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Mycoplasma pneumoniae is an important respiratory pathogen, accounting for up to 25% of community-acquired pneumonia, and is a common cause of hospitalized pneumonia in otherwise healthy adults and children. Mycoplasma pneumoniae isolates can be classified into two main genomic groups (type 1 and type 2) based on sequence variation within the gene encoding the major adhesion molecule P1. Although numerous publications have described real-time PCR assays for the detection of M. pneumoniae, none has been able to discriminate the two genomic types. Here, a real-time PCR assay that can distinguish each type of M. pneumoniae utilizing high-resolution melt-curve analysis is reported. Using this method, 102 isolates obtained from patients from 1965 to the present, including those from recent outbreaks, were typed along with reference strains M129 (type 1) and FH (type 2). The results show that 55 isolates (54%) can be classified as type 1 and 47 isolates (46%) as type 2, and 100% correlation was demonstrated when compared with a standard PCR-restriction fragment length polymorphism typing procedure. Typing of isolates obtained from recent outbreaks in the USA has revealed the presence of both types. This assay provides a rapid, reliable and convenient method for typing M. pneumoniae isolates and may be useful for surveillance purposes and epidemiological investigations, and may provide insight into the biology of M. pneumoniae distribution within populations.

The interaction between hosts and the viruses that infect them is a dynamic one, and a growing literature documents the fact that many viruses have developed mechanisms designed to avoid elimination by the host immune system. One of the immune strategies used by the host and targeted by virus proteins is apoptosis triggered by the cytokine tumor necrosis factor (TNF). Mouse fibroblast LM cells are spontaneously sensitive to TNF. When the wild-type E6 protein from the human papillomavirus type 16 (HPV 16) was expressed in LM cells, the cells became resistant to TNF. This resistance was examined by several means, including cell morphology, the dose- and time-independent response to TNF in a cell death ELISA, trypan blue exclusion, and cell proliferation. The level of p53 did not rise in TNF-treated cells prior to apoptosis, suggesting a p53-independent mechanism. Significant, though not complete, resistance to TNF was also observed following transfection of a plasmid expressing a mutant E6 protein, which is unable to mediate rapid degradation of the p53 tumor suppressor. These results indicate that the HPV 16 E6 protein can protect LM cells from TNF-triggered apoptosis and likely does so by a mechanism other than mediation of p53 degradation.