95
coryneform bacteria obtained from fi sh, a fi sh pathogenic Mycobacterium spp., and
Rothia dentocariosa . Yoshimizu et al. ( 1987 ) noted a cross reaction between a
Pseudomonas and Ren. salmoninarum in iFAT. A 60 kDa heat-shock protein (HSP60)
of Chlamydia psittaci migrated with the 57 kDa protein of Ren. salmoninarum, and
may explain the cross-reactivity of polyclonal renibacterium antiserum (Wood et al.
1995 ). Of course, antisera can be made more specifi c by cross-absorbing with these
organisms. Reinforcing a separate article (Toranzo et al. 1993 ) by using western
blots, Bandín et al. ( 1993 ) reported a common antigen, i.e. the 57 kDa protein,
between Cor. aquaticum, Car. piscicola and Ren. salmoninarum. Also, it was noted
that some isolates of Ren. salmoninarum did not produce the 57 kDa protein (Bandín
et al. 1993 ). This is interesting because using the same strains, McIntosh et al. ( 1996 )
could not fi nd the 57 kDa protein in Cor. aquaticum or Car. piscicola. Also in contrast to Bandín and co-workers, Ren. salmoninarum strain K57 was found to produce
the 57 kDa protein. Brown et al. ( 1995 ) produced evidence that bacteria other than
renibacterium could cross react with antiserum to Ren. salmoninarum. Moreover,
these workers used PCR and confi rmed the conclusion of McIntosh et al. ( 1996 ) that
Cor. aquaticum and Car. piscicola lacked the p57 antigen.
Investment may be placed in the development of monoclonal antibodies, which
should be totally specifi c for Renibacterium (Arakawa et al. 1987 ; Wiens and
Kaattari 1991 ). Evelyn ( 1978 ) contradicted the Utopian opinion of serology, by
reporting that culturing was more sensitive than fl uorescent antibody techniques for
the detection of renibacteria in kidney tissue by a factor of 10:1. This theme was
continued in a later study (Evelyn et al. 1981 ) when experiments were undertaken
to determine whether or not there was correlation between culturing and fl uorescent
antibody based diagnoses of the BKD carrier state. Again, culturing was reported as
more sensitive then fl uorescent antibody methods (Evelyn et al. 1981 ). Nevertheless,
from the work of Paterson and colleagues, it could not be explained what was present in the fi sh which gave a positive fl uorescence test but which could not be cultured. Explanations include the presence of dead cells which retain the ability to
fl uoresce, anaerobes which would require specialized isolation procedures, fastidious aerobes, damaged, dormant or inhibited cells of renibacteria, or even inanimate
particles which microscopically could be mistaken for bacteria. Obviously, caution
is needed in interpreting serological diagnoses. Whenever possible, culturing should
be used for confi rmation.
Molecular Techniques Since the early studies addressing the development of
molecular methods for the detection/diagnosis of BKD, emphasis has moved to a
comparison of the effi cacy of different methods (Sandell and Jacobson 2011 ). León
et al. ( 1994a , b ) published details of a PCR assay using a 149 base pair DNA
sequence, which was sensitive enough to detect 22 renibacterial cells even in tissue,
and of suffi cient specifi city to recognise Ren. salmoninarum but not Aer. hydrophila, Aer. salmonicida, Car. piscicola, Fla. columnare, V. anguillarum, V. ordalii or Y.
ruckeri. Then, this group detailed a 2282 base pair DNA fragment that appeared to
be responsible for internalisation of renibacteria, at least into CHSE-tissue culture
cells (Maulén et al. 1996 ). A nested RT-PCR has shown promise for the detection of
Renibacterium salmoninarum
coryneform bacteria obtained from fi sh, a fi sh pathogenic Mycobacterium spp., and
Rothia dentocariosa . Yoshimizu et al. ( 1987 ) noted a cross reaction between a
Pseudomonas and Ren. salmoninarum in iFAT. A 60 kDa heat-shock protein (HSP60)
of Chlamydia psittaci migrated with the 57 kDa protein of Ren. salmoninarum, and
may explain the cross-reactivity of polyclonal renibacterium antiserum (Wood et al.
1995 ). Of course, antisera can be made more specifi c by cross-absorbing with these
organisms. Reinforcing a separate article (Toranzo et al. 1993 ) by using western
blots, Bandín et al. ( 1993 ) reported a common antigen, i.e. the 57 kDa protein,
between Cor. aquaticum, Car. piscicola and Ren. salmoninarum. Also, it was noted
that some isolates of Ren. salmoninarum did not produce the 57 kDa protein (Bandín
et al. 1993 ). This is interesting because using the same strains, McIntosh et al. ( 1996 )
could not fi nd the 57 kDa protein in Cor. aquaticum or Car. piscicola. Also in contrast to Bandín and co-workers, Ren. salmoninarum strain K57 was found to produce
the 57 kDa protein. Brown et al. ( 1995 ) produced evidence that bacteria other than
renibacterium could cross react with antiserum to Ren. salmoninarum. Moreover,
these workers used PCR and confi rmed the conclusion of McIntosh et al. ( 1996 ) that
Cor. aquaticum and Car. piscicola lacked the p57 antigen.
Investment may be placed in the development of monoclonal antibodies, which
should be totally specifi c for Renibacterium (Arakawa et al. 1987 ; Wiens and
Kaattari 1991 ). Evelyn ( 1978 ) contradicted the Utopian opinion of serology, by
reporting that culturing was more sensitive than fl uorescent antibody techniques for
the detection of renibacteria in kidney tissue by a factor of 10:1. This theme was
continued in a later study (Evelyn et al. 1981 ) when experiments were undertaken
to determine whether or not there was correlation between culturing and fl uorescent
antibody based diagnoses of the BKD carrier state. Again, culturing was reported as
more sensitive then fl uorescent antibody methods (Evelyn et al. 1981 ). Nevertheless,
from the work of Paterson and colleagues, it could not be explained what was present in the fi sh which gave a positive fl uorescence test but which could not be cultured. Explanations include the presence of dead cells which retain the ability to
fl uoresce, anaerobes which would require specialized isolation procedures, fastidious aerobes, damaged, dormant or inhibited cells of renibacteria, or even inanimate
particles which microscopically could be mistaken for bacteria. Obviously, caution
is needed in interpreting serological diagnoses. Whenever possible, culturing should
be used for confi rmation.
Molecular Techniques Since the early studies addressing the development of
molecular methods for the detection/diagnosis of BKD, emphasis has moved to a
comparison of the effi cacy of different methods (Sandell and Jacobson 2011 ). León
et al. ( 1994a , b ) published details of a PCR assay using a 149 base pair DNA
sequence, which was sensitive enough to detect 22 renibacterial cells even in tissue,
and of suffi cient specifi city to recognise Ren. salmoninarum but not Aer. hydrophila, Aer. salmonicida, Car. piscicola, Fla. columnare, V. anguillarum, V. ordalii or Y.
ruckeri. Then, this group detailed a 2282 base pair DNA fragment that appeared to
be responsible for internalisation of renibacteria, at least into CHSE-tissue culture
cells (Maulén et al. 1996 ). A nested RT-PCR has shown promise for the detection of
Renibacterium salmoninarum