The critical role of the microbiology laboratory in infectious disease diagnosis calls for a close, positive working relationship between the physician and the microbiologists who provide enormous value to the health care team. This document, developed by both laboratory and clinical experts, provides information on which tests are valuable and in which contexts, and on tests that add little or no value for diagnostic decisions. Sections are divided into anatomic systems, including Bloodstream Infections and Infections of the Cardiovascular System, Central Nervous System Infections, Ocular Infections, Soft Tissue Infections of the Head and Neck, Upper Respiratory Infections, Lower Respiratory Tract infections, Infections of the Gastrointestinal Tract, Intraabdominal Infections, Bone and Joint Infections, Urinary Tract Infections, Genital Infections, and Skin and Soft Tissue Infections; or into etiologic agent groups, including Tickborne Infections, Viral Syndromes, and Blood and Tissue Parasite Infections. Each section contains introductory concepts, a summary of key points, and detailed tables that list suspected agents; the most reliable tests to order; the samples (and volumes) to collect in order of preference; specimen transport devices, procedures, times, and temperatures; and detailed notes on specific issues regarding the test methods, such as when tests are likely to require a specialized laboratory or have prolonged turnaround times. There is redundancy among the tables and sections, as many agents and assay choices overlap. The document is intended to serve as a reference to guide physicians in choosing tests that will aid them to diagnose infectious diseases in their patients.

Mycobacteria are the causative organisms for diseases such as tuberculosis (TB), leprosy, Buruli ulcer, and pulmonary nontuberculous mycobacterial disease, to name the most important ones. In 2015, globally, almost 10 million people developed TB, and almost half a million patients suffered from its multidrug-resistant form. In 2016, a total of 9,287 new TB cases were reported in the United States. In 2015, there were 174,608 new case of leprosy worldwide. India, Brazil, and Indonesia reported the most leprosy cases. In 2015, the World Health Organization reported 2,037 new cases of Buruli ulcer, with most cases being reported in Africa. Pulmonary nontuberculous mycobacterial disease is an emerging public health challenge. The U.S. National Institutes of Health reported an increase from 20 to 47 cases/100,000 persons (or 8.2% per year) of pulmonary nontuberculous mycobacterial disease among adults aged 65 years or older throughout the United States, with 181,037 national annual cases estimated in 2014. This review describes contemporary methods for the laboratory diagnosis of mycobacterial diseases. Furthermore, the review considers the ever-changing health care delivery system and stresses the laboratory's need to adjust and embrace molecular technologies to provide shorter turnaround times and a higher quality of care for the patients who we serve.

Few studies have assessed the time to blood culture positivity as a predictor of clinical outcome in bloodstream infections (BSIs). The purpose of this study was to evaluate the time to positivity (TTP) of blood cultures in patients with Staphylococcus aureus BSIs and to assess its impact on clinical outcome. We performed a historical cohort study with 91 adult patients with S. aureus BSIs. TTP was defined as the time between the start of incubation and the time that the automated alert signal indicating growth in the culture bottle sounded. Patients with BSIs and TTPs of culture of <12 h (n ‫؍‬ 44) and >12 h (n ‫؍‬ 47) were compared. Septic shock occurred in 13.6% of patients with TTPs of <12 h and in 8.5% of patients with TTP of >12 h (P ‫؍‬ 0.51). A central venous catheter source was more common with a BSI TTP of <12 h (P ‫؍‬ 0.010). Univariate analysis revealed that a Charlson score of >3, the failure of at least one organ (respiratory, cardiovascular, renal, hematologic, or hepatic), infection with methicillin-resistant S. aureus, and TTPs of <12 h were associated with death. Age, gender, an APACHE II score of >20 at BSI onset, inadequate empirical antibiotic therapy, hospital-acquired bacteremia, and endocarditis were not associated with mortality. Multivariate analysis revealed that independent predictors of hospital mortality were a Charlson score of >3 Staphylococcus aureus is a highly common cause of BSIs (27). Moreover, mortality increases when inadequate empirical antibiotic therapy is given to patients with BSIs due to S. aureus (12). The rapid detection of a BSI has an impact on the length of hospitalization (2) and the mortality of bacteremic patients (4).Physicians often make clinical predictions about individual patients (13). However, few studies have formally assessed the bacterial load (9), as indirectly measured as the time to positivity (TTP) of blood cultures, as a predictor of clinical outcome (14, 23).The purpose of this study was to evaluate the association between the time to positivity of blood cultures in patients with S. aureus BSIs and the clinical outcome. MATERIALS AND METHODSSetting. The Virginia Commonwealth University Medical Center (VCUMC) is an 820-bed tertiary-care facility in Richmond, Virginia. The hospital houses nine intensive care units (ICUs), including pediatric ICUs and a burn unit. Approximately 30,000 patients are admitted annually. Study design.Patients with BSIs at VCUMC from 15 December 2003 through 31 December 2004 were identified retrospectively by use of the electronic medical microbiology record. For each case, the time to blood culture positivity was retrieved from the hospital's automated blood culture instrument. The medical microbiology record identified the patient by medical record number so that a retrospective chart review could be conducted. Patients were considered to have had a BSI due to S. aureus if one or more blood cultures were positive for this organism. Each patient was included only once, at the time of the first BSI. Patients less than 1...

The emergence of epidemic multiple-drug-resistant (MDR) strains of Mycobacterium tuberculosis in conjunction with an increase in the number of reported cases of tuberculosis (TB) represents a major public health problem. In light of a recent outbreak of MDR M. tuberculosis at our center, we began the development of a polymerase chain reaction (PCR) assay for the rapid diagnosis of pulmonary TB using two sets of primers, one based on the IS6110 repeated sequence of M. tuberculosis and the other based on the protein antigen b (PAB). Reaction conditions were first optimized as to the appropriate extraction protocol and the concentrations of primer pairs, nucleotides, and MgCl2. Following a preliminary evaluation of the assay with clinical specimens, extraction and amplification procedures were further modified. PAB and IS6110 primers detected between 2 and 23 and 0.023 and 0.23 CFU of M. tuberculosis, respectively, in pooled, M. tuberculosis-negative sputa by our optimized PCR assay. After routine processing for mycobacteria, 734 specimens were subsequently amplified. DNA for amplification was obtained by boiling and beating the sediments with Tween 20. For each reaction, DNA (10 ,ul) was added to an amplification mixture containing 12 pmol of IS6110 primers, 20 pmol of PAB primers, 2 mM MgCl2, 200 JLM nucleotides, and 2.5 U of Taq polymerase and the mixture was then amplified for 40 cycles. The sensitivity and specificity of our PCR assay were 87.2 and 97.7%, respectively. We were unable to interpret the results for seven specimens (1%). In our experience, PCR proved to be a useful rapid diagnostic test for TB in a clinical setting and a valuable epidemiological tool for determining exposure groups in the hospital setting. Our findings also underscore the need for the systematic optimization of PCR assay conditions.

The ability of Staphylococcus epidermidis to transfer antimicrobial resistance to Staphylococcus aureus was tested by mixed culture on filter membranes. Two of six clinical isolates examined were able to transfer resistance to S. aureus strains 879R4RF, RN45ORF, and UM1385RF. Subsequent S.aureus transconjugants resulting from matings with S. epidermidis donors were able to serve as donors to other S. aureus strains at similar frequencies. Cell-free and mitomycin C-induced filtrates of donors and transconjugants showed no plaque-forming ability. Addition of DNase I, citrate, EDTA, calcium chloride, and human sera to mating mixes and agar showed no effect on transfer. Nonviable donor cells were unable to transfer resistance and transfer did not occur at 4°C. Cell-to-cell contact was required since transfer did not occur in broth or when filters of donor and recipient, respectively, were placed back-to-back so cells were not in direct contact. Analysis of DNA from S. epidermidis isolate UM899, its subsequent S. aureus transconjugants, and cured derivatives demonstrated that all resistance markers which transferred resided on plasmids. Mating experiments suggested a central role for the gentamicin plasmid pAM899-1 in the transfer process. It is concluded that our results are consistent with a conjugative transfer of resistance from S. epidermidis to S. aureus analogous to plasmid transfer demonstrated in streptococcal species for plasmids such as pAM,1. This represents a novel mechanism for gene exchange among staphylococci.