268
permitting growth at higher than normal temperatures. If the A-layer is a prerequisite for virulence, it may be assumed that its presence confers advantages on the
bacterial cell in its role as a pathogen. Indeed, several prime functions for the
A-layer have been proposed. Thus, evidence exists that the extracellular layer protects Aer. salmonicida cells from the action of protease (Kay and Trust 1991 ) and
bacteriophage, by shielding its phage receptors (Ishiguro et al. 1981 ). In addition,
the layer may protect the cell from serum complement, insofar as Munn and Trust
( 1984 ) demonstrated that virulent strains (with A-layer) were resistant to complement bacteriocidal activity in the presence (and indeed absence) of specifi c antibody in rainbow trout serum. Other investigations have revealed that hydrophobicity
is conferred upon the bacterial surface by the A-layer (Trust et al. 1983 ; Van Alstine
et al. 1986 ). These workers reported that the hydrophobic A-layer provided Aer.
salmonicida cells with an affi nity for fatty acid esters of polyethylene glycol and an
enhanced ability to associate with rainbow trout and mouse phagocytic monocytes
(macrophages), in the absence of opsonising antibody. Although Trust et al. ( 1983 )
conceded that the advantages to the pathogen of the increased association with macrophages remained to be determined, they suggested as a tentative explanation, the
possibility that Aer. salmonicida is a facultative intracellular pathogen able to survive within phagocytes. Indeed, Munn and Trust ( 1984 ) demonstrated that A-layer
+
bacteria (i.e. bacteria with A-layer) were able to multiply within the principal
phagocytic organs, e.g. the spleen, following experimental infection. Subsequently,
it has become established that Aer. salmonicida is capable of internalization and
replication in macrophages (Ewart et al. 2008 ), where the pathogen is presumed to
be able to resist reactive radicals (Garduño et al. 1997 ). It has been argued that the
surface layer constitutes the fi rst line of defence for Aer. salmonicida, with an inducible catalase and manganese superoxide dismutase as second defensive systems
against macrophage-mediated killing via reactive oxygen species.
The A-layer has also been implicated in a role concerning adhesion to fi sh tissues. By means of in vitro experiments, Parker and Munn ( 1985 ) examined the ability of avirulent (A-layer
− ) cells to adhere to cells of baby hamster kidney and
rainbow trout gonad in tissue culture. Attachment of A-layer
+ Aer. salmonicida to
both types of cells was greater than for the A-layer
− derivative. As a result, Parker
and Munn ( 1985 ) proposed that since attachment to epithelial cells may be the primary step in the pathogenic process, their observations could account for the association of virulence with the presence of an extra outer membrane layer.
Another function of the A-layer is a possible interference with the antibacterial
peptides, namely magainin, cecropins and defensins (Henry and Secombes 2000 ).
To summarise, the accumulating body of evidence indicts the A-layer as a principal virulence determinant, even though its precise functions and the mechanism of
action obviously require further clarifi cation. However, blithe acceptance of an
absolute relationship between virulence and possession of an A-layer must unfortunately be cautioned against. This is in view of reports by Johnson et al. ( 1985 ) and
Ward et al. ( 1985 ) on the occurrence of virulent, auto-agglutinating strains that have
no detectable A-layer. Conversely, Olivier ( 1990 ) recovered non-virulent A-layer
+
isolates. Thus, the association between presence of the extracellular layer and virulence, but not between auto-agglutination and virulence, appears to be open to ques5 Aeromonadaceae Representative (Aeromonas salmonicida)
permitting growth at higher than normal temperatures. If the A-layer is a prerequisite for virulence, it may be assumed that its presence confers advantages on the
bacterial cell in its role as a pathogen. Indeed, several prime functions for the
A-layer have been proposed. Thus, evidence exists that the extracellular layer protects Aer. salmonicida cells from the action of protease (Kay and Trust 1991 ) and
bacteriophage, by shielding its phage receptors (Ishiguro et al. 1981 ). In addition,
the layer may protect the cell from serum complement, insofar as Munn and Trust
( 1984 ) demonstrated that virulent strains (with A-layer) were resistant to complement bacteriocidal activity in the presence (and indeed absence) of specifi c antibody in rainbow trout serum. Other investigations have revealed that hydrophobicity
is conferred upon the bacterial surface by the A-layer (Trust et al. 1983 ; Van Alstine
et al. 1986 ). These workers reported that the hydrophobic A-layer provided Aer.
salmonicida cells with an affi nity for fatty acid esters of polyethylene glycol and an
enhanced ability to associate with rainbow trout and mouse phagocytic monocytes
(macrophages), in the absence of opsonising antibody. Although Trust et al. ( 1983 )
conceded that the advantages to the pathogen of the increased association with macrophages remained to be determined, they suggested as a tentative explanation, the
possibility that Aer. salmonicida is a facultative intracellular pathogen able to survive within phagocytes. Indeed, Munn and Trust ( 1984 ) demonstrated that A-layer
+
bacteria (i.e. bacteria with A-layer) were able to multiply within the principal
phagocytic organs, e.g. the spleen, following experimental infection. Subsequently,
it has become established that Aer. salmonicida is capable of internalization and
replication in macrophages (Ewart et al. 2008 ), where the pathogen is presumed to
be able to resist reactive radicals (Garduño et al. 1997 ). It has been argued that the
surface layer constitutes the fi rst line of defence for Aer. salmonicida, with an inducible catalase and manganese superoxide dismutase as second defensive systems
against macrophage-mediated killing via reactive oxygen species.
The A-layer has also been implicated in a role concerning adhesion to fi sh tissues. By means of in vitro experiments, Parker and Munn ( 1985 ) examined the ability of avirulent (A-layer
− ) cells to adhere to cells of baby hamster kidney and
rainbow trout gonad in tissue culture. Attachment of A-layer
+ Aer. salmonicida to
both types of cells was greater than for the A-layer
− derivative. As a result, Parker
and Munn ( 1985 ) proposed that since attachment to epithelial cells may be the primary step in the pathogenic process, their observations could account for the association of virulence with the presence of an extra outer membrane layer.
Another function of the A-layer is a possible interference with the antibacterial
peptides, namely magainin, cecropins and defensins (Henry and Secombes 2000 ).
To summarise, the accumulating body of evidence indicts the A-layer as a principal virulence determinant, even though its precise functions and the mechanism of
action obviously require further clarifi cation. However, blithe acceptance of an
absolute relationship between virulence and possession of an A-layer must unfortunately be cautioned against. This is in view of reports by Johnson et al. ( 1985 ) and
Ward et al. ( 1985 ) on the occurrence of virulent, auto-agglutinating strains that have
no detectable A-layer. Conversely, Olivier ( 1990 ) recovered non-virulent A-layer
+
isolates. Thus, the association between presence of the extracellular layer and virulence, but not between auto-agglutination and virulence, appears to be open to ques5 Aeromonadaceae Representative (Aeromonas salmonicida)