bSpontaneous bacterial peritonitis (SBP) can be a severe complication occurring in patients with cirrhosis and ascites, with associated mortality often as high as 40%. Traditional diagnostics for SBP rely on culture techniques for proper diagnosis, although recent reports suggest that the presence of bacterial DNA in peritoneal fluid in patients with cirrhosis and ascites is an indicator of SBP. A previously published broad-range PCR (16S PCR) coupled with high-resolution melt analysis (HRMA) was compared with standard culture techniques for diagnosis of SBP in 106 peritoneal fluid samples from patients with suspected SBP. The sensitivity and specificity for 16S PCR for detecting eubacterial DNA compared with those of standard culture techniques were 100% (17/17) and 91.5% (85/89), respectively. Overall, HRMA concordance with species identification was 70.6% (12/17), although the 5 samples that were discordant at the species level were SBP resulting from a polymicrobial infection, and specieslevel identification for polymicrobial infections is outside the capability of HRMA. Both the broad-range 16S PCR and HRMA analysis provide useful diagnostic adjunctive assays for clinicians in detecting and identifying pathogens responsible for SBP. Spontaneous bacterial peritonitis (SBP) is a common and potentially fatal bacterial infection in patients with cirrhosis and ascites, occurring in 10 to 30% of patients, with in-hospital mortality rates ranging from 20 to 30% (1, 2, 6-8, 12). It is secondary to impaired humoral and cellular immune responses that result in indirect intestinal bacterial translocation into the ascitic fluid (1, 2, 6-8, 16, 20). SBP is also associated with a poor long-term prognosis for patients, as mortality rates can reach 50 to 70% at 1 year (2). Given that timely and appropriate antibiotic treatment can improve the clinical outcome, rapid and accurate diagnostic methods for early detection of eubacterial infection responsible for SBP and identification of the causative organisms involved could be particularly useful in acute care settings. Recent attention drawn to the changing microbial and resistance patterns attributed to the increasing use of antibiotic prophylaxis and invasive procedures in such patients further underscores the importance of identifying the causative pathogen to ensure adequate antibiotic coverage (13).Current laboratory diagnosis of SBP is defined as Ն250 polymorphonuclear (PMN) cells/ml and a positive culture from ascitic fluid from the patient (1-13, 15-17, 19-22, 29). Unfortunately, the prolonged turnaround time (1 to 2 days) of culture limits its utility for directing antibiotic selection in acute care settings. Ascitic fluid culture has also been reported to be negative in approximately 20% of patients with clinical manifestations suggestive of SBP and an ascitic PMN count of Ͼ250, so-called culture-negative neutrocytic ascites. On the other hand, a low ascitic PMN count (Ͻ250) with positive culture can also occur in another SBP variant called bacterascites, or monom...

No abstract

) and 100% specificity (95% CI, 93.1 to 100.0%); for bacterial pathogens, 81.8% sensitivity (95% CI, 74.3 to 87.6%) and 73.6% specificity (95% CI, 64.2 to 81.4%); for viral pathogens, 93.3% sensitivity (95% CI, 66.0 to 99.7%) and 97.3% specificity (95% CI, 89.7 to 99.5%); for fungal pathogens, 42.6% sensitivity (95% CI, 29.5 to 56.7%) and 97.8% specificity (95% CI, 91.8 to 99.6%). Our data suggest that RT-PCR-ESI-MS is a useful adjunct to standard culture protocols for rapid detection of both BT and common respiratory pathogens; further study is required for assay validation, especially for fungal detection, and potential implementation. Biothreat (BT) agents are among the most feared of mass terrorism weapons (1-3). In 1970, the World Health Organization estimated that the destruction caused by a theoretical attack with 50 kg of aerosolized Bacillus anthracis, Yersinia pestis, or Francisella tularensis could incur between 150,000 and 250,000 incapacitating casualties and between 19,000 and 100,000 fatalities (4). A more recent analysis by the US Congressional Office of Technology Assessment predicted that 100 kg of Bacillus anthracis has the potential to cause up to 3 million deaths, a mortality rate that would match the predicted lethality of a hydrogen bomb (5).Early recognition of a BT attack is critical to prevent subsequent waves of infected patients but, because initial symptoms are nonspecific and resemble common respiratory infections, rapid identification of the specific pathogen is challenging (6-8). Current algorithms for the detection and identification of BT agents can be time-consuming, as they rely on culture-based methods followed by referral to specialized state and/or national reference laboratories for confirmatory testing (9). However, advances in molecular diagnostics in the past decade have led to the introduction of significantly more-rapid assays, principally using sensitive and specific nucleic acid amplification tests (NAATs), which provide pathogen identification and genotyping capabilities previously not possible with traditional culture-based methods (10). Although a variety of NAATs have been designed and validated for BT detection, the majority of currently available assays are optimized to detect a narrow range of targets (8,11). This technical limitation necessitates a high index of clinical suspicion for a particular agent, limiting practical utility in real-world practice and hindering widespread integration into routine patient care settings.New broad-range PCR assays present a potential solution to this challenge, as they exploit highly conserved bacterial, fungal, and viral genes such as heat shock proteins and RNA polymerases for rapid identification of a large number of target pathogens (both common and rare). Using primers that target conserved genomic motifs flanking variable regions, broad-range assays can generate a diverse array of species-specific amplicons while employing relatively few PCRs. These species-specific amplicons can subsequently be differentiated with ...