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PubMed1992

[Usefulness of microbial investigations in community-acquired pneumonia].

Community acquired pneumonia (CAP) is a common and well known disease, however there is no definite agreement on a common diagnostic-therapeutic strategy. To evaluate the usefulness of microbial investigations in the clinical practice we performed a prospective study on 93 consecutive patients with a diagnosis of CAP. Group I consisted of 46 patients that underwent a diagnostic protocol including sputum, blood cultures and detection of specific antibodies against M. pneumoniae, adenovirus, respiratory syncytial virus, and L. pneumophila. Group II consisted of 47 patients, in which only sputum samples were collected and cultured. No significant differences concerning the aetiologic diagnosis, the outcome and the length of hospitalization were observed in the two groups. The aetiological diagnosis was obtained in 17 patients (18.3%). As result of information obtained from microbiol tests, antibiotic therapy was changed only in 6 patients. Among the prognostic factors only a low albumin level was correlated with the length of hospitalization (p less than 0.01). From our data, the detection of microbial aetiology should not be routinely performed in patients with CAP, but should be reserved only to the severe forms.

Adult↗

PubMed2005

A simple, rapid, sensitive method detecting homoserine lactone (HSL)-related compounds in microbial extracts.

A simple, rapid, sensitive microtiter plate method detecting N-acyl homoserine lactone (HSL)-related compounds was established using an Agrobacterium tumefaciens strain harboring a traG::lacZ/traR reporter gene responsive to HSLs. This strain did not produce its own HSL, but the traG::lacZ reporter gene was induced only when its transcription activator TraR detected a cognate exogenous HSL. Therefore, the assay was expected to be highly specific for HSL-related compounds. Induction of the reporter gene, leading to production of beta-galactosidase enzyme, was measured by using two different beta-galactosidase substrates, X-gal and Galacton-Star, for colorimetric and chemiluminometric detection, respectively. The screen was validated in both the 96-well and 384-well plate formats, and extracts derived from 696 different microbial isolates, mostly unidentified actinomycetes isolated from diverse locations, were tested. Crude extracts of 81 (11.64%) cultures tested positive for HSL-related compounds, and an additional 34 (4.8%) crude extracts showed a moderate to weak signal for HSLs. Data from the fractionated samples, however, suggested a much higher prevalence of HSL signals in these extracts. Of 144 crude extracts fractionated into 10 individual samples at a 10x concentration, 72 (50%) cultures tested positive for HSLs. Six cultures were active only in the crude extract, 18 were active both in crude and one or more of their fractions, and an additional 48 were active in just one or more of their fractions. This finding may be the first to suggest such a high prevalence of HSL-signals found in nature, and a large number of actinomycetes in our collection appeared to produce HSL-related compounds.

4-Butyrolactone↗

PubMed1995

Solid-phase clean-up and thin-layer chromatographic detection of veterinary aminoglycosides.

Chemical methods are needed to confirm the presence of antibiotics detected by microbial inhibition assays in fluids and tissues of farm animals. We have optimized the conditions for the isolation of hygromycin B with a copolymeric bonded solid-phase silica column followed by thin-layer chromatography (TLC) separation and detection of its fluorescence derivative after reaction with fluorescamine. The detection limit of the drug was 50 ng. Serum and plasma samples fortified with hygromycin B were acidified and passed through the copolymerized solid-phase columns previously conditioned with phosphate buffer. Hygromycin B was trapped in the columns and eluted with diethylamine-methanol and analyzed by TLC using acetone-ethanol-ammonium hydroxide as the developing solvent. Hygromycin B bands were derivatized at acidic pH with fluorescamine and visualized under ultraviolet light. Hygromycin B added to bovine plasma was detectable at 25, 50, 100, 250 and 500 ng/ml (ppb). Hygromycin B added to swine serum was detected at 50 ng/ml. However, the serum had to be deproteinized with trichloroacetic acid or acetonitrile prior to solid-phase extraction to gain accurate values. Neomycin and gentamicin (100 ng/ml aqueous solutions) could also be isolated with copolymeric solid-phase columns at a level of 50 ng. Gentamicin, neomycin, gentamicin, spectinomycin, hygromycin B and streptomycin could be separated by TLC, allowing multiresidue detection of these aminoglycosides. The respective RF values of 0.64, 0.56, 0.52, 0.33 and 0.20 indicate the separation of these five compounds. This procedure provides a rapid and sensitive method for the semi-quantitative estimation of aminoglycosides.

Animals↗

PubMed1999

Arsenic in the Meager Creek hot springs environment, British Columbia, Canada.

Levels of arsenic in water from Meager Creek hot springs, British Columbia, Canada, were found to be naturally elevated. Biota including microbial mats, green algae, sedge, cedar, fleabane, monkey flower, moss, mushrooms and lichens, that were expected to be impacted by the water, were analyzed for total levels of arsenic and for arsenic species. The major arsenic species extracted from all samples were arsenate and arsenite, which are toxic forms of arsenic. Additionally, small amounts of arsenosugars X and XI were detected in microbial mats and green algae, implying that cyanobacteria/bacteria, and possibly green algae are capable of synthesizing arsenosugars from arsenate. Low to trace amounts of arsenosugars X and XI were detected in lichens and the fungus Tarzetta cupularis. A large fraction (on average, greater than 50%) of arsenic was not extracted by using methanol/water (1:1) and the chemical and toxicological significance of this arsenic remains unknown.

Arsenic↗

PubMed1986

Centrifugation-augmented solid-phase immunoassay (CASPIA) for the rapid diagnosis of infectious diseases.

We have devised centrifugation-augmented solid-phase immunoassays (CASPIAs) for the detection of microbial pathogens in human body fluids. The CASPIA format combines several features of latex agglutination and solid-phase immunoassay systems to produce an assay system that is inexpensive and simple to perform. We found that CASPIA systems for the detection of rotaviral, group A streptococcal, and Haemophilus influenzae type b antigens were significantly more sensitive than latex agglutination assays and at least as sensitive as enzyme immunoassay systems using analogous reagents. Because CASPIA reactions could either be read macroscopically, recorded with a standard office photocopying device, or quantitated by means of a microplate colorimeter, the assays were applicable under a wide range of clinical and laboratory conditions.

Agglutination Tests↗

PubMed2001

Experimental induction of chronic borreliosis in adult dogs exposed to Borrelia burgdorferi-infected ticks and treated with dexamethasone.

OBJECTIVE: To develop a method to experimentally induce Borrelia burgdorferi infection in young adult dogs. ANIMALS: 22 healthy Beagles. PROCEDURE: All dogs were verified to be free of borreliosis. Twenty 6-month-old dogs were exposed to Borrelia burgdorferi-infected adult ticks and treated with dexamethasone for 5 consecutive days. Two dogs not exposed to ticks were treated with dexamethasone and served as negative-control dogs. Clinical signs, results of microbial culture and polymerase chain reaction (PCR) testing, immunologic responses, and gross and histologic lesions were evaluated 9 months after tick exposure. RESULTS: Predominant clinical signs were episodic pyrexia and lameness in 12 of 20 dogs. Infection with B burgdorferi was detected in microbial cultures of skin biopsy specimens and various tissues obtained during necropsy in 19 of 20 dogs and in all 20 dogs by use of a PCR assay. All 20 exposed dogs seroconverted and developed chronic nonsuppurative arthritis. Three dogs also developed mild focal meningitis, 1 dog developed mild focal encephalitis, and 18 dogs developed perineuritis or rare neuritis. Control dogs were seronegative, had negative results for microbial culture and PCR testing, and did not develop lesions. CONCLUSIONS AND CLINICAL RELEVANCE: Use of this technique successfully induced borreliosis in young dogs. Dogs with experimentally induced borreliosis may be useful in evaluating vaccines, chemotherapeutic agents, and the pathogenesis of borreliosis-induced arthritis.

Animals↗

PubMedDate not supplied

Gowning training: the use of video recording together with microbial assessment.

Following classroom training on proper aseptic gowning technique, clean room operators were filmed while gowning according to standard procedure. Subsequently, the exterior surface of each operator's gown was monitored for microbial contamination. Operators reviewed their personal videotaping with the trainer. Through careful and critical self-observation, the operator's ability to follow the gowning procedure in addition to using good gowning technique was ascertained. Furthermore, the operator's gowning technique was compared with the results of the microbiological monitoring data. A correlation between microbial contamination of the gown surface and improper technique was demonstrated. This correlation enhanced the operator's awareness of the consequences of improper gowning technique. Use of video replay enabled the operator and trainer to identify improper or subtle flaws in technique detected through microbial monitoring but undetected during actual gowning. This training program served to further emphasize the importance of using proper aseptic gowning technique. Clean room operators are more involved in maintaining clean room integrity and their awareness of the potential for contaminating the area by inadequate technique has been enhanced.

Asepsis↗

PubMed2003

Distribution and phylogenetic diversity of the subsurface microbial community in a Japanese epithermal gold mine.

Distribution and phylogenetic diversity of microbial communities in hot, deep underground environments in the Hishikari epithermal gold mine, southern part of Kyushu, Japan, were evaluated using molecular phylogenetic analyses. Samples included drilled cores such as andesitic volcanic rock (0.95-1.78 Ma) and the oceanic sedimentary basement rock of Shimanto-Supergroup (100 Ma), as well as geothermal hot aquifer waters directly collected from two different sites: AW-site (71.5 degrees C, pH 6.19) and XW-site (85.0 degrees C, pH 6.80) at a depth of 350 mbls (meters below land surface). Based on PCR-amplified 16S rRNA gene clone analysis, the microbial communities in the drilled cores and the hot aquifer water from the XW-site consisted largely of the 16S rRNA gene sequences, closely related to the sequences often found in marine environments, while the aquifer water from the AW-site contained 16S rRNA gene sequences representing members of Aquificales, thermophilic methanotrophs within the gamma-subdivision of the Proteobacteria and uncultivated strains within the beta-subdivision of Proteobacteria. The cultivable microbial community detected by enrichment cultivation analysis largely matched that detected by the culture-independent molecular analysis.

Bacteria↗

PubMed1984

Clinical comparison of the Isolator 1.5 microbial tube and the BACTEC radiometric system for detection of bacteremia in children.

The Isolator 1.5 microbial tube (E. I. du Pont de Nemours & Co., Inc., Wilmington , Del.) was compared with the BACTEC radiometric detection system (Johnston Laboratories, Inc., Cockeysville, Md.) for the detection of bacteremia in children. The Isolator 1.5 is a blood culture system designed for small volumes of blood (0.5 to 1.5 ml). The method involves lysis of the cells of the patient and the direct plating of the entire blood lysate on agar media appropriate for the growth of fastidious microorganisms. Of 1,500 paired samples inoculated into the two systems, 68 were positive for 73 clinically significant organisms. The Isolator 1.5 recovered 81% of the positive cultures compared with 84% recovered by the BACTEC system. When paired blood samples with disproportionate volumes were excluded, the Isolator 1.5 detected 3% more positive cultures. More isolates of Streptococcus pneumoniae and Neisseria meningitidis were recovered by the Isolator 1.5, whereas Haemophilus influenzae was recovered most often in the BACTEC bottles (P greater than 0.1). The contamination rates were 8.7 and 3.1% for the Isolator 1.5 and the BACTEC system, respectively. In cultures positive by both systems, the mean time to detection was 4.1 h faster with the Isolator 1.5. The mean time to obtain isolated colonies was 26.6 h faster with the Isolator 1.5. These data indicate the potential value of the Isolator 1.5 microbial tube as a simple, rapid, and sensitive method for the detection of bacteremia in children.

Bacteria↗

PubMed2004

Microbial contamination of femoral head allografts.

OBJECTIVE: To study the incidence of microbial contamination at the bone bank of the United Christian Hospital. DESIGN. Retrospective study. SETTING: Regional hospital, Hong Kong. PATIENTS: A total of 151 patients (33 men and 118 women) who underwent hip arthroplasty surgery and from whom femoral head allografts were retrieved between January 1994 and March 2000; and 81 patients in whom allografts were implanted. MAIN OUTCOME MEASURES: Bone biopsies were taken from the femoral head and used to detect any microbial contamination that might have occurred during removal and after storage. The rates of infection among recipients and donors were also assessed. RESULTS: Of the 151 allografts, 94 non-contaminated allografts were implanted by the end of the study. Fourteen (9.3%) heads showed positive culture results after retrieval and were discarded. Four (4.3%) of the 94 stored allografts that were implanted tested positive for microbial growth, but the recipients of these allografts did not develop any clinical infection. Three (3.2%) had wound infections after implantation of the stored allografts although the grafts had previously been tested negative for any microbial contamination. CONCLUSION: Our centre has a low allograft contamination rate. The wound infection rate among recipients was also low. The culture of a bone biopsy sample is a reliable method to detect contamination of bone grafts. However, the contamination rate among stored allografts should prompt orthopaedics departments to review allograft handling procedures, so as to minimise the chance of contamination.

Anti-Bacterial Agents↗

PubMed2001

Vertical Distribution of Denitrification Potential, Denitrifying Bacteria, and Benzoate Utilization in Intertidal Microbial Mat Communities.

Aspects of denitrification and benzoate degradation were studied in two estuarine microbial mat communities on the California coast by measuring the depth distributions of potential denitrification rates, genetic potential for denitrification, nitrate concentration, benzoate mineralization rates, total bacterial abundance, and abundance of a denitrifying strain (TBD-8b) isolated from one of the sites. Potential denitrification was detected in microbial mat cores from both Elkhorn Slough and Tomales Bay. Maximum denitrification rates were more than two orders of magnitude higher at Elkhorn Slough (3.14 mmol N m -2 d -1) than at Tomales Bay (0.02 mmol N m -2 d -1), and at both sites, the maximum rates occurred in the 0-2 mm depth interval. Ambient pore [NO 3 + NO 2] was substantially higher at Elkhorn Slough than at Tomales Bay. Incorporation and mineralization of benzoate was maximal near the mat surface at Elkhorn Slough. The areal rate of benzoate utilization was 1045 nmol C m -2 d -1, which represented utilization of 70% of the added substrate in 24 h. Total bacterial and TBD-8b abundances were greatest near the surface at both Tomales Bay and Elkhorn Slough, and TBD-8b represented less than 0.2% of the total. Genetic potential for denitrification, quantified by hybridization with a nitrite reductase gene fragment, was present below the mat surface at average levels representing presence of the gene in approximately 10% of the total cells.

Journal Article↗

PubMedDate not supplied

[The leading pathways and factors in the transmission of the causative agents of campylobacteriosis under current conditions].

The authors have studied the epidemiological importance of different paths and factors of the transmission of causative agents in the foci of campylobacteriosis, as well as the character and frequency of microbial associations detected in cases of campylobacteriosis. The family foci of campylobacteriosis have been epidemiologically studied and the detected factors contributing to contacting campylobacteriosis have been analyzed. As revealed in this study, 41.6% of the foci of campylobacteriosis are linked with the active action of the alimentary factor, while 37.9% of the foci are linked with the factor of everyday contacts. High frequency of microbial associations with Campylobacter has been established (20.8%), which makes it possible for sporadic cases and especially outbreaks of campylobacteriosis to be disguised as other infections.

Campylobacter↗

PubMed1993

Biosensors: applications for dairy food industry.

Biosensors are defined as indicators of biological compounds that can be as simple as temperature-sensitive paint or as complex as DNA-RNA probes. Food microbiologists are constantly seeking rapid and reliable automated systems for the detection of biological activity. Biosensors provide sensitive, miniaturized systems that can be used to detect unwanted microbial activity or the presence of a biologically active compound, such as glucose or a pesticide. Immunodiagnostics and enzyme biosensors are two of the leading technologies that have had the greatest impact on the food industry. The use of these two systems has reduced the time for detection of pathogens such as Salmonella to 24 h and has provided detection of biological compounds such as cholesterol or chymotrypsin. The continued development of biosensor technology will soon make available "on-line quality control" of food production, which will not only reduce cost of food production but will also provide greater safety and increased food quality.

Biosensing Techniques↗

PubMed1993

Recent advances of flow cytometry in fundamental and applied microbiology.

This review focuses on the recent applications of flow cytometry (FCM) in microbiological research (1987-mid 1992). It tries to give a scope of the important breakthroughs which occurred in this field during this period. The technical difficulties of microorganism analysis by flow cytometry is briefly appraised. The significance and the limits of the different microbial cell parameters attainable by flow analyses are systematically evaluated: light scatter for cell size and structure, fluorescence measurements for quantification of cellular components, microbial antigen detection and cell physiological activity estimation. Emphasis is given on the new technological advances which appeared in the last two years. The second part of the review is devoted to the analysis of the usefulness of flow cytometric approach in the different fields of microbiology: fundamental studies in microbial physiology, differentiation, microbial ecology and aquatic sciences, medical microbiology, parasitology, microbial pharmacology and biotechnology.

Biotechnology↗

PubMed2002

A new microbial assay for the toxicity detection of contaminated soils.

A comparative study to detect toxicity prior to bioremediation treatment was set in order to investigate dehydrogenase activity inhibition of a common soil bacterium caused by soil contaminated with Cu, Pb, and As. A spectrophotometric test with Pseudomonas fluorescens strain ATCC 13525 utilising the 2,3,5-triphenyl tetrazolium chloride (TTC) reduction by microbial dehydrogenase has been adapted for this purpose. Soil samples are incubated for 48 hours at 30 +/- 1 degrees C in 18-ml tubes in the presence of TTC as an artificial electron-acceptor. The reduced TTC forms a reddish colour substance named triphenyl formazan (TPF), which can be extracted from the microbial cells and measured colorimetrically. The rapid response of biological activity in microorganisms and the reported sensitivity to the toxicants in the contaminated samples are reflected by the TTC reduction method, which is a sensitive tool for toxicity screening of contaminated sites, routine monitoring of bioremediation processes, as well as for feasibility studies of bioremediation treatments, in order to assess whether a specific pollutant or any other substance at a site location could inhibit the microbiological processes.

Arsenic↗

PubMed1996

The effect of a portable HEPA-filtered body exhaust system on airborne microbial contamination in a conventional operating room.

OBJECTIVE: To study the effect of a portable HEPA-filtered air exhaust system (Stackhouse Freedom Surgical Helmet System) on airborne microbial contamination in a modern conventional operating room. DESIGN AND SETTING: Microbial air sampling was done with a two-stage Anderson sampler at the wound site during 46 total joint replacements. All operations were performed by the same surgeon in the same operating room at a large community hospital. RESULTS: In 18 cases done without air exhaust hoods, the number of bacterial and fungal colony-forming units (CFU) ranged from 0.6 to 11.7 (mean, 3.6). Air sampling during 28 operations with the operating team in air exhaust hoods revealed a mean of 3.6 CFU (range, 0 to 11.4). Bacterial CFU averaged 3.4 without hoods and 3.2 with exhaust hoods. Coagulase-negative staphylococci were the most common isolates (48% of isolates with hood, 55% without hood). No infections occurred. CONCLUSION: We concluded that these air exhaust hoods did not lower airborne microbial contamination detectable with this air sampling method, as compared to standard head cover and mask, in a modern conventional operating room.

Air Microbiology↗

PubMed2004

IL-12p70-dependent Th1 induction by human B cells requires combined activation with CD40 ligand and CpG DNA.

The detection of microbial molecules via Toll-like receptors (TLR) in B cells is not well characterized. In this study, we found that both naive and memory B cells lack TLR4 (receptor for LPS) but express TLR9 (receptor for CpG motifs) and produce IL-6, TNF-alpha, and IL-10 upon stimulation with CpG oligonucleotides (ODN), synthetic mimics of microbial DNA. Consistent with the lack of TLR4, purified B cells failed to respond to LPS. Similar to CpG ODN, CD40 ligand (CD40L) alone induced IL-6, TNF-alpha, and IL-10. Production of these cytokines as well as IgM synthesis was synergistically increased when both CpG ODN and CD40L were combined. Unlike IL-6, TNF-alpha, and IL-10, the Th1 cytokine IL-12p70 was detected only when both CpG ODN and CD40L were present, and its induction was independent of B cell receptor cross-linking. CpG ODN did not increase the capacity of CD40L-activated B cells to induce proliferation of naive T cells. However, B cells activated with CpG ODN and CD40L strongly enhanced IFN-gamma production in developing CD4 T cells via IL-12. Together, these results demonstrate that IL-12p70 production in human B cells is under the dual control of microbial stimulation and T cell help. Our findings provide a molecular basis for the potent adjuvant activity of CpG ODN to support humoral immune responses observed in vivo, and for the limited value of LPS.

Adjuvants, Immunologic↗

PubMed2005

Corn grain endosperm type and brown midrib 3 corn silage: ruminal fermentation and N partitioning in lactating cows.

Interactions of endosperm type of corn grain and the brown midrib 3 mutation (bm3) in corn silage on ruminal fermentation and microbial efficiency of lactating dairy cows were evaluated. Eight ruminally and duodenally cannulated cows (72 +/- 8 d in milk; mean +/- SD) were used in a duplicated 4 x 4 Latin square design experiment with a 2 x 2 factorial arrangement of treatments. Treatments were corn grain endosperm type (floury or vitreous) and corn silage type (bm3 or isogenic normal). Diets contained 26% neutral detergent fiber and 30% starch. Increasing ruminal starch digestibility by replacing vitreous corn grain with floury grain reduced mean and minimum ruminal pH. Brown midrib 3 corn silage reduced mean and minimum ruminal pH and increased total volatile fatty acid concentration. Ruminal pH was positively associated with rate of valerate absorption. Although floury endosperm reduced acetate:propionate ratio in both control and bm3 corn silage diets, it had a greater effect on reducing acetate:propionate ratio for control silage compared with bm3 corn silage. Nonammonia N flow to the duodenum did not differ among treatments and no effects of treatment were detected for microbial N and nonammonia, nonmicrobial N flow. Although treatment effects on ruminal fermentation and ruminal pH were observed, few interactions of treatment were detected and treatments did not affect flow of N fractions to the intestines.

Animals↗