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At least 199 records · Page 11Linked to original sources

PubMed2004

[Evaluation of isolation media for the detection of Bartonella henselae--isolation of Bartonella henselae from domestic cats].

Bartonella henselae is a causative agent of cat scratch disease. We preliminarily tested four media for the bacterial growth, including agar plates with sheep, horse or rabbit blood, and chocolate agar. Of these media, rabbit blood and chocolate agar plate were found to be more excellent for the growth than the medium with sheep or horse blood. Blood samples from 60 domestic cats in Yamaguchi Prefecture were then cultured using 7% rabbit blood agar plates and BACTEC9050 (BD), automated blood culture microbial detection system. B. henselae was isolated from six of the 60 (10%) blood samples. Tiny colonies of B. henselae were visible on the agar medium after one week of culture at 35 degrees C in the 5% CO2 atmosphere. BACTEC 9050 detected B. henselae in one of the 10 blood samples and it took two weeks to detect the bacteria automatically, though gram stain failed to show organisms in the blood culture bottle. In conclusion, rabbit blood or chocolate agar and incubation of agar media more than one week and of BACTEC more than two weeks are recommended for the detection of B. henselae.

Agar↗

PubMed2002

Single-tube, nested PCR for the diagnosis of human brucellosis in Kuwait.

The polymerase chain reaction (PCR) offers a sensitive and specific way of detecting microbial DNA in clinical samples. The aims of the present study were to develop an assay, based on a single-tube, nested PCR, for identifying Brucella in samples of human blood and then to explore the use of this test in diagnosis. The primers chosen were derived from IS711, the insertion sequence gene found in all species of Brucella. The assay amplified a 52-bp final product which was detected colorimetrically. The PCR was sensitive and specific, giving positive reactions with 14 strains of Brucella from five species. The lower limit of detection in vitro was 30 organisms. There were no false-positive reactions either with a range of bacteria known to evoke serological cross-reactions with Brucella (Vibrio cholerae, Yersinia enterocolitica, Serratia marcescens, Haemophilus influenzae, Pseudomonas aeruginosa and Escherichia coli K12) or with organisms producing similar clinical syndromes (Mycobacterium tuberculosis and Salmonella typhi). The results of a preliminary field trial of the assay in Kuwait indicate that the assay may be a valuable technique in the diagnosis of human brucellosis, meriting further study with larger numbers of cases. All 28 subjects with brucellosis (diagnosed on the basis of typical clinical features and confirmed by positive serology and, in three cases, by positive blood cultures) were PCR-positive whereas 28 healthy controls and 28 patients with febrile illness attributable to infections other than brucellosis were PCR-negative.

Animals↗

PubMed1994

[Evaluation of the speed of 3 hemoculture systems to detect bacteria in the blood].

BACKGROUND: The speed of three culture blood systems to detect microbial growth was evaluated with 138 strains isolated from blood which belonged to nine species (N. meningitidis, S. aureus, S. pneumoniae, E. coli, E. faecalis, P. aeruginosa, H. influenzae, A. anitratus and Candida spp.). METHODS: An inoculum of 5-50 UFC were added to each bottle: vials 26 plus with Bactec NR-860 (Becton-Dickinson), aerobial vials with Bact/Alert (Organon) and Hemóline biphasic (bioMérieux). Besides, 10 ml of whole human blood from storage were added to each bottle, too. RESULTS: Bact/Alert was faster in detecting bacterial growth than Bactec with differences statistically significants for all of them, except for H. influenzae. On the other hand, Bactec was faster than Hemóline with differences statistically significant for all bacterial species except S. pneumoniae, which was first detected with manual biphasic method. The median time for detecting growth ranged between 10 h for E. faecalis and A. anitratus in Bact/Alert and 36 h for S. pneumoniae with Bactec. CONCLUSIONS: Bact/Alert detected growth earlier than Bactec, and was quicker than Hemóline. However, other factors have to be in account to choose a method. For instance, biphasic method has the advantage of showing growth in solid medium, whereas automatic systems, at the moment, only use liquid media.

Bacteria↗

PubMed2003

Molecular weapons against agricultural vulnerability and the war on terror.

The multiple reports in this issue of the Journal from the Agenda for Action conference, coupled with the analysis by the National Academy of Sciences, the National Research Council, and the Auditor General (UK) on bioterror preparedness and homeland security, highlight the immediate need for rapid disease detection and advanced diagnostic capabilities to protect the public health, animal agriculture, and the numerous associated economies in the United States. In response to the potentially devastating consequences that could arise, there is an acute need for rapid detection of a variety of the lethal foreign animal diseases, such as foot-and-mouth disease virus (FMDV), highly pathogenic strains of avian influenza, classical swine fever, rinderpest, exotic Newcastle disease virus (END), and domestic, vesicular look-alike diseases that include bluetongue, epizootic hemorrhagic disease, vesicular stomatitis, bovine herpes IBR, contagious ecthyma, bovine herpes mammilitis virus, vesicular exanthema, malignant catarrhal fever, and papular stomatitis. Some striking advances are occurring in the creation of rapid technology, including microfluidics, robotics, miniaturization, and biostabilization that are quickly being applied to the development of rapid microbial detection assays. These are now providing important weapons to combat this agricultural vulnerability.

Animal Diseases↗

PubMed1997

Early microbiologic diagnosis of spontaneous bacterial peritonitis with BacT/ALERT.

BACKGROUND/AIMS: Inoculation of ascitic fluid into conventional blood culture bottles is more sensitive than conventional culture in the diagnosis of spontaneous bacterial peritonitis. BacT/ALERT is an automated colorimetric microbial detection system that has been shown to be faster than conventional blood culture bottles in the diagnosis of bacteremia. The aim of the study was to compare the BacT/ALERT system with the conventional culture and the conventional blood culture bottles method in the diagnosis of spontaneous bacterial peritonitis. METHODS: All the ascitic fluid samples from patients with cirrhosis hospitalized in our Department between September 1992 and May 1994 (n=1032) were prospectively evaluated. In all cases, an aliquot of ascitic fluid was sent for Gram's stain and conventional culture, and 20 ml were inoculated at the bedside into blood culture bottles: 10 ml into conventional blood culture bottles and 10 ml into BacT/ALERT. RESULTS: Thirty ascitic fluid infections (23 spontaneous bacterial peritonitis and 7 neutrocytic ascites) and 20 bacterascites were diagnosed. Conventional culture was positive in 10/30 ascitic fluid infections (33.3%), conventional blood culture bottles in 22/30 (73.3%) (p<0.01 compared to conventional culture) and BacT/ALERT in 20/30 (66.6%) (p<0.05 compared to conventional culture, pNS compared to conventional blood culture bottles). The time elapsed for culture positivity was 43.4+/-34.2 h for conventional blood culture bottles and 13.3+/-9.2 h for BacT/ALERT (p<0.001). Thirteen of the 23 cases of spontaneous bacterial peritonitis (56.5%) were detected within the first 12 h with BacT/ALERT, as compared to only three (13%) with conventional blood culture bottles (p<0.03). CONCLUSION: The automated system BacT/ALERT provides an earlier microbiologic diagnosis of spontaneous bacterial peritonitis than conventional blood culture bottles with similar sensitivity.

Anti-Bacterial Agents↗

PubMed2001

Percent G+C profiling accurately reveals diet-related differences in the gastrointestinal microbial community of broiler chickens.

Broiler chickens from eight commercial farms in Southern Finland were analyzed for the structure of their gastrointestinal microbial community by a nonselective DNA-based method, percent G+C-based profiling. The bacteriological impact of the feed source and in-farm whole-wheat amendment of the diet was assessed by percent G+C profiling. Also, a phylogenetic 16S rRNA gene (rDNA)-based study was carried out to aid in interpretation of the percent G+C profiles. This survey showed that most of the 16S rDNA sequences found could not be assigned to any previously known bacterial genus or they represented an unknown species of one of the taxonomically heterogeneous genera, such as Ruminococcus or Clostridium. The data from bacterial community profiling were analyzed by t-test, multiple linear regression, and principal-component statistical approaches. The percent G+C profiling method with appropriate statistical analyses detected microbial community differences smaller than 10% within each 5% increment of the percent G+C profiles. Diet turned out to be the strongest determinant of the cecal bacterial community structure. Both the source of feed and local feed amendment changed the bacteriological profile significantly, whereas profiles of individual farms with identical feed regimens hardly differed from each other. This suggests that the management of typical Finnish farms is relatively uniform or that hygiene on the farm, in fact, has little impact on the structure of the cecal bacterial community. Therefore, feed compounders should have a significant role in the modulation of gut microflora and consequently in prevention of gastrointestinal disorders in farm animals.

Animal Feed↗

PubMed2005

Persistence of Enterococcus faecalis in aquatic environments via surface interactions with copepods.

Several human pathogens and fecal-pollution indicators may persist as viable organisms in natural environments, owing to their ability to activate different types of survival strategies. These strategies include adhesion on both abiotic and biotic surfaces and the entrance to the so-called viable but nonculturable (VBNC) state. In an 18-month survey for the detection of enterococci in both lake water and seawater, C. Signoretto et al. (Appl. Environ. Microbiol. 70:6892-6896, 2004) have shown that Enterococcus faecalis was detected mostly bound to plankton and in the VBNC state. In the present study, we show that in vitro adhesion of E. faecalis to copepods accelerated the entry of cells into the VBNC state relative to that of planktonic bacteria. VBNC E. faecalis cells maintained their adhesive properties to copepods and chitin (the main component of the copepod carapace), though to a reduced extent in comparison with growing cells. Sugar competition experiments showed interference with adhesion to both copepods and chitin by GlcNAc and only to copepods by D-mannose. Four enterococcal cell wall proteins present in both growing and VBNC cells and lipoteichoic acid were shown to be capable of binding chitin. The results indicate that copepods may represent an additional environmental reservoir of enterococci, thus suggesting the advisability of redesigning the protocols currently used for microbial detection during the evaluation of the microbiological quality of environmental samples.

Bacterial Adhesion↗

PubMed2004

Pure shear horizontal SAW biosensor on langasite.

The undetected introduction of pathogens into food or water supplies can produce grave consequences in terms of economic loss and human suffering. Sensitive and selective sensors capable of quickly detecting microbial pathogens are urgently needed to limit the effects of bioterrorist incidents, accidents, or pollution. Shear horizontal surface acoustic wave (SH SAW) devices provide an attractive platform for the design of microbial biosensors that function in liquid media, where Rayleigh-type modes are rapidly attenuated. This paper reports on an exploratory SH SAW delay line designed and fabricated on langasite, La3Ga5SiO14 (LGS), along the novel Euler propagation direction (0 degrees, 22 degrees, 90 degrees). A liquid chamber was fabricated and attached to the top surface, and the device was submitted to liquid and biochemical tests. Moderate (6 dB) additional attenuation of the transmission coefficient, /S21/, was consistently observed when the SH SAW delay line was assembled in the test fixture and submitted to the liquid tests, indicating that LGS is an attractive candidate for liquid sensing. Sensor selectivity can be achieved by integrating the LGS SH SAW delay line with a biochemical recognition layer. A test setup was implemented for the characterization of LGS SH SAW-based biosensors. The delay line response to biomolecule binding was shown by detection of sequential binding of proteins to the SH SAW device delay path. The biotinylated sensor was exposed sequentially to biotin-binding deglycosylated avidin, biotin-modified rabbit IgG, and goat anti-rabbit IgG antibody. As each protein was bound to the sensing surface, marked changes in the delay-line phase were recorded. The reported results demonstrate the capability of these devices to act as biochemical detectors in aqueous solutions, and this work represents the first effort using the novel material LGS in SAW-based biosensor technology.

Acoustics↗

PubMed1999

Variable selection and multivariate methods for the identification of microorganisms by flow cytometry.

BACKGROUND: When exploited fully, flow cytometry can be used to provide multiparametric data for each cell in the sample of interest. While this makes flow cytometry a powerful technique for discriminating between different cell types, the data can be difficult to interpret. Traditionally, dual-parameter plots are used to visualize flow cytometric data, and for a data set consisting of seven parameters, one should examine 21 of these plots. A more efficient method is to reduce the dimensionality of the data (e.g., using unsupervised methods such as principal components analysis) so that fewer graphs need to be examined, or to use supervised multivariate data analysis methods to give a prediction of the identity of the analyzed particles. MATERIALS AND METHODS: We collected multiparametric data sets for microbiological samples stained with six cocktails of fluorescent stains. Multivariate data analysis methods were explored as a means of microbial detection and identification. RESULTS: We show that while all cocktails and all methods gave good accuracy of predictions (>94%), careful selection of both the stains and the analysis method could improve this figure (to > 99% accuracy), even in a data set that was not used in the formation of the supervised multivariate calibration model. CONCLUSIONS: Flow cytometry provides a rapid method of obtaining multiparametric data for distinguishing between microorganisms. Multivariate data analysis methods have an important role to play in extracting the information from the data obtained. Artificial neural networks proved to be the most suitable method of data analysis.

Bacillus subtilis↗

PubMed1997

Different treatment regimens to reduce microbial contamination of purified porcine islets of Langerhans.

BACKGROUND: Different treatment regimens for minimizing microbial contamination of purified porcine islets were analyzed. METHODS: Purified islets, prepared from abattoir-excised pancreata (n=26), were cultured in Hams F-12 with or without antibiotics after islet isolation. Aliquots of pancreas transport media were all contaminated with microbes (100%). Of those islets cultured with no antibiotics, 70% were contaminated, compared with 42% of the islets cultured in penicillin and streptomycin (P/S) (P>0.05). A further 20 consecutive pancreata were then randomized to saline washing (N=10) before islet isolation. Islets were then cultured in either Hams F-12 with P/S (groups C and E) or Hams F-12 with gentamicin, penicillin, and amphotericin B (G/P/A) (groups D and F). RESULTS: None of those purified islets prepared from washed pancreata and then undergoing culture in Hams F-12 with G/P/A (group F) were contaminated. This compared with 30% contamination when islets were cultured in P/S (group E). Neither antibiotic regimen compromised stimulated insulin release (P=0.13). CONCLUSIONS: In conclusion pancreas washing in saline and culture in G/P/A was shown to eradicate detectable microbial contamination of purified porcine islets isolated from abattoir-excised pancreata.

Animals↗

PubMed2004

Microbial DNA induces a host defense reaction of human respiratory epithelial cells.

Epithelial cells represent the initial site of bacterial colonization in the respiratory tract. TLR9 has been identified in B cells and CD 123(+) dendritic cells and found to be involved in the recognition of microbial DNA. It was the aim of the study to investigate the role of TLR9 in the host defense reactions of the respiratory epithelium. Respiratory epithelial cell lines (IHAEo(-), Calu-3) or fully differentiated primary human cells as air-liquid interface cultures were stimulated with bacterial DNA or synthetic oligonucleotides containing CpG motifs (CpG oligodeoxynucleotides). Expression of TLR9, cytokines, and human beta-defensin 2 was determined by quantitative RT-PCR or by ELISA. We found that TLR9 is expressed by respiratory epithelial cell lines and fully differentiated primary epithelial cells at low levels. Stimulation of the above-mentioned cells with bacterial DNA or CpG oligodeoxynucleotide resulted in an inflammatory reaction characterized by a dose-dependent up-regulation of cytokines (IL-6, IL-8) and human beta-defensin 2. Up-regulation of NF-kappaB in epithelial cells in response to the CpG motif containing DNA was inhibited by overexpression of a dominant negative form of MyD88. These results provide clear evidence that the human respiratory epithelium is capable of detecting microbial DNA by TLR9. The respiratory epithelium has an important function in triggering innate immune responses and therefore represents an interesting target for anti-inflammatory therapy.

Adaptor Proteins, Signal Transducing↗

PubMed2005

Optical microbial biosensor for detection of methyl parathion pesticide using Flavobacterium sp. whole cells adsorbed on glass fiber filters as disposable biocomponent.

An optical microbial biosensor was described for the detection of methyl parathion pesticide. Whole cells of Flavobacterium sp. were immobilized by trapping in glass fiber filter and were used as biocomponent along with optic fiber system. Flavobacterium sp. has the organophosphorus hydrolase enzyme, which hydrolyzes the methyl parathion into detectable product p-nitrophenol. The immobilized microbial biocomponent was disposable, cost-effective and showed high reproducibility and uniformity. The detection of methyl parathion by the use of disposable microbial biocomponent with optical biosensor was simple, single step and direct measurement of very low quantity of the sample. The home made reaction vessel was small and needed only 75 microl of sample. A lower detection limit 0.3 microM methyl parathion was estimated from the linear range (4-80 microM) of calibration plot of organophosphorus hydrolase enzymatic assay. The applicability to synthetic methyl parathion spiked samples was demonstrated.

Aryldialkylphosphatase↗

PubMed2002

Molecular sensing of bacteria in plants. The highly conserved RNA-binding motif RNP-1 of bacterial cold shock proteins is recognized as an elicitor signal in tobacco.

To detect microbial infection multicellular organisms have evolved sensing systems for pathogen-associated molecular patterns (PAMPs). Here, we identify bacterial cold shock protein (CSP) as a new such PAMP that acts as a highly active elicitor of defense responses in tobacco. Tobacco cells perceive a conserved domain of CSP and synthetic peptides representing 15 amino acids of this domain-induced responses at subnanomolar concentrations. Central to the elicitor-active domain is the RNP-1 motif KGFGFITP, a motif conserved also in many RNA- and DNA-binding proteins of eukaryotes. Csp15-Nsyl, a peptide representing the domain with highest homology to csp15 in a protein of Nicotiana sylvestris exhibited only weak activity in tobacco cells. Crystallographic and genetic data from the literature show that the RNP-1 domain of bacterial CSPs resides on a protruding loop and exposes a series of aromatic and basic side chains to the surface that are essential for the nucleotide-binding activity of CSPs. Similarly, these side chains were also essential for elicitor activity and replacement of single residues in csp15 with Ala strongly reduced or abolished activity. Most strikingly, csp15-Ala10, a peptide with the RNP-1 motif modified to KGAGFITP, lacked elicitor activity but acted as a competitive antagonist for CSP-related elicitors. Bacteria commonly have a small family of CSP-like proteins including both cold-inducible and noninducible members, and Csp-related elicitor activity was detected in extracts from all bacteria tested. Thus, the CSP domain containing the RNP-1 motif provides a structure characteristic for bacteria in general, and tobacco plants have evolved a highly sensitive chemoperception system to detect this bacterial PAMP.

Agrobacterium tumefaciens↗

PubMed2003

Evaluation of the VITEK 2 system for rapid direct identification and susceptibility testing of gram-negative bacilli from positive blood cultures.

This study explores the possibility of combining the BacT/Alert Microbial Detection System with the VITEK 2 system to achieve rapid bacterial identification and susceptibility testing. Direct inoculation of bacterial suspension to the VITEK 2 ID-GNB card and AST-NO09 card was made by differential centrifugation of blood cultures of organisms with gram-negative enteric bacillus-like morphology. A total of 118 strains were investigated; of these, 97 (82.2%) strains were correctly identified to the species level and 21 (17.8%) strains were not identified; by comparing the results with those of the reference method of API identification systems using a pure culture, it was found that no strain had been misidentified. Among the 21 strains with no identification, 13 (61.9%) strains were nonfermenters. The direct-identification reporting time of VITEK 2 was 3.3 h. Direct testing of susceptibility to 11 antibiotics, i.e., amikacin, cefepime, ceftazidime, ciprofloxacin, gentamicin, imipenem, meropenem, netilmicin, piperacillin, piperacillin-tazobactam, and tobramycin, was also performed by using the broth microdilution (MB) method according to the NCCLS guidelines as a reference. After comparing the MICs of the VITEK 2 system with those obtained by the MB method within +/-twofold dilution, it was determined that the 1,067 organism-antibiotic combinations had an overall correct rate of 97.6% (1,041 combinations). The rates of susceptibility to the 11 antibiotics ranged from 88.7 to 100%, respectively. Only two (0.2%) and four (0.4%) combinations of the susceptibility tests gave very major errors (i.e., reported as sensitive by the VITEK 2 system but shown to be resistant by the MB method) and major errors (i.e., reported as resistant by the VITEK 2 system but shown to be sensitive by the MB method), respectively. The reporting time for the direct testing of susceptibility against the 11 antibiotics for 97 blood culture isolates by the VITEK 2 system ranged from 3.3 to 17.5 h. Compared with conventional methods that require 1 or 2 days, this method can make same-day reporting possible and thus permit better patient management.

Anti-Bacterial Agents↗

PubMed1990

Oscillometric instrument for the non-invasive detection of low-level microbial activity. Part 2: Growth characteristics, detection limits and impedimetric assays.

Instrument performance and application was evaluated with a variety of microbial suspensions. Data were compared with those obtainable from current commercial apparatus in terms of resolution and detection times. Trials with growing and non-growing cultures indicated periodic synchronous ion exchange processes. Media composition, particularly salt and nutrient ratios, remain crucial to early and rapid detection of viable microbial activity at low concentrations. With appropriate conditions very significant improvements in detection times with typical detection thresholds of 300 min for start concentrations of 10(2) are shown. It is concluded that with regard to resolution and stability, the system out-performs current systems and is superior in terms of potential test costs, sample handling and sample transport.

Asparaginase↗

PubMedDate not supplied

Evolution of beetle bioluminescence: the origin of beetle luciferin.

Bioluminescence, the conversion of chemical energy into light in living organisms, is dependent on two principal components, an enzyme luciferase and the substrate luciferin. In beetles, the enzyme luciferase has been extensively studied, with significant enzymological, sequence and structural data now available. Furthermore, the enzyme has been employed in a remarkable number of important applications, from microbial detection and medical imaging to GM gene expression studies. However, there is little information regarding the biosynthesis of beetle luciferin, and here we review the literature and speculate as to its evolutionary origins. Luciferin consists of a benzothiazole moiety attached to a thiazole carboxylic acid moiety, the former being rarely observed in nature but the latter being observed in a broad range of biologically derived molecules. Benzothiazoles are, however, observed in melanogenesis and we speculate as to whether this may be relevant to the understanding of luciferin biosynthesis in beetles. This review examines recent novel insights into beetle luciferin recycling and we assess a range of possible biosynthetic mechanisms.

Animals↗

PubMed2005

Reading the viral signature by Toll-like receptors and other pattern recognition receptors.

Successful host defense against viral infections relies on early production of type I interferon (IFN) and subsequent activation of a cellular cytotoxic response. The acute IFN and inflammatory response against virus infections is mediated by cellular pattern-recognition receptors (PRRs) that recognize specific molecular structures on viral particles or products of viral replication. Toll-like receptors (TLRs) constitute a class of membrane-bound PRRs capable of detecting microbial infections. While TLR2 and TLR4, which were first identified to recognize Gram-positive and Gram-negative bacteria, respectively, sense specific viral proteins on the cell surface, TLRs 3, 7, 8, and 9 serve as receptors for viral nucleic acids in endosomic compartments. In addition to TLRs, cells express cytoplasmic PRRs such as the RNA helicase retinoic acid inducible gene I and the kinase double-stranded RNA-activated protein kinase R, both of which sense dsRNA, a characteristic signature of viral replication, and initiate a protective cellular response. Here we review the recent progress in our understanding of PRRs and viral infections and discuss the molecular and cellular responses evoked by virus-activated PRRs. Finally, we look into what is currently known about the role of PRRs in viral infections in vivo.

Animals↗

PubMed2005

The appropriateness of swab cultures for the release of human allograft tissue.

Surgeries utilizing human allograft tissues have increased dramatically in recent years. With this increase has come a greater reliance on the use of swab culturing to assess allograft tissues for microbial contamination prior to distribution. In contrast to the typical industrial microbiological uses for swabs, the tissue banking industry has relied on swab cultures as a sterility release method for allograft tissues. It has been reported in the literature that swabs have limitations, both in sensitivity and reproducibility, so their suitability as a final sterility release method was evaluated in this study. Two different swab-culturing systems were evaluated (COPAN, EZ Culturette) using human allograft tissues spiked with low levels of multiple bacterial and fungal microorganisms. The average microbial recoveries for all challenge microorganisms for each tissue type and each swab system were calculated. Percent recoveries for each challenge microorganism were also calculated and reported. The results indicated that both swab systems exhibited low and highly variable recoveries from the seeded allograft tissues. Further analysis indicated there was no statistical difference ( proportional, variant=0.05) between the two swab systems. It is the recommendation of the authors that swab culturing not be used to assess relatively low levels of microbial contamination on allografts. Instead, alternative validated microbial detection methods with improved sensitivity and reproducibility should be employed and validated for this critical task.

Cell Culture Techniques↗