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Pseudomonas aeruginosa belong to the special pathogen group capable of causing serious infections, with high mortality rates. The aim of this study was to describe the antibiotic resistance and genomic characteristics of Pseudomonas aeruginosa belonging to international high-risk clone ST235 (GPAE0131 isolate), obtained from hospital wastewater. P. aeruginosa GPAE0131 was isolated from ward tertiary hospital in Brazil and the antibiotic resistance profile was determined by the disc-diffusion method. Genomic characteristics related to antibiotic resistance and virulence factors were evaluated by genomic DNA sequencing on the Illumina MiSeq platform and bioinformatic analysis. GPAE0131 isolate showed resistance to piperacillin-tazobactam, cefepime, ceftazidime, imipenem, meropenem, ciprofloxacin, levofloxacin and tobramycin. Resistome comprehend of resistance genes to ß-lactams (blaVIM-2, blaOXA-4, blaOXA-488, blaPDC-35), aminoglycosides (aph(3')-IIb, aac(6')-IIc, aac(6')-Ib9, aadA1), fosfomycin (fosA), chloramphenicol (catB7) and sulfonamides (sul1). Genome comparisons evidence insertion of blaVIM-2 and blaOXA-4 genes. GPAE0131 isolate was predicted to be pathogenic to humans and several virulence factors were found, including encoding gene for ExoU and exotoxin A. All of these features into a pathogenic international high-risk clone (ST235), classified as critical priority, stands out as public health concern due to the widespread dispersal of human pathogens through wastewater. It is suggested that mitigating measures be implemented, such as the treatment of hospital sewage and the addition of tertiary treatment, to prevent the escape of pathogens at this level into the environment.
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Pseudomonas aeruginosa , Aguas Residuales , Aguas Residuales/microbiología , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/patogenicidad , Humanos , Brasil , Antibacterianos/farmacología , Hospitales , beta-Lactamasas/genética , Factores de Virulencia/genética , GenómicaRESUMEN
Peanut-based products have been associated with Salmonella foodborne outbreaks and/or recalls worldwide. The ability of Salmonella to persist for a long time in a low moisture environment can contribute to this kind of contamination. The objective of this study was to analyse the genome of five S. enterica enterica strains isolated from the peanut supply chain in Brazil, as well as to identify genetic determinants for survival under desiccation and validate these findings by phenotypic test of desiccation stress. The strains were in silico serotyped using the platform SeqSero2 as Miami (M2851), Javiana (M2973), Oranienburg (M2976), Muenster (M624), and Glostrup/Chomedey (M7864); with phylogenomic analysis support. Based on Multilocus Sequence Typing (MLST) the strains were assigned to STs 140, 1674, 321, 174, and 2519. In addition, eight pathogenicity islands were found in all the genomes using the SPIFinder 2.0 (SPI-1, SPI-2, SPI-3, SPI-5, SPI-9, SPI-13, SPI-14). The absence of a SPI-4 may indicate a loss of this island in the surveyed genomes. For the pangenomic analysis, 49 S. enterica genomes were input into the Roary pipeline. The majority of the stress related genes were considered as soft-core genes and were located on the chromosome. A desiccation stress phenotypic test was performed in trypticase soy broth (TSB) with four different water activity (aw) values. M2976 and M7864, both isolated from the peanut samples with the lowest aw, showed the highest OD570nm in TSB aw 0.964 and were statistically different (p < 0.05) from the strain isolated from the peanut sample with the highest aw (0.997). In conclusion, genome analyses have revealed signatures of desiccation adaptation in Salmonella strains, but phenotypic analyses suggested the environment influences the adaptive ability of Salmonella to overcome desiccation stress.
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Arachis , Genoma Bacteriano , Tipificación de Secuencias Multilocus , Filogenia , Salmonella enterica , Arachis/microbiología , Brasil , Salmonella enterica/genética , Salmonella enterica/aislamiento & purificación , Salmonella enterica/clasificación , Microbiología de Alimentos , Islas Genómicas , Desecación , GenómicaRESUMEN
Acinetobacter bereziniae has recently gained medical notoriety due to its emergence as a multidrug resistance and healthcare-associated pathogen. In this study, we report the whole-genome characterization of an A. bereziniae strain (A321) recovered from an infected semiaquatic turtle, as well as a comparative analysis of A. bereziniae strains circulating at the human-animal-environment interface. Strain A321 displayed a multidrug resistance profile to medically important antimicrobials, which was supported by a wide resistome. The novel Tn5393m transposon and a qnrB19-bearing ColE1-like plasmid were identified in A321 strain. Novel OXA-229-like ß-lactamases were detected and expression of OXA-931 demonstrated a 2-64-fold increase in the minimum inhibitory concentration for ß-lactam agents. Comparative genomic analysis revealed that most A. bereziniae strains did not carry any antimicrobial resistance genes (ARGs); however, some strains from China, Brazil, and India harbored six or more ARGs. Furthermore, A. bereziniae strains harbored conserved virulence genes. These results add valuable information regarding the spread of ARGs and mobile genetic elements that could be shared not only between A. bereziniae but also by other bacteria of clinical interest. This study also demonstrates that A. bereziniae can spill over from anthropogenic sources into natural environments and subsequently be transmitted to non-human hosts, making this a potential One Health bacteria that require close surveillance.
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Acinetobacter , Salud Única , Animales , Genómica , Acinetobacter/genética , BrasilRESUMEN
Tumor resistance remains an obstacle to successfully treating oral squamous cell carcinoma (OSCC). Cisplatin is widely used as a cytotoxic drug to treat solid tumors, including advanced OSCC, but with low efficacy due to chemoresistance. Therefore, identifying the pathways that contribute to chemoresistance may show new possibilities for improving the treatment. This work explored the role of the tumor necrosis factor-alpha (TNF-alpha)/NFkB signaling in driving the cisplatin resistance of OSCC and its potential as a pharmacological target to overcome chemoresistance. Differential accessibility analysis demonstrated the enrichment of opened chromatin regions in members of the TNF-alpha/NFkB signaling pathway, and RNA-Seq confirmed the upregulation of TNF-alpha/NFkB signaling in cisplatin-resistant cell lines. NFkB was accumulated in cisplatin-resistant cell lines and in cancer stem cells (CSC), and the administration of TNF-alpha increased the CSC, suggesting that TNF-alpha/NFkB signaling is involved in the accumulation of CSC. TNF-alpha stimulation also increased the histone deacetylases HDAC1 and SIRT1. Cisplatin-resistant cell lines were sensitive to the pharmacological inhibition of NFkB, and low doses of the NFkB inhibitors, CBL0137, and emetine, efficiently reduced the CSC and the levels of SIRT1, increasing histone acetylation. The NFkB inhibitors decreased stemness potential, clonogenicity, migration, and invasion of cisplatin-resistant cell lines. The administration of the emetine significantly reduced the tumor growth of cisplatin-resistant xenograft models, decreasing NFkB and SIRT1, increasing histone acetylation, and decreasing CSC. TNF-alpha/NFkB/SIRT1 signaling regulates the epigenetic machinery by modulating histone acetylation, CSC, and aggressiveness of cisplatin-resistant OSCC and the NFkB inhibition is a potential strategy to treat chemoresistant OSCC.
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Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Neoplasias de la Boca , Humanos , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Línea Celular Tumoral , Cisplatino/farmacología , Cisplatino/uso terapéutico , Resistencia a Antineoplásicos , Emetina/metabolismo , Emetina/uso terapéutico , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Histonas/metabolismo , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias de la Boca/genética , Neoplasias de la Boca/metabolismo , Células Madre Neoplásicas/patología , Sirtuina 1/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Regulación hacia ArribaRESUMEN
Sessile microorganisms are usually recalcitrant to antimicrobial treatments, and it is possible that finding biofilm-related effectors in metatranscriptomics datasets helps to understand mechanisms for bacterial persistence in diverse environments, by revealing protein-encoding genes that are expressed in situ. For this research, selected dairy-associated metatranscriptomics bioprojects were downloaded from the public databases JGI GOLD and NCBI (eight milk and 45 cheese samples), to screen for sequences encoding biofilm-related effectors. Based on the literature, the selected genetic determinants were related to adhesins, BAP, flagellum-related, intraspecific QS (AHL, HK, and RR), interspecific QS (LuxS), and QQ (AHL-acylases, AHL-lactonases). To search for the mRNA sequences encoding for those effector proteins, a custom database was built from UniprotKB, yielding 1,154,446 de-replicated sequences that were indexed in DIAMOND for alignment. The results revealed that in all the dairy-associated metatranscriptomic datasets obtained, there were reads assigned to genes involved with flagella, adhesion, and QS/QQ, but BAP-reads were found only for milk. Significant Pearson correlations (p < 0.05) were observed for transcripts encoding for flagella, RR, histidine kinases, adhesins, and LuxS, although no other significant correlations were found. In conclusion, the rationale used in this study was useful to demonstrate the presence of biofilm-associated effectors in metatranscriptomics datasets, pointing out to possible regulatory mechanisms in action in dairy-related biofilms, which could be targeted in the future to improve food safety.
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Lactobacillaceae presents potential for interspecific Quorum Sensing (QS) in spontaneous cocoa fermentation, correlated with high abundance of luxS. Three Brazilian isolates from cocoa fermentation were characterized by Whole Genome Sequencing and luxS gene was surveyed in their genomes, in comparison with public databases. They were classified as Lactiplantibacillus plantarum, Limosilactobacillus fermentum and Pediococcus acidilactici. LuxS genes were conserved in core genomes of the novel isolates, but in some non-cocoa related Lactic Acid Bacteria (LAB) it was accessory and plasmid-borne. The conservation and horizontal acquisition of luxS reinforces that QS is determinant for bacterial adaptation in several environments, especially taking into account the luxS has been correlated with modulation of bacteriocin production, stress tolerance and biofilm formation. Therefore, in this paper, new clade and species-specific primers were designed for future application for screening of luxS gene in LAB to evaluate the adaptive potential to diverse food fermentations.
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Cacao , Lactobacillales , Limosilactobacillus fermentum , Bacterias/genética , Cacao/genética , Cacao/microbiología , Fermentación , Lactobacillales/genéticaRESUMEN
Spheroids of intestinal cells (Caco-2) were used to evaluate the adhesion/invasion ability of Listeria monocytogenes (pathogen) and Lactobacillus sakei 1 (potential probiotic). Besides, transcriptomic analyses of Caco-2 cells in three dimensional cultures were done, with the aim of revealing possible host-foodborne bacteria interactions. Result of adhesion assay for L. monocytogenes in Caco-2 spheroids was 22.86 ± 0.33%, but it was stimulated in acidic pH (4.5) and by the presence of 2% sucrose (respectively, 32.56 ± 1.35% and 33.25 ± 1.26%). Conversely, the invasion rate of L. monocytogenes was lower at pH 4.5, in comparison with non-stressed controls (18.89 ± 1.05% and 58.65 ± 0.30%, respectively). L. sakei 1 adhered to Caco-2 tridimensional cell culture (27.30 ± 2.64%), with no invasiveness. There were 19 and 21 genes down and upregulated, respectively, in tridimensional Caco-2 cells, upon infection with L. monocytogenes, which involved immunity, apoptosis; cytoprotective responses, cell signalling-regulatory pathways. It was evidenced despite activation or deactivation of several pathways in intestinal cells to counteract infection, the pathogen was able to hijack many host defense mechanisms. On the other hand, the probiotic candidate L. sakei 1 was correlated with decreased transcription of two genes in Caco-2 cells, though it stimulated the expression of 14 others, with diverse roles in immunity, apoptosis, cytoprotective response and cell signalling-regulatory pathways. Our data suggest the use of tridimensional cell culture to mimic the intestinal epithelium is a good model for gathering broad information on the putative mechanisms of interaction between host and bacteria of importance for food safety, which can serve as a basis for further in-depth investigation.
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Técnicas de Cultivo de Célula/métodos , Intestinos/citología , Latilactobacillus sakei/fisiología , Listeria monocytogenes/fisiología , Adhesión Bacteriana , Reactores Biológicos/microbiología , Células CACO-2 , Proliferación Celular , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Concentración de Iones de Hidrógeno , Intestinos/química , Intestinos/microbiología , Probióticos/farmacología , Esferoides Celulares/química , Esferoides Celulares/citologíaRESUMEN
Microbial communities infiltrate the respiratory tract of cystic fibrosis patients, where chronic colonization and infection lead to clinical decline. This report aims to provide an overview of the diversity of bacterial and fungal species from the airway secretion of three young CF patients with severe pulmonary disease. The bacterial and fungal microbiomes were investigated by culture isolation, metataxonomics, and metagenomics shotgun. Virulence factors and antibiotic resistance genes were also explored. A. fumigatus was isolated from cultures and identified in high incidence from patient sputum samples. Candida albicans, Penicillium sp., Hanseniaspora sp., Torulaspora delbrueckii, and Talaromyces amestolkiae were isolated sporadically. Metataxonomics and metagenomics detected fungal reads (Saccharomyces cerevisiae, A. fumigatus, and Schizophyllum sp.) in one sputum sample. The main pathogenic bacteria identified were Staphylococcus aureus, Pseudomonas aeruginosa, Burkholderia cepacia complex, and Achromobacter xylosoxidans. The canonical core CF microbiome is composed of species from the genera Streptococcus, Neisseria, Rothia, Prevotella, and Haemophilus. Thus, the airways of the three young CF patients presented dominant bacterial genera and interindividual variability in microbial community composition and diversity. Additionally, a wide diversity of virulence factors and antibiotic resistance genes were identified in the CF lung microbiomes, which may be linked to the clinical condition of the CF patients. Understanding the microbial community is crucial to improve therapy because it may have the opposite effect, restructuring the pathogenic microbiota. Future studies focusing on the influence of fungi on bacterial diversity and microbial interactions in CF microbiomes will be welcome to fulfill this huge gap of fungal influence on CF physiopathology.
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Fibrosis Quística , Microbiota , Brasil , Fibrosis Quística/complicaciones , Humanos , Pulmón , Esputo , TalaromycesRESUMEN
Dairy foods are complex ecosystems composed of microorganisms from different origins that can affect flavor and safety of final products. The objective of this paper is to assess the in-house microbiota of two Brazilian dairies and to discuss the possible implications of the taxa determined for food protection. In total, 27 samples from dairies were cultured in selective (Baird Parker, de Man, Rogosa and Sharpe) and non-selective (Brain Heart Infusion) media, and the isolates were identified by Sanger sequencing. Moreover, metagenomic DNA was directly extracted from samples and the structure of the bacterial community was determined by massive DNA sequencing followed by bioinformatics analyses. The results showed the majority of isolates belonged to the group of lactic acid bacteria, but Enterobacteriaceae, Staphylococcacceae, Bacillaceae, Pseudomonadaceae and Moraxellaceae were also detected. From the reads obtained in metataxonomics analyses, a heatmap was constructed and the top 20 OTUs (operational taxonomic units) were determined. Besides, 12 most prevalent bacterial taxa were assigned to the core microbiota of the dairies evaluated, which included Thiomonas thermosulfata, Alkalibacillus salilacus, Pseudomonas clemancea, Erythrobacter aquimans, Tetragenococcus doogicus, Macrococcus brunensis, Pseudomonas ludensis, Streptococcus dentinousetti, Serratia entomophila, Vagococcus teuberi, Lactococcus fujiensis and Tolumonas auensis. In conclusion, the results reveal the presence of bacteria that may be related to spoilage and also foodborne diseases, in microbial niches that also present rare taxa, highlighting the importance to consider culture-independent results to evaluate and improve food safety.
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Acinetobacter calcoaceticus-Acinetobacter baumannii complex is considered one of the main causes of hospital-acquired infections. Acinetobacter seifertii was recently characterized within this complex and it has been described as an emergent pathogen associated with bacteremia. The emergence of multidrug-resistant (MDR) bacteria, including Acinetobacter sp., is considered a global public health threat and an environmental problem because MDR bacteria have been spreading from several sources. Therefore, this study aimed to characterize an environmental MDR A. seifertii isolate (SAb133) using whole genome sequencing and a comparative genomic analysis was performed with A. seifertii strains recovered from various sources. The SAb133 isolate was obtained from soil of a corn crop field and presented high MICs for antimicrobials and metals. The comparative genomic analyses revealed ANI values higher than 95% of relatedness with other A. seifertii strains than A. calcoaceticus-A. baumannii complex. Resistome and virulome analyses were also performed and showed different antimicrobial resistance determinants and metal tolerance genes as well as virulence genes related to A. baumannii known virulence genes. In addition, genomic islands, IS elements, plasmids and prophage-related sequences were detected. Comparative genomic analysis showed that MDR A. seifertii SAb133 had a high amount of determinants related to antimicrobial resistance and tolerance to metals, besides the presence of virulence genes. To the best of our knowledge, this is the first report of a whole genome sequence of a MDR A. seifertii isolated from soil. Therefore, this study contributed to a better understanding of the genetic relationship among the few known A. seifertii strains worldwide distributed.