Phenotyping and genotyping methods applied to investigate the relatedness of Brazilian isolates of Enterobacter cloacae

In order to evaluate the resolving power of several typing methods to identify relatedness among Brazilian strains of Enterobacter cloacae, we selected twenty isolates from different patients on three wards of a University Hospital (Orthopedics, Nephrology, and Hematology). Traditional phenotyping methods applied to isolates included biotyping, antibiotic sensitivity, phage-typing, and O-serotyping. Plasmid profile analysis, ribotyping, and macrorestriction analysis by pulsed-field gel electrophoresis (PFGE) were used as genotyping methods. Sero- and phage-typing were not useful since the majority of isolates could not be subtyped by these methods. Biotyping, antibiogram and plasmid profile permitted us to classify the samples into different groups depending on the method used, and consequently were not reliable. Ribotyping and PFGE were significantly correlated with the clinical epidemiological analysis. PFGE did not type strains containing nonspecific DNase. Ribotyping was the most discriminative method for typing Brazilian isolates of E. cloacae.

Random amplified polymorphic DNA profiles as a tool for the characterization of Brazilian keratitis isolates of the genus Acanthamoeba

The genus Acanthamoeba comprises free-living amebae identified as opportunistic pathogens of humans and other animal species. Morphological, biochemical and molecular approaches have shown wide genetic diversity within the genus. In an attempt to determine the genetic relatedness among isolates of Acanthamoeba we analyzed randomly amplified polymorphic DNA (RAPD) profiles of 11 Brazilian isolates from cases of human keratitis and 8 American type culture collection (ATCC) reference strains. We found that ATCC strains belonging to the same species present polymorphic RAPD profiles whereas strains of different species show very similar profiles. Although most Brazilian isolates could not be assigned with certainty to any of the reference species, they could be clustered according to pattern similarities. The results show that RAPD analysis is a useful tool for the rapid characterization of new isolates and the assessment of genetic relatedness of Acanthamoeba spp. A comparison between RAPD analyses and morphological characteristics of cyst stages is also discussed.

The 42-kDa coat protein of Andean potato mottle virus acts as a transcriptional activator in yeast

Interactions of viral proteins play an important role in the virus life cycle, especially in capsid assembly. Andean potato mottle comovirus (APMoV) is a plant RNA virus with a virion formed by two coat proteins (CP42 and CP22). Both APMoV coat protein open reading frames were cloned into pGBT9 and pGAD10, two-hybrid system vectors. HF7c yeast cells transformed with the p9CP42 construct grew on yeast dropout selection media lacking tryptophan and histidine. Clones also exhibited ß-galactosidase activity in both qualitative and quantitative assays. These results suggest that CP42 protein contains an amino acid motif able to activate transcription of His3 and lacZ reporter genes in Saccharomyces cerevisiae. Several deletions of the CP42 gene were cloned into the pGBT9 vector to locate the region involved in this activation. CP42 constructions lacking 12 residues from the C-terminal region and another one with 267 residues deleted from the N-terminus are still able to activate transcription of reporter genes. However, transcription activation was not observed with construction p9CP42deltaC57, which does not contain the last 57 amino acid residues. These results demonstrate that a transcription activation domain is present at the C-terminus of CP42 between residues 267 and 374.

Characterization of bovine respiratory syncytial virus isolated in Brazil

This paper presents the first isolation of bovine respiratory syncytial virus in Brazil and its physicochemical, morphological and molecular characterization. The virus was isolated from 33 samples of nasotracheal secretions, successively inoculated into a Madin-Darby bovine kidney cell culture, which was characterized by physicochemical tests and morphological observation by electron microscopy. The Brazilian sample is an RNA pleomorphic, enveloped, thermolabile and non-hemagglutinating spicular virus. Reverse transcription, followed by nested polymerase chain reaction (nRT-PCR) assay was carried out using oligonucleotides B1, B2A, B3 and B4 for the fusion proteins (F) and B5A, B6A, B7A and B8 for the attachment protein (G). The nRT-PCR-F amplified a fragment of 481 bp corresponding to part of the gene that codes for protein F, whereas nRT-PCR-G amplified a fragment of 371 bp, in agreement with part of the G gene. The virus isolated from Brazilian samples in this study corresponded to the bovine respiratory syncytial virus, and RT-PCR proved to be useful for the diagnosis of bovine clinical samples.

Recombinant E2 glycoprotein of bovine viral diarrhea virus induces a solid humoral neutralizing immune response but fails to confer total protection in cattle

Two recombinant baculoviruses were produced in order to obtain a bovine viral diarrhea virus (BVDV) immunogen: AcNPV/E2 expressing E2 glycoprotein, and AcNPV/E0E1E2 expressing the polyprotein region coding for the three structural proteins of BVDV (E0, E1, and E2). Mice were immunized with Sf9 cells infected with the recombinant baculoviruses in a water in oil formulation and the production of neutralizing antibodies was evaluated. Since E2 elicited higher neutralizing antibody titers than E0-E1-E2 polyprotein, it was selected to immunize cattle. Calves received two doses of recombinant E2 vaccine and were challenged with homologous BVDV 37 days later. The recombinant immunogen induced neutralizing titers which showed a mean value of 1.5 ± 0.27 on the day of challenge and reached a top value of 3.36 ± 0.36, 47 days later (84 days post-vaccination). On the other hand, sera from animals which received mock-infected Sf9 cells did not show neutralizing activity until 25 days post-challenge (62 days post-vaccination), suggesting that these antibodies were produced as a consequence of BVDV challenge. Even when no total protection was observed in cattle, in vitro viral neutralization assays revealed that the recombinant immunogen was able to induce neutralizing antibody synthesis against the homologous strain as well as against heterologous strains in a very efficient way.

Characterization of bacteriophages infecting clinical isolates of Pseudomonas aeruginosa stored in a culture collection

Some clinical isolates of Pseudomonas aeruginosa stored in our culture collection did not grow or grew poorly and showed lysis on the culture plates when removed from the collection and inoculated on MacConkey agar. One hypothesis was that bacteriophages had infected and killed those clinical isolates. To check the best storage conditions to maintain viable P. aeruginosa for a longer time, clinical isolates were stored at various temperatures and were grown monthly. We investigated the presence of phage in 10 clinical isolates of P. aeruginosa stored in our culture collection. Four strains of P. aeruginosa were infected by phages that were characterized by electron microscopy and isolated to assess their ability to infect. The best condition to maintain the viability of the strains during storage was in water at room temperature. Three Siphoviridae and two Myoviridae phages were visualized and characterized by morphology. We confirmed the presence of bacteriophages infecting clinical isolates, and their ability to infect and lyse alternative hosts. Strain PAO1, however, did not show lysis to any phage. Mucoid and multidrug resistant strains of P. aeruginosa showed lysis to 50% of the phages tested.

Sensory evaluation of rice fortified with iron

The objective of this study was to examine sensory differences between conventional rice and iron-fortified Ultra Rice rice (UR) and determine consumer acceptance. Differences between both types of rice were analyzed using the Duo-Trio Test on 37 non-trained judges. The Acceptance Test evaluated general rice appearance, color, aroma and taste by 43 non-trained judges, using a 7-point hedonic scale with extremes ranging from "really disliked" and "really liked." There were no significant differences between the analyzed samples of conventional rice and UR rice using Duo-Trio Test. Mean values from the Acceptance Test for the evaluated attributes (general appearance, color, aroma e taste) were 5.6 and 5.9, corresponding to "liked" and "really liked" according to the hedonic scale, respectively. The iron did not alter the sensory characteristics of the final product, and the iron-fortified rice was well accepted.

Influence of the use of rice bran extract as a source of nutrients on xylitol production

Xylose-to-xylitol bioconversion using 2.5 or 10% (v/v) rice bran extract was performed to verify the influence of this source of nutrients on Candida guilliermondii metabolism. Semisynthetic medium (SM) and sugarcane bagasse hemicellulosic hydrolysate detoxified with ion-exchange resins (HIE) or with alteration in pH combined with adsorption onto activated charcoal (HAC) were fermented in 125 mL Erlenmeyer flasks at 30 ºC and 200 rpm for 72 hours. Activated charcoal supplemented with 2.5% (v/v) rice bran extract was fermented by C. guilliermondii in a MULTIGEN stirred tank reactor using pH 5.0 and 22.9/hour oxygen transfer volumetric coefficient. Higher values of xylitol productivity (0.70, 0.71, and 0.62 g.Lh-1) and xylose-to-xylitol conversion yield (0.71, 0.69, and 0.63 g.g-1) were obtained with 2.5% (v/v) rice bran in semisynthetic medium, ion-exchange resins, and activated charcoal, respectively. Moreover, during batch fermentation, the xylitol volumetric productivity and fermentation efficiency values obtained were 0.53 g.Lh-1 and 61.1%, respectively.

Not any type of rice performs equally to improve lactose-induced diarrhea characteristics in rats: is amylose an antidiarrheal factor?

The effectiveness of different types of rice in relation to their ability to accelerate diarrhea recovering was evaluated in a rat model of osmotic diarrhea (OD). Animals (90-100 g) received protein free diet until reaching up to 20% weight loss, followed by lactose rich diet (LRD) to induce osmotic diarrhea. Rats presenting osmotic diarrhea were divided into 4 groups, which received lactose rich diet for 4 days from 8 am to 8 pm, and one of three experimental products containing 6% rice flour differing in amylose content during the night: high (HA), intermediate (IA), and low (LA). A group fed stock diet containing equivalent amount of lactose was taken as control and allowed to recover spontaneously. Amylose and viscosity (cp at 25 °C, 10 rpm) of final products were determined. Effectiveness was expressed as the ratio between percentages of normal vs. diarrheic stools during the treatment. Fecal characteristics in this rat model improved only as result of feeding high amylose content (HA) type of rice. In this experimental model of osmotic diarrhea in young rats, the antidiarrheal effects of rice were strongly dependent on the type of diet used and appear to be related to its amylose content.

Stability and microbiological quality of rice bran subjected to different heat treatments

Rice bran is a byproduct commonly used for animal feeding; however its nutritional value and potential application in human diet have attracted market interest. Its preservation for safe use is still a challenge, so the objective of this study was to determine the quality of commercially available rice bran samples subjected to different heat treatments (extruding, parboiling, toasting, and microwave oven heating) in order to promote stabilization during storage under room temperature. Rice bran samples were collected from two industries, and each treatment was divided in three parts, each corresponding to three repetitions. All samples were evaluated for moisture content, total microorganisms, mold and yeast counting, hydrolytic rancidity, and lipase activity during 90 days of storage. Most of the heat treatments, including domestic and thermoplastic extrusion, generated products which may be used for human consumption under the tested conditions in terms of physicochemical and microbiological quality. The domestic treatments were more efficient in eliminating microorganisms or keeping them within acceptable limits. The toasted rice bran showed satisfactory results in terms of moisture, hydrolytic rancidity control, and lipase activity.

Physicochemical and sensory profile of rice bran roasted in microwave

The purpose of this study was to evaluate the physical, chemical, and sensory changes in bran from three rice cultivars according to microwave roasting time. This study analyzed three rice cultivars, BRS Sertaneja (S), BRS Primavera (P), and IRGA 417 (I) determining the color parameters (L*, a*, and b *) at 6, 9, 12, 15, and 18 minutes of roasting time. After applying the difference from control test, the rice brans with different characteristics aroma and flavor were selected: S and P roasted for 9 and 15 minutes and IRGA 417 roasted for 9, 12, and 15 minutes. These samples were characterized by Free-Choice Profile descriptive sensory analysis, and their chemical composition was also determined. The longer the roasting process, the higher the roasted flavor intensity and aroma. The IRG 417 cultivar roasted for 12 minutes showed a sweeter flavor and aroma. After roasting, the brans remained rich in protein and lipid and presented higher fiber content and lower reducing sugar and phytic acid content. Microwave roasting for 12 minutes can be a viable option for improving the sensory functional and nutritional characteristics of the rice bran considering its use in food products.

Effect of adding unconventional raw materials on the technological properties of rice fresh pasta

The aim of this study was to develop fettuccini type rice fresh pasta by cold extrusion. To produce the pasta, a 2² Central Composite Rotational Design was used, in which the effects of the addition of pre-gelatinized rice flour - PGRF (0-60%) and modified egg albumin - MEA (0-10%) were studied. The dependent variables were the results of the cooking test and of the instrumental texture. The optimum cooking time for all of the formulations of rice fresh pasta was 3 minutes. MEA had a greater effect on increasing the weight of the pasta when compared to that of PGRF. It was found that with the addition of PGRF increase in loss of solids in cooking water, whereas MEA exerted the opposite effect on this parameter. Moreover, the maximum value of MEA (10%) had an optimum effect on pasta firmness, while PGRF had a negative effect on this parameter. The maximum values of PGRF and MEA reduced the stickiness of the pasta. Based on these results and on the parameters considered as most important, the rice pasta with the best technological characteristics was that with the maximum levels of MEA (10%) and no addition of PGRF (0%). This product was submitted to sensory and microbiological analyses, with good results.

Optimization of extrusion variables for the production of snacks from by-products of rice and soybean

This study aimed to define the process conditions to obtain snacks from the by-products of rice and soybean with physical characteristics suitable for marketing. Therefore, the effects of moisture and extrusion temperature on the expansion and color of the products obtained experimentally obtained were evaluated, and the proximate composition of the by-products and that of the snack with greater desirability were determined. Response surface methodology and rotational central composite design were used, and desirability test based on the regression models adjusted was applied. The most desirable snack, with the highest expansion index (3.39), specific volume (13.5 mL.g-1), and the chromaticity coordinate a* (2.79), was obtained under 12 g.100 g-1 moisture and 85ºC of temperature in the third zone of the extruder. The snack produced under these conditions attained content of protein and lipid content 41 and 64% higher than that of the traditional corn snack. It can be concluded that producing extruded snack made form a mixture of broken grains, rice bran, and soybean okara (81:9:10) is technologically feasible, enabling the development of a new product with good nutritional value that can improve the diet of children, the main consumers of this type of food.

The influence of extraction methods on composition and antioxidant properties of rice bran oil

AbstractThe current study was employed to assess the influence of the different extraction methods on total tocols, γ-oryzanol content, and antioxidant properties of Chiang Mai Black rice, Mali Red rice, and Suphanburi-1 Brown rice bran oil. Rice bran oil (RBO) was extracted by Hexane, Hot pressed, Cold pressed, and Supercritical Fluid Extraction (SFe) methods. High yield of RBO was extracted by hexane and SFe methods. Total and subgroups of tocols, and γ-oryzanol content were determined by HPLC. The hexane extracted sample accounts for high content of γ-oryzanol and tocols. Besides, all of RBO extracts contain a significantly high amount of γ-tocotrienol. In vitro antioxidant assay results indicated that superior quality of oil was recovered by hexane extraction. The temperature in the extraction process also affects the value of the oil. Superior quality of oil was recovered by hexane extraction, in terms of phytochemical contents and antioxidant properties compared to other tested extraction methods. Further, thorough study of factors compromising the quality and quantity of RBO recovery is required for the development of enhanced functional foods and other related products.

Physiological potential of rice seeds treated with rhizobacteria or the insecticide thiamethoxam

Development of new technologies, aiming at increasing productivity in different crops, involves constant research on the effectiveness and application of these techniques in seed treatment. In this study, it was aimed at evaluating physiological potential of rice seeds treated with plant growth-promoting rhizobacteria (PGPR) (strains DFs185, DFs223, DFs306, DFs416), or with two dosages of the insecticide thiamethoxam. The variables assessed were: germination (G); first count of germination (FCG); cold test (CT); length of seedlings aerial parts (LAP), root system (LRS), and total length (TL); emergence speed index (ESI); emergence (E), at 14 days; and dry phytomass (DP). Treatments have had a positive effect on percentage of G, FCG and E. The strain DFs185 has promoted increase in percentage emergence, for five of the six lots assessed. The variables: LAP; LRS; TL; ESI; and DP have undergone low or none influence of treatments; and there has been no toxic effect of rhizobacteria or insecticide thiamethoxam. In the cold test, a negative effect of treatments has been detected. Seed treatment with rhizobacteria, as well as with thiamethoxam, improve quality of low quality rice seeds. The strain DFs185 is promising for treating rice seeds, once it stimulates seed germination and emergence.