Cardiobacterium valvarum, a fastidious Gram-negative bacterium, was detected in the aortic valve of a previously healthy 63-year-old man by broad-range PCR and 16S rRNA gene sequencing. In contrast to the patients in five previously published cases, our patient had neither a congenital bicuspid nor a prosthetic aortic valve. Here, we present a case of C. valvarum native tricuspid aortic valve infective endocarditis and a review of the literature.
- MeSH
- Anti-Bacterial Agents administration & dosage therapeutic use MeSH
- Aortic Valve parasitology pathology MeSH
- Endocarditis, Bacterial drug therapy surgery microbiology MeSH
- Cardiobacterium isolation & purification MeSH
- Cefotaxime administration & dosage therapeutic use MeSH
- Gentamicins administration & dosage therapeutic use MeSH
- Middle Aged MeSH
- Humans MeSH
- Heart Valve Diseases microbiology pathology MeSH
- Polymerase Chain Reaction MeSH
- Check Tag
- Middle Aged MeSH
- Humans MeSH
- Male MeSH
- Publication type
- Case Reports MeSH
- Research Support, Non-U.S. Gov't MeSH
- Review MeSH
OBJECTIVES: Broad-range PCR has the potential to detect virtually any bacterial species via amplification and nucleotide sequencing of a DNA region common to all bacteria. We aimed to evaluate its usefulness and clinical relevance when applied to a wide variety of primary sterile materials. METHODS: A prospective study including 1370 samples (75 heart valves, 151 joint tissue samples, 230 joint aspirates, 848 whole blood samples and 66 culture-negative cerebrospinal fluid samples) were studied by using a commercial PCR system for detecting 16S rDNA (Molzym). The PCR results were compared with culture and were considered to provide added diagnostic value only if the PCR approach revealed new pathogens that were missed by culture. RESULTS: The added value of PCR was evident in 173 of 555 PCR-positive samples (0.126; 0.109-0.144 (proportion from all tested samples; 95% confidence interval)), most frequently in examinations of heart valves (0.56; 0.448-0.672) and joint tissue samples (0.219; 0.153-0.284). In contrast, the lowest rate of PCR with added value was noted for blood samples, regardless of the patient cohort they had been drawn from (nononcologic patients from intensive care: 0.065; 0.043-0.087, haematooncologic children: 0.048; 0.027-0.070). Moreover, PCR missed up to 7.1% of blood culture findings (0.071; 0.048-0.095) regarded as clinically relevant, which was the second highest failure rate after joint tissue samples (0.099; 0.052-0.147). CONCLUSIONS: Broad-range PCR substantially increases detection rate of pathogens, especially from heart valves and joint samples. However, a concurrent risk of false-negative PCR results justifies the need for parallel culture.
- MeSH
- Bacterial Infections diagnosis MeSH
- Bacteriological Techniques methods MeSH
- Molecular Diagnostic Techniques methods MeSH
- Child MeSH
- Adult MeSH
- Infant MeSH
- Middle Aged MeSH
- Humans MeSH
- Adolescent MeSH
- Polymerase Chain Reaction methods MeSH
- Child, Preschool MeSH
- Prospective Studies MeSH
- RNA, Ribosomal, 16S genetics MeSH
- Sequence Analysis, DNA methods MeSH
- Aged, 80 and over MeSH
- Aged MeSH
- Check Tag
- Child MeSH
- Adult MeSH
- Infant MeSH
- Middle Aged MeSH
- Humans MeSH
- Adolescent MeSH
- Male MeSH
- Child, Preschool MeSH
- Aged, 80 and over MeSH
- Aged MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
- Comparative Study MeSH
Fast and accurate detection of causative agents of bloodstream infections remains a challenge of today's microbiology. We compared the performance of cutting-edge technology based on polymerase chain reaction coupled with electrospray ionization-mass spectrometry (PCR/ESI-MS) with that of conventional broad-range 16S rRNA PCR and blood culture to address the current diagnostic possibilities for bloodstream infections. Of 160 blood samples tested, PCR/ESI-MS revealed clinically meaningful microbiological agents in 47 samples that were missed by conventional diagnostic approaches (29.4% of all analyzed samples). Notably, PCR/ESI-MS shortened the time to positivity of the blood culture-positive samples by an average of 34 hr. PCR/ESI-MS technology substantially improved current diagnostic tools and represented an opportunity to make bloodstream infections diagnostics sensitive, accurate, and timely with a broad spectrum of microorganisms covered.
Fast and accurate detection of causative agents of bloodstream infections remains a challenge of today's microbiology. We compared the performance of cutting-edge technology based on polymerase chain reaction coupled with electrospray ionization-mass spectrometry (PCR/ESI-MS) with that of conventional broad-range 16S rRNA PCR and blood culture to address the current diagnostic possibilities for bloodstream infections. Of 160 blood samples tested, PCR/ESI-MS revealed clinically meaningful microbiological agents in 47 samples that were missed by conventional diagnostic approaches (29.4% of all analyzed samples). Notably, PCR/ESI-MS shortened the time to positivity of the blood culture-positive samples by an average of 34 hr. PCR/ESI-MS technology substantially improved current diagnostic tools and represented an opportunity to make bloodstream infections diagnostics sensitive, accurate, and timely with a broad spectrum of microorganisms covered.
- MeSH
- Bacteremia diagnosis microbiology MeSH
- Molecular Diagnostic Techniques methods MeSH
- Adult MeSH
- Spectrometry, Mass, Electrospray Ionization methods MeSH
- Middle Aged MeSH
- Humans MeSH
- Microbiological Techniques MeSH
- Young Adult MeSH
- Polymerase Chain Reaction methods MeSH
- Reproducibility of Results MeSH
- RNA, Ribosomal, 16S MeSH
- Aged, 80 and over MeSH
- Aged MeSH
- Sepsis diagnosis microbiology MeSH
- Check Tag
- Adult MeSH
- Middle Aged MeSH
- Humans MeSH
- Young Adult MeSH
- Male MeSH
- Aged, 80 and over MeSH
- Aged MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
The aging population and increased incidence of severe bacterial infection can lead to sepsis. Interest to early identification of endangered patients and identification of pathogen do not always confirm the infection. To use biomarkers can help in early identification of infection and opportunity to start therapy timeously. All biomarkers were defined in 33 out of 96 patients. Thirty-two (97 %) patients had bacterial infection and 1 (3 %) patient had systemic inflammatory response syndrome (SIRS) without infection. PCR confirmed the infection in 27 cases and blood cultures in 8. Area under curve (AUC) for CD64 was 1.00, meanwhile other biomarkers showed 2-fold smaller AUC for positive infection. CD64 index was associated with bacterial infection (p<0.001) and could be used to confirm assessment of SIRS severity (p=0.037). As regards to our results, limited to only 33 patients, CD64 index served as a good parameter to predict bacterial infection and determine severity. The use of broad range 16S ribosomal RNA (rRNA) PCR proved to be an excellent tool to confirm bloodstream infection. The CD64 index had the highest AUC, which exceeded all the others, and could be used to predict the outcome of broad range 16S rRNA PCR from whole blood. However, C-reactive protein (CRP), procalcitonin (PCT) and sCD14 are much easier and faster to measure, but the values could be elevated in other clinical assessments.
- MeSH
- Bacteria genetics isolation & purification MeSH
- Bacteremia diagnosis MeSH
- Biomarkers analysis MeSH
- Diagnostic Tests, Routine methods MeSH
- DNA, Bacterial genetics MeSH
- Humans MeSH
- Neutrophils chemistry MeSH
- Polymerase Chain Reaction MeSH
- Receptors, IgG analysis MeSH
- DNA, Ribosomal chemistry genetics MeSH
- RNA, Ribosomal, 16S genetics MeSH
- ROC Curve MeSH
- Severity of Illness Index * MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
Cíl: Zavedení, charakterizace a porovnaní dvou metod kvantitativního stanovení viru hepatitidy B-HBV DNA. Obě metody používají totožného principu měření na bázi reál time PCR. První z nich je prováděna pomocí analytického systému LightCycler Roche Diagnostics a druhá na přístroji Rotorgene Corbett Research. Předmětem porovnávání metod byla linearita a pracovní rozsah stanovení, mez detekce, klinická senzitivita a specifičnost obou metod a jejich diagnostická správnost. Materiál: Oběma metodami bylo simultánně analyzováno 68 sér pacientů s diagnostikovanou chronickou hepatitidou B v různých fázích choroby. Výsledky: Regresní analýzou jsme stanovili hodnotu korelačního koeficientu r= 0,995 (p < 0,0001), potvrzující velmi úzký vztah mezi souborem výsledků získaných oběma metodami. Přesnost měření obou metod nepřekračuje hodnotu CV = 15 %. Obě metody disponují vysokým rozsahem linearity měření v intervalu lO 3-lO 8 kopií HBV DNA/ml. Diagnostická správnost srovnávaných metod byla vyhodnocena ROC analýzou. Pro obě metody byla zjištěna hodnota klinické sensitivity 100 %, klinická specifičnost metody LightCycler činila 96 % ve srovnání s 95 % u metody Rotorgene. Klinická efektivita, vyjádřená hodnotou AUC (plochy pod krivkou) byla 0,995 (LightCycler) respektive 0,994 (Rotorgene). Metoda LightCycler vykázala vyšší hodnotu pozitivního pravděpodobnostního poměru + LR = 26 oproti hodnotě 19,5 u Rotorgenu. Závěr: Uvedené údaje vypovídají o velmi vysoké hodnotě klinické užitečnosti obou metod.
Objective: Introduce, characterize and compare two methods of hepatitis virus B DNA quantification in serum. Both methods are based on real time PCR principle. Two measurement systems, LightCycler Roche and Rotorgene Corbett Research were used. We evaluated and compared their precision, linearity, limit of detection, clinical sensitivity, clinical specificity and diagnostic accuracy. Materials: By use of mentioned methods we analysed 68 sera from patients with diagnosed chronic hepatitis B in different stages of disease. Results: Correlation coefficient 0,995 (p < 0,0001) obtained by linear regression analysis shows excellent relationship between results of evaluated methods. The coefficients of variation for repeatability and reproducibility indicate very good measurement precision and are in all cases lower than 15 %. Linearity of both methods has very broad range lO 3 -lO 8copies of HBV DNA/ml. It means that majority of samples may be analysed without repeating after sample dilution. By use of ROC analysis we determined AUC value 0,995, clinical sensitivity 100 %, specificity 96 % and positive likelihood ratio (+LR) 26,0 for LightCycler, resp. AUC = 0,994, specificity 95 %, +LR = 19,5 for Rotorgene. Conclusion: All analytical and clinical characteristics of introduced methods show their usefulness for diagnosis and treatment of chronic hepatitis B.
The isolation of nontuberculous mycobacteria (NTM) from clinical specimens has become very common in recent years. Such organisms are typically environmental and occasionally pathogenic for humans and animals. Standard diagnosis of mycobacterial infections relies on direct examination and culture. However, molecular tools are now available which allow quicker and more accurate diagnosis. Detection of NTM can be performed directly from clinical samples, although identification is mostly carried out after isolation. Sequencing of genomic targets (such as 16S rRNA, ITS, rpoB or hsp65) allows accurate and rapid identification, but has some technical limitations. A brief summary of the molecular methods available for NTM identification and a discussion of the problems associated with the use of sequencing analysis together with a description of available algorithms for NTM identification are the major objectives of this review.
- MeSH
- Algorithms MeSH
- RNA, Bacterial analysis genetics MeSH
- DNA, Bacterial analysis genetics MeSH
- Genome, Bacterial MeSH
- Humans MeSH
- Mycobacterium Infections diagnosis microbiology MeSH
- Nontuberculous Mycobacteria classification genetics isolation & purification MeSH
- Polymerase Chain Reaction methods MeSH
- Reproducibility of Results MeSH
- RNA, Ribosomal, 16S analysis genetics MeSH
- Sequence Analysis, DNA methods MeSH
- Sensitivity and Specificity MeSH
- Bacterial Typing Techniques methods MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Review MeSH
Závěrečná zpráva o řešení grantu Interní grantové agentury MZ ČR
46 l. : il., tab. ; 30 cm
Studie je zaměřena na rychlou diagnostiku bakteriální meningitidy pomocí širokospektrých a druhově specifických primerů. Porovnává senzitivitu a specifitu PCR s metodami latexové aglutinace, mikroskopie a kultivace.; Study is focused on quick PCR diagnostic of bacterial meningitis by broad-range and species-specific primers. The sensitivity and specificity of PCR is compared with other "classic" methods - latex aglutination, microscopy and cultivation.
- MeSH
- Early Diagnosis MeSH
- Molecular Diagnostic Techniques trends MeSH
- Meningitis, Bacterial diagnosis MeSH
- Polymerase Chain Reaction methods utilization MeSH
- Sensitivity and Specificity MeSH
- Serologic Tests MeSH
- Conspectus
- Biochemie. Molekulární biologie. Biofyzika
- NML Fields
- infekční lékařství
- biologie
- NML Publication type
- závěrečné zprávy o řešení grantu IGA MZ ČR
The aim of the study was to establish a diagnostic value for broad-range polymerase chain reaction (br-PCR) and staphylococci-specific multiplex PCR (ssm-PCR) performed on surgical material from patients with staphylococcal infective endocarditis (IE). Data were analysed retrospectively from 60 patients with suspected staphylococcal IE and 59 controls who were surgically treated at three cardiosurgery centres over 4 years. Both PCR tests showed high agreement and could be aggregated. In patients with definite and rejected IE, the clinical sensitivity and specificity of PCR reached 89 and 95%, respectively. Tissue culture (TC) and PCR agreed with blood culture (BC) in 29% and 67% of IE cases. TC helped to determine aetiology in five BC negative cases while PCR aided in nine cases. Out of 52 patients with conclusive staphylococcal IE, 40 were diagnosed with S. aureus and 12 with coagulase-negative staphylococci. PCR was shown to be highly superior to TC in confirming preoperative diagnosis of IE. In addition to aid in culture negative patients, PCR helped to establish or refine aetiology in inconclusive cases. We suggest that simultaneous br-PCR and ssm-PCR performed on surgical material together with histopathology could significantly increase the performance of current Duke criteria.
- MeSH
- Bacteriological Techniques methods MeSH
- Molecular Diagnostic Techniques methods MeSH
- Endocarditis diagnosis microbiology surgery MeSH
- Middle Aged MeSH
- Humans MeSH
- Polymerase Chain Reaction methods MeSH
- Retrospective Studies MeSH
- Sensitivity and Specificity MeSH
- Staphylococcal Infections diagnosis microbiology surgery MeSH
- Staphylococcus classification genetics isolation & purification MeSH
- Check Tag
- Middle Aged MeSH
- Humans MeSH
- Male MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
- Research Support, Non-U.S. Gov't MeSH
