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PubMed1980

Evaluation of two sterility testing methods for intravenous admixtures.

The Addi-Chek Quality Control System (Millipore Corporation) and Ivex-2 Filterset (Abbott Laboratories) were evaluated to determine their effectiveness, applicability, and cost as part of a pharmacy quality-control program. Each method was tested using 50 solutions, 25 of which had been contaminated by inoculation with one of five micro-organisms; the other 25 solutions were used as controls. Aseptic technique was used, and procedures were carried out in a laminar air flow hood. Contaminated solutions were blinded from the person performing the tests. Addi-Chek detected contamination in all the inoculated solutions and in three of the uninoculated solutions. The latter may have been a result of adventitious contamination during the testing procedure. Ivex-2 detected contamination in 24 of the 25 inoculated solutions; no other contamination was found. The effectiveness of the methods in detecting low-level microbial contamination appears comparable. Both methods have been shown to be useful in the pharmacy setting, but Ivex-2 could be used to test for contamination when used as an in-line filter at the patient level. Ivex-2 is less expensive and warrants further evaluation in monitoring for microbial contamination during preparation and administration of intravenous solutions.

Boston↗

PubMed1999

Newer pediatric pathogens.

Newer laboratory techniques have allowed the detection of previously unrecognized microbial agents that are now pathogenic in children, as well as newly recognized manifestations of previously known pathogens. Clinical reports of these newer agents or manifestations are reviewed so that pediatricians can become familiar with these emerging infectious diseases. Physicians need to consider these organisms in the differential diagnosis in appropriate clinical illnesses.

Child↗

PubMed2006

Exercise and Toll-like receptors.

Toll-like receptors (TLRs) are highly conserved trans-membrane proteins that play an important role in the detection and recognition of microbial pathogens. The key product of TLR signalling in antigen presenting cells is the production of inflammatory cytokines and proteins. The TLR pathway plays an important role in mediating whole body inflammation, which has been implicated in the development of chronic disease. An accumulation of chronic, low-grade inflammation is common in individuals that live a sedentary lifestyle; however, the mechanism underlying this connection is not fully understood. There is evidence to show that TLRs may be involved in the link between a sedentary lifestyle, inflammation, and disease. Recent studies have shown that both acute aerobic and chronic resistance exercise resulted in decreased monocyte cell-surface expression of TLRs. Furthermore, a period of chronic exercise training decreases both inflammatory cytokine production and the cell-surface expression of TLR4 on monocytes. These effects may contribute to post-exercise immunodepression and the reported higher susceptibility to infection in athletes. However over the long-term, a decrease in TLR expression may represent a beneficial effect because it decreases the inflammatory capacity of leukocytes, thus altering whole body chronic inflammation. The precise physiological stimulus mediating an exercise-induced decrease in cell-surface TLR expression is not known; however a number of possible signals have been implicated including anti-inflammatory cytokines, stress hormones and heat shock proteins.

Age Factors↗

PubMed1990

[Comparative evaluation of efficiency of various immunologic reactions with Candida antigens in detecting immediate hypersensitivity].

In the dynamic survey of 577 persons having had contacts with yeast-like fungi of the genus Candida the effectiveness of tests for the diagnosis of mycogenic sensitization was made with the use of a number of immunological methods. The basophil degranulation test was shown to have higher sensitivity and specificity in the detection of hypersensitivity to microbial strains used in the production processes of microbiological synthesis, while the use of polysaccharide antigens is more expedient in immunological reactions, such as the precipitation test with the finished product obtained by these processes and with C. albicans. Some prevalence of the level of sensitization in the presence of Candida carriership was established in persons having had contacts with these fungi.

Adult↗

PubMedDate not supplied

[Hygienic and microbiological quality of starters and coagulants used in the production of sheep's milk cheese].

The hygienic level of the raw materials is prevailing in the determination of microbial quality of finished product in food production. Not all alimentary products have the same possibilities of being contaminated. They develop, with a series of intermediate passages, from products that can not be contaminated to products that can be easily attached from micro-organisms. It is essential to locate the points of possible pollution and monitor them with appropriate modalities for a good quality of finished products. The "starters" and "coagulants" of dairy industry may be carrier of microbial pollution if they are improperly handled during the processing and storage. 202 samples of starters and 128 of coagulants, produced by large and small industries have been examined in this work. Total viable count, total Coliforms, anaerobic bacteria and Yeasts have been determined. A quantitative valuation has been also carried out above the lactic bacteria in all samples of starters. The starters results have shown a clean quantitative difference between the industrial products and natural cultures about the lactic microflora value, as well as more high frequency of pollution in samples in which lactic acid bacteria were found below standard. Among the coagulants, no microbial pollution has been detected in the commercial enzymes, whereas serious hygienic problems have shown up in the "rennets"; they have also presented an high anaerobic bacteria number/gr. of product. According to the related data, the AA. think starters and coagulants can have not unfavourable influence on dairy production only is some hygienic operations requirements are followed in their preparation.

Animals↗

PubMed1988

Detection of mycoplasmal antigens in immune complexes from rheumatoid arthritis synovial fluids.

This study was directed towards the detection of suspected antigenic microbial fragments in the immune complex (IC) fraction from chronic inflammatory disorders of the delayed type allergy. Mycoplasmas as the microbial prototype and joint fluids from the rheumatoid host were investigated. Protein-A affinity chromatography was used to isolate the immunoglobulin complex (IgG-IC) in six synovial fluids obtained from rheumatoid arthritis patients. The IgG-IC was digested with pepsin to further purify and obtain F(ab)2 complexes with greater specificity. The F(ab)2 complexes were dissociated and electrophoresed by SDS-PAGE and analyzed by immunoblotting using affinity purified rabbit antisera to six reference strains of human mycoplasmas. The presence of trace amount of mycoplasma antigens in the immune complex fractions was indicated by specific banding with antisera to M. pneumoniae, M. arthritidis, M. hominis, M. fermantans, and M. salivarium in one or more of the six synovial fluid fractions. The ELISA and immunoblot assays of seroconversion in rabbits immunized with the synovial fluid fractions also indicated the presence of mycoplasmal antigens.

Antigen-Antibody Complex↗

PubMedDate not supplied

Evaluation of the ChemScan system for rapid microbiological analysis of pharmaceutical water.

The ChemScan system is a new method for the rapid detection and enumeration of viable microbial cells. It is based on the fluorescent labeling of viable microorganisms collected on a filter membrane and their subsequent automated detection and enumeration by a laser-scanning instrument. The new method was evaluated for the testing of pharmaceutical water by comparison with the standard plate count method. The ChemScan system appeared to be at least as sensitive as the standard method. In some cases the results were equivalent for both methods, but for most water samples the ChemScan results were higher than the standard plate count and sometimes exceeded the latter by an order of magnitude or more.

Drug Industry↗

PubMed2002

Comparative study on microbial removal in immersed membrane filtration (IMF) with and without powdered activated carbon (PAC).

This paper describes an investigation on the effect of microbial removal using IMF for high quality drinking water production. The comparison of IMF and IMF-PAC configuration was carried out in the study to highlight the importance of PAC in the system. The specific objective of this study was to study the effect of PAC adsorption in the IMF-PAC system particularly in removing microbial substances from contaminated raw water. A bench scale IMF-PAC configuration using a flat sheet microfiltration membrane was set up for experimental purposes. Experimentally, the result has shown high removal of microbial substances with the IMF-PAC system compared to IMF. The result of E. coli removal achieved was below the detectable level due to the microbial size, which is bigger than membrane pore size. The addition of PAC has shown a direct effect on total microbial removal. The adsorption of microbial onto PAC surfaces reduced the amount of smaller microbial present in permeate samples. As a conclusion, the configuration of IMF is a promising separation process in removing microbial substances, especially when the system is combined with PAC.

Adsorption↗

PubMed1986

Simplified methods for preparation of microbial fatty acids for analysis by gas chromatography with electron-capture detection.

Analysis of bacterial metabolites and constituents by gas chromatography (GC) with frequency-pulsed electron-capture detection (FPECD) has been suggested as an approach to rapid identification of infection in man. In such methods conversion of analytes to electrophoric derivatives is obligatory. Present methods for analysis of microbial carboxylic acids by GC-FPECD use trichloroethylation. In order to simplify and possibly automate such sample preparation procedures, we investigated pentafluorobenzylation via extractive alkylation and via solid-supported reagents. The sensitivity in terms of minimum amounts of bacteria detectable were determined for the trichloroethyl and pentafluorobenzyl derivatives and results from solid-supported reagents were compared to extractive alkylation.

Alkylation↗

PubMed1999

A novel colorimetric yeast bioassay for detecting trichothecene mycotoxins.

A novel colorimetric microbial bioassay for toxicity has been developed; it shows particular sensitivity to trichothecene mycotoxins. The assay uses inhibition of expression of beta-galactosidase activity within the yeast Kluyveromyces marxianus as a sensitive toxicity indicator, cultures remaining yellow, rather than turning deep green-blue, in the presence of X-gal, a chromogenic substrate. The assay is conducted in standard microtitre plates, permitting small volumes (160 microl) and many replicates, and can be scored either automatically by a plate-reader, or by eye. Factors likely to affect the efficacy of the bioassay, including carbon source, solvents, inoculum cell density, and the use of membrane-modulating agents (MMAs), were assessed. Polymyxin B nonapeptide was the most effective toxicity-enhancing MMA tested, enabling the trichothecene mycotoxin, verrucarin A, to be detected at a concentration of about 1 ng/ml. The assay's reproducibility was examined using polymyxin B sulfate, a cheaper MMA, and another trichothecene mycotoxin, T2 toxin: reproducibility and sensitivity were better for the beta-galactosidase X-gal endpoint than for an alternative chromogenic toxicity indicator, the respiratory substrate 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT).

Biological Assay↗

PubMed1996

Detection of intestinal bacterial translocation using PCR.

Microbial translocation has been suspected to be a major contributing factor in the development of sepsis of unknown origin and multiple organ failure syndrome, but there are currently no tests capable of detecting and quantitating translocation sequentially in humans. The purpose of this study was to develop a sensitive polymerase chain reaction (PCR) test to detect Escherichia coli (E. coli) DNA in the blood of animals after inducing bacterial translocation from the gut. DNA was extracted from blood and primers were used to amplify an 800-bp gene fragment of E. coli by 30-cycle PCR. Detection by southern blotting achieved a sensitivity of 10-100 organisms per 0.3 cc blood. Experimental groups included mice gavaged with 10(10) E. coli followed by 20% body surface area thermal injury, or no injury. Controls included burn only and no treatment groups. Blood was obtained by cardiac puncture 1 hr after burn. Cultures were done on blood samples from all groups. More animals in the burn/gavage group had positive bacterial cultures. All controls were culture negative. E. coli detection by PCR was 100% sensitive in culture positive animals with detection in the gavage/burn group higher than that in all other groups. PCR was negative for all mice without treatment. Several culture negative animals had detectable bacterial DNA by PCR. This highly sensitive and specific method can be used repeatedly to test the blood of patients for the presence of microbial DNA, which could be originating from the gut.

Animals↗

PubMed1996

Species-Specific Detection of Vibrio anguillarum in Marine Aquaculture Environments by Selective Culture and DNA Hybridization.

Methods for specific detection of Vibrio anguillarum in complex microbial communities within diverse marine aquaculture environments were evaluated. A system for the detection of culturable cells based on the combined use of a selective medium and a nonradioactively labeled oligodeoxynucleotide complementary to 16S rRNA was developed. Four hundred fourteen bacterial cultures were evaluated in order to assess the specificity of the method. When both the selective medium and the specific probe gave positive results, the cultures were always identified as V. anguillarum. The selectivity for colony hybridization was 1 V. anguillarum cell in 10,000 total bacterial cells in environmental samples. The utility of the method was also compared with detection by dot blot hybridization of either raw DNA purified from environmental samples or PCR-amplified DNA of 16S rRNA genes, using universal eubacterial primers. The post-PCR hybridization was more sensitive (8 x 10(sup2) cells) than direct hybridization of the whole purified DNA (10(sup6) cells). However, the selective medium-probe combined method was as sensitive as post-PCR hybridization, albeit more specific.

Journal Article↗

PubMed1994

Clinical comparison of isolator and thiol broth with ESP aerobic and anaerobic bottles for recovery of pathogens from blood.

The recovery of pathogens and the speed of their detection were determined for our conventional blood culture system (an Isolator [Wampole] and a 100-ml Thiol bottle [Difco]) compared with automated ESP aerobic and anaerobic bottles (80 ml each; Difco). Each of the four culture devices was inoculated with approximately 10 ml of blood from symptomatic patients weighing more than 80 lb (ca. 36 kg). From 7,070 sets of cultures for 2,841 patients, 607 clinically significant isolates were recovered: 456 (75.1%) from the Isolator, 353 (58.2%) from Thiol, 377 (62.1%) from ESP aerobic bottles, and 346 (57.0%) from ESP anaerobic bottles. Of the 607 isolates, 149 (24.5%) were detected only with the conventional system (Isolator and/or Thiol), and 65 (10.7%) were detected only with the ESP two-bottle system (P < 0.001). Our conventional system allowed for detection of significantly more isolates of members of the family Enterobacteriaceae (P < 0.001), Staphylococcus aureus (P < 0.01), Staphylococcus spp. (coagulase-negative) (P < 0.01), and Enterococcus spp. (P < 0.05), and ESP facilitated detection of significantly more isolates of S. pneumoniae (P < 0.01). When all four devices in a culture set were positive for the same isolate, no microbial species or group was detected significantly earlier ( > or = 24 h) by either blood culture system. The Isolator contamination rate (4.8%) was > or = 6 times the rate for any of the bottles. Of pathogens detected by the Isolator, 50% were recovered in counts of < or = 1.0 CFU/ml and 18% were recovered only as a single colony. The ESP system offered an automated, less labor-intensive blood culture system for which routine subcultures were not required, but the important considerations of culturing large volumes of blood and of obtaining at least two sets from each patient in our population were reemphasized.

Adolescent↗

PubMed1995

Anaerobes in ejaculates of subfertile men.

The clinical significance of micro-organisms in semen samples of asymptomatic subfertile patients is a matter of constant debate. Usually little attention is paid to anaerobic bacteria as they are sensitive to transportation and culturing, and differentiation is difficult, costly and time-consuming. In the present study, special screening was carried out for anaerobes in ejaculates in addition to the routine microbial cultures of genital secretions of both partners. In addition to standard semen analysis and evaluation of sperm ability to penetrate cervical mucus (CM) in vivo (post-coital testing) and in vitro using a standardized test system, semen samples from 126 randomly chosen males of couples with a median duration of infertility of 4 years were examined for colonization with anaerobic bacteria. All couples were without clinical signs or symptoms of genital tract infection. The special care taken for anaerobic growth in semen samples gave a high rate of positive cultures and showed that nearly all ejaculates (99%) were colonized with anaerobic micro-organisms, and potentially pathogenic species were found in 71% of men. This rate was more than four times higher than that obtained with routine cultures and standard transportation (16%). Anaerobic bacterial growth of > or = 10(6) colony forming units (CFU)/ml was seen in 42% (total range 10(3)-10(8) CFU/ ml). In addition, aerobic growth was found in 96% (> or = 10(6) CFU/ml in 21%), potentially pathogenic species in 61% of semen specimens. There were no marked differences in the prevalence of anaerobic micro-organisms in patients with reduced or normal sperm count, motility or morphology. Nor was there any significant difference in anaerobic colonization between samples with impaired or good ability to penetrate CM of female partners (in vivo or in vitro), or the CM of fertile donors in the in-vitro sperm-cervical mucus penetration test (SCMPT) in this asymptomatic group of patients. There was no clear association between microbial colonization and subsequent fertility in vivo within an observation period of 6 months. The results of this study suggest that anaerobic bacteria are often not detected when routine methods for microbial evaluation are used. This should be considered during assisted reproduction and in patients with symptoms of genital tract infection and should lead to further studies in infertile patients where subclinical infection or inflammation is indicated by specific markers in semen samples.

Adult↗

PubMed2006

Pathogen-specific TLR signaling in mucosa: mutual contribution of microbial TLR agonists and virulence factors.

Detection of microorganisms through microbe-associated molecular patterns (MAMP) by Toll-like receptors (TLR) is crucial to trigger protective immunity. In the mucosa, sentinel cells are exposed to MAMP from both pathogens and commensals; however, the TLR response is tightly controlled to avoid inflammation in response to commensals. Uropathogenic Escherichia coli (UPEC) trigger innate responses during urinary tract infection in a TLR4-dependent and CD14-independent manner. UPEC express virulence factors, such as type 1 fimbriae and/or P fimbriae, allowing bacterial attachment to the epithelium. In this issue of the European Journal of Immunology, Fisher et al. show that fimbriae are required to induce a TLR4-specific epithelial response. Depending on the fimbriae expressed by UPEC, different adaptor molecules are involved in TLR4 signaling. These data add to the recent body of evidence suggesting that TLR responses are regulated by co-receptors, such as receptors for virulence factors. In conclusion, the "pathogenic" TLR stimulation provides a novel way for the host to ignore commensal bacteria.

Animals↗

PubMed1986

Simple method to demonstrate radiation-inducible radiation resistance in microbial cells.

A simple method for detection of radiation-inducible radiation resistance was developed by irradiating aliquots (0.01 ml) of cell suspension on agar plates. Part of each experimental plate was subjected to an induction treatment, and subsequent radiation resistance was compared with that of untreated cells on the same plate. The UV radiation resistance of a Micrococcus sp. was increased approximately 1.6 times by an induction treatment. This simple procedure of irradiating cells in a "fixed" position on agar avoided washing, centrifugation, and cell enumeration required in traditional methods.

Agar↗

PubMed1991

Pneumococcal capsular antigen detection and pneumococcal serology in patients with community acquired pneumonia.

BACKGROUND: Methods to determine the microbial cause of community acquired pneumonia include detection of pneumococcal antigen and measurement of pneumococcal capsular antibody response. Their usefulness compared with conventional microbiological techniques was investigated in patients with pneumonia, some of whom had been treated with antibiotics. METHODS: Pneumococcal capsular antigen was detected by latex agglutination in sputum and the results compared prospectively with results of conventional microbiological techniques in 90 patients with community acquired pneumonia. Serum, urine, and pleural fluid samples were also tested for antigen. Serum pneumococcal capsular antibody titres were measured. RESULTS: A diagnosis was established by conventional microbiological techniques in 53 patients, 30 of whom had pneumococcal pneumonia. The sensitivity of antigen detection in first day sputum specimens (n = 18) in those with pneumococcal pneumonia was 94%; antigen was present in 23 of the 27 patients who produced representative sputum on admission and during follow up. The specificity of antigen detection in sputum in patients with non-pneumococcal pneumonia and lung infarction was 87%. Antigen was present in 12 of 25 patients with pneumonia of unknown aetiology who produced representative sputum. Antigen was rarely detected in serum and urine, but was present in pleural fluid in three of four patients with pneumococcal pneumonia and in all four patients with pneumonia of unknown aetiology. Pneumococcal antigen remained detectable in patients treated with antibiotics. Pneumococcal capsular antibody detection was as specific (85%) as antigen detection, but had a lower sensitivity (50%). CONCLUSION: Pneumococcal antigen detection in sputum or pleural fluid is of value in making a rapid diagnosis and provides an additional diagnostic result in patients with pneumococcal pneumonia, especially those receiving antibiotic treatment.

Adult↗

PubMed2006

Genetic Diversity of Microbial Eukaryotes in Anoxic Sediment of the Saline Meromictic Lake Namako-ike (Japan): On the Detection of Anaerobic or Anoxic-tolerant Lineages of Eukaryotes.

Available sequence data on eukaryotic small-subunit ribosomal DNA (SSU rDNA) directly retrieved from various environments have increased recently, and the diversity of microbial eukaryotes (protists) has been shown to be much greater than previously expected. However, the molecular information accumulated to date does still not thoroughly reveal ecological distribution patterns of microbial eukaryotes. In the ongoing challenge to detect anaerobic or anoxic-tolerant lineages of eukaryotes, we directly extracted DNA from the anoxic sediment of a saline meromictic lake, constructed genetic libraries of PCR-amplified SSU rDNA, and performed phylogenetic analyses with the cloned SSU rDNA sequences. Although a few sequences could not be confidently assigned to any major eukaryotic groups in the analyses and are debatable regarding their taxonomic positions, most sequences obtained have affiliations with known major lineages of eukaryotes (Cercozoa, Alveolata, Stramenopiles, and Opisthokonta). Among these sequences, some branched with lineages predominantly composed of uncultured environmental clones retrieved from other anoxic environments, while others were closely related to those of eukaryotic parasites (e.g. Phytomyxea of Cercozoa, Gregarinea of Alveolata, and Ichthyosporea of Opisthokonta).

Anaerobiosis↗