IDENTIFICATION AND SEQUENCING OF THE GROE OPERON AND FLANKING GENES OF LAWSONIA-INTRACELLULARIS - USE IN PHYLOGENY

Proliferative enteropathy (PE) is a complex of diseases of commercial importance to the pig industry. The obligate intracellular bacterium L awsonia intracellularis is consistently associated with PE and pure cu ltures of this bacterium have been used to reproduce PE in pigs, In th is study L, intracellularis bacteria were purified directly from PE-af fected tissue. DNA extracted from purified bacteria was used to constr uct a partial genomic library which was screened using sera from L. in tracellularis-immunized rabbits. Two seroreactive recombinant clones w ere identified, one of which expressed proteins of 10 and 60 kDa, The sequence of the insert from this clone, pISI-2, revealed ORFs with seq uence similarity to the groES/EL operon of Escherichia coli, the 50S r ibosomal proteins L21 and L27 of E, coli, a GTP-binding protein of Bac illus subtilis and a possible protoporphyrinogen oxidase, HemK, of E, coli, Primers designed from unique sequences from the pISI-2 insert am plified DNA from infected, but not non-infected, porcine ilea; the amp licon sequence obtained from tissue-cultured L, intracellularis was id entical to the corresponding sequence in pISI-2, confirming the origin of the clone. The sequence of L, intracellularis GroEL and other GroE L sequences in the databases were used to construct a partial phylogen etic tree. Analysis of the GroEL sequence relationship suggested that L, intracellularis is not significantly related to other organisms who se GroEL sequences are held in the databases and supports previous dat a from 16S sequence analyses suggesting that L, intracellularis is a m ember of a novel group of enteric pathogens.