of transcription termination is essential for proper expression and regulation of bacterial genes. Although Rho has been extensively studied in the model bacteria Escherichia coli (EcRho), the properties of other Rho orthologues in other bacteria are poorly characterized. Here we present the heterologous expression and purification of untagged Rho protein from the diazotrophic environmental bacterium Azospirillum brasilense (AbRho). The AbRho protein was purified to >99% through a simple, reproducible and efficient purification protocol, a two-step chromatography procedure (affinity/gel filtration). By using analytical gel filtration and dynamic light scattering (DLS), we found that AbRho is arranged as an homohexamer as observed in the EcRho orthologue. Secondary structure and enzyme activity of AbRho was also evaluate indicating a properly folded and active protein after purification. Enzymatic assays indicate that AbRho is a RNA-dependent NTPase enzyme.

Genomic landscape of the SARS-CoV-2 pandemic in Brazil suggests an external P.1 variant origin.

Brazil was the epicenter of worldwide pandemics at the peak of its second wave. The genomic/proteomic perspective of the COVID-19 pandemic in Brazil could provide insights to understand the global pandemics behavior. In this study, we track SARS-CoV-2 molecular information in Brazil using real-time bioinformatics and data science strategies to provide a comparative and evolutive panorama of the lineages in the country. SWeeP vectors represented the Brazilian and worldwide genomic/proteomic data from Global Initiative on Sharing Avian Influenza Data (GISAID) between February 2020 and August 2021. Clusters were analyzed and compared with PANGO lineages. Hierarchical clustering provided phylogenetic and evolutionary analyses of the lineages, and we tracked the P.1 (Gamma) variant origin. The genomic diversity based on Chao's estimation allowed us to compare richness and coverage among Brazilian states and other representative countries. We found that epidemics in Brazil occurred in two moments with different genetic profiles. The P.1 lineages emerged in the second wave, which was more aggressive. We could not trace the origin of P.1 from the variants present in Brazil. Instead, we found evidence pointing to its external source and a possible recombinant event that may relate P.1 to a B.1.1.28 variant subset. We discussed the potential application of the pipeline for emerging variants detection and the PANGO terminology stability over time. The diversity analysis showed that the low coverage and unbalanced sequencing among states in Brazil could have allowed the silent entry and dissemination of P.1 and other dangerous variants. This study may help to understand the development and consequences of variants of concern (VOC) entry.

Population-Based Analysis of the Immunoglobulin G Response to Different COVID-19 Vaccines in Brazil.

(1) Background: COVID-19 vaccination in Brazil has been performed mostly with CoronaVac (Sinovac), ChAdOx1-S (AstraZeneca-University of Oxford) and BNT162b2 (Pfizer-BioNTech) vaccines. The titers of IgG antibodies reactive to the SARS-CoV-2 spike protein correlate with vaccine efficacy. Studies comparing vaccine immunogenicity in a real-world scenario are lacking. (2) Methods: We performed a population-based study to analyze the immunoglobulin G response to different COVID-19 vaccines. Citizens older than 18 years (n = 2376) provided personal data, a self-declaration of any previous COVID-19 positive tests and information regarding COVID-19 vaccination: the vaccine popular name and the date of each dose. Blood samples were collected and the levels of IgG reactive to SARS-CoV-2 antigens were determined and compared between different vaccine groups. (3) Results: The seroconversion for anti-spike IgG achieved > 95% by February 2022 and maintained stable until June 2022. Higher anti-spike IgG titers were detected in individuals vaccinated with BNT162b2, followed by ChAdOx1-S and CoronaVac. The anti-spike IgG response was negatively correlated with age and interval after the second dose for the BNT162b2 vaccine. Natural infections boosted anti-spike IgG in those individuals who completed primary vaccination with ChAdOx1-S and CoronaVac, but not with BNT162b2. The levels of anti-spike IgG increased with the number of vaccine doses administered. The application of BNT162b2 as a 3rd booster dose resulted in high anti-spike IgG antibody titers, despite the type of vaccine used during primary vaccination. (4) Conclusions: Our data confirmed the effectiveness of the Brazilian vaccination program. Of the vaccines used in Brazil, BNT162b2 performed better to elicit anti-spike protein IgG after primary vaccination and as a booster dose and thus should be recommended as a booster whenever available. A continuous COVID-19 vaccination program will be required to sustain anti-spike IgG antibodies in the population.

The microbiome of a shell mound: ancient anthropogenic waste as a source of Streptomyces degrading recalcitrant polysaccharides.

Metagenome amplicon DNA sequencing and traditional cell culture techniques are helping to uncover the diversity and the biotechnological potential of prokaryotes in different habitats around the world. It has also had a profound impact on microbial taxonomy in the last decades. Here we used metagenome 16S rDNA amplicon sequencing to reveal the microbiome composition of different layers of an anthropogenic soil collected at a shell mound Sambaqui archeological site. The Samabaqui soil microbiome is mainly composed by phyla Acidobacteria, Rokubacteria, Proteobacteria and Thaumarchaeota. Using culture-dependent analysis we obtained few Streptomyces isolates from the Sambaqui soil. One of the isolates, named Streptomyces sp. S3, was able to grow in minimal medium containing recalcitrant polysaccharides including chitin, xylan, carboxymethylcellulose or microcrystalline cellulose as sole carbon sources. The activities of enzymes degrading these compounds were confirmed in cell free supernatants. The genome sequence revealed not only an arsenal of genes related to polysaccharides degradation but also biosynthetic gene clusters which may be involved in the production of biotechnologically interesting secondary metabolites.

Re-emergence of Gamma-like-II and emergence of Gamma-S:E661D SARS-CoV-2 lineages in the south of Brazil after the 2021 outbreak.

BACKGROUND: We report a genomic surveillance of SARS-CoV-2 lineages circulating in Paraná, southern Brazil, from March 2020 to April 2021. Our analysis, based on 333 genomes, revealed that the first variants detected in the state of Paraná in March 2020 were the B.1.1.33 and B.1.1.28 variants. The variants B.1.1.28 and B.1.1.33 were predominant throughout 2020 until the introduction of the variant P.2 in August 2020 and a variant of concern (VOC), Gamma (P.1), in January 2021. The VOC Gamma, a ramification of the B.1.1.28 lineage first detected in Manaus (northern Brazil), has grown rapidly since December 2020 and was thought to be responsible for the deadly second wave of COVID-19 throughout Brazil. METHODS: The 333 genomic sequences of SARS-CoV-2 from March 2020 to April 2021 were generated as part of the genomic surveillance carried out by Fiocruz in Brazil Genomahcov Fiocruz. SARS-CoV-2 sequencing was performed using representative samples from all geographic areas of Paraná. Phylogenetic analyses were performed using the 333 genomes also included other SARS-CoV-2 genomes from the state of Paraná and other states in Brazil that were deposited in the GISAID. In addition, the time-scaled phylogenetic tree was constructed with up to 3 random sequences of the Gamma variant from each state in Brazil in each month of 2021. In this analysis we also added the sequences identified as the B.1.1.28 lineage of the Amazonas state and and the Gamma-like-II (P.1-like-II) lineage identified in different regions of Brazil. RESULTS: Phylogenetic analyses of the SARS-CoV-2 genomes that were previously classified as the VOC Gamma lineage by WHO/PANGO showed that some genomes from February to April 2021 branched in a monophyletic clade and that these samples grouped together with genomes recently described with the lineage Gamma-like-II. Additionally, a new mutation (E661D) in the spike (S) protein has been identified in nearly 10% of the genomes classified as the VOC Gamma from Paraná in March and April 2021.Finally, we analyzed the correlation between the lineage and the Gamma variant frequency, age group (patients younger or older than 60 years old) and the clinical data of 86 cases from the state of Paraná. CONCLUSIONS: Our results provided a reliable picture of the evolution of the SARS-CoV-2 pandemic in the state of Paraná characterized by the dominance of the Gamma strain, as well as a high frequencies of the Gamma-like-II lineage and the S:E661D mutation. Epidemiological and genomic surveillance efforts should be continued to unveil the biological relevance of the novel mutations detected in the VOC Gamma in Paraná.

The protein-protein interaction network of the Escherichia coli EIIANtr regulatory protein reveals a role in cell motility and metabolic control.

The nitrogen-related PTSNtr system, present in many Proteobacteria including Escherichia coli, acts as a phosphorelay cascade composed of the EINtr, NPr and EIIANtr proteins. Phosphotransfer initiates with phosphoenolpyruvate-dependent EINtr autophosphorylation, the phosphoryl group is then transferred to NPr and finally to a conserved histidine residue on EIIANtr. The reporter metabolites l-glutamine and 2-oxoglutarate reciprocally regulate EINtr autophosphorylation (Lee et al., 2013) and consequently the phosphorylation status of the PTSNtr components is controlled by the availability of nitrogen and carbon. The final phosphate acceptor, EIIANtr, regulates a range of cellular process by acting as the central hub of a complex protein-protein interaction network. Contact between EIIANtr and its target proteins is usually regulated by the EIIANtr phosphorylation status. In this study we performed ligand fishing assays coupled to label-free quantitative proteomics to examine the protein-protein interaction network of E. coli EIIANtr and a phosphomimic variant of the protein. The ligand fishing data, along with phenotypic analysis, indicated that EIIANtr interacts with proteins related to chemotaxis and thereby regulates cell motility. Important metabolic enzymes were also identified as potential EIIANtr binding partners.

Antigen production and development of an indirect ELISA based on the nucleocapsid protein to detect human SARS-CoV-2 seroconversion.

Serological assays are important tools to identify previous exposure to SARS-CoV-2, helping to track COVID-19 cases and determine the level of humoral response to SARS-CoV-2 infections and/or immunization to future vaccines. Here, the SARS-CoV-2 nucleocapsid protein was expressed in Escherichia coli and purified to homogeneity and high yield using a single chromatography step. The purified SARS-CoV-2 nucleocapsid protein was used to develop an indirect enzyme-linked immunosorbent assay for the identification of human SARS-CoV-2 seroconverts. The assay sensitivity and specificity were determined analyzing sera from 140 RT-qPCR-confirmed COVID-19 cases and 210 pre-pandemic controls. The assay operated with 90% sensitivity and 98% specificity; identical accuracies were obtained in head-to-head comparison with a commercial ELISA kit. Antigen-coated plates were stable for up to 3 months at 4 °C. The ELISA method described is ready for mass production and will be an additional tool to track COVID-19 cases.

Transcriptional Responses of Herbaspirillum seropedicae to Environmental Phosphate Concentration.

Herbaspirillum seropedicae is a nitrogen-fixing endophytic bacterium associated with important cereal crops, which promotes plant growth, increasing their productivity. The understanding of the physiological responses of this bacterium to different concentrations of prevailing nutrients as phosphate (Pi) is scarce. In some bacteria, culture media Pi concentration modulates the levels of intracellular polyphosphate (polyP), modifying their cellular fitness. Here, global changes of H. seropedicae SmR1 were evaluated in response to environmental Pi concentrations, based on differential intracellular polyP levels. Cells grown in high-Pi medium (50 mM) maintained high polyP levels in stationary phase, while those grown in sufficient Pi medium (5 mM) degraded it. Through a RNA-seq approach, comparison of transcriptional profiles of H. seropedicae cultures revealed that 670 genes were differentially expressed between both Pi growth conditions, with 57% repressed and 43% induced in the high Pi condition. Molecular and physiological analyses revealed that aspects related to Pi metabolism, biosynthesis of flagella and chemotaxis, energy production, and polyhydroxybutyrate metabolism were induced in the high-Pi condition, while those involved in adhesion and stress response were repressed. The present study demonstrated that variations in environmental Pi concentration affect H. seropedicae traits related to survival and other important physiological characteristics. Since environmental conditions can influence the effectiveness of the plant growth-promoting bacteria, enhancement of bacterial robustness to withstand different stressful situations is an interesting challenge. The obtained data could serve not only to understand the bacterial behavior in respect to changes in rhizospheric Pi gradients but also as a base to design strategies to improve different bacterial features focusing on biotechnological and/or agricultural purposes.

Magnetic Bead-Based Immunoassay Allows Rapid, Inexpensive, and Quantitative Detection of Human SARS-CoV-2 Antibodies.

Immunological methods to detect SARS-CoV-2 seroconversion in humans are important to track COVID-19 cases and the humoral response to SARS-CoV-2 infections and immunization to future vaccines. The aim of this work was to develop a simple chromogenic magnetic bead-based immunoassay which allows rapid, inexpensive, and quantitative detection of human antibodies against SARS-CoV-2 in serum, plasma, or blood. Recombinant 6xHis-tagged SARS-CoV-2 Nucleocapsid protein was mobilized on the surface of Ni2+ magnetic beads and challenged with serum or blood samples obtained from controls or COVID-19 cases. The beads were washed, incubated with anti-human IgG-HPR conjugate, and immersed into a solution containing a chromogenic HPR substrate. Bead transfer and homogenization between solutions was aided by a simple low-cost device. The method was validated by two independent laboratories, and the performance to detect SARS-CoV-2 seroconversion in humans was in the same range as obtained using the gold standard immunoassays ELISA and Luminex, though requiring only a fraction of consumables, instrumentation, time to deliver results, and volume of sample. Furthermore, the results obtained with the method described can be visually interpreted without compromising accuracy as demonstrated by validation at a point-of-care unit. The magnetic bead immunoassay throughput can be customized on demand and is readily adapted to be used with any other 6xHis tagged protein or peptide as antigen to track other diseases.

In silico comparative analysis of Aeromonas Type VI Secretion System.

Aeromonas are bacteria broadly spread in the environment, particularly in aquatic habitats and can induce human infections. Several virulence factors have been described associated with bacterial pathogenicity, such as the Type VI Secretion System (T6SS). This system translocates effector proteins into target cells through a bacteriophage-like contractile structure encoded by tss genes. Here, a total of 446 Aeromonas genome sequences were screened for T6SS and the proteins subjected to in silico analysis. The T6SS-encoding locus was detected in 243 genomes and its genes are encoded in a cluster containing 13 core and 5 accessory genes, in highly conserved synteny. The amino acid residues identity of T6SS proteins ranges from 78 to 98.8%. In most strains, a pair of tssD and tssI is located upstream the cluster (tssD-2, tssI-2) and another pair was detected distant from the cluster (tssD-1, tssI-1). Significant variability was seen in TssI (VgrG) C-terminal region, which was sorted in four groups based on its sequence length and protein domains. TssI containing ADP-ribosyltransferase domain are associated exclusively with TssI-1, while genes coding proteins carrying DUF4123 (a conserved domain of unknown function) were observed downstream tssI-1 or tssI-2 and escort of possible effector proteins. Genes coding proteins containing DUF1910 and DUF1911 domains were located only downstream tssI-2 and might represent a pair of toxin/immunity proteins. Nearly all strains display downstream tssI-3, that codes for a lysozyme family domain protein. These data reveal that Aeromonas T6SS cluster synteny is conserved and the low identity observed for some genes might be due to species heterogeneity or its niche/functionality.

Influence of seasonality on the aerosol microbiome of the Amazon rainforest.

The Amazon rainforest is the world's largest tropical forest, and this biome may be a significant contributor to primary biological aerosol (PBA) emissions on a global scale. These aerosols also play a pivotal role in modulating ecosystem dynamics, dispersing biological material over geographic barriers and influencing climate through radiation absorption, light scattering, or acting as cloud condensation nuclei. Despite their importance, there are limited studies investigating the effect of environmental variables on the bioaerosol composition in the Amazon rainforest. Here we present a 16S rRNA gene-based amplicon sequencing approach to investigate the bacterial microbiome in aerosols of the Amazon rainforest during distinct seasons and at different heights above the ground. Our data revealed that seasonal changes in temperature, relative humidity, and precipitation are the primary drivers of compositional changes in the Amazon rainforest aerosol microbiome. Interestingly, no significant differences were observed in the bacterial community composition of aerosols collected at ground and canopy levels. The core airborne bacterial families present in Amazon aerosol were Enterobacteriaceae, Beijerinckiaceae, Polyangiaceae, Bacillaceae and Ktedonobacteraceae. By correlating the bacterial taxa identified in the aerosol with literature data, we speculate that the phyllosphere may be one possible source of airborne bacteria in the Amazon rainforest. Results of this study indicate that the aerosol microbiota of the Amazon Rainforest are fairly diverse and principally impacted by seasonal changes in temperature and humidity.

The Protein-Protein Interaction Network Reveals a Novel Role of the Signal Transduction Protein PII in the Control of c-di-GMP Homeostasis in Azospirillum brasilense.

The PII family comprises a group of widely distributed signal transduction proteins ubiquitous in prokaryotes and in the chloroplasts of plants. PII proteins sense the levels of key metabolites ATP, ADP, and 2-oxoglutarate, which affect the PII protein structure and thereby the ability of PII to interact with a range of target proteins. Here, we performed multiple ligand fishing assays with the PII protein orthologue GlnZ from the plant growth-promoting nitrogen-fixing bacterium Azospirillum brasilense to identify 37 proteins that are likely to be part of the PII protein-protein interaction network. Among the PII targets identified were enzymes related to nitrogen and fatty acid metabolism, signaling, coenzyme synthesis, RNA catabolism, and transcription. Direct binary PII-target complex was confirmed for 15 protein complexes using pulldown assays with recombinant proteins. Untargeted metabolome analysis showed that PII is required for proper homeostasis of important metabolites. Two enzymes involved in c-di-GMP metabolism were among the identified PII targets. A PII-deficient strain showed reduced c-di-GMP levels and altered aerotaxis and flocculation behavior. These data support that PII acts as a major metabolic hub controlling important enzymes and the homeostasis of key metabolites such as c-di-GMP in response to the prevailing nutritional status.IMPORTANCE The PII proteins sense and integrate important metabolic signals which reflect the cellular nutrition and energy status. Such extraordinary ability was capitalized by nature in such a way that the various PII proteins regulate different facets of metabolism by controlling the activity of a range of target proteins by protein-protein interactions. Here, we determined the PII protein interaction network in the plant growth-promoting nitrogen-fixing bacterium Azospirillum brasilense The interactome data along with metabolome analysis suggest that PII functions as a master metabolic regulator hub. We provide evidence that PII proteins act to regulate c-di-GMP levels in vivo and cell motility and adherence behaviors.

The NADP-dependent malic enzyme MaeB is a central metabolic hub controlled by the acetyl-CoA to CoASH ratio.

Malic enzymes participate in key metabolic processes, the MaeB-like malic enzymes carry a catalytic inactive phosphotransacetylase domain whose function remains elusive. Here we show that acetyl-CoA directly binds and inhibits MaeB-like enzymes with a saturable profile under physiological relevant acetyl-CoA concentrations. A MaeB-like enzyme from the nitrogen-fixing bacterium Azospirillum brasilense, namely AbMaeB1, binds both acetyl-CoA and unesterified CoASH in a way that inhibition of AbMaeB1 by acetyl-CoA is relieved by increasing CoASH concentrations. Hence, AbMaeB1 senses the acetyl-CoA/CoASH ratio. We revisited E. coli MaeB regulation to determine the inhibitory constant for acetyl-CoA. Our data support that the phosphotransacetylase domain of MaeB-like enzymes senses acetyl-CoA to dictate the fate of carbon distribution at the phosphoenol-pyruvate / pyruvate / oxaloacetate metabolic node.

SWeeP: representing large biological sequences datasets in compact vectors.

Vectoral and alignment-free approaches to biological sequence representation have been explored in bioinformatics to efficiently handle big data. Even so, most current methods involve sequence comparisons via alignment-based heuristics and fail when applied to the analysis of large data sets. Here, we present "Spaced Words Projection (SWeeP)", a method for representing biological sequences using relatively small vectors while preserving intersequence comparability. SWeeP uses spaced-words by scanning the sequences and generating indices to create a higher-dimensional vector that is later projected onto a smaller randomly oriented orthonormal base. We constructed phylogenetic trees for all organisms with mitochondrial and bacterial protein data in the NCBI database. SWeeP quickly built complete and accurate trees for these organisms with low computational cost. We compared SWeeP to other alignment-free methods and Sweep was 10 to 100 times quicker than the other techniques. A tool to build SWeeP vectors is available at https://sourceforge.net/projects/spacedwordsprojection/.

Proteomic and Metabolomic Analysis of Azospirillum brasilensentrC Mutant under High and Low Nitrogen Conditions.

Azospirillum brasilense is a diazotrophic microorganism capable of associating with roots of important grasses and cereals, promoting plant growth and increasing crop yields. Nitrogen levels and the Ntr regulatory system control the nitrogen metabolism in A. brasilense. This system comprises the nitrogen regulatory proteins GlnD, which is capable of adding uridylyl groups to the PII proteins, GlnB (PII-1) and GlnZ (PII-2), under limiting nitrogen levels. Under such conditions, the histidine kinase NtrB (nitrogen regulatory protein B) cannot interact with GlnB and phosphorylate NtrC (nitrogen regulatory protein C). The phosphorylated form of NtrC acts as a transcriptional activator of genes involved in the metabolism of alternative nitrogen sources. Considering the key role of NtrC in nitrogen metabolism in A. brasilense, in this work we evaluated the proteomic and metabolomic profiles of the wild-type FP2 strain and its mutant ntrC grown under high and low nitrogen. Analysis of the integrated data identifies novel NtrC targets, including proteins involved in the response against oxidative stress (i.e., glutathione S-transferase and hydroperoxide resistance protein), underlining the importance of NtrC to bacterial survival under oxidative stress conditions.

Virulence characteristics and antimicrobial resistance of Aeromonas veronii biovar sobria 312M, a clinical isolate.

Aeromonas are bacteria widely distributed in the environment, and some species are able to cause infections in humans, of which diarrhea is the most common. The objective of this study was to evaluate the presence of virulence and antimicrobial resistance associated characteristics in A. veronii biovar sobria strain 312M isolated from diarrheal stools. For this, the genome sequencing and phenotypical tests were performed. The draft genome annotation revealed several complete pathways associated with carbon metabolism and a mucin-desulfating sulfatase which may contribute to intestine colonization, and a large number of virulence-associated genes encoding structures associated with adhesion, toxins, and secretion systems. The strain exhibited swimming and swarming motility, biofilm formation, and hemolytic activity. It was resistant to ampicillin, ampicillin/sulbactam, and amoxicillin-clavulanic acid. Although a cphA gene encoding a narrow-spectrum carbapenase was identified in the strain genome, no carbapenemase activity was detected in the antimicrobial susceptibility test. When compared with other A. veronii with complete genomes, the main differences in virulence characteristics are related to lateral flagella and type III and VI secretion systems; the antimicrobial resistance spectrum also varied among strains. The results indicated that A. veronii biovar sobria 312M presents high virulence potential and resistance to limited classes of antimicrobials.

Repetitive genomic elements in Campomanesia xanthocarpa: prospection, characterization and cross amplification of molecular markers.

Repetitive genomic elements were prospected in Campomanesia xanthocarpa, aiming to characterize these elements in a non-model plant species and to develop species-specific microsatellite markers. Approximately 4.12% of the partial genome of C. xanthocarpa is composed of repetitive elements, being retrotransposons the most widely represented. A total of nine polymorphic microsatellite markers were obtained: four nuclear-neutral, two nuclear EST, two plastidial and one mitochondrial. Levels of population genetic diversity of four natural populations of C. xanthocarpa were characterized using these markers. In addition, the cross-species amplification of the microsatellite markers was tested in seven species of tribe Myrteae (Myrtaceae). The characterized microsatellite markers revealed low to moderate levels of genetic diversity (expected heterozygosity range: 0.33-0.57; observed heterozygosity: 0.26-0.74 and number of alleles: 2.25-4.25). Cross-species amplification was successful for all loci, except Cxant76. These nine markers will contribute for studies on genetic diversity, gene flow, plant selection and breeding of this species, towards the conservation of natural populations, as well as its commercial use.

Genome comparison between clinical and environmental strains of Herbaspirillum seropedicae reveals a potential new emerging bacterium adapted to human hosts.

BACKGROUND: Herbaspirillum seropedicae is an environmental ß-proteobacterium that is capable of promoting the growth of economically relevant plants through biological nitrogen fixation and phytohormone production. However, strains of H. seropedicae have been isolated from immunocompromised patients and associated with human infections and deaths. In this work, we sequenced the genomes of two clinical strains of H. seropedicae, AU14040 and AU13965, and compared them with the genomes of strains described as having an environmental origin. RESULTS: Both genomes were closed, indicating a single circular chromosome; however, strain AU13965 also carried a plasmid of 42,977 bp, the first described in the genus Herbaspirillum. Genome comparison revealed that the clinical strains lost the gene sets related to biological nitrogen fixation (nif) and the type 3 secretion system (T3SS), which has been described to be essential for interactions with plants. Comparison of the pan-genomes of clinical and environmental strains revealed different sets of accessorial genes. However, antimicrobial resistance genes were found in the same proportion in all analyzed genomes. The clinical strains also acquired new genes and genomic islands that may be related to host interactions. Among the acquired islands was a cluster of genes related to lipopolysaccharide (LPS) biosynthesis. Although highly conserved in environmental strains, the LPS biosynthesis genes in the two clinical strains presented unique and non-orthologous genes within the genus Herbaspirillum. Furthermore, the AU14040 strain cluster contained the neuABC genes, which are responsible for sialic acid (Neu5Ac) biosynthesis, indicating that this bacterium could add it to its lipopolysaccharide. The Neu5Ac-linked LPS could increase the bacterial resilience in the host aiding in the evasion of the immune system. CONCLUSIONS: Our findings suggest that the lifestyle transition from environment to opportunist led to the loss and acquisition of specific genes allowing adaptations to colonize and survive in new hosts. It is possible that these substitutions may be the starting point for interactions with new hosts.

Uncovering prokaryotic biodiversity within aerosols of the pristine Amazon forest.

Biological aerosols (bioaerosol) are atmospheric particles that act as a dispersion unit of living organisms across the globe thereby affecting the biogeographic distribution of organisms. Despite their importance, there is virtually no knowledge about bioaerosols emitted by pristine forests. Here we provide the very first survey of the prokaryotic community of a bioaerosol collected inside pristine Amazon forest at 2 m above ground. Total atmospheric particles were collected at the Amazon Tall Tower Observatory, subjected to metagenomic DNA extraction and the prokaryotic diversity was determined by 16S rRNA gene amplicon sequencing. A total of 271,577 reads of 250 bp of the 16S rRNA gene amplicon were obtained. Only 27% of the reads could be classified using the 16S SILVA database. Most belonged to Proteobacteria, Actinobacteria and Firmicutes which is in good agreement with other bioaerosol studies. Further inspection of the reads using Blast searches and the 18S SILVA database revealed that most of the dataset was composed of Fungi sequences. The identified microbes suggest that the atmosphere may act as an important gateway to interchange bacteria between plants, soil and water ecosystems.

Genome Analysis of Entomopathogenic Bacillus sp. ABP14 Isolated from a Lignocellulosic Compost.

We report the complete genome sequence of Bacillus sp. strain ABP14 isolated from lignocellulosic compost and selected by its ability in hydrolyzing carboxymethyl cellulose. This strain does not produce a Cry-like protein but showed an insecticidal activity against larvae of Anticarsia gemmatalis (Lepidoptera). Genome-based taxonomic analysis revealed that the ABP14 chromosome is genetically close to Bacillus thuringiensis serovar finitimus YBT020. ABP14 also carries one plasmid which showed no similarity with those from YBT020. Genome analysis of ABP14 identified unique genes related to cell surface structures, cell wall, metabolic competence, and virulence factors that may contribute for its survival and environmental adaptation, as well as its entomopathogenic activity.