Broad-range Bacterial Polymerase Chain Reaction Research Articles

A Dual-Pathogen Mitral Valve Endocarditis Caused by Coxiella burnetii and Streptococcus gordonii-Which Came First?

Infective endocarditis (IE) is still a life-threatening disease with high morbidity and mortality. While usually caused by a single bacterium, poly-microbial infective endocarditis (IE) is rare. Here, we report a (blood-culture-negative) dual pathogen mitral valve IE caused by Coxiella burnetii and Streptococcus gordonii: A 53-year-old woman was presented to an internal medicine department with abdominal pain for further evaluation. Within the diagnostic work up, transthoracic echocardiography (TTE) revealed an irregularly shaped echogenic mass (5 × 13 mm) adherent to the edge of the posterior mitral valve leaflet and protruding into the left atrium. As infected endocarditis was suspected, blood cultures were initially obtained, but they remained negative. Chronic Q fever infection was diagnosed using serologic testing. After the occurrence of cerebral thromboembolic events, the patient was admitted for mitral valve surgery. Intraoperatively, a massively destructed mitral valve with adhering vegetations was noted. Examination of the mitral valve by broad-range bacterial polymerase chain reaction (PCR) and amplicon sequencing confirmed Coxiella burnetii infection and yielded Streptococcus gordonii as the second pathogen. Based on the detailed diagnosis, appropriate antibiotic therapy of both pathogens was initiated, and the patient could be discharged uneventfully on the 11th postoperative day after a successful minimal-invasive mitral valve replacement.

Development of a scoring system to identify high-yield specimens for bacterial broad-range 16S rRNA gene PCR with sequencing at a tertiary care medical center.

Broad-range bacterial polymerase chain reaction with sequencing (BRBPS) provides valuable diagnostic data, especially in cases of culture-negative infections. However, as BRBPS testing demonstrates generally low positivity, cost per impactful result can be high and commonly involves longer turnaround times compared with other methods, targeting use of this assay to high-yield situations remains a challenging goal. Diagnostic stewardship can help alleviate these challenges and increase clinical utility, yet not all laboratories have a dedicated stewardship team, and little formal guidance exists on identifying high-yield samples outside of specific clinical syndromes. In this study, we performed a retrospective review of 86 BRBPS orders from a tertiary care medical center, with a focus on identifying high-yield cases using an infectious markers scoring system, visualized inflammation or organism (VIO) score, to predict return of actionable diagnostic data. Using chart review, we evaluated how results from high VIO score or low VIO score specimens contributed to clinical management. Testing low VIO score samples identified an organism in only 10% of samples, and of these positive results, 33% were considered to represent contamination. Despite negative routine workup and no identified pathogen via BRBPS, broad antimicrobial treatment was continued in 85% of cases with a low VIO score. In contrast, specimens with high VIO scores were more predictably positive by BRBPS, identified organisms that were universally considered pathogens, and provided opportunities to target or de-escalate antimicrobial therapy. This study describes the VIO scoring system to guide the identification of high-yield samples and steward the appropriate use of BRBPS testing.

Disseminated Nocardia in an Immunocompetent Host

A 44-year-old man with no medical comorbidities presented with recurrent inferior vena cava thrombosis and several months of fever, headache, and weight loss. Positron emission tomography–computed tomography was performed, which revealed a 1.5-cm fluorodeoxyglucose avid left lung mass and hypermetabolic soft tissue masses in bilateral inguinal regions (Figure 1). He underwent computed tomography–guided biopsy of the lung and left inguinal masses. Histopathology found clumps of filamentous branching bacilli, consistent with Nocardia (Supplemental Figure, available online at http://www.mayoclinicproceedings.org). Broad-range bacterial polymerase chain reaction (16s rRNA detection) returned positive for Nocardia paucivorans. Subsequent brain magnetic resonance imaging exhibited innumerable brain abscesses (Figure 2). Immunodeficiency work-up was performed and was unremarkable. He began therapy with intravenous trimethoprim-sulfamethoxazole, amikacin, and imipenem.Figure 2Abscesses with surrounding vasogenic edema on T2-weighted fluid-attenuated inversion recovery magnetic resonance imaging.View Large Image Figure ViewerDownload Hi-res image Download (PPT) Nocardia is a filamentous, Gram-positive, partially acid-fast bacterium that is ubiquitous in the environment.1Lerner P.I. Nocardiosis.Clin Infect Dis. 1996; 22: 891-903Crossref PubMed Scopus (499) Google Scholar It is capable of disseminating to almost any organ, but it has a predilection for the lungs and brain. It also has a tendency to recur. Disseminated infection is more common in immunocompromised hosts, but approximately one-third of infections occur in patients who are immunocompetent.2Corti M.E. Villafañe-Fioti M.F. Nocardiosis: a review.Int J Infect Dis. 2003; 7: 243-250Abstract Full Text PDF PubMed Scopus (170) Google Scholar, 3Dominguez D.C. Antony S.J. Actinomyces and Nocardia infections in immunocompromised and nonimmunocompromised patients.J Natl Med Assoc. 1999; 91: 35-39PubMed Google Scholar, 4Sorrell T.C. Mitchell D.H. Iredell J.R. Chen S.C.-A. Nocardia species.in: Bennett J.E. Dolin R. Blaser M.J. Mandell, Douglas, and Bennett's Principles and Practice of Infectious Diseases. 8th ed. Elsevier, Philadelphia, PA2015: 2853-2863Crossref Scopus (16) Google Scholar Treatment for disseminated disease requires induction with intravenous multidrug therapy. Trimethoprim-sulfamethoxazole forms the backbone of induction therapy and is combined with one or both of amikacin and imipenem. Imipenem is required if there is central nervous system involvement. Treatment should be tailored based on susceptibilities, if available. After induction, patients may be transitioned to dual oral therapy with trimethoprim-sulfamethoxazole and either minocycline or amoxicillin-clavulanate. A total of 12 months of therapy is typical for immunocompromised patients and those with central nervous system involvement.4Sorrell T.C. Mitchell D.H. Iredell J.R. Chen S.C.-A. Nocardia species.in: Bennett J.E. Dolin R. Blaser M.J. Mandell, Douglas, and Bennett's Principles and Practice of Infectious Diseases. 8th ed. Elsevier, Philadelphia, PA2015: 2853-2863Crossref Scopus (16) Google Scholar We acknowledge and thank Julie Guerin, MD, and Ayse Tuba Kendi, MD, for interpreting and providing the images of the brain magnetic resonance imaging and positron emission tomography–computed tomography, respectively.

Detection and Identification of Microorganisms by Gene Amplification and Sequencing

Gene amplification and sequencing have led to the discovery of new pathogens as agents of disease and have enabled us to better classify microorganisms from culture. Sequence-based identification of bacteria and fungi using culture is more objective and accurate than conventional methods, especially for classifying unusual microorganisms that are emerging pathogens in immunocompromised hosts. Although a powerful tool, the interpretation of sequence-based classification can be challenging as microbial taxonomy grows more complex, without known clinical correlatives. Additionally, broad-range gene polymerase chain reaction and sequencing have emerged as alternative, culture-independent methods for detecting pathogens from clinical material. The promise of this technique has remained strong, limited mainly by contamination and inadequate sensitivity issues. This review explains sequence-based microbial classification, with emphasis on relating the complex world of microbial taxonomy to a clinical context. Additionally, this review discusses a rational approach to broad-range bacterial polymerase chain reaction and gene sequencing when applied directly to clinical samples.

Primary and opportunistic pathogens associated with meningitis in adults in Bangui, Central African Republic, in relation to human immunodeficiency virus serostatus

To determine the causative organisms and characteristics of patients presenting with meningitis in Bangui in order to provide guidance to physicians for case management. Adults with proven or suspected meningitis were enrolled in this prospective study. Full blood count, blood chemistry, and HIV tests were performed. Cerebrospinal fluid (CSF) was submitted for routine microbiology, chemistry (glucose, protein), and hematology testing. When classical microbiology analyses were negative, a broad-range bacterial polymerase chain reaction (BRBPCR) was used. Of the 276 patients enrolled, 215 (77.9%) were HIV positive. In HIV-positive patients cryptococcal meningitis (CM) was the most common cause of meningitis (39.1%) followed by pyogenic meningitis (PM) (30.7%), mononuclear meningitis (MM) (28.8%), and tuberculous meningitis (TM) (1.4%). In HIV-negative patients, PM was the most common cause (60.7%) followed by MM (37.7%) and CM (1.6%, one case). In-hospital mortality was higher in HIV-positive patients (73/128 = 57%) compared to those HIV negative (3/18 = 16.7%) (p = 0.001). Streptococcus pneumoniae (n = 26) was the most common bacterial diagnosis, mainly in HIV-positive patients (n = 22, 10.2%). Meningococcal meningitis (14 Neisseria meningitidis of group A and one W135) was diagnosed in nine (4.2%) HIV-positive and six (9.8%) HIV-negative patients. Gram-negative rods were isolated from five HIV-positive and two HIV-negative patients, respectively. The bacteria and fungi involved in meningitis did not display high levels of in vitro resistance. Conventional microbiology techniques failed to detect the causative agent in 55 (53.4%) PM cases. Broad-range bacterial PCR detected DNA from S. pneumoniae in three samples, N. meningitidis in two, Escherichia coli in one, Listeria monocytogenes in two and Staphylococcus aureus in one sample. In the CSF of five (three HIV negative and two HIV positive), PCR products were not identified with the oligonucleotide probes specific for the usual species of bacteria found in CSF, or genera commonly considered potential contaminants of clinical samples. Among the MM cases, 77 (90.5%) probable viral meningitis (54 HIV positive and 23 HIV negative) and eight TM (HIV positive) were suspected.

Aetiological diagnosis of brain abscesses and spinal infections: application of broad range bacterial polymerase chain reaction analysis

Objective: To evaluate the usefulness of the broad range bacterial rDNA polymerase chain reaction (PCR) method combined with DNA sequencing in the aetiological diagnosis of intracranial or spinal infections in...

Broad-range bacterial polymerase chain reaction for early detection of bacterial meningitis.

The diagnosis of bacterial meningitis often depends on isolation of bacteria on culture, which may take 24-48 h. DNA amplification techniques could provide rapid diagnosis, which would guide the clinician in antimicrobial therapy decisions. This study determined the clinical utility of polymerase chain reaction (PCR) for the diagnosis of meningitis with use of a broad range of bacterial primers. Seventy-four cerebrospinal fluid specimens obtained from 70 patients were subjected to PCR with use of primers derived from conserved regions of the bacterial 16S RNA gene. The test characteristics for the broad-range bacterial PCR were as follows: sensitivity, 100%; specificity, 98.2%; positive predictive value, 94.4%; and negative predictive value, 100%. Broad-range bacterial PCR may be useful for excluding the diagnosis of meningitis, and the results may influence the decision to initiate or discontinue antimicrobial therapy.

Broad-range bacterial polymerase chain reaction for identification of bacteria in inflamed joints: comment on the article by Wilbrink et al

Broad-range bacterial polymerase chain reaction for identification of bacteria in inflamed joints: comment on the article by Wilbrink et al