Two Methods of Measuring Cryptococcus neoformans Fungal Burden in Macrophages.

The ability of C. neoformans to survive and replicate within host phagocytes enables it to evade the immune system and allows for persistence of the infection. As such, measuring fungal burden of C. neoformans strains-and indeed how drug treatments can influence fungal burden-provides important information about C. neoformans pathogenesis. In this chapter, we describe two methods that may be used to appraise fungal burden: a standard end-point colony-formation assay for calculating the average number of yeast per host cell and a fluorescence microscopy-based method that may be used to measure changes in fungal burden in individual living macrophages in real time.

Evaluación antifúngica del ozono contra Candida albicans: estudio in vitro / Antifungal evaluation of ozone against Candida albicans: in vitro study

Introducción: la Candida albicans (C. albicans) es un patógeno fúngico que puede causar infecciones superficiales o potencialmente mortales. Los biofilms de C. albicans muestran rasgos fenotípicos únicos, el más destacado es su notable resistencia a una amplia variedad de agentes antimicóticos. Una de las alternativas para inhibir el crecimiento de este microorganismo es el ozono debido a sus propiedades bactericidas, fungicidas y virucidas; sin embargo, escasa información ha sido reportada en C. albicans. Objetivo: el objetivo de este estudio fue evaluar el efecto fungicida del ozono en C. albicans. Material y métodos: la metodología consistió en agregar ozono a tubos de ensayo con medios de caldo nutritivo en diversas concentraciones y tiempos de ozonización. El efecto fungicida fue determinado con la determinación del número de colonias de C. albicans en agar nutritivo a través de procedimiento microbiológicos estandarizados por triplicado. Resultados: todas las muestras con ozono mostraron adecuados niveles de inhibición de crecimiento del microorganismo. Además, el efecto fungicida del ozono se encontró para ser significativamente dependiente del tiempo de ozonización y de la concentración. Conclusión: el uso de terapia con ozono podría tener potencial en el control de infecciones micóticas causadas por la presencia de C. albicans (AU)

Introduction: Candida albicans (C. albicans) is a fungal pathogen that can cause superficial or life-threatening infections. Biofilms of C. albicans display unique phenotypic traits, the most prominent being their remarkable resistance to a wide variety of antifungal agents. One of the alternatives to inhibit the growth of this microorganism is ozone due to its bactericidal, fungicidal and virucidal properties; however, little information has been reported on C. albicans. Objective: the objective of this study was to evaluate the fungicidal effect of ozone on C. albicans. Material and methods: the methodology consisted in adding ozone to test tubes with nutrient broth media in various concentrations and ozonation times. The fungicidal effect was determined by determining the number of colonies of C. albicans in nutrient agar through standardized microbiological procedures in triplicate. Results: all the ozone samples showed adequate levels of growth inhibition of the microorganism. Furthermore, the fungicidal effect of ozone was found to be significantly dependent on ozonation time and concentration. Conclusion: the use of ozone therapy could have potential in the control of fungal infections caused by the presence of C. albicans (AU)

Análisis microbiológico a corto y largo plazo del material usado para esterilizar instrumental odontológico / Short and long-term microbiological analysis of the material for sterilize dental instruments

Introducción: el material para empaquetar el instrumental odontológico, como pueden ser bolsas de tela, papel o plástico, es usado por profesionales de la salud; sin embargo, es necesario esclarecer la efectividad de cada uno y determinar el tiempo que permanece estéril luego del procedimiento. Objetivo: identificar la eficacia de tela, plástico y papel como materiales para esterilizar instrumental a corto y largo plazo. Material y métodos: se realizaron cultivos sólidos y líquidos de instrumental esterilizado en tres materiales y con diferentes tiempos de postesterilización. Se incubaron a 36 oC por 72 horas en condiciones aerobias y anaerobias. Los resultados se analizaron usando una prueba de Kruskal-Wallis, seguida de una prueba de Dunn. Resultados: los resultados mostraron que inmediatamente después del proceso de esterilización, los tres materiales son efectivos (Kruskal-Wallis test, p = 0.2752), 24 horas (p = 0.2492), siete (p = 0.0509) y 14 días (p = 0.0006). Veinticuatro horas posterior a la esterilización la tela no es efectiva, el plástico disminuye su efectividad y el papel sigue siendo efectivo. Conclusión: en nuestros resultados, el papel es la mejor opción para esterilizar instrumental (AU)

Introduction: material such as cloth, paper or plastic bags to wrap dental instruments is used by health professionals, however, it is necessary to clarify the effectiveness of each one and determine if it remains sterile after the procedure. Objective: to determine the effectiveness of cloth, plastic and paper as materials to sterilize dental instruments in the short and long term. Material and methods: we carry out solid and liquid cultures of sterilized instruments in three materials, at different post-sterilization times, incubated at 36 oC for 72 hours under aerobic and anaerobic conditions, and the results were analyzed using a Kruskal-Wallis test, followed by from a Dunn's test. Results: our results showed that immediately after the sterilization process the three materials are effective (Kruskal-Wallis; p = 0.2752), 24 hours (p = 0.2492), 7 (p = 0.0509) and 14 (p = 0.0006) days. Twenty-four hours after the cloth is not effective, plastic decreases its effectiveness and paper remain effective. Conclusion: in our results, paper is the best option to sterilize dental instruments (AU)

Estudio de supervivencia de Streptococcus mutans en un tipo de fomite / Survival study of Streptococcus mutans in a type of fomite

Objetivo: Evaluar la supervivencia de Streptococcus mutans (S.mutans)en un tipo de fómite. Método: Se reactivó una cepa de S.mutans ATCC25175 criopre-servada en agar TYCSB. El inóculo se estandarizó en PBS buffer hasta obtener turbidez equivalente al 0,5 de Mc Farland y un OD = 0.01 por espectrofotome-tría. Bloques plásticos de 2cm2/superficie fueron seleccionados como fómites. La descontaminación de los bloques se realizó por inmersión en alcohol etílico 70% v/v durante 10 minutos, los que fueron secados en cabina de seguridad biológica. La conta-minación de los mismos se realizó por inmersión en inóculo estandarizado durante 10 minutos. Los blo-ques contaminados se extrajeron y depositaron so-bre placas de Petri estériles hasta cumplir los tiem-pos propuestos (T0-T4 con intervalos de 30 minutos). A cada tiempo, los bloques fueron eluidos en 20 ml de buffer PBS y agitados en vortex durante 30 segun-dos. 100 µl de cada eluato fueron sembrados por dis-persión en agar TYCSB e incubados en anaerobiosis por 48 horas a 37°C. El recuento de colonias (UFC/ml) se realizó bajo lupa estereoscópica 50X. Resulta-dos: El recuento inicial de S.mutans fue de 7,8 X 106(DS+1,7 X 106) UFC/ml y para cada tiempo de estu-dio fue de: T0=3.25 X 104 (DS+1.9 X 103); T1=2.63X104 (DS+4,50E+03); T2= 1.85 X 104 (DS+9,45E+02); T3=1.93 X103(DS+1,29E+03) y T4=1.2X103 (DS+7,21x102). Conclusión: En los rangos de tiempos establecidos, la cepa de S.mutans ensayada permaneció viable sobre la superficie plástica (AU)

Aim: To evaluate the survival time of Streptococcus mutans (S.mutans) in a type of fomites. Method: A strain of cryopreserved S.mutans ATCC 25175 was reactivated in TYCSB agar. The inoculum was standardized in the PBS buffer to obtain turbidity equivalent to 0.5 Mc Farland and OD = 0.01 by spectrophotometry. Plastic blocks of 2 cm2 /surface were selected as fomites. Decontamination of the blocks was carried out for 10 minutes by immersion in ethyl alcohol 70% v/v, which were dried in a biosafety chamber. Contamination was carried out by immersion in standardized inoculum for 10 minutes. The contaminated blocks were extracted and put on sterile Petri dishes until the proposed times were met (T0-T4 at 30-minute intervals). At each time, the blocks were eluted in 20 ml of PBS buffer and vortexed for 30 seconds. 100 µl of each eluate were dispersed on TYCSB agar and incubated anaerobically for 48 hours at 37°C. Colony count (CFU/ml) was performed under a 50X stereoscopic magnifying glass. Results: The initial S.mutans count was 7,8 X 106 (DS+1,7 X 106) CFU/ml and for each study time was: T0=3.25 X 104 (DS+1.9 X 103); T1=2.63X104 (DS+4,50E+03); T2= 1.85 X 104 (DS+9,45E+02); T3=1.93 X103(DS+1,29E+03) y T4=1.2X103 (DS+7,21x102). Conclusion: Within the established time ranges, the tested S.mutans strain remained viable on the plastic surface (AU))

Large-scale comparison of E. coli levels determined by culture and a qPCR method (EPA Draft Method C) in Michigan towards the implementation of rapid, multi-site beach testing.

Fecal pollution remains a challenge for water quality managers at Great Lakes and inland recreational beaches. The fecal indicator of choice at these beaches is typically Escherichia coli (E. coli), determined by culture-based methods that require over 18 h to obtain results. Researchers at the United States Environmental Protection Agency (EPA) have developed a rapid E. coli qPCR methodology (EPA Draft Method C) that can provide same-day results for improving public health protection with demonstrated sensitivity, specificity, and data acceptance criteria. However, limited information is currently available to compare the occurrence of E. coli determined by cultivation and by EPA Draft Method C (Method C). This study provides a large-scale data collection effort to compare the occurrence of E. coli determined by these alternative methods at more than 100 Michigan recreational beach and other sites using the complete set of quantitative data pairings and selected subsets of the data and sites meeting various eligibility requirements. Simple linear regression analyses of composite (pooled) data indicated a correlation between results of the E. coli monitoring approaches for each of the multi-site datasets as evidenced by Pearson R-squared values ranging from 0.452 to 0.641. Theoretical Method C threshold values, expressed as mean log10 target gene copies per reaction, that corresponded to an established E. coli culture method water quality standard of 300 MPN or CFU /100 mL varied only from 1.817 to 1.908 for the different datasets using this model. Different modeling and derivation approaches that incorporated within and between-site variability in the estimates also gave Method C threshold values in this range but only when relatively well-correlated datasets were used to minimize the error. A hypothetical exercise to evaluate the frequency of water impairments based on theoretical qPCR thresholds corresponding to the E. coli water quality standard for culture methods suggested that the methods may provide the same beach notification outcomes over 90% of the time with Method C results differing from culture method results that indicated acceptable and unacceptable water quality at overall rates of 1.9% and 6.6%, respectively. Results from this study provide useful information about the relationships between E. coli determined by culture and qPCR methods across many diverse freshwater sites and should facilitate efforts to implement qPCR-based E. coli detection for rapid recreational water quality monitoring on a large scale in the State of Michigan.

Hongos anamorfos biodegradan los libros del Área Histórica de la Universidad Central del Ecuador (UCE) / Anamorphous fungi biodegrade the books of the Historical Area from the Central University of Ecuador (CUE)

Determinamos los géneros de hongos anamorfos que contaminan los libros del área de cuarentena y limpieza, dentro del Área Histórica de la Universidad Central del Ecuador (UCE). Realizamos un hisopado aleatorio a una muestra representativa de 50 de estos libros de acuerdo a una Tabla militarizada estándar. También hisopamos como muestra preferencial a 21 libros gravemente contaminados con hongos. Los hisopados tuvieron una superficie de 5x5 cm, friccionando en la pasta, el borde y el interior de estos libros. Las 213 muestras tomadas fueron inoculadas en medio de cultivo Agar Malta. Los medios fueron incubados a una temperatura de 28°C durante 7 días. Realizamos observaciones por microscopía a 40 y 100x además de usar literatura especializada para la identificación hasta el nivel de género de hongos anamorfos. Los géneros más abundantes en este estudio fueron Penicillium (80,2%) y Mucor (8,1%). (AU)

We determined the genera of anamorphic fungi that contaminate the books in the quarantine and cleaning area, within the Historical Area of the Central University of Ecuador (CUE). We performed a random swab on a representative sample of 50 of these books according to a standard militarized Table. We also swabbed as a preferential sample 21 books seriously contaminated with fungi. The swabs had a surface area of 5x5 cm, rubbing on the paste, the edge and the interior of these books. The 213 samples taken were inoculated in Agar Malta culture medium. The media were incubated at a temperature of 28° C for 7 days. We made observations by microscopy at 40 and 100x in addition to using specialized literature for the identification down to the genus level of anamorphic fungi. The most abundant genus in this study were Penicillium(80,2%) and Mucor(8,1%). (AU)

Recuento de Streptococcus mutans sobre restauraciones realizadas con adhesivo con y sin carga de nanopartículas de óxido de zinc y cobre / Streptococcus mutans count on restorations made with adhesive with and without loading zinc and copper oxide nanoparticles

Objetivo: Determinar las propiedades antimicrobianas de la incorporación de nanopartículas de óxido de zinc y cobre en un adhesivo de grabado y lavado total sobre Streptococcus mutans en pacientes con restauraciones de resina compuesta confeccionadas con adhesivo cargado. Métodos: Estudio experimental, randomizado, la muestra estuvo conformada por 25 pacientes, de ambos sexos, pertenecientes al posgrado de Ortodoncia de la Facultad de Odontología de la Universidad de Chile, en los cuales se confirmó presencia de Streptococcus mutans en saliva. Se confeccionaron restauraciones de resina compuesta oclusales, en premolares superiores con indicación de exodoncia por el tratamiento de ortodoncia, con adhesivo cargado (cuya composición fue 5/0,2 por ciento ZnO y Cu, respectivamente) y control (sin presencia de nanopartículas en su composición), según el listado de aleatorización. Se tomaron muestras microbiológicas en tres tiempos con la técnica de la cubeta (antes, 1 semana y 4 semanas posterior a la confección de las restauraciones). Se obtuvieron, aislaron e identificaron colonias de Streptococcus mutans a partir de las muestras obtenidas. Se usó el test de Mann-Whitney mediante el paquete estadístico SPSS v.21 Resultados: El promedio del recuento de UFC de Streptococcus mutans en el grupo experimental fue mayor posterior a la confección de las restauraciones de resina compuesta. Los resultados de la identificación molecular por PCR demuestran la presencia de Streptococcus mutans en 20 de 25 muestras. Conclusiones: No existen diferencias en el recuento de Streptococcus mutans antes y después de la aplicación del adhesivo sobre las restauraciones de resina compuesta(AU)

Objective: To determine the antimicrobial properties of the incorporation of zinc and copper oxide nanoparticles in an etching and total wash adhesive on Streptococcus mutans in patients with composite resin restorations made with loaded adhesive. Methods: Experimental and randomized trial, the sample were 25 patients, of both sexes, belonging to the FOUCH Orthodontic postgraduate program, in whom the presence of Streptococcus mutans in saliva was confirmed. Occlusal composite resin restorations were made in upper premolars with indication of extraction by orthodontic treatment, with loaded adhesive (whose composition is 5 / 0.2% ZnO and Cu respectively) and control (without the presence of nanoparticles in their composition), according to the scrambling listing. Microbiological samples were taken in three stages with the cuvette technique (before, 1 week and 4 weeks after the restoration was made). Colonies of Streptococcus mutans were obtained, isolated and identified from the samples obtained. The statistical analysis used the SPSS v.21 software, the data was analyzed by Mann Whitney test Results: The average CFU count of Streptococcus mutans in the experimental group (adhesive modified with zinc oxide and copper nanoparticles) was higher after the fabrication of composite resin restorations. The results of molecular identification by PCR demonstrate the presence of Streptococcus mutans in 20 of 25 samples. Conclusions: There are no differences in the count of Streptococcus mutans before and after the application of the adhesive on the composite resin restorations(AU)

Eficacia del instrumento XP-endo Finisher y del sistema EndoActivator en la reducción/eliminación del biofilm bacteriano: un ensayo ex vivo / Ability of the XP-endo Finisher instrument and the EndoActivator system to reduce/eliminate of the bacterial biofilm: an ex vivo assay

Objetivos: Comparar ex vivo la eficacia del instrumento XP-endo Finisher y del sistema EndoActivator en la reducción/eliminación del biofilm microbiano en conductos radiculares infectados. Materiales y métodos: Se utilizaron 23 premolares inferiores humanos extraídos cuya longitud fue estandarizada en 17 mm. Todos los conductos se prepararon con el sistema WaveOne Gold Medium (#35.06). Los dientes se esterilizaron, se inocularon con Enterococcus faecalis y se separaron en dos grupos experimentales de 10 piezas cada uno. De los 3 dientes remanentes, 1 fue utilizado como control positivo y 2, como controles negativos. En el grupo 1, las soluciones irrigantes se agitaron con XP-endo Finisher. En el grupo 2, se utilizó EndoActivator. Se tomaron muestras antes de la contaminación, luego de esta y después de la agitación de los irrigantes mediante conos de papel estériles. La carga microbiana fue sembrada en agar sangre y los conos se cultivaron en caldo tripteína de soja. La remoción de la carga microbiana se determinó por la presencia o ausencia de turbiedad del medio. Las unidades formadoras de colonias (UFC) remanentes se cuantificaron y los resultados se categorizaron como R1 (≤10 UFC) o R2 (>10 UFC). Los datos fueron analizados mediante la prueba de Fisher. Resultados: No hubo diferencias significativas entre XP-endo Finisher y EndoActivator (P>0,05). El número de usos no influyó sobre la capacidad operativa de ambos instrumentos (AU)

Aim: To compare ex vivo the effectiveness of the XP-endo Finisher and the EndoActivator in biofilm reduction/ removal from infected root canals. Materials and methods: Twenty three extracted human single-rooted lower premolars were selected and standardised to 17 mm in length. All the canals were prepared with WaveOne Gold Medium reciprocating files (#35.06). The teeth were autoclaved and inoculated with Enterococcus faecalis. The infected teeth were then assigned to 2 experimental groups of 10 teeth each according to the final irrigation/agitation protocol. Of the three remaining teeth, one was used as a positive control, and the other two were used as negative controls. In Group 1 the irrigating solutions were agitated with XP-endo Finisher while in Group 2 the EndoActivator was used. All root canals were sampled before and after contamination, and again after irrigant agitation with sterile paper points. The microbial load was spread on blood agar plates and the paper points were cultured in sterile trypticase soy broth. The removal of the microbial load was determined by visual observation of the turbidity of the media and by quantification of the number of colony-forming units (UFC). The results were categorized as R1 (≤10 UFC) or R2 (>10 UFC). Data were analysed by the Fisher's exact test at P<0.05. Results: No significant differences was found between XP-endo Finisher and EndoActivator (P>0.05) regarding their effectiveness in the reduction/removal of the microbial biofilm. The number of uses of both instruments did not affect their operative performance (AU) Conclusion: XPF and EA were both equally effective for microbial biofilm reduction/removal from ex vivo infected root canals (AU)

Suppressing of milk-borne pathogenic using new water-soluble chitosan-azidopropanoic acid conjugate: Targeting milk-preservation quality improvement.

Dairy animals are major reservoirs for many milk-borne pathogens (MBPs) such as Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli O157:H7). Thus, dairy industries dedicate most of their processes to eliminate or minimize microbial contamination. Although pasteurization may offer an ideal solution for microbial decontamination; nevertheless, it may negatively impact organoleptic and nutritive values of dairy products. In this context, this work aimed to develop an innovative strategy, to tackle this challenge, based on the chemical preservation of milk. In this endeavor, we have succeeded to design a new safe multifunctional bio-preservative based on natural antimicrobials (water-soluble chitosan (WSC) and 2-azidopropanoic acid (APA), (WSC-APA) conjugate). Interestingly, the minimum inhibitory concentrations (MICs) of WSC-APA have the capacity to completely suppress the proliferation of E. coli O157:H7 and S. aureus cells (100% bacterial reduction) in refrigerated milk samples during 20 and 24 h, respectively. Moreover, no Staphylococci species could be detected in refrigerated milk samples remediated with WSC-APA (0.25 mg/mL) after 6 storage days. Meanwhile, the coliform count has reduced by 99.7% in conjugate-treated milk samples during 10 storage days. Thus, new bio-preservative (WSA-APA) can be safely used to increase the shelf-life of milk without sacrificing its organoleptic and nutritive values.

Prevalence of Staphylococcus aureus colonization in patients for total joint arthroplasty in South Africa.

BACKGROUND: Periprosthetic joint infections (PJIs) are a major source of morbidity and mortality for patients undergoing total joint arthroplasty (TJA). Staphylococcus aureus (S. aureus) colonization is an independent, modifiable risk factor for periprosthetic joint infections. Post-operative infections are reported to be ten times greater in S. aureus carriers than in non-carriers in developed countries though recorded data is lacking for the developing world. This study aims to determine the prevalence of S. aureus colonization in patients awaiting TJA in South Africa. METHODS: We prospectively assessed 119 patients awaiting total knee arthroplasty and total hip arthroplasty between May and October 2016. We screened three separate anatomical sites on each patient for S. aureus. Patients with positive cultures were treated with intranasal mupirocin ointment and chlorhexidine body wash. Univariate and comparative statistical analyses to determine risk factors for colonization was conducted using t tests, Fisher's exact tests, and chi-square analyses. RESULTS: The overall prevalence of methicillin-sensitive S. aureus colonization was 31.9% (n = 38). There were no patients colonized with methicillin-resistant S. aureus. Nasal swabs returned a yield of 81.6% (n = 31), with groin swabs and axillary swabs at 39.5% (n = 15) and 28.9% (n = 11), respectively. Eradication was successful in 94.74% (n = 36) after 5 days treatment. All patients (100%) were decolonized after counseling and repeat eradication treatment. The overall complication rate was 7.6% (n = 9). The 30-day readmission rate in the S. aureus-colonized group was 7.9% (n = 3) as opposed to 7.4% (n = 6) in the non-colonized cohort. There were no 60- and 90-day readmissions and no cases were revised at a mean follow-up of 2.26 years. CONCLUSIONS: The rate of S. aureus colonization in patients undergoing elective TJA in a developing country was 31.9% and is equivalent to reported rates in developed countries. Eradication treatment with combined intranasal mupirocin ointment and chlorhexidine body wash is a successful treatment modality. A larger cohort of patients is recommended to determine risk factors and post-operative septic sequelae in this population group.

Viability assessment of Bifidobacterium longum ATCC 15707 on non-dairy foods using quantitative fluorescence microscopy.

This study demonstrates an effective technique for separating and purifying viable bacteria from samples that interfere with viability staining. The viability of Bifidobacterium longum ATCC 15707 was assessed using Percoll Buoyant Density Gradient Centrifugation (PBDC) to separate bacteria from complex non-dairy food matrices and Quantitative Fluorescence Microscopy (QFM) to determine individual cells using LIVE/DEAD BacLight bacterial viability staining. Water agar (3%) was used to retain cells of B. longum and offered a lower fluorescence background with BacLight viability staining, compared with fixation on polycarbonate (PC) black membrane. The effect of drying temperatures and non-dairy foods on viability of B. longum was assessed. B. longum coated on oat, peanut or raisin was separated by filtration, low- and high-speed centrifugation, flotation and sedimentation buoyant density centrifugation. Purified cells were subsequently deposited on water agar for rehydration followed by LIVE/DEAD BacLight viability staining and enumeration. Conventional plate counting was also conducted to compare viability results. Finally, this method was applied to assess cell membrane damages of B. longum incorporated onto non-dairy foods during 24 h drying. Furthermore, viability assessment of B. longum coated onto oat, peanut, or raisin was much lower by plate counting compared to viability staining. Drying appeared to have a greater impact when viability was assessed by plate counting compared to viability staining. IMPORTANCE: Enumeration of viable beneficial bacteria from function foods presents a significant bottleneck for product development and quality control. Interference with microscopic and/or fluorescent techniques by ingredients, time required to incubate plated microbes, and the transient nature of the colony forming unit make rapid assessment of viable bacteria difficult. Viability assessment of Bifidobacterium longum ATCC 15707 by Percoll Buoyant Density Gradient Centrifugation with LIVE/DEAD BacLight viability staining on water agar (3%) was in agreement with serial dilution enumeration. Without the need for incubation viability assessment by staining provided a more rapid means to assess the impact of drying on the viability of B. longum coated onto oat, peanut or raisin.

Effect of bisphenol-A (BPA) on placental biomarkers for inflammation, neurodevelopment and oxidative stress.

Background Bisphenol-A (BPA) is a widespread pollutant whose effects on pregnant women are poorly understood. Therefore, we investigated the effects of BPA on basal and bacteria-stimulated production of proinflammatory cytokines [interleukin (IL)-1ß, tumor necrosis factor-α (TNF-α) and IL-6], anti-inflammatory mediators [soluble glycoprotein 130 (sgp) 130, heme oxidase-1 (HO-1) and IL-10] and biomarkers for neurodevelopment [brain-derived neurotrophic factor (BDNF)], and oxidative stress [8-isoprostane (8-IsoP)] by the placenta. Methods Placental explant cultures were treated with BPA (0-10,000 nM) in the presence or absence of 107 colony-forming unit (CFU)/mL heat-killed Escherichia coli for 24 h. Biomarker concentrations in conditioned medium were quantified by the enzyme-linked immunosorbent assay (ELISA). Results Under basal conditions, IL-1ß and IL-6 production was enhanced by BPA in a dose-dependent manner. Sgp130, a soluble receptor that reduces IL-6 bioactivity, was suppressed by BPA at 1000-10,000 nM. BPA also enhanced BDNF production at 1000 and 10,000 nM, and 8-IsoP expression at 10 and 100 nM. For bacteria-treated cultures, BPA increased IL-6 production at 100 nM and reduced sgp130 at 1000 nM but had no effect on IL-1ß, TNF-α, BDNF, HO-1, 8-IsoP or IL-10 production. Conclusion BPA may increase placental inflammation by promoting IL-1ß and IL-6 but inhibiting sgp130. It may also disrupt oxidative balance and neurodevelopment by increasing 8-IsoP and BDNF production.

Evaluation of the complexity of indoor air in hospital wards based on PM2.5, real-time PCR, adenosine triphosphate bioluminescence assay, microbial culture and mass spectrometry.

BACKGROUND: The aim of this study was to establish a set of assessment methods suitable for evaluating the complex indoor environment of hospital wards and to ascertain the composition of bacteria and microbial ecology of hospital wards. METHODS: Colony-forming units (CFUs), PM2.5 detection, real-time PCR, and adenosine triphosphate (ATP) bioluminescence assay were employed to evaluate the complexity of indoor air in 18 wards of nine departments in a hospital and two student dormitories in a university. Subsequently, the microbial samples were quantified and identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). RESULTS: Although the studied indices were relatively independent, the PM2.5 content was correlated with bacterial CFUs determined by passive sedimentation method, bacterial and fungal counts measured by real-time PCR, and ATP bioluminescence assay. The composition of microorganisms in the air of hospital wards differed from that in the air of student dormitories. The dominant genera in hospital wards were Staphylococcus (39.4%), Micrococcus (21.9%), Corynebacterium (11.7%), Kocuria (4.4%), Bacillus (2.9%), Streptococcus (1.6%), Moraxella (1.6%), and Enterococcus (1.3%), and the microbial ecology differed between Respiration Dept. III and other hospital departments. Additionally, 11.1 and 27.3% of bacteria in hospital wards and student dormitories were not identified, respectively. CONCLUSIONS: Assessment of environmental quality of hospital wards should be based on comprehensive analysis with multiple indicators. There may be imbalances in the microbial diversity in the hospital wards, therefore, monitoring of the environmental quality of hospitals is important in the prevention of nosocomial infections.

Intracellular Assays to Monitor Survival and Growth of Yersinia pestis Within Macrophages.

Yersinia pestis is able to survive and replicate within macrophages, while also being able to live in the extracellular milieu of the host. Assays that facilitate better understanding of how Y. pestis survives intracellularly and subverts normal host antimicrobial defenses require the ability to monitor intracellular Y. pestis survival and replication. In this chapter three different assays for monitoring intracellular survival and replication will be described, along with the formulas and methods to quantify and present the acquired data. These assays are fundamental to answering a multitude of questions pertaining to which bacterial factors are important for intracellular survival. Additionally, these assays can be used, with modifications, for other intracellular pathogens of interest. The first assay discussed will be the conventional bacterial enumeration assay, which quantifies bacterial numbers directly through a classic colony forming units (CFU) assay. Quantifying bacterial burden through CFU determination allows for differentiation between intracellular/cell-associated bacteria and extracellular bacteria. However, CFU determination is laborious, does not allow for direct kinetic monitoring of bacterial growth, and is difficult to adapt to high throughput assays. Bioluminescence bioreporters that use luciferase to monitor bacterial numbers allow for simple, plate reader-based, real-time kinetic monitoring of bacterial growth that is amendable to high throughput techniques. Finally, we will describe live cell microscopy using fluorescent bioreporters, which allows for monitoring of bacterial replication in individual cells and the possibility to visualize interactions between bacterial and host proteins during intracellular infection.

Effect of a shielded continuous ultraviolet-C air disinfection device on reduction of air and surface microbial contamination in a pediatric oncology outpatient care unit.

BACKGROUND: For a clean hospital environment, we evaluated whether ultraviolet-C (UV-C) air disinfection reduces airborne and surface microbial contamination in an outpatient pediatric oncology center. METHODS: A pre- and post-intervention study compared 6 test locations, where continuous shielded UV-C air disinfection devices were installed, with 10 control locations without UV-C. Pre- and post-intervention air and surface samples were collected for bacterial and fungal cultures. Percent changes in colony forming unit (CFU) counts in the test and control locations were compared. RESULTS: Mean bacterial CFU count per cubic meter air and per surface contact plates decreased by 27% (P = .219) and 37% (P = .01), respectively, in test locations compared to 40% (P = .054) and 30% (P = .006) reductions in control locations. Mean fungal CFU count per cubic meter air and per surface contact plates increased by 14% (P = .156) and 19% (P = .048), respectively, in test locations compared to 24% (P = .409) and 2% (P = .34) increases in control locations. CONCLUSIONS: There were no consistent statistically significant differences in the air and surface culture results between test locations where UV-C devices were installed and control locations. The effectiveness of UV-C air disinfection in reducing air and surface microbial contamination in outpatient clinical areas where immunocompromised children are encountered was not proven.

Assessment of stability and fluctuations of cultured lower airway bacterial communities in people with cystic fibrosis.

BACKGROUND: Routine clinical culture detects a subset of the cystic fibrosis (CF) airways microbiota based on culture-independent (molecular) methods. This study aimed to determine how extended sputum culture of viable bacteria changes over time in relation to clinical status and predicts exacerbations. METHODS: Sputa from patients at a baseline stable and up to three subsequent time-points were analysed by extended-quantitative culture; aerobe/anaerobe densities, ecological indexes and community structure were assessed together with clinical outcomes. RESULTS: Eighty patients were prospectively recruited. Sputa were successfully collected and cultured at 199/267 (74.5%) study visits. Eighty-two sputa from 25 patients comprised a complete sample-set for longitudinal analyses. Bacterial density, ecological indexes and clinical outcomes were unchanged in 18 patients with three sequential stable visits. Conversely, in 7 patients who had an exacerbation, total bacterial and aerobe densities differed over four study visits (P < .001) with this difference particularly apparent between the baseline visit and completion of acute antibiotic treatment where a decrease in density was observed. Bacterial communities were more similar within than between patients but stable patients had the least variation in community structure over time. Using logistic regression in a further analysis, baseline features in 37 patients without compared to 15 patients with a subsequent exacerbation showed that clinical measures rather than bacterial density or ecological indexes were independent predictors of an exacerbation. CONCLUSIONS: Greater fluctuation in the viable bacterial community during treatment of an exacerbation than between stable visits was observed. Extended-quantitative culture did not provide prognostic information of a future exacerbation.

Effect of using ceiling-mounted systems for imaging in hybrid operating rooms on the level of colony-forming units during surgery.

BACKGROUND: Large imaging systems in hybrid operating rooms are used increasingly. However, the effect of these ceiling-mounted imaging systems on air quality during surgical procedures has not been studied to date. AIM: To evaluate the level of colony-forming units (cfu)/m3 near the surgical wound and near the instrument table during surgery. METHODS: Measurements were performed in four hybrid operating rooms at four different hospitals. During surgical procedures, at least three samples were taken with active slit air samplers. At the end of the day's surgical schedule, simulations involving movement of the ceiling-mounted system were also performed. The threshold value for the average cfu level during a surgical procedure was set at ≤10 cfu/m3, and for individual samples was set at ≤30 cfu/m3. RESULTS: The median value near the surgical wound was 1 cfu/m3 and at the instrument table was 2 cfu/m3. However, in one hybrid operating room (two procedures out of 16) where the instrument table was not positioned directly under the unidirectional flow (UDF) system, the threshold value for the average cfu level at the instrument table was exceeded. For one of these procedures, the maximum value for an individual sample was also exceeded. CONCLUSIONS: A ceiling-mounted imaging system in combination with a UDF system can result in cfu levels near the surgical wound and at the instrument table that are well below the threshold value of 10 cfu/m3 during surgery. If the instrument table is not positioned directly under the UDF system, the cfu level is higher.

Subgingival microbiological profile of periodontitis patients in Dominican Republic

The aim of this study was to evaluate the deproteinization of primary enamel by analyzing etching pattern types, with and without the application of 5% NaOCl before acid etching with 37% H3PO4. Fifteen extracted human primary molars were randomly selected for the present in vitro study; 1mm x 1mm blocks were prepared and divided into two groups (n = 21). These groups were treated as follows: Group AAcid Etching with 37% H3PO4 gel for 15 s; Group B5% NaOCl for 60 s + Acid Etching with 37% H3PO4for 15 s. The specimens were prepared for scanning electron microscopy analysis. The images were evaluated for quality types I and II etching of the enamel surface using ImageJ software. Datasets were checked for normality by KolgomorvSmirnov test and the nonparametric unpaired MannWhitney test was applied. The mean surface area of type I and II etching pattern values was 1922.314 µm2for Group A and 3840.473 µm2Group B. We conclude that deproteinization with 5% NaOCl prior to acid etching can be used to increase the area of adhesion and the quality of the etching pattern (AU)

El objetivo del estudio fue evaluar la desproteinización del esmalte primario a través de los tipos de patrones de grabado, con y sin NaOCl 5% utilizado antes del grabado ácido con H3PO4 37%. Quince dientes primarios humanos extraídos se seleccionaron al azar para el presente estudio in vitro, se prepararon bloques de 1mm x 1 mm y se dividieron en dos grupos (n = 21). Estos grupos se trataron de la siguiente manera: Grupo A: Grabado ácido con H3PO4 37% en gel durante 15 segundos; Grupo B: NaOCl 5% durante 60 segundos + Grabado ácido con H3PO4 37% durante 15 segundos. Las muestras se prepararon para el análisis de microscopía electrónica de barrido. Las imágenes obtenidas se evaluaron principalmente por la calidad de los grabados tipo I y II de la superficie del esmalte primario, utilizando el software Image J. Los datos se analizaron en cuanto a su normalidad mediante la prueba de KolgomorvSmirnov, se utilizó pruebas no paramétricas: Prueba de MannWhitney no pareada. Como resultado, se encontró que el área de superficie media de los valores de patrón de grabado de tipo I y II para el Grupo A era 1922,314 µm2 y el Grupo B era 3840,473 µm2. Finalmente, llegamos a la conclusión de que se puede usar la desproteinización con NaOCl 5% antes del grabado ácido para aumentar el área de adhesión y la calidad del patrón de grabado (AU)

Fog, phenolic acids and UV-A light irradiation: A new antimicrobial treatment for decontamination of fresh produce.

This study evaluates synergistic interactions of food grade phenolic acids (gallic and ferulic acid) and UV-A light to achieve decontamination of fresh produce using a fog to improve dispersion of the phenolic acids on produce surface. Nonvirulent strains of Escherichia coli O157:H7 and Listeria innocua were used as model bacteria and spinach was selected as a model fresh produce. Synergistic combination of a fog deposited phenolic acid and a UV-A light treatment achieved reduction in bacterial plate count up to 2 log CFU/cm2 independently of the initial load of the bacteria (104 or 106 CFU/cm2). Following the treatment, fog deposited gallic and ferulic acid could be easily removed from the surface of produce by immersion in water and the treatment did not significantly alter the total endogenous phenolic content of spinach. The treatment also did not affect the texture, but impacted the color of the spinach leaves on a Hunter's Lab scale although the visual color changes were small. Overall, this technology may aid in developing alternative approaches for decontamination processes using food grade compounds.

Monitoring bacterial communities in ε-Polylysine-treated bighead carp (Aristichthys nobilis) fillets using culture-dependent and culture-independent techniques.

The present study investigated the effect of ε-Polylysine on bacterial communities, sensorial, and chemical properties [total volatile basic nitrogen (TVB-N), biogenic amines, and breakdown products of adenosine triphosphate] of bighead carp (Aristichthys nobilis) fillets stored at 4 ±â€¯1 °C. Bacterial communities were explored by the culture-dependent method and the high-throughput sequencing targeting on 16S rRNA genes. The results showed that the major genera in spoiled control samples were Aeromonas, Pseudomonas, Shewanella, and Acinetobacter. ε-Polylysine inhibited the growth of Pseudomonas, Shewanella, and Acinetobacter. Consequently, Aeromonas and Janthinobacterium were dominant in spoiled treated samples. The sensorial shelf-life of the control and treated groups were 8 days and 10 days, respectively. Furthermore, due to the inhibitory effect of ε-Polylysine on bacteria, chemical changes of the treated group were slower, reflecting as lower concentrations of TVB-N, putrescine, cadaverine, and hypoxanthine, and higher contents of inosine 5'-monophosphate and hypoxanthine riboside at the end of storage. In conclusion, ε-Polylysine altered the bacterial communities and delayed quality deterioration of bighead carp fillets during chilled storage.