Chemical Composition and Antimicrobial Effectiveness of Ocimum gratissimum L. Essential Oil Against Multidrug-Resistant Isolates of Staphylococcus aureus and Escherichia coli.

The study investigated the antimicrobial activity of the essential oil extract of Ocimum gratissimum L. (EOOG) against multiresistant microorganisms in planktonic and biofilm form. Hydrodistillation was used to obtain the EOOG, and the analysis of chemical composition was done by gas chromatography coupled with mass spectrometry (GC/MS) and flame ionization detection (GC/FID). EOOG biological activity was verified against isolates of Staphylococcus aureus and Escherichia coli, using four strains for each species. The antibacterial action of EOOG was determined by disk diffusion, microdilution (MIC/MBC), growth curve under sub-MIC exposure, and the combinatorial activity with ciprofloxacin (CIP) and oxacillin (OXA) were determined by checkerboard assay. The EOOG antibiofilm action was performed against the established biofilm and analyzed by crystal violet, colony-forming unit count, and SEM analyses. EOOG yielded 1.66% w/w, with eugenol as the major component (74.83%). The MIC was 1000 µg/mL for the most tested strains. The growth curve showed a lag phase delay for both species, mainly S. aureus, and reduced the growth level of E. coli by half. The combination of EOOG with OXA and CIP led to an additive action for S. aureus. A significant reduction in biofilm biomass and cell viability was verified for S. aureus and E. coli. In conclusion, EOOG has relevant potential as a natural alternative to treat infections caused by multiresistant strains.

In vitro resistance of Enterobacter cloacae isolated from fresh seafood to colistin

Abstract INTRODUCTION: Enterobacter cloacae is a clinically important bacterium from the Enterobacteriaceae family. This study evaluated resistance of E. cloacae strains from fish (n=14) and shrimp (n=9) to colistin. METHODS: Biochemical identification and antimicrobial susceptibility tests were carried out in an automated Vitek®2 instrument. RESULTS: Colistin resistance was observed in 21.4% and 66.7% of the strains from fish and shrimp, respectively. We observed minimum inhibitory concentrations of ≥16 mg/L and ≤5 mg/L in 8 and 15 of all strains, respectively. CONCLUSIONS: Fish and shrimp can carry drug-resistant enterobacteria, which can be of clinical interest.

Plectranthus amboinicus essential oil and carvacrol bioactive against planktonic and biofilm of oxacillin- and vancomycin-resistant Staphylococcus aureus.

BACKGROUND: The emergence of multidrug-resistant bacteria is a worldwide concern and in order to find an alternative to this problem, the occurrence of antimicrobial compounds in Plectranthus amboinicus essential oil was investigated. Thus, this study aims to determine susceptibility of Staphylococcus aureus isolated from food to antibiotics, P. amboinicus essential oil (PAEO) and carvacrol. METHODS: Leaves and stem of P. amboinicus were used for extraction of essential oil (PAEO) by hydrodistillation technique and EO chemical analysis was performed by gas chromatography coupled to a mass spectrometer. S. aureus strains (n = 35) isolated from food and S. aureus ATCC 6538 were used to evaluate the antimicrobial and antibiofilm activity of PAEO and carvacrol. All strains (n = 35) were submitted to antimicrobial susceptibility profile by disk diffusion method. Determination of MIC and MBC was performed by microdilution technique and antibiofilm activity was determined by microtiter-plate technique with crystal violet assay and counting viable cells in Colony Forming Units (CFU). RESULTS: Carvacrol (88.17%) was the major component in the PAEO. Antibiotic resistance was detected in 28 S. aureus strains (80%) and 12 strains (34.3%) were oxacillin and vancomycin-resistant (OVRSA). From the 28 resistant strains, 7 (25%) showed resistance plasmid of 12,000 bp. All strains (n = 35) were sensitive to PAEO and carvacrol, with inhibition zones ranging from 16 to 38 mm and 23 to 42 mm, respectively. The lowest MIC (0.25 mg mL-1) and MBC (0.5 mg mL-1) values were observed when carvacrol was used against OVRSA. When a 0.5 mg mL-1 concentration of PAEO and carvacrol was used, no viable cells were found on S. aureus biofilm. CONCLUSION: The antibacterial effect of carvacrol and PAEO proves to be a possible alternative against planktonic forms and staphylococcal biofilm.

In Vitro Antibacterial and Antibiofilm Activity of Lippia alba Essential Oil, Citral, and Carvone against Staphylococcus aureus.

In vitro antimicrobial and antibiofilm activities of the Lippia alba essential oil and its major components (citral and carvone) against Staphylococcus aureus were investigated. Essential oils (LA1EO, LA2EO, and LA3EO) were extracted from the aerial parts of three L. alba specimens by hydrodistillation and analyzed by gas chromatography coupled to a mass spectrometer. Minimum Inhibitory Concentrations (MIC) and Minimum Bacterial Concentration (MBC) were determined by the microdilution method. For the antibiofilm assays, the biomass formation in the biofilm was evaluated by the microtiter-plate technique with the crystal violet (CV) assay and the viability of the bacterial cells was analyzed. All oils and their major components presented antibacterial activity, and the lowest MIC and MBC values were 0.5 mg mL-1 when LA1EO and citral were used. Potential inhibition (100%) of S. aureus biofilm formation at the concentration of 0.5 mg mL-1 of all EOs was observed. However, the elimination of biofilm cells was confirmed at concentrations of 1 mg mL-1, 2 mg mL-1, 2 mg mL-1, and 0.5 mg mL-1 for LA1EO, LA2EO, LA3EO, and citral, respectively. The results obtained in the present research point to the promising antibacterial and antibiofilm potential of L. alba EOs against S. aureus, a species of recognized clinical interest.

RAW TROPICAL OYSTERS AS VEHICLES FOR MULTIDRUG-RESISTANT Vibrio parahaemolyticus / Ostras tropicais cruas como fonte de Vibrio parahaemolyticus multirresistentes

The following study aimed to determine the antimicrobial susceptibility profile of Vibrio parahaemolyticus strains from fresh and frozen oysters Crassostrea rhizophorae sold in Fortaleza-Brazil. An antibiogram was performed on 87 isolates using nine antibiotics: gentamicin (Gen 10 µg), ampicillin (Amp 10 µg), penicillin G (Pen 10U), ciprofloxacin (Cip 5 µg), chloramphenicol (Chl 30 µg), nalidixic acid (Nal 30 µg), tetracycline (Tet 30 µg), vancomycin (Van 30 µg) and erythromycin (Ery 15 µg). All strains were resistant to at least one antibiotic, and 85 (97.7%) were multi-resistant, with predominance of the Van+ Pen+Amp resistance profile (n = 46). Plasmid resistance to Pen, Amp and Ery was detected. Thus, the risk that raw oyster consumption poses to the health of consumers is highlighted, due to the fact that these bivalves may host antibacterial-resistant microorganisms.

O presente estudo objetivou determinar o perfil de suscetibilidade a antimicrobianos de cepas de Vibrio parahaemolyticus oriundas de ostras “in natura” e congeladas comercializadas em Fortaleza-Brasil. Oitenta e sete (87) cepas foram submetidas ao antibiograma com emprego de nove antibióticos: gentamicina (Gen 10 µg), ampicilina (Amp 10 µg), penicilina G (Pen 10U), ciprofloxacin (Cip 5 µg), cloranfenicol (Clo 30 µg), ácido nalidíxico (Nal 30 µg), tetraciclina (Tet 30 µg), vancomicina (Van 30 µg) e eritromicina (Eri 15 µg). Todas as cepas mostram-se resistentes a pelo menos um antibiótico, e 85 (97,7%) apresentaram multirresistência, com predomínio do perfil Van+ Pen+Amp (n = 46). Foi detectada resistência plasmidial a Pen, Amp e Eri. Dessa forma, o risco que o consumo de ostras cruas representa para a saúde dos consumidores merece ser destacado, uma vez que esses bivalves podem ser veículos de transmissão de micro organismos multirresistentes a fármacos antibacterianos.

Multiple enzymatic profiles of Vibrio parahaemolyticus strains isolated from oysters / Multiple enzymatic profiles of Vibrio parahaemolyticus strains isolated from oysters.

The enzymatic characterization of vibrios has been used as a virulence indicator of sanitary interest. The objective of this study was to determine the enzymatic profile of Vibrio parahaemolyticus strains (n=70) isolated from Crassostrea rhizophorae oysters. The strains were examined for the presence of gelatinase (GEL), caseinase (CAS), elastase (ELAS), phospholipase (PHOS), lipase (LIP), amilase (AML) and DNase. All enzymes, except elastase, were detected in more than 60

of the strains. The most recurrent enzymatic profiles were AML + DNase + PHOS + GEL + LIP (n=16; 22.9

) and AML + CAS + DNase + PHOS + GEL + LIP (n=21; 30

). Considering the fact that exoenzyme production by vibrios is closely related to virulence, one must be aware of the bacteriological risk posed to human health by the consumption of raw or undercooked oysters.

Hemolytic and urease activities in vibrios isolated from fresh and frozen oysters

INTRODUCTION: The present study aimed to survey the Vibrio microbiota of oysters (Crassostrea rhizophorae) obtained from restaurants in Fortaleza, State of Ceará, Brazil, and to identify virulence factors. METHODS: The isolated vibrios were submitted to biochemical identification and were tested for hemolytic and urease activities. RESULTS: The isolated strains belonged to 13 species, with predominance of Vibrio mimicus. Of the strain isolates only from fresh samples, 20.5% and 2.8% showed hemolytic and urease activities, respectively. CONCLUSIONS: The findings support the little-publicized claim that Vibrio species other than V. parahaemolyticus and V. vulnificus can represent a health risk to public health.

Multiple enzymatic profiles of Vibrio parahaemolyticus strains isolated from oysters.

The enzymatic characterization of vibrios has been used as a virulence indicator of sanitary interest. The objective of this study was to determine the enzymatic profile of Vibrio parahaemolyticus strains (n=70) isolated from Crassostrea rhizophorae oysters. The strains were examined for the presence of gelatinase (GEL), caseinase (CAS), elastase (ELAS), phospholipase (PHOS), lipase (LIP), amilase (AML) and DNase. All enzymes, except elastase, were detected in more than 60% of the strains. The most recurrent enzymatic profiles were AML + DNase + PHOS + GEL + LIP (n=16; 22.9%) and AML + CAS + DNase + PHOS + GEL + LIP (n=21; 30%). Considering the fact that exoenzyme production by vibrios is closely related to virulence, one must be aware of the bacteriological risk posed to human health by the consumption of raw or undercooked oysters.

Uso de soro de leite bovino para elaboração de meios de cultura para Staphylococcus aureus / Culture media made by bovine whey for Staphylococcus aureus growth

Foi investigada a eficiência do cultivo de Staphylococcus aureus ATCC 25923 em meios experimentais elaborados com soro de leite bovino. Foram preparados meios líquido (caldo soro – CS) e sólido (ágar soro – AS) suplementados com cloreto de sódio (CS+NaCl e AS+NaCl) e extrato de levedura (CS+Lev eAS+Lev). No meio AS, foi testado o enriquecimento com extrato de levedura (0,1%) e diluição em água(40%) (AS+H2O+Lev). Os meios comerciais de caldo infusão cérebro coração (BHI) e ágar Baird-Parker(BP) foram utilizados como controle. A eficiência dos meios experimentais foi avaliada por determinação da biomassa e contagem de colônias, utilizando-se a cepa S. aureus ATCC 25923. Em todos os testes, o crescimento bacteriano foi compatível à quantificação observada nos meios utilizados como controle e houve desenvolvimento de colônias com características morfológicas típicas de S. aureus, principalmente no meio AS+H2O+Lev. Concluiu-se que o soro de leite bovino pode ser utilizado como componente alternativo na elaboração de meio de cultura para S. aureus.

Staphylococcus coagulase-positiva e enterobactérias em camarão Litopenaeus vannamei comercializado in natura / Coagulase-positive staphylococci and enterobacteria in fresh shrimp Litopenaeus vannamei

A qualidade microbiológica de camarões (Litopenaeus vannamei) comercializados in natura em Fortaleza(CE) foi investigada por meio de quantificação de Staphylococcus coagulase-positiva (Sph CP), coliformes totais (CT) e termotolerantes (CTT) e também pela detecção de Salmonella e isolamento de Escherichiacoli. Das 24 amostras adquiridas no comércio varejista, quatro (16,7 por cento) apresentaram índices de Sph CP acima de 103/g. Não foi detectada Salmonella nas amostras analisadas. O número de amostras positivas para CT, CTT e E. coli foi, respectivamente, de 23 (95,8 por cento), 13 (54,2 por cento) e 6 (25 por cento). De acordo com a legislação vigente no Brasil, as quatro amostras com contagens de Sph CP > 103/g são consideradas impróprias para o consumo. Ademais, chama-se a atenção para a ocorrência de E. coli em 25 por cento das amostras analisadas, uma vez que essa bactéria é indicadora de contaminação fecal.

Coliformes, salmonella e staphylococcus coagulase positiva em hortaliças utilizada na merenda escolar da rede pública de ensino de Sobral, CE / Coliforms, salmonella and staphylococcus coagulase positive in vegetables used in school lunches in public schools in Sobral, CE

Foram pesquisadas 24 amostras de hortaliças utilizadas na merenda da rede de escola pública de Sobral – Ceará. A pesquisa retrata a contagem e identificação de bactérias do grupo dos coliformes, Staphylococcus coagulase positiva e pesquisa de Salmonella. Das 24 amostras 62,5% apresentaram contaminação por Coliformes termotolerantes acima da permitida pela ANVISA. Cerca de 14 cepas de Coliformes termotolerantes foram isoladas sendo 14,3% Citrobacter diversus e 85,7% Enterobacter agglomerans. A quantificação de coliformes totais variou de <1,8 a >160 x 10, não sendo detectada a presença de Salmonella. A contagem de Staphylococcus variou de 250 a 20 x 10 UFC/g, desses foram isoladas 32 cepas, sendo 9 de Staphylococcus coagulase positiva e 23 de Staphylococcus coagulase negativa.

Staphylococcus coagulase positiva e coliformes em leite servido em merenda escolar / Staphylococcus coagulase positive and coliforms in milk served in school feeding

O leite é um dos alimentos mais servidos na merenda escolar, sendo considerado de alto valor nutricional devido aos seus constituintes, o que o torna um excelente meio nutritivo para o crescimento de bactérias. Foram analisadas 15 amostras de leite servido na merenda de uma escola pública na cidade de Sobral, CE sendo 5 amostras de leite em pó diluído em água, 5 pasteurizado e 5 UHT (ultra alta temperatura). A avaliação microbiológica foi concernente à determinação do Número Mais Provável (NMP) de coliformes totais (CT) e termotolerantes (CTT), presença de Salmonella, identificação e contagem de Staphylococcus. (...) Detectou-se a presença de Staphylococcus coagulase positiva e ausência de Salmonella em todas as amostras.

Bactérias entéricas presentes em amostras de água e camarão marinho litopenaeus vannamei oriundos de fazendas de cultivo no estado do Ceará, Brasil / Enteric bacteria in water samples and marine shrimp litopenaeus vannamei cultured in the state of Ceará, Brazil

Foram realizados a quantificação de coliformes totais (CT) e termotolerantes (CTT), isolamento e identificação de coliformes, e pesquisa de Salmonella em 28 amostras de água e 28 de camarão da espécie Litopenaeus vannamei, oriundas de duas fazendas de cultivo localizadas no Estado do Ceará, Brasil. Nenhuma amostra de água apresentou índice de CTT acima do limite de 2.500/100 mL preconizado pela legislação para águas salobras destinadas ao cultivo de organismos para fins de consumo. O Número Mais Provável (NMP/g) de CTT das amostras de camarão variou de <3 a 2,9 x 104. A maior frequência de isolamento de coliformes nas amostras de água e camarão foi a da espécie Escherichia coli. Em apenas três (5,35%), das 56 amostras analisadas, foi detectada a presença de Salmonella sorovar Newport e S. Saintpaul. Apesar do baixo índice de CTT e da baixa incidência de salmonela, a presença dessas bactérias entéricas em ambientes de cultivo de peneídeos é preocupante, uma vez que podem provocar infecções em humanos.

Samples of water (n = 28) and Litopenaeus vannamei (n = 28) from two shrimp farms in Ceará state, Brazil were evaluated for total coliforms (TC), total thermotolerant coliforms (TTC), coliform species diversity and Salmonella. No water sample presented TTC levels above the maximum level (2,500 MPN/100 mL) established by regulation for brackish water aquaculture producing seafood for human consumption. The most probable number (MPN) of TTC in shrimp samples ranged from <3 to 2.9 x 104 CFU/g. The coliform species most frequently isolated from water and shrimp was Escherichia coli. Only three (5.35%) of the 56 samples tested were positive for Salmonella (Newport and Saintpaul serovars). In spite of the low TTC levels observed, the presence of potentially pathogenic enteric bacteria in shrimp culture is a disquieting finding.

Antibacterial effect (in vitro) of Moringa oleifera and Annona muricata against Gram positive and Gram negative bacteria / Efeito antibacteriano (in vitro) de Moringa oleifera (moringa) e Annona muricata (graviola) frente a bactérias Gram-negativas e Gram-positiva

Antibacterial effects of aqueous and ethanolic extracts of seeds of moringa (Moringa oleifera) and pods of soursop (Annona muricata) in the concentration of 1:5 and 1:10 in volumes 50, 100, 150 and 200 µL were examined against Staphylococcus aureus, Vibrio cholerae, Escherichia coli (isolated from the organism and the aquatic environment) and Salmonella Enteritidis. Antibacterial activity (inhibition halo > 13 mm) against S. aureus, V. cholerae and E. coli isolated from the whiteleg shrimp, Litopenaeus vannmaei, was detected in aqueous and ethanolic extracts of moringa. E. coli isolated from tilapiafish, Oreochromis niloticus, was sensitive to the ethanolic extract of moringa. The aqueous extracts of soursop showed an antibacterial effect against S. aureus and V. cholerae, but the antibacterial activity by the ethanol extracts of this plant was not demonstrated.

Para avaliação do efeito bactericida frente à Staphylococcus aureus, Vibrio cholerae, Escherichia coli (isolada de pescados e ambiente aquático) e Salmonella Enteretidis, foram testados extratos aquosos e etanólicos de sementes de moringa (Moringa oleifera) e casca de graviola (Annona muricata) na concentração de 1:5 e 1:10, nos volumes de 50, 100, 150 e 200 µL. Os resultados mostraram efeito antibacteriano (halo de inibição > 13mm) dos extratos aquosos e etanólicos de moringa frente a S. aureus, V. cholerae e E. coli isoladas de camarão cinza Litopenaeus vannmaei. A cepa de E. coli isolada do pescado Oreochromis niloticus apresentou sensibilidade frente ao extrato etanólico de moringa. Os extratos aquosos de graviola apresentaram efeito bactericida frente a S. aureus e V. cholerae, entretanto, os extratos etanólicos dessa planta não mostraram atividade antibacteriana.

Perfil de suscetibilidade a antibióticos de Staphylococcus coagulase positivo isolados de manipuladores de merenda escolar / Profile antibiotic susceptibility of coagulase positive isolates handlers school meals

O presente estudo teve como objetivo a detecção e a verificação da suscetibilidade a antimicrobianos de Staphylococcus isolados de 4 manipuladores de alimentos de 2 escolas da rede pública de ensino da cidade de Sobral-Ceará. De cada manipulador foram obtidas amostras das cavidades oral e nasal, cabelo e mão direita. Para a pesquisa de estafilococos foi utilizado o meio de Baird-Parker (Difco) enriquecido com solução de telurito a 1% e emulsão de gema de ovo a 50%. Foram tomadas 15 cepas de Staphylococcus coagulase positivo para a realização de antibiograma , sendo testados 8 antibióticos. Foi detectado Staphylococcus concomitantemente em todas as áreas anatômicas amostradas de todos os manipuladores avaliados...

Raw oysters can be a risk for infections

The aims of this study were to count and identify sucrose positive and negative vibrios isolated from cultivated Crassostrea rhizophorae oysters during their growing cycle. Every month for 12 months, 10 to 18 oysters were collected for study. Collections occurred at the Center for Studies of Coastal Aquaculture (CSCA), which is associated with the Institute of Marine Science, Labomar, located in Euzebio, Ceará, Brazil. Approximately 150 oysters and their intervalvular liquor were studied. Vibrio Standard Plates Counts (SPC) from oyster meat and their intervalvular liquor varied from 25 to 59,000,000 CFU/g. For most of the 12 months of the oysters' life, it was possible to identify Vibrio parahaemolyticus. Vibrio carchariae was identified in four collections. Among other isolated species, the most important, considering public health risks, was V. vulnificus, although only one strain was confirmed. We concluded that retail purchased oysters should never be eaten raw or undercooked because many species of the genus Vibrio are known to be pathogenic to humans and live naturally on and in shellfish throughout their life cycle.

Enterobactérias em pescado oriundo da Lagoa da Fazenda, Sobral, CE / Enterobacteria in fish from the Lagoa da Fazenda, Sobral, CE

Foram analisadas 46 amostras do pescado Oreochromis niloticus capturadas na Lagoa da Fazenda, em Sobral, CE. As análises foram referentes à enumeração, isolamento e identificação de coliformes fecais (CF) e determinação do perfil e suscetibilidade dos isolados de Escherichia coli e antimicrobianos. Das 46 amostras analisadas, 25 (54 por cento) apresentaram um índice de CF variando de 2 a 460NMP/g. Foram isoladas 50 cepas, sendo 37 (74 por cento) identificadas como E. Coli, 8 (16 por cento) como Enterobacter sp. e 5 (10 por cento) como Klebsiella sp. Das 11 cepas de E. Coli testadas quanto à suscetibilidade aos 11 antibióticos, 9(81,8 por cento) apresentaram sensibilidade a todos os antibióticos testados.

Análise bacteriológica de merenda escolar servida em um colégio estadual de Sobral, Ceará / Bacteriological analysis of school feeding served in a state college in Sobral, Ceará

Foram analisadas 31 amostras de merenda escolar coletadas em um colégio da rede estadual no município de Sobral (CE). A avaliação bacteriológica foi concernente à quantificação de coliformes totais (CT) e fecais (CF), pesquisa de Staphylococcus e Salmonella. Das 31 amostras analisadas, 58 por cento apresentaram contaminação por CT variando de 21 a 18.000NMP/mL, e 35,5 por cento com CF variando de 2 a 2.900NMP/mL. Foi detectada a presença de Staphylococcus coagulase negativa em 35,5 por cento das amostras e ausência de Salmonella em todas as amostras.

Susceptibilidade"in vitro" a antimicrobianos de estirpes de Vibrio spp isoladas de camarões (Litopenaeus vannamei) e de água de criação destes animais provenientes de uma fazenda de camarões no Ceará - Nota prévia / In vitro antimicrobial susceptibility of Vibrio spp isolated from shrimp (Litopenaeus vannamei) and water from ponds of a shrimp farm in Ceará

Foram feitos ensaios de susceptibilidade a antibióticos em 48 cepas de Vibrio isoladas do cultivo do Litopenaeus vannamei no Ceará. Para os testes de difusão foram utilizados 11 antibióticos. A espécie que apresentou maior percentagem de resistência aos antimicrobianos foi V. cholerae, onde 33,33% das 12 cepas testadas mostraram-se resistentes a sulfazotrim, 25% a ampicilina e 33,33% a ceftriaxona.

Suscetibility assays were performed for 48 Vibrio strains isolated from pond-reared Litopenaeus vannamei in Ceará, Brazil. The diffusion assays tested 11 antibiotics. The most resistant species was V. cholerae: of 12 strains tested, one third was resistant to sulfamethoxazole-trimethoprim, one fourth to ampicillin and one third to ceftriaxone.