You are on page 1of 28

427

CAB INTERNATIONAL 1999. Fish Diseases and Disorders, Volume 3:


Viral, Bacterial and Fungal Infections (eds P.T.K. Woo and D.W. Bruno)
11
Motile Aeromonads
(Aeromonas hydrophila)
T. Aoki
Laboratory of Genetics and Biochemistry, Department of Aquatic Biosciences,
Tokyo University of Fisheries, Konan 4-5-7, Minato-ku, Tokyo 108, Japan.
INTRODUCTION
Motile aeromonads of the Aeromonas hydrophila complex cause a haemorrhagic
septicaemia in fish (Bullock et al., 1971; Egusa, 1978; Schperclaus et al.,
1992). This bacterium has been observed in numerous species of freshwater fish
and occasionally in marine fish and in amphibians, reptiles, cattle and humans
throughout the world (Bullock et al., 1971; Khardori and Fainstein, 1988).
However, the most significant diseases occur in cultured freshwater fish. The
bacterium is distributed widely in fresh water and bottom sediments containing
organic material, as well as in the intestinal tract of fish (Aoki, 1974; Egusa,
1978; Hazen et al., 1978; Seidler et al., 1980; Kaper et al., 1981; Van der Kooij
and Hijnen, 1988; Sugita et al., 1994, Dumontet et al., 1996).
Infectious abdominal dropsy in common carp has been attributed to the
A. hydrophila group (Aeromonas punctata) and was first described by
Schperclaus (1930), who reported on this condition in cultured, wild and
stocked carp in central and eastern Europe. The causative agent has since been
shown to be rhabdovirus carpio (spring viraemia of carp) (Fijan, 1972; Wolf,
1988). During the 1960s, outbreaks of red fin disease, caused by A. hydrophila,
occurred frequently in cultured eels in Japan (Hoshina, 1962; Egusa, 1978)
(Fig. 11.1), with concurrent infections by Saprolegnia parasitica (Egusa, 1978).
Currently, only sporadic outbreaks of A. hydrophila occur in cultured eels.
Aeromonas hydrophila is typically recognized as an opportunistic pathogen or
secondary invader (Austin and Austin, 1987).
Conversely, there have been reports of A. hydrophila acting as a primary
pathogen in fish. Isolates differ greatly in their pathogenicity with some strains
being highly virulent and others non-virulent. Eddy (1960) and Kou (1972a)
reported that non-virulent or weakly pathogenic strains did not produce gas and
acetone from glucose. Wakabayashi et al. (1981) recognized common and
identical biochemical characteristics in virulent strains, in particular the
production of elastase and enzymes involved in lysis of Staphylococcus. These
428
T. Aoki
characteristics were identical to the A. hydrophila biovar. hydrophila, following
classification by Popoff and Vron (1976). Economically, the disease attributed
to these bacteria is of greatest importance in cultured freshwater fish.
Recent advances in biochemistry, molecular biology and virulence factors
associated with A. hydrophila have led to new understanding of this bacterial
group. These are reviewed in this chapter.
THE DISEASE AND AGENT
Species of fish affected and geographical distribution of
Aeromonas hydrophila
Most cultured and wild freshwater fish are susceptible to infection by
A. hydrophila, but particularly cold-water fish, such as brown trout (Salmo
trutta), rainbow trout (Oncorhynchus mykiss), chinook salmon (Oncorhynchus
tshawytscha), ayu (Plecoglossus altivelis), carp (Cyprinus carpio), channel
catfish (Ictalurus punctatus), clariid catfish (Clarias batrachus) (Fig. 11.2),
Japanese eel (Anguilla japonica), American eel (Anguilla rostrata), gizzard
shad (Dorosoma cepedianum), goldfish (Carassius auratus), golden shiner
(Notemigonus crysoleucas), snakehead fish (Ophicephalus striatus) and tilapia
(Tilapia nilotica) (Bullock et al., 1971; Egusa, 1978; Saitanu, 1986).
Fig. 11.1. Eel (Anguilla japonica) with red-fin disease (haemorrhagic septicaemia) caused by
Aeromonas hydrophila (courtesy of Dr Teruo Miyazaki).
429 Motile Aeromonads
The disease
Diseased fish usually display cutaneous haemorrhage of the fins and trunk, and
the condition is often refered to as red fin disease (Hoshina, 1962) (Fig. 11.1).
The bacteria multiply inside the intestine, causing a haemorrhagic mucous
desquamative catarrh. Toxic metabolites of A. hydrophila are absorbed from the
intestine and induce a toxaemia. Capillary haemorrhage occurs in the dermis of
fins and trunk and in the submucosa of the stomach. Hepatic cells and epithelia
of renal tubules show degeneration. Glomeruli are destroyed and the tissue
becomes haemorrhagic, with exudates of serum and fibrin (Miyazaki and Jo,
1985; Miyazaki and Kaige, 1985) (Figs 11.311.5).
European carp infected with A. hydrophila show severe tail and fin rot and
visible haemorrhage and ulceration of the body surface. Widespread
proliferation of bacteria is usually observed in the intestine. In some reports
(Fijan, 1972; Wolf, 1988), the histopathological phenomena associated with the
rhabdovirus infection haemorrhagic septicaemia of carp have been erroneously
attributed to motile aeromonads (Bullock et al., 1971). Aeromonas hydrophila is
widely distributed in the intestinal tract of cultured fish and the water and
sediments of freshwater ponds which are rich in organic materials. Virulent
strains of A. hydrophila in these environments are possible sources of infection.
Outbreaks of disease are usually associated with a change in environmental
conditions. Stressors, including overcrowding, high temperature, a sudden
change of temperature, rough handling, transfer of fish, low dissolved oxygen,
Fig. 11.2. Clariid catfish (Clarias batrachus) with ulcerative form of haemorrhagic septicaemia
caused by Aeromonas hydrophila (courtesy of Dr Kriengsag Saitanu).
430
T. Aoki
Fig. 11.3. Aeromonas hydrophila infection in J apanese eel. Intestine shows bacterial
multiplication in the contents and mucousdesquamative catarrh (haematoxylin and eosin stain)
(courtesy of Dr Teruo Miyazaki).
Fig. 11.4. Aeromonas hydrophila infection in J apanese eel. Skin shows capillary haemorrhage
in the dermal loose connective tissue and the thinned epithelium (haematoxylin and eosin stain)
(courtesy of Dr Teruo Miyazaki).
431 Motile Aeromonads
poor nutritional status and fungal or parasitic infection, contribute to physiological
changes and heighten susceptibility to infection. This applies to fish of all ages.
Causative bacterium
There are three mesophilic motile aeromonad species in Bergeys Manual of
Systematic Bacteriology: A. hydrophila, Aeromonas caviae and Aeromonas
sobria (Popoff et al., 1981; Popoff, 1984). Currently, the following additional
Aeromonas species have been recognized: A. allosaccharophila (Martinez-
Murcia et al., 1992a; Esteve et al., 1995a), A. encheleia (Esteve et al., 1995b),
A. eteropelogenes (Collins et al., 1993), A. eurenophila (Martinez-Murcia et al.,
1992b), A. ichthiosmia (Collins et al., 1993), A. jandaei (Carnahan et al., 1991a),
A. media (Allen et al., 1983), A. schuberti (Hickman-Brenner et al., 1988),
A. trota (Carnahan et al., 1991b) and A. veronii (Hickman-Brenner et al., 1987).
The similarity of the above-mentioned ten species of motile aeromonads was
determined as a dendogram, using numerical taxonomy (Carnahan and Joseph,
1993). Millership (1996) reviewed characteristics for the differentiation of the
common phenotypes of these species in aeromonads. The genus Aeromonas has
been classified into the family Vibrionaceae; however, Colwell et al. (1986)
proposed that these aeromonads constituted a separate family, the Aero-
monadaceae.
Fig. 11.5. Aeromonas hydrophila infection in J apanese eel. Kidney shows destroyed glomeruli
accompanying exudation of serum and fibrin. Epithelia of renal tubules show necrosis and
atrophy (haematoxylin and eosin stain) (courtesy of Dr Teruo Miyazaki).
432
T. Aoki
Kou (1972a, 1973) and Wakabayashi et al. (1981) recognized that almost all
pathogenic strains of motile aeromonads relevant to aquaculture were
encompassed within A. hydrophila biovar. hydrophila, proposed by Popoff and
Vron (1976).
Aeromonas hydrophila is a Gram-negative rod-shaped bacterium and is
motile, due to a monotrichous polar flagellum. The bacterium measures 0.31.0
m in diameter and 1.03.5 m in length. It has no spore stage or capsule. The
optimum growth temperature is 28C, but growth can occur at 37C. Colonies
on nutrient agar are white to pale pink, round and convex, with entire margins.
The biochemical characteristics are shown in Table 11.1. Cytochrome oxidase
Table 11.1. Biological characteristics of Aeromonas hydrophila.
Characteristics Response
Indole production in 1% peptone water +
Aesculin hydrolysis +
Growth in potassium cyanide (KCN) broth +
L-Histidine and L-arginine utilization +
L-Arabinose utilization +
Acetoin from glucose (Voges-Proskauer (VP) reaction) +
H
2
S from cysteine +
Oxidase +
Cytochrome oxidase +
Catalase +
Methyl red (MR) experiment d
Acetylmethylcarbinol production +
2,3-Butanediol production +
2,3-Butanediol dehydrogenase +
-Galactosidase production +
Phosphatase +
Nitrate reduction +
Urease
Malonate
Gelatin liquefaction +
Casein digestion +
Loeffler serum digestion +
Starch hydrolysis +
Lipase +
Lecithinase +
Glucuronate utilization +
Ornithine decarboxylase
DNAse +
RNAse +
Haemolysis +
Carbohydrate decomposition
Adonitol
Aesculin d
Arabinose d
Cellobiose d
Dextrin +
Dulcitol
Fructose +
Galactose +
433 Motile Aeromonads
and catalase reactions are positive. It is a facultative anaerobe, fermenting
carbohydrates to acid, or acid and gas. Aeromonas hydrophila is resistant to the
vibriostatic agent O/129 (phosphate: 2,4-diamino-6,7-diisopropylpteridine
phosphate) 150 g, reduces nitrates to nitrite, is unable to grow in media
containing 6.5% sodium chloride (NaCl) and is generally resistant to ampicillin
and carbenicillin. The guanine plus cytosine (G + C) content of the deoxy-
ribonucleic acid (DNA) is 5763% (MacInnes et al., 1979).
Aeromonas hydrophila contains thermostable O, thermolabile K and
flagellar H antigens. Serologically, the O antigen of A. hydrophila is hetero-
geneous (Sakazaki and Shimada, 1984; Janda et al., 1994, 1996). Different
serotypes have been observed from various sources of fish, isolated in different
years and places (Eddy, 1960; Bullock, 1966). Interestingly, a common antigen
has been found among virulent strains (Kou, 1972b; Leblanc et al., 1981;
Merino et al., 1996). Protein fingerprints do not correlate with biochemical
characteristics. Both phenotype and protein fingerprints show clustering of
epizootiologically related isolates (Millership and Want, 1993).
The outer membrane component protein of A. hydrophila isolates is variable
(Aoki and Holland, 1985), but Maruvada et al. (1992) detected species-specific
polypeptides of the outer membrane from A. hydrophila, and Wilcox et al.
(1992) suggested that outer membrane protein profiles were useful for
confirming the identity of A. hydrophila. Howard et al. (1993) cloned eight exe
genes of extracellular protein secretion and membrane assembly from
A. hydrophila and found them to have a high similarity with the extracellular
secretion operons of a number of different Gram-negative bacteria. Shaw and
Hodder (1978) showed that O-polysaccharides were remarkably similar
Glucose +
Glycerol +
Glycogen +
Inositol
Inulin
Lactose d
Maltose +
Mannitol +
Mannose +
Melezitose
Raffinose d
Rhamnose d
Salicin d
Sorbitol d
Sorbose
Starch +
Sucrose d
Trehalose +
Xylose
+, Typically positive; , typically negative; d, differs among strains; H
2
S,
hydrogen sulphide; DNAse, deoxyribonuclease; RNAse, ribonuclease.
Table 11.1. Continued
Characteristics Response
434
T. Aoki
structures in motile Aeromonas species, including A. hydrophila.
MacInnes et al. (1979) investigated the DNA homology of 17 strains of
A. hydrophila, which had been collected from various sources, using
A. hydrophila ATCC7966 as a reference strain. The percentage homology DNA
ranged from 39 to 100%, with a mean value of 64.7%. Aeromonas hydrophila
does not seem to show any significant divergence among the 17 strains
investigated. The 16S ribosomal DNA (rDNA) from ten species of Aeromonas
was sequenced to analyse relatedness (Martinez-Murcia et al., 1992b).
Homology for 1052 nucleotides of 16S rDNAs from ten species of Aeromonas
exhibited very high levels, ranging from 98 to 100%. The nucleotide sequences
of 16S rDNA of A. hydrophila are therefore useful in species identification. East
and Collins (1993) showed that the region encoding 23S ribonucleic acid (RNA)
from A. hydrophila was identical to that of the gamma division of Proteo-
bacteria, Escherichia coli and Plesiomonas shigelloides. Small-subunit ribo-
somal RNA (rRNA) sequences of Aeromonas were examined for a phylogenetic
analysis (Ruimy et al., 1994).
Aeromonas hydrophila strains possessing a common plasmid, with a
molecular size of 2.528 MDa, were isolated by Toranzo et al. (1983). Small
plasmids from 2.6 to 6 MDa were found in A. hydrophila isolated in Malaysia
(Ansary et al., 1992). Cryptic plasmids were also detected from A. hydrophila
strains from freshwater fish and fresh water (Noterdaeme et al., 1991). No
relationship between their virulence or biochemical characteristics and the
presence of plasmids was found (Brown et al., 1997).
DIAGNOSTIC METHODS
The gross signs of disease are haemorrhagic septicaemia and fin rot. The
aetiological agent can be grown on brainheart infusion medium, tryptosoy agar
(tryptone soya agar), nutrient agar and MacConkey agar with incubation at
2025C for 2448 h. Numerous selective media have been developed for the
isolation and presumptive identification of A. hydrophila or motile aeromonads
(Moyer, 1996), including RimlerShotts medium (Shotts and Rimler, 1973),
modified peptone beef-extract glycogen agar (McCoy and Seidler, 1973),
RippeyCabelli (membrane filter method (mA)) agar (Rippey and Cabelli,
1979), MacConkeys agar supplemented with trehalose (Kaper et al., 1981) and
starchampicillin agar (Palumbo et al., 1985). Davis and Sizemore (1981)
reported that Rimler and Shotts medium and RippeyCabelli agar were not
suitable for A. hydrophila. Arcos et al. (1988) compared six media for selective
isolation of A. hydrophila and showed that mA agar gave the best recovery rate
and also an acceptable specificity, but its selectivity was low.
An API-20E test strip is used widely for identification of the
Enterobacteriaceae (Kaper et al., 1979). Toranzo et al. (1986) indicated that the
importance of biochemical characteristics must be backed up by standardized
testing.
For diagnosis, the aetiological agent is isolated on nutrient agar at 30C
from kidney of a fish with haemorrhagic septicaemia. Isolates are motile
435 Motile Aeromonads
Gram-negative bacilli. Wakabongo et al. (1992) found that only four tests
aesculin hydrolysis, acetoin production, lysine decarboxylation and gas from
glucose were sufficient to distinguish A. hydrophila, A. caviae and A. sobria.
A number of A. hydrophila-lytic bacteriopahges were isolated from river
water, river mud, sewage and human clinical origin (e.g. stool, urine) and phage
typing was attempted (Chow and Rouf, 1983; Demarta and Peduzzi, 1984;
Altwegg et al., 1988; Merino et al., 1990; Fukuyama et al., 1991, 1992). A
comprehensive phage-typing system for A. hydrophila will provide a useful tool
for epizootiological and ecological studies.
Aeromonas hydrophila has been identified by the gel-diffusion technique
(Bullock, 1966), direct fluorescent antibody technique (Lewis and Allison,
1971), indirect fluorescent antibody technique (Lewis and Savage, 1972),
immunoblotted sodium dodecylsulphate (SDS)-polyacrylamide gel electro-
phoresis (PAGE) (Mulla and Millership, 1993) and enzyme-linked
immunosorbent assay (Merino et al., 1993). These methods are of limited value,
because many different serological types of A. hydrophila are distributed in fish
farms (Eddy, 1960; Bullock, 1966).
Lucchini and Altwegg (1992) differentiated ribotyping of restriction
genomic DNAs of aeromonads, using different fragments of a 16S rDNA gene of
E. coli as a probe and achieved identification of most Aeromonas strains to
species level. This method is easier than DNADNA hybridization, because only
minimal amounts of genomic DNA are needed and several strains can be
analysed on a single gel. Pulsed-field gel electrophoresis is a rapid and
discriminatory technique for the typing of A. hydrophila where a common origin
of infection is suspected (Talon et al., 1996).
Deoxyribonucleic acid probe hybridization technology is now becoming
available for the direct detection and identification of microorganisms. The
colony hybridization technique (Grunstein and Hogness, 1975) can be applied
easily to identify and count the causative organism. Fish-pathogenic bacteria,
such as Vibrio anguillarum (Hirono et al., 1996) and Pasteurella piscicida (Zhao
and Aoki, 1992a), can be detected by probe hybridization with their specific
DNA nucleotide sequence or cryptic plasmid. However, there are many common
DNA fragments between A. hydrophila and Aeromonas salmonicida (Miyata et
al., 1995), which makes it unlikely that this probe technique will be successful
for this species.
The sensitivity obtained using hybridization with non-radiolabelled probes
is lower than with radiolabelled probes (Zhao and Aoki, 1992a). However,
radiolabelled probes are generally unacceptable for use in diagnosis and further
development of detection procedures based on non-radiolabelling is still
required.
The polymerase chain reaction (PCR) (Mullis and Faloona, 1987) is useful
for detecting fish pathogens from diseased fish and their environment. This
technique is available for A. salmonicida (Miyata et al., 1996), Edwardsiella
tarda (Aoki and Hirono, 1995), P. piscicida (Aoki et al., 1997) and
V. anguillarum (Hirono et al., 1996). However, recently, Cascn et al. (1996)
found a specific PCR primer set for the detection of A. hydrophila hybridization
group 1.
436
T. Aoki
The DNA fingerprinting method, AFLP (amplified fragment length
polymorphism), is a valuable high-resolution genotype tool for classification of
Aeromonas species (Huys et al., 1996).
CONTROL AND TREATMENT
Transmission of the disease
The aetiological agent is transmitted horizontally but not vertically. It is
distributed widely in water and sediments of ponds and can be transmitted by
discharge from the intestinal tract and external lesions on the skin (Aoki, 1974;
Egusa, 1978; Hazen et al., 1978; Seidler et al., 1980; Kaper et al., 1981; Van der
Kooij and Hijnen, 1988; Sugita et al., 1994; Dumontet et al., 1996). Parasite
damage and fungal infection of the epidemics may allow entry and spread of
bacterial infection (Egusa, 1978).
Aeromonas hydrophila has been recognized as a pathogen not only of
amphibians, reptiles and snakes but also cattle and humans (Eddy, 1960;
Khardori and Fainstein, 1988). It has been implicated as the causative agent of
clinical infections in humans, including septicaemia and peritonitis (Janda and
Abbott, 1996). Recently, its role as a psychrotrophic spoiler of meat, seafood and
vegetables has been recognized (Palumbo, 1996). No confirmed cases of
A. hydrophila food poisoning have been reported, but its association with
gastrointestinal illness suggests that it plays a role in food-borne disease. The
epidemiological relationship among A. hydrophila isolated from fish, human and
environmental resources is particularly difficult to assess.
Aeromonas hydrophila is cosmopolitan in distribution. Infection has been
seen in many freshwater fish, e.g. ayu, carp, channel catfish, eel, gizzard shad,
salmon, snakehead fish and trout (Bullock et al., 1971; Egusa, 1978; Saitanu,
1986).
CONTROL OF DISEASE
In general, bacterial infection occurs when fish are physiologically stressed or
sanitation is poor. Good hygiene, periodic drying and disinfection of ponds are
important in prevention. Avoidance of overcrowding, low oxygen and rough
handling are the best methods of prevention.
Chemotherapy
Chemotherapeutic agents are used for the treatment of A. hydrophila in fish
farms (Aoki, 1992). Isolates of A. hydrophila have been found to be sensitive to
chloramphenicol, florfenicol, tetracycline, sulphonamide, nitrofuran derivatives
and pyridonecarboxylic acids (Aoki and Egusa, 1971; Endo et al., 1973; Katae
et al., 1979; Fukui et al., 1987). Fluorinated analogue, tetracycline derivatives,
437 Motile Aeromonads
nitrofuran derivatives, sulphonamide and pyridonecarboxylic acids are effective
in oral treatments (Austin and Austin, 1987). A review by the Fisheries Agency
of the Japanese government on chemotherapeutic use against bacterial infections
in cultured fish has been carried out (Okamoto, 1992). The use of chlor-
amphenicol and almost all nitrofuran derivatives in fish have been restricted in
Japan since 1983.
The extensive use of antibacterial agents has led to an increase in resistant
strains of A. hydrophila. Drug-resistant strains carrying transferable R plasmids
appeared in 1971 in cultured eel and multiple resistant strains carrying R
plasmids are now distributed widely in cultured amago, ayu, carp and eel in
Japan (Aoki et al., 1971, 1972; Aoki and Watanabe, 1973; Aoki, 1974, 1975).
Almost all strains have intrinsic resistance to ampicillin, and drug-resistant
strains carried transferable R plasmid, which encoded resistance to sulpho-
namide and tetracycline and to chloramphenicol, sulphonamide and
streptomycin (Akashi and Aoki, 1986). Drug-resistant strains have been isolated
from the water, as well as from the intestinal tracts of cultured fish (Aoki, 1974,
1975) in France, Ireland (Hedges et al., 1985), Malaysia (Ansary et al., 1992;
Son et al., 1997), Taiwan (Kou and Chung, 1980) and the USA (Shotts et al.,
1976). Recently, drug-resistant strains have been isolated from cultured
snakehead (O. striatus) in Thailand (Aoki et al., 1990). The DNA of transferable
R plasmids was classified into incompatibility (Inc) (Couturier et al., 1988)
groups AC, which had been detected in different areas, and resistance to
sulphonamide and tetracycline and showed high homology with each other
(Akashi and Aoki, 1986). R plasmids classified into Inc group U, with resistance
to chloramphenicol, sulphonamide and streptomycin, showed homology within
a specific species and with R plasmids from A. salmonicida. Aoki (1988)
demonstrated that A. hydrophila strains carrying R plasmids having identical
DNA structures are widely distributed in cultured freshwater fish in various
areas.
Tetracycline-resistance (tet) genes on the R plasmids of Gram-negative
bacteria have been classified on the basis of their DNA structure into six classes;
Tet A, B, C, D, E, F and G (Levy, 1988; Zhao and Aoki, 1992b). The tet gene of
the R plasmids detected from A. hydrophila, which were isolated in Japan, was
classified into Tet D class (Aoki and Takahashi, 1987). Class D and E
tetracycline-resistance determinants were found in resistant strains of
A. hydrophila isolated in catfish ponds in the USA (DePaola and Roberts, 1995).
A class D tetracycline-resistance determinant of RA1 from A. hydrophila
contained the resistance (tet A) and repressor (tet R) genes. The tet A gene
encoded 394 amino acids, with a calculated molecular size of 41.1 kDa (Varela
and Griffith, 1993), and the tet R encoded 218 amino acids, with a calculated
molecular size of 24.4 kDa (Unger et al., 1984).
The 40 MDa plasmid encoded with resistance to carbenicillin and
novobiocin also regulated adherence and production of a potent haemolysin
(Hanes and Chandler, 1993).
438
T. Aoki
PATHOGENESIS AND IMMUNITY
A variety of possible virulence factors of A. hydrophila have been suggested,
including lipopolysaccharides (endotoxins), extracellular products (ECP),
siderophores, the ability of attachment to host cells and surface proteins. The
ECP include a cytotoxin, enterotoxin, haemolysins, protease, haemagglutinin
and acetyl cholinesterase (Cahill, 1990; Gosling, 1996; Howard et al., 1996).
Aeromonas hydrophila enters through the epithelium of the intestinal tract of
fish. Enterotoxins of A. hydrophila cause fluid to accumulate in ligated rabbit
ileal loops. Enterotoxins are divided into two types, cytotonic and cytotoxic.
Boulanger et al. (1977) isolated two different haemolysins, an - and a
-haemolysin, both of which have been implicated in the pathogenesis of
infection. Allan and Stevenson (1981) investigated the production of protease
and haemolysin in the ECP of A. hydrophila strains and showed a close
correlation between the quantity of haemolysin and toxicity to fish. Two
biologically similar but immunologically distinct haemolysins were purified by
Asao et al. (1986) and Kozaki et al. (1987, 1989). These haemolysins (cytotoxic
enterotoxin) had enterotoxic activity and caused fluid accumulation in infant
mice, in mouse intestine and in rabbit ligated ileal loops. Purified haemolysin
also displayed cytotoxicity to Vero cells and lethal toxicity to mice. The
molecular weight of the haemolysin was estimated at 48,00050,000 kDa and
biological activity was inactivated with heating for 5 min at 65C (Asao et al.,
1984).
Cross-reaction of A. hydrophila haemolysin and cholera toxin has been
reported, and a specific synthetic oligonucleotide probe for regions of A + B
subunits of cholera toxin was found to hybridize with chromosomal DNA from
some strains of A. hydrophila. The enterotoxin purified by Rose et al. (1989) had
a molecular size of 52,000, but only 25 residues of the N-terminal sequences
were found to be identical to the haemolysin of A. hydrophila (aerolysin)
(Howard et al., 1987). The amino acid residues differed significantly between
enterotoxin and aerolysin over the vast majority of the protein. The structure of
proaerolysin was determined by X-ray crystallography at 2.8 (Parker et al.,
1994). The aerolysin secretion system indicates that it is a haemolysin which is
directed outside the cell through periplasm (Wong and Buckley, 1993).
Five haemolysin genes (AHH1, AHH2, AHH3, AHH4 and AHH5) were
cloned from A. hydrophila into a E. coli vector (Aoki and Hirono, 1991; Hirono
and Aoki, 1991; Hirono et al., 1992). Each was classified into three groups,
depending on their nucleotide sequences. AHH1 belonged to one group (group
1), which contained part of a homologous sequence of the haemolysin genes of
Vibrio cholerae El Tor and Vibrio vulnificus (Rader and Murphy, 1988). There
were two highly conservative regions, and the location of the cysteine residue
was conserved. These regions may be important in the control of haemolytic
activity (Fig. 11.6). The other group (group 2) included AHH3, AHH4 and
AHH5, the previously reported aerolysin gene from A. hydrophila and A. trota
(Howard et al., 1987; Husslein et al., 1988) (Fig. 11.7). Chopra et al. (1993)
cloned a cytolytic enterotoxin gene from A. hydrophila. The gene was also
classified into group 2 by the nucleotide sequence. The remaining gene, AHH2,
439 Motile Aeromonads
is placed in group 3. From colony hybridization analysis, using the cloned
haemolysin genes, it was found that AHH1 (group 1) and AHH5 (group 2) were
widely distributed among aeromonads (Table 11.2). It is interesting that all
tested strains of A. salmonicida have AHH1 and AHH5 genes. One strain of A.
hydrophila has two or three haemolysin genes (Hirono and Aoki, 1991; Hirono
et al., 1992).
A significant qualitative as well as quantitative difference in the protease
components of ECP was produced by A. hydrophila and A. sobria, which were
pathogenic for fish (Nieto and Ellis, 1991). The role of protease in the virulence
of A. hydrophila is currently controversial. Wakabayashi et al. (1981) described
most of the virulent strains of A. hydrophila biovar. hydrophila as having a high
proteolytic activity. A single protease purified from A. hydrophila was lethal to
carp and was dermonecrotic to guinea-pigs. Thune et al. (1982) and Lallier et al.
(1984) found A. hydrophila protease to be lethal. Lallier et al. (1984) also
showed that both virulent and weakly virulent strains were dermonecrotic in the
guinea-pig, and a dermonecrotic factor was observed in sonicated cells of an
A. hydrophila strain isolated from eel (Shimizu, 1968). Chabot and Thune
Fig. 11.6. Comparison of deduced amino acid sequences among the AHH1, ASH4 and Vibrio
cholerae El Tor haemolysin (Rader and Murphy, 1988). The same amino acid residues are
indicated as on the aligned sequences. VCH, V. cholerae El Tor haemolysin.
440
T. Aoki
(1991) characterized three proteases and found no correlation between either
qualitative or quantitative protease production and virulence in age-0 channel
catfish. A metalloprotease with a molecular size of 38 kDa and a serine
proteinase with a molecular size of 22 kDa was purified from A. hydrophila
strain B52. These were stable at 56C for 10 min and had a lethal effect in fish,
with a median lethal dose (LD
50
) of 150 ng g
1
. Only the serine protease
possessed cytotoxic activity (Rodriguez et al., 1992). Leung and Stevenson
(1988) found two distinct types of extracellular protease: thermostable
Fig. 11.7. Comparison of deduced amino acid sequences among the AHH3, AHH4, AHH5,
ASH3, ASA1, Aeromonas hydrophila aerolysin (Howard et al., 1987) and A. trota aerolysin
(Husslein et al., 1988). The same amino acid residues are indicated as on the aligned sequences.
AHAER, A. hydrophila aerolysin; ATAER, A. trota aerolysin.
441 Motile Aeromonads
metalloprotease and thermolabile serine protease. The relationship between
these and the two purified by Nieto and Ellis (1986) is not known, but they were
also lethal to fish. Protease from A. hydrophila enhanced haemolysin activity
(Howard and Buckley, 1985), but Lallier et al. (1984) found that purified
haemolysin from A. hydrophila was not lethal to fish. A novel zinc-proteinase
was purified and characterized from A. hydrophila (Loewy et al., 1993), and
Rivero et al. (1990) cloned an extracellular protease gene from A. hydrophila.
Aeromonas hydrophila strains, which had haemolytic activity, enterotoxin
productivity and cytotoxic ability, were not virulent for rainbow trout (Santos et
al., 1988).
Yadav et al. (1992) noted that the fish cell lines, especially the BB (brown
bullhead, Ictalurus nebulosus) cells were sensitive targets for A. hydrophila
cytotoxins, even when assayed in conditions vastly different from those of
mammalian cells. Cytotoxin-producing strains were frequently associated with
epizootic ulcerative syndrome (EUS)-infected fish, compared with healthy fish.
Cytotoxic A. hydrophila strains have a role in the pathogenicity and progression
of EUS (Yadav et al., 1992).

Table 11.2. Colony hybridization analysis of Aeromonas species A.


hydrophila, A. sobria, A. caviae, A. veronii and A. salmonicida, using AHH1,
AHH5, ASH1 and ASA1 DNA probes.
Sources DNA probes
(no. of tested
Strains strains) AHH1 AHH5 ASH1 ASA1
Aeromonas Canned milk (1) 1* (1) 1 (1) 0 (0) 0 (0)
hydrophila Human (14) 9 (9) 4 (11) 0 (0) 8 (8)
Fish (31) 15 (16) 11 (23) 3 (3) 21 (26)
Environment (5) 5 (5) 4 (5) 0 (0) 5 (5)
Turtle (1) 1 (1) 1 (1) 0 (0) 1 (1)
Total (52) 31 (32) 21 (41) 3 (3) 35 (40)
Aeromonas sobria Human (18) 2 (3) 3 (7) 0 (0) 18 (18)
Fish (4) 0 (1) 1 (3) 0 (0) 4 (4)
Environment (2) 1 (1) 0 (2) 0 (0) 2 (2)
Bovine (1) 0 (0) 0 (0) 0 (0) 1 (1)
Frog (1) 0 (0) 0 (1) 0 (0) 1 (1)
Total (26) 3 (5) 4 (12) 0 (0) 26 (26)
Aeromonas caviae Fish (6) 2 (2) 1 (4) 0 (0) 6 (6)
Environment (4) 2 (2) 1 (2) 0 (0) 3 (3)
Pig (1) 1 (1) 1 (1) 0 (0) 1 (1)
Total (11) 5 (5) 3 (7) 0 (0) 10 (10)
Aeromonas veronii Human (1) 0 (0) 0 (0) 0 (0) 1 (1)
Aeromonas Salmonid 104 (104) 104 (104) 1 (1) 104 (104)
salmonicida fish (104)
*Hybridized number using high-stringency condition.

Hybridized number using low-stringency condition.


442
T. Aoki
Aoki and Holland (1985) observed that iron-binding proteins with a
molecular size of 6870 kDa in A. hydrophila were induced under iron-limiting
conditions. Enterobactin, the catecholate siderophore produced by a strain of
A. hydrophila (Andrus and Payne, 1983), and amonabactin, a novel phenolate
siderophore in A. hydrophila 495A2, were identified (Barghouthi et al., 1989,
1991). Esteve and Amaro (1991) found the hydroxamate-type siderophores
produced by A. hydrophila. The iron-uptake system was similar in A. hydrophila
and A. sobria isolated from European eels (Anguilla anguilla) and in
A. salmonicida. These authors suggested that siderophore produced by
A. hydrophila could be important and play a role in virulence for acquisition of
iron from the host. Recently, the amonabactins were synthesized and their
spectroscopic properties were elucidated (Telford et al., 1994).
The receptor cell surface of A. hydrophila can bind to iron-containing
proteins lactoferrin, transferrin, ferritin, cytochrome C and haemin of the
host (Kishore et al., 1991; Ascencio et al., 1992), demonstrating a relationship
between lactoferrin binding and siderophore production by the bacteria. A
carbohydrate-reactive outer-membrane protein, which may contribute as an
adhesive mechanism, was isolated from A. hydrophila strain A6 (Quinn et al.,
1993, 1994). This protein is related to the colonization strategies of aeromonads
associated with human enteric disease. However, the protein has not been proved
to relate to pathogenicity in fish.
Two cytotonic enterotoxin genes were cloned from A. hydrophila; one of the
enterotoxins was heat-labile at 56C, while the other was heat-stable (Chopra
et al., 1994).
The role of haemolytic activity and cytotoxic activity in contributing to
pathogenicity to fish still needs to be elucidated.
The attachment of a pathogen to the epithelial tissue is the first step in the
host disease process. The W pili of A. hydrophila strain Ae6 is the colonization
factor for the intestine, as well as a haemagglutinin (Hokama and Nakasone,
1990; Hokama et al., 1990). Haemagglutinating activity was also detected in
A. hydrophila (Atkinson and Trust, 1980; Corral et al., 1990). Aeromonas
hydrophila exhibited aggregative adherence to HEp-2 cells (Neves et al., 1994),
fish tissue culture and mucus-coated glass slides (Krovacek et al., 1987).
Adherence and haemagglutination were not significantly correlated with
virulence in fish (Corral et al., 1990).
De Meuron and Peduzzi (1979) have suggested that the K-antigen is a
pathogenicity factor. Virulent strains of A. hydrophila possessed an S-layer on
the cell surface (Dooley et al., 1986; Dooley and Trust, 1988; Ford and Thune,
1991, 1992).
Recently, Rodriquez et al. (1993) purified acetylcholinesterase from ECP
of A. hydrophila and showed it to be lethal to fish. Glycerophospholipid
: cholesterol acyltransferase (GCAT) and lipopolysaccharide complex enhanced
the lethal exotoxicity and cytolysin of A. salmonicida (Lee and Ellis, 1990). A
gene encoding GCAT was cloned from A. hydrophila (Thornton et al., 1988).
Experimental vaccination for prophylaxis against infection of A. hydrophila
has been examined (Stevenson, 1988). Fish immunized either intramuscularly or
intraperitoneally with vaccine showed protection against challenge. The
443 Motile Aeromonads
agglutinating antibody titre increased in the serum of immunized fish (Song
et al., 1976; Ruangpan et al., 1986; Karunasagar et al., 1991).
Immersion vaccination of channel catfish using polyvalent sonicated
antigens of A. hydrophila provides protection (Thune and Plumb, 1982). Lamers
et al. (1985) noted that agglutinating antibody was recognized in the serum of
carp immunized with A. hydrophila bacterin, following a second immersion with
this vaccine. However, fish vaccinated by immersion or orally showed
questionable protection.
Catfish immunized intraperitoneally by injection with the acid extract of the
S-layer protein of A. hydrophila were protected from the homologous, virulent
strain (Ford and Thune, 1992). Serological types of A. hydrophila are
heterogeneous and a polyvalent vaccine is thought to be necessary for
prevention of the infection.
TOPICS FOR FURTHER STUDY
There is significant interest in the pathogenicity of A. hydrophila to fish. Several
virulence factors, including the production of endotoxin, ECP, siderophores and
surface proteins, and the ability of attachment to host cells, are under study, but
their relative importance has not yet been elucidated. Cahill (1990) considers
that proteases are most important in fish pathogenicity, but further study on
adhesins, siderophores and surface proteins is needed to understand their role in
the pathogenesis of disease caused by A. hydrophila.
REFERENCES
Akashi, A. and Aoki, T. (1986) Characterization of transferable R plasmids from
Aeromonas hydrophila. Bulletin of the Japanese Society of Scientific Fisheries 52,
649655.
Allan, B.J. and Stevenson, R.M.W. (1981) Extracellular virulence factors of Aeromonas
hydrophila in fish infections. Canadian Journal of Microbiology 27, 11141122.
Allen, D.A., Austin, B. and Colwell, R.R. (1983) Aeromonas media, a new species
isolated from river water. International Journal of Systematic Bacteriology 33,
599604.
Altwegg, M., Altwegg-Bissig, R., Demarta, A., Peduzzi, R., Reeves, M.W. and
Swaminathan, B. (1988) Comparison of four typing methods for Aeromonas
species. Journal of Diarrhoeal Diseases Research 6, 8894.
Andrus, C. and Payne, S.M. (1983) In: The 83rd Annual Meeting of the American Society
for Microbiology, New Orleans, Louisiana, March 1983. American Soceity for
Microbiology, Washington, DC, Abstract D13, p. 61.
Ansary, A., Haneef, R.M., Torres, J.L. and Yadav, M. (1992) Plasmids and antibiotic
resistance in Aeromonas hydrophila isolated in Malaysia from healthy and diseased
fish. Journal of Fish Diseases 15, 191196.
Aoki, T. (1974) Studies of drug-resistant bacteria isolated from water of carp-ponds and
intestinal tracts of carp [in Japanese]. Bulletin of the Japanese Society of Scientific
Fisheries 40, 247254.
444
T. Aoki
Aoki, T. (1975) Effects of chemotherapeutics on bacterial ecology in the water of ponds
and the intestinal tracts of cultured fish, ayu (Plecoglossus altivelis). Japanese
Journal of Microbiology 19, 712.
Aoki, T. (1988) Drug-resistant plasmids from fish pathogens. Microbiological Sciences
5, 219223.
Aoki, T. (1992) Chemotherapy and drug resistance in fish farms in Japan. In: Shariff, M.,
Subasinghe, R.P. and Arthur, J.R. (eds) Diseases in Asian Aquaculture, Vol. I. Fish
Health Section, Asian Fisheries Society, Manila, Philippines, pp. 519529.
Aoki, T. and Egusa, S. (1971) Drug sensitivity of Aeromonas liquefaciens isolated from
freshwater fishes. Bulletin of the Japanese Society of Scientific Fisheries 37,
176185.
Aoki, T. and Hirono, I. (1991) Cloning and characterization of the haemolysin
determinants from Aeromonas hydrophila. Journal of Fish Diseases 14, 303312.
Aoki, T. and Hirono, I. (1995) Detection of the fish-pathogenic bacteria Edwardsiella
tarda by polymerase chain reaction. In: Kuo, C.-M., Wu, J.-L. Hwang, P.-P. (eds)
Proceedings of the International Symposium on Biotechnology Application in
Aquaculture. Asian Fisheries Society Special Publication No. 10, Asian Fisheries
Society, Manila, pp. 135146.
Aoki, T. and Holland, B.I. (1985) The outer membrane proteins of the fish pathogens
Aeromonas hydrophila, Aeromonas salmonicida and Edwardsiella tarda. FEMS
Microbiology Letters 27, 299305.
Aoki, T. and Takahashi, A. (1987) Class D tetracycline resistance determinants of R
plasmids from the fish pathogens Aeromonas hydrophila, Edwardsiella tarda, and
Pasteurella piscicida. Antimicrobial Agents and Chemotherapy 31, 12781280.
Aoki, T. and Watanabe, T. (1973) Studies of drug-resistant bacteria isolated from
eel-pond water and intestinal tracts of the eel (Anguilla japonica and Anguilla
anguilla) [in Japanese]. Bulletin of the Japanese Society of Scientific Fisheries 39,
121130.
Aoki, T., Egusa, S., Ogata, Y. and Watanabe, T. (1971) Detection of resistance factors in
fish pathogen Aeromonas liquefaciens. Journal of General Microbiology 65,
343349.
Aoki, T., Egusa, S., Yada, C. and Watanabe, T. (1972) Studies of drug resistance and R
factors in bacteria from pond-cultured salmonids. Japanese Journal of
Microbiology 16, 233238.
Aoki, T., Umeda, T., Takami, K., Kitao, T., Saitanu, K., Chongthaleong, A. and
Punyaratabandhu, P. (1990) Drug-resistant Aeromonas hydrophila in Thailand. In:
Hirano, R. and Hanyu, I. (eds) The Second Asian Fisheries Forum. Asian Fishereies
Society, Manila, pp. 693696.
Aoki, T., Ikeda, D., Katagiri, T. and Hirono, I. (1997) Rapid detection of the fish-
pathogenic bacterium Pasteurella piscicida by polymerase chain reaction targetting
nucleotide sequences of the species-specific plasmid pZP1. Fish Pathology 32,
143151.
Arcos, M.L., de Vicente, A., Moriigo, M.A., Romero, P. and Borrego, J.J. (1988)
Evaluation of several selective media for recovery of Aeromonas hydrophila from
polluted waters. Applied and Environmental Microbiology 54, 27862792.
Asao, T., Kinoshita, Y., Kozaki, S., Uemura, T. and Sakaguchi, G. (1984) Purification
and some properties of Aeromonas hydrophila hemolysin. Infection and Immunity
46, 122127.
Asao, T., Kozaki, S., Kato, K., Kinoshita, Y., Otsu, K., Uemura, T. and Sakaguti, G.
(1986) Purification and characterizaition of an Aeromonas hydrophila hemolysin.
Journal of Clinical Microbiology 24, 228232.
445 Motile Aeromonads
Ascencio, F., Ljungh, A. and Wadstrom, T. (1992) Characterization of lactoferrin bind-
ing by Aeromonas hydrophila. Applied and Environmental Microbiology 58, 4247.
Atkinson, H.M. and Trust, T.J. (1980) Hemagglutination properties and adherence
ability of Aeromonas hydrophila. Infection and Immunity 27, 938946.
Austin, B. and Austin D.A. (1987) Bacterial Fish Pathogens: Disease in Farmed and
Wild Fish. Ellis Horwood, Chichester, UK.
Barghouthi, S., Young, R., Olson, M.O.J., Arceneaux, J.E.L., Clem, L.W. and Byers,
B.R. (1989) Amonabactin, a novel tryptophan- or phenylalanine-containing
phenolate siderophore in Aeromonas hydrophila. Journal of Bacteriology 171,
18111816.
Barghouthi, S., Payne, S.M., Arceneaux, J.E.L. and Byers, B.R. (1991) Cloning,
mutagenesis, and nucleotide sequence of a siderophore biosynthetic gene (amoA)
from Aeromonas hydrophila. Journal of Bacteriology 173, 51215128.
Boulanger, Y., Lallier, R. and Cousineau, G. (1977) Isolation of enterotoxigenic
Aeromonas from fish. Canadian Journal of Microbiology 23, 11611164.
Brown, R.L., Sanderson, K. and Kirov, S.M. (1997) Plasmids and Aeromonas virulence.
FEMS Immunology and Medical Microbiology 17, 217223.
Bullock, G.L. (1966) Precipitins and agglutinin reaction of aeromonads isolated from
fish and other sources. Bulletin de lOffice International des Epizooties 65,
805824.
Bullock, G.L., Conroy, D.A. amd Snieszko, S.F. (1971) Septicemic diseases caused by
motile aeromnads and pseudomonads. In: Snieszko, S.F. and Axelrod, H.R. (eds)
Diseases of Fishes. Book 2A: Bacterial Diseases of Fishes. TFH Publications,
Neptune, New Jersey, pp. 2141.
Cahill, M.M. (1990) Virulence factors in motile Aeromonas species. Journal of Applied
Bacteriology 69, 116.
Carnahan, A.M. and Joseph, S.W. (1993) Systematic assessment of geographically and
clinically diverse aeromonads. Systematic Applied Microbiology 16, 7284.
Carnahan, A., Fanning, G.R. and Joseph, S.W. (1991a) Aeromonas jandaei (formerly
genospecies DNA group 9 A. sobria), a new sucrose-negative species isolated from
clinical specimens. Journal of Clinical Microbiology 29, 560564.
Carnahan, A.M., Chakraborty, T., Fanning, G.R., Verma, D., Ali, A., Janda, J.M. and
Joseph, S.W. (1991b) Aeromonas trota sp. nov., an ampicillin-susceptible species
isoalted from clinical specimens. Journal of Clinical Microbiology 29, 12061210.
Cascn, A., Anguita, J., Hernanz, C., Snchez, M., Fernndez, M. and Naharro, G.
(1996) Identification of Aeromonas hydrophila hybridization group 1 by PCR
assays. Applied and Environmental Microbiology 62, 11671170.
Chabot, D.J. and Thune, R.L. (1991) Proteases of the Aeromonas hydrophila complex:
identification, characterization and relation to virulence in channel catfish,
Ictalurus punctatus (Rafinesque). Journal of Fish Diseases 14, 171183.
Chopra, A.K., Houston, C.W., Peterson, J.W. and Jin, G.-F. (1993) Cloning, expression,
and sequence analysis of a cytolytic enterotoxin gene from Aeromonas hydrophila.
Canadian Journal of Microbiology 39, 513523.
Chopra, A.K., Pham, R. and Houston, C.W. (1994) Cloning and expression of putative
cytotoxinic enterotoxin-encoding genes from Aeromonas hydrophila. Gene 139,
8791.
Chow, M.S. and Rouf, M.A. (1983) Isolation and partial characterization of two
Aeromonas hydrophila bacteriophages. Applied and Environmental Microbiology
45, 16701676,
Collins, M.D., Martinez-Murcia, A.J. and Cai, J. (1993) Aeromonas enteropelogenes and
Aeromonas ichthiosmia are identical to Aeromonas trota and Aeromonas veronii,
446
T. Aoki
respectively, as revealed by small-subunit rRNA sequence analysis. International
Journal of Systematic Bacteriology 43, 855856.
Colwell, R.R., MacDonell, M.T. and Deley, J.D. (1986) Proposal to recognize the family
Aeromonadaceae fam. nov. International Journal of Systematic Bacteriology 36,
473477.
Corral, F.D., Shotts, E.B. and Brown, J. (1990) Adherence, haemagglutinination and cell
surface characteristics of motile aeromonads virulent for fish. Journal of Fish
Diseases 13, 255268.
Couturier, M., Bex, F., Bergquist, P.L. and Maas, W.K. (1988) Identification and
classification of bacterial plasmids. Microbiological Reviews 52, 375395.
Davis, J.W. and Sizemore, R.K. (1981) Nonselectivity of RimlerShotts medium for
Aeromonas hydrophila in estuarine environments. Applied and Environmental
Microbiology 42, 544545.
Demarta, A. and Peduzzi, R. (1984) Etude pidmiologique des Aeromonas par
lysotypie. Estratto dalla Rivista Italiana di Piscicoltura e Ittiopatologia 19,
148155.
de Meuron, P.A. and Peduzzi, R. (1979) Characterization of Aeromonas strains isolated
from fresh water fishes and various reptiles. Zentralblatt fr Venterinarmedizin (B)
26, 153167.
DePaola, A. and Roberts, M.C. (1995) Class D and E tetracycline resistance
determinants in Gram-negative bacteria from catfish ponds. Molecular and Cellular
Probes 9, 311313.
Dooley, J.S.G. and Trust, T.J. (1988) Surface protein composition of Aeromonas
hydrophila strains virulent for fish: identification of a surface array protein. Journal
of Bacteriology 170, 499506.
Dooley, J.S.G., Lallier, R. and Trust, T.J. (1986) Antigenic structure of Aeromonas
hydrophila. Veterinary Immunology and Immunopathology 12, 339344.
Dumontet, S., Krovacek, K., Baloda, S.B., Grottoli, R., Pasquale, V. and Vanucci, S.
(1996) Ecological relationship between Aeromonas and Vibrio spp. and planktonic
copepods in the coastal marine environment in southern Italy. Comparative
Immunology, Microbiology and Infectious Diseases 19, 245254.
East, A.K. and Collins, M.D. (1993) Molecular characterization of DNA encoding 23S
rRNA and 16S-23S rRNA intergenic spacer regions of Aeromonas hydrophila.
FEMS Microbiology Letters 106, 129134.
Eddy, B.P. (1960) Cephalotrichous, fermentative Gram-negative bacteria; the genus
Aeromonas. Journal of Applied Bacteriology 23, 216249.
Egusa, S. (1978) Infectious Diseases of Fish [in Japanese]. Kouseisha Kouseikaku,
Tokyo, 554 pp.
Endo, T., Ogishima, K., Hayasaka, H., Kaneko, S. and Ohshima, S. (1973) Application
of oxolinic acid as a chemotherapeutic agent against infections diseases in fishes I.
Antibacterial activity, chemotherapeutic effects and pharmacokinetics of oxolinic
acid fishes [in Japanese]. Bulletin of the Japanese Society of Scientific Fisheries 39,
165171.
Esteve, C. and Amaro, C. (1991) Siderophore production in Aeromonas spp. isolated
from European eel, Anguilla anguilla L. Journal of Fish Diseases 14, 423427.
Esteve, C., Gutirrez, M.C. and Ventosa, A. (1995a) DNA relatedness among Aeromonas
allosaccharophila strains and DNA hybridization groups of the genus Aeromonas.
International Journal of Systematic Bacteriology 45, 390391.
Esteve, C., Gutirrez, M.C. and Ventosa, A. (1995b) Aeromonas encheleia sp. nov.,
isolated from European eels. International Journal of Systematic Bacteriology 45,
462466.
447 Motile Aeromonads
Fijan, N.N. (1972) Infectious dropsy in carp a disease complex. In: Mawdesley, L.E.
(ed.) Diseases of Fish. Symposia of the Zoological Society of London, No. 30,
Academic Press, London, pp. 3951.
Ford, L.A. and Thune, R.L. (1991) S-layer positive motile aeromonads isolated from
channel catfish. Journal of Wildlife Diseases 27, 557561.
Ford, L.A. and Thune, R.L. (1992) Immunization of channel catfish with a crude, acid-
extracted preparation of motile aeromonad S-layer protein. Biomedical Letters 47,
335362.
Fukui, H., Fujihara, Y. and Kano, T. (1987) In vitro and in vivo antibacterial activities of
florfenicol, a new fluorinated analog of thiamphenicol, against fish pathogens. Fish
Pathology 22, 201207.
Fukuyama, M., Kamimura, T., Itoh, T., Hara, M., Tabuchi, K., Murata, M. and Kohzaki,
K. (1991) Studies on motile Aeromonas infection: 2. Development of a
bacteriophage typing system for motile Aeromonas [in Japanese]. Kansenshogaku
Zasshi 65, 813819.
Fukuyama, M., Kawakami, K., Imagawa, Y., Itoh, T., Hara, M. and Tabuchi, K. (1992)
Studies on motile Aeromonas infection: 3. Phage typing of motile Aeromonas
isolated from patients with diarrhoea [in Japanese]. Kansenshogaku Zasshi 66,
628631.
Gosling, P.J. (1996) Pathogenic mechanisms. In: Austin, B., Altwegg, M., Gosling, P.J.
and Joseph, P. (eds) The Genus Aeromonas. John Wiley and Sons, Chichester, pp.
245265.
Grunstein, M. and Hogness, D.S. (1975) Colony hybridization: a method for the
isolation of cloned DNAs that contain a specific gene. Proceedings of the National
Academy of Sciences of the USA 72, 39613965.
Hanes, D.E. and Chandler, D.K.F. (1993) The role of a 40-megadalton plasmid in the
adherence and hemolytic properties of Aeromonas hydrophila. Microbial
Pathogenesis 15, 313317.
Hazen, T.C., Fliermans, C.B., Hirsch, R.P. and Esch, G.W. (1978) Prevalence and
distribution of Aeromonas hydrophila in the United States. Applied and
Environmental Microbiology 36, 731738.
Hedges, R.W., Smith, P. and Brazil, G. (1985) Resistance plasmids of aeromonads.
Journal of General Microbiology 131, 20912095.
Hickman-Brenner, F.W., MacDonald, K.L., Steigerwalt, A.G., Fanning, G.R., Brenner,
D.J. and Farmer, J.J., III (1987) Aeromonas veronii, a new ornithine decarboxylase-
positive species that may cause diarrhea. Journal of Clinical Microbiology 25,
900906.
Hickman-Brenner, F.W., Fanning, G.R., Arduino, M.J., Brenner, D.J. and Farmer J.J., III
(1988) Aeromonas schubertii, a new mannitol-negative species found in human
clinical specimens. Journal of Clinical Microbiology 26, 15611564.
Hirono, I. and Aoki, T. (1991) Nucleotide sequence and expression of an extracellular
hemolysin gene of Aeromonas hydrophila. Microbial Pathogenesis 11, 189197.
Hirono, I., Aoki, T., Asao, T. and Kozaki, S. (1992) Nucleotide sequences and
characterization of haemolysin genes from Aeromonas hydrophila and Aeromonas
sobria. Microbial Pathogenesis 13, 433446.
Hirono, I., Masuda, T. and Aoki, T. (1996) Cloning and detection of the hemolysin gene
of Vibrio anguillarum. Microbial Pathogenesis 21, 173182.
Hokama, A. and Nakasone, N. (1990) Pili of Aeromonas hydrophila: purification,
characterization, and biological role. Microbiology and Immunology 34, 8398.
Hokama, A., Honma, Y. and Nakasone, N. (1990) Pili of an Aeromonas hydrophila strain
as a possible colonization factor. Microbiology and Immunology 34, 901915.
448
T. Aoki
Hoshina, T. (1962) Studies on Red-fin Disease of Eel [in Japanese]. Special Research
Report of Tokyo University of Fisheries, No. 6, Tokyo, 105 pp.
Howard, S.P. and Buckley, J.T. (1985) Activation of the hole-forming toxin aerolysin by
extracellular processing. Journal of Bacteriology 163, 336340.
Howard, S.P., Garland, W.J., Green, M.J. and Buckley, T. (1987) Nucleotide sequence of
the gene for the hole-forming toxin aerolysin of Aeromonas hydrophila. Journal of
Bacteriology 169, 28692871.
Howard, S.P., Critch, J. and Bedi, A. (1993) Isolation and analysis of eight exe genes and
their involvement in extracellular secretion and outer membrane assembly in
Aeromonas hydrophila. Journal of Bacteriology 175, 66956703.
Howard, S.P., Macintyre, S. and Buckley, J.T. (1996) Toxins. In: Austin, B., Altwegg,
M., Gosling, P.J. and Joseph, P. (eds) The Genus Aeromonas. John Wiley and Sons,
Chichester, pp. 267286.
Husslein, V., Huhle, B., Jarchau, T., Lurz, R., Goebel, W. and Chakraborty, T. (1988)
Nucleotide sequence and transcriptional analysis of the aerCaerA region of
Aeromonas sobria encoding aerolysin and its regulatory region. Molecular
Microbiology 2, 507517.
Huys, G., Coopman, R., Janssen, P. and Kersters, K. (1996) High-resolution genotypic
analysis of the genus Aeromonas by AFLP fingerprinting. International Journal of
Systematic Bacteriology 46, 572580.
Janda, J.M. and Abbott, S.L. (1996) Human pathogens. In: Austin, B., Altwegg, M.,
Gosling, P.J. and Joseph, P. (eds) The Genus Aeromonas. John Wiley and Sons,
Chichester, pp. 149173.
Janda, J.M., Guthertz, L.S., Kokka, R.P. and Shimada, T. (1994) Aeromonas species in
septicemia: laboratory characteristics and clinical observations. Clinical Infectious
Diseases 19, 7783.
Janda, J.M., Abbott, S.L., Khashe, S., Kellogg, G.H., and Shimada, T. (1996) Further
studies on biochemical characteristics and serologic properties of the genus
Aeromonas. Journal of Clinical Microbiology 34, 19301933.
Kaper, J., Seidler, R.J., Lockman, H. and Colwell, R.R. (1979) Medium for the
presumptive identification of Aeromonas hydrophila and Enterobacteriaceae.
Applied and Environmental Microbiology 38, 10231026.
Kaper, J.B., Lockman, H., Colwell, R. and Joseph, S.W. (1981) Aeromonas hydrophila:
ecology and toxigenicity of isolates from an estuary. Journal of Applied
Bacteriology 50, 359377.
Karunasagar, I., Rosalind, G. and Karunasagar, I. (1991) Immunological response of the
Indian major carps to Aeromonas hydrophila vaccine. Journal of Fish Diseases 14,
413417.
Katae, H., Kouno, K., Takase, Y., Miyazaki, H., Hashimoto, M. and Shimizu, M. (1979)
The evaluation of piromidic acid as an antibiotic in fish: an in vitro and in vivo
study. Journal of Fish Diseases 2, 321335.
Khardori, N. and Fainstein, V. (1988) Aeromonas and Plesiomonas as etiological agents.
Annual Review of Microbiology 42, 395419.
Kishore, A.R., Erdei, J., Naidu, S.S., Falsen, E., Forsgren, A. and Naidu, A.S. (1991)
Specific binding of lactoferrin to Aeromonas hydrophila. FEMS Microbiology
Letters 67, 115119.
Kou, G.H. (1972a) Studies on the occurrence and biochemical properties of virulent and
avirulent strains of freshwater fish pathogen, Aeromonas liquefaciens. Journal of
the Fisheries Society of Taiwan 1, 813.
Kou, G.H. (1972b) Study on the fish pathogen, Aeromonas liquefaciens I [in Chinese].
Aquaculture 2, 2233.
449 Motile Aeromonads
Kou, G.H. (1973) Study on the fish pathogen, Aeromonas liquefaciens II. The
connections between pathogenic properties and the activities of toxic substances.
Journal of the Fisheries Society of Taiwan 2, 4246.
Kou, G.H. and Chung, H.Y. (1980) Drug Resistant R
+
Bacteria in Eel-culture Pond [in
Chinese]. Reports on Fish Disease Research, III, CAPD Fisheries Series No. 3,
Republic of China, 8 pp.
Kozaki, S., Kurokawa, A., Asao, T., Kato, K., Uemura, T. and Sakaguchi, G. (1987)
Enzyme-linked immunosorbent assay for Aeromonas hydrophila hemolysins.
FEMS Microbiology Letters 41, 147151.
Kozaki, S., Asao, T., Kamata, Y. and Sakaguchi, G. (1989) Characterization of
Aeromonas sobria hemolysin by use of monoclonal antibodies against Aeromonas
hydrophila hemolysin. Journal of Clinical Microbiology 27, 17821786.
Krovacek, K., Faris, A., Ahne, W. and Mnsson, I. (1987) Adhesion of Aeromonas
hydrophila and Vibrio anguillarum to fish cells and to mucus-coated glass slides.
FEMS Microbiology Letters 42, 8589.
Lallier, R., Bernard, F. and Lalonde, G. (1984) Difference in the extracellular products of
two strains of Aeromonas hydrophila virulent and weakly virulent for fish.
Canadian Journal of Microbiology 30, 900904.
Lamers, C.H.J., de Haas, M.J.H. and van Muiswinkel, W.B. (1985) The reaction of the
immune systen of fish to vaccination: development of immunological memory in
carp, Cyprinus carpio L., following direct immersion in Aeromonas hydrophila
bacterin. Journal of Fish Diseases 8, 256262.
Leblanc, D., Mittal, K.R., Oliver, G. and Lallier, R. (1981) Serogrouping of motile
Aeromonas species isolated from healthy and moribund fish. Applied and
Environmental Microbiology 42, 5660.
Lee, K.K. and Ellis, A.E. (1990) Glycerophospholipid : cholesterol acyltransferase
complexed with lipopolysaccharide (LPS) stabilizes and enhances toxicity of the
enzyme. Journal of Bacteriology 172, 53825393.
Leung, K.Y. and Stevenson, R.M.W. (1988) Tn5-induced protease-deficient strains of
Aeromonas hydrophila with reduced virulence for fish. Infection and Immunity 56,
26392644.
Levy, S.B. (1988) Tetracycline resistance determinants are widespread. ASM News 54,
418421.
Lewis, D.H. and Allison, T.C. (1971) An immunofluorescent technique for detecting
Aeromonas liquefaciens in fish utilized in lunar exposure studies. Transactions of
the American Fisheries Society 100, 575578.
Lewis, D.H. and Savage, N.L. (1972) Detection of antibody to Aeromonas liquefaciens
in fish by indirect fluorescent antibody technique. Journal of Fisheries Research
Board of Canada 29, 211212.
Loewy, A.G., Santer, U.V., Wieczorek, M., Blodgett, J.K., Jones, S.W. and Cheronis, J.C.
(1993) Purification and characterization of a novel zinc-proteinase from cultures of
Aeromonas hydrophila. Journal of Biological Chemistry 268, 90719078.
Lucchini, G.M. and Altwegg, M. (1992) rRNA gene restriction patterns as taxonomic
tools for the genus Aeromonas. International Journal of Systematic Bacteriology
42, 384389.
McCoy, R.H. and Seidler, R.J. (1973) Potential pathogens in the environment: isolation,
enumeration, and identification of seven genera of intestinal bacteria associated
with small green pet turtles. Applied Microbiology 25, 534538.
MacInnes, J.I., Trust, T.J. and Crosa, J.H. (1979) Deoxyribonucleic acid relationships
among members of the genus Aeromonas. Canadian Journal of Microbiology 25,
579586.
450
T. Aoki
Martinez-Murcia, A.J., Esteve, C., Garay, E. and Collins, M.D. (1992a) Aeromonas
allosaccharophila sp. nov., a new mesophilic member of the genus Aeromonas.
FEMS Microbiology Letters 91, 199206.
Martinez-Murcia, A.J., Benlloch, S. and Collins, M.D. (1992b) Phylogenetic
interrelationships of members of the genera Aeromonas and Plesiomonas as
determined by 16S ribosomal DNA sequencing: lack of congruence with results of
DNADNA hybridization. International Journal of Systematic Bacteriology 42,
412421.
Maruvada, R., Das, P., Ghosh, A.N., Pal, S.C. and Nair, G.B. (1992) Electrophoretic
mobility and immunoblot analysis of the outer membrane proteins of Aeromonas
hydrophila, A. sobria and A. caviae. Microbios 71, 105113.
Merino, S., Camprubi, S. and Toms, J.M. (1990) Isolated and partial characterization of
bacteriophages PM2 from Aeromonas hydrophila. FEMS Microbiology Letters 68,
239244.
Merino, S., Camprubi, S. and Toms, J.M. (1993) Detection of Aeromonas hydrophila in
food with an enzyme-linked immunosorbent assay. Journal of Applied Bacteriology
74, 149154.
Merino, S., Rubires, X., Aguilar, A. and Toms, J.M. (1996) The O:34-antigen
lipopolysaccharide as an adhesin in Aeromonas hydrophila. FEMS Microbiology
Letters 139, 97101.
Millership, S.E. (1996) Identification. In: Austin, B., Altwegg, M., Gosling, P.J. and
Joseph, P. (eds) The Genus Aeromonas. John Wiley and Sons, Chichester, pp.
84107.
Millership, S.E. and Want, S.V. (1993) Characterization of strains of Aeromonas spp. by
phenotype and whole-cell protein fingerprint. Journal of Medical Microbiology 39,
107113.
Miyata, M., Aoki, T., Inglis, V., Yoshida, T. and Endo, M. (1995) RAPD analysis of
Aeromonas salmonicida and Aeromonas hydrophila. Journal of Applied
Bacteriology 79, 181185.
Miyata, M., Inglis, V. and Aoki, T. (1996) Rapid identification of Aeromonas
salmonicida subspecies salmonicida by the polymerase chain reaction. Aquaculture
141, 1324.
Miyazaki, T. and Jo, Y. (1985) A histopathological study of motile aeromonad disease in
ayu. Fish Pathology 20, 5559.
Miyazaki, T. and Kaige N. (1985) A histopathological study on motile aeromonad
disease of crucian carp. Fish Pathology 21, 181185.
Moyer, N.P. (1996) Isolation and enumeration of aeromonads. In: Austin, B., Altwegg,
M., Gosling, P.J. and Joseph, P. (eds) The Genus Aeromonas. John Wiley and Sons,
Chichester, pp. 3984.
Mulla, R. and Millership, S. (1993) Typing of Aeromonas spp. by numerical analysis of
immunoblotted SDS-PAGE gels. Journal of Medical Microbiology 39, 325333.
Mullis, K.B. and Faloona, F.A. (1987) Specific synthesis of DNA in vitro via a
polymerase-catalyzed chain reaction. In: Wu, R. (ed.) Methods in Enzymology, Vol.
155. Academic Press, San Diego, pp. 335350.
Neves, M.S., Nunes, M.P. and Milhomen, A.M. (1994) Aeromonas species exhibit
aggregative adherence to HEp-2 cells. Journal of Clinical Microbiology 32,
11301131.
Nieto, T.P. and Ellis, A.E. (1986) Characterization of extracellular metallo- and serine-
proteases of Aeromonas hydrophila (strain B
51
). Journal of General Microbiology
132, 19751979.
Nieto, T.P. and Ellis, A.E. (1991) Heterogeneity of extracellular proteases produced by
451 Motile Aeromonads
different isolates of Aeromonas hydrophila and A. sobria pathogenic for fish.
Journal of Fish Diseases 14, 229235.
Noterdaeme, L., Bigawa, S., Willems, K.A. and Ollievier, F. (1991) Biochemical and
physiological characteristics and plasmid profiles of Aeromons hydrophila strains,
isolated from freshwater fish and from fresh water. Journal of Fish Diseases 14,
313321.
Okamoto, A. (1992) Restrictions on the use of drugs in aquaculture in Japan. In:
Chemotherapy in Aquaculture: From Theory to Reality. Office International des
Epizooties, Paris, pp. 109114.
Palumbo, S.A. (1996) The Aeromonas hydrophila group in food. In: Austin, B., Altwegg,
M., Gosling, P.J. and Joseph, P. (eds) The Genus Aeromonas. John Wiley and Sons,
Chichester, pp. 287310.
Palumbo, S.A., Maxino, F., Williams, A.C., Buchanan, R.L. and Thayer, D.W. (1985)
Starchampicillin agar for the quantitative detection of Aeromonas hydrophila.
Applied and Environmental Microbiology 50, 10271030.
Parker, M.W., Buckley, J.T., Postma, J.P.M., Tucker, A.D., Leonard, K., Pattus, F. and
Tsernoglou, D. (1994) Structure of the Aeromonas toxin proaerolysin in its water-
soluble and membrane-channel states. Nature 367, 292295.
Popoff, M. (1984) Genus III. Aeromonas Kluyver and van Niel 1936, 398
AL
. In: Krieg,
N.R. (ed.) Bergeys Manual of Systematic Bacteriology, Vol. 1. Williams and
Wilkins, Baltimore, pp. 545548.
Popoff, M. and Vron, M. (1976) A taxonomic study of the Aeromonas hydrophila
Aeromonas punctata group. Journal of General Microbiology 94, 1122.
Popoff, M.Y., Coynault, C., Kiredjian, M. and Lemelin, M. (1981) Polynucleotide
sequence relatedness among motile Aeromonas species. Current Microbiology 5,
109114.
Quinn, D.M., Wong, C.Y.F., Atkinson, H.M. and Flower, R.L.P. (1993) Isolation of
carbohydrate-reactive outer membrane proteins of Aeromonas hydrophila. Infection
and Immunity 61, 371377.
Quinn, D.M., Atkinson, H.M., Bretag, A.H., Tester, M., Trust, T.J., Wong, C.Y.F. and
Flower, R.L.P. (1994) Carbohydrate-reactive, pore-forming outer membrane
proteins of Aeromonas hydrophila. Infection and Immunity 62, 40544058.
Rader, A.E. and Murphy, J.R. (1988) Nucleotide sequences and comparison of the
hemolysin determinants of Vibrio cholerae EI Tor RV79 (Hly
+
) and RV79 (Hly

)
and classical 569B (Hly

). Infection and Immunity 56, 14141419.


Rippey, S.R. and Cabelli, V.J. (1979) Membrane filter procedure for enumeration of
Aeromonas hydrophila in fresh waters. Applied and Environmental Microbiology
38, 108113.
Rivero, O., Anguita, J., Paniagua, C. and Naharro, G. (1990) Molecular cloning and
characterization of an extracellular protease gene from Aeromonas hydrophila.
Journal of Bacteriology 172, 39053908.
Rodriguez, L.A., Ellis, A.E. and Nieto, T.P. (1992) Purification and characterisation of
an extracellular metalloprotease, serine protease and haemolysin of Aeromonas
hydrophila strain B
32
: all are lethal for fish. Microbial Pathogenesis 13, 1724.
Rodriguez, L.A., Fernandez, A.I.G. and Nieto, T.P. (1993) Production of the lethal
acetylcholinesterase toxin by different Aeromonas hydrophila strains. Journal of
Fish Diseases 16, 7378.
Rose, J.M., Houston, C.W., Coppenhaver, D.H., Dixon, J.D. and Kurosky, A. (1989)
Purification and chemical characterization of a cholera toxin-cross-reactive
cytolytic enterotoxin produced by a human isolate of Aeromonas hydrophila.
Infection and Immunity 57, 11651169.
452
T. Aoki
Ruangpan, L., Kitao, T. and Yoshida, T. (1986) Protective efficacy of Aeromonas
hydrophila vaccines in the Nile tilapia. Veterinary Immunology and
Immunopathology 12, 345350.
Ruimy, R., Breittmayer, V., Elbaze, P., Lafay, B., Boussemart, O., Gautheir, M. and
Christen, R. (1994) Phylogenetic analysis and assessment of the genera Vibrio,
Photobacterium, Aeromonas, and Plesiomonas deduced from small-subunit rRNA
sequences. International Journal of Systematic Bacteriology 44, 416426.
Saitanu, K. (1986) Aeromonas hydrophila infections in Thailand. In: Maclean, J.L.,
Dizon, L.B. and Hosillos, L.V. (eds) The First Asian Fisheries Forum. Asian
Fisheries Society, Manila, Philippines. pp. 231234.
Sakazaki, R. and Shimada, T. (1984) O-serogrouping scheme for mesophilic Aeromonas
strains. Japanese Journal of Medical Science and Biology 37, 247255.
Santos, Y., Toranzo, A.E., Barja, J.L., Nieto, T.P. and Villa, T.G. (1988) Virulence
properties and enterotoxin of Aeromonas strains isolated from fish. Infection and
Immunity 56, 32853293.
Schperclaus, W. (1930) Pseudomonas punctata als Krankheitserreger bei Fischen.
Zeitung fr Fischerei 28, 289370.
Schperclaus, W., Kulow, H. and Schreckenbach, K. (1992) Infectious abdominal
dropsy. In: Schperclaus, W. (ed.) Fish Diseases, Vol. 1. Akademie-Verlag, Berlin,
pp. 401458.
Seidler, R.J., Allen, D.A., Lockman, H., Colwell, R.R., Joseph, S.W. and Daily, O.P.
(1980) Isolation, enumeration, and characterization of Aeromonas from polluted
waters encountered in diving operations. Applied and Environmental Microbiology
39, 10101018.
Shaw, D.H. and Hodder, H.J. (1978) Lipopolysaccharides of the motile aeromonads:
core oligosaccharide analysis as an aid to taxonomic classification. Canadian
Journal of Microbiology 24, 864868.
Shimizu, T. (1968) Studies on pathogenic properties of Aeromonas liquefaciens IV.
Necrotic factor for eel and guinea pig. Bulletin of the Japanese Society of Scientific
Fisheries 35, 613618.
Shotts, E.B. and Rimler, R. (1973) Medium for the isolation of Aeromonas hydrophila.
Applied Microbiology 26, 550553.
Shotts, E.B., Jr, Vanderwork, V.L. and Campbell, L.M. (1976) Occurrence of R factors
associated with Aeromonas hydrophila isolates from aquarium fish and waters.
Journal of the Fisheries Research Board of Canada 33, 736740.
Son, R., Rusul, G., Sahilah, A.M., Zainuri, A., Raha, A.R. and Salmah, I. (1997)
Antibiotic resistance and plasmid profile of Aeromonas hydrophila isolates from
cultured fish, tilapia (Tilapia mossambica). Letters in Applied Microbiology 24,
479482.
Song, Y.-L., Chen, S.-N. and Kou, G.-H. (1976) Agglutinating antibodies production and
protection in eel (Anguilla japonica) inoculated with Aeromonas hydrophila (A.
liquefaciens) antigens. Journal of the Fisheries Society of Taiwan 4, 2529.
Stevenson, R.M.W. (1988) Vaccination against Aeromonas hydrophila. In: Ellis, A.E.
(ed.) Fish Vaccine. Academic Press, London, pp. 112123.
Sugita, H., Nakamura, T., Tanaka, K. and Deguchi, Y. (1994) Identification of
Aeromonas species isolated from freshwater fish with the microplate hybridization
method. Applied and Environmental Microbiology 60, 30363038.
Talon, D., Dupont, M.J., Lesne, J., Thouverez, M. and Michel-Briand, Y. (1996) Pulsed-
field gel electrophoresis as an epidemiological tool for clonal identification of
Aeromonas hydrophila. Journal of Applied Bacteriology 80, 277282.
Telford, J.R., Leary, J.A., Tunstad, L.M.G., Byers, B.R. and Raymond, K.N. (1994)
453 Motile Aeromonads
Amonabactin: characterization of a series of siderophores from Aeromonas
hydrophila. Journal of American Chemical Society 116, 44994500.
Thornton, J., Howard, S.P. and Buckley, J.T. (1988) Molecular cloning of a
phospholipidcholesterol acyltransferase from Aeromonas hydrophila: sequence
homologies with lecithincholesterol acyltransferase and other lipases. Biochemica
et Biophysica Acta 959, 153159.
Thune, R.L. and Plumb, J.A. (1982) Effect of delivery method and antigen preparation
on the production of antibodies against Aeromonas hydrophila in channel catfish.
Progressive Fish Culturist 44, 5354.
Thune, R.L., Graham, T.E., Riddle, L.M. and Amborski, R.L. (1982) Extracellular
proteases from Aeromonas hydrophila: partial purification and effects on age-0
channel catfish. Transactions of the American Fisheries Society 111, 749754.
Toranzo, A.E., Barja, J.L. Colwell, R.R. and Hetrick, F.M. (1983) Characterization of
plasmids in bacterial fish pathogens. Infection and Immunity 39, 184192.
Toranzo, A.E., Santos, Y., Nieto, T.P. and Barja, J.L. (1986) Evaluation of different assay
systems for identification of environmental Aeromonas strains. Applied and
Environmental Microbiology 51, 652656.
Unger, B., Klock, G. and Hillen, W. (1984) Nucleotide sequence of the repressor gene of
the RA1 tetracycline resistance determinant: structural and functional comparison
with three related Tet repressor genes. Nucleic Acids Research 12, 76937703.
Van der Kooij, D. and Hijnen, W.A.M. (1988) Nutritional versatility and growth kinetics
of an Aeromonas hydrophila strain isolated from drinking water. Applied and
Environmental Microbiology 54, 28422851.
Varela, M.F. and Griffith, J.K. (1993) Nucleotide and deduced protein sequences of the
class D tetracycline resistance determinant: relationship to other antimicrobial
transport proteins. Antimicrobial Agents and Chemotherapy 37, 12531258.
Wakabayashi, H., Kanai, K., Hsu, T.-C. and Egusa, S. (1981) Pathogenic activities of
Aeromonas hydrophila biovar. hydrophila (Chester) Popoff and Veron, 1976 to
fishes. Fish Pathology 15, 319325.
Wakabongo, M., Bortey, E., Meier, F.A. and Dalton, H.P. (1992) Rapid identification of
motile Aeromonas. Diagnostic Microbiology and Infectious Disease 15, 511515.
Wilcox, M.H., Cook, A.M., Thickett, K.J., Eley, A. and Spencer, R.C. (1992) Phenotypic
methods for speciating clinical Aeromonas isolates. Journal of Clinical Pathology
45, 10791083.
Wolf, K. (1988) Fish Viruses and Fish Viral Diseases. Cornell University of Press,
Ithaca, New York.
Wong, K.R. and Buckley, J.T. (1993) Aeromonas spp. can secrete Escherichia coli
alkaline phosphatase into the culture supernatant, and its release requires a
functional general secretion pathway. Molecular Microbiology 9, 955963.
Yadav, M., Indira, G. and Ansary, A. (1992) Cytotoxin elaboration by Aeromonas
hydrophila isolated from fish with epizootic ulcerative syndrome. Journal of Fish
Diseases 15, 183189.
Zhao, J. and Aoki, T. (1992a) A specific DNA hybridization probe for detection of
Pasteurella piscicida. Diseases of Aquatic Organisms 7, 203210.
Zhao, J. and Aoki, T. (1992b) Nucleotide sequence analysis of the class G tetracycline
resistance determinant from Vibrio anguillarum. Microbiology and Immunology 36,
10511060.