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PubMed1997

Monoclonal antibodies for the detection of desialylation of erythrocyte membranes during haemolytic disease and haemolytic uraemic syndrome caused by the in vivo action of microbial neuraminidase.

Especially in childhood, the in vivo action of microbial neuraminidase may cause haemolytic anaemia or life-threatening haemolytic uraemic syndrome. The exposure of the Thomsen-Friedenreich (T) crypto-antigen and T-antigen polyagglutinability of erythrocytes has been described as the first sign of toxic cleavage of N-acetylneuraminic acid (Neu5Ac) from sialoglycoproteins of cell membranes. This phenomenon may, however, be too unspecific to initiate treatment for toxin elimination. The present study investigated the diagnostic effectiveness of a panel of three monoclonal antibodies (mcabs) for the estimation of the clinical significance of neuraminidase action in vivo. Depending on the amount of Neu5Ac released, the mcabs I-C4, II-Q9 and III-Y12 recognized different epitopes on erythrocyte asialoglycophorin. In 1345 patients, the mcab II-09 detected cleavage of Neu5Ac in 32 children who had T-antigen polyagglutinability and mild to moderate haemolytic anaemia. However, only 10 patients, whose erythrocytes were agglutinated by the mcabs III-Y12 or I-C4, developed severe haemolysis, thrombocytopenia, and finally the life-threatening haemolytic uraemic syndrome (p<0.0002). In conclusion, these mcabs provided an early marker of the in vivo action of neuraminidase. Two different degrees of erythrocyte desialylation, as defined by these mcabs, are suggested to reflect the severity of toxin-associated disease.

Anemia, Hemolytic↗

PubMed1994

Biodegradation of naphthenic acids by microbial populations indigenous to oil sands tailings.

Organic acids, similar in structure to naphthenic acids, have been associated with the acute toxicity of tailings produced by the oil sands industry in northeastern Alberta, Canada. Bacterial cultures enriched from oil sands tailings were found to utilize as their sole carbon source both a commercial mixture of naphthenic acids and a mixture of organic acids extracted from oil sands tailings. Gas chromatographic analysis of both the commercial naphthenic acids and the extracted organic acids revealed an unresolved "hump" formed by the presence of many overlapping peaks. Microbial activity directed against the commercial mixture of naphthenic acids converted approximately 50% of organic carbon into CO2 and resulted in a reduction in many of the gas chromatographic peaks associated with this mixture. Acute toxicity testing utilizing the Microtox test revealed a complete absence of detectable toxicity following the biodegradation of the naphthenic acids. Microbial activity mineralized approximately 20% of the organic carbon present in the extracted organic acids mixture, although there was no indication of a reduction in any gas chromatographic peaks with biodegradation. Microbial attack on the organic acids mixture reduced acute toxicity to approximately one half of the original level. Respirometric measurements of microbial activity within microcosms containing oil sands tailings were used to provide further evidence that the indigenous microbial community could biodegrade naphthenic acids and components within the extracted organic acids mixture.

Acinetobacter calcoaceticus↗

PubMed2005

Technetium-99m ciprofloxacin imaging in the diagnosis of postsurgical bony infection and evaluation of the response to antibiotic therapy: A case report.

The use of radiolabelled antibiotics is fast emerging as a promising diagnostic test for the detection of infective lesions, because of their specific binding to the bacterial component. Ciprofloxacin is a broad-spectrum antibiotic that has been used as a radiolabelled antibiotic in both the diagnosis of infections in rabbits and in clinical trials on humans. The diagnosis of skeletal microbial infections remains a challenge, especially in orthopaedic patients with implants. We present a case report of a patient with undiagnosed and unrelenting pain in the lower third of the left leg, which had persisted for 6 months. A novel scintigraphy, which was developed in-house using a 99mTC ciprofloxacin single vial kit, was used for diagnosing the active bacterial infection. A 99mTC methylene diphosphonate bone scan was also performed to locate precisely the site of the lesion. Gradually increasing concentrations of both the radiotracers confirmed the presence of active bacterial infection in the distal third of the left tibia. Follow-up scanning after antibiotic therapy showed the disappearance of the pooling of 99mTC ciprofloxacin, indicating a good response. Therefore, 99mTC ciprofloxacin has good diagnostic and prognostic applications in deep-seated covert skeletal microbial infections. However, the persistence of a focal concentration detected by the 99mTC methylene diphosphonate scan was attributed to the inflammatory and not the infective pathology.

Adult↗

PubMed1998

Evaluation of microbial hazards during processing of Spanish prepared Flamenquín.

Flamenquín is a traditional, prepared, frozen meat product from the south of Spain made with minced pork, chicken, and cooked ham. Since it is a prepared raw meat product some microbial hazards could be associated with the process of making it. Microbiological analyses have been performed throughout the various steps of processing over a 1-year period to evaluate microbial hazards in the commercial process. High levels of microorganisms were observed all through the processing of this product, the mincing and mixing steps being where major microbial contamination was observed. Pathogenic bacteria such as Staphylococcus aureus, Clostridium perfringens, Escherichia coli and Pseudomonas aeruginosa were detected during processing. Raw materials and food handlers were the principal sources of microbial contamination. A modification of processing to include a heating step after mincing and mixing and an improvement in hygiene practices could eliminate the microbial hazards. Both modifications should be noted for the implementation of a hazard analysis of critical control points (HACCP) program in commercial flamenquín processing.

Clostridium perfringens↗

PubMed1983

Effects of environmental toxicants on metabolic activity of natural microbial communities.

Two methods of measuring microbial activity were used to study the effects of toxicants on natural microbial communities. The methods were compared for suitability for toxicity testing, sensitivity, and adaptability to field applications. This study included measurements of the incorporation of C-labeled acetate into microbial lipids and microbial glucosidase activity. Activities were measured per unit biomass, determined as lipid phosphate. The effects of various organic and inorganic toxicants on various natural microbial communities were studied. Both methods were useful in detecting toxicity, and their comparative sensitivities varied with the system studied. In one system, the methods showed approximately the same sensitivities in testing the effects of metals, but the acetate incorporation method was more sensitive in detecting the toxicity of organic compounds. The incorporation method was used to study the effects of a point source of pollution on the microbiota of a receiving stream. Toxic doses were found to be two orders of magnitude higher in sediments than in water taken from the same site, indicating chelation or adsorption of the toxicant by the sediment. The microbiota taken from below a point source outfall was 2 to 100 times more resistant to the toxicants tested than was that taken from above the outfall. Downstream filtrates in most cases had an inhibitory effect on the natural microbiota taken from above the pollution source. The microbial methods were compared with commonly used bioassay methods, using higher organisms, and were found to be similar in ability to detect comparative toxicities of compounds, but were less sensitive than methods which use standard media because of the influences of environmental factors.

Journal Article↗

PubMed2006

Microbiological assessments of compost toilets: in situ measurements and laboratory studies on the survival of fecal microbial indicators using sentinel chambers.

Compost toilet systems were assessed for their ability to reduce microbial indicators and pathogens. Bacterial pathogens were not detected in any samples indicating a low survival rate in composting feces and/or an initial low occurrence. Indicator bacteria showed large variations with no clear trend of lower bacterial numbers after longer storage. In controlled composting experiments, thermophilic conditions were only reached when amendments were made (grass and a sugar solution). Even then it was impossible to ensure a homogenous temperature in the composting fecal material and therefore difficult to achieve a uniform reduction and killing of indicator organisms. Presumptive thermotolerant coliforms, Salmonella typhimurium Phage 28 B and eggs of Ascaridia galli, proved useful as indicators. However, regrowth was detected for enterococci and total numbers of bacteria grown at 36 degrees C. These indicator parameters may therefore overestimate the level of other (pathogenic) bacteria present in the material and can not be recommended for use as reliable indicator organisms in composting toilet systems. The addition of indicator bacteria to fecal material contained in semi-permeable capsules proved to be a useful technique to ensure that microorganisms were contained in a small test volume.

Animals↗

PubMed1992

Enumeration of subgingival species on primary isolation plates using colony lifts.

This study evaluated the feasibility of using a colony lift method and DNA probes to enumerate bacterial species cultured on primary isolation plates. Fourteen digoxigenin-labeled whole chromosomal DNA probes representing 12 subgingival species were validated by hybridization with colony lifts prepared from 249 reference strains of 51 species grown on Trypticase soy agar plates supplemented with 5% sheep blood. Colonies of reference strains were lifted onto Nytran filters from plates and treated to lyse cells, remove cellular proteins, denature and fix microbial DNA to the filters. Positive reactions were detected with an anti-digoxigenin antibody conjugated to alkaline phosphatase and revealed by bromo-chloro-indolyl phosphate and nitroblue tetrazolium. Cross-reactions were not observed for 13/14 probes, but 2 strains of Streptococcus mitis reacted with the probe to Streptococcus sanguis II. Subgingival plaque samples were taken by means of a sterile curette from mesiobuccal surfaces of teeth present in each of 26 subjects with differing periodontal disease states. Samples were dispersed, diluted, plated and incubated anaerobically for 7 d at 35 degrees C. Colonies were lifted as described above. Filters were cut into sections and hybridized with the 14 digoxigenin-labeled DNA probes. The probes were used to enumerate the test species and the total number of isolates was determined in 711 plaque samples. The colony lift method and DNA probes provided a sensitive, economical and quantitative method for enumerating cultivable microbial species in subgingival plaque samples. In addition, the amplification provided by growing the organisms on agar plates facilitated determination of numbers of organisms in small plaque samples, such as those from healthy sites.

Actinomyces↗

PubMed2001

Protocol for detection of biofilms on needleless connectors attached to central venous catheters.

Central venous catheter needleless connectors (NCs) have been shown to develop microbial contamination. A protocol was developed for the collection, processing, and examination of NCs to detect and measure biofilms on these devices. Sixty-three percent of 24 NCs collected from a bone marrow transplant center contained biofilms comprised primarily of coagulase-negative staphylococci.

Biofilms↗

PubMed1999

Bacterial respiration of arsenic and selenium.

Oxyanions of arsenic and selenium can be used in microbial anaerobic respiration as terminal electron acceptors. The detection of arsenate and selenate respiring bacteria in numerous pristine and contaminated environments and their rapid appearance in enrichment culture suggest that they are widespread and metabolically active in nature. Although the bacterial species that have been isolated and characterized are still few in number, they are scattered throughout the bacterial domain and include Gram-positive bacteria, beta, gamma and epsilon Proteobacteria and the sole member of a deeply branching lineage of the bacteria, Chrysiogenes arsenatus. The oxidation of a number of organic substrates (i.e. acetate, lactate, pyruvate, glycerol, ethanol) or hydrogen can be coupled to the reduction of arsenate and selenate, but the actual donor used varies from species to species. Both periplasmic and membrane-associated arsenate and selenate reductases have been characterized. Although the number of subunits and molecular masses differs, they all contain molybdenum. The extent of the environmental impact on the transformation and mobilization of arsenic and selenium by microbial dissimilatory processes is only now being fully appreciated.

Adenosine Triphosphatases↗

PubMed2006

Evaluation of the in-situ aerobic cometabolism of chlorinated ethenes by toluene-utilizing microorganisms using push-pull tests.

A series of transport, biostimulation, and activity push-pull tests were performed under induced and natural gradient conditions in a trichloroethene (TCE) and cis-dichloroethene (c-DCE) contaminated aquifer. Transport tests demonstrated the feasibility of injecting and recovering complex solute mixtures from the aquifer. During the biostimulation tests, decreases in toluene concentration and the production of o-cresol as an intermediate oxidation product indicated the presence of toluene-utilizing microorganisms. Activity tests demonstrated that the stimulated microbial community had the ability to transform injected c-DCE and trans-dichloroethene (t-DCE) at similar zero-order rates. Injected isobutene was oxidized to isobutene oxide, which indicated that a toluene ortho-monooxygenase enzyme system was likely responsible for the observed c-DCE and t-DCE transformations. c-DCE zero-order transformation rates in drift push-pull tests were similar to those obtained from traditional push-pull tests (about 0.1 microM/h). Analysis of drift test data using first-order kinetic analysis resulted in similar conclusions as those obtained using zero-order kinetic analyses. When 1-butyne, an inhibitor of toluene ortho-monooxygenase, was added to injected test solutions, the oxidation of toluene, and the transformation of isobutene, c-DCE, and t-DCE were inhibited. The results illustrate how a series of push-pull tests can be used in combination to detect, quantify and confirm in-situ cometabolic microbial transformations.

Aerobiosis↗

PubMed1994

Molecular detection of Bacteroides forsythus in human periodontitis.

The usefulness of a digoxigenin-labeled genomic DNA probe for the detection of subgingival Bacteroides forsythus was examined. In addition, the arbitrarily primed polymerase chain reaction (AP-PCR) was used to delineate the genetic diversity of B. forsythus periodontal isolates. The DNA probe detected 10(3) B. forsythus cells and yielded a strong signal at 10(4) cells. It reacted with B. forsythus ATCC 43037T and 44 clinical isolates and showed no detectable reactivity with 75 strains of 24 other oral microbial species. In comparison to culture, the DNA probe in a dot-blot method demonstrated a sensitivity of 88.8% and a specificity of 38.4% (accuracy, 72.5%). By colony-blotting on primary plates, a sensitivity of 98.1% and a specificity of 53.8% (accuracy, 82.5%) were obtained. B. forsythus was detected in 449 (73.1%) of 614 periodontitis patients. The occurrence of the organism was closely associated with Porphyromonas gingivalis, both species being present in 54.8% and absent in 22.2% of 270 study samples. AP-PCR identified 24 B. forsythus genotypes among 27 test strains. This study demonstrated the utility of a non-radioactive genomic probe for direct detection of B. forsythus in subgingival specimens. The species showed a considerable degree of genetic diversity. DNA analysis may help to determine the role of B. forsythus in periodontal disease and its mode of transmission among exposed individuals.

Adolescent↗

PubMed2002

Cultivating the uncultured.

The recent application of molecular phylogeny to environmental samples has resulted in the discovery of an abundance of unique and previously unrecognized microorganisms. The vast majority of this microbial diversity has proved refractory to cultivation. Here, we describe a universal method that provides access to this immense reservoir of untapped microbial diversity. This technique combines encapsulation of cells in gel microdroplets for massively parallel microbial cultivation under low nutrient flux conditions, followed by flow cytometry to detect microdroplets containing microcolonies. The ability to grow and study previously uncultured organisms in pure culture will enhance our understanding of microbial physiology and metabolic adaptation and will provide new sources of microbial metabolites. We show that this technology can be applied to samples from several different environments, including seawater and soil.

Bacteria↗

PubMed2005

Pathways and metabolites of microbial degradation of selected acidic pharmaceutical and their occurrence in municipal wastewater treated by a membrane bioreactor.

Laboratory degradation tests with five acidic pharmaceuticals using activated sludge as inocculum under aerobic conditions were performed and microbial metabolites were analysed by liquid chromatography-mass spectrometry (LC-MS). Ketoprofen was partly mineralized as a sole source of carbon and energy and the metabolites determined by LC-MS suggest microbial ketoprofen degradation to proceed along the pathway known for biphenyls and related compounds. Bezafibrate, naproxen and ibuprofen were degraded only cometabolically whereas no transformation was obtained for diclofenac. Some biodegradation intermediates in these batch tests could be tentatively identified by means of LC-MS. The first step in microbial bezafibrate degradation appears to be the hydrolytic cleavage of the amide bond, generating well degradable 4-chlorobenzoic acid as one of the hydrolysis products. As previously found for mammals, ether cleavage and formation of desmethylnaproxen was the initial step in microbial degradation of naproxen. Two isomers of hydroxy-ibuprofen were detected as intermediates in the mineralization of ibuprofen. Laboratory studies suggest that naproxen and ibuprofen can be fully mineralized whereas more stable metabolites occur in microbial ketoprofen and bezafibrate transformation, that may deserve further attention. A LC-MS method for the trace analysis of these metabolites in water was developed and applied to municipal wastewater. Municipal wastewater treatment by a membrane bioreactor may gradually improve the removal of these pharmaceuticals.

Aerobiosis↗

PubMed2005

A multivariate statistical approach to spatial representation of groundwater contamination using hydrochemistry and microbial community profiles.

Managers of landfill sites are faced with enormous challenges when attempting to detect and delineate leachate plumes with a limited number of monitoring wells, assess spatial and temporal trends for hundreds of contaminants, and design long-term monitoring (LTM) strategies. Subsurface microbial ecology is a unique source of data that has been historically underutilized in LTM groundwater designs. This paper provides a methodology for utilizing qualitative and quantitative information (specifically, multiple water quality measurements and genome-based data) from a landfill leachate contaminated aquifer in Banisveld, The Netherlands, to improve the estimation of parameters of concern. We used a principal component analysis (PCA) to reduce nonindependent hydrochemistry data, Bacteria and Archaea community profiles from 16S rDNA denaturing gradient gel electrophoresis (DGGE), into six statistically independent variables, representing the majority of the original dataset variances. The PCA scores grouped samples based on the degree or class of contamination and were similar over considerable horizontal distances. Incorporation of the principal component scores with traditional subsurface information using cokriging improved the understanding of the contaminated area by reducing error variances and increasing detection efficiency. Combining these multiple types of data (e.g., genome-based information, hydrochemistry, borings) may be extremely useful at landfill or other LTM sites for designing cost-effective strategies to detect and monitor contaminants.

Archaea↗

PubMed2002

A heteroduplex method for detection of targeted sub-populations of bacterial communities.

We describe a simple method, based on heteroduplex mobility analysis of 16S rDNA fragments, for targeted detection of sub-populations of bacteria within diverse microbial communities. A small (ca. 200 bp) polymorphic fragment of the bacterial 16S rRNA gene was amplified from sample DNA using universal primers. Sample products were hybridised with a fluorescently labelled fragment amplified from a selected 'reporter' organism representing the target group. The resulting products were resolved and the labelled heteroduplex pairs detected on non-denaturing gels using automated DNA detection technology. A model, based on analysis of samples with known 16S rDNA sequences, demonstrates that heteroduplex mobility is inversely correlated with genetic distance and that beyond 26% genetic difference, heteroduplex products are not detected. The utility of the method was tested by field studies in which stream biofilms could be characterised by heteroduplex profiles generated with heterotrophic and autotrophic reporter organisms representing target groups.

Bacteria↗

PubMed2005

Value of surveillance blood cultures in neutropenic patients--a pilot study.

In this prospective pilot study we evaluated the efficacy of blood cultures (BC) in detecting bacteremia and fungemia prior to the occurrence of infectious signs. Between February 2003 and July 2003, BC were performed twice weekly in neutropenic hematological patients using blood drawn from central venous catheters. Microbial growth prior to the onset of infectious symptoms was detected in 3 of 45 neutropenic episodes in 39 patients and led to modifications in patient management. These results suggest that further prospective studies are warranted to determine the clinical usefulness of surveillance with BC in neutropenic patients.

Adult↗

PubMedDate not supplied

[Antilactoferrin activity of microorganisms].

The detection rate of the antilactoferrin sign and the level of its manifestation in 165 strains of different microbial species, isolated from patients with inflammatory diseases and intestinal dysbacteriosis, were analyzed. The detection rate of antilactoferrin activity was 43 - 90% for Escherichia coli strains, 20 - 86% for Staphylococcus aureus, 60 - 100% for Klebsiella pneumoniae, 76 - 78% for Candida albicans, isolated from different biotopes of man. Most frequently and with high levels of manifestation this sign was registered in strains isolated from the reproductive tract of women. The detection rate of this sign and the level of its manifestation in bacteria of the vaginal and cervical microbiocenosis, isolated from patients, were higher in comparison with healthy persons. The inverse dependence between the level of the antilactoferrin activity of microflora and the content of lactoferrin during the inflammatory process in women was established.

Biological Factors↗

PubMed1992

Yersinia-triggered reactive arthritis. Use of polymerase chain reaction and immunocytochemical staining in the detection of bacterial components from synovial specimens.

OBJECTIVE: To investigate whether microbial DNA is present in synovial specimens from patients with Yersinia-triggered reactive arthritis. METHODS: Synovial specimens from 13 patients with Yersinia enterocolitica O:3-triggered reactive arthritis and from 16 control patients were studied using polymerase chain reaction and immunocytochemical staining techniques. RESULTS: Yersinia chromosomal DNA was not found in any of the synovial specimens from Yersinia-triggered arthritis patients or controls, whereas with immunocytochemical techniques, Yersinia antigens were observed in synovial specimens from all of the patients with Yersinia-triggered reactive arthritis. CONCLUSION: Only stable bacterial degradation products, not whole bacteria, are present at the site of inflammation in Yersinia-triggered reactive arthritis.

Adult↗