PubMed Health⌕ Search

Biomedical subjects

Franklin R Cockerill

Publications and source records attributed to Franklin R Cockerill.

At least 19 recordsLinked to original sources
PubMed2006

Bronchoscopy in ventilator-associated pneumonia: agreement of calibrated loop and serial dilution.

RATIONALE: Although the serial dilution technique for quantitative culture of bronchoalveolar fluid is considered to be the gold standard for the diagnosis of ventilator-associated pneumonia, it is more labor intensive than the calibrated loop technique. OBJECTIVE: We sought to determine the agreement between the calibrated loop and serial dilution techniques in the diagnosis of ventilator-associated pneumonia. METHODS: We prospectively measured bacterial colony counts by the serial dilution and calibrated loop techniques in 121 bronchoalveolar lavage samples of 104 patients with suspected ventilator-associated pneumonia. MEASUREMENTS AND MAIN RESULTS: At the time of bronchoscopy, patients had received mechanical ventilation for a median of 8 d. Patients were receiving antibiotics when 90 of the 121 (74.4%) bronchoalveolar samples were obtained. The colony counts of 13 bacterial isolates were too numerous to count by the calibrated loop technique; by serial dilution technique, their counts ranged from 4.70 to 6.74 log10 cfu/ml. Fifty other bacteria had paired colony counts measured by each of the two techniques: the bias (95% confidence interval) between the two techniques was -0.380 (-0.665 to -0.095) log10 cfu/ml, with precision of 1.002 log10 cfu/ml and 95% limits of agreement of -2.344 to 1.584 log10 cfu/ml. Using the threshold of 4 log10 cfu/ml as a criterion for the diagnosis of ventilator-associated pneumonia, there was discordance only for one bacterial organism between the two techniques. CONCLUSIONS: The calibrated loop technique can be used for the diagnosis of ventilator-associated pneumonia using bronchoalveolar lavage fluid.

Anti-Bacterial Agents↗
PubMed2006

Unusual aerobic and anaerobic bacteria associated with prosthetic joint infections.

The association of certain microorganisms, such as Staphylococcus epidermidis, Staphylococcus aureus, and beta-hemolytic streptococci, with prosthetic joint infection (PJI) has been recognized for many years. To our knowledge, a systematic review of the presentation and management of less commonly encountered species of coagulase-negative staphylococci, nutritional-variant streptococci, aerobic non-spore and spore forming Gram-positive or anaerobic bacteria is not available. We therefore sought to provide a comprehensive literature review of PJI due to these microorganisms that will provide a valuable and quick reference for clinicians caring for these patients. We conducted a Medline search of all case reports and case series of PJI due to unusual aerobic and anaerobic bacteria. The presentation, surgical, and medical management strategies were reviewed. Appropriate medical and surgical management of such infections is complex and evolving as newer diagnostic tests, surgical techniques and antimicrobials become available. Management of patients with these infections requires close collaboration between the orthopaedic surgeon, infectious disease specialist and microbiology laboratory.

Bacteria, Aerobic↗
PubMed2006

Fungi, mycobacteria, zoonotic and other organisms in prosthetic joint infection.

A systematic review of the presentation and management of prosthetic joint infection (PJI) due to zoonotic microorganisms, fungi, mycobacteria and other unusual microorganisms is not available. We therefore sought to provide a resource for the practicing orthopaedic surgeon which will serve as a guide for making appropriate decisions in managing such rare, but potentially problematic conditions. We conducted a Medline search of all case reports of PJI due to these unusual microorganisms. Our review of the current literature showed the diagnosis of PJI due to zoonotic microorganisms, fungi, mycobacteria and other unusual microorganisms typically necessitates specialized diagnostic tests. Maintaining a high index of suspicion in diagnosis of such unusual microorganisms and requesting the appropriate laboratory tests at the time of surgical débridement is crucial for determining the microbiologic etiology of these infections. The appropriate medical and surgical management of such infections is complex and often requires the use of special antimicrobials with which the orthopaedist may not be familiar. Collaboration with an infectious disease specialist is advisable when treating these infections.

Humans↗
PubMed2006

Sonication of explanted prosthetic components in bags for diagnosis of prosthetic joint infection is associated with risk of contamination.

Explanted orthopedic implants from 54 patients with aseptic failure and 24 patients with prosthetic knee or hip infection were sonicated in polyethylene bags. The sensitivities of periprosthetic tissue and sonicate fluid cultures for the diagnosis of prosthetic joint infection were 54% and 75%, whereas the specificities were 98% and 87%, respectively. Sonication in bags improved bacterial recovery from the surface of orthopedic implants; however, it lacked specificity, due to bag leakage.

Adult↗
PubMed2005

Detection of Tropheryma whipplei DNA in clinical specimens by LightCycler real-time PCR.

A real-time PCR method using the LightCycler (Roche Applied Science, Indianapolis, IN) was compared to a conventional PCR assay for the detection of Tropheryma whipplei in 321 clinical specimens. The LightCycler method had sensitivity and specificity comparable to the conventional method but required considerably less labor and time (3 h versus 3 to 5 days).

Actinomycetales↗
PubMed2005

Use of the Roche LightCycler Strep B assay for detection of group B Streptococcus from vaginal and rectal swabs.

The results for a real-time PCR assay, using the LightCycler Strep B analyte-specific reagents (Roche Diagnostics Corporation, Indianapolis, Ind.), were compared to a direct plate method combined with a broth enrichment culture method for detection of group B streptococcus colonization in pregnant women. Two separate evaluations were conducted using two different automated nucleic extraction instruments, the MagNA Pure LC instrument (Roche Diagnostics Corporation) and the lower-capacity MagNA Pure Compact instrument (Roche Diagnostics Corporation). The sensitivities, specificities, and positive and negative predictive values for the different evaluation methods were as follow: for the LightCycler Strep B assay with MagNA Pure LC, 100, 97, 90, and 100%, respectively; for the LightCycler Strep B assay with MagNA Pure Compact, 92.5, 99, 97, and 97.5%, respectively. The LightCycler Strep B assay combined with either MagNA Pure LC or MagNA Pure Compact extraction is a suitable method for detecting group B streptococcus colonization in pregnant women. An advantage of the LightCycler assay over culture is the considerably reduced turnaround time for results.

Carrier State↗
PubMed2004

Evidence of nanobacterial-like structures in calcified human arteries and cardiac valves.

Mechanisms mediating vascular calcification remain incompletely understood. Nanometer scale objects hypothesized to be a type of bacteria (nanobacteria) are associated with calcified geological specimens, human kidney stones, and psammona bodies in ovarian cancer. Experiments were designed to evaluate human vascular tissue for the presence of similar nanometer-scale objects. Calcified human aneurysms (n = 8), carotid plaques (n = 2), femoral arterial plaques (n = 2), and cardiac valves (n = 2) and noncalcified aneurysms from patients with bicuspid aortic valve disease (n = 2) were collected as surgical waste from the Heart Hospital of Austin, Austin, Texas, and Mayo Clinic, Rochester, Minnesota. Whole mounts or adjacent sections from each specimen were examined by electron microscopy, stained for calcium phosphate, or stained with a commercially available antibody (8D10). Filtered (0.2 microm) homogenates of aneurysms were cultured and costained with 8D10 antibody followed by PicoGreen to detect DNA or incubated with [3H]uridine. Staining for calcium phosphate was heterogeneously distributed within all calcified tissues. Immunological staining with 8D10 was also heterogeneously distributed in areas with and without calcium phosphate. Analysis of areas with positive immunostaining identified spheres ranging in size from 30 to 100 nm with a spectral pattern of calcium and phosphorus (high-energy dispersive spectroscopy). Nanosized particles cultured from calcified but not from noncalcified aneurysms were recognized by a DNA-specific dye and incorporated radiolabeled uridine, and, after decalcification, they appeared via electron microscopy to contain cell walls. Therefore, nanometer-scale particles similar to those described as nanobacteria isolated from geological specimens and human kidney stones can be visualized in and cultured from calcified human cardiovascular tissue.

Aged↗
PubMed2004

Purification and characterization of Mycobacterium tuberculosis KatG, KatG(S315T), and Mycobacterium bovis KatG(R463L).

Isoniazid, a first-line antibiotic used for the treatment of tuberculosis, is a prodrug that requires activation by the Mycobacterium tuberculosis enzyme KatG. The KatG(S315T) mutation causes isoniazid resistance while the KatG(R463L) variation is thought to be a polymorphism. Much of the work to date focused on isoniazid activation by KatG has utilized recombinant enzyme overexpressed in Escherichia coli. In this work, native KatG and KatG(S315T) were purified from M. tuberculosis, and KatG(R463L) was purified from Mycobacterium bovis. The native molecular weight, enzymatic activity, optical, resonance Raman, and EPR spectra, K(D) for isoniazid binding, and isoniazid oxidation rates were measured and compared for each native enzyme. Further, the properties of the native enzymes were compared and contrasted with those reported for recombinant KatG, KatG(S315T), and KatG(R463L) in order to assess the ability of the recombinant enzymes to act as good models for the native enzymes.

Amino Acid Substitution↗
PubMed2003

Evaluation of the MagNA pure LC instrument for extraction of hepatitis C virus RNA for the COBAS AMPLICOR Hepatitis C Virus Test, version 2.0.

The COBAS AMPLICOR system has played a major role in the transition of molecular diagnostics from research to routine clinical laboratory use by automating the nucleic acid amplification and detection processes. However, sample preparation remains a labor-intensive portion of the procedure. In this study, we evaluated the performance of the COBAS AMPLICOR Hepatitis C Virus Test, version 2.0 (Roche Molecular Systems, Branchburg, N.J.) following manual hepatitis C virus (HCV) RNA extraction versus automated extraction with the MagNA Pure LC instrument (Roche Applied Science, Indianapolis, Ind.). Parallel replicate testing was performed with standard dilutions of 100, 75, 60, and 0 HCV IU/ml and 153 clinical specimens. An analytical sensitivity of 75 IU/ml was achieved with either the manual or the standard-volume (200 microl) automated extraction methodologies (25 of 26 [96.2%]; 95% confidence interval [95% CI], 80.4 to 99.9), whereas the clinical sensitivity and specificity were both 100% with either extraction method. A large-volume (1 ml) automated extraction method was also evaluated with standard dilutions of 40, 25, 10, and 0 IU/ml and the same 153 clinical specimens. The analytical sensitivity of the COBAS AMPLICOR assay with the large-volume extraction method was 25 HCV IU/ml (26 of 26 [100%]; 95% CI, 86.8 to 100), whereas the clinical sensitivity and specificity were both 100%. The MagNA Pure LC instrument is a versatile, labor-saving platform capable of integration with minimal modification of the existing assay procedure. The increased sensitivity of the COBAS AMPLICOR Hepatitis C Virus Test, version 2.0 performed in conjunction with large-volume HCV RNA extraction may be important in HCV diagnostic testing as new therapeutic strategies evolve.

Hepacivirus↗
PubMed2003

Application of rapid-cycle real-time polymerase chain reaction for diagnostic testing in the clinical microbiology laboratory.

CONTEXT: Rapid-cycle real-time polymerase chain reaction (PCR) technology combines rapid thermocycling with real-time fluorescent probe detection of amplified target nucleic acids. OBJECTIVES: To review and compare the method of rapid-cycle real-time PCR to conventional PCR methods. To describe the application of rapid-cycle real-time PCR for diagnostic testing in the microbiology laboratory. DATA SELECTION: Information is presented from published literature as well as from personal experience at the Mayo Clinical Microbiology Laboratory (Rochester, Minn). CONCLUSIONS: Compared to conventional PCR methods, rapid-cycle real-time PCR diagnostics are much faster and easier to perform, and, because both PCR and probe detection occur in the same reaction vessel, the possibility of amplified product (amplicon) contamination is lessened. Furthermore, compared to conventional culture-based or direct antigen detection methods, rapid-cycle real-time PCR assays are frequently more sensitive and much more rapid techniques for detecting or quantifying microorganisms in human samples and for identifying genes or mutations in pathogens associated with antimicrobial resistance.

Bacteria↗
PubMed2002

Comparison of line probe assay and DNA sequencing of 5' untranslated region for genotyping hepatitis C virus: description of novel line probe patterns.

We compared a commercial line probe assay (INNO-LiPA HCV II, Innogenetics, N.V., Ghent, Belgium, distributed by Bayer Diagnostics) to an in-house 5' untranslated region direct DNA sequencing method for genotyping hepatitis C virus (HCV). Initial evaluation demonstrated that the INNO-LiPA HCV II assay and sequencing assay assigned the same genotype for 110/132 (83.3%) patient specimens (98 subtype and 12 genotype only identifications). Following the initial evaluation, the INNO-LiPA HCV II assay was used routinely to genotype HCV from patient specimens submitted to our laboratory for genotyping (n = 1,739). During this second part of the study, novel line probe patterns have been noted and interpreted using the in-house direct sequencing assay. Reactivity at bands 1, 2, 3, 4, 5 and 8 (n = 4) or 1, 2, 3, 4, 6 and 7 (n = 2) represented HCV genotype 1. Reactivity at bands 1, 2, 5 and 9 (n = 1) represented HCV genotype 2. Reactivity at bands 1, 2, 5, 9 and 16 (n = 1) represented HCV genotype 4. Reactivity at bands 1, 2, 5, 9, 10, 11 (weak band) and 12 (n = 118) most likely represented HCV genotype 2b. This information should be of use to INNO-LiPA HCV II assay users.

5' Untranslated Regions↗
PubMed2002

Clinical features of patients with novel Yersinia species.

Our purpose was to describe the clinical features of patients with novel Yersinia species. Between 1985 and 1999; 194 patients had yersinia species isolated from stool specimens, 38 (20%) had non-Yersinia enterocolitica species; 12 (32%) had Yersinia intermedia, 7 (18%) Yersinia fredericksenii, 3 (8%) Yersinia kristensenii, and the remaining 16 (42%) were unclassified non-Yersinia enterocolitica species. The most common presenting symptom was diarrhea alone in 10 (26.3%) patients. Symptoms persisted for >1 month in 54% of cases; 21% had symptoms for <1 week; 18 (47%) patients were taking corticosteroids, acid suppressants, and/or antibiotics when Yersinia was isolated. An immunocompromised state was present in 11 (29%) patients. An immunocompromised host and administration of acid suppressants predicted persistence of symptoms for >1 month. Most [27 (71%)] patients received no treatment; 5 (13%) received antibiotics. In conclusion, novel non-Yersinia enterocolitica species, including Yersinia intermedia, Y. fredericksenii, and Y. kristensenii may represent up to 20% of all Yersinia isolates. Diarrhea is the most common symptom.

Adolescent↗
PubMed2002

Oxidative stress increases susceptibility of Mycobacterium tuberculosis to isoniazid.

Isoniazid is a first-line antibiotic used in the treatment of infections caused by Mycobacterium tuberculosis. Isoniazid is a prodrug requiring oxidative activation by the catalase-peroxidase hemoprotein, KatG. Resistance to isoniazid can be obtained by point mutations in the katG gene, with one of the most common being a threonine-for-serine substitution at position 315 (S315T). The S315T mutation is found in more than 50% of isoniazid-resistant clinical isolates and results in an approximately 200-fold increase in the MIC of isoniazid compared to that for M. tuberculosis H37Rv. In the present study we investigated the hypothesis that superoxide plays a role in KatG-mediated isoniazid activation. Plumbagin and clofazimine, compounds capable of generating superoxide anion, resulted in a lower MIC of isoniazid for M. tuberculosis H37Rv and a strain carrying the S315T mutation. These agents did not cause as great of an increase in isoniazid susceptibility in the mutant strain when the susceptibilities were assessed by using the inhibitory concentration that causes a 50% decrease in growth. These results provide evidence that superoxide can play a role in isoniazid activation. Since clofazimine alone has antitubercular activity, the observation of synergism between clofazimine and isoniazid raises the interesting possibility of using both drugs in combination to treat M. tuberculosis infections.

Antitubercular Agents↗
PubMed2002

Multiplex LightCycler PCR assay for detection and differentiation of Bordetella pertussis and Bordetella parapertussis in nasopharyngeal specimens.

A rapid real-time multiplex PCR assay for detecting and differentiating Bordetella pertussis and Bordetella parapertussis in nasopharyngeal swabs was developed. This assay (LC-PCR-IS) targets the insertion sequences IS481 and IS1001 of B. pertussis and B. parapertussis, respectively, and is performed using the LightCycler (Roche Molecular Biochemicals, Indianapolis, Ind.). The analytical sensitivity is less than one organism per reaction. Results for Bordetella culture and/or direct fluorescent antibody testing and a second LightCycler PCR assay (target, pertussis toxin gene) were compared to results of the LC-PCR-IS assay for 111 nasopharyngeal swabs submitted for pertussis testing. Of the specimens, 12 were positive (9 B. pertussis and 3 B. parapertussis) and 68 specimens were negative by all methods. Three other specimens were positive for B. pertussis by at least two of the methods (including the LC-PCR-IS assay), and another 28 specimens were positive for B. pertussis by the LC-PCR-IS assay only. No specimens were negative by the LC-PCR-IS assay and positive by the other methods. A conventional PCR method (target, IS481) was also compared to the LC-PCR-IS assay for a different group of nasopharyngeal swab specimens (n = 96): 44 specimens were positive and 41 specimens were negative for B. pertussis with both PCR methods. Nine specimens were positive for B. pertussis by the LC-PCR-IS assay and negative by the conventional PCR assay, and two specimens were positive for B. pertussis by the conventional PCR assay and negative by the LC-PCR-IS assay. Positivity of the two assays was not significantly different (P = 0.0654). The insertion sequence IS481 is also present in Bordetella holmesii; specimens containing B. holmesii may yield false-positive results. The LC-PCR-IS assay takes approximately 45 min to complete post-nucleic acid extraction, compared to 24 h for the conventional PCR assay previously used in our laboratory. The LC-PCR-IS assay is easier to perform than the conventional PCR assay, and the closed system decreases the chance of contamination. All of these characteristics represent a significant improvement in the detection of B. pertussis and B. parapertussis in nasopharyngeal specimens.

Bordetella↗
PubMed2002

Comparison of the Borrelia DotBlot G, MarDx, and VIDAS enzyme immunoassays for detecting immunoglobulin G antibodies to Borrelia burgdorferi in human serum.

Three enzyme immunoassays, Borrelia DotBlot G (GenBio, San Diego, Calif.), MarDx EIA (MarDx Diagnostics, Inc., Carlsbad, Calif.), and VIDAS (bioMérieux, St. Louis, Mo.) were compared for their ability to detect immunoglobulin G antibodies to Borrelia burgdorferi in 100 human serum samples. The "gold standard" positive result for each of these samples was determined by Western immunoblot analysis (MarDx Marblot Test System) (n = 99) or clinical signs and symptoms suggestive of Lyme disease with other laboratory results positive for B. burgdorferi (n = 1). Based on these criteria for a gold standard positive result, 29 of the 100 samples tested were considered true positives and 71 were considered true negatives. The following sensitivities and specificities were noted, respectively, for each method: Borrelia DotBlot G, 93 and 90%; MarDx, 100 and 35%; and VIDAS, 100 and 92%. Because of high sensitivity and specificity and ease of use, the VIDAS test is an appealing method, especially for laboratories that perform high volumes of tests. The high sensitivity but low specificity of the MarDx method compared to the VIDAS and Borrelia DotBlot G methods requires that Western blot confirmatory tests be performed frequently. The Borrelia DotBlot G method has acceptable specificity but appears to lack sensitivity when compared to the VIDAS and MarDx methods.

Antibodies, Bacterial↗
PubMed2002

Detection of Bacillus anthracis DNA by LightCycler PCR.

Anthrax is a zoonotic disease that is also well recognized as a potential agent of bioterrorism. Routine culture and biochemical testing methods are useful for the identification of Bacillus anthracis, but a definitive identification may take 24 to 48 h or longer and may require that specimens be referred to another laboratory. Virulent isolates of B. anthracis contain two plasmids (pX01 and pX02) with unique targets that allow the rapid and specific identification of B. anthracis by PCR. We developed a rapid-cycle real-time PCR detection assay for B. anthracis that utilizes the LightCycler instrument (LightCycler Bacillus anthracis kit; Roche Applied Science, Indianapolis, Ind.). PCR primers and probes were designed to identify gene sequences specific for both the protective antigen (plasmid pX01) and the encapsulation B protein (plasmid pX02). The assays (amplification and probe confirmation) can be completed in less than 1 h. The gene encoding the protective antigen (pagA) was detected in 29 of 29 virulent B. anthracis strains, and the gene encoding the capsular protein B (capB) was detected in 28 of 29 of the same strains. Three avirulent strains containing only pX01 or pX02, and therefore only pagA or pagB genes, could be detected and differentiated from virulent strains. The assays were specific for B. anthracis: the results were negative for 57 bacterial strains representing a broad range of organisms, including Bacillus species other than anthracis (n = 31) and other non-Bacillus species (n = 26). The analytical sensitivity demonstrated with target DNA cloned into control plasmids was 1 copy per microl of sample. The LightCycler Bacillus anthracis assay appears to be a suitable method for rapid identification of cultured isolates of B. anthracis. Additional clinical studies are required to determine the usefulness of this test for the rapid identification of B. anthracis directly from human specimens.

Antigens, Bacterial↗