Marlei Gomes da Silva

In vitro activity of Brazilian medicinal plants, naturally occurring naphthoquinones and their analogues, against methicillin-resistant Staphylococcus aureus.

Fourteen extracts from Brazilian traditional medicinal plants used to treat infectious diseases were used to look for potential antimicrobial activity against multiresistant bacteria of medical importance. Staphylococcus aureus strains were susceptible to extracts of Punica granatum and Tabebuia avellanedae. The minimum inhibitory concentrations (MICs) of the total extracts and of additional fractions of these plants were determined by employing strains of methicillin-resistant (MRSA) and -sensitive (MSSA) S. aureus, including isolates of the PFGE clone A, which is prevalent in Brazil and two ATCC reference strains. A mixture of ellagitannins isolated from P. granatum and two naphthoquinones isolated from T. avellanedae demonstrated antibacterial activity against all S. aureus strains tested. Semi-synthetic furanonaphthoquinones (FNQs) showed lower MICs than those exhibited by natural occurring naphthoquinones. The results indicate that these natural products can be effective potential candidates for the development of new strategies to treat MRSA infections.

Antimicrobial Ellagitannin of Punica granatum Fruits

The ethyl acetate extract of Punica granatum fruits was fractionated by chromatographic techniques to afford the ellagitannin punicalagin. The substance was found to be active against methicillin-resistant Staphylococcus aureus strains and was identified by HPLC/UV and 1HNMR. The antibacterial assays which guided the isolation of the tannin were conducted using the disc diffusion method. Minimum inhibitory concentration (MIC) was determined by the dilution method according to NCCLS (National Committee for Clinical Laboratory Standards) procedure.

Bacteria in the apical root canal of teeth with primary apical periodontitis

OBJECTIVE Bacteria settled in the apical root canal are in a privileged position to inflict damage to the periradicular tissues. Therefore, the species identified in this region can be of special relevance for the pathogenesis of apical periodontitis. This study investigated the occurrence and levels of several bacterial taxa in the apical root canal of teeth with apical periodontitis. STUDY DESIGN DNA extracts from samples taken from the apical part of the root canal of extracted teeth evincing chronic apical periodontitis lesions served as templates for analysis of the presence and levels of 28 bacterial species/phylotypes using a 16S ribosomal RNA gene-based reverse-capture checkerboard hybridization assay. RESULTS Bacterial DNA was detected in 19 out of 20 samples. Detected taxa included Pseudoramibacter alactolyticus (32%), Bacteroidetes clone X083 (26%), Streptococcus species (21%), Olsenella uli (10.5%), Synergistes clone BA121 (10.5%), Fusobacterium nucleatum (10.5%), Porphyromonas endodontalis (10.5%), Dialister clone BS016 (5%), Filifactor alocis (5%), Parvimonas micra (5%), and Treponema denticola (5%). Of these, only Bacteroidetes clone X083 and Synergistes clone BA121 were found at levels above 10(5). CONCLUSION Occurrence of these bacterial taxa in the apical part of infected root canals indicates their potential pathogenetic role in the etiology of apical periodontitis.

Prevalence and Clonal Analysis of Porphyromonas gingivalis in Primary Endodontic Infections

This study investigated the prevalence of Porphyromonas gingivalis in 62 teeth with primary endodontic infections by using a species-specific 16S rRNA gene-based nested polymerase chain reaction assay. P. gingivalis isolates recovered from 2 infected root canals were also analyzed for clonal diversity by using arbitrarily primed PCR. Overall, P. gingivalis was found in 48% of the samples. This species was specifically detected in 36% of canals of teeth with chronic apical periodontitis, in 46% of the cases of acute apical periodontitis, and in 67% of acute apical abscesses. P. gingivalis was significantly more frequent in abscess aspirates than in canals of teeth with chronic apical periodontitis (P < .05). Typing of colonies retrieved from 2 infected canals revealed 2 clones per individual. These findings confirmed that P. gingivalis can be an important endodontic pathogen, mostly associated with acute abscesses, and demonstrated that different clonal types of this species can colonize the root canal in the same individual.

Non-tuberculous mycobacteria I: one year clinical isolates identification in Tertiary Hospital Aids Reference Center, Rio de Janeiro, Brazil, in pre highly active antiretroviral therapy era

The aim of this study was to determine the prevalence of non-tuberculous mycobacteria (NTM) isolates at University Hospital, Reference Center for Aids in Rio de Janeiro, Brazil, during one year. We used standard biochemical tests for species identification and IS1245 PCR amplification was applied as a Mycobacterium avium specific identification marker. Four hundred and four specimens from 233 patients yielded acid-fast bacilli growth. M. tuberculosis was identified in 85% of the patients and NTM in 15%. NTM disseminated infection was a common event correlated with human immunodeficiency virus (HIV) infected patients and only in HIV negative patients the source of NTM was non sterile site. M. avium complex (MAC) was biochemically identified in 57.8% (49/83) of NTM isolates, most of them from sterile sites (75.5%), and in 94% (46/49) the IS 1245 marker specific for M. avium was present. Twenty NTM strains showed a MAC biochemical pattern with the exception of a urease-positive (99% of MAC are urease-negative), however IS1245 was detected in 96% of the strains leading to their identification as M. avium. In this group differences in NTM source was not significant. The second most frequently isolated NTM was identified as M. scrofulaceum (7.2%), followed by M. terrae (3.6%), M. gordonae (2.4%), M. chelonae (1.2%), M. fortuitum (1.2%) and one strain which could not be identified. All were IS1245 negative except for one strain identified as M. scrofulaceum. It is interesting to note that non-sterile sites were the major source of these isolates (92.8%). Our finding indicated that M. avium is still the major atypical species among in the MAC isolates recovered from Brazilian Aids patients without highty active antiretroviral therapy schema. Some discrepancies were seen between the identification methods and further investigations must be done to better characterize NTM isolates using other phenotypic and genotypic methods.

RFLP patterns and risk factors for recent tuberculosis transmission among hospitalized tuberculosis patients in Rio de Janeiro, Brazil

Isolates of Mycobacterium tuberculosis from 120 tuberculosis patients seen in the 12 months ending September 1994 at 2 tertiary-care centres in Rio de Janeiro were characterized by IS6110 restriction fragment length polymorphism (RFLP) analysis. Ninety-seven patients (81%) had isolates with unique RFLP patterns, while 23 patients (19%) had isolates that belonged to 11 different RFLP cluster patterns. The strains from the latter patients were distributed among 1 group of 3 patients and 10 groups of 2 patients each. The cluster-pattern strains were not associated with gender, age, HIV infection, type of residence, living in shelter, homelessness or previous history of tuberculosis. However, clustering was strongly associated with multidrug resistance (P = 0.006). These data suggest that recent exogenous transmission may be important for the development of new cases of multidrug-resistant disease in patients attending tertiary-care centres in Rio de Janeiro, Brazil.

Statins Increase Rifampin Mycobactericidal Effect

ABSTRACT Mycobacterium leprae and Mycobacterium tuberculosis antimicrobial resistance has been followed with great concern during the last years, while the need for new drugs able to control leprosy and tuberculosis, mainly due to extensively drug-resistant tuberculosis (XDR-TB), is pressing. Our group recently showed that M. leprae is able to induce lipid body biogenesis and cholesterol accumulation in macrophages and Schwann cells, facilitating its viability and replication. Considering these previous results, we investigated the efficacies of two statins on the intracellular viability of mycobacteria within the macrophage, as well as the effect of atorvastatin on M. leprae infections in BALB/c mice. We observed that intracellular mycobacteria viability decreased markedly after incubation with both statins, but atorvastatin showed the best inhibitory effect when combined with rifampin. Using Shepards model, we observed with atorvastatin an efficacy in controlling M. leprae and inflammatory infiltrate in the BALB/c footpad, in a serum cholesterol level-dependent way. We conclude that statins contribute to macrophage-bactericidal activity against Mycobacterium bovis, M. leprae, and M. tuberculosis. It is likely that the association of statins with the actual multidrug therapy effectively reduces mycobacterial viability and tissue lesion in leprosy and tuberculosis patients, although epidemiological studies are still needed for confirmation.

Relationship Between Fcγ Receptor and Interleukin-1 Gene Polymorphisms and Post-treatment Apical Periodontitis

INTRODUCTION Genetic polymorphisms have been reported to act as modifiers of diverse diseases and, as such, might theoretically influence the severity and response to treatment of apical periodontitis. The purpose of this study was to investigate the association of Fcgamma receptor and interleukin (IL)-1 gene polymorphisms with post-treatment apical periodontitis in Brazilian individuals. METHODS The study population consisted of 18 patients with post-treatment apical periodontitis and 44 individuals with root canal-treated teeth exhibiting healthy/healing periradicular tissues (controls). Patients were typed for the following genes (alleles): FcgammaRIIA (R131 or H131), FcgammaRIIIB (NA1 or NA2), IL-1A (1 or 2), and IL-1B (1 or 2). RESULTS No significant statistical differences were observed for all specific genotypes and almost all allele carriage rates of the test genes as well as combinations thereof with regard to association with disease (P > .05). Actually, only 2 genetic conditions were found to be associated with post-treatment apical periodontitis: carriage of allele H131 of the FcgammaRIIa gene (P = .04) and a combination of this allele with allele NA2 of the FcgammaRIIIb gene (P < .01). CONCLUSIONS Data from the present study suggest that some conditions associated with polymorphism of Fcgamma receptor genes might influence the patients response to endodontic treatment of teeth with apical periodontitis.

Multidrug-Resistant Nontuberculous Mycobacteria Isolated from Cystic Fibrosis Patients

ABSTRACT Worldwide, nontuberculous mycobacteria (NTM) have become emergent pathogens of pulmonary infections in cystic fibrosis (CF) patients, with an estimated prevalence ranging from 5 to 20%. This work investigated the presence of NTM in sputum samples of 129 CF patients (2 to 18 years old) submitted to longitudinal clinical supervision at a regional reference center in Rio de Janeiro, Brazil. From June 2009 to March 2012, 36 NTM isolates recovered from 10 (7.75%) out of 129 children were obtained. Molecular identification of NTM was performed by using PCR restriction analysis targeting the hsp65 gene (PRA-hsp65) and sequencing of the rpoB gene, and susceptibility tests were performed that followed Clinical and Laboratory Standards Institute recommendations. For evaluating the genotypic diversity, pulsed-field gel electrophoresis (PFGE) and/or enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR) was performed. The species identified were Mycobacterium abscessus subsp. bolletii (n = 24), M. abscessus subsp. abscessus (n = 6), Mycobacterium fortuitum (n = 3), Mycobacterium marseillense (n = 2), and Mycobacterium timonense (n = 1). Most of the isolates presented resistance to five or more of the antimicrobials tested. Typing profiles were mainly patient specific. The PFGE profiles indicated the presence of two clonal groups for M. abscessus subsp. abscessus and five clonal groups for M. abscesssus subsp. bolletii, with just one clone detected in two patients. Given the observed multidrug resistance patterns and the possibility of transmission between patients, we suggest the implementation of continuous and routine investigation of NTM infection or colonization in CF patients, including countries with a high burden of tuberculosis disease.

Mycobacterium massiliense BRA100 strain recovered from postsurgical infections: resistance to high concentrations of glutaraldehyde and alternative solutions for high level disinfection

PURPOSE To evaluate the minimum inhibitory concentration (MIC) of GTA against these microorganisms and alternative disinfectants for high-level disinfection (HLD). METHODS Reference mycobacteria and clinical M. massiliense strains were included in this study. Active cultures were submitted to susceptibility qualitative tests with GTA dilutions (ranging from 1.5% to 8%), and commercial orthophthaldehyde (OPA) and peracetic acid (PA)-based solutions, during the period of exposure as recommended by National Agency of Sanitary Surveillance for HLD. RESULTS All reference and M. massiliense non-BRA100 strains, recovered from sputum, were susceptible to any GTA concentration, OPA and PA solutions. M. massiliense BRA100 strains presented MIC of 8% GTA and were susceptible to OPA and PA. CONCLUSION M. massiliense BRA100 strain is resistant to high GTA concentrations (up to 7%), which proves that this product is non-effective against specific rapidly growing mycobacteria and should be substituted by OPA or PA-based solutions for HLD.