An up-date on the differentiation of Brachyspira species from pigs with nox-PCR-based restriction fragment length polymorphism.

Different Brachyspira (B.) species colonize the porcine intestinal tract, some of which are pathogens of significant clinical and economic importance. In 2002 we published a novel molecular method for differentiation of Brachyspira species from pigs based on the amplification of the nox-gene and the generation of species-specific restriction patterns (nox-RFLP) using the enzymes BfmI and DpnII (Rohde et al., 2002). We applied this method for identification in addition to biochemical testing in doubtful cases until 2008. Since 2009 we have used it as the first line method of identification. The current study documents the results of examining 2050 Brachyspira isolates collected from January 2009 to December 2011. In addition to identifying isolates with previously described patterns, four novel restriction fragment length patterns were observed, and isolates with these patterns could be assigned to the species B. intermedia and the B. innocens/murdochii complex on the basis of their phenotypic properties and by nox-sequence analysis. In 2007 a potentially new Brachyspira species, "B. suanatina", was described in Swedish pigs (Råsbäck et al., 2007). From the published nox-gene sequence it could be expected that this Brachypira species should show a new restriction pattern making nox-RFLP a suitable technique for identification of "B. suanatina". In this study the new restriction fragment length pattern could be demonstrated in one of the strains described by Råsbäck et al. (AN4859/03). Nevertheless, no isolates with this new pattern corresponding to "B. suanatina" were identified amongst the 2050 Brachyspira isolates examined from northern Germany.