Larry G. Reimer

Update on detection of bacteremia and fungemia.

The presence of microorganisms in a patients blood is a critical determinant of the severity of the patients illness. Equally important, the laboratory isolation and identification of a microorganism present in blood determine the etiologic agent of infection, especially when the site of infection is localized and difficult to access. This review addresses the pathophysiology and clinical characteristics of bacteremia, fungemia, and sepsis; diagnostic strategies and critical factors in the detection of positive blood cultures; characteristics of manual and instrument approaches to bacteremia detection; approaches for isolating specific microorganisms associated with positive blood cultures; and rapid methods for the identification of microorganisms in blood cultures.

Relevance of the Number of Positive Bottles in Determining Clinical Significance of Coagulase-Negative Staphylococci in Blood Cultures

ABSTRACT Coagulase-negative staphylococci (CNS) are the most commonly isolated contaminants from blood cultures, yet they frequently cause true infections. Determining the clinical significance of CNS is difficult, and clinicians often consider the number of positive bottles within a set of blood culture bottles in their assessment. Therefore, in three separate studies, we counted the number of positive bottles within blood culture sets comprising two, three, or four bottles in order to predict whether or not CNS were clinically significant isolates (CSI) in adult patients with suspected sepsis. Each culture was evaluated by independent, published clinical criteria to determine its clinical importance. Of 486 positive sets that included two adequately filled bottles, 127 (26%) CNS were CSI, 329 (67%) were contaminants, and 30 (6%) were indeterminate as a cause of sepsis. Among CSI, 39 and 61% were isolated from one and two bottles, respectively. The positive predictive value for sepsis was 18% when one bottle was positive and 37% when both bottles were positive. Of 235 positive sets that included three adequately filled bottles, 81 (34%) were CSI, 109 (46%) were contaminants, and 45 (19%) were indeterminate as a cause of sepsis. Of CSI, 43, 38, and 19% were found in one, two, and three bottles, respectively. The positive predictive value for sepsis was 28, 52, and 30% when one, two and three bottles were positive. Of 303 positive blood culture sets that included four adequately filled bottles, 64 (21%) were considered CSI, 197 (65%) were contaminants, and 42 (14%) were indeterminate as a cause of sepsis. Of CSI, 27, 28, 19, and 27% were found in one, two, three, and four bottles, respectively. The positive predictive value for sepsis was 11, 30, 34, and 37% when one, two, three, and four bottles were positive. We conclude that the number of culture bottles positive in a given culture set cannot reliably predict the clinical significance of the CNS isolated and, therefore, should not be used as a criterion for determining whether or not an isolate represents true infection or contamination.

Role of the Microbiology Laboratory in the Diagnosis of Lower Respiratory Tract Infections

The appropriate use of the clinical microbiology laboratory for diagnosing lower respiratory tract infections is controversial. As in clinical care, it is crucial to categorize the presenting illness properly as acute bronchitis, an acute exacerbation of chronic bronchitis, community-acquired pneumonia, or nosocomial pneumonia if diagnostic efforts to establish a microbial etiology are to be productive for the individual patient and affordable to society. The greatest potential benefit of microbiological investigations lies in the etiologic diagnosis of pneumonia. For community-acquired pneumonia, evaluation of a gram-stained smear of sputum in terms of both quality and microorganisms present can help guide initial therapy as well as aid interpretation of subsequent culture results. As discussed in this review, the role of the clinical microbiology laboratory in the etiologic diagnosis of nosocomial and complicated pneumonias is more extensive and, in addition to evaluation of respiratory secretions, may include cultures of blood, pleural fluid, and specimens obtained by bronchoscopy. However, a prerequisite for the use of all currently available tests is their deployment for patients with clinical and radiographic evidence of pneumonia.

Human Immunodeficiency Virus Type 1 Drug Resistance Testing: a Comparison of Three Sequence-Based Methods

ABSTRACT The use of genotypic assays for determining drug resistance in human immunodeficiency virus (HIV) type 1 (HIV-1)-infected patients is increasing. These tests lack standardization and validation. The aim of this study was to evaluate several tests used for the determination of HIV-1 drug resistance. Two genotypic tests, the Visible Genetics TruGene HIV-1 Genotyping Kit and the Applied Biosystems HIV Genotyping System, were compared using 22 clinical samples. Genotyping results were also obtained from an independent reference laboratory. The Visible Genetics and Applied Biosystems genotyping tests identified similar mutations when differences in the drug databases and reference strains were taken into account, and 19 of 21 samples were equivalent. The concordance between the two assays was 99% (249 of 252 mutation sites). Mutations identified by the reference laboratory varied the most among those identified by the three genotypic tests, possibly because of differences in the databases. The concordance of the reference laboratory results with the results of the other two assays was 80% (201 of 252). Samples with 500 to 750 HIV RNA copies/ml could be sequenced by the Visible Genetics and Applied Biosystems assays using 1 ml of input. The Visible Genetics and Applied Biosystems assays both generated an accurate sequence. However, the throughput of the Visible Genetics assay is more limited and may require additional instruments. The two assays differ technically but are similar in overall complexity. Data analysis in the two assays is straightforward, but only the reports provided by Visible Genetics contain information relating mutations to drug resistance. HIV drug resistance genotyping by sequencing is a complex technology which presents a challenge for analysis, interpretation, and reporting.

Recovery of clinically important microorganisms from the BacT/Alert blood culture system does not require testing for seven days

Recently, we published a comparison of the BacT/Alert blood culture system with the BACTEC 660/730 nonradiometric blood culture system using blood inocula of 5 ml per bottle. By reanalyzing data collected during that study, we found that, for true-positive isolates causing bacteremia or fungemia, 363 (97.6%) of 376 and 341 (97.7%) of 349 isolates were recovered by the end of day 5 of testing, and 364 (97.9%) of 376 and 343 (98.3%) of 349 isolates were recovered by the end of day 6 of testing for aerobic and anaerobic bottles, respectively. Most isolates recovered on days 6 (24 of 27) and 7 (20 of 25) of testing were either contaminants or indeterminate as a cause of sepsis. When used as recommended by the manufacturer, only six (1.3%) of 464 clinically important isolates recovered on test days 6-7 would have gone undetected had testing been limited to 5 days and four (0.9%) of 464 had testing been limited to 6 days. We conclude that BacT/Alert bottles can be tested for as few as 5 days and then discarded with minimal loss of true-positive isolates and maximal reduction of contaminants.

Effect of antimicrobials on blood cultures in endocarditis

To study the effect of antimicrobials on bacterial growth in blood cultures, we used both simulated blood cultures and cultures obtained from rabbits with experimental endocarditis. Four strains of bacteria were incubated individually with six antimicrobials in nine blood culture media. Positivity rates varied with the ratio of the antimicrobial concentration to the MIC of the organism: 161 of 162 cultures (99%) were positive when the ratio was less than 1/10; 52 of 108 (48%) were positive when the ratio was between 1/10 and one; and none of 54 were positive when the ratio was greater than one. Endocarditis was produced in 28 rabbits with either E. coli, P. aeruginosa, S. aureus, or viridans streptococcus. Following a single dose of an antimicrobial, blood was taken for culture in eight media. Only for viridans streptococcus did recovery rates vary significantly in different media. Recovery rates for this organism in two supplemented peptone broths (78% and 89%) and in hypertonic supplemented peptone (78%) were each higher than in thioglycolate (22%), Columbia (22%), Bactec aerobic and anaerobic (11%), and trypticase soy broths (11%) (p less than 0.05 for each pair). Growth of bacteria in blood cultures containing antimicrobials depended on the ratio of the antimicrobial concentration to the MIC and, for viridans streptococcus, the blood culture medium.

Controlled Comparison of BacT/ALERT FAN Aerobic Medium and BACTEC Fungal Blood Culture Medium for Detection of Fungemia

ABSTRACT Yeasts are an increasingly common cause of nosocomial bloodstream infections. Methods for their detection are many; controlled comparisons are few. The vented FAN aerobic blood culture medium has been shown to be superior to the standard BacT/ALERT aerobic medium for the detection of fungemia as well as bacteremia. The BACTEC selective fungal medium (FM) (BD Biosciences, Sparks, Md.) allowed detection of more episodes of fungemia than did a resin-containing medium with equal volumes of blood cultured. Therefore, we compared vented FAN to FM for the ability to recover fungi from the blood of patients who were at increased risk of having fungemia. From 5,109 cultures processed for which both FAN and FM bottles were adequately filled, fungi were recovered from 87 cultures. Of these, 47 were detected with both bottles, 12 were detected with FAN only, and 28 were detected with FM only (P < 0.05). FAN was the first bottle positive for 36 of the 47 cultures for which both bottles yielded the same fungus, whereas the FM bottle was the first bottle positive for 11 cultures (P < 0.001). A total of 54 episodes of fungemia were identified, with 40 detected by both media, 4 detected only by FAN, and 10 detected only by FM (P value, not significant). We conclude that the vented FAN aerobic bottle is comparable to the FM bottle for detection of episodes of yeast infection but has the added benefit of detecting bacteria.

Acute Infection With Sin Nombre Hantavirus without Pulmonary Edema

Acute infection with Sin Nombre virus has been associated with development of hantavirus cardiopulmonary syndrome (HCPS), a severe cardiopulmonary illness with respiratory failure and shock. We present two cases of Sin Nombre hantavirus infections that did not lead to marked pulmonary complications in two otherwise healthy young adults from Utah and California. Sin Nombre virus causes a wider spectrum of disease severity than has been previously reported.

Absence of detectable antibody in a patient infected with human immunodeficiency virus.

Infection with human immunodeficiency virus (HIV) is routinely and easily diagnosed with use of enzyme immunoassay (EIA) test kits. We describe an unusual patient who developed AIDS despite testing negative for antibodies to HIV 35 times over a 4-year period. HIV infection was confirmed by the results of p24-antigen assays and polymerase chain reaction amplification of proviral DNA. Sequence analysis of the virus demonstrated that it was closely related to a strain obtained from the patients sexual partner. The explanation for this patients persistently negative EIA results is unclear. However, this case does suggest that physicians who treat patients with AIDS-defining conditions but for whom standard HIV antibody testing is negative should consider the possibility that HIV infection is present and may be identified by additional testing procedures.

Susceptibility of beta-hemolytic streptococci to nine antimicrobial agents among four medical centers in Salt Lake City, Utah, USA

A multicenter study was performed to evaluate the susceptibility of beta-hemolytic streptococci to nine antimicrobial agents. MICs were performed in cation-supplemented Mueller-Hinton broth with 3.5% lysed sheep red blood cells according to NCCLS guidelines. A total of 646 isolates were tested: 300 (46%) group A; 170 (26%) group B; 38 (6%) group C, 35 (5%) group F; 83 (17%) group G; and 20 (3%) nongroupable. Six percent of the total isolates were resistant to one or more of the antibiotics tested. Approximately 7% of 387 strains from the University of Utah Hospital and Clinics were resistant to erythromycin. Four isolates were resistant to clindamycin. Six strains (3%) from Primary Childrens Medical Center (207 tested) were resistant to one or more of the macrolides. Resistance was rare at the LDS Hospital and the Salt Lake Veterans Affairs Hospital. Overall, resistance among beta-hemolytic streptococci in this geographic location does not seem to be a significant problem, except at the tertiary care university hospital.