Rapid identification of mycobacteria and drug-resistant Mycobacterium tuberculosis by use of a single multiplex PCR and DNA sequencing.

Tuberculosis (TB) remains a significant global health problem for which rapid diagnosis is critical to both treatment and control. This report describes a multiplex PCR method, the Mycobacterial IDentification and Drug Resistance Screen (MID-DRS) assay, which allows identification of members of the Mycobacterium tuberculosis complex (MTBC) and the simultaneous amplification of targets for sequencing-based drug resistance screening of rifampin-resistant (rifampin(r)), isoniazid(r), and pyrazinamide(r) TB. Additionally, the same multiplex reaction amplifies a specific 16S rRNA gene target for rapid identification of M. avium complex (MAC) and a region of the heat shock protein 65 gene (hsp65) for further DNA sequencing-based confirmation or identification of other mycobacterial species. Comparison of preliminary results generated with MID-DRS versus culture-based methods for a total of 188 bacterial isolates demonstrated MID-DRS sensitivity and specificity as 100% and 96.8% for MTBC identification; 100% and 98.3% for MAC identification; 97.4% and 98.7% for rifampin(r) TB identification; 60.6% and 100% for isoniazid(r) TB identification; and 75.0% and 98.1% for pyrazinamide(r) TB identification. The performance of the MID-DRS was also tested on acid-fast-bacterium (AFB)-positive clinical specimens, resulting in sensitivity and specificity of 100% and 78.6% for detection of MTBC and 100% and 97.8% for detection of MAC. In conclusion, use of the MID-DRS reduces the time necessary for initial identification and drug resistance screening of TB specimens to as little as 2 days. Since all targets needed for completing the assay are included in a single PCR amplification step, assay costs, preparation time, and risks due to user errors are also reduced.

Three outbreaks of foodborne botulism caused by unsafe home canning of vegetables--Ohio and Washington, 2008 and 2009.

Foodborne botulism is a potentially fatal paralytic illness caused by ingestion of neurotoxin produced by the spore-forming bacterium Clostridium botulinum. Historically, home-canned vegetables have been the most common cause of botulism outbreaks in the United States. During 2008 and 2009, the Centers for Disease Control and Prevention (CDC) and state and local health departments in Ohio and Washington State investigated three outbreaks caused by unsafe home canning of vegetables. We analyzed CDC surveillance data for background on food vehicles that caused botulism outbreaks from 1999 to 2008. For the three outbreaks described, patients and their family members were interviewed and foods were collected. Laboratory testing of clinical and food samples was done at the respective state public health laboratories. From 1999 to 2008, 116 outbreaks of foodborne botulism were reported. Of the 48 outbreaks caused by home-prepared foods from the contiguous United States, 38% (18) were from home-canned vegetables. Three outbreaks of Type A botulism occurred in Ohio and Washington in September 2008, January 2009, and June 2009. Home-canned vegetables (green beans, green bean and carrot blend, and asparagus) served at family meals were confirmed as the source of each outbreak. In each instance, home canners did not follow canning instructions, did not use pressure cookers, ignored signs of food spoilage, and were unaware of the risk of botulism from consuming improperly preserved vegetables. Home-canned vegetables remain a leading cause of foodborne botulism. These outbreaks illustrate critical areas of concern in current home canning and food preparation knowledge and practices. Similar gaps were identified in a 2005 national survey of U.S. adults. Botulism prevention efforts should include targeted educational outreach to home canners.

Fatal pulmonary infection associated with a novel organism, "para-streptomyces abscessus".

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 intra-histiocytic 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 "Para-streptomyces abscessus."

Clinical isolates of Staphylococcus intermedius masquerading as methicillin-resistant Staphylococcus aureus.

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 beta-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.

Prosthetic joint infection due to "Helcococcus pyogenes" [corrected].

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.

Nocardia veterana, a new emerging pathogen.

Nocardia veterana is a newly described species named after the veteran's 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 beta-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.