02802nas a2200349   4500000000100000008004100001260001300042653001800055653001900073653001100092653002500103653002800128653002500156653003100181653003000212653000900242653002400251100001400275700001600289700001400305700001400319700001200333700001200345700001300357700001300370700001700383245012900400300001100529490000700540520189100547022001402438       1992                            d  c1992 May10aBase Sequence10aDNA, Bacterial10aHumans10aLeprosy, lepromatous10aMolecular Sequence Data10aMycobacterium leprae10aNucleic Acid Hybridization10apolymerase chain reaction10aSkin10aSpecies Specificity1 aArnoldi J1 aSchlüter C1 aDuchrow M1 aHübner L1 aErnst M1 aTeske A1 aFlad H D1 aGerdes J1 aBöttger E C00aSpecies-specific assessment of Mycobacterium leprae in skin biopsies by in situ hybridization and polymerase chain reaction.  a618-230 v663 a<p>Conventional histopathologic diagnosis of mycobacterial infections are limited to the determination of "acid-fast bacilli". A species-specific diagnosis is thus far impossible. In addition, routine microbiologic assessments of mycobacteria suffer from the major drawback that a species-specific diagnosis is extremely time-consuming and in several cases even impossible. As Mycobacterium leprae cannot be cultured in vitro, we tried to specifically target this obligate intracellular parasite by in situ hybridization and polymerase chain reaction (PCR) techniques. For this purpose we used a 22 mer oligonucleotide probe recognizing a species-specific sequence of the 16S rRNA of Mycobacterium leprae. Using an immunoenzymatic detection method for in situ hybridization we were able to specifically assess Mycobacterium leprae (a) in long-term cultured macrophages in vitro infected with different mycobacteria species and (b) in frozen sections of skin biopsies obtained from patients suffering from lepromatous leprosy. These results could be confirmed and extended by PCR experiments in which we used conserved oligonucleotide primers for 16S rRNA to amplify bacterial DNA isolated from different eubacterial species and from fresh-frozen as well as from formalin-fixed, paraffin-embedded and routinely processed mycobacteria-infected tissues. Upon Southern blot analysis, the Mycobacterium leprae-specific oligonucleotide probe exclusively hybridized with PCR products obtained from Mycobacterium leprae-containing samples (including paraffin sections), but not with PCR products obtained from samples containing other mycobacterial species. As species-specific oligonucleotide probes targeted at rRNA are described for a variety of mycobacterial species, these methods may be generally applied for a rapid species-specific assessment of mycobacteria in histologic material.</p>  a0023-6837