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AIMS: To evaluate the prevalence, molecular characteristics, antimicrobial susceptibility, and epithelial invasion of Streptococcus agalactiae strains isolated from pregnant women and newborns in Rio de Janeiro, Brazil. METHODS AND RESULTS: A total of 67 S. agalactiae isolates, 48 isolates from pregnant women and 19 from neonates, were analyzed. Capsular type Ia and V were predominant (35.8%/each). The multilocus sequence typing analysis revealed the presence of 19 STs grouped into 6 clonal complexes with prevalence of CC17/40.3% and CC23/34.3%. The lmb and iag virulence genes were found in 100% of isolates. Four S. agalactiae strains, belonging to CC17/ST1249 and CC23/ST23, were able to adhere to A549 respiratory epithelial cells. Antimicrobial resistance was verified mainly to tetracycline (85%), erythromycin (70.8%), and clindamycin (58.3%). Four S. agalactiae isolates were multidrug resistant. The resistance genes tested were found in 92.5% of isolates for tetM, 58.2% for ermB, 28.4% for mefAE, and 10.4% for tetO. CONCLUSION: The study showed a high prevalence of virulence and antimicrobial genes in S. agalactiae strains isolated from pregnant women and newborns, supporting the idea that continued surveillance is necessary to identify risk factors and perform long-term follow-up in pregnant women and neonates in Rio de Janeiro.
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Antibacterianos , Células Epiteliais , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Infecções Estreptocócicas , Streptococcus agalactiae , Streptococcus agalactiae/genética , Streptococcus agalactiae/efeitos dos fármacos , Streptococcus agalactiae/isolamento & purificação , Feminino , Humanos , Brasil , Gravidez , Infecções Estreptocócicas/microbiologia , Antibacterianos/farmacologia , Recém-Nascido , Células Epiteliais/microbiologia , Farmacorresistência Bacteriana/genética , Adulto , Fatores de Virulência/genética , Complicações Infecciosas na Gravidez/microbiologia , Farmacorresistência Bacteriana Múltipla/genética , Virulência/genéticaRESUMO
Hospital bloodstream infection (BSI) caused by methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of morbidity and mortality and is frequently related to invasive procedures and medically complex patients. An important feature of MRSA is the clonal structure of its population. Specific MRSA clones may differ in their pathogenic, epidemiological, and antimicrobial resistance profiles. Whole-genome sequencing is currently the most robust and discriminatory technique for tracking hypervirulent/well-adapted MRSA clones. However, it remains an expensive and time-consuming technique that requires specialized personnel. In this work, we describe a pangenome protocol, based on binary matrix (1,0) of open reading frames (ORFs), that can be used to quickly find diagnostic, apomorphic sequence mutations that can serve as biomarkers. We use this technique to create a diagnostic screen for MRSA isolates circulating in the Rio de Janeiro metropolitan area, the RdJ clone, which is prevalent in BSI. The method described here has 100% specificity and sensitivity, eliminating the need to use genomic sequencing for clonal identification. The protocol used is relatively simple and all the steps, formulas and commands used are described in this work, such that this strategy can also be used to identify other MRSA clones and even clones from other bacterial species.
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Streptococcus pyogenes is known to be associated with a variety of infections, from pharyngitis to necrotizing fasciitis (flesh-eating disease). S. pyogenes of the ST62/emm87 lineage is recognized as one of the most frequently isolated lineages of invasive infections caused by this bacterium, which may be involved in hospital outbreaks and cluster infections. Despite this, comparative genomic and phylogenomic studies have not yet been carried out for this lineage. Thus, its virulence and antimicrobial susceptibility profiles are mostly unknown, as are the genetic relationships and evolutionary traits involving this lineage. Previously, a strain of S. pyogenes ST62/emm87 (37-97) was characterized in our lab for its ability to generate antibiotic-persistent cells, and therapeutic failure in severe invasive infections caused by this bacterial species is well-reported in the scientific literature. In this work, we analyzed genomic and phylogenomic characteristics and evaluated the virulence and resistance profiles of ST62/emm87 S. pyogenes from Brazil and international sources. Here we show that strains that form this lineage (ST62/emm87) are internationally spread, involved in invasive outbreaks, and share important virulence profiles with the most common emm types of S. pyogenes, such as emm1, emm3, emm12, and emm69, which are associated with most invasive infections caused by this bacterial species in the USA and Europe. Accordingly, the continued increase of ST62/emm87 in severe S. pyogenes diseases should not be underestimated.
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99mTc-EDDA/HYNIC-TOC is an easily available and cheaper radionuclide that could be used for somatostatin-receptor-based imaging of neuroendocrine tumours (NETs). We aimed to evaluate the diagnostic performance of 99mTc-EDDA/HYNIC-TOC compared to111In-DTPA-octreotide in patients (pts) with NETs. We performed a prospective diagnostic study including pts with biopsy-confirmed NET and at least one visible lesion at conventional imaging. Two independent nuclear medicine physicians evaluated pts who underwent 99mTc and 111In scans and images. The primary outcome was comparative diagnostic accuracy of 99mTc and 111In. Secondary outcomes include safety. Nine pts were included and performed 14 paired scans. Overall, 126 lesions were identified. 99mTc demonstrated superior sensitivity both when all images were analysed (93.7, 95% CI 88.1% - 96.8% versus 74.8%, 95% CI 66.6 - 81.6%, p < 0.001) and when liver-specific images were analysed (97.8%, 95% CI 92.7% - 99.5% versus 85.1%, 95% CI 76.6% - 91.0%, p < 0.001). 99mTc was also associated with a lower negative likelihood ratio (LR) (0.002, 95% CI 0.009 - 0.1 versus 0.19, 95% CI 0.12 - 0.42, p = 0.009) when evaluating hepatic lesions. Adverse events happened in 3 pts after 111In and in 2 pts after 99mTc, all grade 1. The 99mTc demonstrated a higher sensitivity overall and a better negative LR in liver-specific images compared to 111In in pts with NETs. Our findings suggest that 99mTc is an alternative to 111In and is especially useful in ruling out liver metastases. NCT02691078.
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BACKGROUND: Typing of staphylococcal cassette chromosome mec (SCCmec) elements is commonly used for studies on the molecular epidemiology of MRSA. OBJECTIVES: To perform an investigation centred on uncovering the reasons for misclassification of MRSA clonal complex 5 (CC5) SCCmec type II clinical isolates in our laboratory. METHODS: MRSA isolates from CC5 were subjected to WGS and SCCmec typing. RESULTS: This investigation led to the discovery that the classification failure was due to an insertion of IS1272 carrying the fabI gene on a transposable element (TnSha1) that confers increased MIC to the biocide triclosan. Genomic analysis revealed that fabI was present in 25% of the CC5 MRSA isolates sampled. The frequency of TnSha1 in our collection was much higher than that observed among publicly available genomes (0.8%; nâ=â24/3142 CC5 genomes). Phylogenetic analyses revealed that genomes in different CC5 clades carry TnSha1 inserted in different integration sites, suggesting that this transposon has entered CC5 MRSA genomes on multiple occasions. In at least two genotypes, ST5-SCCmecII-t539 and ST5-SCCmecII-t2666, TnSha1 seems to have entered prior to their divergence. CONCLUSIONS: Our work highlights an important misclassification problem of SCCmecII in isolates harbouring TnSha1 when Boye's method is used for typing, which could have important implications for molecular epidemiology of MRSA. The importance of increased-MIC phenotype is still a matter of controversy that deserves more study given the widespread use of triclosan in many countries. Our results suggest expanding prevalence that may indicate strong selection for this phenotype.
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Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Triclosan , Humanos , Infecções Estafilocócicas/epidemiologia , Triclosan/farmacologia , Testes de Sensibilidade Microbiana , Filogenia , DNA Bacteriano/genética , CromossomosRESUMO
We typed 600 methicillin-resistant Staphylococcus aureus (MRSA) isolates collected in 51 hospitals in the Rio de Janeiro, Brazil, metropolitan area during 2014-2017. We found that multiple new clonal complex (CC) 5 sequence types had replaced previously dominant MRSA lineages in hospitals. Whole-genome analysis of 208 isolates revealed an emerging sublineage of multidrug-resistant MRSA, sequence type 105, staphylococcal cassette chromosome mec II, spa t002, which we designated the Rio de Janeiro (RdJ) clone. Using molecular clock analysis, we hypothesized that this lineage began to expand in the Rio de Janeiro metropolitan area in 2009. Multivariate analysis supported an association between bloodstream infections and the CC5 lineage that includes the RdJ clone. Compared with other closely related isolates, representative isolates of the RdJ clone more effectively evaded immune function related to monocytic cells, as evidenced by decreased phagocytosis rate and increased numbers of viable unphagocytosed (free) bacteria after in vitro exposure to monocytes.
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Bacteriemia , Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Bacteriemia/epidemiologia , Brasil/epidemiologia , Humanos , Staphylococcus aureus Resistente à Meticilina/genética , Monócitos , Infecções Estafilocócicas/epidemiologiaRESUMO
BACKGROUND AND OBJECTIVES: Staphylococcus aureus is an important pathogen and a frequent cause of infections associated with biofilm production in implantable medical devices. Biofilm production can be induced by sub-inhibitory concentrations (sub-MICs) of certain antibiotics, but few studies have researched this occurrence in S. aureus. In this study, we investigated the effect of sub-MICs of rifampicin and minocycline on biofilm production by five clinical and five non-clinical S. aureus isolates. METHODS: Microtiter Plate assay and Congo Red Agar Test were used to analyze the biofilm production. The biofilm composition was evaluated by the detachment assay with sodium metaperiodate and proteinase K. RESULTS: Rifampicin sub-MICs induced very high biofilm formation in seven isolates that were non-producers in Tryptic Soy Broth. In one producer isolate, the biofilm formation level was not affected by sub-MICs of this drug. Sub-MICs of minocycline did not induce biofilm production in all isolates tested and in two producer isolates, instead, MIC/2 and MIC/4 inhibited biofilm production. The results of the drugs in combination were similar to those with rifampicin alone. The biofilm matrix was identified as polysaccharide, except for one producer isolate, classified as proteinaceous. Polysaccharide biofilm producer isolates, when grown on Congo Red Agar without sucrose, but with sub-MICs of rifampicin, showed results in agreement with those obtained in Microtiter Plate Test. CONCLUSION: The high biofilm production induced by sub-MICs of rifampicin has potential clinical relevance, because this is one of the drugs commonly used in the impregnation of catheters. In addition, it is used adjunctively to treat certain S. aureus infections.
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AbstractPrevious studies by us demonstrated the antidepressant-like and antinociceptive effects of lipophilic extracts and dimeric acyl-phloroglucinols from species of the genus Hypericum native to Southern Brazil. Uliginosin B and HC1 (an enriched phloroglucinol fraction from Hypericum caprifoliatum) are able to inhibit monoamine synaptosomal uptake without binding to the monoaminergic sites on neuronal transporters, unlike classical antidepressants. The current study aimed at investigating the action of H. caprifoliatum Cham. & Schltdl. and Hypericum polyanthemum Klotzsch ex Reichardt, Hypericaceae, cyclohexane extracts and their main component, HC1 and uliginosin B, on G protein coupled receptors by using the [35S]-guanosine-5′-O-(3-thio)triphosphate ([35S]-GTPγS) binding assay, which reveals the G protein activity. The antidepressant-like effect of acute (one or three treatments within 24 h) and repeated (five days with and without a three day wash-out) treatments with the cyclohexane extracts was evaluated using the rat forced swimming test. The [35S]-GTPγS binding to monoamines and opioid receptors stimulated by agonists was performed ex vivo in brain membranes of rats acutely or repeatedly treated with the cyclohexane extracts. The effect of HC1 and Uliginosin B on [35S]-GTPγS binding assay was performed by direct incubation with brain membranes in the absence of agonists. Their antidepressant-like effect was evaluated through the mice forced swimming test. The extracts, HC1 and Uliginosin B showed antidepressant-like effect in the forced swimming test. The acute treatments with extracts increased the [35S]-GTPγS binding stimulated by the monoamines, while after five days of treatment the [35S]-GTPγS binding was reduced even after three day wash-out. These effects are not due to HC1 or Uliginosin B interaction with the receptors, since direct incubation with these phloroglucinols did not affect [35S]-GTPγS binding to membranes. Our findings indicate that H. caprifoliatum and H. polyanthemumextracts bring about adaptive changes in monoamine receptors, which reinforces their antidepressant-like profile.
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Valeriana glechomifolia, a native species from southern Brazil, presents antidepressant-like activity and diene valepotriates (VAL) contribute to the pharmacological properties of the genus. It is known that depression can develop on an inflammation background in vulnerable patients and antidepressants present anti-inflammatory properties. We investigated the effects of VAL (10 mg/kg, p.o.) on sickness and depressive-like behaviors as well as proinflammatory cytokines (IL-1ß and TNF-α) and BDNF expression in the cortex of mice exposed to a 5 min swimming session (as a stressful stimulus) 30 min before the E. coli LPS injection (600 µg/kg, i.p.). The forced swim + LPS induced sickness and depressive-like behaviors, increased the cortical expression of IL-1ß and TNF-α, and decreased BDNF expression. VAL was orally administered to mice 1 h before (pretreatment) or 5 h after (posttreatment) E. coli LPS injection. The pretreatment with VAL restored the behavioral alterations and the expression of cortical proinflammatory cytokines in LPS-injected animals but had no effects on BDNF expression, while the posttreatment rescued only behavioral alterations. Our results demonstrate for the first time the positive effects of VAL in an experimental model of depression associated with inflammation, providing new data on the range of action of these molecules.
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The forced swim test (FST) is widely used to evaluate the antidepressant-like activity of compounds and is sensitive to stimuli that cause depression-like behaviors in rodents. The immobility behavior observed during the test has been considered to represent behavioral despair. In addition, some studies suggest that the FST impairs rats' performance on cognitive tests, but these findings have rarely been explored. Thus, we investigated the effects of the FST on behavioral tests related to neuropsychiatric diseases that involve different cognitive components: novel object recognition (NOR), the object location test (OLT) and prepulse inhibition (PPI). Brain-derived neurotrophic factor (BDNF) levels in the frontal cortex and hippocampus were evaluated. The rats were forced to swim twice (15-min session followed by a 5-min session 24h later) and underwent cognitive tests 24h after the last swimming exposure. The FST impaired the rats' performance on the OLT and reduced the PPI and acoustic startle responses, whereas the NOR was not affected. The cognitive impairments were not correlated with an immobility behavior profile, but a significant negative correlation between the frontal BDNF levels and immobility behavior was identified. These findings suggest a protective role of BDNF against behavioral despair and demonstrate a deleterious effect of the FST on spatial memory and pre-attentive processes, which point to the FST as a tool to induce cognitive impairments analogous to those observed in depression and in other neuropsychiatric disorders.