Bacterial colonization in hemodialysis temporary dual lumen catheters: A prospective study

Aims. The use of hemodialysis temporary dual-lumen catheters is often complicated by infections, which may be a significant cause of death among patients with end stage renal disease (ESRD). The aim of this study was to assess the incidence of bacteremia and bacterial colonization related to non-tunneled, non-cuffed, dual-lumen temporary catheters in patients with ESRD submitted to hemodialysis. Methods. This study included 29 patients with ESRD. After catheter implantation, patients were monitored throughout the period of catheter permanence by means of blood samples collected weekly from a peripheral vein. Bacteria were isolated and identified according to CLSI recommendations. When catheters were removed for any reason, their tips were evaluated microbiologically. Results. A total of 194 blood samples from the 29 patients implanted with 55 catheters were analyzed. Of these, 15.5% (30 samples) demonstrated bacterial growth, principally Staphylococcus epidermidis (64.5%). Twenty patients (68.9%) presented at least one positive blood culture during follow-up. The median time for catheter colonization was 18.5 days (95% CI: 16.8-30.3). Of the 55 catheters implanted, 28 (50.9%) showed bacterial colonization, corresponding to 23.4 episodes/1000 catheter/days and 9.2 episodes of bacteremia /1000 catheter/days. Fifteen of 28 catheter tips analyzed showed bacterial growth (53.5%). In 14 of these (93.3%), there was agreement between the isolates from the catheter tip and blood cultures. Of 24 episodes of positive blood cultures from 20 different patients in 17 episodes (70.8%), the patients showed no clinical signs or symptoms of bacteremia. Conclusions. The high incidence of catheter colonization, the correlation between blood and catheter tip cultures, and the occurrence of frequent cases of asymptomatic bacteremia justify the proposal of routine peripheral blood collections to monitor patients undergoing hemodialysis with temporary dual-lumen catheters.

Bacterial leakage along the implant-abutment interface of premachined or cast components

In recent clinical studies, contamination of the inner parts of dental implants through bacterial penetration along the implant components has been observed. The aim of the present in-vitro study was to investigate leakage of Fusobacterium. nucleatum through the interface between implants and premachined or cast abutments. Both premachined (n = 10) and cast (n = 10) implant abutment assemblies were inoculated with 3.0 mu L of microbial inoculum. The assemblies were completely immersed in 5.0 mL of tryptic soy broth culture medium to observe leakage at the implant-abutment interface after 14 days of anaerobic incubation. Bacterial growth in the medium, indicative of microbial leakage, was found only in 1 out of 9 samples (11.1%) in each group. Both premachined and cast abutments connected to external hexagonal implants provide low percentages of bacterial leakage through the interface in in vitro unloaded conditions if the manufacturer`s instructions and casting procedures are properly followed.

Antibacterial activity of root canal filling materials for primary teeth: zinc oxide and eugenol cement, Calen paste thickened with zinc oxide, Sealapex and EndoREZ

This study evaluated in vitro the antibacterial activity of 4 root canal filling materials for primary teeth - zinc oxide and eugenol cement (ZOE), Calen paste thickened with zinc oxide (Calen/ZO), Sealapex sealer and EndoREZ sealer - against 5 bacterial strains commonly found in endodontic infections (Kocuria rhizophila, Enterococcus faecalis, Streptococcus mutans, Escherichia coli and Staphylococcus aureus) using the agar diffusion test (agar-well technique). Calen paste, 1% chlorhexidine digluconate (CHX) and distilled water served as controls. Seven wells per dish were made at equidistant points and immediately filled with the test and control materials. After incubation of the plates at 37oC for 24 h, the diameter of the zones of bacterial growth inhibition produced around the wells was measured (in mm) with a digital caliper under reflected light. Data were analyzed statistically by analysis of variance and Tukey's post-hoc test (?=0.05). There were statistically significant differences (p<0.0001) among the zones of bacterial growth inhibition produced by the different materials against all target microorganisms. K. rhizophila was inhibited more effectively (p<0.05) by ZOE, while Calen/ZO had its highest antibacterial activity against E. faecalis (p<0.05). S. mutans was inhibited by Calen/ZO, Sealapex and ZOE in the same intensity (p>0.05). E. coli was inhibited more effectively (p<0.05) by ZOE, followed by Calen/ZO and Sealapex. Calen/ZO and ZOE were equally effective (p>0.05) against S. aureus, while Sealapex had the lowest antibacterial efficacy (p<0.05) against this microorganism. EndoREZ presented antibacterial activity only against K. rhizophila and S. aureus. The Calen paste and Calen/ZO produced larger zones of inhibition than 1% CHX when the marker microorganism was E faecalis. In conclusion, the in vitro antibacterial activity of the 4 root canal filling materials for primary teeth against bacterial strains commonly found in endodontic infections can be presented in a decreasing order of efficacy as follows: ZOE>Calen/ZO>Sealapex>EndoREZ.

Blood Meal-Derived Heme Decreases ROS Levels in the Midgut of Aedes aegypti and Allows Proliferation of Intestinal Microbiota

The presence of bacteria in the midgut of mosquitoes antagonizes infectious agents, such as Dengue and Plasmodium, acting as a negative factor in the vectorial competence of the mosquito. Therefore, knowledge of the molecular mechanisms involved in the control of midgut microbiota could help in the development of new tools to reduce transmission. We hypothesized that toxic reactive oxygen species (ROS) generated by epithelial cells control bacterial growth in the midgut of Aedes aegypti, the vector of Yellow fever and Dengue viruses. We show that ROS are continuously present in the midgut of sugar-fed (SF) mosquitoes and a blood-meal immediately decreased ROS through a mechanism involving heme-mediated activation of PKC. This event occurred in parallel with an expansion of gut bacteria. Treatment of sugar-fed mosquitoes with increased concentrations of heme led to a dose dependent decrease in ROS levels and a consequent increase in midgut endogenous bacteria. In addition, gene silencing of dual oxidase (Duox) reduced ROS levels and also increased gut flora. Using a model of bacterial oral infection in the gut, we show that the absence of ROS resulted in decreased mosquito resistance to infection, increased midgut epithelial damage, transcriptional modulation of immune-related genes and mortality. As heme is a pro-oxidant molecule released in large amounts upon hemoglobin degradation, oxidative killing of bacteria in the gut would represent a burden to the insect, thereby creating an extra oxidative challenge to the mosquito. We propose that a controlled decrease in ROS levels in the midgut of Aedes aegypti is an adaptation to compensate for the ingestion of heme.

Evaluation of two semi-selective media to detect Curtobacterium flaccumfaciens pv. flaccumfaciens in bean seeds

Evaluation of two semi-selective media to detect Curtobacterium flaccumfaciens pv. flaccumfaciens in bean seeds This study aimed to compare the effectiveness of the semi-selective MSCFF and modified CNS culture media in detecting Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff) in bean seeds, using the streak and spread plate techniques. Four 500 g subsamples, obtained from two samples of bean seeds, were immersed in 600 mL of sterile distilled water for 18 h at 5 degrees C. Suspensions were picked and transferred to plates with both culture media. Plates were then incubated at 28 degrees C, and bacterial growth on both media was evaluated 72 and 144 hours later, compared to the growth of a Cff reference strain. Both media revealed the presence of Cff colonies. Typical colonies were isolated for PCR analyses and pathogenicity tests on tobacco leaves. A characteristic Cff growth on MSCFF medium was observed for the seed samples, for the two plate techniques used, in both evaluations. On the modified CNS culture medium, the bacterial growth was only detected in seed samples after 144 hours of incubation, regardless of the plate technique used. The results showed Cff grew faster on the MSCFF semi-selective culture medium. Bacterial isolates tested were identified as Cff by both PCR analyses and a positive tobacco hypersensitivity reaction.

Short communication: Identification of subclinical cow mastitis pathogens in milk by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Subclinical mastitis is a common and easily disseminated disease in dairy herds. Its routine diagnosis via bacterial culture and biochemical identification is a difficult and time-consuming process. In this work, we show that matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) allows bacterial identification with high confidence and speed (1 d for bacterial growth and analysis). With the use of MALDI-TOF MS, 33 bacterial culture isolates from milk of different dairy cows from several farms were analyzed, and the results were compared with those obtained by classical biochemical methods. This proof-of-concept case demonstrates the reliability of MALDI-TOF MS bacterial identification, and its increased selectivity as illustrated by the additional identification of coagulase-negative Staphylococcus species and mixed bacterial cultures. Matrix-assisted laser desorption-ionization mass spectrometry considerably accelerates the diagnosis of mastitis pathogens, especially in cases of subclinical mastitis. More immediate and efficient animal management strategies for mastitis and milk quality control in the dairy industry can therefore be applied.

Direct and transdentinal antibacterial activity of chlorhexidine

Purpose: To evaluate the antibacterial effect of different chlorhexidine (CHX) concentrations against Streptococcus mutans using the agar-diffusion method with and without human dentin discs placed between the bacteria and the test substances. Methods: For the direct application (agar-well technique), a base layer containing 15 mL of BHI agar and 300 mu L. of S. mutans inoculum (10(9) cfu/mL) was prepared in Petri dishes. Six wells per dish were made at equidistant points and immediately filled with CHX gels (0.12%, 0.2%, 1% and 2%), 35% phosphoric acid and pure natrosol (n=6 wells/substance). Paper discs soaked in sterile distilled water served as control group (n=6). For the indirect application (transdentinal diffusion), 0.2 mm- and 0.5 mm-thick human dentin discs (36 discs/thickness) had the hydraulic conductance determined, which allowed the homogeneous allocation of them to the experimental and control groups. The discs were placed at equidistant points on the Petri dishes containing BHI with the S. mutans inoculum (six discs per dish; one per substance) with the pulpal side in contact with the bacteria. In the discs treated with CHX gels, dentin surface was etched with H(3)PO(4) and rinsed with distilled water before CHX gel application for 1 minute. After both direct and indirect application, the dishes were incubated for 24 hours and the bacterial growth inhibition zones formed around the wells and dentin discs were measured. Data were analyzed statistically by the non-parametric Kruskal-Wallis and Mann-Whitney tests at 5% significance level. Results: In the direct test, all CHX concentrations presented a dose-dependent antibacterial activity against S. mutans. In the indirect test, there were statistically significant differences (P< 0.05) among all groups and the largest microbial growth inhibition zones were observed when 2% CHX was applied on 0.2 mm-thick discs (P< 0.05). It was concluded that all evaluated CHX gels exhibited both direct and transdentinal antibacterial activity against S. mutans. This effect of CHX was strongly influenced by the CHX concentration as well as the dentin barrier thickness. (Am J Dent 2010;23:255-259).

Evaluation of the pH and electrical conductivity in milk from jersey cows during the first month of lactation

In order to assess the influence of the colostrum period on pH and, electrical conductivity, we collected 418 milk samples from 127 Jersey cows. The samples were collected from healthy udders that did not present any bacterial growth in the microbiological examination. They were divided into eight groups as follows < 1/2 day; 1/2 and 1 degrees day; 2 degrees day; 3 degrees day; 4 degrees and 5 degrees day; 6 degrees and 7 degrees day; 8 degrees to 15 degrees day; 16 degrees to 30 degrees days of lactation. The samples were collected before milking and the following analyses were conducted: pH, electrical conductivity. In the first 24 hours of lactation, there was an reduction in electrical conductivity value, associated with an increase in pH value. We observed that transition of secretion from colostrum to milk, occurs during the first week of lactation; from 6(rd) day of lactation for pH value and 3(th) day for electrical conductivity value. We recommend the use the following figures as normal ranges for the first 24 hours of lactation (colostrum period): pH <= 6,51 and electrical conductivity <= 6,33 mS/cm; while for the interval between 2(nd) and 7(th) days of lactation (transition from colostrum to milk) we suggest the use of the values as normal ranges: pH <= 6,66 and electrical conductivity <= 5,93 mS/cm.

Apoptosis of macrophages during pulmonary Mycobacterium bovis infection: correlation with intracellular bacillary load and cytokine levels

P>Apoptosis of macrophages infected with pathogenic mycobacteria is an alternative host defence capable of removing the environment supporting bacterial growth. In this work the influence of virulence and bacterial load on apoptosis of alveolar macrophages during the initial phase of infection by Mycobacterium bovis was investigated. BALB/c mice were infected intratracheally with high or low doses of the virulent (ATCC19274) or attenuated (bacillus Calmette-Guerin Moreau) strains of M. bovis. The frequency of macrophage apoptosis, the growth of mycobacteria in macrophages, and the in situ levels of the cytokines tumour necrosis factor-alpha (TNF-alpha), interleukin-10 (IL-10) and IL-12 and of the anti-apoptotic protein Bcl-2 were measured at day 3 and day 7 post-infection. An increase of macrophage apoptosis was observed after infection with both strains but the virulent strain induced less apoptosis than the attenuated strain. On the 3rd day after infection with the virulent strain macrophage apoptosis was reduced in the high-dose group, while on the 7th day post-infection macrophage apoptosis was reduced in the low-dose group. Inhibition of apoptosis was correlated with increased production of IL-10, reduced production of TNF-alpha and increased production of Bcl-2. In addition, the production of IL-12 was reduced at points where the lowest levels of macrophage apoptosis were observed. Our results indicate that virulent mycobacteria are able to modulate macrophage apoptosis to an extent dependent on the intracellular bacterial burden, which benefits its intracellular growth and dissemination to adjacent cells.

Influence of organic fertilization on the number of culturable diazotrophic endophytic bacteria isolated from sugarcane

The numbers of culturable diazotrophic endophytic bacteria (CDEB) from roots stems and leaves of sugarcane submitted to organic inorganic or no fertilization were compared In order to determine the size of the N(2) fixing populations the Most Probable Number technique (MPN) was used The quantification of diazotrophic bacteria by using the acetylene reduction assay (ARA) was more accurate than observing the bacterial growth in the vials to confirm N(2) fixing capability the detection of gene nifH was performed on a sample of 105 Isolated bacteria The production of extracellular enzymes involved in the penetration of the plants by the bacteria was also studied The results showed that organic fertilization enhances the number of CDEB when compared with conventional fertilization used throughout the growing season The maximum number of bacteria was detected in the roots Roots and stems presented the greatest number of CDEB in the middle of the cropping season and in leaves numbers varied according to the treatment Using two pairs of primers and two different methods the nifH gene was found in 104 of the 105 tested isolates Larger amounts of pectinase were released by isolates from sugarcane treated with conventional fertilizers (66%) whereas larger amounts of cellulase were released by strains isolated from sugarcane treated with organic fertilizers (80%) (C) 2010 Elsevier Masson SAS All rights reserved

Culture and exopolysaccharide production from sugarcane molasses by Gordonia polyisoprenivorans CCT 7137, isolated from contaminated groundwater in Brazil

Gordonia polyisoprenivorans CCT 7137 was isolated from groundwater contaminated with leachate in an old controlled landfill (Sauo Paulo, Brazil), and cultured in GYM medium at different concentrations of sugarcane molasses (2%, 6%, and 10%). The strain growth was analyzed by monitoring the viable cell counts (c.f.u. mL(-1)) and optical density and EPS production was evaluated at the end of the exponential phase and 24 h after it. The analysis of the viable cell counts showed that the medium that most favored bacterial growth was not the one that favored EPS production. The control medium (GYM) was the one that most favored the strain growth, at the maximum specific growth rate of 0.232 h(-1). Differences in bacterial growth when cultured at three different concentrations of molasses were not observed. Production of EPS, in all culture media used, began during the exponential phase and continued during the growth stationary phase. The highest total EPS production, after 24 h of stationary phase, was observed in 6% molasses medium (172.86 g L(-1)) and 10% (139.47 g L(-1)) and the specific total EPS production was higher in 10% molasses medium (39.03 x10(-11)g c.f.u.(-1)). After the exponential phase, in 2%, 6%, and 10% molasses media, a higher percentage of free exopolysaccharides (EPS) was observed, representing 88.4%, 62.4%, and 64.2% of the total, respectively. A different result was observed in pattern medium, which presented EPS made up of higher percentage of capsular EPS (66.4% of the total). This work is the first study on EPS production by G. polyisoprenivorans strain in GYM medium and in medium utilizing sugarcane molasses as the sole nutrient source and suggests its potential use for EPS production by G. polyisoprenivorans CCT 7137 aiming at application in biotechnological processes.

Preparation and characterization of the novels terpolymers of poly-{trans-[RuCl(2)(vpy)(4)]-styrene-divinylbenzene} and styrene-divinylbenzene-vinylpiridine impregnated with silver nanoparticles

This paper reports the preparation and characterization of poly-{trans-[RuCl(2)(vpy)(4)]-styrene-divinylbenzene} and styrene-divinylbenzene-vinylpiridine filled with nanosilver. Theses materials were synthesized by non aqueous polymerization through a chemical reaction using benzoyl peroxide as the initiator. The nanosilver was obtained from chemical reduction using NaBH(4) as reducing agent and sodium citrate as stabilizer. The nanometric dimension of nanosilver was monitored by UV-visible and confirmed through TEM. The morphology was characterized by SEM and the thermal properties were done by TGA and DSC. The antimicrobial action of the polymers impregnated with nanosilver was evaluated using both microorganisms, Staphylococcus aureus and Escherichia coli. The antimicrobial activity of the poly-{trans-[RuCl(2)(vpy)(4)]-styrene-divinylbenzene} filled with nanosilver was confirmed by the presence of an inhibition halo of the bacterial growth in seeded culture media, but was not confirmed to the styrene-divinylbenzene-vinylpiridine. The present work suggest that trans - [RuCl(2)(vpy)(4)] complex facilitate the release of silver ion from the media.

Bacterial soil community in a Brazilian sugarcane field

The assessment of bacterial communities in soil gives insight into microbial behavior under prevailing environmental conditions. In this context, we assessed the composition of soil bacterial communities in a Brazilian sugarcane experimental field. The experimental design encompassed plots containing common sugarcane (variety SP80-1842) and its transgenic form (IMI-1 - imazapyr herbicide resistant). Plants were grown in such field plots in a completely randomized design with three treatments, which addressed the factors transgene and imazapyr herbicide application. Soil samples were taken at three developmental stages during plant growth and analyzed using 16S ribosomal RNA (rRNA)-based PCR-denaturing gradient gel electrophoresis (PCR-DGGE) and clone libraries. PCR-DGGE fingerprints obtained for the total bacterial community and specific bacterial groups - Actinobacteria, Alphaproteobacteria and Betaproteobacteria - revealed that the structure of these assemblages did not differ over time and among treatments. Nevertheless, slight differences among 16S rRNA gene clone libraries constructed from each treatment could be observed at particular cut-off levels. Altogether, the libraries encompassed a total of eleven bacterial phyla and the candidate divisions TM7 and OP10. Clone sequences affiliated with the Proteobacteria, Actinobacteria, Firmicutes and Acidobacteria were, in this order, most abundant. Accurate phylogenetic analyses were performed for the phyla Acidobacteria and Verrucomicrobia, revealing the structures of these groups, which are still poorly understood as to their importance for soil functioning and sustainability under agricultural practices.

Genetic diversity and plant-growth related features of Burkholderia spp. from sugarcane roots

Brazil is the largest sugarcane producer in the world, mainly due to the development of different management strategies. Recently, microbial-plant related studies revealed that bacterial isolates belonging to the genus Burkholderia are mainly associated with this plant and are responsible for a range of physiological activity. In this study, we properly evaluate the physiological activity and genetic diversity of endophytic and rhizospheric Burkholderia spp. isolates from sugarcane roots grown in the field in Brazil. In total, 39 isolates previously identified as Burkholderia spp. were firstly evaluated for the capability to fix nitrogen, produce siderophores, solubilise inorganic phosphates, produce indole-acetic acid and inhibit sugarcane phytopathogens in vitro. These results revealed that all isolates present at least two positive evaluated activities. Furthermore, a phylogenetic study was carried out using 16S rRNA and gyrB genes revealing that most of the isolates were affiliated with the Burkholderia cepacia complex. Hence, a clear separation given by endophytic or rhizospheric niche occupation was not observed. These results presented an overview about Burkholderia spp. isolates from sugarcane roots and supply information about the physiological activity and genetic diversity of this genus, given direction for further studies related to achieve more sustainable cultivation of sugarcane.

Effect of bacterial inoculation, plant genotype and developmental stage on root-associated and endophytic bacterial communities in potato (Solanum tuberosum)

Beneficial bacteria interact with plants by colonizing the rhizosphere and roots followed by further spread through the inner tissues, resulting in endophytic colonization. The major factors contributing to these interactions are not always well understood for most bacterial and plant species. It is believed that specific bacterial functions are required for plant colonization, but also from the plant side specific features are needed, such as plant genotype (cultivar) and developmental stage. Via multivariate analysis we present a quantification of the roles of these components on the composition of root-associated and endophytic bacterial communities in potato plants, by weighing the effects of bacterial inoculation, plant genotype and developmental stage. Spontaneous rifampicin resistant mutants of two bacterial endophytes, Paenibacillus sp. strain E119 and Methylobacterium mesophilicum strain SR1.6/6, were introduced into potato plants of three different cultivars (Eersteling, Robijn and Karnico). Densities of both strains in, or attached to potato plants were measured by selective plating, while the effects of bacterial inoculation, plant genotype and developmental stage on the composition of bacterial, Alphaproteobacterial and Paenibacillus species were determined by PCR-denaturing gradient gel-electrophoresis (DGGE). Multivariate analyses revealed that the composition of bacterial communities was mainly driven by cultivar type and plant developmental stage, while Alphaproteobacterial and Paenibacillus communities were mainly influenced by bacterial inoculation. These results are important for better understanding the effects of bacterial inoculations to plants and their possible effects on the indigenous bacterial communities in relation with other plant factors such as genotype and growth stage.