627
calculated as <2 × 10
3 cells (Birkbeck et al. 2004 ). The nature of the antigens to
which animals respond has been addressed by Kuzuk et al . ( 1996 ), who used rickettsias purifi ed from CSE cells by differential and Percoll density gradient centrifugation and rabbit antiserum. The conclusion was that the rabbit antiserum reacted
with four protein and two carbohydrate (core region of the LPS) surface expressed
antigens of 65, 60, 54, 51, 16 and ~11 kDa. However, a complication arises as a
report of a subsequent study by Barnes et al . ( 1998 ), who concluded that the major
antigens were of 56, 30 and 20 kDa. Nevertheless by using rickettsial suspensions,
attempts were made to infect coho salmon via the gills, intestine (by anal intubation) and skin with the data revealing that use of all sites led to infection. However,
intact skin (injury facilitated invasion) and gills were found to be the most effective
entry sites, followed by intestine (Smith et al. 1999 , 2004 ). Isolations have been
made from the brain (of coho salmon), and it is reasoned that this may well be an
important location of the pathogen in the host (Skarmeta et al. 2000 ). Experimental
evidence has supported the possibility of vertical transmission when after the examination of male and female broodstock, the pathogen was detected by immunofl uorescence in milt and the coelomic fl uid in 14/15 of the fi sh (Larenas et al. 2003 ).
Subsequently, the pathogen was detected in the resultant fry, albeit without evidence
of clinical disease. After in vitro infection of ova, the organism was seen by SEM to
be attached to the surface (of the ovum) by apparent membrane extensions; these
were reasoned to allow the later entry into the ovum (Larenas et al. 2003 ).
Evidence from the use of cell cultures points to the pathogen replicating within
the cytoplasmic vesicles of cells although the mechanisms for survival and replication are unclear. However, Isla et al . ( 2014 ) determined that during the infection
process involving the SHK-1 cell line, Pis. salmonis increased signifi cantly the
expression of ClpB and BipA protein, which were recognised as heat shock and
virulence proteins, respectively. These would permit Pis. salmonis to survive the
hostile conditions of the macrophages, thus evading cellular degradation, and
enabling intracellular replication (Isla et al. 2014 ). The pathogen has been recognized to possess several mechanisms for acquiring iron, which points to the ability
of the organism to obtain ferrous and ferric iron from various sources (Pulgar et al.
2015 ). Clathrin has been determined to be required for internalization, and actin
cytoskeleton have a role throughout the infective process (Ramirez et al. 2015 ).
Cytotoxicity by thermostable ECPs has been described (Rojas et al. 2013 ).
Disease Control
Vaccine Development Attempts have been made to develop vaccines. In one
study, formalised cells (10
6.7 TCID 50 /ml) administered by i.p. led to the development
of good protection in a fi eld trial with coho salmon (Smith et al. 1995 ). Heat inactivated (100 °C for 30 min) and formalised whole cell suspensions containing 10
9
cells/ml gave commendable protection with RPS of 71 and 50 %, respectively, when
applied intraperitoneally in adjuvant to Atlantic salmon (Birkbeck et al. 2004 ). An
approach using oral vaccination by feeding every 3 days for 30 days at 6 mg of
Piscirickettsiaceae Representative
calculated as <2 × 10
3 cells (Birkbeck et al. 2004 ). The nature of the antigens to
which animals respond has been addressed by Kuzuk et al . ( 1996 ), who used rickettsias purifi ed from CSE cells by differential and Percoll density gradient centrifugation and rabbit antiserum. The conclusion was that the rabbit antiserum reacted
with four protein and two carbohydrate (core region of the LPS) surface expressed
antigens of 65, 60, 54, 51, 16 and ~11 kDa. However, a complication arises as a
report of a subsequent study by Barnes et al . ( 1998 ), who concluded that the major
antigens were of 56, 30 and 20 kDa. Nevertheless by using rickettsial suspensions,
attempts were made to infect coho salmon via the gills, intestine (by anal intubation) and skin with the data revealing that use of all sites led to infection. However,
intact skin (injury facilitated invasion) and gills were found to be the most effective
entry sites, followed by intestine (Smith et al. 1999 , 2004 ). Isolations have been
made from the brain (of coho salmon), and it is reasoned that this may well be an
important location of the pathogen in the host (Skarmeta et al. 2000 ). Experimental
evidence has supported the possibility of vertical transmission when after the examination of male and female broodstock, the pathogen was detected by immunofl uorescence in milt and the coelomic fl uid in 14/15 of the fi sh (Larenas et al. 2003 ).
Subsequently, the pathogen was detected in the resultant fry, albeit without evidence
of clinical disease. After in vitro infection of ova, the organism was seen by SEM to
be attached to the surface (of the ovum) by apparent membrane extensions; these
were reasoned to allow the later entry into the ovum (Larenas et al. 2003 ).
Evidence from the use of cell cultures points to the pathogen replicating within
the cytoplasmic vesicles of cells although the mechanisms for survival and replication are unclear. However, Isla et al . ( 2014 ) determined that during the infection
process involving the SHK-1 cell line, Pis. salmonis increased signifi cantly the
expression of ClpB and BipA protein, which were recognised as heat shock and
virulence proteins, respectively. These would permit Pis. salmonis to survive the
hostile conditions of the macrophages, thus evading cellular degradation, and
enabling intracellular replication (Isla et al. 2014 ). The pathogen has been recognized to possess several mechanisms for acquiring iron, which points to the ability
of the organism to obtain ferrous and ferric iron from various sources (Pulgar et al.
2015 ). Clathrin has been determined to be required for internalization, and actin
cytoskeleton have a role throughout the infective process (Ramirez et al. 2015 ).
Cytotoxicity by thermostable ECPs has been described (Rojas et al. 2013 ).
Disease Control
Vaccine Development Attempts have been made to develop vaccines. In one
study, formalised cells (10
6.7 TCID 50 /ml) administered by i.p. led to the development
of good protection in a fi eld trial with coho salmon (Smith et al. 1995 ). Heat inactivated (100 °C for 30 min) and formalised whole cell suspensions containing 10
9
cells/ml gave commendable protection with RPS of 71 and 50 %, respectively, when
applied intraperitoneally in adjuvant to Atlantic salmon (Birkbeck et al. 2004 ). An
approach using oral vaccination by feeding every 3 days for 30 days at 6 mg of
Piscirickettsiaceae Representative