Jennifer Prentice

Clinical Isolates of Staphylococcus intermedius Masquerading as Methicillin-Resistant Staphylococcus aureus

ABSTRACT Staphylococcus intermedius is a zoonotic organism that can be associated with human disease. We report two separate cases of S. intermedius infection in which a false-positive rapid penicillin binding protein 2a latex test in conjunction with the phenotypic properties of β-hemolysis and coagulase positivity allowed the clinical isolates to masquerade as methicillin-resistant Staphylococcus aureus. 16S rRNA gene sequencing and the absence of mecA revealed the strains to be methicillin-susceptible S. intermedius.

Real-Time Quantitative Reverse Transcription PCR for Monitoring of Blood-Stage Plasmodium falciparum Infections in Malaria Human Challenge Trials

To detect pre-patent parasitemia, we developed a real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) for the asexual 18S ribosomal RNA (rRNAs) of Plasmodium falciparum. Total nucleic acids extracted from whole blood were combined with control RNA and tested by qRT-PCR. The assay quantified > 98.7% of parasite-containing samples to ±0.5 log(10) parasites/mL of the nominal value without false positives. The analytical sensitivity was ≥ 20 parasites/mL. The coefficient of variation was 0.6% and 1.8% within runs and 1.6% and 4.0% between runs for high and low parasitemia specimens, respectively. Using this assay, we determined that A-type 18S rRNAs are stably expressed at 1 × 10(4) copies per ring-stage parasite. When used to monitor experimental P. falciparum infection of human volunteers, the assay detected blood-stage infections 3.7 days earlier on average than thick blood smears. This validated, internally controlled qRT-PCR method also uses a small (50 μL) sample volume requiring minimal pre-analytical handling, making it useful for clinical trials.

Characterization of a Novel Group of Mycobacteria and Proposal of Mycobacterium sherrisii sp. nov.

ABSTRACT We describe here the characterization of five isolates of Mycobacterium simiae-like organisms representing a novel group based on whole-cell fatty acid analysis and genotypic evaluation. Two of the five isolates in this study, W55 and W58, were previously considered to belong to M. simiae serotype 2. Analysis of cellular fatty acids by gas-liquid chromatography indicated a close clustering of this group, which was well differentiated from the other M. simiae-like species. Molecular characterization was performed by nucleic acid sequencing of the small subunit rRNA gene and the gene encoding the 65-kDa heat shock protein and genomic DNA hybridization. Sequence analysis of the entire 16S rRNA gene showed a unique sequence most closely related to those of M. triplex and M. simiae. The hsp65 partial gene sequence was identical for the five isolates, with 97% identity to the M. simiae type strain. However, qualitative whole genomic DNA hybridization analysis confirmed that this group is genetically distinct from M. simiae and M. triplex. Antimicrobial susceptibilities for this group resemble those of M. simiae and M. lentiflavum. We conclude that this group represents a unique Mycobacterium species for which we propose the name Mycobacterium sherrisii sp. nov.

Nocardia veterana, a New Emerging Pathogen

ABSTRACT Nocardia veterana is a newly described species named after the veterans hospital where it was first isolated. This initial type strain was not thought to be clinically significant. We describe three cases of pulmonary disease attributable to N. veterana: two cases in patients presenting with multiple pulmonary nodules in a setting of immunocompromise and one case of exacerbation of chronic pulmonary disease. The isolates were susceptible to ampicillin, imipenem, gentamicin, amikacin, and trimethoprim-sulfamethoxazole and had reduced susceptibilities to ceftriaxone, cefotaxime, minocycline, and ciprofloxacin. The MICs of amoxicillin-clavulanate were higher than that of ampicillin alone, and the bacteria produced a β-lactamase detectable only after induction with clavulanic acid. Phenotypically, the isolates could not be characterized beyond the Nocardia genus level. All three isolates were definitively identified as N. veterana by PCR and sequencing of the 16S rRNA gene. On the basis of their susceptibility and restriction enzyme analysis profiles, our findings indicate that they could potentially be misidentified as N. nova. These cases illustrate the pathogenic potential of this newly described species and emphasize the importance of accurate identification of Nocardia isolates to the species level by integrated use of phenotypic and genotypic methods.

Molecular Diagnosis of Cystoisosporiasis Using Extended-Range PCR Screening

The differential diagnosis of diarrhea in immunocompromised patients encompasses many intestinal parasites including the coccidian Cystoisospora belli. Gastrointestinal infection with C. belli leads to cystoisosporiasis with diarrhea and, depending on host immune status, can cause extraintestinal disease. C. belli is usually diagnosed by examination of stool or intestinal biopsy specimens; however, the organism may be undetected using these test methods. Thus, more sensitive molecular tools for detection of pathogenic parasites are desirable. Herein is described a patient with AIDS who had persistent diarrhea of unknown cause. Microscopic examinations of stool and ileal biopsy specimens were initially unremarkable for any specific pathogen. Screening of DNA extracted from biopsy material using extended-range PCR primers recognizing conserved DNA sequences found in many fungi and parasites revealed infection with C. belli, which was confirmed at repeat histologic analysis. Extended-range PCR screening was used because the differential diagnosis was broad and other tools were not applied, yet this molecular approach led to the appropriate diagnosis and treatment of the condition. Thus, this approach offers a promising test for diagnosis of parasitic diseases that elude diagnosis using conventional methods.

Prosthetic Joint Infection Due to “Helcococcus pyogenica”

ABSTRACT Helcococci have previously been associated with the colonization of ulcers and infections of the skin and soft tissues. We describe a case of prosthetic joint infection due to a previously undescribed organism that is genetically most closely related to Helcococcus.

Molecular analysis of sarcoidosis lymph nodes for microorganisms: a case–control study with clinical correlates

Introduction Sarcoidosis is an incurable, chronic granulomatous disease primarily involving the lungs and lymph nodes of unknown aetiology, treated with non-specific anti-inflammatory/immunosuppressive drugs. Persistently symptomatic patients worsen with a disabling, potentially fatal clinical course. To determine a possible infectious cause, we correlated in a case-control study the clinical information with the presence of bacterial DNA in sarcoidosis mediastinal lymph nodes compared with control lymph nodes resected during cancer surgery. Methods We retrospectively studied formalin-fixed, paraffin-embedded, mediastinal lymph nodes from 30 patients with sarcoidosis and 30 control patients with lung cancer. Nucleic acids were extracted from nodes, evaluated by ribosomal RNA PCR for bacterial 16S ribosomal DNA and the results were sequenced and compared with a bacterial sequence library. Clinical information was correlated. Results 11/30 (36.7%) of lymph nodes from patients with sarcoidosis had detectable bacterial DNA, significantly more than control patient lymph nodes (2/30, 6.7%), p=0.00516. At presentation, 19/30 (63.3%) patients with sarcoidosis were symptomatic including all patients with detectable bacterial DNA. Radiographically, there were 18 stage I and 12 stage II patients. All stage II patients were symptomatic and 75% had PCR-detectable bacteria. After a mean follow-up of 52.8±32.8 months, all patients with PCR-detectable bacteria in this series were persistently symptomatic requiring treatment. Discussion 36.6% of patients with sarcoidosis had detectable bacterial DNA on presentation, all of these patients were quite symptomatic and most were radiographically advanced stage II. These findings suggest that bacterial DNA-positive, symptomatic patients have more aggressive sarcoidosis that persists long term and might benefit from antimicrobial treatment directed against this presumed chronic granulomatous infection.

Systematic Assessment of Culture Review as a Tool to Assess Errors in the Clinical Microbiology Laboratory

CONTEXT Daily supervisory review is a common practice in microbiology laboratories; however, there are no publications describing errors corrected by this practice. OBJECTIVE To determine (1) the correction rates for routinely reviewed positive cultures, (2) the correction rates for negative cultures, and (3) the types of corrections that are found, including the number with potential clinical significance. DESIGN We prospectively assessed errors identified during culture report review for all positive (10-month period) and negative (1-month period) cultures at a single, university-based clinical microbiology laboratory in the United States. Errors were classified using predefined categories, and total and per category error rates were determined. A chi(2) test was used to assess significant differences between error rates. RESULTS A total of 112,108 culture reports were examined; 914 reports required a total of 1043 corrections. Of 101,703 positive culture reports, 786 (0.8%) required 900 corrections, 302 (0.3%) of which were potentially clinically significant. Of 10,405 negative culture reports, 128 (1.2%) required 143 corrections, 5 (0.05%) of which were potentially clinically significant. The rate of potentially clinically significant errors was significantly higher among positive versus negative culture reports (P < .001). Errors from positive culture reports most commonly involved susceptibility (374 [42%]), reporting (275 [31%]), and identification workup (217 [24%]). Most potentially significant errors from positive culture reports involved susceptibility testing (n = 253) and specimens from wound or lower respiratory tract (P < .001). CONCLUSIONS Review of culture reports from positive cultures from nonsterile sites with special attention to antimicrobial susceptibility testing and reporting would be most likely to detect potentially significant errors within the clinical microbiology laboratory.

Fatal Pulmonary Infection Associated with a Novel Organism, “Parastreptomyces abscessus”

ABSTRACT Actinomycetes are increasingly recognized as pathogenic in the immunocompromised host. We isolated an asporogenous, nonmotile, aerobic gram-positive rod from a transplant recipient with a fatal pulmonary infection. The pathology was similar to that associated with Rhodococcus equi, including intrahistiocytic localization. The organism was relatively inert in standard biochemical tests. 16S rRNA gene sequencing indicated a potentially unique organism most closely related to the genus Streptomyces, for which we propose the name “Parastreptomyces abscessus.”