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PubMed1998

Intraorifice sealing of gutta-percha obturated root canals to prevent coronal microleakage.

A study was conducted to evaluate Cavit, Intermediate Restorative Material, and Super-EBA as intraorifice filling materials to prevent coronal microleakage. Root canal instrumentation and obturation was done on 74 extracted single-rooted teeth. Three and one-half millimeters of the gutta-percha was removed from the coronal aspect of the root canal and replaced with one of the three filling materials. The teeth were suspended in scintillation vials containing trypticase soy broth, and human saliva was added to the pulp chambers. Microbial penetration was detected as an increase in turbidity of the broth corresponding to bacterial growth. At the end of 90 days, the results showed that 15% of the Cavit-filled orifices leaked, whereas 35% of the Intermediate Restorative Material and Super-EBA-filled orifices leaked. The gutta-percha obturated root canals that received an intraorifice filling material leaked significantly less than the obturated, unsealed control group--all of which leaked in < 49 days.

Calcium Sulfate↗

PubMed2003

Detection of pathogenic bacteria in shellfish using multiplex PCR followed by CovaLink NH microwell plate sandwich hybridization.

Outbreak of diseases associated with consumption of raw shellfish especially oysters is a major concern to the seafood industry and public health agencies. A multiplex PCR amplification of targeted gene segments followed by DNA-DNA sandwich hybridization was optimized to detect the etiologic agents. First, a multiplex PCR amplification of hns, spvB, vvh, ctx and tl was developed enabling simultaneous detection of total Salmonella enterica serotype Typhimurium, Vibrio vulnificus, Vibrio cholerae and Vibrio parahaemolyticus from both pure cultures and seeded oysters. Amplicons were then subjected to a colorimetric CovaLink NH microwell plate sandwich hybridization using phosphorylated and biotinlylated oligonucleotide probes, the nucleotide sequences of which were located internal to the amplified DNA. The results from the hybridization with the multiplexed PCR amplified DNA exhibited a high signal/noise ratio ranging between 14.1 and 43.2 measured at 405 nm wavelength. The sensitivity of detection for each pathogen was 10(2) cells/g of oyster tissue homogenate. The results from this study showed that the combination of the multiplex PCR with a colorimetric microwell plate sandwich hybridization assay permits a specific, sensitive, and reproducible system for the detection of the microbial pathogens in shellfish, thereby improving the microbiological safety of shellfish to consumers.

Animals↗

PubMed2003

Microbiology of liver abscesses and the predictive value of abscess gram stain and associated blood cultures.

Although rare, pyogenic liver abscesses are potentially fatal. We evaluated the predictive value of Gram stain of liver abscess aspirates and temporally associated blood cultures. Gram stains detected bacteria in 79% of the liver abscesses tested. The sensitivity and specificity of Gram stain of the liver abscesses were 90% and 100% for Gram-positive cocci (GPC) and 52% and 94% for Gram-negative bacilli (GNB). The sensitivities of the blood cultures for any GPC and GNB present in the liver abscess were 30% and 39%, respectively. Although, Gram stains and blood cultures offer incomplete detection of the microbial contents of pyogenic liver abscesses, both tests should always accompany liver abscess cultures.

Bacteriological Techniques↗

PubMed2001

Incidence of Chlamydia trachomatis and other potential pathogens in neonatal conjunctivitis.

OBJECTIVE: Ocular infection in neonatology is a permanent and important health problem. To improve primary attention, prevention, and control, the study of the potential bacterial etiology of all consecutive cases of conjunctivitis was incorporated as a regular procedure in primary care from July 1995 to December 1998. MATERIALS AND METHODS: Prof. A. Posadas Hospital (Great Buenos Aires) has an average of 4294 births per year. This report analyzes the results obtained in 332 infants (age range, 0-30 d) with conjunctivitis. Clinical conjunctivitis was diagnosed in inpatients and outpatients by the same specialized staff. Isolation and characterization of bacteria were done by conventional microbiologic methods, including specific search for Neisseria gonorrhoeae and Chlamydia trachomatis. Chlamydia trachomatis was studied by antigen immunodetection and polymerase chain reaction, and genotyped by restriction fragment length polymorphism. RESULTS: Conjunctivitis had an incidence (cases per 1000 live births) of 39.6 in 1995, 25.3 in 1996, 15.4 in 1997, and 15.2 in 1998. Microbial growth was detected in 167 (50.3%) of 332 cases. Ocular C. trachomatis infection was detected in 26 cases (7.83%). Five of seven isolates in tissue cultures belonged to type E and two to type G. Bacteria from respiratory ecology were the main isolates: Haemophilus influenzae (16.9%), Streptococcus pneumoniae (12.3%), and Staphylococcus aureus (8.7%). Haemophilus influenzae isolates were not serotyped and 17.2% of them were b-lactamase producers. In 15 cases both H. influenzae and S. pneumoniae were isolated together. Of S. pneumoniae, 4.9% were oxacillin resistant. CONCLUSIONS: There has been a decline in the total number of cases of neonatal conjunctivitis, but the disease is still an important health problem. Chlamydia trachomatis also shows a decreasing profile with an incidence of (cases per 1000 live births) 4.39 in 1995, 1.85 in 1996, 1.01 in 1997, and 0.78 in 1998, and a tendency to show more incidence in spring-summer and significant accumulation of cases in babies between 7 and 9 days of age. Haemophilus influenzae alone (12.3%) or associated with S. pneumoniae (4.5%) appears as a prevalent potential bacterial pathogen. A significant accumulation of H. influenzae and S. pneumoniae cases occurs in winter. In 47.6% of cases, there was no bacterial growth. No significant seasonal differences in percentage of negative cultures or among the three-day age groups were detected. Neisseria gonorrhoeae was not found associated with ophthalmia neonatorum in this series.

Age Factors↗

PubMed2003

Marker and reporter genes: illuminating tools for environmental microbiologists.

Fluorescent and luminescent marker and reporter genes provide easily detectable phenotypes to microbial cells and are therefore valuable tools for the study of microorganisms in the environment. Although these tools are becoming widely adopted, there are still issues that remain to be solved, such as the dependence of the reporter output on the physiological status of the cell. Eventually it might be the use of marker and reporter genes themselves that will contribute towards better understanding of the physiological status of specific microbial populations in nature.

Bacteria↗

PubMed1997

Biodiversity within hot spring microbial mat communities: molecular monitoring of enrichment cultures.

We have begun to examine the basis for incongruence between hot spring microbial mat populations detected by cultivation or by 16S rRNA methods. We used denaturing gradient gel electrophoresis (DGGE) to monitor enrichments and isolates plated therefrom. At near extincting inoculum dilutions we observed Chloroflexus-like and cyanobacterial populations whose 16S rRNA sequences have been detected in the 'New Pit' Spring Chloroflexus mat and the Octopus Spring cyanobacterial mat. Cyanobacterial populations enriched from 44 to 54 degrees C and 56 to 63 degrees C samples at near habitat temperatures were similar to those previously detected in mat samples of comparable temperatures. However, a lower temperature enrichment from the higher temperature sample selected for the populations found in the lower temperature sample. Three Thermus populations detected by both DGGE and isolation exemplify even more how enrichment may bias our view of community structure. The most abundant population was adapted to the habitat temperature (50 degrees C), while populations adapted to 65 degrees C and 70 degrees C were 10(2)- and 10(4)-fold less abundant, respectively. However, enrichment at 70 degrees C favored the least abundant strain. Inoculum dilution and incubation at the habitat temperature favored the more numerically relevant populations. We enriched many other aerobic chemoorganotrophic populations at various inoculum dilutions and substrate concentrations, most of whose 16S rRNA sequences have not been detected in mats. A common feature of numerically relevant cyanobacterial, Chloroflexus-like and aerobic chemorganotrophic populations, is that they grow poorly and resist cultivation on solidified medium, suggesting plating bias, and that the medium composition and incubation conditions may not reflect the natural microenvironments these populations inhabit.

Bacteria, Aerobic↗

PubMed2004

Bacterial disease resistance in Arabidopsis through flagellin perception.

Plants and animals recognize microbial invaders by detecting pathogen-associated molecular patterns (PAMPs) such as flagellin. However, the importance of flagellin perception for disease resistance has, until now, not been demonstrated. Here we show that treatment of plants with flg22, a peptide representing the elicitor-active epitope of flagellin, induces the expression of numerous defence-related genes and triggers resistance to pathogenic bacteria in wild-type plants, but not in plants carrying mutations in the flagellin receptor gene FLS2. This induced resistance seems to be independent of salicylic acid, jasmonic acid and ethylene signalling. Wild-type and fls2 mutants both display enhanced resistance when treated with crude bacterial extracts, even devoid of elicitor-active flagellin, indicating the existence of functional perception systems for PAMPs other than flagellin. Although fls2 mutant plants are as susceptible as the wild type when bacteria are infiltrated into leaves, they are more susceptible to the pathogen Pseudomonas syringae pv. tomato DC3000 when it is sprayed on the leaf surface. Thus, flagellin perception restricts bacterial invasion, probably at an early step, and contributes to the plant's disease resistance.

Arabidopsis↗

PubMed2002

Field and pulping performances of transgenic trees with altered lignification.

The agronomic and pulping performance of transgenic trees with altered lignin has been evaluated in duplicated, long-term field trials. Poplars expressing cinnamyl alcohol dehydrogenase (CAD) or caffeate/5-hydroxy-ferulate O-methyltransferase (COMT) antisense transgenes were grown for four years at two sites, in France and England. The trees remained healthy throughout the trial. Growth indicators and interactions with insects were normal. No changes in soil microbial communities were detected beneath the transgenic trees. The expected modifications to lignin were maintained in the transgenics over four years, at both sites. Kraft pulping of tree trunks showed that the reduced-CAD lines had improved characteristics, allowing easier delignification, using smaller amounts of chemicals, while yielding more high-quality pulp. This work highlights the potential of engineering wood quality for more environmentally benign papermaking without interfering with tree growth or fitness.

Alcohol Oxidoreductases↗

PubMed1999

Case report. Kerion Celsi effectively treated with terbinafine. Characteristics of kerion Celsi in the elderly in Japan.

A 75-year-old non-working male living in Sagamihara, Kanagawa Prefecture, had erythematous plaques with scales associated with follicular pustules in the head area extending from the occipital to right temporal regions about 1 month prior to his initial visit, when hair loss increased. The diagnosis was kerion Celsi. Trichophyton rubrum was isolated from scales and tissues taken from lesions in the head. Histopathological examinations showed irregular epidermal thickening with dense cell infiltration from the dermis to subcutaneous adipose tissues. Granulomatous reactions involving neutrophils, histiocytes and giant cells were seen mainly in the hair follicles. Periodic acid-Schiff (PAS) and Grocott-positive microbial elements were detected in the horny layer, and inside and outside the hair follicles. Pustules disappeared 1 week after starting the oral treatment with terbinafine (125 mg day-1). A cure was achieved 2 weeks after starting the treatment, with only slight scales remaining. No recurrence has been observed to date. Terbinafine was thought to be very effective and safe for kerion Celsi. We reviewed 27 cases of kerion Celsi reported in patients, aged at least 70 years, in Japan and found that the major characteristics of this disease in Japan include the following: (1) female cases outnumber male cases; (2) the causative organism was T. rubrum in 14 of 27 patients (51.9%); and (3) topical application of steroids often induces this disease in patients with superficial tinea capitis.

Administration, Oral↗

PubMed2002

The characteristics of infections in crush syndrome.

OBJECTIVE: To analyze the results of clinical and bacteriological examinations of crush syndrome patients admitted to our institute after the Marmara earthquake. METHODS: Data were collected retrospectively from patients' files. Forty patients are included. Their mean age was 31.5 +/- 13.5 years and 18 were male. Their mean time under the rubble was 13.1 +/- 14.4 h. Fasciotomies were performed on 41 extremities of 30 patients. One hundred and twelve (mean 2.9 +/- 1.9 samples/patient) bacteriological samples were collected from wounds (51), blood (23), urine (25) and catheters (13). RESULTS: Microbial growth was detected in 67 samples from 38 (95%) patients. Non-fermenting Gram-negative bacilli, Gram-positive cocci, Enterobacteriaciace and yeast-like fungi were isolated in 67%, 17%, 12% and 4% of the samples, respectively. Acinetobacter (36%) and Pseudomonas aeruginosa (21%), the major bacterial isolates from wound infections, were resistant to carbapenems and sensitive to quinolones. As the hospitalization period increased, other infections supervened. Gram-positive cocci and non-fermenting Gram-negative bacilli were detected in six blood and seven catheter samples and methicillin-resistant Staphylococcus aureus was the major isolate. Nine (22%) of the patients died due to sepsis despite all supportive therapies. CONCLUSIONS: Infections are still major factors in crush syndrome-related deaths.

Adolescent↗

PubMed2003

Interactions between coagulation and inflammation.

The coagulation system evolved as a mechanism to limit the loss of vital elements from the internal milieu following mechanical injury to the circulatory system. The innate immune system developed as a rapid response system to detect and clear microbial invaders that have breached the integument of multicellular organisms. The coagulation and innate immunity systems coevolved from a common ancestral substrate early in eukaryotic development, and these 2 systems retain a highly integrated and coregulated circuitry of signals and control elements that defend the host following tissue injury and microbial invasion. Advances in the understanding of the molecular mechanisms that control clotting and inflammation have led to the successful development of recombinant human activated protein C for the treatment of human septic shock. Optimal use of this and other anticoagulant agents in sepsis will necessitate further basic research into the critical linkage between coagulation and innate immunity.

Blood Coagulation↗

PubMed2001

Infectious complications following T-cell depleted hematopoietic stem-cell transplantation.

BACKGROUND: Although sepsis is a common complication during stem-cell transplantation, the prevalence of infections after hematopoietic recovery is less well known. METHODS: We undertook a retrospective analysis of infectious episodes in patients who underwent allogeneic BM (n = 77) or PBSC (n = 29) grafting from HLA identical siblings. T-cell depletion of the stem-cell grafts with anti CD 52 (CAMPATH-1) Abs was employed for the prevention of GvHD. RESULTS: Patients' median age was 30 (4-54) years. Antibiotic prophylaxis was with oral amphotericin, ofloxacin and i.v. or oral acyclovir. Fever was treated empirically with a third generation cephalosporin and aminoglucosides until results of microbiological cultures became available. Six patients died of graft failure. GvHD was observed in 18% but in no case was it > Grade II. Only seven patients did not develop pyrexia during the initial admission or within 60 days following graft infusion. Median duration of pyrexia was 10 (range 2-49) days. A microbial source was detected in 42% and it was Gram (+) in 86%, Gram (-) in 11% and fungal in 3%. In 16 patients, indwelling venous catheters were removed due to severe infection. Subsequent to the recovery of the blood parameters, the most prevalent infection was by herpes varicella/zoster in 20; another 17 developed herpes simplex. In total 40/102 were re-hospitalized for pyrexia, which in four cases was of unknown origin. Bacterial infections with Staphylococcus Aureus and S. Epidermitis were seen in 10 and seven patients respectively. CMV was detected in seven patients. Thirteen patients died of sepsis and in 10, it was related to GvHD or graft failure. Another 20 died following recurrence of the malignancy. Overall, 39 patients died and 63% survived at a median DFS of 1992 (range 623-5092) days. DISCUSSION: We conclude that during the initial neutropenic period the dominant infections are by Gram (+) organisms, often associated with indwelling catheters. Once the BM has recovered, the main morbidity is by viral infections, but Gram+ organisms still remain common bacterial pathogens.

Adolescent↗

PubMed2003

Near-infrared detection of potential evidence for microscopic organisms on Europa.

The possibility of an ocean within the icy shell of Jupiter's moon Europa has established that world as a primary candidate in the search for extraterrestrial life within our Solar System. This paper evaluates the potential to detect evidence for microbial life by comparing laboratory studies of terrestrial microorganisms with measurements from the Galileo Near Infrared Imaging Spectrometer (NIMS). If the interior of Europa at one time harbored life, some evidence may remain in the surface materials. Examination of laboratory spectra of terrestrial extremophiles measured at cryogenic temperatures reveals distorted, asymmetric nearinfrared absorption features due to water of hydration. The band centers, widths, and shapes of these features closely match those observed in the Europa spectra. These features are strongest in reddish-brown, disrupted terrains such as linea and chaos regions. Narrow spectral features due to amide bonds in the microbe proteins provide a means of constraining the abundances of such materials using the NIMS data. The NIMS data of disrupted terrains exhibit distorted, asymmetric near-infrared absorption features consistent with the presence of water ice, sulfuric acid octahydrate, hydrated salts, and possibly as much as 0.2 mg cm(-3) of carbonaceous material that could be of biological origin. However, inherent noise in the observations and limitations of spectral sampling must be taken into account when discussing these findings.

Extraterrestrial Environment↗

PubMed2025

Seed2LP: seed inference in metabolic networks for reverse ecology applications.

MOTIVATION: A challenging problem in microbiology is to determine nutritional requirements of microorganisms and culture them, especially for the microbial dark matter detected solely with culture-independent methods. The latter foster an increasing amount of genomic sequences that can be explored with reverse ecology approaches to raise hypotheses on the corresponding populations. Building upon genome-scale metabolic networks (GSMNs) obtained from genome annotations, metabolic models predict contextualized phenotypes using nutrient information. RESULTS: We developed the tool Seed2LP, addressing the inverse problem of predicting source nutrients, or seeds, from a GSMN and a metabolic objective. The originality of Seed2LP is its hybrid model, combining a scalable and discrete Boolean approximation of metabolic activity, with the numerically accurate flux balance analysis (FBA). Seed inference is highly customizable, with multiple search and solving modes, exploring the search space of external and internal metabolites combinations. Application to a benchmark of 107 curated GSMNs highlights the usefulness of a logic modelling method over a graph-based approach to predict seeds, and the relevance of hybrid solving to satisfy FBA constraints. Focusing on the dependency between metabolism and environment, Seed2LP is a computational support contributing to address the multifactorial challenge of culturing possibly uncultured microorganisms. AVAILABILITY AND IMPLEMENTATION: Seed2LP is available on https://github.com/bioasp/seed2lp.

Metabolic Networks and Pathways↗

PubMed2005

Toll-like receptors in innate immunity.

Functional characterization of Toll-like receptors (TLRs) has established that innate immunity is a skillful system that detects invasion of microbial pathogens. Recognition of microbial components by TLRs initiates signal transduction pathways, which triggers expression of genes. These gene products control innate immune responses and further instruct development of antigen-specific acquired immunity. TLR signaling pathways are finely regulated by TIR domain-containing adaptors, such as MyD88, TIRAP/Mal, TRIF and TRAM. Differential utilization of these TIR domain-containing adaptors provides specificity of individual TLR-mediated signaling pathways. Several mechanisms have been elucidated that negatively control TLR signaling pathways, and thereby prevent overactivation of innate immunity leading to fatal immune disorders. The involvement of TLR-mediated pathways in autoimmune and inflammatory diseases has been proposed. Thus, TLR-mediated activation of innate immunity controls not only host defense against pathogens but also immune disorders.

Animals↗

PubMed1987

Enzyme-linked synthetic oligonucleotide probes: non-radioactive detection of enterotoxigenic Escherichia coli in faecal specimens.

Synthetic oligonucleotides, complementary to unique sequences in the heat stable enterotoxin gene of Escherichia coli specific for humans, were prepared with a 30-atom spacer arm and a 3' terminal sulfhydryl group which was coupled to bromoacetyl-derivatized alkaline phosphatase. The resulting direct enzyme-linked oligonucleotide probes, containing one enzyme molecule per oligonucleotide, successfully diagnosed enterotoxigenic Escherichia coli in clinical specimens by using a modified colony hybridization method and a colorimetric assay. The procedure is rapid, simple and reliable with a sensitivity equivalent to that using 5'-terminally labelled [32p]-oligonucleotide probes. The results indicate that the enzyme-labelled oligonucleotide probes should be applicable to the routine diagnosis of enterotoxigenic Escherichia coli and possess the potential for the detection of other microbial pathogens.

Alkaline Phosphatase↗

PubMed2002

An assessment of microbial coronal leakage of temporary filling materials in endodontically treated teeth.

This in vitro study evaluated the microbial leakage of Cavit, IRM, and Dyract when used as temporary filling materials after root canal treatment. The degree of coronal leakage was assessed by using a microbiological marker consisting of Streptococcus faecalis and Candida albicans. For each of the two organisms, a set of 15 maxillary premolars were prepared chemomechanically and obturated with thermoplasticized gutta-percha. A 3.5-mm thick layer of one of the three temporary filling materials was inserted in the access cavities of the teeth from each group (each group was compromised of five teeth). The control teeth (four positive and four negative) lacked any filling material over the gutta-percha, whereas the orifice and the apical foramen of the negative control were completely sealed with nail polish. Each tooth was placed in a well of a 24-well tissue culture plate and embedded in trypticase soy broth and 0.5% Bactoagar. An organism suspension was inoculated in the access cavity, and microbial penetration was detected as an increase in turbidity of the broth. At the end of 30 days, the results showed that all positive control teeth leaked within 1 week, whereas those that served as negative control remained uncontaminated throughout the test period. With both organisms, IRM started to leak after 10 days, whereas Cavit and Dyract leaked after 2 weeks.

Analysis of Variance↗

PubMed2006

Characterization and quantification of ammonia-oxidizing bacteria in eutrophic coastal marine sediments using polyphasic molecular approaches and immunofluorescence staining.

Tokyo Bay, a eutrophic bay in Japan, receives nutrients from wastewater plants and other urban diffuse sources via river input. A transect was conducted along a line from the Arakawa River into Tokyo Bay to investigate the ecological relationship between the river outflow and the distribution, abundance and population structure of ammonia-oxidizing bacteria (AOB). Five surficial marine sediments were collected and analysed with polyphasic approaches. Heterogeneity and genetic diversity of beta-AOB populations were examined using restriction fragment length polymorphism (RFLP) analysis of 16S rRNA and amoA genes. A shift of the microbial community was detected in samples along the transect. Both 16S rRNA and amoA genes generated polymorphisms in the restriction profiles that were distinguishable at each sampling site. Two 16S rRNA gene libraries were constructed using the reverse transcription polymerase chain reaction (RT-PCR) method to determine the major ammonia oxidizers maintaining high cellular rRNA content. Two major groups were observed in the Nitrosomonas lineage; no Nitrosospira were detected. The effort to isolate novel AOB was successful; the isolate dominated in the gene libraries. For quantitative analysis, a real-time PCR assay targeting the 16S rRNA gene was developed. The population sizes of beta-AOB ranged from 1.6 x 10(7) to 3.0 x 10(8) cells g(-1) in dry sediments, which corresponded to 0.1-1.1% of the total bacterial population. An immunofluorescence staining using anti-hydroxylamine oxidoreductase (HAO) antibody was also tested to obtain complementary data. The population sizes of ammonia oxidizers ranged between 2.4 x 10(8) and 1.2 x 10(9) cells g(-1) of dry sediments, which corresponded to 1.2-4.3% of the total bacterial fraction. Ammonia-oxidizing bacteria cell numbers deduced by the two methods were correlated (R = 0.79, P < 0.01). In both methods, the number of AOB increased with the distance from the river mouth; ammonia-oxidizing bacteria were most numerous at B30, where the ammonium concentration in the porewater was markedly lower and the nitrite concentration was slightly higher than nearby sites. These results reveal spatial distribution and shifts in the population structure of AOB corresponding to nutrients and organic inputs from the river run-off and phytoplankton bloom.

Ammonia↗