Microbial culture and checkerboard DNA-DNA hybridization assessment of bacteria in root canals of primary teeth pre- and post-endodontic therapy with a calcium hydroxide/chlorhexidine paste

Aim. To investigate the root canal microbiota of primary teeth with apical periodontitis and the in vivo antimicrobial effects of a calcium hydroxide/chlorhexidine paste used as root canal dressing. Design. Baseline samples were collected from 30 root canals of primary teeth with apical periodontitis. Then, the root canals were filled with a calcium hydroxide paste containing 1% chlorhexidine for 14 days and the second bacteriologic samples were taken prior to root canal filling. Samples were submitted to microbiologic culture procedure to detect root canal bacteria and processed for checkerboard DNA-DNA hybridization. Results. Baseline microbial culture revealed high prevalence and cfu number of anaerobic, black-pigmented bacteroides, Streptococcus, and aerobic microorganisms. Following root canal dressing, the overall number of cfu was dramatically diminished compared to initial contamination (P < 0.05), although prevalence did not change (P > 0.05). Of 35 probes used for checkerboard DNA-DNA hybridization, 31 (88.57%) were present at baseline, and following root canal dressing, the number of positive probes reduced to 13 (37.14%). Similarly, the number of bacterial cells diminished folowing application of calcium hydroxide/chlorhexidine root canal dressing (P = 0.006). Conclusion. Apical periodontitis is caused by a polymicrobial infection, and a calcium hydroxide/chlorhexidine paste is effective in reducing the number of bacteria inside root canals when applied as a root canal dressing.

Dynamics of Aeromonas species isolated from wastewater treatment system

Aeromonas are widely distributed in the aquatic environment, and are considered to be emerging organisms that can produce a series of virulence factors. The present study was carried out in a sanitary sewage stabilization pond treatment system, located in Lins, State of Sao Paulo, Brazil. Most probable number was applied for estimation of the genus Aeromonas. Colony isolation was carried out on blood agar ampicillin and confirmed by biochemical characterization. Aeromonas species were isolated in 72.4% of influent samples, and in 55.2 and 48.3% of effluent from anaerobic and facultative lagoons, respectively. Thirteen Aeromonas species were isolated, representing most of the recognized species of these organisms. Even though it was possible to observe a tendency of decrease, total elimination of these organisms from the studied system was not achieved. Understanding of the pathogenic organism`s dynamics in wastewater treatment systems with a reuse potential is especially important because of the risk it represents.

Development and Evaluation of a Novel pH Indicator Biodegradable Film Based on Cassava Starch

A pH indicator film based on cassava starch plasticized with sucrose and inverted sugar and incorporated with grape and spinach extracts as pH indicator sources (anthocyanin and chlorophyll) has been developed, and its packaging properties have been assessed. A second-order central composite design (2(2)) with three central points and four star points was used to evaluate the mechanical properties (tensile strength, tensile strength at break, and elongation at break percentage), moisture barrier, and microstructure of the films, and its potential as a pH indicator packaging. The films were prepared by the casting technique and conditioned under controlled conditions (75% relative humidity and 23 degrees C), at least 4 days before the analyses. The materials were exposed to different pH solutions (0, 2, 7, 10, and 14) and their color parameters (L*, a*, b*, and haze) were measured by transmittance. Grape and spinach extracts have affected the material characterization. Film properties (mechanical properties and moisture barrier) were strongly influenced by extract concentration presenting lower results than for the control. Films containing a higher concentration of grape extract presented a greater color change at different pH`s suggesting that anthocyanins are more effective as pH indicators than chlorophyll or the mixture of both extracts. (C) 2010 Wiley Periodicals, Inc. J Appl Polym Sci 120: 1069-1079,2011

Tibouchina pulchra (Cham.) Cogn., a native Atlantic Forest species, as a bio-indicator of ozone: Visible injury

Tibouchina pulchra saplings were exposed to carbon filtered air (CF), ambient non-filtered air (NF) and ambient non-filtered air + 40 ppb ozone (NF + O-3) 8 h per day during two months. The AOT40 values at the end of the experiment were 48, 910 and 12,895 ppb h(-1), respectively, for the three treatments. After 25 days of exposure (AOT40=3871 ppb h(-1)), interveinal red stippling appeared in plants in the NF + O-3 chamber. In the NF chamber, symptoms were observed only after 60 days of exposure (AOT40 = 910 ppb h(-1)). After 60 days, injured leaves per plant corresponded to 19% in NF + O-3 and 1% in the NF treatment; and the average leaf area injured was 7% within the NF + O-3 and 0.2% within the NF treatment. The extent of leaf area injured (leaf injury index) was mostly explained by the accumulated exposure of ozone (r(2) = 0.89; p < 0.05). (C) 2007 Elsevier Ltd. All rights reserved.

A new type of Brazilian propolis: Prenylated benzophenones in propolis from Amazon and effects against cariogenic bacteria

Ethanol extracts of four propolis samples (E1-E4) from Manaus (Brazilian Amazon) were analysed by HPLC/DAD/ESI-MS/MS and GC/EIMS. The major constituents of E2 and E4 were analysed by NMR ((1)H and (13)C) and ESI/MS/MS. The main constituents of E2 and E4 are polyprenylated benzophenones: 7-epi-nemorosone, 7-epi-clusianone (major E4 constituents), xanthochymol and gambogenone (major E2 constituents), making up a chemical profile so far unreported for Brazilian propolis. Aristhophenone, methyl insigninone, 18-ethyloxy-17-hydroxy-17,18-dihydroscrobiculatone B, and derivatives of dimethyl weddellianone A and B, propolones, and a scrobiculatone derivative, were detected as minor constituents. Triterpenoids (beta-amyrins, beta-amyrenone, lupeol and lupenone) were ubiquitous and predominant in El and E3. The extracts E2 and E4 were highly active against the cariogenic bacteria Streptococcus mitis, Streptococcus mutans and Streptococcus salivarius. E2 was more active than E4, probably due to a higher content of 2-epi-nemorosone, while the latter was richer in di-hydroxylated compounds. (C) 2010 Elsevier Ltd. All rights reserved.

Annual profile of fecal androgen and glucocorticoid levels in free-living male American kestrels from southern mid-latitude areas

Fecal samples and behavioral data were collected at a fortnightly basis during 11 months period from free-living male American kestrels living in southeast Brazil (22 degrees S latitude). The aim was to investigate the seasonal changes in testicular and adrenal steroidogenic activity and their correlation to reproductive behaviors and environmental factors. The results revealed that monthly mean of fecal glucocorticoid metabolites in May and June were higher than those estimated in November. in parallel, monthly mean of androgen metabolites in September was higher than those from January to April and from October to November. Molt took place from January to March, whereas copulation was observed from June to October but peaked in September. Nest activity and food transfer to females occurred predominantly in October, and parental behavior was noticed only in November. Territorial aggressions were rare and scattered throughout the year. Multiple regression analysis revealed that fecal androgen levels are predicted by photoperiod and copulation, while fecal glucocorticoid levels are only predicted by photoperiod. Bivariate correlations showed that fecal androgen metabolites were positively correlated with fecal glucocorticoid metabolites and copulation, but negatively correlated with molt. Additionally, copulation was positively correlated with food transfer to females and nest activity, but negatively correlated with molt. These findings suggest that male American kestrels living in southeast Brazil exhibit significant seasonal changes in fecal androgen and glucocorticoid concentrations, which seem to be stimulated by decreasing daylength but not by rainfall or temperature. (C) 2009 Elsevier Inc. All rights reserved.

Influence of Radiotransmitters on Fecal Glucocorticoid Levels of Free-Ranging Male American Kestrels

Although radiotelemetry is considered a valuable technique for ornithological field studies, several assumptions have been made about the impact that transmitters may have on the estimation of behavioral, ecological, and reproductive parameters. To assess the potential effects of backpack radiotransmitters, we captured and assigned 8 male American kestrels (Falco sparverius) into 2 groups: radiotagged (n = 6) and control individuals (leg-banded, n = 2). Thereafter, we collected feces approximately 2 hours after capture (day -1), and subsequently during days 0 (releasing day), 4, 7, 15, 30, 40, and 55. Prior to fecal analysis, we validated the corticosterone enzyme immunoassay using standard procedures (e. g., parallelism, dose-response curve), and we confirmed physiological significance of fecal glucocorticoid metabolites through adrenocorticotropin challenge, which induced an increase of 4-fold (446.10 +/- 60.73 ng/g) above baseline (114.27 +/- 15.23 ng/g) within 4 hours (P < 0.001). Both groups exhibited a significant increase in fecal glucocorticoids during day 0 (P < 0.001), but concentrations returned to preattachment values within 4 days. Fecal glucocorticoid concentrations did not differ between samples of radiotagged and leg-banded kestrels (P > 0.05). In spite of the small number of monitored subjects, these findings suggested that radiotransmitters did not affect adrenocortical activity in these male American kestrels. (JOURNAL OF WILDLIFE MANAGEMENT 73(5): 772-778; 2009)

Tyrosine hydroxylase immunoreactivity as indicator of sympathetic activity: simultaneous evaluation in different tissues of hypertensive rats

Burgi K, Cavalleri MT, Alves AS, Britto LRG, Antunes VR, Michelini LC. Tyrosine hydroxylase immunoreactivity as indicator of sympathetic activity: simultaneous evaluation in different tissues of hypertensive rats. Am J Physiol Regul Integr Comp Physiol 300: R264-R271, 2011. First published December 9, 2010; doi: 10.1152/ajpregu.00687.2009.-Vasomotor control by the sympathetic nervous system presents substantial heterogeneity within different tissues, providing appropriate homeostatic responses to maintain basal/stimulated cardiovascular function both at normal and pathological conditions. The availability of a reproducible technique for simultaneous measurement of sympathetic drive to different tissues is of great interest to uncover regional patterns of sympathetic nerve activity (SNA). We propose the association of tyrosine hydroxylase immunoreactivity (THir) with image analysis to quantify norepinephrine (NE) content within nerve terminals in arteries/arterioles as a good index for regional sympathetic outflow. THir was measured in fixed arterioles of kidney, heart, and skeletal muscle of WistarKyoto rats (WKY) and spontaneously hypertensive rats (SHR) (123 +/- 2 and 181 +/- 4 mmHg, 300 +/- 8 and 352 +/- 8 beats/min, respectively). There was a differential THir distribution in both groups: higher THir was observed in the kidney and skeletal muscle (similar to 3-4-fold vs. heart arterioles) of WKY; in SHR, THir was increased in the kidney and heart (2.4- and 5.3-fold vs. WKY, respectively) with no change in the skeletal muscle arterioles. Observed THir changes were confirmed by either: 1) determination of NE content (high-performance liquid chromatography) in fresh tissues (SHR vs. WKY): +34% and +17% in kidney and heart, respectively, with no change in the skeletal muscle; 2) direct recording of renal (RSNA) and lumbar SNA (LSNA) in anesthetized rats, showing increased RSNA but unchanged LSNA in SHR vs. WKY. THir in skeletal muscle arterioles, NE content in femoral artery, and LSNA were simultaneously reduced by exercise training in the WKY group. Results indicate that THir is a valuable technique to simultaneously evaluate regional patterns of sympathetic activity.

Vibrios dominate as culturable nitrogen-fixing bacteria of the Brazilian coral Mussismilia hispida

Taxonomic characterization was performed on the putative N-2-fixing microbiota associated with the coral species Mussismilia hispida, and with its sympatric species Palythoa caribaeorum, P. variabilis, and Zoanthus solanderi, off the coast of Sao Sebastiao (Sao Paulo State, Brazil). The 95 isolates belonged to the Gammaproteobacteria according to the 16S rDNA gene sequences. In order to identify the isolates unambiguously, pyrH gene sequencing was carried out. The majority of the isolates (n = 76) fell within the Vibrio core group, with the highest gene sequence similarity being towards Vibrio harveyi and Vibrio alginolyticus. Nineteen representative isolates belonging to V. harveyi (n = 7), V. alginolyticus (n = 8), V. campbellii (n = 3), and V parahaemolyticus (n = 1) were capable of growing six successive times in nitrogen-free medium and some of them showed strong nitrogenase activity by means of the acetylene reduction assay (ARA). It was concluded that nitrogen fixation is a common phenotypic trait among Vibrio species of the core group. The fact that different Vibrio species can fix N, might explain why they are so abundant in the mucus of different coral species. (C) 2008 Published by Elsevier GmbH.

Occurrence of herpes simplex virus 1 and three periodontal bacteria in patients with chronic periodontitis and necrotic pulp

Viral and bacterial associations appear to be implicated in the development of periodontal infections. Little information is available describing the periodontopathic agents in root canals with necrotic pulp. In this study, the occurrence and the combinations among herpes simplex virus type 1 (HSV-1) and Dialister pneumosintes, Tannerella forsythia.. and Treponema denticola in patients with chronic periodontitis and necrotic pulp were evaluated. Clinical samples from healthy subjects and patients with periodontal or pulp infections were analyzed using a nested polymerase chain reaction PCR to detect HSV and PCR to detect the 3 periodontal bacteria. The presence of Tannerella forsythia and Treponema denticola was observed in healthy, periodontitis, and necrotic pulp patients. HSV was observed in periodontitis and necrotic pulp patients, and no healthy subject harbored D. pneumosintes or HSV. The occurrence of Tannerella forsythia was not statistically significant in patients with necrotic pulp (P = 0.704). Periodontal bacteria were observed varying from 10.3% to 20.7% in periodontitis and necrotic pulp patients. The presence of Treponema denticola - HSV association was predominant in patients showing necrotic pulp (24.1%); however, HSV alone was observed in one patient with periodontitis and in another patient with necrotic pulp. The presence of double association among bacteria or bacteria - HSV could indicate a role in both periodontitis and necrotic pulp, and Tannerella forsythia - Treponenta denticola - HSV and Tannerella forsythia - D. pneumosintes - Treponema denticola - HSV associations might be important in periodontitis.

Screening for endophytic nitrogen-fixing bacteria in Brazilian sugar cane varieties used in organic farming and description of Stenotrophomonas pavanii sp. nov.

A Gram-negative, rod-shaped, non-spore-forming and nitrogen-fixing bacterium, designated ICB 89(T), was isolated from stems of a Brazilian sugar cane variety widely used in organic farming. 16S rRNA gene sequence analysis revealed that strain ICB 89(T) belonged to the genus Stenotrophomonas and was most closely related to Stenotrophomonas maltophilia LMG 958(T), Stenotrophomonas rhizophila LMG 22075(T), Stenotrophomonas nitritireducens L2(T), [Pseudomonas] geniculata ATCC 19374(T), [Pseudomonas] hibiscicola ATCC 19867(T) and [Pseudomonas] beteli ATCC 19861(T). DNA-DNA hybridization together with chemotaxonomic data and biochemical characteristics allowed the differentiation of strain ICB 89(T) from its nearest phylogenetic neighbours. Therefore, strain ICB 89(T) represents a novel species, for which the name Stenotrophomonas pavanii sp. nov. is proposed. The type strain is ICB 89(T) (=CBMAI 564(T) =LMG 25348(T)).

Presence of periodontopathic bacteria in coronary arteries from patients with chronic periodontitis

In this study the presence of periodontopathic pathogens in atheromatous plaques removed from coronary arteries of patients with chronic periodontitis and periodontally healthy subjects by PCR was detected. Our results indicate a significant association between the presence of Porphyromonas gingivalis and atheromas, and the periodontal bacteria in oral biofilm may find a way to reach arteries. (C) 2010 Elsevier Ltd. All rights reserved.

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

Quantitative detection of periodontopathic bacteria in atherosclerotic plaques from coronary arteries

Oral pathogens, including periodontopathic bacteria, are thought to be aetiological factors in the development of cardiovascular disease. In this study, the presence of Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum-periodonticum-simiae group, Porphyromonas gingivalis, Prevotella intermedia, Prevotella nigrescens and Tannerella forsythia in atheromatous plaques from coronary arteries was determined by real-time PCR. Forty-four patients displaying cardiovascular disease were submitted to periodontal examination and endarterectomy of coronary arteries. Approximately 60-100 mg atherosclerotic tissue was removed surgically and DNA was obtained. Quantitative detection of periodontopathic bacteria was performed using universal and species-specific TaqMan probe/primer sets. Total bacterial and periodontopathic bacterial DNA were found in 94.9 and 92.3 %, respectively, of the atheromatous plaques from periodontitis patients, and in 80.0 and 20.0%, respectively, of atherosclerotic tissues from periodontally healthy subjects. All periodontal bacteria except for the F. nucleatum-periodonticum-simiae group were detected, and their DNA represented 47.3 % of the total bacterial DNA obtained from periodontitis; patients. Porphyromonas gingivalis, A. actinomycetemcomitans and Prevotella intermedia were detected most often. The presence of two or more periodontal species could be observed in 64.1 % of the samples. In addition, even in samples in which a single periodontal species was detected, additional unidentified microbial DNA could be observed. The significant number of periodontopathic bacterial DNA species in atherosclerotic tissue samples from patients with periodontitis suggests that the presence of these micro-organisms in coronary lesions is not coincidental and that they may in fact contribute to the development of vascular diseases.

Screening of bacteria to produce polyhydroxyalkanoates from xylose

Although xylose is a major constituent of lignocellulosic feedstock and the second most abundant sugar in nature, only 22% of 3,152 screened bacterial isolates showed significant growth in xylose in 24 h. Of those 684, only 24% accumulated polyhydroxyalkanoates after 72 h. A mangrove isolate, identified as Bacillus sp. MA3.3, yielded the best results in literature thus far for Gram-positive strains in experiments with glucose and xylose as the sole carbon source. When glucose or xylose were supplied, poly-3-hydroxybutyrate (PHB) contents of cell dry weight were, respectively, 62 and 64%, PHB yield 0.25 and 0.24 g g(-1) and PHB productivity (P(PHB)) 0.10 and 0.06 g l(-1) h(-1). This 40% P(PHB) difference may be related to the theoretical ATP production per 3-hydroxybutyrate (3HB) monomer calculated as 3 mol mol(-1) for xylose, less than half of the ATP/3HB produced from glucose (7 mol mol(-1)). In PHB production using sugar mixtures, all parameters were strongly reduced due to carbon catabolite repression. PHB production using Gram-positive strains is particularly interesting for medical applications because these bacteria do not produce lipopolysaccharide endotoxins which can induce immunogenic reactions. Moreover, the combination of inexpensive substrates and products of more value may lead to the economical sustainability of industrial PHB production.