Detection and identification of Variola virus in fixed human tissue after prolonged archival storage.

Smallpox disease has been eradicated from the human population since 1979, but is again a concern because of its potential use as an agent of bioterrorism or biowarfare. World Health Organization-sanctioned repositories of infectious Variola virus are known to occur in both Russia and the United States, but many believe other undeclared and unregulated sources of the virus could exist. Thus, validation of improved methods for definitive identification of smallpox virus in diagnostic specimens is urgently needed. In this paper, we describe the discovery of suspected Variola infected human tissue, fixed and preserved for decades in largely unknown solutions, and the use of routine histology, electron microscopy, and ultimately DNA extraction and fluorogenic 5' nuclease (TaqMan) assays for its identification and confirmation.

Identification of virulence genes in uropathogenic Escherichia coli by multiplex polymerase chain reaction and their association with infectivity in mice.

OBJECTIVES: To use multiplex polymerase chain reaction (PCR) to screen a large number of Escherichia coli clinical isolates for virulence factor genes and to evaluate the importance of several known factors in the etiology of urinary tract infection. METHODS: Eighty-six E. coli isolates from urine or vaginal or rectal swabs of patients with recurrent urinary tract infection were screened for P fimbria (pap), hemolysin (hly), aerobactin (aer), cytotoxic necrotizing factor 1 (cnf1), S fimbria (sfa), and afimbrial adhesion I (afaI) genes by multiplex PCR. The phenotype of the strains was determined for type 1 fimbriae and O antigen serotype. The infectivity of 11 strains with different combinations of virulence factors was tested using a mouse model of unobstructed urinary tract infection. RESULTS: Type 1 fimbriae were present in 81 of the 86 strains and was the only virulence factor in approximately one third of the isolates. Genes for hly, aer, cnf1, sfa, or pap were present in approximately one fourth of the strains; afaI was present less frequently. A positive type 1 fimbriae phenotype was common to all strains that induced a bladder infection in mice. CONCLUSIONS: Multiplex PCR methods can be effectively applied to studies that require genetic screening of numerous E. coli uropathogens. Where phenotypic information was available, it was consistent with genotypes identified by PCR. Infectivity studies showed that the presence of the type 1 fimbriae gene in an E. coli isolate was required to establish a bladder infection. Other genes that were not identified in this study may also be required in mice and humans.