Evaluation of commercial probiotic lactic cultures against biofilm formation by Cronobacter sakazakii

BACKGROUND/AIMS: Cronobacter sakazakii, an emergent pathogen is considered as a major concern to infants and neonates fed on reconstituted powdered infant milk formula. In conjunction with many other factors, biofilm forming capacity adds to its pathogenic potential. In view of the facts that infants are at highest risk to C. sakazakii infections, and emerging antibiotic resistance among pathogens, it is imperative to evaluate probiotic cultures for their efficacy against C. sakazakii. Therefore, pure probiotic strains were isolated from commercial probiotic products and tested for their antimicrobial and anti-biofilm activities against C. sakazakii. METHODS: A total of 6 probiotic strains were tested for their antibiotic susceptibility followed by antimicrobial activity using cell-free supernatant (CFS) against C. sakazakii. The inhibitory activity of CFS against biofilm formation by C. sakazakii was determined using standard crystal violet assay and microscopic observations. RESULTS: All the probiotic strains were sensitive to ampicillin, tetracycline, vancomycin and carbenicillin whereas most of the strains were resistant to erythromycin and novobiocin. Four of the 6 probiotic derived CFS possessed antimicrobial activity against C. sakazakii at a level of 40 μL. A higher biofilm inhibitory activity (>80%) was observed at initial stages of biofilm formation with weaker activity during longer incubation upto 48 hours (50%–60%). CONCLUSIONS: The study indicated the efficacy of isolated commercial probiotics strains as potential inhibitor of biofilm formation by C. sakazakii and could be further explored for novel bioactive molecules to limit the emerging infections of C. sakazakii.

Genetic characterization of methicillin-resistant Staphylococcus aureus from humans and animals within the community / 대한수의학회지

Methicillin-resistant Staphylococcus (S.) aureus (MRSA) is one of the most important nosocomial pathogens worldwide and the emergence of this strain has become a major clinical problem. In this study, we investigated the prevalence of MRSA and their genetic characteristics in 69 S. aureus isolated from humans and animals. In human isolates, higher antimicrobial resistance rates were observed against penicillin (80.6%), followed by erythromycin (11.9%) and tetracycline (9.0%). All of them were susceptible to clindamycin, enrofloxacin, novobiocin, pirlimycin, trimethoprim/sulfamethoxazole and vancomycin. The resistance patterns in animal isolates were similar to those of human isolates. Two (2.9%) MRSA strains were isolated from human (n = 1) and animal (n = 1), and these isolates were confirmed as carrying the mecA gene. One isolate originating from human was resistant to 7 drugs and the other isolate derived from animal was resistant to 11 drugs. Staphylococcal cassette chromosome mec (SCCmec) variant IIIB was identified in animal isolate but SCCmec type of an isolate from human was not exactly determined. Two MRSA isolates showed unrelated PFGE pattern between them. Our results indicated although the frequency of MRSA isolates from humans and animals was low, a continuous surveillance and monitoring should be called for to prevent the contamination and spread of MRSA in the community. To our knowledge, this is the first time that SCCmec type variant IIIB was detected from animals in Korea.

Suscetibilidade à novobiocina na identificação de amostras de Staphylococcus coagulase negativos (SCoN) isolados de hemoculturas / Susceptibility to the novobiocin in the identification of isolates of Staphylococcus negative (SCoN) isolated from blood cultures

Tradicionalmente o Laboratório de Microbiologia Clínica utiliza a prova de suscetibilidade a novobiocina para distinguir as espécies clinicamente significativas de SCoN, entre elas o Staphylococcus saprophyticus. Devido ao aumento destes microrganismos nas infecções relacionadas à assistência à saúde, este estudo teve como objetivo relatar duas bacteremias por SCoN resistentes a novobiocina ocorridas em maio e setembro de 2006, em um hospital geral, na cidade de São Paulo. Primeiramente o teste fenotípico apontou resistência a novobiocina, mas com padrões distintos de identificação ao S. saprophyticus. Na identificação convencional, asamostras fermentaram trealose, manitol e manose, sendo positivas nos testes da urease e fosfatase alcalina. No sistema semi-automatizado, a confirmação da espécie apontou o Staphylococcus cohnii subsp. urealyticus com 99,99 de probabilidade. No teste de disco difusão, os isolados mostraram-se resistentes à oxacilina, mas suscetível a cefoxitina, vancomicina e teicoplanina. Houve confirmação pela metodologia do Etest® mostrando CIM para oxacilina superior a 256 μg/ml, e suscetibilidade a vancomicina e a teicoplanina.A reação da PCR confirmou a presença do gene mecA nos isolados. Estes dados demonstram a importância dos SCoN isolados em hemoculturas, sendo necessária uma correta identificação destes microrganismos.

Antibiotic resistance profile: isolation and characterization of clinical isolates of staphylococci from patients with community-acquired skin infections

Although most bacterial infections of the skin bear out to be minor in nature, a few such dermatologic entities are major, to the spot of yet being fatal. The mortality rate is usually up to 30% to 50% and depends upon the type of infection, original disease, and resistant type. In this study hundred and five bacterial strains were isolated from skin wounds, burns and acne patients from hospitals at different locations in the cosmopolitan city of Karachi. These bacterial strains were identified by conventional methods. Seventy two percent [72%] of total isolated organisms were found to be Staphylococcus aureus while the remaining thirty three percent [33%] were Staphylococcus epidermidis. The antibiotic resistance of identified organisms was carried out by disc-diffusion method with commercially available disc of five antibiotics having different mode of actions such as cell wall synthesis inhibitors, membrane permeability alternatives and DNA synthesis inhibitors. Staphylococcus aureus show more resistant to these antibiotics as compared to Staphylococcus epidermidis. The most effective antibiotic for Staphylococcus aureus is vancomycin showing 80.5% efficacy, then methicillin with 68.0% efficacy, erythromycin with 55.6% efficacy, novobiocin with 54.1% efficacy and then bacitracin with 25.0% efficacy. The most effective antibiotic for Staphylococcus epidermidis is methicillin showing 84.8% efficacy, then vancomycin with 81.2% efficacy, novobiocin with 63.6% efficacy, erythromycin with 42.4% efficacy and then bacitracin with 27.8% efficacy

Assessment of human DNA repair (NER) capacity with DNA repair rate (DRR) by comet assay / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>Alkaline comet assay was used to evaluate DNA repair (nucleotide excision repair, NER) capacity of human fresh lymphocytes from 12 young healthy non-smokers (6 males and 6 females).</p><p><b>METHODS</b>Lymphocytes were exposed to UV-C (254 nm) at the dose rate of 1.5 J/m2/sec. Novobiocin (NOV) and aphidicolin (APC), DNA repair inhibitors, were utilized to imitate the deficiency of DNA repair capacity at the incision and ligation steps of NER. Lymphocytes from each donor were divided into three grougs: UVC group, UVC plus NOV group, and UVC plus APC group. DNA single strand breaks were detected in UVC irradiated cells incubated for 0, 30, 60, 90, 120, 180, and 240 min after UVC irradiation. DNA repair rate (DRR) served as an indicator of DNA repair capacity.</p><p><b>RESULTS</b>The results indicated that the maximum DNA damage (i.e. maximum tail length) in the UVC group mainly appeared at 90 min. The ranges of DRRs in the UVC group were 62.84%-98.71%. Average DRR value was 81.84%. The DRR difference between males and females was not significant (P < 0.05). However, the average DRR value in the UVC plus NOV group and the UVC plus APC group was 52.98% and 39.57% respectively, which were significantly lower than that in the UVC group (P < 0.01).</p><p><b>CONCLUSION</b>The comet assay is a rapid, simple and sensitive screening test to assess individual DNA repair (NER) capacity. It is suggested that the time to detect DNA single strand breaks in comet assay should include 0 (before UV irradiation), 90 and 240 min after exposure to 1.5 J x m(-2) UVC at least. The DRR, as an indicator, can represent the individual DNA repair capacity in comet assay.</p>

Detecting DNA repair capacity of human lymphocytes exposed to ultraviolet C with comet assay / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To assess DNA repair capacity of human lymphocytes with comet assay.</p><p><b>METHODS</b>Fresh lymphocytes form twelve 26-year old donors (6 males, 6 females) were exposed to ultraviolet C (UVC, 254 nm) at the dose rate of 1.5 J/m(2). The lymphocytes of each donor were divided into three parts: UVC group, UVC + aphidicolin (APC) group, UVC + novobiocin (NOV) group. DNA single strand breaks were detected with comet assay in UVC-irradiated cells and unirradiated cells incubated for 30, 60, 90, 120, 180 and 240 min. DNA repair rate (DRR) was calculated and served as an indicator of DNA repair capacity.</p><p><b>RESULTS</b>The maximum average comet tail length (MTL) in three groups appeared 90 min after UVC exposure. The DRR range of UVC group was 81.84% (62.84% - 98.71%); There was no significant difference in DRR between males and females (P > 0.05). However, the average DRRs of UVC + NOV group and UVC + APC group (52.98% and 39.57% respectively) were significantly lower than that of UVC group (P < 0.01).</p><p><b>CONCLUSION</b>Comet assay is a rapid and simple screening test to assess DNA repair capacity. DRR, as an indicator, may express the individual DNA repair capacity.</p>

Enzymatic characterization of Mycobacterium smegmatis ADP-ribosyltransferase

ADP-ribosyltransferase (ADPRT) catalyzes the reaction in which the ADP-ribose moiety of beta-NAD+ is transferred to specific amino acid residues in target proteins. The ADPRT of Mycobacterium smegmatis has been known to inactivate rifampin through ADP-ribosylation. However, the enzymatic characteristics and functions of the enzyme have not been elucidated yet. In this study, the ADPRT-glutathione S-transferase (GST) fusion protein was expressed in Escherichia coli and enzymatic characteristics of the fusion protein were investigated. ADPRT-GST fusion protein was an ADPribosyltransferase that had no NAD glycohydrolase activity. ADPRT-GST fusion protein showed no self-inactivation phenomenon that is a universal nature for all NAD glycohydrolases and is important in regulating its activity. ADPRT activity of the enzyme was decreased by novobiocin and isonicotinic acid hydrazide. These results suggest that Mycobacterium smegmatis ADPRT could be regulated by a different way from other NADases and involved in bacterial physiological process through a post-translational modification of cytosolic proteins.

Novobiocin inhibits angiogenesis and shows synergistic effect with vincristine / 药学学报

<p><b>AIM</b>To study the anti-angiogenic activity of novobiocin and its mechanism of action.</p><p><b>METHODS</b>The anti-angiogenic activity of novobiocin was determined using chick embryo chorioallantoic membrane(CAM) assay. MTT assay, zymography and related assays were used to observe the effects of drugs on bovine aorta endothelial cells and human pulmonary carcinoma PG cells.</p><p><b>RESULTS</b>Novobiocin at the doses of 100 and 200 micrograms/egg inhibited angiogenesis by 31.6% and 68.7% in CAM, respectively. The combination of novobiocin and vincristine enhanced the anti-angiogenic effect. Novobiocin inhibited the proliferation of bovine aortic endothelial cells in a concentration-dependent manner. In addition, novobiocin suppressed MMP-2 secretion, migration, and tube formation of endothelial cells. As determined by MTT assay, novobiocin in combination with vincristine displayed synergistic effect on the proliferation of PG cells,</p><p><b>CONCLUSION</b>This study demonstrates that novobiocin is active in suppressing angiogenesis and the anti-angiogenic activity may be enhanced by combination with vincristine. The anti-angiogenic activity of novobiocin may be related, at least in part, to its inhibition of cell proliferation, cell migration, tube formation and secretion of matrix metalloproteinases.</p>

Susceptibilidad a antimicrobianos de cepas de estafilococos coagulasa positivos aisladas de mastitis bovina en la cuenca lechera central de la Argentina / Antimicrobial sensitivity of coagulase-positive staphylococcal strains isolated from bovine mastitis in the central dairy catchment area of Argentina

The activity of antimicrobial agents frequently used for treating bovine mastitis was determined against 101 coagulase-positive staphylococci isolated from bovine mammary secretion. The isolates were obtained from 39 dairy farms located in the central dairy area of Argentina. The disk diffusion method was used and the following antimicrobial agents were tested: penicillin, ampicillin, oxacillin, cephacetrile, penicillin + novobiocin, erythromycin, pirlimycin, novobiocin and neomycin. The highest levels of resistance were observed against penicillin and ampicillin (47.6), while the lowest against erythromycin (2), pirlimycin (4) and neomycin (2.9). No resistant strains against oxacillin, cephacetrile and penicillin + novobiocin were detected.

Contaminación bacteriana y presencia de Sinorhizobium meliloti en aguas de los canales de riego del río Neuquén / Bacterial contamination and presence of Sinorhizobium meliloti in irrigation canal water from the NeuquÚn River

A survey of the changes in populations of heterotrophic bacteria, coliform microorganisms and S. meliloti was conducted in samples taken from the water irrigation channels of the NeuquÚn River (Argentina). Fifty-six water samples were collected during the spring-summer seasons of 1997-1999 years. Both the heterotrophic plate count bacterial and the number of coliforms oscillated between 110-5050 CFU/ml and 8-1400 CFU/100 ml, respectively, during the period this study was carried out. Fecal coliforms were detected in 91.1 of the water samples investigated. Moreover, the results showed that S. meliloti capable of nodulating alfalfa (Medicago sativa L.) Cuf 101 were present in 68 of the water samples and in effectiveness studies, no isolate out of 25 evaluated could be classified as superior N fixers. That is, they did not produce plants equal in weight to nitrate-grown plants (KNO3 0.05). All the S. meliloti strains were resistant to novobiocin and bacitracin, while 72 of the microsymbionts demonstrated resistance to between seven and ten antibiotics. Results presented in this study showed that irrigation waters of the NeuquÚn river could act as dispersal agents of both ineffective S. meliloti strains and thermotolerant coliform bacteria.

Influence of novobiocin on gama-irradiation G0-lymphocytes as analyzed by cytogenetic endpoints

Experiments with novobiocin (NB) post-treatment were performed to verify its effect on the frequencies of micronuclei (MN) and chromosomal aberrations (CA) induced by g-irradiation (0.75, 1.5 and 3.0 Gy) in human lymphocytes at G0-phase. The frequencies of MN significantly decreased by 44 and 50 per cent, for the treatment with NB 50 µg/ml (30-min pulse) after radiation doses of 1.5 and 3.0 Gy, respectively. However, CA frequencies were not significantly affected. No significant effect on CA was observed when lymphocyte cultures were exposed to a single dose of 2.0 Gy at the G0-phase and posttreated with 25 µg/ml NB for three hours either immediately after irradiation (G0-phase) or after 24 h (S-phase). The significant suppressive effect of NB on MN frequencies supports the hypothesis that NB interaction with chromatin increases access to DNA repair enzymes.

Significância etiológica e características de estafilococos coagulase-negativa isolados de processos infecciosos em recém-nascidos / Etiologic significance and characteristics of coagulase-negative staphylococci isolated from infectious among newborns

Os estafilococos coagulase-negativa (ECN), embora reconhecidos como saprófitas por muito tempo, têm emergido como agentes etiológicos de uma série de infecçöes, sendo atualmente os mais freqüentemente isolados em infecçöes hospitalares. Esses microrganismos apresentam elevado risco potencial de bacteremia nosocomial entre recém-nascidos com baixo peso ao nascimento, os quais geralmente säo imunologicamente imaturos e freqüentemente requerem procedimentos invasivos para a adminstraçäo de substâncias nutritivas e medicamentosas. O aumento da incidência de bacteremia nosocomial por ECN em neonatos nos últimos anos, tem sido também associado ao aumento da sobrevivência de crianças prematuras com peso ao nascimento menor que 1.500 g e à sua longa permanência no ambiente hospitalar. Contudo, como os ECN fazem parte da flora normal da pele, freqüentemente contaminam espécimes clínicos, sendo muitas vezes negligenciados por esse aspecto quanto à sua importância etiológica. Tendo em vista essas características, este estudo teve como objetivos avaliar a significância etiológica de estafilococos coagulase-negativa isolados de processos infecciosos em recém-nascidos da Unidade neonatal do Hospital das Clínicas da Faculdade de Medicina de Botucatu e a caracterizaçäo das linhagens quanto aos fatores de virulência e sensibilidade às drogas. As linhagens de ECN isoladas foram classificadas em significativas, suspeitas e contaminantes com base em uma série de dados clínicos e laboratoriais. Das 117 linhagens de ECN isoladas, 51 (43,6 por cento) foram classificadas como significativas, nove (7,7 por cento) como suspeitas e 57 (48,7 por cento) como contaminantes. Das 45 crianças com infecçäo por ECN, 37 (82,2 por cento) eram prematuras e 22 (48,9 por cento) com peso ao nascimento < 1500g. A maioria das crianças com infecçäo por ECN estavam submetidas a dois ou mais procedimentos invasivos (77,8 por cento), incluindo o uso de catéter (88,9 por cento), nutriçäo parenteral (66,7 por cento) e ventilaçäo mecânica (57,8 por cento). O Staphylococcus epidermidis foi a espécie mais frequentemente isolada (77,8 por cento) e mais associada com infecçäo (90,2 por cento) do que com contaminaçäo (68,4 por cento). Outras espécies de ECN, incluindo uma linhagem de S. haemolyticus, duas linhagens de S. lugdunensis, uma linhagem de S. simulans e uma linhagem de S. xylosis, também foram isoladas de crianças com evidência clínica de pneumonia, enterocolite necrosante ou sepse...

Aislamiento de Staphylococcus xylosus, asociado con vulvo-vaginitis pediatrica / Isolation of Staphylococcus xylosus associated with pediatric vulvovaginitis

A traves de procedimientos bacteriologicos sencillos pero conlcuyentes, se aisla el 13-07-1994 estaphylococcus excylosus en un proceso cronico de vulvo-vaginitis (muestra 442/1994) siendo la base de este procedimiento el esquema propuesto por Kloos y Schleifer. El antibiograma (KIRBI-BAUER) demostro sensibilidad a Rifampicina, Cefradina, Ciprofloxacina, Gentamicina, Oxacilina y resistencia a la ampicilina, lincomicina y eritromicina, en concordancia con otros resultados internacionales previos. Es pertinente desarrollar este tipo de pruebas en Bolivia para delimitar el espectro epidemiologicos de este y otros ESTAPHYLOCOCCUS COAGULASA NEGATIVOS (CNS)

Utilizaçäo do meio de cultura novobiocina-nitrato-sacarose (NNS) no diagnóstico presuntivo de staphylococcus saprophyticus / Use of NNS (Novobiocin-Nitrate-Sucrose) medium to differentiate staphylococcus saprophyticus from other coagulase-negative staphylococci

O meio de cultura NNS, constituído de púrpura de bromocresol, nitrato de potássio, sacarose e novobiocina, foi utilizado na diferenciaçäo de Staphylococcus saprophyticus isolados de urina. Na formulaçäo do meio NNS, a resistência à novobiocina, característica normalmente utilizada no diagnóstico presuntivo de Staphylococcus saprophyticus, foi associada a sua capacidade de utilizar a sacarose e à ausência da enzima nitrato-redutase, visando a diferenciaçäo deste microorganismo dos demais estafilococos coagulase negativa, inclusive os igualmente resistentes à novobiocina (STAPHYLOCOCCUS COHNII, STAPHYLOCOCCUS XYLOSUS E STAPHYLOCOCCUS SCIURI). Os Staphylococcus Cohnii näo utilizam a sacarose (88% das cepas), enquanto os Staphylococcus xylosus e Staphylococcus sciuri reduzem nitrato a nitrito em 100 e 80% das cepas, respectivamente. De 74 cepas de estafilococos coagulase negativa isoladas de urina, 49 (66,2%) cepas de Staphylococcus saprophyticus foram diferenciadas pelo meio NNs, tendo sido obtidos resultados de valor predicativo de resultado positivo de 100%, sensibilidade de 95,9%, especificidade de 100% e eficiência de 97,2%...

Acción sobre haemophilus influenzae de su ácido desoxirribonucléico transformante irradiado in vitro con luz ultravioleta a -70- / Action on haemophilus influenzae of its transforming deoxyribonucleic acid irradited in vitro with ultraviolet light at -70-C

La irradiación con luz ultravioleta a - 70-C de DNA transformante desnaturalizado, el cual fue posteriormente sonicado y renaturalizado con DNA no irradiado, así como el DNA no irradiado y sometido a los otros tratamientos, provocaron un fuerte efecto letal sobre H. influenzae, el cual fue detecto al efectuarse la cuenta viable de las mezclas de transformación, encontrándose la disminución de la viabilidad de las células de un 98% para el caso del DNA irradiado y de un 97.2% para el DNA sin irradiar. Pensamos que la letalidad pueda deberse, tanto a la integración al genóforo de la célula receptora de los marcadores letales provados por la luz UV, como a la activación de un fago defectuoso causada por la penetración del DNA transformante irradiado o no con luz UV. La irradiación con luz UV del DNA transformante desnaturalizado, el cual fue renaturalizado consifo mismo, hizo aumentar ligeramente el número de mutantes resistentes a Kanamicina en células competentes de H. influenzae. Al someterse a lisis sónica del DNA irradiado, antes de renaturalizarse con DNA no irradiado, el resultaod obtenido fue aproximadamente el mismo que en el caso anterior. Sin embargo, cuando el número de mutantes resistentes a novobiocina se corrigió por la cuenta viable de la mezcla de transformación y el efecto mutagenético se cuantificó con relación a las frecuencias de mutación, encontramos un aumento de dichas frecuencias de 74 veces respecto a los testigos de células no tratadas con DNA. Este efecto puede, en principio, deberse a la integración en el genoma receptor de las lesiones mutagenéticas causadas por la irradiación con luz UV a - 70-C del DNA transformante, mas la afirmación concluyente de esto, requerirá la demostración de que dicha integración se lleva a cabo. Proponemos que estas lesiones mutagenéticas pudieran ser del tipo 5-timinil-5, 6-dihidrotimina. Fue interesante que del SNA no irradiado con luz UV, provocara también un aumento aunque menor al producido por el DNA irradiado, de las frecuencias de mutación. La explicación de este hecho, necesitará de un análisis experimental detallado posterior, utilizando mutantes rec y uvr de H. influenzae

Identificaçäo presuntiva de Staphylococcus saprophyticus através de método automatizado / Presumptive identification of Staphylococcus saprophyticus using an automated method

Os autores propöem novo método automatizado para verificaçäo do comportamento de estafilococos coagulase negativa à novobiocina, através do sistema MS-2 (Abbott Laboratories, Diagnostics Div., Irving, Texas). A nova técnica automatizada possibilita o teste de um maior número de espécimes em um tempo médio de 99 minutos, apresentando 100% de concordância com o método tradicional de cultura