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Enterococci are members of the microbiota of humans and other animals. They can also be found in the environment, associated with food, healthcare infections, and hospital settings. Due to their wide distribution, they are inserted in the One Health context. The selective pressure caused by the extensive use of antimicrobial agents in humans, animals, and agriculture has increased the frequency of resistance to various drugs among enterococcal species. CRISPR-Cas system, an important prokaryotic defense mechanism against the entry of mobile genetic elements, may prevent the acquisition of genes involved in antimicrobial resistance and virulence. This system has been increasingly used as a gene editing tool, which can be used as a way to recognize and inactivate genes of interest. Here, we conduct a review on CRISPR systems found in enterococci, considering their occurrence, structure and organization, mechanisms of action and use as a genetic engineering technology. Type II-A CRISPR-Cas systems were shown to be the most frequent among enterococcal species, and the orphan CRISPR2 was the most commonly found system (54.1%) among enterococcal species, especially in Enterococcus faecalis. Distribution of CRISPR systems varied among species. CRISPR systems had 1 to 20 spacers, with size between 23 and 37 bp and direct repeat sequences from 25 to 37 bp. Several applications of the CRISPR-Cas biotechnology have been described in enterococci, mostly in vitro, using this editing tool to target resistance- and virulence-related genes., (© 2024. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.)

Sugar-induced metabolic imbalances are a major health problem since an excessive consumption of saccharides has been linked to greater obesity rates at a global level. Sucrose, a disaccharide composed of 50% glucose and 50% fructose, is commonly used in the food industry and found in a range of fast, restaurant, and processed foods. Herein, we investigated the effects of a TRPC4/TRPC5 blocker, ML204, in the metabolic imbalances triggered by early exposure to sucrose-enriched diet in mice. TRPC4 and TRPC5 belong to the family of non-selective Ca +2 channels known as transient receptor potential channels. High-sucrose (HS)-fed animals with hyperglycaemia and dyslipidaemia, were accompanied by increased body mass index. mesenteric adipose tissue accumulation with larger diameter cells and hepatic steatosis in comparison to those fed normal diet. HS mice also exhibited enhanced adipose, liver, and pancreas TNFα and VEGF levels. ML204 exacerbated hyperglycaemia, dyslipidaemia, fat tissue deposition, hepatic steatosis, and adipose tissue and liver TNFα in HS-fed mice. Normal mice treated with the blocker had greater hepatic steatosis and adipose tissue cell numbers/diameter than those receiving vehicle, but showed no significant changes in tissue inflammation, glucose, and lipid levels. The results indicate that TRPC4/TRPC5 protect against the metabolic imbalances caused by HS ingestion.

Introduction: The aac(6')-Ib-cr and bla KPC genes are spreading among Enterobacteriaceae species, including Providencia stuartii, in some countries of world., Methods: These genes were investigated in 28 P. stuartii isolates from a public hospital in Recife, Pernambuco, Brazil, by PCR and sequencing., Results: The aac(6')-Ib-cr gene was detected in 16 resistant isolates, and the bla KPC gene was seen in 14., Conclusions: The presence of these genes in P. stuartii multi- and extensively drug-resistant isolates indicates that the resistance arsenal of this species is increasing, thus limiting the therapeutic options.

Introduction: Pseudomonas aeruginosa is an opportunistic pathogen associated with healthcare-related infections, affecting mainly patients with underlying diseases and immunosuppression. This microorganism has several virulence mechanisms that favour its pathogenesis, including the production of biofilm. This study aimed to analyze the phenotypic production of biofilms, the occurrence of quorum sensing (QS) genes, and the clonal profile of clinical isolates of P. aeruginosa from colonized/infected patients in a tertiary hospital in Recife-PE., Methods: We obtained 21 isolates that were classified as infection isolates (II), and 10 colonization isolates (CI). The phenotypic analysis for biofilm production was performed quantitatively. The QS genes were detected by specific PCRs, and the clonal profile was assessed using ERIC-PCR., Results: Of the 31 isolates, 58.1 % (18/31) were biofilm producers, of which 70 % (7/10) were CI and classified as weakly adherent; 52.4 % (11/21) of the II produced biofilms, and were classified as weak (38.1 %, (8/21)), moderate (9.5 %, (2/21)), and strongly adherent (4.8 %, (1/21)). All isolates harbored the QS genes analyzed. In the clonal analysis, 26 distinct genetic profiles were identified, highlighting the presence of a clone in four samples, i.e., one infection isolate, and 3 colonization isolates., Conclusions: The detection of biofilm formation is important in P. aeruginosa in addition to the identification of colonization and infection isolates, especially from complex environments such as ICUs. Further, we define a strategy for monitoring and analyzing P. aeruginosa strains that can potentially cause infections in hospitalized patients.

Acinetobacter baumannii and Pseudomonas aeruginosa are the most relevant Gram-negative bacteria associated with hospital and opportunistic infections. This study aimed to evaluate the dynamics of drug-resistant A . baumannii and P . aeruginosa and biofilm formers from two public hospitals in northeastern Brazil. One hundred isolates (35 from A. baumannii and 65 from P. aeruginosa ) were identified using the automated Vitek ® 2 Compact method (bioMérieux) and confirmed using the MALDI-TOF (MS) mass spectrometry technique. Molecular experiments were performed by polymerase chain reaction (PCR) to detect the frequency of bla KPC , bla IMP , bla VIM , and bla SHV genes. The biofilm formation potential was evaluated using crystal violet in Luria Bertani Miller and trypticase soy broth culture media under the following conditions: at standard concentration, one quarter (25%) of the standard concentration and supplemented with 1% glucose. In addition, the genetic diversity of the isolates was verified by the ERIC-PCR technique. Isolates presented distinct resistance profiles with a high level of beta-lactam resistance. The highest index of genes detected was bla KPC (60%), followed by bla SHV (39%), bla VIM (8%), and bla IMP (1%). All the isolates were sensitive to the polymyxins tested and formed biofilms at different intensities. Twelve clones of A . baumannii and eight of P . aeruginosa were identified, of which few were indicative of intra- and interhospital dissemination. This study reveals the dispersion dynamics of these isolates in the hospital environment. The results demonstrate the importance of monitoring programs to combat the spread of these pathogens.

Introduction: The increasing reports of vancomycin-resistant Staphylococcus strains (VRS) haves caused concern worldwide, from the laboratory detection to patient management. This study aimed to identify the occurrence of VRS strains among healthcare professionals from a university hospital., Methods: A total of 102 Staphylococcus sp. isolates from healthcare professionals, obtained in a previous study were evaluated according to standard techniques for VRS detection., Results: After screening inoculation of plates containing 6µg/ml of vancomycin, 19 resistant isolates were identified. The susceptibility profile to other antimicrobials revealed 18 multidrug resistant isolates. The minimum inhibitory concentration (MIC) was determined by E-test and broth microdilution. According to E-tests, of 19 isolates grown in BHI-V6, four isolates presented MIC ≥ 128 µg/ml, seven with MIC ranging from 4 to 8 µg/ml, and eight with MIC ≤ 2µg/ml. By broth microdilution, 14 isolates presented MIC ≤ 2 µg/ml and five with MIC ≥ 16µg/ml. The presence of the gene vanA was determined by PCR in the five resistant isolates, and this gene was detected in one of the strains. Furthermore, among the 19 strains, the gene mecA was found in 13 (39,4%) isolates, including the strain carrying the gene vanA., Conclusions: Based on these results, we highlight the presence of one strain carrying both vanA and the mecA genes, as well as multidrug-resistant strains colonizing healthcare professionals, and their importance as potential vectors to spread strains carrying resistance genes in the hospital environment.

Introduction: Biofilm production is an important mechanism for the survival of Pseudomonas aeruginosa and its relationship with antimicrobial resistance represents a challenge for patient therapeutics. P. aeruginosa is an opportunistic pathogen frequently associated to nosocomial infections, especially in imunocompromised hosts., Objectives: Analyze the phenotypic biofilm production in P. aeruginosa isolates, describe clonal profiles, and analyze quorum sensing (QS) genes and the occurrence of mutations in the LasR protein of non-biofilm producing isolates., Methods: Isolates were tested for biofilm production by measuring cells adherence to the microtiter plates. Clonal profile analysis was carried out through ERIC-PCR, QS genes were by specific PCR., Results: The results showed that 77.5% of the isolates were considered biofilm producers. The results of genotyping showed 38 distinct genetic profiles. As for the occurrence of the genes, 100% of the isolates presented the lasR, rhlI and rhlR genes, and 97.5%, presented the lasI gene. In this study nine isolates were not biofilm producers. However, all presented the QS genes. Amplicons related to genes were sequenced in three of the nine non-biofilm-producing isolates (all presenting different genetic similarity profile) and aligned to the sequences of those genes in P. aeruginosa strain PAO1 (standard biofilm-producing strain). Alignment analysis showed an insertion of three nucleotides (T, C and G) causing the addition of an amino acid valine in the sequence of the LasR protein, in position 53., Conclusion: The modeling of the resulting LasR protein showed a conformational change in its structure, suggesting that this might be the reason why these isolates are unable to produce biofilm., (Copyright © 2018 Sociedade Brasileira de Infectologia. Published by Elsevier Editora Ltda. All rights reserved.)

Objective: To phenotypically evaluate biofilm production by Pseudomonas aeruginosa clinically isolated from patients with ventilator-associated pneumonia., Methods: Twenty clinical isolates of P. aeruginosa were analyzed, 19 of which were from clinical samples of tracheal aspirate, and one was from a bronchoalveolar lavage sample. The evaluation of the capacity of P. aeruginosa to produce biofilm was verified using two techniques, one qualitative and the other quantitative., Results: The qualitative technique showed that only 15% of the isolates were considered biofilm producers, while the quantitative technique showed that 75% of the isolates were biofilm producers. The biofilm isolates presented the following susceptibility profile: 53.3% were multidrug-resistant, and 46.7% were multidrug-sensitive., Conclusion: The quantitative technique was more effective than the qualitative technique for the detection of biofilm production. For the bacterial population analyzed, biofilm production was independent of the susceptibility profile of the bacteria, demonstrating that the therapeutic failure could be related to biofilm production, as it prevented the destruction of the bacteria present in this structure, causing complications of pneumonia associated with mechanical ventilation, including extrapulmonary infections, and making it difficult to treat the infection.

Pseudomonas aeruginosa, Acinetobacter spp. and Klebsiella spp. are three of the pathogens most frequently involved in infections of cancer patients, and the production of β -lactamases is a major mechanism of resistance due to its wide diversity of existing enzymes. Therefore, the aim of the present study was to investigate the microbiological profile and data related to patients and infections, and to search for β -lactamase genes in bacterial isolates from hospitalized cancer patients in a hospital in Recife, Pernambuco, Brazil. A total of 169 isolates were recovered between 2012 and 2014, of which 58 were P. aeruginosa, 36 were Acinetobacter spp. and 75 were Klebsiella spp. A high percentage of carbapenem resistance was observed in P. aeruginosa and Acinetobacter spp. Among the carbapenem-resistant bacteria, the blaSPM-1 gene was detected in P. aeruginosa (35.5 %) and Acinetobacter spp. (3.8 %), while blaKPC was detected in P. aeruginosa (25.8 %) only. Among the third- and fourth-generation cephalosporin-resistant strains, in Klebsiella spp. we detected the genes blaTEM (30.6 %), blaCTX-M (58.3 %) and blaKPC (5.6 %), and in Acinetobacter spp. only blaTEM (25.9 %). This the first report of an Acinetobacter baumannii blaSPM-1 gene carrier that has been isolated in Brazil. The most frequent cancer types were bowel tumour [14.8 %; 95 % confidence interval (CI95 %) 9.8-21.1 %], breast cancer (13.6 %; CI95 % 8.8-19.7 %) and prostate cancer (11.2%; CI95 % 6.9-17.0 %). These results therefore provide knowledge of susceptibility profile and resistance mechanisms and thus can contribute to the strategic formulation of hospital infection control plans and the rational use of antimicrobials, reducing mortality from infection levels in cancer patients.