Establishment of a saliva donor selection for in vitro biofilm growth / Seleção de doadores de saliva para crescimento de biofilme in vitro

Introdução: O emprego de biofilmes polimicrobianos, utilizando a saliva como inóculo, é um modelo promissor para o estudo de biofilmes cariogênicos in vitro. Entretanto, ainda não existe uma padronização para seleção de doadores de saliva. Objetivo: O objetivo deste estudo foi estabelecer uma metodologia para seleção de doadores de saliva utilizando fatores salivares microbianos e características in vitro do biofilme. Material e método: Para doação de saliva foram selecionados vinte voluntários. Os voluntários permaneceram 24 horas sem escovar os dentes e ficaram em jejum por 2 horas antes da coleta da saliva. Foram avaliados os seguintes parâmetros: viabilidade das bactérias anaeróbias totais e mutans streptococci; concentração inibitória mínima (CIM) e concentração bactericida mínima (CBM) da clorexidina; capacidade de formação de biofilme por meio da biomassa; e a suscetibilidade dos biofilmes à clorexidina. Resultado: A viabilidade bacteriana da saliva, a capacidade de formação de biofilme e a suscetibilidade do biofilme à clorexidina foram apresentadas como média e intervalo de confiança (95%). A diferença entre a viabilidade do biofilme (mutans streptococci e bactérias totais) após tratamento com NaCl 0,9% e diacetato de clorexidina 0,2% foi comparada pelo teste t de Student com nível de significância estabelecido em 5%. A viabilidade total de bactérias anaeróbias (mediana) foi de 7,28 log 1+UFC/mL (unidades formadoras de colônia/mL). A viabilidade dos mutans streptococci na saliva apresentou mediana de 5,47 log 1+UFC/mL. Para capacidade de formação de biofilme a mediana da biomassa foi de 0,1172 A570. Conclusão: O tratamento com clorexidina reduziu significativamente os mutans streptococci e a viabilidade total das bactérias. A metodologia para seleção do doador de saliva foi estabelecida com sucesso.

Introduction: The utilization of polymicrobial biofilms, with saliva as an inoculum, represents a promising model for in vitro studies on cariogenic biofilms. However, there is still no standardization for selecting saliva donors. Objective: The aim of this study is to establish a methodology for the selection of saliva donors using microbial salivary factors and in vitro biofilm characteristics. Material and method: For saliva donation, twenty volunteers were selected. Volunteers remained 24 h without brushing their teeth and fasted for 2 h before saliva collection. The following parameters were evaluated: total anaerobic bacteria and mutans streptococci viability; minimum inhibitory concentration (MIC) and minimum bactericide concentration (MBC) of chlorhexidine; biofilm forming capacity by biomass assessment; and the susceptibility of biofilms to chlorhexidine. Result: Saliva bacterial viability, biofilm forming capacity and biofilm susceptibility to chlorhexidine were presented as mean and confidence interval (95%). The difference between biofilm (mutans streptococci and Total bacteria) viability after treatment with NaCl 0.9% and 0.2% chlorhexidine diacetate was compared using the Student t-test with a significance level established at 5%. Total anaerobic bacteria viability (median) was 7.28 log 1+CFU/mL (colony forming units/ mL). Mutans streptococci viability in the saliva showed a median of 5.47 log 1+CFU/mL. Biofilm forming capacity showed that biomass had a median of 0.1172 A570. Conclusion: Treatment with chlorhexidine significantly reduced mutans streptococci and total bacteria viability. The methodology for the selection of the saliva donor was successfully established.

Effects of nutrient additives and incubation period on sporulation and viability of the entomopathogenic fungus, Metarhizium anisopliae (Hypocreales: Clavicipitaceae)

Aims@#Metarhizium anisopliae is an entomopathogenic fungus (EPF) that exists naturally in the environment and potentially be used as a biological control agent against many insect pests. This study aims to evaluate the effect of nutrient additives on the yield and viability of M. anisopliae spore and to determine the optimum incubation period for maximum spore production.@*Methodology and results@# In this study, M. anisopliae was cultivated by solid-state fermentation using rice as a growth medium. Three different nutrient additives were examined which aimed to maximize the production of M. anisopliae spores. Among the three nutrient additives evaluated, yeast (1.84 ± 0.04 g) supported better growth and spore production than molasses (0.58 ± 0.04 g) and palm oil (0.47 ± 0.09 g). The incubation period between 2-6 weeks produced higher spore yield (0.97 ± 0.02 g spores) at week 4 with a better spore viability (86.30 ± 0.45%) at week 2. @*Conclusion, significance and impact of study@#Hence, it is suggested that the optimum incubation period is between 2 and 6 weeks after inoculation, and M. anisopliae could be mass produced in large quantities on rice substrate with the addition of yeast as the nutrient additives.

Awareness and precaution attitude of dentists as regards to risks associated with exposure to COVID-19 / Conscientização e atitude de precaução dos dentistas em relação aos riscos associados à exposição a COVID-19

Objective: Dentists need a high level of awareness to limit the spread of COVID-19 (Coronavirus disease 2019). This study aimed to evaluate the level of awareness and attitude regarding the risk associated with dental procedures among dentists. Material and Methods: An online questionnaire was submitted to dentists between April- May 2020. The questionnaire form included questions related to demographic data, the transmission characterization of SARS-CoV-2, and treatment of COVID-19 patients. Data were analyzed using IBM SPSS V23 and chi-square test (p ≤ 0.05). Results: A total of 3825 participants (29.1± 7.6 years) were included. In COVID-19, the riskiest dental branch in terms of the risk of contamination through saliva was considered to be Periodontics (32.2%), while the least risky branch was Orthodontics (0.2%). Specific dental treatment procedures considered at high risk of contamination were tooth preparation (69.4%), scaling and root planing (63.5%), filling (53.4%), and pulpectomy (40.5%). The parameters of the study that differed according to gender and professional status were the viability of the virus, the risk assessment, saliva contamination risk, and aerosol-generating activities for COVID-19 (p < 0.05). Conclusion: Dentists were aware of the risk assessment and extra precautionary methods. However, they had limited knowledge about the viability of the virus. Dentists should be aware of recommended approaches and update their knowledge about COVID-19 to limit the spread of the disease. Since dentistry is an area suitable for the transmission of the COVID-19, the fact that dentists have information about the viability of this virus will be lifesaving in clinical applications (AU)

Objetivo: Os dentistas precisam de um alto nível de conhecimento para limitar a disseminação de COVID-19 (doença por coronavírus 2019). Este estudo teve como objetivo avaliar o nível de consciência e atitude em relação ao risco associado aos procedimentos odontológicos entre os dentistas. Material e Métodos: Um questionário online foi submetido aos dentistas entre abril e maio de 2020. O formulário do questionário incluía questões relacionadas aos dados demográficos, à caracterização da transmissão da SARS-CoV-2 e ao tratamento de pacientes com COVID-19. Os dados foram analisados usando IBM SPSS V23 e teste do qui-quadrado (p ≤ 0,05). Resultados:Um total de 3825 participantes (29,1 ± 7,6 anos) foram incluídos. Na COVID-19, o ramo odontológico de maior risco em termos de risco de contaminação pela saliva foi considerado a Periodontia (32,2%), enquanto o ramo de menor risco foi a Ortodontia (0,2%). Os procedimentos odontológicos específicos considerados de alto risco de contaminação foram preparo dentário (69,4%), raspagem e alisamento radicular (63,5%), obturação (53,4%) e pulpectomia (40,5%). Os parâmetros do estudo que diferiram de acordo com sexo e status profissional foram a viabilidade do vírus, a avaliação de risco, risco de contaminação da saliva e atividades geradoras de aerossol para COVID-19 (p <0,05). Conclusão: Os dentistas estão cientes da avaliação de risco e dos métodos de precaução extra. No entanto, eles tinham conhecimento limitado sobre a viabilidade do vírus. Os dentistas devem estar cientes das abordagens recomendadas e atualizar seus conhecimentos sobre COVID-19 para limitar a propagação da doença. Como a odontologia é uma área propícia para a transmissão da COVID-19, se dentistas tiverem informações sobre a viabilidade desse vírus poderão salvar vidas em aplicações clínicas. (AU)

SARS-CoV-2 em superfícies: persistência e medidas preventivas: uma revisão sistemática / SARS-CoV-2 in Surfaces: persistence and prevention measures: a systematic review / SARS-CoV-2 en Superficies: persistencia y medidas preventivas: una revisión sistemática

Objetivo: verificar a persistência do SARS-CoV-2 nas diferentes superfícies e medidas preventivas contra a transmissão do vírus. Método: revisão sistemática norteada pelo método PRISMA. Foram utilizadas as bases de buscas PubMed e LILACS de janeiro a junho de 2020, com os descritores: "2019-nCOV" OR "SARS-CoV-2" OR "COVID-19" AND "transmission" OR "transmission route" AND "viability" AND "surface" OR "inanimate surface" AND "prevention". As informações extraídas foram autor/ano, país, tipo de publicação, nome da revista, idioma, país da publicação e base de dados. Resultados: foram identificadas 178 publicações, com exclusão de 164 artigos, nove por idioma, 12 por outras doenças e/ou patógenos e 143 pelo título e/ou resumo. Foram incluídos 14 artigos qualitativos, oito artigos de revisões narrativas, uma comunicação breve, dois artigos originais e um editorial. Treze artigos foram publicados em inglês e um em português. Conclusão: coronavírus humanos (HCoV 229E) podem se manter em diferentes superfícies durante duas horas até nove dias. Baixas temperaturas e reduzida umidade relativa do ar favorecem a sobrevivência do SARS-CoV-2, sendo mais estável em plásticos e aço inoxidável do que em cobre e papelão. A recomendação é higienização de superfícies e mãos com água, sabão ou higienizadores à base de álcool.(AU)

Objective: to verify the persistence of SARS-CoV-2 on different types of surfaces and the preventive measures against the transmission of the virus. Method: a systematic review was carried out, using the PRISMA method. The PubMed and LILACS databases from January to June 2020 were used, with the following descriptors: "2019-nCOV" OR "SARS-CoV-2" OR "COVID-19" AND "transmission" OR "transmission route" AND "viability" AND "surface" OR "inanimate surface" AND "prevention". Information extracted was author/year, country, type of publication, journal name, language, country of publication and database. Results: 178 publications were identified. 164 articles were excluded, nine by language, 12 by other diseases and/or pathogens and 143 by title and/or abstract. 14 qualitative articles were included, eight articles of narrative reviews, one short communication, two original articles and one editorial. Thirteen articles were published in English and one in Portuguese. Conclusion: human coronaviruses (HCoV 229E) can persist on different surfaces for two hours up to nine days. Low temperatures and low relative humidity of the air favor the survival of SARS-CoV-2, which is more stable on plastics and on stainless steel than on copper and cardboard. The recommendation is frequent surface and hand hygiene with water, soap or alcohol-based rubs.(AU)

Objetivo: verificar la persistencia del SARS-CoV-2 en diferentes superficies y las medidas preventivas contra la transmisión del virus. Método: se realizó una revisión sistemática, utilizando el método PRISMA. Se utilizaron las bases de datos de búsqueda de PubMed y LILACS de enero a junio de 2020, con los descriptores: "2019-nCOV" O "SARS-CoV-2" O "COVID-19" Y "transmisión" O "ruta de transmisión" Y "viabilidad" Y "superficie" O "superficie inanimada" Y "prevención". Las informaciones extraídas fueron autor / año, país, tipo de publicación, nombre de la revista, idioma, país de publicación y base de datos. Resultados: se identificaron 178 publicaciones. Se excluyeron 164 artículos, nueve por idioma, 12 por otras enfermedades y/o patógenos y 143 por título y/o resumen, incluidos 14 artículos cualitativos, ocho artículos de revisiones narrativas, una comunicación breve, dos artículos originales y uno editorial. Se publicaron trece artículos en inglés y uno en portugués. Conclusión: los coronavirus humanos (HCoV 229E) pueden matenerse en diferentes superficies durante dos horas hasta nueve días. Las bajas temperaturas y la reducida humedad relativa del aire favorecen la supervivencia del SARS-CoV-2, siendo más estable en plásticos y acero inoxidable que en cobre y carton. La recomendación es limpiar superficies y manos con agua, jabón o limpiadores a base de alcohol.(AU)

Development of a new protocol for freeze-drying preservation of Pseudoalteromonas nigrifaciens and its protective effect on other marine bacteria

Background: Freeze-drying is known as one of the best methods to preserve bacterial strains. Protectant is the key factor affecting the survival rate of freeze-dried strains. In addition, salinity, bacterial suspension concentration, drying time, and other factors can also affect the survival rate of strains to varying degrees. At present, there are relatively few studies on freeze-drying preservation of marine bacteria. In the present study, we performed the freeze-drying protectant screening and optimized the preservation conditions for Pseudoalteromonas nigrifaciens, which is widely distributed in marine environment. The protective effects of the screened protectants were verified by 18 other marine bacterial strains. Results: The results indicated that the combination of 5.0% (w/v) lactose, 5.0% (w/v) mannitol, 5.0% (w/v) trehalose, 10.0% (w/v) skim milk powder, 0.5% (w/v) ascorbic acid and 0.5% (w/v) gelatin was the best choice for the preservation of P. nigrifaciens. The suggested salinity and concentration of initial cell suspension were 10 g/L NaCl and 1.0 × 109 CFU/mL, respectively. Furthermore, stationary-phase cells were the best choice for the freeze-drying process. The highest survival rate of P. nigrifaciens reached 52.8% when using 5­10% (w/v) skim milk as rehydration medium. Moreover, the other 18 marine strains belonging to Pseudoalteromonas, Vibrio, Photobacterium, Planomicrobium, Edwardsiella, Enterococcus, Bacillus, and Saccharomyces were freezedried under the abovementioned conditions. Their survival rates were 2.3­95.1%. Conclusion: Collectively, our results supported that the protectant mixture and parameters were beneficial for lyophilization of marine bacteria

Effects of microbial conditioning and temperature on the leaf-litter shredding activity of Phylloicussp

Few studies try to explain the effects in tropical lotic ecosystems of an increase in water temperature on the shredding activity of invertebrate shredders, particularly in association with the quality of the leaf litter and the degree of litter conditioning. Therefore, the aims of this study were as follows: i) to better understand how this key invertebrate shredder group affects the decomposition of different species of leaf litter under gradual increases in temperature and microbial conditioning; and ii) to verify the possible consequences on leaf mass loss (LML). Three species of leaf litter were used in two experiments. Inexperiment I, the litters of three species (Protium spruceanum, Richeria grandisand Ingalaurina) at three conditioning levels (1, 7, 14 days) were tested under five different temperatures (20, 22, 24, 26 and28°C). In experiment II, the leaf litters of three species were used, without conditioning, under four temperatures (20, 22, 26 and27°C). The shredding performed by Phylloicussp. was largely dependent on the lignin and cellulose concentrations in each leaf species, independent of conditioning. The presence or absence of conditioning may cause the shredders to use different energy compensation strategies in response to the temperature increases.

Survival of microencapsulated Lactobacillus casei (prepared by vibration technology) in fruit juice during cold storage

Background: Foods including probiotics are considered "functional foods." As an alternative to dairy products, we investigated the behavior of Lactobacillus casei when exposed to low-pH fruit juice. Juices of fruits such as pineapple, raspberry, and orange were assessed. Free and microencapsulated forms of L. casei were compared, and the viability of the probiotic was evaluated under storage at 4°C for 28 d. Microbiological analyses were carried out to ensure a safe and healthy product for consumers who look for foods with probiotics from sources other than dairy. Results: Low pH affected L. casei survival during storage depending on the type of fruit juice. In the case of pineapple juice, some microcapsules were broken, but microcapsules recovered at the end of the storage period had 100% viability (2.3 × 107 CFU/g spheres). In the case of orange juice, more than 91% viability (5.5 × 106 CFU/g spheres) was found. In raspberry juice, viability decreased rapidly, disappearing at the end of the storage period, which was caused by the absorption of high concentrations of anthocyanin inside microcapsules more than low pH. Conclusion: Low pH affected the survival of L. casei under refrigeration; even when they were microencapsulated, acidic conditions impacted their viability. Although pH affects viability, its value is very sensitive and will depend on the type of fruit juice and its composition. Some fruit juices contain compounds used as substrates for Lactobacillus and other compounds with antimicrobial effects.

Evaluación de la viabilidad de Legionella pneumophila en muestras de agua / Evaluation of the viability of Legionella pneumophila in water samples

Introducción: Legionella pneumophila se sitúa entre los principales agentes causales de neumonía adquirida en la comunidad y de origen nosocomial. La inhalación de aerosoles potencialmente contaminados con la bacteria, producto de la colonización de redes y otros sistemas que utilizan agua, representa un peligro para la salud de los individuos expuestos. Objetivo: evaluar la viabilidad de L. pneumophila en muestras de agua almacenadas en diferentes intervalos de tiempo para el diagnóstico por cultivo microbiológico de Legionella spp. Métodos: Se contaminaron artificialmente muestras de agua con dos cepas de L. pneumophila de serogrupos diferentes y la conformación de una mezcla de ellas, para un total de 15 muestras. Los frascos contaminados fueron procesados a las 24 h, 72 h, 7 días, 14 días y 21 días. Se realizó cultivo microbiológico según ISO 11731: 2004 y PNO 03-013: 2015. Resultados: Se demostró viabilidad de la bacteria en muestras almacenadas hasta 21 días. El método de concentración por filtración resultó tener los mayores recobrados del microorganismo. Conclusiones: El tiempo de almacenamiento de las muestras afecta la viabilidad de L. pneumophila. Sienta las bases para estudios posteriores de robustez del diagnóstico de L. pneumophila como parte del servicio que presta el Centro de Investigaciones Científicas de la Defensa Civil en los programas de prevención y control Legionella spp. en instalaciones de interés turístico e industrial(AU)

Introduction: Legionella pneumophila is one of the main causative agents of community- and hospital-acquired pneumonia. Inhalation of sprays potentially contaminated with the bacterium, due to the colonization of networks and other systems using water, is a hazard to the health of exposed individuals. Objective: Evaluate the viability of L. pneumophila in samples of water stored at various time intervals for the microbiological culture diagnosis of Legionella spp. Methods: Water samples were artificially contaminated with two strains of L. pneumophila from different serogroups and a mixture of them, for a total of 15 samples. The contaminated vessels were processed at 24 h, 72 h, 7 d, 14 d and 21 d. Microbiological culture was performed in compliance with ISO 11731: 2004 and PNO 03-013: 2015. Results: The bacterium was found to be viable in samples stored up to 21 days. The filtration concentration method obtained the greatest amount of the microorganism. Conclusions: Storage time of the samples affects the viability of L. pneumophila. The study lays the foundations for further research about the validity of L. pneumophila diagnosis as part of the service offered by the Civil Defense Scientific Research Center in Legionella spp. prevention and control programs for tourist and industrial facilities(AU)

Viabilidade de culturas probióticas em queijos tipo Minas Frescal / Viability of probiotic cultures in cheese Minas Frescal type

Os consumidores estão mais exigentes com a qualidade dos alimentos, e diante deste cenário, a adição de bactérias probióticas com apelo na saúde é uma tendência promissora, principalmente em produtos lácteos como queijo Minas Frescal. Objetivou-se verificar a viabilidade da adição de L. acidophilus LA-5 e B. animalis subsp. lactis BB-12 em queijo Minas Frescal, sob refrigeração a 4ºC, no 0, 7, 14 e 21 dias de armazenamento. As amostras foram inoculadas em ágar MRS e incubadas em anaerobiose a 35°C ± 2ºC por 48 horas. A população de células viáveis de bactérias probióticas se manteve nos níveis mínimos exigido pela legislação (10(8)-10(9) UFC g-1) durante todo o tempo de prateleira, o que evidencia a qualidade do queijo Minas Frescal como matriz alimentícia adequada e como um alimento potencialmente probiótico.

The bactericidal effect of an atmospheric-pressure plasma jet on Porphyromonas gingivalis biofilms on sandblasted and acid-etched titanium discs

PURPOSE: Direct application of atmospheric-pressure plasma jets (APPJs) has been established as an effective method of microbial decontamination. This study aimed to investigate the bactericidal effect of direct application of an APPJ using helium gas (He-APPJ) on Porphyromonas gingivalis biofilms on sandblasted and acid-etched (SLA) titanium discs. METHODS: On the SLA discs covered by P. gingivalis biofilms, an APPJ with helium (He) as a discharge gas was applied at 3 different time intervals (0, 3, and 5 minutes). To evaluate the effect of the plasma itself, the He gas–only group was used as the control group. The bactericidal effect of the He-APPJ was determined by the number of colony-forming units. Bacterial viability was observed by confocal laser scanning microscopy (CLSM), and bacterial morphology was examined by scanning electron microscopy (SEM). RESULTS: As the plasma treatment time increased, the amount of P. gingivalis decreased, and the difference was statistically significant. In the SEM images, compared to the control group, the bacterial biofilm structure on SLA discs treated by the He-APPJ for more than 3 minutes was destroyed. In addition, the CLSM images showed consistent results. Even in sites distant from the area of direct He-APPJ exposure, decontamination effects were observed in both SEM and CLSM images. CONCLUSIONS: He-APPJ application was effective in removing P. gingivalis biofilm on SLA titanium discs in an in vitro experiment.

Comparison between static and semi-dynamic models for microcosm biofilm formation on dentin

Abstract Microcosm biofilm has been applied to induce carious lesions in dentin. However, no study has been done to compare the impact of the type of model for providing nutrients to microcosm biofilm formation on dentin. Objective This study compared the performance of two kinds of models (static and semi-dynamic) on the biofilm formation and the development of dentin carious lesions. Material and Methods In both models, biofilm was produced using inoculum from pooled human saliva mixed with McBain saliva for the first 8 h (5% CO2 and 37°C). Afterwards, for the static model, the samples were placed in 24-wells microplate containing McBain saliva with 0.2% sucrose, which was replaced at 24 h. In the semi-dynamic model, the samples were submitted to artificial mouth system with continuous flow of McBain saliva with 0.2% sucrose (0.15 ml/min, 37°C) for 10 h a day (for the other 14 h, no flow was applied, similarly to the static model). After 5 days, biofilm viability was measured by fluorescence and dentin demineralization by transverse microradiography. Results Biofilm viability was significantly lower for the static compared with semi-dynamic model, while dentin demineralization was significantly higher for the first one (p<0.05). The static model was able to produce a higher number of typical subsurface lesions compared with the semi-dynamic model (p<0.05). Conclusions The type of model (static and semi-dynamic) applied in the microcosm biofilm may have influence on it's viability and the severity/profile of dentin carious lesions.

Effect of hydroalcoholic extract of Myracrodruon urundeuva All. and Qualea grandiflora Mart. leaves on the viability and activity of microcosm biofilm and on enamel demineralization

Abstract Objectives: The aim of this study was to assess the effect of Myracrodruon urundeuva All. and Qualea grandiflora Mart. leaves hydroalcoholic extracts on viability and metabolism of a microcosm biofilm and on enamel demineralization prevention. Methodology: Microcosm biofilm was produced on bovine enamel using inoculum from pooled human saliva mixed with McBain saliva, under 0.2% sucrose exposure, for 14 days. The biofilm was daily-treated with the extracts for 1 min. At the end, it was analyzed with respect to viability by fluorescence, CFU counting and extracellular polysaccharides (phenol-sulphuric acid colorimetric assay) and lactic acid (enzymatic assay) production. The demineralization was measured by TMR. The data were compared using ANOVA or Kruskal-Wallis (p<0.05). Results: M. urundeuva All. at 100, 10 and 0.1 μg/mL and Q. grandiflora Mart. at 100 and 0.1 μg/mL reduced biofilm viability similarly to positive control (chlorhexidine) and significantly more than the negative-vehicle control (35% ethanol). M. urundeuva at 1000, 100 and 0.1 μg/mL were able to reduce both lactobacilli and mutans streptococci CFU counting, while Q. grandiflora (1000 and 1.0 μg/mL) significantly reduced mutans streptococci CFU counting. On the other hand, the natural extracts were unable to significantly reduce extracellular polysaccharides and lactic acid productions neither the development of enamel carious lesions. Conclusions: The extracts showed antimicrobial properties on microcosm biofilm, however, they had no effect on biofilm metabolism and caries protection.

Survival of Salmonella spp. in minced meat packaged under vacuum and modified atmosphere

Abstract The effect of different modified atmosphere packaging regimes on the behavior of Salmonella spp. on minced meat was studied. Minced meat was experimentally contaminated with a Salmonella spp. cocktail (S. Enteritidis, S. Typhimurium, S. Infantis and S. Arizonae), packaged under vacuum or modified atmosphere with initial headspaces containing 20%O2/50%CO2/30%N2 and 20%O2/30%CO2/50%N2) and stored at 3 ± 1 °C for 12 days. Samples were analyzed for Salmonella spp., viable and lactic acid bacteria count every third day. Salmonella spp. counts decreased during storage in all packaging types, with reductions of about 1.5 log CFU/g. A significant difference (p < 0.01) was noted between Salmonella spp. counts in meat packaged in vacuum and modified atmospheres, although there was no significant difference in Salmonella spp. count between meat packaged in 50%CO2, and meat packaged in 30%CO2. At the end of the study, there were significant differences (p < 0.01; p < 0.05) in total viable and lactic acid bacterial counts between meat packaged in vacuum and modified atmosphere, and the lowest counts were noted in meat packaged in modified atmosphere with 50%CO2.

Cryopreservation at -75 °C of Agaricus subrufescens on wheat grains with sucrose

Abstract Agaricus subrufescens is a basidiomycete which is studied because of its medicinal and gastronomic importance; however, less attention has been paid to its preservation. This study aimed to evaluate the effect of sucrose addition to substrate and cryotube on the viability of Agaricus subrufescens cryopreserved at -20 °C and at -75 °C for one and two years. Zero, 10% or 20% sucrose was added to potato dextrose agar or wheat grain. The mycelia were cryopreserved in the absence of cryoprotectant or with sucrose solutions at 15%, 30% or 45%. After one or two years at -75 °C or at -20 °C, mycelia were thawed and evaluated about viability, initial time of growth, colony diameter and genomic stability. Cryopreservation at -20 °C is not effective to keep mycelial viability of this fungus. Cryopreservation at -75 °C is effective when sucrose is used in substrates and/or cryotubes. Without sucrose, cryopreservation at -75 °C is effective only when wheat grains are used. Physiological characteristic as mycelial colony diameter is negatively affected when potato dextrose agar is used and unaffected when wheat grain is used after two-year cryopreservation at -75 °C. The fungus genome does not show alteration after two-year cryopreservation at -75 °C.

Long-term cryopreservation of basidiomycetes

Abstract Basidiomycetes have several biotechnological and industrial applications such as enzyme production, bioremediation, pharmaceutical and functional food production. Due to climatic features, the preservation of several basidiomycetes is threatened, and to guarantee the preservation of this genetic resource, the development of long-term preservation techniques is necessary once there is no universal protocol for the cryopreservation of basidiomycetes. Cryopreservation is a technique in which microorganisms are submitted to ultralow temperatures. Therefore, this study aimed to collect information on the main conditions for long-term cryopreservation of basidiomycetes in the last 20 years. Scientific articles on cryopreservation of basidiomycetes published from 1997 to 2016, were researched, and only the studies on two intervals of cryopreservation were considered: from 1 to 2 years and for longer than 2 years. The analyzed conditions of basidiomycete cryopreservation were: most studied genera, cryopreservation temperature, substrate, cryoprotectant (and preservation substrate), cryopreservation period, thawing temperature and cultivation medium after thawing, physiological and genetic stability of basidiomycetes after thawing in cryopreservation. In this review, the viability of the main cryopreservation conditions of basidiomycetes studied in the last 20 years are presented and discussed.

Biofilm formation by Staphylococcus aureus and Salmonella spp. under mono and dual-species conditions and their sensitivity to cetrimonium bromide, peracetic acid and sodium hypochlorite

Abstract The aim of this study was evaluated the biofilm formation by Staphylococcus aureus 4E and Salmonella spp. under mono and dual-species biofilms, onto stainless steel 316 (SS) and polypropylene B (PP), and their sensitivity to cetrimonium bromide, peracetic acid and sodium hypochlorite. The biofilms were developed by immersion of the surfaces in TSB by 10 d at 37 °C. The results showed that in monospecies biofilms the type of surface not affected the cellular density (p > 0.05). However, in dual-species biofilms on PP the adhesion of Salmonella spp. was favored, 7.61 ± 0.13 Log10 CFU/cm2, compared with monospecies biofilms onto the same surface, 5.91 ± 0.44 Log10 CFU/cm2 (p < 0.05). The mono and dual-species biofilms were subjected to disinfection treatments; and the most effective disinfectant was peracetic acid (3500 ppm), reducing by more than 5 Log10 CFU/cm2, while the least effective was cetrimonium bromide. In addition, S. aureus 4E and Salmonella spp. were more resistant to the disinfectants in mono than in dual-species biofilms (p < 0.05). Therefore, the interspecies interactions between S. aureus 4E and Salmonella spp. had a negative effect on the antimicrobial resistance of each microorganism, compared with the monospecies biofilms.

Development and characterization of an innovative synbiotic fermented beverage based on vegetable soybean

Abstract Soymilk was produced from vegetable soybean and fermented by probiotics (Lactobacillus acidophilus La-5, Bifidobacterium animalis Bb-12) in co-culture with Streptococcus thermophilus. The composition of the fermented beverage and oligosaccharides content were determined. The effect of fructooligosaccharides and inulin on the fermentation time and viability of probiotic microorganisms throughout 28 days of storage at 5 °C were evaluated. The soymilk from vegetable soybeans was fermented in just 3.2 h, when pH reached 4.8. Fermentation reduced the contents of stachyose and raffinose in soymilk. Prebiotics had no effect on acidification rate and on viability of B. animalis and S. thermophilus in the fermented beverage. The viable counts of B. animalis Bb-12 remained above 108 CFU mL-1 in the fermented soymilk during 28 days of storage at 5 °C while L. acidophilus La-5 was decreased by 1 log CFU mL-1. The fermented soymilk from vegetable soybeans showed to be a good food matrix to deliver probiotic bacteria, as well as a soy product with a lower content of non-digestible oligosaccharides.

Effect of storage temperature on Streptococcus mutans viability / Influência da temperatura de armazenamento na viabilidade de Streptococcus mutans

Introduction: Proper storage conditions and maintenance of viable biological material plays an important role in microbiological research, allowing for the opportunity to conduct future studies. Objective: To evaluate the viability of Streptococcus mutans strains that were previously grown and stored under different temperatures for approximately eight years. Material and method: In this study, we evaluated 393 bacterial isolates that were stored in a freezer at -80°C (G1) and 200 isolates stored in a freezer at -20°C (G2). Aliquots of each sample were plated on blood agar and mitis-salivarius bacitracin sucrose agar-solidified medium. After incubating under microaerophilic conditions in an incubator at 37°C for 72 hours, the presence, morphology and purity of bacterial growth was observed. The data were analyzed by means of descriptive statistics. Result: Microbial viability was observed in almost all samples (99.7%) in G1, whereas all isolates stored at -20°C were considered inviable. Conclusion: The viability of S. mutans is influenced by the storage temperature of the samples, and the strains remain viable when stored under ideal temperature conditions (-80°C), even when stored for a long period of time.

Introdução: O correto armazenamento e manutenção adequada de material biológico viável representam importante papel nas pesquisas microbiológicas em virtude da oportunidade de desenvolvimento de pesquisas futuras. Objetivo: Avaliar a viabilidade de Streptococcus mutans (S. mutans ) previamente cultivados e armazenados sob diferentes temperaturas há aproximadamente oito anos. Material e método: Foram avaliados 393 isolados bacterianos armazenados em freezer a -80°C (G1) e 200 em freezer a -20°C (G2). Alíquotas de cada amostra foram semeadas em meios de cultura ágar sangue e ágar Mitis Salivarius Bacitracina Sacarose. Após incubação sob condições de microaerofilia em estufa a 37°C, durante 72 horas, observou-se: presença, morfologia e pureza do crescimento bacteriano. Os dados obtidos foram analisados por meio de estatística descritiva. Resultado: A viabilidade microbiana foi observada em praticamente todas as amostras (99,7%) em G1. Por outro lado, todas aquelas estocadas a -20°C foram consideradas inviáveis. Conclusão: A viabilidade de S. mutans é influenciada pela temperatura de armazenamento das amostras, sendo que as cepas permanecem viáveis quando estocadas em condições ideais de temperatura (-80°C), mesmo quando armazenadas por longo período de tempo.

Removal and killing of multispecies endodontic biofilms by N-acetylcysteine

ABSTRACT Removal of bacterial biofilm from the root canal system is essential for the management of endodontic disease. Here we evaluated the antibacterial effect of N-acetylcysteine (NAC), a potent antioxidant and mucolytic agent, against mature multispecies endodontic biofilms consisting of Actinomyces naeslundii, Lactobacillus salivarius, Streptococcus mutans and Enterococcus faecalis on sterile human dentin blocks. The biofilms were exposed to NAC (25, 50 and 100 mg/mL), saturated calcium hydroxide or 2% chlorhexidine solution for 7 days, then examined by scanning electron microscopy. The biofilm viability was measured by viable cell counts and ATP-bioluminescence assay. NAC showed greater efficacy in biofilm cell removal and killing than the other root canal medicaments. Furthermore, 100 mg/mL NAC disrupted the mature multispecies endodontic biofilms completely. These results demonstrate the potential use of NAC in root canal treatment.

Encapsulation of specific Salmonella Enteritidis phage f3αSE on alginate-spheres as a method for protection and dosification

Background: Bacteriophages have been proposed as an alternative to control pathogenic bacteria resistant to antibiotics. However, they are not extensively used due to different factors such as vulnerability under environmental conditions and the lack of efficient administration methods. A potential solution is the encapsulation of bacteriophages in hydrogel polymers to increase their viability and as a controlled release method. This work describes the use of alginate-Ca+2 matrixes as mechanisms for protection and dosification of the phage f3αSE which has been successfully used to prevent infections produced by Salmonella Enteritidis. Results: The viability of the pure phage is reduced in near 100% after 1-h incubation at pH 2 or 3. However, the encapsulated phage remains active in 80, 6% at pH 3, while no differences were observed at pH 2, 4 or 7. Exposition of f3αSE to different T° showed that the viability of this phage decreased with increased T° to near 15% at 60°C, while the encapsulated phage remains with 50% viability at same temperature. Finally, the encapsulation of phages showed to extend their presence for 100 h in the medium compared to non-encapsulated phages in a water flow system, which simulate automatic birdbath used in poultry industry, maintaining the phage concentration between 102 and 104 PFU/mL during 250 h. Conclusions: Encapsulation in alginate-Ca+2 spheres can be a good alternative to extend viability of phages and can be used as a phage method dosification method in water flow systems.