Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome

Background. From shotgun libraries used for the genomic sequencing of the phytopathogenic bacterium Xanthomonas axonopodis pv. citri (XAC), clones that were representative of the largest possible number of coding sequences (CDSs) were selected to create a DNA microarray platform on glass slides (XACarray). The creation of the XACarray allowed for the establishment of a tool that is capable of providing data for the analysis of global genome expression in this organism. Findings. The inserts from the selected clones were amplified by PCR with the universal oligonucleotide primers M13R and M13F. The obtained products were purified and fixed in duplicate on glass slides specific for use in DNA microarrays. The number of spots on the microarray totaled 6,144 and included 768 positive controls and 624 negative controls per slide. Validation of the platform was performed through hybridization of total DNA probes from XAC labeled with different fluorophores, Cy3 and Cy5. In this validation assay, 86% of all PCR products fixed on the glass slides were confirmed to present a hybridization signal greater than twice the standard deviation of the deviation of the global median signal-to-noise ration. Conclusions. Our validation of the XACArray platform using DNA-DNA hybridization revealed that it can be used to evaluate the expression of 2,365 individual CDSs from all major functional categories, which corresponds to 52.7% of the annotated CDSs of the XAC genome. As a proof of concept, we used this platform in a previously work to verify the absence of genomic regions that could not be detected by sequencing in related strains of Xanthomonas. © 2010 Moreira et al; licensee BioMed Central Ltd.

Formação de biofilme cristalino de Proteus mirabilis em cateter urológico

The indwelling urethral catheter has an important role for patients with urinary retention, bladder obstruction, neurological damage and other diseases. Urine contains minerals which precipitate in alkaline pH, crystallize and block the urological catheter. The crystallization of the ionic components of urine occurs in the presence of urease, an enzyme produced by Proteus mirabilis. This bacterium adheres to inanimate surfaces and forms biofilms. The aim of this study was to investigate the formation of crystalline biofilm on the luminal surface of siliconized latex catheters by means of scanning electron microscope, after channeling artificial urine infected with Proteus mirabilis. The experiment was performed in vitro using a dynamic flow system. The artificial urine compounds were salts of calcium, magnesium, phosphates, urea and egg albumin, and it was infected with Proteus mirabilis ATCC 25933. The urine flow was stopped after crystallization of the ionic components. Crystallization was observed after alkalinization of urine. Scanning electron microscopy showed the presence of crystals and morphologies typical of bacilli embedded in an amorphous mass on the internal lumen of the catheter. The present study showed that catheter encrustation may limit the use of long-term indwelling catheter. © SBEB - Sociedade Brasileira de Engenharia Biomédica.

Bacillus Thuringiensis mutant increase activity against Apodoptera Frugiperda larvae

Bacillus thuringiensis is a Gram-positive bacterium which main characteristic is the production of Cry proteins, that is toxic to some insects. These proteins, when ingested by susceptible insects, become active causing their death. In nature, it is possible to found B. thuringiensis strains which produce these proteins, but they differ in productivity (some of these isolates are more productive then others), and as to the toxicity levels of the produced proteins. Two B. thuringiensis strains that were highly effective against Spodoptera frugiperda larvae were chosen to verifying genetic mutation implication on Cry proteins productivity. One strain with a prolific spores production, while the other one only produced small amounts of spores. A genomic mutant library of these two isolates was, separately, constructed by genome Tn-5 transposon random insertion. Data analysis showed that mutation had a direct effect on the spores production, inducing an increase as well as a decrease in the production, according to the different strain observed. These results indicate, for the first time, that it is possible to use the described technique with B. thuringiensis, as well as the possibility to genetically breeding this bacteria. Another possibility introduced here is the possibility to do functional genetic studies mediated by mutagenesis in this bacterium.

Capacidade predatória de chrysoperla externa sobre bemisia tabaci biótipo b expostas ao fungo metarhizium anisopliae

The objective of this study was to determine the influence of Metarhizium anisopliae on the predatory capacity and functional response of Chrysoperla externa fed Bemisia tabaci B biotype rearing cotton. The stock rearings of B. tabaci B biotype and C. externa were maintained in cages according to preexisting methodologies. Upon hatching, predator larvae were fed on eggs of Ephestia küehniella until their 3rd instar and then fed on whitefly nymphs in density exceeding their intake capacity (150 nymphs/individual). The nymphs were offered on cotton leaf disks (5 cm diameter), 24 hours after the treatments application following: T1: Control (water); T2: M. anisopliae at the concentration of 105 conidia cm-3; T3: 10 8 conidia cm-3 origined from Metharril® formulation of the 109 viable conidia cm-3 M. anisopliae active principle. The predatory capacity was obtained by the difference between the numbers of nymphs supplied and nymphs left. Five different nymphal densities (130, 160, 190, 220, and 250) were selected for the functional response study. The number of nymphs preyed was recorded after 24, 48, 72, and 96 hours. The M. anisopliae entomopathogenic fungi did not influence predatory capacity of C. externa which ranged from 70.6 individuals at the highest concentration to 87.1 in the control. Conclued that the C. externa predatory capacity was not affected by fungi M. anisopliae concentrations. The predator showed a type-II functional response after 24 hours, and a type-I response after 48, 72s, and 96 hours independent from fungi concentrations.

Avaliação da qualidade da água purificada em farmácias magistrais da região de São José do Rio Preto, SP

Water is the raw material used most in the production of diverse pharmaceutical forms and, being a constituent of the formulation itself, is subject to a number of physico-chemical and microbiological specifications. In addition, it is indispensable for laboratory tests and the cleaning of equipment and apparatus. The aim of this study was to ascertain the degree of physicochemical and microbiological contamination of purified water used in compounding pharmacies in the city of São José do Rio Preto, SP, Brazil. Samples were taken as recommended in the USP Pharmacopeia, with careful aseptic technique, and sent immediately the to quality control laboratory. Physicochemical properties were analyzed, including appearance, pH, conductivity, residue after evaporation, ammonia, calcium, chloride, heavy metals, sulfate and oxidizable substances, and microbiological tests were performed: total aerobic microbial count and detection of total and thermotolerant coliforms and Pseudomonas aeruginosa. Results showed that some parameters did not conform to the standards, especially pH, conductivity, inorganic impurities, oxidizable substances and microbiological test data, in 10%, 17%, 10%, 14% and 20% of the analyzed samples, respectively, This points to the need for greater care in the production and/or storage of purified water in these pharmaceutical establishments.

Presence of a human diarrheagenic escherichia coli clone in captivity kept psittacidaes

Bacterial cultures of cloaca swabs from 86 captivity kept psittacidaes revealed 17 Escherichia coli bearing birds sharing strains which, on the basis of enterobacterial repetitive intergenic consensus (ERIC) PCR analysis, proved to be genetically similar. Further, triplex PCR specific for the genetic markers chuA, yjaA, and TSPE4.C2 was used to assign the strains to the E. coli reference collection (EcoR) B2 group. One strain of each, from the enteropathogenic (EPEC), enteroaggregative (EAEC) and Shiga toxin (STEC) E. coli pathovars were found among these isolates. © Marietto-Gonçalves et al.; Licensee Bentham Open.

Quercetin reduces Staphylococcus aureus interaction with neutrophils

Flavonoids, including quercetin, have been reported to modulate the ability of Staphylococcus aureus to adhere to host tissue without exhibiting direct antibacterial activity. In the present study, we evaluated the interaction of S. aureus pretreated with 40 μg/mL of quercetin with neutrophils to assay oxidative burst stimulation, using luminol-amplified chemiluminescence. S. aureus pre-incubated with subinhibitory concentration of quercetin induced significantly less light emission by neutrophils than did untreated bacteria. The results of the present study demonstrate that quercetin decreases S. aureus uptake by neutrophils.

Detection of SPM and IMP metallo-β-lactamases in clinical specimens of Pseudomonas aeruginosa from a Brazilian public tertiary hospital

Phenotypic and genotypic SPM and IMP metallo-β-lactamases (MBL) detection and also the determination of minimal inhibitory concentrations (MIC) to imipenem, meropenem and ceftazidime were evaluated in 47 multidrug-resistant Pseudomonas aeruginosa isolates from clinical specimens. Polymerase chain reaction detected 14 positive samples to either blaSPM or blaIMP genes, while the best phenotypic assay (ceftazidime substrate and mercaptopropionic acid inhibitor) detected 13 of these samples. Imipenem, meropenem and ceftazidime MICs were higher for MBL positive compared to MBL negative isolates. We describe here the SPM and IMP MBL findings in clinical specimens of P. aeruginosa from the University Hospital of Botucatu Medical School, São Paulo, Brazil, that reinforce local studies showing the high spreading of blaSPM and blaIMP genes among Brazilian clinical isolates. © 2011 Elsevier Editora Ltda.

Reduction of bacterial adhesion to biocompatible polymer surfaces via plasma processing

Plasma processing of the surfaces of biomaterials is interesting because it enables modification of the characteristics of a surface without affecting bulk properties. In addition, the results are strongly influenced by the conditions of the treatment. Therefore, by adjusting the plasma parameters it is possible to tailor the surface properties to best fulfill the requirements of a given application. In this work, polyurethane substrates have been subjected to sulfur hexafluoride glow discharge plasmas. The influences of different SF 6 plasma exposure times and pressures on the adhesion of Staphylococcus aureus and Pseudomonas aeruginosa to the polymer have been investigated. The wettability and surface free energy have been evaluated via contact angle measurements. At low pressure (6.7 Pa) the contact angle decreases with increasing exposure time in the 180 s to 540 s interval, but at higher pressure (13.3 Pa) it increases as a function of the same variable. Bacterial adhesion has been quantified from in vitro experiments by determining the growth of colonies on Petri dishes treated with agar nutrient. It has been observed that the surface properties play an important role in microbe adhesion. For instance, the density of adhered P. aeruginosa decreased as the surface contact angle increased. S. aureus preferred to adhere to hydrophobic surfaces. © 2011 by Begell House, Inc.

Gastrointestinal changes associated to heart failure

Over the last decade, several studies were conducted on the gastrointestinal changes associated to chronic heart failure. This article presents a literature review on the physiopathology and clinical consequences of pathological digestive changes of heart failure patients. Structural and functional abnormalities of the gastrointestinal tract, such as edema of absorptive mucosa and intestinal bacterial overgrowth, have been leading to serious clinical consequences. Some of these consequences are cardiac cachexia, systemic inflammatory activation and anemia. These conditions, alone or in combination, may lead to worsening of the pre-existing ventricular dysfunction. Although currently there is no therapy specifically earmarked for gastrointestinal changes associated to heart failure, the understanding of digestive abnormalities is germane for the prevention and management of systemic consequences.

Qualidade microbiológica da água e histopatologia de brânquias de peixes provenientes de pisciculturas do município de itapecuru-mirim, Estado do Maranhão

The present study evaluated the microbiological water quality and tissue lesions in gills from Nile tilapia (Oreochromis niloticus) and hybrid tambacu (Colossoma macropomum female x Piaractus mesopotamicus male). For this, water and gills were collected from fish farming at six locations in Itapecuru- Mirim County, Maranhão State. Microbiological water analyses revealed contamination by total coliforms, Escherichia coli and heterotrophic bacteria. In the gills, we observed a diversity of Gram-positive and Gramnegative bacteria. The tissue lesions were: lamellar fusion, interlamellar hyperplasia, sub-epithelial edema and telangiectasia. Inflammatory lesions were not observed. Significant statistical difference (p > 0.05) was not detected when comparing different gills lesions during rainy and dry season. The correlation between lesion and pond type was statistically different (p < 0.05) for lamellar fusion and interlamellar hyperplasia which occurred more frequently at ground ponds. Regarding the frequency of lesions in the different fish species, there was statistical difference (p < 0.05), and the tambacu was more sensitive to lamellar fusion while tilapia was more sensitive for the other lesions. In relation to the sampling stations, there was statistical difference for all the gill lesions. In conclusion, tissue lesions are nonspecific and function as a defense mechanism against polluted aquatic environments, without infectious character.

Antimicrobial effects of ozonated water on the sanitization of dental instruments contaminated with E. coli, S. aureus, C. albicans, or the spores of B. atrophaeus

Objectives: Ozone has been used as an alternative method for the decontamination of water, food, equipment and instruments. The objective of this study was to evaluate the antimicrobial effects of ozonated water on the sanitization of dental instruments that were contaminated by Escherichia coli, Staphylococcus aureus, Candida albicans and the spores of Bacillus atrophaeus. Methods: A total of one hundred and twenty standardized samples of diamond dental burs were experimentally contaminated with E. coli (ATCC 25922), S. aureus (ATCC 6538) and C. albicans (ATCC 18804) and the spores of B. atrophaeus (ATCC 6633) for 30min. After the contamination, the samples were exposed to ozonated water (10mg/L O3) for 10 or 30min. The control group was composed of samples that were exposed to distilled water for 30min. After the exposure to the ozonated water, 0.1mL aliquots were seeded onto BHI agar to count the colony-forming units per milliliter (CFU/mL) of E. coli, S. aureus, and B. atrophaeus. Sabouraud dextrose agar was used to count the CFU/mL of C. albicans. The results were subjected to an analysis of variance and the Tukey test. Results: For all of the microorganisms studied, the ozonated water reduced the number of CFU/mL after 10 and 30. min of sanitization, and this microbial reduction was dependent on the duration of the exposure to the ozonated water. E. coli exhibited the greatest reduction in CFU/mL (2.72-3.78. log) followed by S. aureus (2.14-3.19. log), C. albicans (1.44-2.14. log) and the spores of B. atrophaeus (1.01-1.98. log). Conclusion: The ozonated water was effective in reducing the CFU of E. coli, S. aureus, C. albicans and B. atrophaeus spores, suggesting that ozonated water can be used for the sanitization of dental instruments. © 2012 King Saud Bin Abdulaziz University for Health Sciences.

Entomopathogens isolated from invasive ants and tests of their pathogenicity

Some ant species cause severe ecological and health impact in urban areas. Many attempts have been tested to control such species, although they do not always succeed. Biological control is an alternative to chemical control and has gained great prominence in research, and fungi and nematodes are among the successful organisms controlling insects. This study aimed to clarify some questions regarding the biological control of ants. Invasive ant species in Brazil had their nests evaluated for the presence of entomopathogens. Isolated entomopathogens were later applied in colonies of Monomorium floricola under laboratory conditions to evaluate their effectiveness and the behavior of the ant colonies after treatment. The entomopathogenic nematodes Heterorhabditis sp. and Steinernema sp. and the fungi Beauveria bassiana, Metarhizium anisopliae, and Paecilomyces sp. were isolated from the invasive ant nests. M. floricola colonies treated with Steinernema sp. and Heterorhabditis sp. showed a higher mortality of workers than control. The fungus Beauveria bassiana caused higher mortality of M. floricola workers. However, no colony reduction or elimination was observed in any treatment. The defensive behaviors of ants, such as grooming behavior and colony budding, must be considered when using fungi and nematodes for biological control of ants. Copyright © 2012 Maria Fernanda Miori de Zarzuela et al.

Validation of microbiological assay for determination of cefuroxime in injectable preparations

The validation of a microbiological assay, applying agar diffusion method for determination of the active of cefuroxime in power for injection, is described. Using a strain of Micrococcus luteus ATCC 9341 as the test organism, cefuroxime was measured in concentrations ranging from 30.0 to 120.0 μg/mL. The method validation showed that it is linear (r = 0.9999), precise (relative standard deviation = 0.37%) and accurate (it measured the added quantities). Microbiological assay is satisfactory for quantitation of cefuroxime in powder for injection and the validity of the proposed bioassay, which is a simple and a useful alternative methodology for cefuroxime determination in routine quality control.

Mineral water: A microbiological approach

The microbiological quality of bottled mineral water of various domestic brands sold in Brazil was investigated, with particular focus on the heterotrophic plate count (HPC). Neither total coliforms nor Escherichia coli were found in any 1.5 L bottle samples. Total coliforms were found in 2.9% of the small bottles, while in 20 L bottles the presence of total coliforms and E. coli was demonstrated in 15.5 and 2.4% of samples, respectively. Pseudomonas aeruginosa was detected in 4.3, 4.5 and 9.5% of small, 1.5 and 20 L bottles, respectively. In 36.4% of the samples of 1.5 L bottles, the HPC was above 500 cfu/mL. This percentage of samples with an HPC above 500 cfu/mL increased to 52.0 and 61.9% in small and 20 L bottles, respectively. Higher contamination by total coliforms, E. coli, P. aeruginosa and HPCs occurred in 20 L bottles. In conclusion, several samples in this study were outside the international quality standard for mineral water and the large number of samples with high HPCs shows that more work must be done on the use of HPC in mineral water and the damaging effects that these microorganisms may cause to humans. The bottled mineral water was confirmed as a particularly important public health problem, due to the poor microbiological quality of the products that are marketed. © IWA Publishing 2012.