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 Lawsonia intracellularis is consistently associated with PE and pure cultures of this bacterium have been used to reproduce PE in pigs. In this study L. intracellularis bacteria were purified directly from PE-affected tissue. DNA extracted from purified bacteria was used to construct a partial genomic library which was screened using sera from L. intracellularis-immunized rabbits. Two seroreactive recombinant clones were identified, one of which expressed proteins of 10 and 60 kDa. The sequence of the insert from this clone, pISI-2, revealed ORFs with sequence similarity to the groES/EL operon of Escherichia coli, the 505 ribosomal proteins L21 and L27 of E. coli, a GTP-binding protein of Bacillus subtilis and a possible protoporphyrinogen oxidase, HemK, of E. coli. Primers designed from unique sequences from the pISI-2 insert amplified DNA from infected, but not non-infected, porcine ilea; the amplicon sequence obtained from tissue-cultured L. intracellularis was identical to the corresponding sequence in pISI-2, confirming the origin of the clone. The sequence of L. intracellularis GroEL and other GroEL sequences in the databases were used to construct a partial phylogenetic tree. Analysis of the GroEL sequence relationship suggested that L. intracellularis is not significantly related to other organisms whose GroEL sequences are held in the databases and supports previous data from 16S sequence analyses suggesting that L. intracellularis is a member of a novel group of enteric pathogens.
Dale
C Jane H
CJH
Microbiology and Immunology Department, University of MelbourneRoyal Parade, Parkville, Victoria 3052Australia.|Victorian Institute of Animal Science475 Mickleham Road, Attwood, Victoria 3049Australia.
Moses
Eric K
EK
Victorian Institute of Animal Science475 Mickleham Road, Attwood, Victoria 3049Australia.
Ong
Chin-Chui
CC
Microbiology and Immunology Department, University of MelbourneRoyal Parade, Parkville, Victoria 3052Australia.|Victorian Institute of Animal Science475 Mickleham Road, Attwood, Victoria 3049Australia.
Morrow
Chris J
CJ
Victorian Institute of Animal Science475 Mickleham Road, Attwood, Victoria 3049Australia.
Reed
Michael B
MB
Microbiology and Immunology Department, University of MelbourneRoyal Parade, Parkville, Victoria 3052Australia.|Victorian Institute of Animal Science475 Mickleham Road, Attwood, Victoria 3049Australia.
Hasse
Dete
D
Victorian Institute of Animal Science475 Mickleham Road, Attwood, Victoria 3049Australia.
Strugnell
Richard A
RA
Microbiology and Immunology Department, University of MelbourneRoyal Parade, Parkville, Victoria 3052Australia.
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics*; D018834:Chaperonin 60 / Q000235:genetics; D018833:Chaperonins; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial / Q000235:genetics*; D015698:Genomic Library; D006090:Gram-Negative Bacteria / Q000145:classification / Q000235:genetics*; D006360:Heat-Shock Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009876:Operon / Q000235:genetics*; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013552:Swine
D001426:Bacterial Proteins; D018834:Chaperonin 60; D029968:Escherichia coli Proteins; C028715:GroE protein, E coli; D006360:Heat-Shock Proteins; D018833:Chaperonins