RESUMO
OBJECTIVES: The objective of this study was to evaluate the efficiency of SB-20 M culture medium to perform differential morphological identification of S. mutans and S. sobrinus compared to biochemical identification and to proteomic identification by the MALDI-TOF mass spectrometry technique. MATERIAL AND METHODS: Unstimulated saliva samples from 266 dental students were seeded on SB-20 M culture medium by the wooden spatula technique. After incubation, S. mutans and S. sobrinus colonies were identified by stereomicroscopy based on their differential morphological characteristics. Following these procedures, 135 colonies with characteristic morphology of S. mutans (89 colonies) and S. sobrinus (46 colonies) were randomly selected, submitted to biochemical identification (biotyping) and proteomic identification by the MALDI-TOF mass spectrometry technique. The results were compared using the Kappa test, with a 5% significance level. RESULTS: All (100%) S. mutans colonies were correctly identified after culture in SB-20 M medium compared to biotyping and proteomic identification. For S. sobrinus, morphological identification in SB-20 M medium was correct for 43 colonies (93.5%) compared to biotyping and proteomic identification. However, there was no statistically significant difference when comparing the capacity to identify S. mutans and S. sobrinus of the three techniques (p < 0.001; K = 0.951). CONCLUSIONS: It was concluded that the SB-20 M culture medium for morphological identification of S. mutans and S. sobrinus was highly reliable, being comparable to the MALDI-TOF mass spectrometry technique. CLINICAL RELEVANCE: The efficiency evaluation of identification methods of S. mutans and S. sobrinus is clinically relevant in order to determine caries risk and activity of patients.
Assuntos
Cárie Dentária , Proteômica , Streptococcus mutans , Streptococcus sobrinus , Cárie Dentária/microbiologia , Humanos , Saliva , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Streptococcus mutans/isolamento & purificação , Streptococcus sobrinus/isolamento & purificaçãoRESUMO
In this paper we investigated the antibacterial activity of a methanolic extract of Rosmarinus officinalis L. and their main constituents, carnosic acid and rosmarinic acid, against 37 nosocomial strains of multidrug-resistant bacteria. Results obtained showed that both the rosemary extract and carnosic acid inhibited all clinical isolates of Staphylococcus aureus methicillin-resistant and Enterococcus faecalis gentamicin and streptomycin-resistant bacteria examined (MICs 60 ug/mL vs. 200 ug/mL, respectively). Rosemary extract showed MIC values between 400 and 1600 ug/ml against the Gram-negative multidrug-resistant bacteria: Escherichia coli, Proteus mirabilis, Enterobacter cloacae, Pseudomonas aeruginosa, Morganella morganii and Providencia stuartii, while carnosic acid showed MIC of 120 to 240 ug/mL. Bactericidal effect of carnosic acid against S. aureus and E. faecalis was observed at their MIC value, while 2 x MIC to 4 x MIC were needed to kill Gram-negative bacteria. Rosmarinic acid showed a narrow spectrum of action against a few Gram-negative clinical isolates. Our findings suggest that carnosic acid would be a good lead candidate useful in counteracting drug-resistant infections.
En este trabajo evaluamos la actividad antibacteriana de un extracto metanólico de Rosmarinus officinalis L. y sus principales componentes el ácido carnósico y ácido rosmarínico, contra 37 cepas de bacterias multirresistentes nosocomiales. Los resultados muestran que el extracto de romero y el ácido carnósico, inhibieron las bacterias Gram-positivas Staphylococcus aureus resistentes a meticilina y Enterococcus faecalis resistentes a gentamicina y estreptomicina (CIM 200 ug/mL y 60 ug/mL, respectivamente). El extracto de romero inhibió los Gram negativos multirresistentes: Escherichia coli, Proteus mirabilis, Enterobacter cloacae, Pseudomonas aeruginosa, Morganella morganii y Providencia stuartii (CIM 400 a 1600 ug/mL), mientras que el ácido carnósico mostró valores de CIM entre 120 a 240 ug/mL. El ácido carnósico mostró actividad bactericida contra S. aureus y E. faecalis a su CIM, mientras que 2 a 4 X CIM se requirieron para matar las bacterias Gram-negativas. El ácido rosmarínico mostró inhibió unos pocos aislados clínicos Gram-negativos. Estos hallazgos sugieren que el ácido carnósico puede ser de utilidad contra infecciones bacterianas multirresistentes a antibióticos.
Assuntos
Antibacterianos/farmacologia , Bactérias , Cinamatos/farmacologia , Abietanos/farmacologia , Extratos Vegetais/farmacologia , Rosmarinus/química , Bactérias/isolamento & purificação , Cinamatos/análise , Depsídeos/análise , Depsídeos/farmacologia , Abietanos/análise , Extratos Vegetais/química , Farmacorresistência Bacteriana Múltipla , Testes de Sensibilidade Microbiana , RosmarinusRESUMO
In the last years, Enterobacteriaceae such as Klebsiella pneumoniae, Proteus mirabilis and Escherichia coli, have acquired resistance to third-generation cephalosporins (C3G) because of the presence of plasmid-mediated AmpC ß-lactamases. The aim of this work was to detect plasmid AmpC enzymes and to investigate the predominant types in our region. Between March and July 2009, 733 consecutive isolates of Enterobacteriaceae derived from hospitals and outpatient centers were studied. Susceptibility testing was performed by disk diffusion; one P. mirabilis and three E. coli strains showed resistance to cephamycins (cefoxitin) and C3G. An E-test to determine MIC and a synergy test by aminophenylboronic disks were performed. Enzymatic activity against cefoxitin was confirmed by a microbiological assay. A polymerase chain reaction (PCR) for the detection of plasmid-mediated ampC genes of different groups was performed and a 462-bp amplicon was obtained when using primers directed against the CIT group; the obtained sequences were compared to blaCMY-2 sequences, showing 100% identity. The emergence of CMY-2-type plasmid-mediated AmpC ß-lactamases indicated the importance of implementing systematic monitoring of these resistances to avoid potential clinical and epidemiological consequences.
Assuntos
Proteínas de Bactérias/análise , Escherichia coli/enzimologia , Proteus mirabilis/enzimologia , Fatores R/genética , beta-Lactamases/análise , Sequência de Aminoácidos , Argentina , Proteínas de Bactérias/genética , Cefoxitina/farmacologia , Resistência às Cefalosporinas/genética , Cefalosporinas/farmacologia , DNA Bacteriano/genética , Enterobacteriaceae/enzimologia , Enterobacteriaceae/genética , Enterobacteriaceae/isolamento & purificação , Infecções por Enterobacteriaceae/microbiologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/microbiologia , Humanos , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , Infecções por Proteus/microbiologia , Proteus mirabilis/efeitos dos fármacos , Proteus mirabilis/genética , Proteus mirabilis/crescimento & desenvolvimento , Proteus mirabilis/isolamento & purificação , Homologia de Sequência de Aminoácidos , Infecções Urinárias/microbiologia , beta-Lactamases/química , beta-Lactamases/genéticaRESUMO
El propósito del presente trabajo fue determinar la potencia antimicrobiana de extractos alcohólicos de plantas utilizadas popularmente en Argentina como antisépticos y antiinflamatorios: Dasyphyllum diaconthoides, Erythrina cristagalli, Larrea cuneifolia, Larrea divaricata, Phytolacca dioica, Pithecoctenium cynanchoides, Prosopanche americana, Schinus molle, Schkuhria pinnata, Senna aphylla y Solidago chilensis. La inhibición del crecimiento bacteriano se determinó a través de ensayos de difusión en agar, macrodilución en medio sólido y microdilución en medio líquido frente a 47 aislamientos clínicos multirresistentes a antibióticos, obtenidos de pacientes de un hospital de Tucumán, Argentina: Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Enterobacter cloacae, Serratia marcescens, Morganella morganii, Acinetobacter baumannii, Pseudomonas aeruginosa, Stenotrophomonas maltophilia. De acuerdo con los valores de concentración inhibitoria mínima (CIM), tres de las once especies ensayadas fueron las más activas: L. divaricata, L. cuneifolia, y S. aphylla (CIM de 25 a 200 µg/mL). P. mirabilis, A. baumanni y S. maltophilia fueron las cepas más susceptibles con valores de CIM entre 25 y 50 µg/mL seguido por P. aeruginosa con valores de CIM de 50 a 100 µg/mL. Los valores de concentración bactericida mínima (CBM) fueron semejantes o dos veces superiores a los valores de CIM. Mediante ensayos bioautográficos se comprobó que los extractos más activos presentaban al menos dos principios antimicrobianos. Análisis fitoquímicos indican que estos compuestos son de naturaleza fenólica. Los resultados obtenidos justificarían el uso de estos extractos para el tratamiento de infecciones bacterianas, especialmente aquellas de origen dérmico.
The present study was conducted to investigate antimicrobial activity of alcoholic extracts of Argentine medicinal plant species (Dasyphyllum diacanthoides, Erythrina cristagalli, Larrea cuneifolia, Larrea divaricata, Phytolacca dioica, Pithecoctenium cynanchoides, Prosopanche americana, Schinus molle, Schkuhria pinnata, Senna aphylla and Solidago chilensis) against multidrug resistant human pathogen gram negative bacteria isolated from a Hospital in Tucumán, Argentina. Inhibition of bacterial growth was investigated using disc diffusion, agar macrodilution and broth microdilution methods against multiresistant clinical isolates of nine different specie of gram negative bacteria: Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Enterobacter cloacae, Serratia marcescens, Morganella morganii, Acinetobacter baumannii, Pseudomonas aeruginosa, Stenotrophomonas maltophilia. A significant antimicrobial activity was found in three of the eleven plant species studied. Based on minimal inhibitory concentration (MIC) values, three plant species, L. divaricata, L. cuneifolia, and S. aphylla were the most potent ones with MIC values between 25-200 µg/mL. Overall, P. mirabilis, M. morganii and P. aeruginosa isolates were the most susceptible to these extracts with MIC values of 25 to 50 µg/mL. All extracts showed significant inhibitory activities on bacteria growth in a dose phenolic compound-dependent fashion. The minimal bactericidal concentration (MBC) values were identical to the MIC values or twofold higher than the corresponding MIC. Contact bioautography indicated that crude extracts possess several major antibacterial components. Phytochemical screening showed that the bioactive compounds correspond to polyphenols. Investigations are in progress to purify the bioactive principles.
Assuntos
Plantas Medicinais , Plantas Medicinais/fisiologia , Plantas Medicinais/microbiologia , Argentina , Plantas Medicinais/citologia , Resistência Microbiana a Medicamentos , Schinus molle , Larrea , Erythrina , Anti-InfecciososRESUMO
The in vitro activity of piperacillin-tazobactam and several antibacterial drugs commonly used in Argentinean hospitals for the treatment of severe infections was determined against selected but consecutively isolated strains from clinical specimens recovered from hospitalized patients at 17 different hospitals from 9 Argentinean cities from different geographic areas during the period November 2001-March 2002. Out of 418 Enterobacteriaceae included in the Study 84% were susceptible to piperacillin-tazobactam. ESBLs putative producers were isolated at an extremely high rate since among those isolates obtained from patients with hospital acquired infections 56% of Klebsiella pneumoniae, 32% of Proteus mirabilis and 25% Escherichia coli were phenotypically considered as ESBLs producers Notably P.mirabilis is not considered by for screening for ESBL producers. ESBLs producers were 100% susceptible to imipenem and 70% were susceptible to piperacillin-tazobactam whereas more than 50% were resistant to levofloxacin. The isolates considered as amp C beta lactamase putative producers showed 99% susceptibility to carbapenems while 26.7% were resistant to piperacillin-tazobactam and 38.4% to levofloxacin. Noteworthy only 4% of the Enterobacteriaceae isolates were resistant to amikacin. Piperacillin-tazobactam was the most active agent against Pseudomonas aeruginosa isolates (MIC(90): 128 microg/ml; 78% susceptibility) but showed poor activity against Acinetobacter spp (MIC(90):>256 microg/ml; 21.7% susceptibility). Only 41.7% Acinetobacter spp isolates were susceptible to ampicillin-sulbactam. Piperacillin-tazobactam inhibited 100% of Haemophilus influenzae isolates (MIC(90) < 0.25 microg/ml) but only 16.6% of them were ampicillin resistant. The activity of piperacillin-tazobactam against oxacillin susceptible Staphylococcus aureus or coagulase negative staphylococci was excellent (MIC(90) 2 microg/ml; 100% susceptibility). Out of 150 enterococci 12 isolates (8%) were identified as E.faecium and only three isolates (2%), 2 E.faecium and 1 E.faecalis were vancomycin resistant. All the enterococci isolates were susceptible to linezolid. Piperacillin-tazobactam showed excellent activity (MIC(90) 2 microg/ml; 92% susceptibility). Regarding pneumococci all the isolates showed MICs of 16 microg/ml for piperacillin-tazobactam. Among 34 viridans group streptococci only 67% were penicillin susceptible and 85.2% ceftriaxone susceptible whereas piperacillin-tazobactam was very active (MIC(90) 4 microg/ml).Piperacillin-tazobactam is therefore a very interesting antibacterial drug to be used, preferably in combination (IE: amikacin-vancomycin) for the empiric treatment of severe infections occurring in hospitalized patients in Argentina. Caution must be taken for infections due to ESBL producers considering that the inoculum effect MICs can affect MIC values.