In vitro susceptibility to fosfomycin in clinical and environmental extended-spectrum beta-lactamase producing and/or ciprofloxacin-non-susceptible Escherichia coli isolates.

Extended-spectrum beta-lactamase producing and ciprofloxacin-non-susceptible Escherichia coli are clinical and environmental issues. We evaluated the susceptibility profile of fosfomycin in non-susceptible E. coli isolated from urine and the environment. We measured the activity of fosfomycin against 319 and 36 E. coli strains from urine and environmental isolates, respectively, collected from rivers. Fosfomycin resistance profiles were investigated using the minimal inhibitory concentration (MIC), according to the Clinical and Laboratory Standards Institute (CLSI) and the European Committee for Antimicrobial Susceptibility Testing (EUCAST) guidelines. Antibiotic susceptibility testing revealed that 5% and 6.6% of urine samples were non-susceptible to fosfomycin according to CLSI and EUCAST guidelines, respectively. The fosfomycin MIC50/90 was 0.5/4 mg/L. Of the 36 E. coli isolates from river water, 11.1% and 13,8% were non-susceptible to fosfomycin according to CLSI and EUCAST, respectively (range ≤0.25 ≥512 mg/L). All the isolates with MIC ≥512 mg/L for fosfomycin showed the fosA3 gene. Fosfomycin resistance was more frequent in the environment than in clinical samples.

In vitro susceptibility to fosfomycin in clinical and environmental extended-spectrum beta-lactamase producing and/or ciprofloxacin-non-susceptible Escherichia coli isolates

ABSTRACT Extended-spectrum beta-lactamase producing and ciprofloxacin-non-susceptible Escherichia coli are clinical and environmental issues. We evaluated the susceptibility profile of fosfomycin in non-susceptible E. coli isolated from urine and the environment. We measured the activity of fosfomycin against 319 and 36 E. coli strains from urine and environmental isolates, respectively, collected from rivers. Fosfomycin resistance profiles were investigated using the minimal inhibitory concentration (MIC), according to the Clinical and Laboratory Standards Institute (CLSI) and the European Committee for Antimicrobial Susceptibility Testing (EUCAST) guidelines. Antibiotic susceptibility testing revealed that 5% and 6.6% of urine samples were non-susceptible to fosfomycin according to CLSI and EUCAST guidelines, respectively. The fosfomycin MIC50/90 was 0.5/4 mg/L. Of the 36 E. coli isolates from river water, 11.1% and 13,8% were non-susceptible to fosfomycin according to CLSI and EUCAST, respectively (range ≤0.25 ≥512 mg/L). All the isolates with MIC ≥512 mg/L for fosfomycin showed the fosA3 gene. Fosfomycin resistance was more frequent in the environment than in clinical samples.

Rare genetic variants involved in multisystem inflammatory syndrome in children: a multicenter Brazilian cohort study.

Introduction: Despite the existing data on the Multisystem Inflammatory Syndrome in Children (MIS-C), the factors that determine these patients evolution remain elusive. Answers may lie, at least in part, in genetics. It is currently under investigation that MIS-C patients may have an underlying innate error of immunity (IEI), whether of monogenic, digenic, or even oligogenic origin. Methods: To further investigate this hypothesis, 30 patients with MIS-C were submitted to whole exome sequencing. Results: Analyses of genes associated with MIS-C, MIS-A, severe covid-19, and Kawasaki disease identified twenty-nine patients with rare potentially damaging variants (50 variants were identified in 38 different genes), including those previously described in IFNA21 and IFIH1 genes, new variants in genes previously described in MIS-C patients (KMT2D, CFB, and PRF1), and variants in genes newly associated to MIS-C such as APOL1, TNFRSF13B, and G6PD. In addition, gene ontology enrichment pointed to the involvement of thirteen major pathways, including complement system, hematopoiesis, immune system development, and type II interferon signaling, that were not yet reported in MIS-C. Discussion: These data strongly indicate that different gene families may favor MIS- C development. Larger cohort studies with healthy controls and other omics approaches, such as proteomics and RNAseq, will be precious to better understanding the disease dynamics.

TgKDAC4: A Unique Deacetylase of Toxoplasma's Apicoplast.

Toxoplasma gondii is an obligate intracellular parasite of the phylum Apicomplexa and causes toxoplasmosis infections, a disease that affects a quarter of the world's population and has no effective cure. Epigenetic regulation is one of the mechanisms controlling gene expression and plays an essential role in all organisms. Lysine deacetylases (KDACs) act as epigenetic regulators affecting gene silencing in many eukaryotes. Here, we focus on TgKDAC4, an enzyme unique to apicomplexan parasites, and a class IV KDAC, the least-studied class of deacetylases so far. This enzyme shares only a portion of the specific KDAC domain with other organisms. Phylogenetic analysis from the TgKDAC4 domain shows a putative prokaryotic origin. Surprisingly, TgKDAC4 is located in the apicoplast, making it the only KDAC found in this organelle to date. Transmission electron microscopy assays confirmed the presence of TgKDAC4 in the periphery of the apicoplast. We identified possible targets or/and partners of TgKDAC4 by immunoprecipitation assays followed by mass spectrometry analysis, including TgCPN60 and TgGAPDH2, both located at the apicoplast and containing acetylation sites. Understanding how the protein works could provide new insights into the metabolism of the apicoplast, an essential organelle for parasite survival.

Modulation of Klebsiella pneumoniae Outer Membrane Vesicle Protein Cargo under Antibiotic Treatment.

Klebsiella pneumoniae is a nosocomial pathogen and an important propagator of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains. Like other Gram-negative bacteria, they secrete outer membrane vesicles (OMVs) that distribute virulence and resistance factors. Here, we subjected a K. pneumoniae-XDR to subinhibitory concentrations of meropenem, amikacin, polymyxin B, and a combination of these agents to evaluate changes in the protein cargo of OMVs through liquid chromatography-tandem mass spectrometry (LC-MS/MS). Genome sequencing of the clinical isolate K. pneumoniae strain HCD1 (KpHCD1) revealed the presence of 41 resistance genes and 159 virulence factors. We identified 64 proteins in KpHCD1-OMVs modulated with different antibiotic treatments involved in processing genetic information, environmental information, cell envelope formation, energy metabolism, and drug resistance. The OMV proteome expression profile suggests that OMVs may be associated with pathogenicity, survival, stress response, and resistance dissemination.

TgKDAC4: a unique deacetylase of toxoplasma's apicoplast

Toxoplasma gondii is an obligate intracellular parasite of the phylum Apicomplexa and causes toxoplasmosis infections, a disease that affects a quarter of the world’s population and has no effective cure. Epigenetic regulation is one of the mechanisms controlling gene expression and plays an essential role in all organisms. Lysine deacetylases (KDACs) act as epigenetic regulators affecting gene silencing in many eukaryotes. Here, we focus on TgKDAC4, an enzyme unique to apicomplexan parasites, and a class IV KDAC, the least-studied class of deacetylases so far. This enzyme shares only a portion of the specific KDAC domain with other organisms. Phylogenetic analysis from the TgKDAC4 domain shows a putative prokaryotic origin. Surprisingly, TgKDAC4 is located in the apicoplast, making it the only KDAC found in this organelle to date. Transmission electron microscopy assays confirmed the presence of TgKDAC4 in the periphery of the apicoplast. We identified possible targets or/and partners of TgKDAC4 by immunoprecipitation assays followed by mass spectrometry analysis, including TgCPN60 and TgGAPDH2, both located at the apicoplast and containing acetylation sites. Understanding how the protein works could provide new insights into the metabolism of the apicoplast, an essential organelle for parasite survival.

Multiple introductions and country-wide spread of DENV-2 genotype II (Cosmopolitan) in Brazil.

Dengue virus serotype 2, genotype Cosmopolitan (DENV-2-GII), is one of the most widespread DENV strains globally. In the USA, DENV-2 epidemics have been dominated by DENV-2 genotype Asian-American (DENV-2-GIII), and the first cases of DENV-2-GII were only described in 2019, in Peru, and in 2021 in Brazil. To gain new information about the circulation of DENV-2-GII in Brazil, we sequenced 237 DENV-2 confirmed cases sampled between March 2021 and March 2023 and revealed that DENV-2-GII is already present in all geographic regions of Brazil. The phylogeographic analysis inferred that DENV-2-GII was introduced at least four times in Brazil, between May 2020 and August 2022, generating multiple clades that spread throughout the country with different success. Despite multiple introductions of DENV-2-GII, analysis of the country-wide laboratory surveillance data showed that the Brazilian dengue epidemic in 2022 was dominated by DENV-1 in most states. We hypothesize that massive circulation of DENV-2-GIII in previous years in Brazil might have created a population immune barrier against symptomatic homotypic reinfections by DENV-2-GII, leading to sustained cryptic circulation in asymptomatic cases and localized outbreaks of this new genotype. In summary, our study stresses the importance of arboviral genomic surveillance to close monitoring and better understanding the potential impact of DENV-2-GII in the coming years.

Role of Efflux Pumps on Antimicrobial Resistance in Pseudomonas aeruginosa.

Antimicrobial resistance is an old and silent pandemic. Resistant organisms emerge in parallel with new antibiotics, leading to a major global public health crisis over time. Antibiotic resistance may be due to different mechanisms and against different classes of drugs. These mechanisms are usually found in the same organism, giving rise to multidrug-resistant (MDR) and extensively drug-resistant (XDR) bacteria. One resistance mechanism that is closely associated with the emergence of MDR and XDR bacteria is the efflux of drugs since the same pump can transport different classes of drugs. In Gram-negative bacteria, efflux pumps are present in two configurations: a transmembrane protein anchored in the inner membrane and a complex formed by three proteins. The tripartite complex has a transmembrane protein present in the inner membrane, a periplasmic protein, and a porin associated with the outer membrane. In Pseudomonas aeruginosa, one of the main pathogens associated with respiratory tract infections, four main sets of efflux pumps have been associated with antibiotic resistance: MexAB-OprM, MexXY, MexCD-OprJ, and MexEF-OprN. In this review, the function, structure, and regulation of these efflux pumps in P. aeruginosa and their actions as resistance mechanisms are discussed. Finally, a brief discussion on the potential of efflux pumps in P. aeruginosa as a target for new drugs is presented.

Lower airway microbiota and decreasing lung function in young Brazilian cystic fibrosis patients with pulmonary Staphylococcus and Pseudomonas infection.

Cystic fibrosis (CF) is a genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator gene that leads to respiratory complications and mortality. Studies have shown shifts in the respiratory microbiota during disease progression in individuals with CF. In addition, CF patients experience short cycles of acute intermittent aggravations of symptoms called pulmonary exacerbations, which may be characterized by a decrease in lung function and weight loss. The resident microbiota become imbalanced, promoting biofilm formation, and reducing the effectiveness of therapy. The aim of this study was to monitor patients aged 8-23 years with CF to evaluate their lower respiratory microbiota using 16S rRNA sequencing. The most predominant pathogens observed in microbiota, Staphylococcus (Staph) and Pseudomonas (Pseud) were correlated with clinical variables, and the in vitro capacity of biofilm formation for these pathogens was tested. A group of 34 patients was followed up for 84 days, and 306 sputum samples were collected and sequenced. Clustering of microbiota by predominant pathogen showed that children with more Staph had reduced forced expiratory volume in one second (FEV1) and forced vital capacity (FVC) compared to children with Pseud. Furthermore, the patients' clinical condition was consistent with the results of pulmonary function. More patients with pulmonary exacerbation were observed in the Staph group than in the Pseud group, as confirmed by lower body mass index and pulmonary function. Additionally, prediction of bacterial functional profiles identified genes encoding key enzymes involved in virulence pathways in the Pseud group. Importantly, this study is the first Brazilian study to assess the lower respiratory microbiota in a significant group of young CF patients. In this sense, the data collected for this study on the microbiota of children in Brazil with CF provide a valuable contribution to the knowledge in the field.

Emergence and Spread of the SARS-CoV-2 Variant of Concern Delta across Different Brazilian Regions.

The SARS-CoV-2 variant of concern (VOC) Delta was first detected in India in October 2020. The first imported cases of the Delta variant in Brazil were identified in April 2021 in the southern region, followed by more cases in different regions during the following months. By early September 2021, Delta was already the dominant variant in the southeastern (87%), southern (73%), and northeastern (52%) Brazilian regions. This study aimed to understand the spatiotemporal dissemination dynamics of Delta in Brazil. To this end, we employed a combination of maximum likelihood (ML) and Bayesian methods to reconstruct the evolutionary relationship of 2,264 VOC Delta complete genomes (482 from this study) recovered across 21 of the 27 Brazilian federal units. Our phylogeographic analyses identified three major transmission clusters of Delta in Brazil. The clade BR-I (n = 1,560) arose in Rio de Janeiro in late April 2021 and was the major cluster behind the dissemination of the VOC Delta in the southeastern, northeastern, northern, and central-western regions. The AY.101 lineage (n = 207) that arose in the Paraná state in late April 2021 and aggregated the largest fraction of sampled genomes from the southern region. Lastly, the AY.46.3 lineage emerged in Brazil in the São Paulo state in early June 2021 and remained mostly restricted to this state. In the rapid turnover of viral variants characteristic of the SARS-CoV-2 pandemic, Brazilian regions seem to occupy different stages of an increasing prevalence of the VOC Delta in their epidemic profiles. This process demands continuous genomic and epidemiological surveillance toward identifying and mitigating new introductions, limiting their dissemination, and preventing the establishment of more significant outbreaks in a population already heavily affected by the COVID-19 pandemic. IMPORTANCE Amid the SARS-CoV-2 continuously changing epidemic profile, this study details the space-time dynamics of the emergence of the Delta lineage across Brazilian territories, pointing out its multiple introductions in the country and its most prevalent sublineages. Some of these sublineages have their emergence, alongside their genomic composition and geographic distribution, detailed here for the first time. A special focus is given to the emergence process of Delta outside the country's south and southeast regions, the most populated and subjects of most published SARS-CoV-2 studies in Brazil. In summary, the study allows a better comprehension of the evolution process of a SARS-CoV-2 lineage that would be associated with a significant recrudescence of the pandemic in Brazil.

Resensitization of Fosfomycin-Resistant Escherichia coli Using the CRISPR System.

Antimicrobial resistance is a public health burden with worldwide impacts and was recently identified as one of the major causes of death in 2019. Fosfomycin is an antibiotic commonly used to treat urinary tract infections, and resistance to it in Enterobacteriaceae is mainly due to the metalloenzyme FosA3 encoded by the fosA3 gene. In this work, we adapted a CRISPR-Cas9 system named pRE-FOSA3 to restore the sensitivity of a fosA3+ Escherichia coli strain. The fosA3+ E. coli strain was generated by transforming synthetic fosA3 into a nonpathogenic E. coli TOP10. To mediate the fosA3 disruption, two guide RNAs (gRNAs) were selected that used conserved regions within the fosA3 sequence of more than 700 fosA3+ E. coli isolates, and the resensitization plasmid pRE-FOSA3 was assembled by cloning the gRNA into pCas9. gRNA_195 exhibited 100% efficiency in resensitizing the bacteria to fosfomycin. Additionally, the edited strain lost the ampicillin resistance encoded in the same plasmid containing the synthetic fosA3 gene, despite not being the CRISPR-Cas9 target, indicating plasmid clearance. The in vitro analysis presented here points to a path that can be explored to assist the development of effective alternative methods of treatment against fosA3+ bacteria.

Phylogenetic-based inference reveals distinct transmission dynamics of SARS-CoV-2 lineages Gamma and P.2 in Brazil.

The COVID-19 epidemic in Brazil experienced two major lineage replacements until mid-2021. The first was driven by lineage P.2, in late 2020, and the second by lineage Gamma, in early 2021. To understand how these SARS-CoV-2 lineages spread in Brazil, we analyzed 11,724 genomes collected throughout the country between September 2020 and April 2021. Our findings indicate that lineage P.2 probably emerged in July 2020 in the Rio de Janeiro state and Gamma in November 2020 in the Amazonas state. Both states were the main hubs of viral disseminations to other Brazilian locations. We estimate that Gamma was 1.56-3.06 times more transmissible than P.2 in Rio de Janeiro and that the median effective reproductive number (Re) of Gamma varied according to the geographic context (Re = 1.59-3.55). In summary, our findings support that lineage Gamma was more transmissible and spread faster than P.2 in Brazil.

High Genomic Identity between Clinical and Environmental Strains of Herbaspirillum frisingense Suggests Pre-Adaptation to Different Hosts and Intrinsic Resistance to Multiple Drugs.

The genus Herbaspirillum is widely studied for its ability to associate with grasses and to perform biological nitrogen fixation. However, the bacteria of the Herbaspirillum genus have frequently been isolated from clinical samples. Understanding the genomic characteristics that allow these bacteria to switch environments and become able to colonize human hosts is essential for monitoring emerging pathogens and predicting outbreaks. In this work, we describe the sequencing, assembly, and annotation of the genome of H. frisingense AU14559 isolated from the sputum of patients with cystic fibrosis, and its comparison with the genomes of the uropathogenic strain VT-16-41 and the environmental strains GSF30, BH-1, IAC152, and SG826. The genes responsible for biological nitrogen fixation were absent from all strains except for GSF30. On the other hand, genes encoding virulence and host interaction factors were mostly shared with environmental strains. We also identified a large set of intrinsic antibiotic resistance genes that were shared across all strains. Unlike other strains, in addition to unique genomic islands, AU14559 has a mutation that renders the biosynthesis of rhamnose and its incorporation into the exopolysaccharide unfeasible. These data suggest that H. frisingense has characteristics that provide it with the metabolic diversity needed to infect and colonize human hosts.

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.

Origin and evolution of nonulosonic acid synthases and their relationship with bacterial pathogenicity revealed by a large-scale phylogenetic analysis.

Nonulosonic acids (NulOs) are a group of nine-carbon monosaccharides with different functions in nature. N-acetylneuraminic acid (Neu5Ac) is the most common NulO. It covers the membrane surface of all human cells and is a central molecule in the process of self-recognition via SIGLECS receptors. Some pathogenic bacteria escape the immune system by copying the sialylation of the host cell membrane. Neu5Ac production in these bacteria is catalysed by the enzyme NeuB. Some bacteria can also produce other NulOs named pseudaminic and legionaminic acids, through the NeuB homologues PseI and LegI, respectively. In Opisthokonta eukaryotes, the biosynthesis of Neu5Ac is catalysed by the enzyme NanS. In this study, we used publicly available data of sequences of NulOs synthases to investigate its distribution within the three domains of life and its relationship with pathogenic bacteria. We mined the KEGG database and found 425 NeuB sequences. Most NeuB sequences (58.74 %) from the KEGG orthology database were classified as from environmental bacteria; however, sequences from pathogenic bacteria showed higher conservation and prevalence of a specific domain named SAF. Using the HMM profile we identified 13 941 NulO synthase sequences in UniProt. Phylogenetic analysis of these sequences showed that the synthases were divided into three main groups that can be related to the lifestyle of these bacteria: (I) predominantly environmental, (II) intermediate and (III) predominantly pathogenic. NeuB was widely distributed in the groups. However, LegI and PseI were more concentrated in groups II and III, respectively. We also found that PseI appeared later in the evolutionary process, derived from NeuB. We use this same methodology to retrieve sialic acid synthase sequences from Archaea and Eukarya. A large-scale phylogenetic analysis showed that while the Archaea sequences are spread across the tree, the eukaryotic NanS sequences were grouped in a specific branch in group II. None of the bacterial NanS sequences grouped with the eukaryotic branch. The analysis of conserved residues showed that the synthases of Archaea and Eukarya present a mutation in one of the three catalytic residues, an E134D change, related to a Neisseria meningitidis reference sequence. We also found that the conservation profile is higher between NeuB of pathogenic bacteria and NanS of eukaryotes than between NeuB of environmental bacteria and NanS of eukaryotes. Our large-scale analysis brings new perspectives on the evolution of NulOs synthases, suggesting their presence in the last common universal ancestor.

Comparative Genomics Provides Insights into the Taxonomy of Azoarcus and Reveals Separate Origins of Nif Genes in the Proposed Azoarcus and Aromatoleum Genera.

Among other attributes, the Betaproteobacterial genus Azoarcus has biotechnological importance for plant growth-promotion and remediation of petroleum waste-polluted water and soils. It comprises at least two phylogenetically distinct groups. The "plant-associated" group includes strains that are isolated from the rhizosphere or root interior of the C4 plant Kallar Grass, but also strains from soil and/or water; all are considered to be obligate aerobes and all are diazotrophic. The other group (now partly incorporated into the new genus Aromatoleum) comprises a diverse range of species and strains that live in water or soil that is contaminated with petroleum and/or aromatic compounds; all are facultative or obligate anaerobes. Some are diazotrophs. A comparative genome analysis of 32 genomes from 30 Azoarcus-Aromatoleum strains was performed in order to delineate generic boundaries more precisely than the single gene, 16S rRNA, that has been commonly used in bacterial taxonomy. The origin of diazotrophy in Azoarcus-Aromatoleum was also investigated by comparing full-length sequences of nif genes, and by physiological measurements of nitrogenase activity using the acetylene reduction assay. Based on average nucleotide identity (ANI) and whole genome analyses, three major groups could be discerned: (i) Azoarcus comprising Az. communis, Az. indigens and Az. olearius, and two unnamed species complexes, (ii) Aromatoleum Group 1 comprising Ar. anaerobium, Ar. aromaticum, Ar. bremense, and Ar. buckelii, and (iii) Aromatoleum Group 2 comprising Ar. diolicum, Ar. evansii, Ar. petrolei, Ar. toluclasticum, Ar. tolulyticum, Ar. toluolicum, and Ar. toluvorans. Single strain lineages such as Azoarcus sp. KH32C, Az. pumilus, and Az. taiwanensis were also revealed. Full length sequences of nif-cluster genes revealed two groups of diazotrophs in Azoarcus-Aromatoleum with nif being derived from Dechloromonas in Azoarcus sensu stricto (and two Thauera strains) and from Azospira in Aromatoleum Group 2. Diazotrophy was confirmed in several strains, and for the first time in Az. communis LMG5514, Azoarcus sp. TTM-91 and Ar. toluolicum TT. In terms of ecology, with the exception of a few plant-associated strains in Azoarcus (s.s.), across the group, most strains/species are found in soil and water (often contaminated with petroleum or related aromatic compounds), sewage sludge, and seawater. The possession of nar, nap, nir, nor, and nos genes by most Azoarcus-Aromatoleum strains suggests that they have the potential to derive energy through anaerobic nitrate respiration, so this ability cannot be usefully used as a phenotypic marker to distinguish genera. However, the possession of bzd genes indicating the ability to degrade benzoate anaerobically plus the type of diazotrophy (aerobic vs. anaerobic) could, after confirmation of their functionality, be considered as distinguishing phenotypes in any new generic delineations. The taxonomy of the Azoarcus-Aromatoleum group should be revisited; retaining the generic name Azoarcus for its entirety, or creating additional genera are both possible outcomes.

Cross-Kingdom Extracellular Vesicles EV-RNA Communication as a Mechanism for Host-Pathogen Interaction.

The extracellular vesicle (EVs) traffic has been highlighted as a very important pathway of cellular communication. EVs are produced by prokaryotes and eukaryotes organisms and can carry molecules to help maintain homeostasis, responding to general disbalance, infections, and allowing rapid modulation of the immune system. In the context of infection, EVs from both the host and the pathogen have been identified as playing roles in the recruitment of immunological molecules that can lead to the resolution of the infection or the host's defeat. Bacterial vesicles RNA cargo play roles in the host cell by regulating gene expression and modulating immune response. In fungi the RNA molecules present in EVs are diverse and participate in communication between the host and pathogenic fungi. Little is known about how cross-kingdom sRNA trafficking occurs, although in recent years, there has been an increase in studies that relate EV participation in sRNA delivery. This review aims to elucidate and update the reader concerning the role of extracellular vesicles, with emphasis in the RNA content. We describe the EVs during infection from the host point-of-view, as well as the bacteria and fungi pathogens producing EVs that help the establishment of the disease.

Metagenomic analysis of the bacterial microbiota associated with cultured oysters (Crassostrea sp.) in estuarine environments.

In this work, we identified the bacterial microbiota associated with farmed oystersin estuarine regions of four states in the north eastern region of Brazil. During the drought and rainy seasons, for eight months, twenty oysters were sampled seasonally from seven different marine farms. In the laboratory, DNA extraction, amplification, and sequencing of the 16S rRNA gene were performed to establish the taxonomic units. We identified 106 genera of bacteria belonging to 103 families, 70 orders, 39 classes, and 21 phyla. Out of the total, 40 of the genera represented bacteria potentially pathogenic to humans; of these, nine are known to cause foodborne diseases and six are potentially pathogenic to oysters. The most prevalent genera were Mycoplasma, Propionigenium, Psychrilyobacter, and Arcobacter. The results indicate the need for more systematic monitoring of bacteria of the genus Mycoplasma in oyster farming operations in the Brazilian north eastern region. Currently, Mycoplasma is not one of the microorganisms analysed and monitored by order of Brazilian legislation during the oyster production and/or commercialization process, even though this genus was the most prevalent at all sampling points and presents pathogenic potential both for oysters and for consumers.

Molecular relationships of Campomanesia xanthocarpa within Myrtaceae based on the complete plastome sequence and on the plastid ycf2 gene.

Plastomes are very informative structures for comparative phylogenetic and evolutionary analyses. We sequenced and analyzed the complete plastome of Campomanesia xanthocarpa and compared its gene order, structure, and evolutionary characteristics within Myrtaceae. Analyzing 48 species of Myrtaceae, we identified six genes representing 'hotspots' of variability within the plastomes (ycf2, atpA, rpoC2, pcbE, ndhH and rps16), and performed phylogenetic analyses based on: (i) the ycf2 gene, (ii) all the six genes identified as 'hotspots' of variability, and (iii) the genes identified as 'hotspots' of variability, except the ycf2 gene. The structure, gene order, and gene content of the C. xanthocarpa plastome are similar to other Myrtaceae species. Phylogenetic analyses revealed the ycf2 gene as a promissing region for barcoding within this family, having also a robust phylogenetic signal. The synonymous and nonsynonymous substitution rates and the Ka/Ks ratio revealed low values for the ycf2 gene among C. xanthocarpa and the other 47 analyzed species of Myrtaceae, with moderate purifying selection acting on this gene. The average nucleotide identity (ANI) analysis of the whole plastomes produced phylogenetic trees supporting the monophyly of three Myrtaceae tribes. The findings of this study provide support for planning conservation, breeding, and biotechnological programs for this species.

Resistome analysis of bloodstream infection bacterial genomes reveals a specific set of proteins involved in antibiotic resistance and drug efflux.

Bacterial resistance to antibiotics is a global public health problem. Its association with bloodstream infections is even more severe and may easily evolve to sepsis. To improve our response to these bacteria, it is essential to gather thorough knowledge on the main pathogens along with the main mechanisms of resistance they carry. In this paper, we performed a large meta-analysis of 3872 bacterial genomes isolated from blood samples, from which we identified 71 745 antibiotic resistance genes (ARGs). Taxonomic analysis showed that Proteobacteria and Firmicutes phyla, and the species Klebsiella pneumoniae and Staphylococcus aureus were the most represented. Comparison of ARGs with the Resfams database showed that the main mechanism of antibiotic resistance is mediated by efflux pumps. Clustering analysis between resistome of blood and soil-isolated bacteria showed that there is low identity between transport and efflux proteins between bacteria from these environments. Furthermore, a correlation analysis among all features showed that K. pneumoniae and S. aureus formed two well-defined clusters related to the resistance mechanisms, proteins and antibiotics. A retrospective analysis has shown that the average number of ARGs per genome has gradually increased. The results demonstrate the importance of comprehensive studies to understand the antibiotic resistance phenomenon.