Viral diseases 1

3
CHAPTER THREE Bacterial diseases
Eleonor V. Alapide-Tendencia and Leobert D. de la Peña

Diseases caused by bacteria may cause heavy mortality in both wild and cul-
tured fish and crustaceans. Bacteria are found everywhere in the aquatic envi-
ronment. Most bacterial disease agents are part of the normal microflora of the
marine environment and are generally considered as secondary or opportunis-
tic pathogens. Almost all fish bacterial pathogens are capable of independent
existence outside the fish. There are only a few obligatory pathogens. Even
these, however, are capable of living for a long time in the tissues of their host
without causing injury. Clinical infections and disease usually occur only after
the onset of some major changes in the physiology or body of the host. Thus, to
understand bacterial diseases of fish, one must understand the relationship of
bacteria with their host and with their environment.
As in all animal production systems, bacterial disease is one of the major prob-
lems facing production, development and expansion of the aquaculture indus-
try. The control of disease is particularly difficult because fish are often farmed
in systems where production is dependent on natural environmental condi-
tions. Changes or deterioration in the aquatic environment cause most of the
bacterial diseases encountered, and environmental effects give rise to many
other adverse conditions. A second major constraint on disease control is the
relatively limited number of therapeutic agents available for the control of bac-
terial disease agents. Even recommended therapies and preventive measures
pose limitations. Their application to aquatic animals is often difficult in actual
practice, and sometimes impossible to implement.
Outbreaks of major bacterial diseases in aquaculture can be significantly re- duced if
proper attention is paid to good husbandry practices and the mainte- nance of
optimum environmental conditions, especially water quality. Another important
consideration involves identifying the predisposing factors that may lead to a
disease state. Once predisposing factors are identified, appropriate corrective
measures should be initiated in the culture system.
Most bacterial disease show similar signs, especially in fishes. Bacterial infec-
tion may appear on the skin or fins of fish, exoskeleton or appendages of crus-
taceans, in the muscles and in the internal organs. In nearly all cases, red spots,
brown or black spots, or necrotic tissues can be observed. Inflammation may
also occur. Proper identification of the causative agent is important to ensure
successful treatment.
WHAT ARE BACTERIA?

Bacteria are not visible to the naked eye. These microorganisms are of very
small dimensions, usually between 0.5 and 10 microns (µm). But, when bacte-
a b
ria multiply in great numbers on a solid medium, they form visible colonies
representing millions or billions of individual cells. The cells can be seen only
under a microscope from a smear stained with a dye on a microscope slide.
Bacteria differ from other cells in that they are prokaryotic (lacking a nuclear
c membrane). The nucleus occupies the center of the cell. All its genetic material
is linked in a single chromosome. The cytoplasm is densely packed with RNA
and is finely granular because of the presence of ribosome. The nucleus/ cyto-
plasm complex is packaged in a complex envelope or integument. Its inner-
most layer is the thin cytoplasmic membrane (plasmalemma). Outside the
membrane is a rigid cell wall. Some bacterial pathogens develop a capsule out-
d e side of the cell wall, which is usually associated with the virulence or infective
ability of the organism. Many of the pathogenic bacteria are flagellated and a
few have no flagella for locomotion. Some move by body flexing or gliding.
Some bacteria produce enzymes called extracellular products or ECP, which
are associated with the microorganism’s virulence. Extracellular products are
highly toxic to fish and crustaceans and they contain proteases, hemolysins,
exohemagglutinins and cytotoxins.
Figure 3-1. The different shapes and ar-
rangements of bacteria: (a) cocci in Bacteria reproduce asexually by binary fission. That is, they multiply by an
streptococcal arrangement, (b) cocci in elongation of the cell followed by a division.
staphylococcal arrangement, (c) bacilli,
(d) several kinds of spirilla, and (e) The most common method used to detect the presence of bacteria is by gram
comma-shaped vibrios stain. The gram stain classifies bacteria into two groups: the gram positive and
the gram negative. Gram-positive bacteria are those that possess a thick pepti-
doglycan cell wall which will retain the initial crystal violet stain during wash-
ing with 95% alcohol. Gram-negative bacteria are those that possess a uni-
molecular peptidoglycan cell wall bounded on one side by the cytoplasmic
membrane and on the other side by the outer membrane; such cells are decol-
orized by 95% alcohol and take up the secondary stain. To identify bacterium,
a pure culture should be obtained, containing a single species and not a mix-
ture of different kinds of bacteria.
In classifying bacteria, one needs to pay attention to the cell shape. There are
three distinct cell forms: cocci, baccili and spiral (Fig.3-1). Cocci are spherical
cells and exist in several patterns or groupings which are specific to the genus.
Paired cocci are called diplococcus, while those in long chains are called strep-
tococcus. Irregularly grouped cocci are called staphylococcus. Longer and cy-
lindrical bacteria are known as bacilli or rods. Cells that are between the coc-
cus and the bacillus in shape are called coccobacilli. The short, curved rods are
the vibrios. When more than one curvature is observed, it is called spirilla.
Most bacteria that cause disease in fish and crustaceans are rod-shaped. Figure
3-1 shows the different shapes and arrangements of bacteria.
The shape, size and color of a given colony are also important in identification. The
bacterial colony surface texture, whether rough, smooth or mucoid, should

26 Health Management in Aquaculture

 also be observed. The same species can form rough or smooth colonies, de-
pending on environmental conditions and the virulence or infective ability of
the strain. Smoother colonies are often more virulent or harmful. Considering
the large number of bacteria known to exist, gram stain reaction and morpho-
logical form are not enough to identify bacterial species. To identify a bacte-
rium, the different physiological or bodily and biochemical properties it pos-
sesses must first be known through a series of tests.

IDENTIFYING THE REAL CAUSE OF A DISEASE; KOCH’S POSTULATES

Not all bacteria present in the body of a fish or crustacean are pathogenic or
may cause disease. Some bacteria may be harmless or even beneficial. By care-
fully noting the observations suggested here, one might be able to tell if the
isolated bacterium suspected of causing the disease is the causative organism.
Koch, a German physician and bacteriologist, enunciated the following criteria
in 1891 to establish unequivocally a causal relationship between an organism
and a specific disease.
• The organism should be found in all cases of the disease in question, and its
distribution in the body should be in accordance with the lesions observed.
• The organism should be cultivated outside the body of the host, in pure
culture, for several generations.
• The organism so isolated should reproduce the disease when introduced
into other susceptible animals.
• The organism must be reisolated from the experimentally-infected animal.

Virulence Determinants To produce disease, microorganisms must be able to:

• Enter the host;
• Multiply under the physical and chemical conditions of the host tissues;
• Interfere with the action of humoral (in body fluids) and cellular defense
mechanisms of the host; and
• Damage tissues thereby producing the unpleasant and possibly lethal ef-
fects.
Bacteria are ubiquitous. This means that they can be found or are present al-
most everywhere in the aquatic environment. The actual role of these microor-
ganisms may vary from being beneficial to that of being a secondary opportu-
nistic invader, attacking only when the host is weakened or injured, or a pri-
mary pathogen that may cause the death of the species.
In this chapter, the important bacterial diseases of fish and crustaceans are discussed
as well as the different diagnostic methods, gross signs and preven- tive measures
and treatments of these diseases.

Bacterial diseases 27
IMPORTANT BACTERIAL DISEASES OF FISH

Columnaris Disease CAUSATIVE AGENT:

Flavobacteriu m colu mnare (previously named as Flexibacter colu mnaris)
SPECIES AFFECTED:
Ayu (Plecoglossus altivelis) , tilapia (Oreochromis niloticus), carp (Cyprinus
sp .), channel catfish (Ictalurus punctatus) , goldfish (Carassius auratus), rohu
(Labeo rohita)
GROSS SIGNS:
The first indication of infection is generally the appearance of a white spot on
some part of the head, gills, fin or body. A zone with a distinct reddish tinge
usually surrounds this. The lesions are circular as if spreading from a single
focus towards all directions at the same rate. On the gills, the lesions are more
necrotic. On the skin, they develop into hemorrhagic ulcers, with an overlying
seroma of bacterial cell and necrotic tissue. Histologically, there is epidermal
spongiosis, necrosis, and ulcerations.
EFFECTS ON HOST:
F. colu mnare is an opportunistic pathogen widely distributed in the water. The
disease does not usually occur as spontaneous infection but results from inju-
ries to the fish, or physical and nutritional deficiencies. Outbreaks are affected
by factors such as temperature and stress. Dissolved cations such as sodium,
potassium, calcium and magnesium enhance their infectivity. F. columnare at-
tacks fish primarily through the gills or abraded epidermal areas. Once the
pathogen is established, proteolytic enzymes break down the skin and muscle
to open necrotic lesions. The bacterium appear systematically after extensive
tissue necrosis. Gill lesions may cause respiratory difficulty and the fish eventu-
ally dies. Body lesions are subject to secondary infection by other microorgan-
isms. Fish that survive the infection may become carriers of the disease.
DIAGNOSIS:
Columnaris disease can be presumptively diagnosed from disease signs on the
skin and gills of the host and from squash preparations made from scrapings of
the affected areas. In wet mount preparations of infected tissues, the bacteria
show a slow gliding movement, gathering into characteristic column-like
masses that give the disease its name. Microscopic examination of lesions
shows the presence of long, slender, possibly filamentous, rod-shaped, gram-
negative bacteria. The growth of F. colu mnare on solid media is usually charac-
terized by yellow-green, flat and rough spreading colonies that adhere to the
media. F. columnare can also be detected from fish and water using indirect
fluorescent antibody technique (IFAT).

28 Health Management in Aquaculture

 PREVENTION AND CONTROL:

• vaccination
• environmental manipulation like lowering water temperature
• addition of competitive bacteria like Citrobacter fecundii and Aeromonas
hydrophila
• copper sulfate dip at 40 mg/L for 20 min or 500 mg/L for 1 min
• oxolinic acid dip at 1 mg/L for 24 h
• sulphamerazine and oxytetracycline at 220 mg/ kg/ day for 10 days fol-
lowed by 50 to 75 mg/ kg/ day for 10 days.

Edwardsiella Septicaemia or CAUSATIVE AGENT:

Edwardsiellosis
Edwardsiella tarda
SPECIES AFFECTED:
Tilapia (Oreochromis niloticus), channel catfish (Ictalurus punctatus) , mullet
(Mugil cephalus) , carp (Cyprinus carpio)
GROSS SIGNS:
Edwardsiella tarda infection manifests itself by the presence of small, 3-5 mm
cutaneous or skin lesions located dorso-laterally (from along the
back to the side) on the body. These lesions progress into abscesses
and develop obvious convex swollen areas. The skin loses pigmen-
tation. A foul smelling gas is emitted when the skin is incised. The
lesion contains large amounts of necrotized or dead tissue. Inter-
nally, there is generalized hyperemia (Fig. 3-2) and enlargement of
the liver and kidney. Histologically, the lesion is characterized by
focal necrosis, often extending from muscle, haemopoietic tissue
and liver parenchyma (Fig. 3-3) to perforate the abdominal wall.
EFFECTS ON HOST:
E. tarda infection usually occurs during the warm, summer
Figure 3-2. Oreochromis niloticus with Edward- months. The source of E. tarda is presumably the intestinal con-
siella tarda infection showing white nodules on tents of carrier animals such as snakes, fish (eel and catfish), and
theliver
some amphibians and reptiles. High temperature, poor water qual-
ity and crowding may contribute to the onset and severity of the
disease. Affected fish lose mobility of the caudal or tail portion of
the body. E. tarda infection may cause lesions in the dermis, mus-
culature and visceral organs of the host. Skin lesions when incised
emit a foul smelling gas. The lesion contains large amount of necro-
tized or dead tissue.
DIAGNOSIS:
The bacterium is easily isolated from muscle and internal organs of
clinically diseased fish on most general-purpose media such as
Figure 3-3. Focal necrosis in the liver of O. niloticus
brain heart infusion agar (BHIA) and tryptic soy agar (TSA). Small
infected with Edwardsiella tarda . (Hematoxylin and punctate colonies develop in 24-48 h on inoculated media.
Eosin stain, 25X)

Bacterial diseases 29
 PREVENTION AND CONTROL :

• Improve water quality.

• Reduce stocking density.
• Apply oxytetracycline at 55 mg/ kg fish for 10 days.

Vibriosis CAUSATIVE AGENT:

Vibrio alginolyticus, V. anguillarum and V. vulnificus
SPECIES AFFECTED:
Grouper (Epinephelus sp.) , rabbitfish (Siganus sp.), milkfish (Chanos
chanos) , seabass (Lates calcarifer) , sea bream (Sparus aurata)
GROSS SIGNS:
The first signs of the disease are usually anorexia or loss of appetite,
with darkening either of the whole fish or of particular areas of the
Figure 3-4. Vibrio infected grouper with hemor-
rhagic and necrotic fins. Hemorrhagic lesionso n dorsum or back. Other common signs of vibriosis are hemorrhagic
the dorsal body part can also be observed spot on different parts of the body including necrotic fins (Fig. 3-4),
eye opacity (Fig. 3-5) and exophthalmia (Fig. 3-6). The perachute
condition results in death without any other clinical signs except oc-
casional periorbital or abdominal oedema. Chronically infected fish
generally exhibit very pale gills and large granulating lesions deep in
the muscle (Fig. 3-7a; 3-7b). In hatcheries, the presence of red spots
in tanks is a sign of Vibrio infection.
EFFECTS ON HOSTS:
Vibriosis usually occurs in the warm summer months, especially
when the stocking densities are high, and the salinities and organic
Figure 3-5. Eye opacity in milkfish fingerlings
infected with Vibrio parahaemolyticus loads are also high. Stressed fish are more prone to Vibrio infection.
When an outbreak occurs, mortalities of 50% or higher can be ob-
served in young fish. In older fish, losses may be lower, but infected
fish do not feed or grow. When harvested, fish may have large ne-
crotic lesions in the middle of the muscle mass.
DIAGNOSIS:
Figure 3-6 Bilateral exophthalmia in fish with Squash preparations of kidney, liver, spleen, necrotic muscle tissue
bacterial infection and other organs reveal the bacterium. The pathogen can usually be
isolated from infected organs in pure culture using standard bacterio-
logical media, such as BHIA, nutrient agar (NA) and TSA, provided
they contain 1-2% sodium chloride. Thiosulphate citrate bile salt
agar (TCBS) is a medium that selectively promotes growth of patho-
genic vibrios while inhibiting other bacteria.
PREVENTION AND CONTROL:
• Maintain good water quality, good husbandry procedures and
lower stocking densities.
• Apply oxytetracycline at 77 mg/ kg of fish or nitrofurazone at 56
mg/ kg of fish for 10 days.
Figure 3-7a. Dermo-muscular lesion in mullet • Vaccinate.
with vibrio infection

30 Health Management in Aquaculture

Figure 3-7b. Hyperemia and blood clot
in the abdominal cavity of mullet wtih
vibrio infection

Motile Aeromonad Septicemia CAUSATIVE AGENT:

Aeromonas hydrophila, A. caviae, and A. sobria.
Species affected:
Tilapia (Oreochromis niloticus), milkfish (Chanos chanos) , goldfish (Carassius
auratus) , catfish (Clarias batrachus) , snakehead (Ophicephalus striatus) , goby
(Glossogobius guirus), climbing perch (Anabas testudineus) , gourami (Tricho-
gaster sp.), mullet (Mugil cephalus)
GROSS SIGNS:
External signs of motile aeromonad disease vary from darkening in color, en-
largement of the abdominal area (Fig. 3-8a) to an extensive superficial redden-
ing of a large area of the body (Fig. 3-9), often with necrosis of fins or tail and
extensive ulceration over a considerable portion of the flanks or dorsum. The
ulcers are usually shallow and the surface may go brown as it necrotizes or
decays (Fig. 3-10). Other disease signs are scale loss, mouth sores,

Figure 3-8a. Oreochromis

niloticus with Aeromonas
hydrophila infection. Note
the enlarged abdominal cav-
ity

Figure 3-8b (far right). The

same O. niloticus with hype-
remia and enlarged abdomi-
nal cavity due to A. hydro-
phila

Figure 3-9. Hemorrhagic le-

sion in milkfish infected
with Aeromonas hydrophila

Figure 3-10 (far right).

Dermo-muscular lesion in
catfish experimentally
in- fected with A.
hydrophila obtained from
EUS fish

Bacterial diseases 31
 exophthalmia, and eye opacity. Internally, there may be dropsy (Fig. 3-8b),
hyperemia, and the congestion of the internal organs.
EFFECTS ON HOST:
The organisms are usually transmitted through the mouth but may also enter
through the skin or gill abrasions. The organisms multiply in the intestine or at
the site of invasion and are spread throughout the body by the bloodstream.
Internally, there may be ascitic fluid, anemia and damaged internal organs
which may lead to mortalities. Mortality as high as 80% may occur among
physically stressed, nutritionally deficient, anoxious or injured young fish.
Older fishes are less susceptible to motile aeromonads, although 20 to 35%
mortalities are not common.
DETECTION AND CULTIVATION:
Squash preparation of the kidney is useful when searching for the etiological
agent of the disease. The organisms appear as rod-shaped bacteria, a few are
coccoids or short rods in form, usually in single or pairs but rarely in short
chains or filaments. They grow well on most common laboratory media such
as BHIA, TSA and NA.
PREVENTION AND CONTROL:
• Avoid overcrowding of fish in holding facilities.

Pseudomonad Septicemia or CAUSATIVE AGENT:

Red Spot Disease
Pseudomonas fluorescens, P. anguilliseptica, and P. chlororaphis.
SPECIES AFFECTED:
Milkfish (Chanos chanos) , goldfish (Carassius auratus), tilapia (Oreochromis
niloticus)
GROSS SIGNS:
The external disease signs of pseudomonad septicaemia are similar to those caused
by other gram-negative bacterial pathogen of fish. The disease causes small
hemorrhages in the skin around the mouth and opercula and along the ventral or
abdominal surfaces. The body surface may ooze blood and slime in severe cases but
there is no reddening of the fins and anus.
EFFECTS ON HOST:
Pseudomonas spp. enters the host either through the oral route or through bro-
ken or abraded skin and damaged gills. The organism is carried throughout the
fish body by the blood stream. The bacteria and their toxin destroy body tis-
sues, organs and functions. Dysfunctions of the different body organs may lead
to mortality of up to 70%.
DIAGNOSIS:
The organisms can usually be isolated from the kidney and other internal or-
gans of affected fish, as well as from the lesion. They grow well on most com-
mon laboratory media such as BHIA, TSA and NA. GSP agar or Pseudomonas-

32 Health Management in Aquaculture

 Aeromonas Selective Agar is a medium that selectively promotes growth of
Aeromonas and Pseudomonas, and inhibits growth of other bacteria.
PREVENTION AND CONTROL:
• Maintain proper stock management procedures, ensure water quality and
reduce stocking density.
• Transfer in a tank and raise temperature to 26-27°C and maintain for about
2 weeks.

Streptococcal Infection CAUSATIVE AGENT:

Streptococcus sp.
Species affected:
Seabass (Lates calcarifer) , tilapia (Oreochromis niloticus), rabbitfish (Siganus
guttatus) , ayu (Plecoglossus altivelis)
GROSS SIGNS:
Clinical signs vary among species of affected fish. However, erratic swimming,
darkening of body color, unilateral or bilateral exopthalmia, corneal opacity,
hemorrhages on the opercula and the bases of the fins and ulceration of body
surface are the most common clinical signs. The hemorrhagic lesions, which
gradually extend and ulcerate to release decayed material, are generally raised
and have a darkened zone around them. The lesions are more superficial than
in furunculosis or vibriosis.
EFFECTS ON HOST:
Infected fish have difficulty ventilating, and lose the ability to orient them-
selves in the water. The eye becomes opaque and necrotic, conditions that can
result to blindness. Fish swim in a spiralling motion. Fish are able to respond to
stimuli, but have little control over movements. The spleen and kidney become
enlarged. Dysfunction of the damaged internal organs may lead to mortalities.
DIAGNOSIS:
The pathogen grows easily on tryptic soy agar supplemented with 0.5% glu-
cose, brain heart infusion agar, Todd-Hewitt broth agar and horse agar. Colo-
nies develop after 24-48 h of incubation at 20-30°C. The colonies on agar plates
appear small (0.5-1mm diameter), yellowish, translucent, rounded and slightly
raised.
PREVENTION AND CONTROL:
• Avoid overcrowding, overfeeding and unnecessary handling or transport.
• Remove and slaughter promptly all moribund fish in ponds or net cages at
early stage of infection to prevent outbreak or reduce severity of disease.
• Apply erythromycin at 25-50mg/ kg body weight of fish for 4-7 days.

Bacterial diseases 33
 Mycobacteriosis or Piscine CAUSATIVE AGENT:
Tuberculosis
Mycobacterium marinum, M. fortuitum and M. chelonae
SPECIES AFFECTED:
Siamese fighting fish (Betta splendens)
GROSS SIGNS:
Piscine mycobacteriosis is a systemic, chronic, progressive disease presenting
various clinical features depending upon species and ecological conditions.
Listlessness, anorexia, emaciation, exopthalmia, skin discoloration and exter-
nal lesions ranging from scale loss to nodules, ulcers and fin necrosis are signs
of advancing infection. Gross internal pathology of mycobacteriosis show gray-
white lesions of various sizes in most organs and tissues, but the kidney and
liver are most often involved.
EFFECTS ON HOST:
Mycobacteriosis is a chronic progressive disease. The disease may take several
years to progress from the asymptomatic state to clinical illness. Initially the
pigment will fade, and the fish appear sluggish with loss of appetite. Skin ul-
cers will develop. Fin and tail rot and loss of scale may also be seen. Nodules
may form in the muscle and internal organs, which may lead to emaciation or
edema or peritonitis. Infection may spread to the skeleton, in which deformi-
ties become apparent. Mortalities will then be observed.
DIAGNOSIS:
Primary isolation of fish mycobacteria is best achieved using Ogawa and
Lowenstein-Jensen media. Subcultures develop at 28°C within 3-5 days on
these media. On Ogawa medium, the cultures appear creamy in the dark but
brilliant yellow color when exposed to light. Cultures may not always be ob-
tained even from fish showing unequivocal evidence of infection. Mycobacte-
ria may also be isolated occasionally on general-purpose bacteriological media
such as tryptic soy agar, or brain heart infusion agar, provided that a large
inoculum is used. All fish mycobacterium have been cultured at 20-30°C for 2
to 30 days. The isolates are strongly acid-fast, rod-shaped, weakly gram-posi-
tive, cord forming, non-motile and non-spore forming. Optimum temperature
for bacterial growth is between 15°C to 37°C, but the isolates grow best at
28°C.
PREVENTION AND CONTROL:
• Sanitation, disinfection, and destruction of carrier fishes are the primary
methods of controlling mycobacteriosis.
• Avoid feeding fish with contaminated fish products. Pasteurize food before
use.
• Apply chloramine B or T at 10 mg/ l for 24 h.

34 Health Management in Aquaculture

BACTERIAL DISEASES OF CRUSTACEANS

Bacterial infections of cultured crustaceans occur as: bacterial fouling of sur- faces,
cuticular or subcuticular localized infections, and internal or systemic infections.

Bacterial Fouling of Surfaces Filamentous Bacterial Disease

CAUSATIVE AGENT:
Leucothrix sp., Thiothrix sp., Flexibacter sp., Cytophaga sp., Flavobacteriu m sp.
SPECIES AFFECTED:
Penaeus monodon, P. merguiensis, P. indicus
GROSS SIGNS:
Presence of fine, colorless, thread-like growth on the body surface (Fig. 3-11)
and gills as seen under a microscope.
EFFECTS ON HOST:
Infected eggs show a thick mat of filaments on the surface, which may inter-
fere with respiration or hatching. In larvae and postlarvae, normal respiration,
feeding, locomotion, and molting may be seriously impaired, resulting in
slower growth rates, retarded development and eventually death. However,
larval shrimps are less prone to infestations due to rapid succession of molts
throughout the different larval stages. Frequent molting does not allow ad-
equate time for the bacteria to accumulate on the exoskeleton. In larger
shrimps, it may result in respiratory distress. Mortality is due to hypoxia. Dis-
ease onset is associated with high organic loads in culture water, low dis-
solved oxygen levels and added stress from molting. If left untreated in inten-
sive culture systems, accumulative mortality may reach 80% or more within a
few days to a few weeks of onset of disease signs.
DIAGNOSIS:
Direct microscopic examination of wet mounts of larvae or
postlarvae, appendages and gill filaments excised from juvenile or
adult shrimp, and of filamentous organisms attached to external
surfaces of the cuticle.
PREVENTION AND CONTROL:
• Maintain good water quality with optimum dissolved oxygen
and low organic matter levels.
• Apply Cutrine Plus at 0.15 ppm copper in 24 h flowthrough
Figure 3-11. Strands of filamentous bacterium
treatments
Leucothrix sp. on heavily infested gills of juvenile • Apply 0.5 ppm copper in 4 to 6-h static treatments for PL 2 and
Penaeus monodon. At left is the protozoan
Zoothamnium (fresh mount, 200x)
older.

Bacterial diseases 35
 Cuticular or Subcuticular Shell Disease, Brown/Black Spot, Black Rot/Erosion, Blisters, Necrosis of
Localized Infections Appendages
CAUSATIVE AGENT:
Shell-degrading bacteria belonging to Vibrio, Aeromonas, and Pseudomonas
groups
Species affected:
Penaeus monodon, P. merguiensis, P. indicus
GROSS SIGNS:
The disease manifests itself as brownish to black, single or multiple, eroded
areas on the general body cuticle (Fig. 3-12a, 3-12b), append- ages, and
gills.
In larval and postlarval stages, the affected appendage shows a ciga-
rette butt-like appearance (Fig. 3-13). Blister containing cyanotic ge-
latinous fluid may develop on the carapace and abdominal segment.
The blister may extend to the underside of the ventro-lateral section of
the carapace creating a bulge on the underside.
EFFECTS ON HOST:

Figure 3-12a. Shell disease on the carapace of Infection usually starts at sites of punctures or injuries caused by the
Penaeus monodon telson or rostrum, in cracks on the abdominal segment from sudden
flexure of the shrimp body, or from other damage caused by cannibal-
ism. Another infection site is the cuticle colonized by a large number
of bacteria. The bacteria produce extracellular lipases, proteases, and
chitinases, which together erode the multiple-layered cuticle, resulting
in the development of the disease. The progressive destruction of the
cuticle also provides a route of entry for secondary pathogens like
fungi or opportunistic bacteria. Such infections may become lethal
because of osmotic imbalances, molting problems, secondary fungal
infection and a generalized septicemia. The affected shrimp becomes
susceptible to cannibalism or dies from stress and energy exhaustion.
The disease is associated with trauma to the cuticle (e.g. heavy aera-
Figure 3-12b. Shell disease on the abdominal seg-
ment of Penaeus monodon tion), conditions that encourage a high number of bacteria in the cul-
ture water (e.g. poor hatchery hygiene, high organic loads or contami-
nated algae) and undefined nutritional and environmental stressors.
DIAGNOSIS:
Diseased penaeids are examined for appearance of multifocal mela-
nized cuticular lesions on the cuticle or the general body surface, the
appendages, or the gills. Diagnosis may also be made by bacteriologi-
cal (isolation, purification and identification) and serological (slide ag-
glutination) methods.
PREVENTION AND CONTROL:
Figure 3-13. Penaeus monodon post larvae with • Maintain good water quality and use nutritionally adequate diets.
necrotic pleopods. Necrotic area appears like ciga-
rette butt • Keep organic load of the water at low levels by removing sediments

36 Health Management in Aquaculture

 which harbor high numbers of bacteria.
• Minimize handling and overcrowding and reduce other forms of stress.
• Avoid injuries to the exoskeleton of the shrimps to prevent the development
of primary portals of entry.
• Induce molting, which eliminates the condition, but not when underlying
tissues are damaged.

Internal or Systemic Infections Luminous Bacterial Disease

CAUSATIVE AGENT:
Vibrio harveyi (Fig. 3-14) and V. splendidus
SPECIES AFFECTED:
Penaeus monodon, P. merguiensis, and P. indicus
STAGES AFFECTED:
Eggs, larvae, postlarvae, juveniles and adults
GROSS SIGNS:
Shrimps become weak and opaque-white. Affected shrimps often swim to the
pond surface and edges. Heavily infected shrimps in tanks and ponds show a
continuous greenish glow when observed in total darkness. When viewed un-
der the microscope, the hemocoel and internal tissues appear densely packed
with active bacteria.
EFFECTS ON HOST:
The hepatopancreas is the target organ of infection. Histopathology shows se-
vere inflammation in and around hepatopancreatic tubules of the entire organ.
In larger animals, melanized lesions are found in the proximal region of the
hepatopancreas. These lesions affect the digestive function of the organ as the
necrotic parts become non-functional. Total necrosis and dysfunction lead to
death, while partial dysfunction causes slow growth as not all tubules function
in digestion, absorption and storage. Systemic infections result in mortality of
up to 100%.
DIAGNOSIS:
The disease may be detected by bacteriological (isolation, purification and
identification); histological (demonstration of rod-shaped bacteria in tissues of
diseased shrimp); and serological [slide agglutination, fluorescent antibody
technique (FAT) and enzyme linked immunosorbent assay (ELISA) using spe- cific
antibodies] methods.
PREVENTION AND CONTROL:
• Disinfect incoming water and use filtration equipment to prevent entry of
luminous bacteria into the hatchery system.
Figure 3-14. Agar culture of the lumi- • Use only previously disinfected water during spawning and rearing.
nous bacteria Vibrio harveyi. Photo
taken in total darkness • Wash eggs.

Bacterial diseases 37
 • Siphon out sediments and debris from the tank bottom.
• Disinfect infected stock first before discarding.
• Wash and disinfect hatchery paraphernalia after each larval rearing period.
• Use microbially mature or aged seawater.
• Apply commercially available probiotics to maintain ecological balance
within the system.
• Use immunoprophylaxis or vaccination.
• Monitor bacterial population and diversity in the intake and rearing waters
of the shrimp pond.
• Apply commercially available probiotics.
• Use low salinity rearing water and reservoirs.
• Practice crop rotation.
• Install greenwater culture system and other system modifications.
• The disease may be prevented by rigorous water management.
• Apply antibiotics and other antibacterial substances only as the last resort.

Non-luminous Vibrios

CAUSATIVE AGENT:
Vibrio parahaemolyticus, V. alginolyticus, V. anguillarum, V. vulni- ficus,
V. damsela, V. fluvialis and V. penaeicida.
SPECIES AFFECTED:
Penaeus monodon, P. vannamei, P. japonicus
GROSS SIGNS:

Figure 3-15. Penaeus monodon with melanized

Affected shrimp may show erratic or disoriented swimming alternat-
cuticular lesion on the body surface due to bacte- ing with periods of lethargy. There is loss of appetite. General signs of
rial infection. Whitening of the necrotic muscle severe stress include opaqueness of abdominal muscle (Fig. 3-15), an-
at the 6th abdominal segment can also be
observed
orexia and expansion of chromatophores.
In larval and early postlarval shrimp, signs include melanization, ne-
crosis of appendage tips and the presence of large numbers of swarm-
ing bacteria visible in the hemocoel of moribund or weak shrimp. Due
to loss of appetite, fecal strands cannot be observed and gut is empty.
Hepatopancreas of affected shrimp showed varying degrees of inflam-
a hf
mation, hemocytic infiltration and fibriosis (Fig. 3-16).
EFFECTS ON HOST:

m Mortality in some instances is nearly 100% of affected population. Majority

f of cases of vibriosis is secondary in nature, occurring as a result of other
primary conditions, including other infectious diseases, nutritional diseases,
Figure 3-16. Hepatopancreas of Penaeus extreme stress, wounds, etc.
monodon infected with Vibrio parahaemolyticus .
Note the presence of atrophied tubules (a) and DIAGNOSIS:
melanized tubules (m). The intertubular spacesare
infiltrated with hemocytes (hf) and connec- tive Infection may be detected by bacteriological (isolation, purification
tissues (f). (Hematoxylin and Eosin stain,100x) and identification), histological (demonstration of rod-shaped bacteria

38 Health Management in Aquaculture

 in tissues of diseased shrimp), and serological (slide agglutination, FAT and ELISA
using specific antibodies) methods; and by Polymerase Chain Reaction (PCR).
PREVENTION AND CONTROL:
• Maintain good water quality and use nutritionally adequate diets.
• Minimize handling and overcrowding; reduce effects of other forms of
stress.
• Apply commercially available probiotics; use low salinity rearing water and
reservoirs; practice crop rotation and install greenwater culture system and
other system modifications.
• Perform immunoprophylaxis or vaccination.
• The disease can be treated with rigorous water management.
• Apply antibiotics and antibacterial substances only as a last resort.

SUMMARY

Fish and crustaceans that are not weakened by poor environmental conditions,
or by other causes, such as parasitic infestation, nutritional deficiency, han-
dling stress, or chemical intoxication, are more resistant to bacterial infections.
This is due to the presence of a large amount of bactericidal substances in the
blood, which helps overcome infections. So, the best precaution against the
occurrence of bacterial infections is to provide the fish with optimum environ-
mental conditions, adequate amounts of the right kinds of food and avoidance
of stress, including overcrowding. Vaccination / immunization and genetic ma-
nipulation (i.e., the development of specific pathogen resistant fry) are also
some ways of preventing bacterial diseases. The use of antibiotics should al-
ways be an option of the last resort.