Restriction Endonuclease DNA Analysis of Leptospira interrogans Serovars Icterohaemorrhagiae and Copenhageni

Abstract

Leptospira interrogans serovar icterohaemorrhagiae strains Ictero No. I and RGA and serovar copenhageni strains M20, Shiromizu and Shibaura were examined by restriction endonuclease DNA analysis. Fifteen endonucleases (AluI, BamHI, BglII, EcoRI, HaeIII, Hhal, HindIII, KpnI, PstI, SacI, SaiI, SmaI, StyI, XbaI and XhoI) were used as the digesting enzymes.Strain Ictero No. I showed endonuclease cleavage patterns which differed from those of the other four strains only when it was digested with enzymes Kpnl and HindIII. When digested with Kpnl, an extra band of about 5 .4 kb was clearly produced, and when digested with HindIII, an extra band of about 25 kb was produced. When the other 13 enzymes were used, no differences were found between the endonuclease cleavage patterns among the five strains. Moreover, strains RGA, M20, Shiromizu and Shibaura could not be distinguished by the restriction endonuclease DNA analysis using all 15 endonucleases. In addition , six newly isolated leptospires from patients with leptospirosis and from Rattus norvegicus were compared with the Ictero No. I and M20 strains, by restriction endonuclease DNA analysis using enzymes KpnI and HindIII. Three leptospires belonging to serovar icterohaemorrhagiae showed the same endonuclease cleavage patterns as the M20 strain. The other three strains, which belong to serovar copenhageni, showed almost the same endonuclease cleavage patterns as the M20 strain; only the Kaijima 702 strain produced an extra band which was not identical to the Ictero No. Ispecific extra band when digested with HindIII. The leptospiral restriction endonuclease DNA analysis has revealed taxonomic structures that are unrecognized by serology alone.

Genome conservation in isolates of Leptospira interrogans

“…2), which are very difficult to distinguish by serological methods (20) and which have identical PFGE Notl restriction profiles. These two serovars have been reported previously to have identical EcoRI or other endonuclease restriction patterns (28,34) and identical DNA-rRNA hybridization patterns (26). They must therefore be extremely closely related genetically.…”

The Number of Large Ribosomal RNA Genes in Leptospira interrogans and Leptospira biflexa

“…Recently, new methods for classifying leptospires have been developed. Restriction endonuclease DNA analysis is one of the accurate and reliable new procedures and classification of leptospires by this procedure has been reported (14)(15)(16)(21)(22)(23). However, whole-cell DNA digests with enzymes generate various restriction fragments, since the genome size of a common eubacterium is over 109 daltons.…”

Diversidad genética de aislamientos peruanos de Leptospira spp. mediante electroforesis en gel de campo pulsado

“…Una de las limitaciones que tiene la técnica es la de no poder diferenciar entre los serovares Icterohaemorrhagiae y Copenhageni; sin embargo, su cercana relación genética y la incapacidad de las técnicas serológicas y moleculares para diferenciarlas ya han sido reportadas previamente (32,33) . Asimismo, la PFGE solo permite identificar especies en caso de que el perfil del aislamiento se encuentre en la base de cepas referenciales.…”