Evaluation of a BioFire multiplex PCR panel for detection of joint infections using retrospective and prospectively collected specimens.

The BioFire Joint Infection Panel (JI panel) is a newly FDA-approved multiplex PCR assay for detection of common bone and joint pathogens with 39 targets which include select Gram-positive and Gram-negative bacteria, yeast, and antimicrobial resistance genes. We evaluated the performance of the JI panel in detecting joint infections in our patient population. Sixty-three frozen, residual joint fluid specimens were retrospectively tested using the JI panel. An additional 104 residual joint fluid specimens were de-identified and prospectively tested within 1 week of collection. Results from routine bacterial cultures were used as the reference standard, which included inoculation to agar plates and blood culture bottles. For the frozen specimens, the JI panel showed a positive percent agreement (PPA) of 92.8% and a negative percent agreement (NPA) of 97.1%. PPA was 71.4% and NPA was 94.8% for fresh specimens. A total of 12 discrepancies were observed among the 167 specimens tested. The JI panel demonstrated good overall agreement with routine culture for the detection of joint infections and may improve timely diagnosis when used in conjunction with bacterial culture. However, potential false-positive and false-negative results were observed in both retrospective and prospective testing of specimens.IMPORTANCEThe BioFire JI panel is a new commercially available multiplex PCR assay for detecting common pathogens causing bone and joint infections. The test is performed directly on joint fluids with a fast turnaround time of 1 hour. Our study shows that while the JI panel overall shows good agreement with routine culture, discrepancies were observed in 7% of cases and results should be interpreted with appropriate clinical context.

Clinicopathologic Evaluation and Molecular Profiling of Recurrent Stage IA Endometrial Endometrioid Carcinoma: A Case-control Study.

Most low-grade, early-stage endometrial endometrioid carcinomas (EEC) have an excellent prognosis; however, recurrences occur in a small subset with several studies reporting an increase in CTNNB1 exon 3 mutations in this population. Herein we evaluated 10 recurrent low-grade (FIGO 1 or 2), early-stage (FIGO IA) EECs matched to 10 nonrecurrent EECs to further characterize their clinicopathologic features and molecular phenotype. Cases were matched to controls based on size, grade, and depth of invasion. All tumors were evaluated for specific clinicopathologic parameters followed by next-generation sequencing using a 1213 gene panel. Recurrent EECs demonstrated no significant clinicopathologic differences when compared with nonrecurrent EECs, in terms of age, body mass index, pattern of invasion, presence of endometrial atypical hyperplasia/endometrioid intraepithelial neoplasia, associated metaplastic changes, peritumoral lymphocytes, mitoses, and tumor-infiltrating lymphocytes. Both cohorts also showed a similar number of pathogenic mutations, including CTNNB1 exon 3 mutations, as well as tumor mutational burden and microsatellite profiles. Although in this particular study, the lack of correlation between CTNNB1 exon 3 mutation and recurrence might be secondary to a small sample size, it also suggests the presence of other contributing factors. Thus, it helps set the foundation for larger series incorporating whole genome, transcriptome, proteome, and epigenome analyses to answer this clinically important question.

Expect the unexpected: endocarditis caused by Legionella feeleii.

We report a fatal case of Legionella feeleii endocarditis in a post-lung transplant patient. The diagnosis was delayed, as routine microbiological testing of nonrespiratory specimens does not account for extrapulmonary Legionella, and urine antigen testing only reliably detects Legionella pneumophila serogroup 1. This case also illustrates the utility of molecular sequencing for blood culture-negative endocarditis.

Fecal metabolite profiling identifies liver transplant recipients at risk for postoperative infection.

Metabolites produced by the intestinal microbiome modulate mucosal immune defenses and optimize epithelial barrier function. Intestinal dysbiosis, including loss of intestinal microbiome diversity and expansion of antibiotic-resistant pathobionts, is accompanied by changes in fecal metabolite concentrations and increased incidence of systemic infection. Laboratory tests that quantify intestinal dysbiosis, however, have yet to be incorporated into clinical practice. We quantified fecal metabolites in 107 patients undergoing liver transplantation (LT) and correlated these with fecal microbiome compositions, pathobiont expansion, and postoperative infections. Consistent with experimental studies implicating microbiome-derived metabolites with host-mediated antimicrobial defenses, reduced fecal concentrations of short- and branched-chain fatty acids, secondary bile acids, and tryptophan metabolites correlate with compositional microbiome dysbiosis in LT patients and the relative risk of postoperative infection. Our findings demonstrate that fecal metabolite profiling can identify LT patients at increased risk of postoperative infection and may provide guideposts for microbiome-targeted therapies.

Reduced immunomodulatory metabolite concentrations in peri-transplant fecal samples from heart allograft recipients.

Background: Emerging evidence is revealing the impact of the gut microbiome on hematopoietic and solid organ transplantation. Prior studies postulate that this influence is mediated by bioactive metabolites produced by gut-dwelling commensal bacteria. However, gut microbial metabolite production has not previously been measured among heart transplant (HT) recipients. Methods: In order to investigate the potential influence of the gut microbiome and its metabolites on HT, we analyzed the composition and metabolite production of the fecal microbiome among 48 HT recipients at the time of HT. Results: Compared to 20 healthy donors, HT recipients have significantly reduced alpha, i.e. within-sample, microbiota diversity, with significantly lower abundances of key anaerobic commensal bacteria and higher abundances of potentially pathogenic taxa that have been correlated with adverse outcomes in other forms of transplantation. HT recipients have a wide range of microbiota-derived fecal metabolite concentrations, with significantly reduced levels of immune modulatory metabolites such as short chain fatty acids and secondary bile acids compared to healthy donors. These differences were likely due to disease severity and prior antibiotic exposures but were not explained by other demographic or clinical factors. Conclusions: Key potentially immune modulatory gut microbial metabolites are quantifiable and significantly reduced among HT recipients compared to healthy donors. Further study is needed to understand whether this wide range of gut microbial dysbiosis and metabolite alterations impact clinical outcomes and if they can be used as predictive biomarkers or manipulated to improve transplant outcomes.

Bifidobacteria metabolize lactulose to optimize gut metabolites and prevent systemic infection in patients with liver disease.

Progression of chronic liver disease is precipitated by hepatocyte loss, inflammation and fibrosis. This process results in the loss of critical hepatic functions, increasing morbidity and the risk of infection. Medical interventions that treat complications of hepatic failure, including antibiotic administration for systemic infections and lactulose treatment for hepatic encephalopathy, can impact gut microbiome composition and metabolite production. Here, using shotgun metagenomic sequencing and targeted metabolomic analyses on 847 faecal samples from 262 patients with acute or chronic liver disease, we demonstrate that patients hospitalized for liver disease have reduced microbiome diversity and a paucity of bioactive metabolites, including short-chain fatty acids and bile acid derivatives, that impact immune defences and epithelial barrier integrity. We find that patients treated with the orally administered but non-absorbable disaccharide lactulose have increased densities of intestinal bifidobacteria and reduced incidence of systemic infections and mortality. Bifidobacteria metabolize lactulose, produce high concentrations of acetate and acidify the gut lumen in humans and mice, which, in combination, can reduce the growth of antibiotic-resistant bacteria such as vancomycin-resistant Enterococcus faecium in vitro. Our studies suggest that lactulose and bifidobacteria serve as a synbiotic to reduce rates of infection in patients with severe liver disease.

Evaluating specimen pooling for Neisseria gonorrhoeae, Chlamydia trachomatis, and Mycoplasma genitalium screening in asymptomatic men who have sex with men.

BACKGROUND: Screening for Neisseria gonorrhoeae and Chlamydia trachomatis is recommended for individuals at high-risk for sexually transmitted infections (STIs), while the role of Mycoplasma genitalium screening is still unclear. We evaluated whether specimen pooling is an effective alternative for sexually transmitted infection testing during resource shortages. METHODS: This 2-year prospective study enrolled 135 asymptomatic patients from a community outreach site who identified as men who have sex with men. Oropharyngeal, urine, and/or rectal swab specimens were pooled and tested using the Aptima Combo 2 assay for Neisseria gonorrhoeae and Chlamydia trachomatis. The Aptima Mycoplasma genitalium assay was performed on 34 patients whose specimens had sufficient sample volume for additional testing. Results from pooled specimens were compared to results obtained by individual-site testing. RESULTS: The positive percent agreement (PPA) between pooled testing and individual-site testing was 93.8% and negative percent agreement (NPA) was 100% for Neisseria gonorrhoeae. For Chlamydia trachomatis, the PPA was 85.7% and NPA was 99.2%. A PPA of 72.7% and NPA of 95.7% were observed for Mycoplasma genitalium. Discrepancy analysis revealed the majority of false-negative pools were from failure to detect positive rectal samples. CONCLUSIONS: Specimen pooling reduces sensitivity of the Aptima assays for Chlamydia trachomatis, Neisseria gonorrhoeae, and Mycoplasma genitalium and thus should be used with caution.

Evaluation of the EUROIMMUN Anti-SARS-CoV-2 ELISA Assay for detection of IgA and IgG antibodies.

As the Coronavirus 2019 (COVID-19) pandemic evolves, the development of immunoassays to help determine exposure and potentially predict immunity has become a pressing priority. In this report we present the performance of the EUROIMMUN enzyme-linked immunosorbent assay (ELISA) for semi-quantitative detection of IgA and IgG antibodies in serum and plasma samples using recombinant S1 domain of the SARS-CoV-2 spike protein as antigen. Specimens from patients, with and without COVID-19 infection, were tested at the University of Chicago Clinical Microbiology and Immunology Laboratory. Of 86 samples from SARS-CoV-2 PCR-negative patients, including 28 samples positive for common human coronavirus strains, 76 tested negative and 10 tested positive for IgA (88.4% agreement, 95% CI: 79.9-93.6) while 84 tested negative and 2 tested positive for IgG (97.7% agreement, 95% CI: 91.9-99.6). Of 82 samples from SARS-CoV-2 PCR-positive patients, 14 tested negative and 68 tested positive for IgA (82.9% agreement, 95% CI: 73.4-89.5) while 27 tested negative and 55 tested positive for IgG (67.1% agreement, 95% CI: 56.3-76.3). Of samples collected ≥4 days after positive PCR, 38 of 42 (90.5% agreement, 95% CI: 77.9-96.2) were positive for IgA, and 42 of 42 (100% agreement, 95% CI: 91.6-100) were positive for IgG, respectively. The EUROIMMUN Anti-SARS-CoV-2 ELISA Assay demonstrated good sensitivity for detection of IgA and excellent sensitivity for detection of IgG antibodies from samples collected ≥4 days, after COVID-19 diagnosis by PCR. This assay demonstrated good specificity for IgA and excellent specificity for IgG and demonstrated only borderline cross reaction in 2 of the 28 samples from patients with common human coronaviruses infection, types NL63 and OC43.