625
Chap. 12 ) or Mueller-Hinton agar. However, a marine-based broth medium with
L-cysteine has been reported to enable the successful culture of Pis. salmonis from
infected fi sh with incubation at 18 °C for 6 days (Yañez et al. 2012 ). A blood and
serum-free medium was described leading to growth of Piscirickettsia salmonis at
17–19 °C (optimally at 25 °C) , in which the cell mass retained pathogenicity
(Henriquez et al. 2013 ).
The problem of purifying the bacteria from tissue culture cells was addressed by
use of 30 % percoll in which bacteriophage like particles were observed by TEM
(Yuksel et al. 2001 ) and resolved by use of iodixanol (= Optiprep) as substrate for
differential centrifugation gradients which together with DNase1 digestion led to
suffi cient pure, i.e. 99 %, bacteria for DNA work (Henríquez et al. 2003 ).
Characteristics of the Pathogen
A single isolate, designated LF-89, was studied in detail by Fryer et al . ( 1992 ).
The 16S rRNA conformed to the gamma subdivision of the Proteobacteria.
Moreover, LF-89 did not show any specifi c relationship to any of 450 bacterial 16S
rRNA sequences held on fi le. Nevertheless, similarities were apparent with
Wolbachia persica (similarity = 86.3 %) and Coxiella burnetii (similarity = 87.5 %)
than to representatives of Ehrlichia, Rickettsia or Rochalimaea. In short, it was
deemed that the salmonid pathogen was suffi ciently novel to warrant description in
a new genus of the family Rickettsiaceae. The organism recovered from white sea
bass was reported to have a 96.3–98.7 % 16S rDNA homology with Pis. salmonis
(Arkush et al. 2005 ), which is low for a confi rmed identity.
Analysis of 16S rRNA revealed that Irish isolates formed two groupings whereas
Canadian, Norwegian and Scottish cultures clustered together (Reid et al. 2004 ).
The possibility of genetic differences between isolates was examined with a view to
explaining reasons for differences in virulence and mortality rates. By electrophoretic analysis of the internal transcribed spacer region of 11 Chilean isolates, two
groupings were recognised (Casanova et al. 2003 ).
Box 11.14: Piscirickettsia salmonis
The pathogen is a pleomorphic, non-motile Gram-negative predominantly
coccoid (and ring forms) organism of variable size (0.5 × 1.5–2.0 μm), occurring intracellularly as individuals, pairs or groups. Electron microscopy
reveals that each organism is bound by two membrane layers; a characteristic
trait of the Rickettsiales, and possibly the tribe Erlichiae.
Piscirickettsiaceae Representative
Chap. 12 ) or Mueller-Hinton agar. However, a marine-based broth medium with
L-cysteine has been reported to enable the successful culture of Pis. salmonis from
infected fi sh with incubation at 18 °C for 6 days (Yañez et al. 2012 ). A blood and
serum-free medium was described leading to growth of Piscirickettsia salmonis at
17–19 °C (optimally at 25 °C) , in which the cell mass retained pathogenicity
(Henriquez et al. 2013 ).
The problem of purifying the bacteria from tissue culture cells was addressed by
use of 30 % percoll in which bacteriophage like particles were observed by TEM
(Yuksel et al. 2001 ) and resolved by use of iodixanol (= Optiprep) as substrate for
differential centrifugation gradients which together with DNase1 digestion led to
suffi cient pure, i.e. 99 %, bacteria for DNA work (Henríquez et al. 2003 ).
Characteristics of the Pathogen
A single isolate, designated LF-89, was studied in detail by Fryer et al . ( 1992 ).
The 16S rRNA conformed to the gamma subdivision of the Proteobacteria.
Moreover, LF-89 did not show any specifi c relationship to any of 450 bacterial 16S
rRNA sequences held on fi le. Nevertheless, similarities were apparent with
Wolbachia persica (similarity = 86.3 %) and Coxiella burnetii (similarity = 87.5 %)
than to representatives of Ehrlichia, Rickettsia or Rochalimaea. In short, it was
deemed that the salmonid pathogen was suffi ciently novel to warrant description in
a new genus of the family Rickettsiaceae. The organism recovered from white sea
bass was reported to have a 96.3–98.7 % 16S rDNA homology with Pis. salmonis
(Arkush et al. 2005 ), which is low for a confi rmed identity.
Analysis of 16S rRNA revealed that Irish isolates formed two groupings whereas
Canadian, Norwegian and Scottish cultures clustered together (Reid et al. 2004 ).
The possibility of genetic differences between isolates was examined with a view to
explaining reasons for differences in virulence and mortality rates. By electrophoretic analysis of the internal transcribed spacer region of 11 Chilean isolates, two
groupings were recognised (Casanova et al. 2003 ).
Box 11.14: Piscirickettsia salmonis
The pathogen is a pleomorphic, non-motile Gram-negative predominantly
coccoid (and ring forms) organism of variable size (0.5 × 1.5–2.0 μm), occurring intracellularly as individuals, pairs or groups. Electron microscopy
reveals that each organism is bound by two membrane layers; a characteristic
trait of the Rickettsiales, and possibly the tribe Erlichiae.
Piscirickettsiaceae Representative