Subgingival bacterial community profiles in HIV-infected Brazilian adults with chronic periodontitis.

BACKGROUND AND OBJECTIVE: To compare the subgingival microbial diversity between non-HIV-infected and HIV-infected individuals with chronic periodontitis using denaturing gradient gel electrophoresis (DGGE). MATERIAL AND METHODS: Thirty-two patients were selected: 11 were HIV-infected and 21 were non-HIV-infected, and all had chronic periodontitis. Periodontal measurements included probing depth, clinical attachment level, visible supragingival biofilm and bleeding on probing. Subgingival biofilm samples were collected from periodontal sites (50% with probing depth ≤ 4 mm and 50% with probing depth ≥ 5 mm) and whole-genomic-amplified DNA was obtained. The DNA samples were subjected to amplification of a 16S rRNA gene fragment using universal bacterial primers, followed by DGGE analysis of the amplified gene sequences. RESULTS: The non-HIV-infected group presented higher mean full-mouth visible supragingival biofilm (p = 0.004), bleeding on probing (p = 0.006), probing depth (p < 0.001) and clinical attachment level (p = 0.001) in comparison with the HIV-infected group. DGGE analysis revealed 81 distinct bands from all 33 individuals. Banding profiles revealed a higher diversity of the bacterial communities in the subgingival biofilm of HIV-infected patients with chronic periodontitis. Moreover, cluster and principal component analyses demonstrated that the bacterial community profiles differed between these two conditions. High interindividual and intra-individual variability in banding profiles were observed for both groups. CONCLUSION: HIV-infected patients with chronic periodontitis present greater subgingival microbial diversity. In addition, the bacterial communities associated with HIV-infected and non-HIV-infected individuals are different in structure.

Reliability of the MicroScan WalkAway PC21 panel in identifying and detecting oxacillin resistance in clinical coagulase-negative staphylococci strains.

The purpose of this study was to determine the reliability of the MicroScan WalkAway PosCombo21 (PC21) system for the identification of coagulase-negative staphylococci (CNS) strains and the detection of oxacillin resistance. Using molecular and phenotypic methods, 196 clinical strains were evaluated. The automated system demonstrated 100 % reliability for the identification of the clinical strains Staphylococcus haemolyticus, Staphylococcus hominis and Staphylococcus cohnii; 98.03 % reliability for the identification of Staphylococcus epidermidis; 70 % reliability for the identification of Staphylococcus lugdunensis; 40 % reliability for the identification of Staphylococcus warneri; and 28.57 % reliability for the identification of Staphylococcus capitis, but no reliability for the identification of Staphylococcus auricularis, Staphylococcus simulans and Staphylococcus xylosus. We concluded that the automated system provides accurate results for the more common CNS species but often fails to accurately identify less prevalent species. For the detection of oxacillin resistance, the automated system showed 100 % specificity and 90.22 % sensitivity. Thus, the PC21 panel detects oxacillin-resistant strains, but is limited by the heteroresistance that is observed when using most phenotypic methods.

Antimicrobial synergism against different lineages of methicillin-resistant Staphylococcus aureus carrying SCCmec IV.

AIM: To evaluate the synergistic activity of antimicrobial drugs against lineages of methicillin-resistant Staphylococcus aureus (MRSA) carrying SCCmec IV. The biofilm production and related genes were also detected. METHODS AND RESULTS: Forty two MRSA isolates were tested for biofilm production and related genes. Biofilm/biomass susceptibility to gentamicin (G), linezolid (L), rifampicin (R) and vancomycin (V) was determined for six isolates from three lineages prevalent in Rio de Janeiro hospitals in concentrations ranging from 0·25 to 64 µg ml(-1). Biomass was evaluated by microtitre plate test and number of viable cells (CFU cm(-2)) and inspected by epifluorescence microscopy. All isolates presented the icaA and sasG genes, but only 38% were biofilm producers. There were 50 and 45% biomass reductions when concentrations ≥4 µg ml(-1) of R or L and ≥16 µg ml(-1) of G or V, respectively, were used. Synergism tests produced a 55% biomass reduction with R(2µgml-1) + G(16µgml-1), R(2µgml-1) + L(2µgml-1), R(2µgml-1) + V(4µgml-1), and L(2µgml-1) + V(4µgml-1). Number of viable cells was reduced from 2 to 3 logs with R(2µgml-1) + L(2µgml-1) and R(2µgml-1) + V(4µgml-1). CONCLUSIONS: Synergisms involving R plus L and R plus V caused important reductions in biofilm/biomass and the number of viable cells. Drug combinations should be considered in the chemotherapies of MRSA-SCCmec IV infections. SIGNIFICANCE AND IMPACT OF THE STUDY: Biofilms in MRSA infections restrict the clinical choice of antimicrobials. Thus, knowledge of the best options for monotherapy and drug synergisms could improve clinical results.

Antibacterial effect of coffee: calcium concentration in a culture containing teeth/biofilm exposed to Coffea Canephora aqueous extract.

UNLABELLED: This study determined the changes of calcium concentration in a medium containing teeth/biofilm exposed to Coffea canephora extract (CCE). Enamel fragments were randomly fixed into two 24-well polystyrene plates containing BHI. Pooled human saliva was added to form biofilm on fragments. Specimens were divided into treatment groups (G, n = 8 per group) and treated with 50 µl daily for 1 min per week, as follows: G1, 20% CCE; G2, Milli-Q water (negative control); G3, antibiotic (positive control). Six fragments represented the blank control (G4). The calcium content was observed at baseline, 4 and 7 days of treatment by atomic-absorption spectrophotometry. Cross-sectional hardness of enamel was a demineralization indicator. Calcium increased in the medium after 4 and 7 days of treatment in G1 (3·80 ± 1·3 mg l(-1) and 4·93 ± 2·1 mg l(-1) , respectively) and G3 (4th day = 5·7 ± 1·8 mg l(-1) ; 7th day = 6·7 ± 3·5 mg l(-1) ) (P > 0·05). Calcium from G2 decreased after 7 days, which was different from G3 (P < 0·05). The lower calcium content, at the end of the experiment, was represented by G4, 2·16 ± 0·2 mg l(-1) . The increase in calcium after treatment with CCE is probably due to its antibacterial effect, which caused the bacterial lysis and consequent release of calcium in the medium. SIGNIFICANCE AND IMPACT OF THE STUDY: This study revealed an inhibitory action of Coffea canephora against dental biofilm. This coffee species caused bacterial lysis and consequent release of calcium into the medium. Furthermore, the advantage of coffee as an antibacterial beverage is that it is consumed in a concentrated form (6-10%) as opposed to various medicinal infusions that have shown such effect in vitro and are usually consumed at 1-2%. Therefore, a light roasted C. canephora aqueous extract can be considered as a potential anticariogenic substance.

Increase in methicillin-resistant Staphylococcus spp. colonization among pregnant individuals during COVID-19 pandemic.

Methicillin-resistant Staphylococcus (MRS) has been associated with neonatal infections, with colonization of the anovaginal tract being the main source of vertical transmission. The COVID-19 pandemic has altered the frequency of antibiotic usage, potentially contributing to changes in the dynamics of bacterial agents colonizing humans. Here we determined MRS colonization rates among pregnant individuals attending a single maternity in Rio de Janeiro, Brazil before (January 2019-March 2020) and during (May 2020-March 2021) the COVID-19 pandemic. Anovaginal samples (n = 806 [521 samples before and 285 during the pandemic]) were streaked onto chromogenic media. Colonies were identified by MALDI-TOF MS. Detection of mecA gene and SCCmec typing were assessed by PCR and antimicrobial susceptibility testing was done according to CLSI guidelines. After the onset of the pandemic, MRS colonization rates increased significantly (p < 0.05) from 8.6% (45) to 54.7% (156). Overall, 215 (26.6%) MRS isolates were detected, of which S. haemolyticus was the most prevalent species (MRSH, 84.2%; 181 isolates). SCCmec type V was the most frequent among MRS (63.3%; 136), and 31.6% (68) of MRS strains had a non-typeable SCCmec, due to new combinations of ccr and mecA complexes. Among MRS strains, 41.9% (90) were resistant to at least 3 different classes of antimicrobial agents, and 60% (54) of them were S. haemolyticus harboring SCCmec V. MRS colonization rates and the emergence of multidrug-resistant variants detected in this study indicate the need for continuing surveillance of this important pathogen within maternal and child populations.

Reduced-order modeling and analysis of dynamic cerebral autoregulation via diffusion maps.

Objective.A data-driven technique for parsimonious modeling and analysis of dynamic cerebral autoregulation (DCA) is developed based on the concept of diffusion maps. Specifically, first, a state-space description of DCA dynamics is considered based on arterial blood pressure, cerebral blood flow velocity, and their time derivatives. Next, an eigenvalue analysis of the Markov matrix of a random walk on a graph over the dataset domain yields a low-dimensional representation of the intrinsic dynamics. Further dimension reduction is made possible by accounting only for the two most significant eigenvalues. The value of their ratio indicates whether the underlying system is governed by active or hypoactive dynamics, indicating healthy or impaired DCA function, respectively. We assessed the reliability of the technique by considering healthy individuals and patients with unilateral internal carotid artery (ICA) stenosis or occlusion. We computed the sensitivity of the technique to detect the presumed side-to-side difference in the DCA function of the second group (assuming hypoactive dynamics on the occluded or stenotic side), using McNemar's chi square test. The results were compared with transfer function analysis (TFA). The performance of the two methods was also compared under the assumption of missing data.Main results.Both diffusion maps and TFA suggested a physiological side-to-side difference in the DCA of ICA stenosis or occlusion patients with a sensitivity of 81% and 71%, respectively. Further, both two methods suggested the difference between the occluded or stenotic side and any two sides of the healthy group. However, the diffusion maps captured additional difference between the unoccluded side and the healthy group, that TFA did not. Furthermore, compared to TFA, diffusion maps exhibited superior performance when subject to missing data.Significance.The eigenvalues ratio derived using the diffusion maps technique can be potentially used as a reliable and robust biomarker for assessing how active the intrinsic dynamics of the autoregulation is and for indicating healthy versus impaired DCA function.

Staphylococcus haemolyticus as an important hospital pathogen and carrier of methicillin resistance genes.

Phenotypic and molecular methods were used to characterize the antibiotic resistance of 64 clinical isolates of Staphylococcus haemolyticus. By PCR of the mecA gene, 87% were found to be methicillin resistant. Approximately 55% harbored staphylococcal cassette chromosome mec element (SCCmec) type V, and only one SCCmec type IV. Many isolates (75%) displayed multiresistance, and pulsotype analysis showed a high diversity.

Rhabdias filicaudalis n. sp. (Nematoda: Rhabdiasidae) from the snake Spilotes pullatus (Serpentes: Colubridae) in Brazil.

Rhabdias filicaudalis n. sp. (Nematoda: Rhabdiasidae) from the lung of Spilotes pullatus (Serpentes: Colubridae) is described. The host snake was captured in the municipality of Avaré, São Paulo State, Brazil. Rhabdias filicaudalis n. sp. differs from all other species by the combination of the following characters: straight body, truncated anterior end, six weakly developed lips arranged in two opposite groups of three, pre-equatorial vulva, oesophagus length/body length ratio (%) 4.9-7.5 (5.8 +/- 0.6), nerve ring distance from anterior end/oesophagus length ratio (%) 36.9-61.1 (49.8 +/- 6.4), tail length/body length ratio (%) 3.0-5.5 (4.0 +/- 0.5), vulva distance from anterior end/body length ratio (%) 39.9-51.7 (45.9 +/- 3.5), and a cuticular filiform tail tip.

Morphological and molecular characterization of Strongyloides ophidiae (Nematoda, Strongyloididae).

The aim of the present study is to report morphological data from parasitic female, rhabditoid and filarioid larvae, free-living female worms and eggs of Strongyloides ophidiae (Nematoda, Strongyloididae). In addition, a molecular DNA analysis was carried out using a pool of eight S. ophidiae parasitic females. Samples were obtained from the small intestine of Oxyrhopus guibei (Serpentes, Colubridae) collected in the municipality of Lençóis Paulista, State of São Paulo, Brazil. DNA amplification by polymerase chain reaction (PCR) resulted in a 350 bp band for samples containing S. ophidiae and Strongyloides venezuelensis DNA. Strongyloides ophidiae nucleotide sequence analysis showed 98% similarity with Strongyloides procyonis and 97% with Strongyloides cebus, Strongyloides stercoralis, Strongyloides fuelleborni and Strongyloides sp. from snakes.

Joint time-frequency analysis of dynamic cerebral autoregulation using generalized harmonic wavelets.

OBJECTIVE: To develop a joint time-frequency analysis technique based on generalized harmonic wavelets (GHWs) for dynamic cerebral autoregulation (DCA) performance quantification. APPROACH: We considered two groups of human subjects to develop and validate the method: 55 healthy volunteers and 35 stroke-free subjects with unilateral internal carotid artery stenosis (CAS). We determined the mean and coherence-weighted average of the phase shift (PS) of appropriately defined GHW-based transfer functions, based on data points over the joint time-frequency domain. We compared agreement of standard transfer function analysis (TFA) and GHW analyses in healthy subjects using Bland-Altman plots. We assessed sensitivity of each metric to detect the presumed side-to-side difference in DCA function in CAS subjects (with decreased PS on the occluded side), using McNemar's chi square test to compare each metric to the standard TFA approach. An alternative Morlet wavelet-based approach was also considered. MAIN RESULTS: The GHW and TFA methods exhibited strong agreement in healthy subjects. Among CAS subjects, GHW metrics outperformed TFA and Morlet wavelet-based approaches in identifying expected side-to-side differences: TFA sensitivity was 40.0% (95%CI 23.9-57.9), Morlet 60.0% (95%CI 42.1-76.1), and GHW >70% for both metrics (GHW mean PS sensitivity 74.3, 95%CI 56.7-87.5, p  = 0.0027 versus TFA; GHW coherence-weighted PS sensitivity 71.4, 95%CI 53.7-85.4, p  = 0.0009 versus TFA). SIGNIFICANCE: In comparison to the widely used stationary Fourier transform-based TFA and to Morlet wavelet-based analysis, our data suggest that the GHW-based analysis performs better in identifying DCA asymmetry between the two cerebral hemispheres in patients with high grade unilateral carotid stenosis. Our method may provide enhanced confidence in employing DCA metrics as a sensitive diagnostic tool for detecting impaired DCA function in a variety of pathological settings.

Pseudomonas aeruginosa epidemic strain carrying bla(SPM) metallo-beta-lactamase detected in Rio de Janeiro, Brazil.

The present study was designed to characterize beta-lactamase genes and evaluate polymerase chain reaction (PCR) typing for multidrug-resistant Pseudomonas aeruginosa pulsed-field gel electrophoresis (PFGE) genotype A isolates from Rio de Janeiro, Brazil, collected between April 1999 and March 2000 and one additional isolate collected in June 2002. As reported previously, all of the genotype A isolates produced non-characterized metallo-beta-lactamase. These isolates (22) were screened for the bla(SPM) gene by PCR and dot-blotting. Isolates were typed by PCR fingerprinting with primers RAPD-1, 272, 208, 1290, ERIC-1 and ERIC-2. The bla(SPM) gene was detected in 18 (82%) of the 22 isolates. PCR fingerprinting gave results that correlated with PFGE, except with primer 1290. In Rio de Janeiro and other Brazilian states, nearly all SPM-producing P. aeruginosa isolates belong to a single PFGE type accounting for a large proportion of drug-resistant P. aeruginosa hospital infections. RAPD PCR fingerprinting may be a useful technique to screen for an epidemic multidrug-resistant strain in Brazil.

Emergence of high-level mupirocin resistance in methicillin-resistant Staphylococcus aureus isolated from Brazilian university hospitals.

Surveillance for methicillin-resistant Staphylococcus aureus (MRSA) was implemented in Rio de Janeiro and Uberlândia University Hospitals, which had different policies on use of mupirocin. One hundred fourteen multiresistant MRSA strains were isolated from 62 patients. Mupirocin resistance was observed in 63% of strains in Rio de Janeiro, where there was extensive use of topical mupirocin, and 6.1% in Uberlândia, where its use was rare.

Detection of ileS-2 gene encoding mupirocin resistance in methicillin-resistant Staphylococcus aureus by multiplex PCR.

The presence of the ileS-2 gene, responsible for mupirocin resistance, in clinical isolates of methicillin-resistant Staphylococcus aureus was determined by multiplex polymerase chain reaction. Three pairs of primers were used, which yielded specific fragments of femA (encoding a unique feature of S. aureus), mecA (encoding resistance to methicillin) and ileS-2 genes. The multiplex polymerase chain reaction system is an easy and time-saving technique that, together with a rapid method for DNA extraction by boiling, may be incorporated as a routine analysis in clinical diagnostic laboratories.

Meticillin-resistant Staphylococcus aureus: spread of specific lineages among patients in different wards at a Brazilian teaching hospital.

This study aimed to characterize meticillin-resistant Staphylococcus aureus (MRSA) lineages circulating in a Brazilian teaching hospital. MRSA isolates from nasal swabs were evaluated to assess antimicrobial susceptibility, staphylococcal cassette chromosome mec (SCCmec), Panton-Valentine leucocidin status, pulsed-field gel electrophoresis profile and multi-locus sequence type (MLST) analysis. Eighty-three MRSA isolates were analysed. SCCmec III (43.4%) and IV (49.4%) were predominant. ST1-IV (USA400) was more common in internal medicine (P = 0.002) whereas 'clone M' (SCCmec III) was more common in the medical and surgical intensive care unit (P = 0.004), and all isolates were ST5-IV (USA800) in dermatology (P < 0.001). These data improved the understanding of the MRSA epidemiology inside the hospital and helped to establish effective control measures.

SubMICs of penicillin and erythromycin enhance biofilm formation and hydrophobicity of Corynebacterium diphtheriae strains.

Subinhibitory concentrations (subMICs) of antibiotics may alter bacterial surface properties and change microbial physiology. This study aimed to investigate the effect of a subMIC (⅛ MIC) of penicillin (PEN) and erythromycin (ERY) on bacterial morphology, haemagglutinating activity, cell-surface hydrophobicity (CSH) and biofilm formation on glass and polystyrene surfaces, as well as the distribution of cell-surface acidic anionic residues of Corynebacterium diphtheriae strains (HC01 tox(-) strain; CDC-E8392 and 241 tox(+) strains). All micro-organisms tested were susceptible to PEN and ERY. Growth in the presence of PEN induced bacterial filamentation, whereas subMIC of ERY caused cell-size reduction of strains 241 and CDC-E8392. Adherence to human erythrocytes was reduced after growth in the presence of ERY, while CSH was increased by a subMIC of both antibiotics in bacterial adherence to n-hexadecane assays. Conversely, antibiotic inhibition of biofilm formation was not observed. All strains enhanced biofilm formation on glass after treatment with ERY, while only strain 241 increased glass adherence after cultivation in the presence of PEN. Biofilm production on polystyrene surfaces was improved by ⅛ MIC of ERY. After growth in the presence of both antimicrobial agents, strains 241 and CDC-E8392 exhibited anionic surface charges with focal distribution. In conclusion, subMICs of PEN and ERY modified bacterial surface properties and enhanced not only biofilm formation but also cell-surface hydrophobicity. Antibiotic-induced biofilm formation may contribute to the inconsistent success of antimicrobial therapy for C. diphtheriae infections.

Loss of lingual sensitivity and slightly increased size signaling schwannoma in a patient with mixed conjunctive tissue disease.

The purpose of this study was to describe an unusual case of a lingual schwannoma associated with a mixed connective tissue disease (MCTD). A case report. A lingual schwannoma with loss of lingual sensitivity and slightly increased size in an 18-year-old patient with MCTD was correctly diagnosed through a biopsy and no reoccurrence was observed one year after the surgical removal of the tumor and sensitivity returned 3 months after surgery. This case was considered uncommon, making the clinical diagnosis challenging in view of the diversity of possibilities for its differential diagnosis, thus showing the importance of a biopsy to confirm the diagnosis and long term follow up in such cases.

Short communication: polymicrobial community in teeth associated with severe early-childhood caries.

AIM: This was to characterise the microbial diversity in the complex dental plaque of children with severe early-childhood caries (S-ECC), using the denaturing gel gradient electrophoresis (DGGE) technique. METHODS: The DGGE technique was used as a diagnostic tool to analyse samples from the oral cavity of a patient with S-ECC. Dental plaque samples from a 3-year-old child with S-ECC were taken from the primary central maxillary incisor (biofilm on vestibular surface), primary maxillary molar (biofilm on vestibular surface), primary central maxillary incisor (dentine), primary maxillary molar (dentine) and saliva and then analysed by PCR-DGGE. RESULTS: Three bands occurred in all samples, moreover, 86% of similarity was observed in the pattern of bands between incisor and molar teeth biofilm samples, including four similar bands. CONCLUSION: DGGE is a valuable tool for differentiating the microbial composition of the oral plaque in S-ECC children.

Inhibitory properties of Coffea canephora extract against oral bacteria and its effect on demineralisation of deciduous teeth.

OBJECTIVES: The antibacterial activity of Coffea canephora extract was evaluated in vitro against Streptococcus mutans and Streptococcus sobrinus. The viability of planktonic cells was analysed by susceptibility tests (MIC and MBC) and time-kill assays. The effect of the extract on dental demineralisation was also investigated. METHODS: Primary 1st molar fragments (n=24) were inoculated with a saliva pool and sustained in a multiple plaque growth system for 10 days to form biofilm. The biofilm was treated with light roasted C. canephora extract at 20%, Milli-Q water (negative control) and chlorhexidine (positive control) once a day, during a week. Blank controls comprised fragments without treatment. Biofilm pH was monitored in the last day of treatment. Changes in tooth mineralisation were assessed by cross-sectional microhardness (CSMH) test. RESULTS: MIC and MBC for S. mutans were 7±2 mg/mL and 160±0 mg/mL, respectively, showing no activity for S. sobrinus. The extract produced a 4-log reduction in the number of colonies of S. mutans after 3-h treatment (p<0.05) with undiluted extract (20%) and MBC concentration (16%). There was no difference among negative/blank controls and coffee plaque pH. Differences between CSMH values of dental fragments subjected to the coffee extract and to chlorhexidine were not significant. At depths up to 30 µm from the enamel surface, coffee extract and chlorhexidine promoted higher CSMH values when compared to blank/negative controls (p<0.05). CONCLUSION: Our data suggest that light roasted C. canephora extract is beneficial as an anticariogenic substance.

Typing of Staphylococcus aureus from surgical site infections: comparison of pulsed-field gel electrophoresis (PFGE) and PCR technique using repetitive extragenic palindromic (rep) and Tn916-Shine-Dalgarno (TnSD) target sequences.

Staphylococcus aureus continues to be the main cause of surgical site infections. DNA typing is useful for studying this type of infection and establishing control programs within hospitals. In this study 19 S. aureus strains were isolated from surgical site infections of 19 patients, between August and December 1994 at the Rio de Janeiro University Hospital. The strains were typed by pulsed-field gel electrophoresis (PFGE) and by two polymerase chain reaction techniques targeting the repetitive extragenic palindromic and Tn916-Shine Dalgarno sequences. Analysis of the PFGE patterns divided the collection into 15 types, while PCR techniques identified 11 distinct strain patterns. There were two clusters, 1 of four strains and 1 of two strains with related PFGE and PCR patterns. Of the remaining strains, 10 were clustered in 5 PCR patterns but their PFGE patterns showed 4 to 6 different bands, and they were considered to be possibly related. The comparison of the S. aureus typing systems in the present study indicated that the PCR methods are useful for initial screening of genetically related isolates, but strains with identical PCR fingerprint need to be typed with PFGE for detailed strain differentiation.