Synergism of Phyllanthus niruni Extract with Gentamicin against Methicillin Resistant Staphylococcus aureus

Aims: This study aimed to find out any synergism of gentamicin with the solvent extracts of small tropical herb Phyllanthus niruri to combat methicillin resistant Staphylococcus aureus. Methodology: Bioactive constituents of Phyllanthus niruri were extracted by macerating ground dry powder of the leaves in water, n-hexane, chloroform, methanol and ethanol for 48-72 h followed by filtration and evaporation of solvents. Microdilution method was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of these extracts. The synergistic effects between gentamicin and the extracts were evaluated by the checkerboard assay to calculate the fractional inhibitory concentration index (FICI). In all cases, ten hospital associated MRSA strains were used. Results: The MIC of aqueous and methanolic extracts of P. niruri against different MRSA strains varies from 3.125 mg/ml to 12.5 mg/ml. For the MRSA strain the combination of methanolic extract with gentamicin decreased the MIC of extract from 6.25 mg/ml to 0.2 mg/ml and the MIC of gentamicin from 2048 µg/ml to 256 µg/ml showing a strong synergistic effect with a fractional inhibitory concentration index of 0.157. Steroids, flavonoids, phenolic compounds and quinones identified in the extracts may play role in synergistic relation. Conclusion: The present investigation shows that bioactive constituents from Phyllanthus niruri have an excellent synergy with gentamicin against MRSA and can be further explored as an alternative anti-staphylococcal agent.

In vitro combination susceptibility testing for detecting clinical isolates of Acinetobacter baumannii by five kinds of antibiotics with sulbactam / 临床检验杂志

Objective To investigate the in vitro antibacterial activity of single cefoperazone,ceftazidime,imipenem,tigecycline and colistin and their combination with sulbactam against clinical isolates of Acinetobacter baumannii (A.baumannii).Methods Twentythree meropenem-resistant A.baumannii strains and 21 meropenem-sensitive strains from the Study for Monitoring Antimicrobial Resistance Trends (SMART) during 2011 and 2012 were collected,and their combination susceptibility tests were performed by the checkerboard dilution method.The fractional inhibitory concentration (FIC) index was calculated to evaluate the combination effect of antibiotics.Results No antagonism effect was observed for all of combinations.The combination of sulbactam with cefoperazone or tigecycline mainly produced synergistic effect on A.baumannii,and the strains with FIC index ≤0.5 accounted for 56.8％ and 50.0％,respectively.The combination of sulbactam with imipenem or colistin showed synergistic or partially synergistic effect on A.baumannii,and the strains with FIC index ＜ 1 accounted for 61.4％ and 52.3％,respectively.However,the combination of sulbactan with ceftazidime didn't show any interation,and the strains with FIC index ≥4 accounted for 63.6％.Conclusion The combination of sulbactam with cefoperazone has the best synergistic effect on A.baumannii,especially on carbapenem-sensitive A.baumannii.The combination of sulbactam with imipenem or tigecycline may enhance the antibacterial activity on carbapenem-resistant A.baumannii.The combination of sulbactam with imipenem or tigecycline may be helpful for the treatment of carbapenem-resistant A.baumannii infection.

Synergistic antifungal activity of the lipophilic fraction of Hypericum carinatum and fluconazole

ABSTRACT Hypericum species, Hypericaceae, are recognized as a source of therapeutical agents. Purified fractions and isolated compounds have been shown antimicrobial activity. As the indiscriminate use of antifungals and the increase of infections caused by emerging species are leading to the search of new alternative treatments, the aim of this study was to continue the study with Hypericum carinatum Griseb. lipophilic fraction, rich in phloroglucinol derivatives, investigating the effect of its association with fluconazole against emerging yeasts (Candida krusei, C. famata, C. parapsilosis and Cryptococcus neoformans). The synergistic activity between H. carinatum lipophilic fraction and fluconazole was assessed by two methodologies for multiple dose–response analysis: checkerboard and isobologram. Regarding synergistic experiments, the effect of the association was higher than the effect of fluconazole alone against Candida krusei and C. famata isolates (MIC fluconazole decreased about eight and four folds, respectively), suggesting that, somehow, H. carinatum lipophilic fraction compounds are facilitating the action of this drug. On the other hand, when tested against Cryptococcus neoformans and C. parapsilosis, fluconazole showed better results than the association. Thus, against Candida krusei and C. famata, the lipophilic fraction of H. carinatum was able to reduce the MIC values of fluconazole and could be considered as a potential alternative to be used against emerging yeast species.

Papel de cepas de Streptococcus mutans e Bifidobacterium spp. na etiologia ou proteção contra doenças bucais / The role of Streptococcus mutans strains and Bifidobacterium spp. in the etiology or protection against oral diseases

A boca é um ecossistema complexo composto por centenas de espécies microbianas que apresentam características genotípicas e fenotípicas distintas que permitem sua adaptação e sobrevivência às adversidades desse ambiente. Streptococcus mutans é a espécie bacteriana mais relacionada à etiologia da cárie dentária, devido principalmente ao seu potencial acidogênico, acidúrico e na aderência e formação de biofilme dental. Outras espécies bacterianas, como as bifidobactérias, foram detectadas em diversos sítios bucais incluindo lesões cariosas iniciais ou cavitadas e estão sendo relacionadas à etiologia da cárie precoce da infância (CPI). Contrariamente, efeito benéfico na prevenção da doença periodontal tem sido observado para as bifidobactérias pelo fato dessas espécies competirem e interferirem na aderência e formação do biofilme de patógenos periodontais. Assim, ambiguidade em relação ao papel das bifidobactérias na etiologia ou prevenção de doenças bucais está sendo observada na literatura. Este trabalho foi dividido em três capítulos. Os objetivos foram: 1) avaliar a diversidade genotípica e as características fenotípicas de cepas de S. mutans, isoladas do biofilme dental de crianças CPI e CPI-severa (CPI-S) em comparação com crianças livres de cárie (LC); 2) avaliar a capacidade de produzir e tolerar ácidos, de formar biofilme e de induzir lesões iniciais de cárie in vitro por espécies de bifidobactérias comparadas à de espécies bacterianas já reconhecidas no contexto da cárie dentária e 3) investigar o efeito antagonista in vitro de algumas espécies de bifidobactérias sobre biofilmes de Fusobacterium nucleatum, Porphyromonas gingivalis e Streptococcus oralis. No capítulo 1, cepas de S. mutans de amostras de biofilme de crianças com CPI, CPI-S e LC foram isoladas em meio Agar Mitis Salivarius com bacitracina e avaliadas geneticamente pelo método de AP-PCR (Reação em Cadeia da Polimerase usando Iniciadores Arbitrários) e fenotipicamente pelos métodos de...

The mouth is a complex ecosystem composed of hundreds of microbial species with different genotypic and phenotypic characteristics that allow their adaptation and survival in the adversity of that environment. Streptococcus mutans is the bacterial species more closely related to the etiology of dental caries, mainly due to its acidogenic and aciduric potential and adherence and formation of dental biofilm. Other bacterial species, such as bifidobacteria, have been detected in oral sites including initial and cavitated carious lesions and have been related to the etiology of early childhood caries (ECC). In contrast, beneficial effect of bifidobacteria in the prevention of periodontal disease has been observed because they compete and interfere with the adherence and biofilm formation by periodontal pathogens. Thus, ambiguity regarding the role of bifidobacteria in the etiology or prevention of oral diseases has been observed in the literature. This work was divided into three chapters. The objectives were: 1) to evaluate the genotypic diversity and phenotypic characteristics of S. mutans strains isolated from dental biofilm of ECC and severe-ECC (S-ECC) children compared to caries-free children (CF); 2) to evaluate the ability to produce and tolerate acids, to form biofilm and inducing in vitro initial carious lesions by species of bifidobacteria compared to the bacterial species already recognized in the context of dental caries and 3) to investigate the in vitro antagonistic effect of some bifidobacteria species on biofilms of Fusobacterium nucleatum, Porphyromonas gingivalis and Streptococcus oralis. In Chapter 1, S. mutans strains from biofilm samples of children with ECC, S-ECC and CF were isolated in Agar Mitis Salivarius with bacitracin and genetically evaluated by AP-PCR method (Arbitrary Primers - Polymerase Chain Reaction) and phenotypically by acidogenicity methods, verifying the final pH of the cultures after exposure to high glucose concentration...

Comparison of two methods testing synergistic action of Reduning and cefoperazone sodium/sulbactam sodium on extensive drug resistant Acinetobacter bauman / 国际检验医学杂志

Objective To observe the results of broth dilution method and disc diffusion method to test the synergistic effect of Reduning and cefoperazone sodium / sulbactam sodium(SCF) on extensive drug resistant Acinetobacter bauman (XDR-AB) in vitro environment ,and compare their compliance to guide the clinical medication .Methods A total of 12 strains of XDR-AB from infec-tion patients in our hospital in 2015 were collected ,the strain was sub cultured .Firstly ,observe the minimum inhibitory concentra-tion (MIC) of SCF and Reduning on XDR-AB alone and in combination by broth dilution method .And then judge the synergy effects through calculation .Secondly ,the inhibition ring diameter and the synergy effects was detected using the disc diffusion meth-od .Results The MIC of Reduning and SCF in combination on XDR-AB was declined compared with them alone .The Fractional in-hibitory concentration of Reduning and SCF in combination on XDR-AB were equal or less than 0 .5 ,they had synergistic effect on XDR-AB .The inhibition ring diameter of Reduning was 10 mm tested by disk diffusion method .Different strains of XDR-AB on SCF bacteriostatic annulus diameter difference ,5 strains were 15 mm ,3 strains were 16 mm ,and 4 strains were 17 mm .Reduning and SCF appeared synergistic effect according to the inhibition ring diameter expanded when they effected on XDR -AB in combina-tion .Conclusion In vitro ,Reduning combined with SCF on XDR-AB has good synergistic effect .Compared with broth microdilution checkerboard dilution method ,disk diffusion method is more simple and convenient ,but it has a certain subjective on judging re-sults ,which is better to operate by experienced person .

Antibacterial activity of three newly-synthesized chalcones & synergism with antibiotics against clinical isolates of methicillin-resistant Staphylococcus aureus.

Background & objectives: Multidrug-resistance of methicillin-resistant Staphylococcus aureus (MRSA) is a serious therapeutical problem. Chalcones belong to a group of naturally occurring flavonoids, usually found in various plant species, and have potent antibacterial, antiviral and antifungal activities. The goal of this study was to evaluate the antibacterial effect of three newly-synthesized chalcones against clinical isolates of MRSA, and their synergism with β-lactam and non- β-lactam antibiotics. Methods: Antimicrobial activity of the three newly-synthesized chalcones was tested against 19 clinical isolates of MRSA and a laboratory control strain of MRSA (ATCC 43300). The synergism with β-lactams: cefotaxime (CFX), ceftriaxone (CTX), and non-β-lactam antibiotics: ciprofloxacin (CIP), gentamicin (GEN) and trimethoprim/sulphamethoxazole (TMP-SMX) was investigated by checkerboard method. Results: All evaluated compounds showed significant anti-MRSA activity with MIC values from 25-200 μg/ml. Observed synergism with antibiotics demonstrated that chalcones significantly enhanced the efficacy of CIP, GEN and TMP-SMX. Interpretation & conclusions: oOur study demonstrated that three newly-synthesized chalcones exhibited significant anti-MRSA effect and synergism with non-β-lactam antibiotics. The most effective compound was 1,3-Bis-(2-hydroxy-phenyl)-propenone. Our results provide useful information for future research of possible application of chalcones in combination with conventional anti-MRSA therapy as promising new antimicrobial agents.

Identificação de patógenos da cavidade bucal no lavado broncoalveolar de pacientes sob ventilação mecânica / Identification of pathogens of oral cavity in bronchoalveolar lavage of patients under invasive mechanical ventilation

Pacientes críticos internados em Unidade de Terapia Intensiva (UTI), em sua maioria, requerem auxílio da Ventilacão Mecânica Invasiva (VMI). A análise da diversidade microbiana é fundamental para identificação de espécies, ainda desconhecidas, responsáveis pela Pneumonia Associada a Ventilação Mecânica (PAVM). Os objetivos deste estudo foram: a) Estudar a diversidade microbiana presente na cavidade oral e no BAL (mini-BAL) de pacientes internados em UTI sob VMI, através da técnica Checkerboard DNA-DNA Hybridization; b) Avaliar se o perfil microbiano presente na cavidade oral está presente no Lavado Bronco Alveolar (mini-BAL), em diferentes períodos de tempo da VMI (12 h, 48 h, 96 h) e na extubação; c) Detectar a presença de espécies bacterianas circulantes na corrente sanguínea de pacientes internados em UTI sob VMI através dos resultados de hemocultura. Participaram do estudo 10 pacientes críticos internados em UTI sob VMI. As amostras foram coletadas no dorso da língua, espaço subgengival, no BAL, no aspirado endotraqueal e cânula de intubação orotraqueal. Foi possível identificar a presença de espécies bacterianas no BAL/aspirado endotraqueal, inclusive com aumento da carga bacteriana com o tempo prolongado de Intubação orotraqueal (IOT), 48 h e 96 h; e em relação à mucosa oral e espaço subgengival também foi possível observar já nas primeiras 12 h a presença de espécies bacterianas no BAL, estes achados apresentaram associação com PAVM e com resultados de hemocultura positiva. As bactérias isoladas na hemocultura foram S auerus, S epidermides, P aeruginosa. Houve associação de febre com PAVM e hemocultura positiva. Concluiu-se que durante a IOT, espécies bacterianas migram rapidamente para as vias aéreas inferiores, podendo contribuir para a fisiopatogenia da PAVM, houve associação de PAVM com Enterococcus faecalis, Fusobacterium periodonticum, Gemella morbillorum, Neisseria mucosa, Propionibacterium, Prevotella melaninogenica...

Critical patients admitted to intensive care unit (ICU), in their majority, require Invasive Mechanical Ventilation assistance (MV). The analysis of microbial diversity is fundamental to identifying species, as yet unknown, responsible for Pneumonia associated with mechanical ventilation (VAP). The objectives of this study were: to Study microbial diversity) present in the oral cavity and in the BAL (mini-BAL) of patients hospitalized in ICU under VM by Checkerboard DNA-DNA Hybridization technique; b) assess whether the microbial profile present in the oral cavity is present in Broncho Alveolar (mini-BAL), in different time periods of the MV (12 h, 48 ‘h, 96 h) and on extubation; c) detect the presence of bacterial species circulating in the bloodstream of patients hospitalized in ICU under VM through the results of blood cultures. Participated in this study 10 critical patients admitted to ICU under MV. The samples were collected on the dorsum of the tongue, subgingival space, in BAL, endotracheal aspirate and orotracheal intubation cannula. It was possible to identify the presence of bacterial species in the BAL/endotracheal aspirate, including with increased bacterial load with the extended time of orotraqueall Intubation (OTI), 48 h and 96 h; and in relation to the oral mucosa and subgingival space has also been possible to observe in the first 12 h the presence of bacterial species in the BAL, these findings showed association with VAP and with positive blood culture results. The isolated bacteria in blood culture were S auerus, S epidermides, P aeruginosa. There was association with fever and blood culture positive PAVM. It was concluded that during the OTI, bacterial species migrate quickly to the lower airways and may contribute to the pathophysiology of VAP. There were associations between VAP and Enterococcus faecalis, Fusobacterium periodonticum, Gemella morbillorum, Neisseria mucosa, Propionibacterium, Prevotella melaninogenica...

Remodelação alveolar e perfil bacteriano em implantes instalados em diferentes níveis relacionados à crista óssea / Bone remodeling and bacterial profile in dental implants inserted in different levels in relation to the crestal bone

A substituição clínica de dentes naturais perdidos por implantes osteointegrados tem representado uma das primeiras opções terapêuticas para a reabilitação de pacientes total ou parcialmente edêntulos. Apesar dos excelentes índices de sucesso demonstrados pelas restaurações implanto-suportadas, alguns fatores permanecem não esclarecidos, principalmente no que diz respeito à remodelação óssea ao redor dos implantes osteointegrados. Desta forma, o objetivo deste estudo foi avaliar a relação do nível de instalação dos implantes dentários com os parâmetros clínicos, com a remodelação óssea peri-implantar e com a colonização bacteriana, em implantes de plataforma regular, submetidos à carga imediata. Implantes de plataforma regular foram instalados em dois diferentes níveis em relação à crista óssea – ao nível ósseo e supra ósseo (1 mm). No total, trinta e cinco implantes em 9 pacientes (idade média de 62,4 ± 11,2 anos) foram avaliados radiograficamente no momento da instalação dos implantes (T1) e 6 meses após (T2), momento no qual também foram feitas análises clínicas e coleta de amostras para o teste microbiológico. Nos exames radiográficos foram analisadas a perda óssea, a partir de mensurações lineares da distância entre um ponto fixo do componente protético e o ponto mais coronário do contato osso-implante, e a densidade óptica alveolar obtida a partir de regiões ósseas de interesse (ROIs). As análises clínicas consistiram na avaliação da profundidade de sondagem e na mensuração do volume do fluido gengival peri-implantar. O perfil bacteriano dos sítios avaliados foi caracterizado por meio do método de análise de checkerboard DNA-DNA hybridization. Os testes estatísticos realizados mostraram não haver relação entre o nível de instalação dos implantes em relação à crista óssea e a remodelação óssea alveolar, tanto com relação ...

The replacement of missing teeth with osteointegrated dental implants represents one of the first therapeutic options for the rehabilitation of totally or partially edentulous patients. In spite of clinical success rates for implant-supported restorations, some factors remain nuclear, especially those related to alveolar bone remodeling around osteointegrated implants. Thus, the aim of this study was to evaluate the relation between the installation level of dental implants with bone remodeling and bacterial colonization, in regular platform implants, under an immediate loading protocol. Dental implants with regular platform were inserted in different levels related to the bone crest – on the level of bone crest and above the bone crest. In total, thirty-five implants in 9 patients (mean age 62,4 ± 11,2 years) were radiographically evaluated in the moment of implant installation (T1) and 6 months after (T2), when clinical analysis and sample collecting for microbiologic test were performed. In radiographic exams were analyzed the marginal bone loss, from linear measurements of the distance between a fixed point of prosthetic components and the most coronal point of bone-implant contact, and optical alveolar density, from bone regions of interest (ROIs). Clinical analysis consisted of probing depth evaluation and peri-implant crevicular fluid volume mensuration. The bacterial profile of selected sites was characterized by checkerboard DNA-DNA hybridization method. Statistic tests showed no relation between implant insertion levels and alveolar bone remodeling, with relation to marginal bone loss (p = 0,725) and optical alveolar density (p = 0,975). It was not also possible to establish a correlation among alveolar ...

Identificação de patógenos da cavidade bucal no lavado broncoalveolar de pacientes sob ventilação mecânica / Identification of pathogens of oral cavity in bronchoalveolar lavage of patients under invasive mechanical ventilation

Pacientes críticos internados em Unidade de Terapia Intensiva (UTI), em sua maioria, requerem auxílio da Ventilacão Mecânica Invasiva (VMI). A análise da diversidade microbiana é fundamental para identificação de espécies, ainda desconhecidas, responsáveis pela Pneumonia Associada a Ventilação Mecânica (PAVM). Os objetivos deste estudo foram: a) Estudar a diversidade microbiana presente na cavidade oral e no BAL (mini-BAL) de pacientes internados em UTI sob VMI, através da técnica Checkerboard DNA-DNA Hybridization; b) Avaliar se o perfil microbiano presente na cavidade oral está presente no Lavado Bronco Alveolar (mini-BAL), em diferentes períodos de tempo da VMI (12 h, 48 h, 96 h) e na extubação; c) Detectar a presença de espécies bacterianas circulantes na corrente sanguínea de pacientes internados em UTI sob VMI através dos resultados de hemocultura. Participaram do estudo 10 pacientes críticos internados em UTI sob VMI. As amostras foram coletadas no dorso da língua, espaço subgengival, no BAL, no aspirado endotraqueal e cânula de intubação orotraqueal. Foi possível identificar a presença de espécies bacterianas no BAL/aspirado endotraqueal, inclusive com aumento da carga bacteriana com o tempo prolongado de Intubação orotraqueal (IOT), 48 h e 96 h; e em relação à mucosa oral e espaço subgengival também foi possível observar já nas primeiras 12 h a presença de espécies bacterianas no BAL, estes achados apresentaram associação com PAVM e com resultados de hemocultura positiva. As bactérias isoladas na hemocultura foram S auerus, S epidermides, P aeruginosa. Houve associação de febre com PAVM e hemocultura positiva. Concluiu-se que durante a IOT, espécies bacterianas migram rapidamente para as vias aéreas inferiores, podendo contribuir para a fisiopatogenia da PAVM, houve associação de PAVM com Enterococcus faecalis, Fusobacterium periodonticum, Gemella morbillorum, Neisseria mucosa, Propionibacterium, Prevotella melaninogenica, ...

Critical patients admitted to intensive care unit (ICU), in their majority, require Invasive Mechanical Ventilation assistance (MV). The analysis of microbial diversity is fundamental to identifying species, as yet unknown, responsible for Pneumonia associated with mechanical ventilation (VAP). The objectives of this study were: to Study microbial diversity) present in the oral cavity and in the BAL (mini-BAL) of patients hospitalized in ICU under VM by Checkerboard DNA-DNA Hybridization technique; b) assess whether the microbial profile present in the oral cavity is present in Broncho Alveolar (mini-BAL), in different time periods of the MV (12 h, 48 ‘h, 96 h) and on extubation; c) detect the presence of bacterial species circulating in the bloodstream of patients hospitalized in ICU under VM through the results of blood cultures. Participated in this study 10 critical patients admitted to ICU under MV. The samples were collected on the dorsum of the tongue, subgingival space, in BAL, endotracheal aspirate and orotracheal intubation cannula. It was possible to identify the presence of bacterial species in the BAL/endotracheal aspirate, including with increased bacterial load with the extended time of orotraqueall Intubation (OTI), 48 h and 96 h; and in relation to the oral mucosa and subgingival space has also been possible to observe in the first 12 h the presence of bacterial species in the BAL, these findings showed association with VAP and with positive blood culture results. The isolated bacteria in blood culture were S auerus, S epidermides, P aeruginosa. There was association with fever and blood culture positive PAVM. It was concluded that during the OTI, bacterial species migrate quickly to the lower airways and may contribute to the pathophysiology of VAP. There were associations between VAP and Enterococcus faecalis, Fusobacterium periodonticum, Gemella morbillorum, Neisseria mucosa, Propionibacterium, Prevotella melaninogenica, ...

Análise da diversidade microbiana em infecções endodônticas persistentes / Microbial diversity analysis in persistent root canal infections

O presente trabalho teve por objetivo investigar a microbiota de canais radiculares relacionadas ao insucesso do tratamento endodôntico, buscando a identificação e a quantificação destes micro-organismos. Foram selecionados 36 dentes com infecção endodôntica persistente. O material obturador foi removido do canal radicular e amostras microbiológicas foram coletadas dos canais com o auxílio de limas tipo Hedstrõen e cones de papel absorvente estéril. A técnica do Checkerboard DNA-DNA hybridization foi utilizada para detecção de até 79 espécies bacterianas em cada amostra, utilizando sondas de DNA específicas. Os dados microbiológicos foram expressos em percentagem média (prevalência), proporção e nível médio de cada espécie em cada amostra. Os testes t independente e de correlação de Pearson foram usados para correlacionar a contagem das bactérias testadas com os dados clínicos (p≤ 0,05). Foi encontrada uma média de 11 espécies por amostra. E. faecium (36%), S. epidermidis (36%), E. saburreum (28%), P. micra (28%), S. sanguis (28%), C. sputigena (28%), L. buccalis (28%), E. faecalis (28%) e S. warneri (28%) foram as espécies mais prevalentes, e as espécies encontradas em níveis médios mais altos foram E. faecium, D. pneumosintes, S. epidermidis, H. pylori e C. sputigena. T. socranskii (3%), F. periodonticum (3%), C. gingivalis (3%), S. ixodetis (3%) apresentaram prevalências mais baixas. E. faecium e S. epidermidis apresentaram os maiores valores de prevalência, níveis médios e proporção. Não houve correlação entre a microbiota detectada nas amostras com os sinais e sintomas clínicos apresentados pelos pacientes, porém nas lesões periapicais de maior área foi detectada contagem significativamente maior de bacilos e espécies Gram-negativas (p<0,05). Baseado nos resultados obtidos é possível concluir que a microbiota presente em dentes...

The present study investigated the composition of the root canal microbiota in endodontic failures, aiming to identify and quantify these microorganisms. Thirty six teeth with persistent endodontic infection were selected. The root-filling materials were removed and microbiological samples were taken from the root canals with a Hedstrõen-type file and sterile paper points. The Checkerboard DNA-DNA hybridization technique was used for the detection of 79 bacterial species in each sample, using specific DNA probes. Microbiological data were express in mean prevalence, proportions and levels of each species in each sample. t independent test and Pearson correlation test were use to correlate bacterial counts and clinical conditions (p≤ 0,05). There were found a mean of 11 different species per sample. E. faecium (36%), S. epidermidis (36%), E. saburreum (28%), P. micra (28%), S. sanguis (28%), C. sputigena (28%), L. buccalis (28%), E. faecalis (28%) and S. warneri (28%) were the most prevalent species, and the species found in highest mean levels were E. faecium, D. pneumosintes, S. epidermidis, H. pylori and C. sputigena. T. socranskii (3%), F. periodonticum (3%), C. gingivalis (3%) and S. ixodetis (3%) were found in low prevalence. E. faecium and S. epidermidis presented the highest values of prevalence, means levels and proportions. No correlation was found between the detected microbiota and clinical findings; however in periapical lesions with highest areas, higher levels of rods and Gram-negative species were detected (p<0.05). Based on these results it may be concluded that the microbiota in teeth with persistent apical periodontitis presents a mixed and complex profile, and periapical lesions with larger area might be high associated with higher counts of rods and Gram-negative species...

Análise da diversidade microbiana em infecções endodônticas persistentes / Microbial diversity analysis in persistent root canal infections

O presente trabalho teve por objetivo investigar a microbiota de canais radiculares relacionadas ao insucesso do tratamento endodôntico, buscando a identificação e a quantificação destes micro-organismos. Foram selecionados 36 dentes com infecção endodôntica persistente. O material obturador foi removido do canal radicular e amostras microbiológicas foram coletadas dos canais com o auxílio de limas tipo Hedströen e cones de papel absorvente estéril. A técnica do Checkerboard DNA-DNA hybridization foi utilizada para detecção de até 79 espécies bacterianas em cada amostra, utilizando sondas de DNA específicas. Os dados microbiológicos foram expressos em percentagem média (prevalência), proporção e nível médio de cada espécie em cada amostra. Os testes t independente e de correlação de Pearson foram usados para correlacionar a contagem das bactérias testadas com os dados clínicos (p≤ 0,05). Foi encontrada uma média de 11 espécies por amostra. E. faecium (36%), S. epidermidis (36%), E. saburreum (28%), P. micra (28%), S. sanguis (28%), C. sputigena (28%), L. buccalis (28%), E. faecalis (28%) e S. warneri (28%) foram as espécies mais prevalentes, e as espécies encontradas em níveis médios mais altos foram E. faecium, D. pneumosintes, S. epidermidis, H. pylori e C. sputigena. T. socranskii (3%), F. periodonticum (3%), C. gingivalis (3%), S. ixodetis (3%) apresentaram prevalências mais baixas. E. faecium e S. epidermidis apresentaram os maiores valores de prevalência, níveis médios e proporção. Não houve correlação entre a microbiota detectada nas amostras com os sinais e sintomas clínicos apresentados pelos pacientes, porém nas lesões periapicais de maior área foi detectada contagem significativamente maior de bacilos e espécies Gram-negativas (p<0,05). Baseado nos resultados obtidos é possível concluir que a microbiota presente em dentes com ...

The present study investigated the composition of the root canal microbiota in endodontic failures, aiming to identify and quantify these microorganisms. Thirty six teeth with persistent endodontic infection were selected. The root-filling materials were removed and microbiological samples were taken from the root canals with a Hedströen-type file and sterile paper points. The Checkerboard DNA-DNA hybridization technique was used for the detection of 79 bacterial species in each sample, using specific DNA probes. Microbiological data were express in mean prevalence, proportions and levels of each species in each sample. t independent test and Pearson correlation test were use to correlate bacterial counts and clinical conditions (p≤ 0,05). There were found a mean of 11 different species per sample. E. faecium (36%), S. epidermidis (36%), E. saburreum (28%), P. micra (28%), S. sanguis (28%), C. sputigena (28%), L. buccalis (28%), E. faecalis (28%) and S. warneri (28%) were the most prevalent species, and the species found in highest mean levels were E. faecium, D. pneumosintes, S. epidermidis, H. pylori and C. sputigena. T. socranskii (3%), F. periodonticum (3%), C. gingivalis (3%) and S. ixodetis (3%) were found in low prevalence. E. faecium and S. epidermidis presented the highest values of prevalence, means levels and proportions. No correlation was found between the detected microbiota and clinical findings; however in periapical lesions with highest areas, higher levels of rods and Gram-negative species were detected (p<0.05). Based on these results it may be concluded that the microbiota in teeth with persistent apical periodontitis presents a mixed and complex profile, and periapical lesions with larger area might be high associated with higher counts of rods and Gram-negative species.

Combined antimicrobial susceptibility test against pan-drug-resistant Acinetobacter baumannii with E-test and microdilution checkerboard assay / 中华微生物学和免疫学杂志

Objective To evaluate the activity of antibiotics against pan-drug-resistant (PDR) Acinetobacter baumannii by combination antimicrobial susceptibility test in viro with epsilometric methods (Etest method) and microdilution checkerboard (CB method),and to detect a good correlation between timekill curve with the above mentioned two assays.Methods Thirty-one clinical isolates of PDR Acinetobacter baumannii were selected for mono and combination antimicrobial susceptibility test in vitro by E-test and CB method,then a comparison was conducted between the test results and the time-kill curve.Mono drugs involved tigecycline,colistin,imipenem and amikacin,and combinations involved two of drugs above,and three drugs involved imipenem/tigecycline,plus amikacin combination.Results Synergistic effect was detected in imipenem plus colistin and tigecycline plus imipenem combination.A high comparability was revealed between the E-test method with antimicrobial drugs added into the culture medium and the time-kill curves.Synergy in the combination of imipenem/tigecycline,plus amikacin was detected by the CB method and time-kill curves.Conclusion The results showed that the effect of specific combination of antibiotics against PDR Acinetobacter baumannii could be predicted by testing their synergistic effect with combination antimicrobial susceptibility test.

Evaluation of the synergistic potential of vancomycin combined with other antimicrobial agents against methicillin-resistant Staphylococcus aureus and coagulase-negative Staphylococcus spp strains

Methicillin-resistant Staphylococcus aureus (MRSA) and coagulase-negative Staphylococcus spp (CNS) are the most common pathogens that cause serious long term infections in patients. Despite the existence of new antimicrobial agents, such as linezolid, vancomycin (VAN) remains the standard therapy for the treatment of infections caused by these multidrug-resistant strains. However, the use of VAN has been associated with a high frequency of therapeutic failures in some clinical scenarios, mainly with decreasing concentration of VAN. This work aims to evaluate the synergic potential of VAN plus sulfamethoxazole/trimethoprim (SXT), VAN plus rifampin (RIF) and VAN plus imipenem (IPM) in sub-minimum inhibitory concentrations against 22 clinical strains of MRSA and CNS. The checkerboard method showed synergism of VAN/RIF and VAN/SXT against two and three of the 22 strains, respectively. The combination of VAN with IPM showed synergistic effects against 21 out of 22 strains by the E-test method. Four strains were analyzed by the time-kill curve method and synergistic activity was observed with VAN/SXT, VAN/RIF and especially VAN/IPM in sub-inhibitory concentrations. It would be interesting to determine if synergy occurs in vivo. Evidence of in vivo synergy could lead to a reduction of the standard VAN dosage or treatment time.

In vitro efficacy of Meropenem and Sulbactam combination against Acinetobacter baumannii / 中华检验医学杂志

Objective To evaluate the synergy effect of Meropenem and Sulbactam combination against Meropenem-resistant and Meropenem-susceptible A.baumannii in vitro and optimize combination ratio of Meropenem and Sulbactam to achieve best synergy effect.Methods Evaluating the synergy effect of Meropenem and Sulbaetam combination through microdilution checkerboard method against Meropenemresistant and Meropenem-susceptible A.baumannii,isolated from inpatients of Chinese hospitals.Assessing the synergy effect of combination in different ratios of Meropenem to Sulbactam.Results The checkerboard method with the combination of Meropenem and Sulbactam demonstrated 25.0％ ( 10/40 ) synergism,67.5％ (28/40) partial synergism,7.5％ (3/40) additive,no indifference and antagonism in Meropenemsusceptible isolates,and 27.5％ (11/40) synergism,40.0％ (16/40) partial synergism,25.0％(10/40) additive,no indifference and antagonism in Meropenem-resistant isolates.Eleven Meropenemresistant isolates which showed synergism in synergy test were tested for MICs of combination of Meropenem and Sulbactam,using ratios of 4∶ 1,2∶ 1,1∶1 and 1∶2,and the MIC90 were 64∶ 16,64∶ 32,32∶32,32∶64 μg/ml,respectively.Conclusions Meropenem and Sulbactam combination show synergism or partial synergism against most A.baumannii isolates.The optimal ration of combination for clinical use may be 1∶ 1.

In vitro antifungal activity and cytotoxic effect of essential oils and extracts of medicinal and aromatic plants against Candida krusei and Aspergillus fumigatus / Atividade antifúngica in vitro e os efeitos citotóxicos de óleos essenciais e extratos de plantas medicinais e aromáticas contra Candida krusei e Aspergillus fumigatus

The plants are usually used in traditional medicine as antimicrobial agents and their essential oils and extracts have been known to possess antifungal activity. The aim of this study was to evaluate in vitro the activity of 32 essential oils and 29 extracts against C. krusei and A. fumigatus as well as the cytotoxic effect on Vero cells. Time-kill curve and interaction between antifungals and the most active sample against C. krusei, was also evaluated. The oils from C. ambrosioides and the extract of M. cucullata showed antifungal activity against C. krusei (GM-MIC 7.82 and 31.25 µg/mL, respectively). L. citriodora was actives against C. krusei and A. fumigates (GM-MIC = 99.21 µg/mL and 62.5 µg/mL respectively). Time-kill assays done with C. ambrosioides oil showed fungicidal activity at 4x MIC. The interaction of C. ambrosioides oil with itraconazole and amphotericin B was tested following the chequerboard technique. No interaction was detected for the combination of C. ambrosioides oil with amphotericin B and itraconazole (FICI range = 1.03-1.06 and 1.03-1.00, respectively). Cytotoxicity assays for all samples were carried out with MTT. Only the oil from Hedyosmun sp. and L. dulcis were cytotoxic.

As plantas são geralmente utilizadas na medicina tradicional como agentes antimicrobianos e seus óleos essenciais e extratos foram conhecidos por possuir atividade antifúngica. O objetivo deste estudo foi avaliar in vitro a atividade de 32 óleos essenciais e 29 extratos contra Candida krusei e Aspergillus fumigatus, bem como o efeito citotóxico em células Vero. A curva do tempo-morte e a interação entre antifúngicos e Chenopodium ambrosioidese do extrato de Myrcia cucullata mostraram atividade antifúngica contra C. krusei (geometric means of the minimal inhibitory concentration [GM-MIC] 7,82 e 31,25 µg/mL, respectivamente). Lippia citriodora foi ativa contra C. krusei e A. fumigatus (GM-CIM = 99,21 µg/mL e 62,5 µg/mL, respectivamente). Os testes de tempo-morte feitos com óleo de C. ambrosioides mostraram atividade fungicida em 4x MIC. A interação do óleo C. ambrosioides com itraconazol e anfotericina B foi testada pela técnica de xadrez. Nenhuma interação foi detectada pela combinação do óleo C. ambrosioides com anfotericina B e itraconazol (intervalo fractional inhibitory index [FICI] = 1,03-1,06 e 1,03-1,00, respectivamente). Os ensaios de citotoxicidade para todas as amostras foram realizadas com MTT. Apenas os óleos Hedyosmun sp. e L. dulcis foram citotóxicos.

In Vitro Effects of Combined Antibiotics against Multidrug-resistant Pseudomonas aeruginosa / 대한내과학회지

BACKGROUND/AIMS: The inhibitory effects of the combination of beta-lactam with ciprofloxacin or amikacin against clinical isolates of multidrug-resistant Pseudomonas aeruginosa were evaluated. METHODS: This study examined ten isolates with variable levels of resistance to ceftazidime, cefepime, piperacillin/tazobactam, meropenem, ciprofloxacin, and amikacin. The efficacy of the combined antibiotics was studied using a checkerboard method or in vitro killing assay. RESULTS: The combination of ceftazidime, cefepime, aztreonam, piperacillin-tazobactam, or meropenem with amikacin showed synergistic effects for all of the strains regardless of the minimum inhibitory concentration (MIC) of amikacin, but combination with ciprofloxacin showed a synergistic effect for the isolate with a low MIC of ciprofloxacin by the checkerboard method. The isolates with a high MIC of ciprofloxacin showed an indifferent effect in combination with beta-lactam and ciprofloxacin. The in vitro killing assay showed that meropenem with ciprofloxacin acted synergistically for the isolates with a MIC of 16 microgram/mL of ciprofloxacin. However, amikacin showed synergistic effects with meropenem for the isolates with high-level resistance against amikacin, i.e., up to an MIC of 128 microgram/mL. Contrary to the checkerboard method results, no synergy was observed for the combination of ceftazidime/piperacillin-tazobactam and amikacin. CONCLUSIONS: Meropenem with amikacin can be the first choice for infections caused by multidrug-resistant P. aeruginosa when the level of resistance is not known.

Uso de la técnica de hibridización checkerboard ADN-ADN para la identificación de la microbiota periimplantaria: revisión bibliográfica / Use of checkerboard DNA-DNA hybridization technique for periimplantitis microbiota identification: literature review

El término Periimplantitis hace referencia a la condición de enfermedad en los tejidos de soporte de los implantes bucales. Su etiología es multifactorial, aunque la biopelícula microbiana desempeña un papel esencial en la etiopatogenia de la enfermedad. A través de técnicas de cultivo, se han identificado algunas bacterias implicadas en la etiopatogenia de las periodontopatías, entre estas, especies pertenecientes a los Géneros Fusobacterium, Prevotella y Porphyromonas. El desarrollo de técnicas de biologia molecular permite la identificación de especies bacterianas que antes no eran referidas como parte de la microbiota responsable de la patogénesis periodontal. La detección de la microbiota presente en los sacos periodontales que se originan alrededor de los implantes es necesaria para el establecimiento de la conducta terapéutica a ser instruida, más aún si se toma den consideración que las alteraciones de las estructuras de soporte están directamente relacionadas con muchos de los microorganismos presentes y constituyen una de las causas más frecuentes de fracaso del tratamiento rehabilitador con implantes. El objetivo de este artículo es presentar una revisión bibliográfica de la técnica de hibridización checkerboard ADN-ADN para la identificación de los microorganismos más frecuentemente asociados a la peri-implantitis

The periimplantitis term is characterized as a disease that affects the support implant tissues. It has a multifactorial etiology, with an important role of the biofilm on the periodontal diseases. The culture techniques use possibilited to identify bacterias responsible for periodontal alterations, as Fusobacterium, Prevotella and Porphyromonas ssp. The development of the molecular biology techniques possibilited the identification of some bacterial species, that there were not related in the literature and there were responsible for periodontal pathogenesis. The microbiota present on the periodontal pockets in overdentures may contribute in the orientation of the therapeutic procedures, considering that the structural alterations on the periodontal tissues are straightly related to microorganism presence, and it constitutes one of the causes that affect the oral rehabilitation treatment with implants. This, the aim of this study was to present a literature review of DNA-DNA hybridization technique in the identification of the microorganisms more related to periimplantitis

DNA Checkerboard Method for Bacterial Pathogen Identification in Oral Diseases / Método de Chequeo de DNA para la Identificación de Bacterias Patógenas en las Enfermedades de la Cavidad Oral

This work aim to show by literature review the principal characteristics of the DNA checkerboard method for bacterial pathogens identification in oral diseases, showing the most varieties uses and applications of this technique.

Este trabajo tiene como objetivo, presentar en una revisión de la literatura, las principales características del método de chequeo del DNA para la identificación de bacterias patógenas en la cavidad oral, mostrando las diferentes utilizaciones y aplicaciones de esta técnica.

In vitro activity of ciprofloxacin or Amikacin combined with ?-lactams against Multidrug-Resistant Pseudomonas aeruginosa Strains / 中国实用内科杂志

Objective The aim of this study was to evaluate the in vitro activity of Ciprofloxacin(CIP)or Amikacin(AK)combined with four clinically-used antipseudomonas ?-lactams against multidrug-resistant Pseudomonas aeruginosa strains.Methods Determining the MICs of six antibiotics to 35 clinically isolated Pseudomonas aeruginosa strains which have different resistance patterns by 2-fold broth dilution method;determining the in vitro activity of Ciprofloxacin or Amikacin combined with Ceftazidime(CAZ),Imipenem(IMP),Meropenem(MEM)and Piperacillin/Tazobactam(TZP)by broth checkerboard method.Results The combinations of Ciprofloxacin or Amikacin with four ?-lactams have mainly synergistic or additive effect,the synergistic rate of AK-group(AK+CAZ:71.43%;AK+IMP:42.86%;AK+MEM:54.29%;AK+TZP:85.71%)was higher than that of CIP-group(CIP+CAZ:20%;CIP+IMP:25.71%;CIP+MEM:14.29%;CIP+TZP:31.43%).The combination of CIP+TZP has the highest synergistic rate in CIP-group;the combination of AK+TZP has the highest synergistic rate in AK-group.None of the combinations had an antagonistic effect.Conclusion the combination of ciprofloxacin or amikacin with ?-lactams have good in vitro activity to multidrug-resistant Pseudomonas aeruginosa strains.

Antibacterial Activity of Water Soluble Components of Elfvingia applanata Alone and in Combinations with Quinolones

A preparation of water soluble components(EA) was made from carpophores of Elfvingia applanata(Pers.) Karst and its in vitro antibacterial activity on a number of bacterial species was examined by macrobroth dilution assay. Among 16 species of bacteria tested, the most potent antibacterial activity was observed against Staphylococcus epiderrnidis and Proteus vulgaris, of which MICs were 1.25 mg/ml. To investigate the antibacterial effects in combinations of EA with quinolone antibiotics, such as ciprofloxacin, enoxacin, lomefloxacin, norfloxacin, and ofloxacin, the fractional inhibitory concentrations(FICs) and the fractional inhibitory concentration indices(FICIs) for four bacterial strains were determined by macrobroth dilution checkerboard assay. Combinations of EA and quinolones exhibited either additive or indifferent effects of antibacterial activity in most instances. However, both synergistic and antagonistic effects were not observed in any cases.