Fish & Invertebrate Bacteriology

Question 1

Describe the general principles of treating bacterial infections in fish.

What type of bacteria cause most infections in fish?

Are any bacteria transmitted vertically?

What demographics are fish are most likely to get sick?

What are the risk factors for developing bacterial infections?

What are the typical signs a fish has a bacterial infection?

How are these infections diagnosed? Are special media needed?

What antibiotics can be used in food fish?

What other treatments can be provided?

Answer 1

Bacterial Diseases (C4)

• Most bacterial disease are due to opportunistic infections by ubiquitous bacteria – often gram-negative bacilli.
  
  • Morbidity and mortality often secondary to stressors
    
    • Increased exposure such as from poor biosecurity increases probability of transmission
  • Systemic infection most common, but local infections may be seen
• Gram positive bacilli (Erysipelothrix, mycobacterium, Nocardia, Renibacterium) and gram-positive cocci (Enterococcus, Lactococcus, Streptococcus, Vagococcus)
• Horizontal transmission seen with all bacteria, and vertical important for Renibacterium salmoninarum and Flavobacterium psychrophilum
  
  • Many ubiquitous in environment and normal components of protective biofilm
• Juvenile and geriatric fish are more likely to show clinical signs
• Risk factors: high stocking density, inappropriate social structure, inappropriate environments, aggression/displacement, poor water quality (low DO, high ammonia), contaminants, inappropriate water temperature, poor nutrition, recent handling, recent transport, recent spawning, other disease (especially if damage physical defenses – Ichthyophthirius multifillis), and immune suppressive therapy (copper)
• Acute infections often present with erythema, petechiae, skin ulcers and fin erosions. Ulcerative keratitis or uveitis may be seen
• Chronic infections may present with poor BCS and change in coloration
• Culture useful for diagnosis
  
  • Routine media (trypticase soy agar, brain-heart infusion agar or MacConkey agar) useful for Aeromonas, Pseudomonas, Vibrio, Edwardsiella, Streptococcus, and Yersinia
    
    • Selective media needed for Flavobacterium, Renibacterium and Francisella
  • Molecular testing and sequencing allows for definitive species identification. Identification based on phenotype may not be reliable for bacteria in fish.
  • Husbandry management: increase aeration (DO target 95-100%), resolve water quality, resolve temp issues, increase water flow rate/turnover, reduce social stressors (barriers), consider adding Vit C, minimize stressors.
    
    • Higher doses of UV reported fro some bacteria (Flavobacterium and Renibacterium). UV and ozone are synergistic and lower applied doses can be used when combined. Most effective after mechanical and biological filtration.
  • For food fish, florfenicol, oxytetracycline and sulfadimethoxine/ormetoprim are FDA approved for certain species – must follow label instructions.
    
    • Low dose hypersalinity (1-3 g/L) can be used to reduce osmotic stress in freshwater fish
    • Anti-inflammatory doses of steroids may limit overstimulation of the immune system and improve prognosis
  • Prevention – reduce or resolve stressors (especially after transport or at permissive water temps), reduce exposure/transmission, and reduce severity of disease (Vit C, immune stimulants – beta glucans, allicin/crushed garlic, propolis ethanol extract)
  • Zoonosis – highest risk is from direct wound inoculation, most fish bacterial pathogens common in environment and transmission possible through water contact with mucosal surfaces, open wounds, or food

Question 2

Describe septicemia in fish.

What bacteria usually cause issues in marine fish? What about freshwater and brackish fish?

What conditions are associated with outbreaks and infection?

What lesions are typically present on necropsy?

Answer 2

• Bacterial Disease (ZP)
• Gram-negative bacteria, septicemia
  
  • Vibrio alginolyticus, Vibrio (Listonella) anguillarum (red pest of eels), V. ordalii, Allivibrio salmonicida (Hitra dz), V. vulnificus, and Photobacterium damselae subsp damselae.
    
    • Warm and cold water marine spp.
  • Aeromonas hydrophila (red pest), A. veroni, A. caviae, A. sobria, Pseudomonas fluorescens, and others.
    
    • FW and brackish water spp, typically warm water.
  • P. damselae piscicida is not motile but included here for association with sepsis.
  • Rising environmental temperatures are important in pathogenesis.
  • Spring epizootics not uncommon due to rising temps.
  • Vibrio can similarly be dependent on host and environment, but can occur in absent of recognized predisposing factors.
  • Infection associated with stressful events.
  • Necropsy – multifocal hemorrhage, nephron and splenomegaly, cutaneous ulcers.
  • Necrotizing dermatitis and vasculitis common and characteristic.
    
    • Progression to deep ulceration or cavitary necrotizing myositis.
    • Multicentric granulomas are the typical presentation for chronic P. damselae piscicida infection (pseudotuberculosis).
    • Diagnosis dependent on visualization of typical septicemia lesions, ID of organism.

Question 3

What is the etiologic agent of furunculosis in fish?

What species get typical furunculosis? What species get atypical furunculosis?

What are the typical clinical signs? How do the atypical infections differ? What are mortalities like?

What are the typical lesions on necropsy?

How are these infections treated?

How can they be prevented?

Answer 3

• Aeromonas salmonicida (CGFM)
  
  • Aeromonas salmonicida subsp salmonicida (typical furunculosis) causes furunculosis. Other subspecies can cause atypical furunculosis in a variety of species.
    
    • Both cause septicemia – skin ulcer and hemorrhages are common clinical signs
  • Commonly associated with rhabdivirus carpio (infectious dropsy of carp)
  • Nonmotile gram negative bacilli
  • Horizontal transmission (ingestion and contact). Can survive for months in freshwater, but days in saltwater
  • Typical furunculosis – wild and cultured freshwater salmonids – brown trout, Atlantic salmon, Pacific salmon
  • Atypical furunculosis – affects variety of freshwater and saltwater teleosts – salmonids, sand eels, halibut, goldfish, koi, Atlantic cod
  • Permissive water temp often > 72F
  • Signs
    
    • Peracute death in juvenile fish, lethargy, inappetence, erythema, ecchymoses (base of fins), hemorrhages, skin ulcers, fin erosions, boils (furuncles /nodules) – not common, ulcers, skin darkening, gill pallor, generalized edema, coelomic distention.
    • High mortality – up to 90%
  • Diagnosis:
    
    • Cytology histology and culture – short gram-negative bacilli (from blood smear, effusion, tissue cytology or histo)
    • Necropsy – petechiae, congestion, serosanguinous coelomic effusion, focal necrosis, tissue pallor
    • Molecular techniques have limited ability to differentiate between species and subspecies
  • Treatment
    
    • Antibiotics based on C/S. Oxytetracycline, sulfadimethoxine/ormetoprim, and florfenicol approved for salmonids with furunculosis
    • Potassium permanganate immersion can be used if infection limited to skin
  • Prevention
    
    • Vaccines for typical furunculosis in salmonids – usually by IP injection. Vaccines with mineral oil adjuvants best but associated with adverse reactions (coelomic pigmentation, adhesions, granulomas)
    • Immune stimulants (allicin, glucans, chitosan, and probiotics) can increase resistance to A. salmonicida.

Problem 47 – Furunculosis, Goldfish Ulcer Disease, Carp Erythrodermatitis – Aeromonas salmonicida

• Host: Salmonids, cyprinids
  
  • Atlantic salmon are most susceptible – rainbow trout most resistant – furunculosis is how it presents in them
  • Cyprinids are more likely to get the ulcerative form of disease
• Clinical Signs/Pathology:
  
  • Acute: hemorrhage around fins and gills, organs
  • Chronic: focal areas of swelling, hemorrhage, and tissue necrosis in the muscles
  • Crater-like furuncles, septicemia, liquefactive necrosis
• Diagnostics:
  
  • Pure culture in affected tissue
  • Gram negative, non-motile rod
• Treatment:
  
  • Vaccine available
  • Oxytetracycline, furazolidone, oxolinic acids, sulfonamides – many isolates are resistant
  • Fluoroquinolones and florfenicol are commonly effective
  • Bacteria can survive off the fish for about 3 weeks – need to have good disinfection
• Aeromonas salmonicida (ZP)
• Gram negative, nonmotile, short rod, obligate parasite of fish.
• Causes furunculosis in salmonids.
  
  • Atypical A. salmonicida including 3 additional subspecies – S. smithia, S. achromogenes, S. masoucida, and others cause dz in cyprinids, salmonids, FW and SW.
• Associated with goldfish ulcer dz, carp erythrodermatitis, ulcer disease of flounder.
• Carrier fish crucial to maintenance of dz.
  
  • Transmission via direct contact, contaminated water, ingestion of infected material.
  • Peracute to acute septicemia.
    
    • Fingerlings through adults.
  • Subacute to chronic form is furunculosis.
    
    • Older individuals with partial immunity or spp with greater resistance.
  • Atypical infections assoc with ulcerative rather than septic dz.
  • Gross septicemia – fin base and visceral hemorrhage, splenomegaly.
    
    • Histo – large intravascular aggregates of bacteria, little to no inflammation.
  • Gross furunculosis – extensive cavitary or hemorrhagic lesions in skin and muscle, distort external contours of body.
    
    • Histo – multicentric necrosis, hemorrhage, inflammation.
    • Deep liquefactive necrosis and hemorrhage of body wall muscle, cutaneous ulceration.
    • Ulcerations will be contiguous with deep cavitary lesions.
  • Atypical infections – cutaneous ulcers of various severity, dermal edema.
  • ID of organism with selective bacterial culture or PCR necessary for definitive dx.

Question 4

What is the etiologic agent of motile aeromonad septicemia?

What fish are typically affected?

How is this transmitted?

What are the typical clinical signs? What is the mortality like?

What lesions are present on necropsy?

What medications are typically used?

Answer 4

• Motile Aeromonad Septicemia
  
  • Several Aeromonas species are opportunists that can cause motile aeromonad septicemia – most commonly reported bacterial infection of fish
  • Readily grow in culture – may be infection or contaminant thus can misdiagnose primary disease.
  • Common in the environment and many aeromonad species nonpathogenic
    
    • Motile gram negative bacilli
  • Amphibians and reptiles may act as carrier or vectors. Transmission horizontal, through ingestion and contact.
  • Predominantly found in cultured and wild freshwater fish – carp, goldfish, channel catfish, tilpia, bass. Less common in marine teleosts.
  • Permissive water temp often 50-72 or higher.
  • Signs
    
    • Inappetence, lethargy, erythema, petechiae, skin ulcers, fin erosins, cutaneous edema, boils (furuncles), exophthalmos, coelomic distention, gill pallor, neurologic signs.
    • Mortality typically low but can reach 100% in fry or fingerlings
  • Diagnosis – based on multiple consistent findings (clinical signs, cytology, necropsy, histology, and culture). Short gram negative bacilli may be seen on blood smears, effusions, tissue cytology, or histology.
    
    • Necropsy – petechie, congestion, coelomic effusion, tissue pallor, organomegaly, focal necrosis
    • Culture of affected tissues or blood routinely
  • Treatment – Antibiotic therapy based on C/S. Oxytetracycline approved for catfish and salmonids. Enrofloxacin, ceftazidime, amikacin, florfenicol and TMS commonly used in aquariums.
  • Prevention – motile aeromonads ubiquitous and cannot be eliminated – prevention focused on reducing stressors and severity of disease. Immune stimulants may be helpful if given prior to infection.
  • Zoonosis – some species are zoonotic (foodborne, and waterborne diarrhea) from consumption of raw or inadequately cooked or processed fish
    
    • Low risk if healthy immune system, no damage to skin barrier and good food hygiene

Problem 46 – Motile Aeromonad Infections – Aeromonas hydrophila

• Host: all fish are susceptible
• Epidemiology:
  
  • Several Aeromonas pathogens – A. hydrophila is the most significant
  • Often isolated from mucosal surfaces & internal organs of healthy fish
  • Higher prevalence in polluted waters
  • Have zoonotic potential – usually through puncture wounds or ingestion of infected fish
• Clinical Signs & Pathology:
  
  • Superficial to deep skin lesions
  • Start with hemorrhage and necrosis and may progress to reddish or gray ulcers with muscle necrosis and eventually progressing to hemorrhagic septicemia
• Diagnostics:
  
  • Presence of free melanin or lipofuscin from ruptured melanomacrophage centers is characteristic of septicemia
  • Kidney is best sample for cutlure
• Treatments:
  
  • Outbreaks often resolve with water quality corrections
  • Oxytetracycline and nifurpirinol have successfully controlled some outbreaks – sulfas are commonly used for resistant strain

Question 5

Describe the effects of vibriosis and photobacteriosis in marine fish.

What species are commonly affected?

Is this a normal part of their flora?

What are the typical clinical signs?

How is it diagnosed? What samples should be taken?

How is it treated?

Are they zoonotic?

Answer 5

• Vibriosis
  
  • Caused by a variety of bacteria within the genus vibrio – some primary pathogens but most ubiquitous in the environment
  • Causes bacterial septicemia in a wide range of fish species
  • Aliivibrio salmonicida common in salmon and cod aquaculture in Canada, Norway and UK
  • Etiology – over 20 Vibrio spp – Vibrio (Listonella, Beneckea) anguillarum causes saltwater furunculosis
    
    • Part of normal fish intestinal flora which closely resembles populations in the water. Common commensals in elasmobranch tissues and blood and are present in light-emitting organs of marine fish and cephalopods.
    • Curved to spiral gram-negative bacilli
  • Transmission – horizontal and likely also vertical transmission. Aquatic invertebrates may act as carriers or vectors
  • Disease is possible in any marine or brackish water fish (including elasmobranchs)
    
    • Commonly reported in Salmonidae (salmon and trout), bass, cod, yellowtail, seahorses, requiem sharks. Occasionally reported in freshwater fish.
  • Permissive temp high for most Vibrio spp and low for Aliivibrio salmonicida.
  • Signs
    
    • Lethargy, inappetence, skin darkening, pallor, erythema, petechiae, ecchymosis, fin erosions, coelomic distention (ascites or organomegaly), or pinch coelom (loss of BCS), corneal edema, keratitis, exophthalmos, gill pallor, neurologic or respiratory signs
  • Diagnosis – based on multiple consistent findings (clinical signs, cytology, necropsy, histology, culture) – not culture alone
    
    • Short curved to spiral shaped gram-negative bacilli seen on blood smear, effusions, tissue cytology and histology.
    • Necropsy – petechiae, congestion, coelomic effusion, tissue pallor, focal necrosis (esp kidney and spleen) with intralesional gram negative bacilli.
    • Culture of affected tissues (kidney and spleen) or blood often successful.
      
      • Culture with routine or selective mediation (thiosulfate citrate bile salts). Improved success with 10 g/L NaCl in marine spp.
      • Blood culture may be positive in healthy elasmobranch
      • PCR recommended for species identification
  • Management – in aquaculture, regulations may require movement restrictions. Reduction of stressors or other diseases sometimes enough to control signs.
    
    • Antibiotics based on c/s
    • Healthy, diverse microbiome may reduce exposure to more pathogenic Vibrio spp
    • Commercial immersion vaccines available for some spp (V. anguillarum and V. ordalii in salmonids)
  • Zoonosis
    
    • Predominately foodborne through ingestion of raw or inadequately cooked shellfish (oysters) or contaminated food/water – signs include nausea, vomiting, abdominal cramps, pyrexia
    • Transmission possible through inoculation into wounds (local infection à sepsis)
    • The species that causes cholera (Vibrio cholerae) is not found in fish
    • Low risk if healthy immune system, no damage to skin barrier and good food hygiene

Problem 50 – Salt Water Furunculosis, Vibriosis– Vibrio spp.

• Host: Marine and estuarine fish
• Clinical Signs/Pathology:
  
  • V. anguillarum - Hemorrhage of skin, fins, tail, internal organs, abdominal distension
  • V. salmonicida - Coelomic sanguineous fluid, swim bladder hemorrhage, internal viscera, skin lesions
  • Many other vibrio species
  • Septicemic disease, renal, splenic necrosis, cardiomyopathy
• Diagnostics:
  
  • Culture
  • Gram negative, short (usually curved) motile rods
• Treatment:
  
  • Oxytetracycline, nitrofurans, sulfonamides
  • There are vaccines for certain vibrios

Problem 51 – Pasteurellosis, Pseudotuberculosis, Photobacteriosis – Photobacterium damselae

• Host: Warm and temperate marine fish
• Clinical Signs/Pathology:
  
  • Small hemorrhages on operculum or base of fin, abnormal skin color
  • Enlarged spleen, kidney in the acute form – multiple white foci on these organs in chronic form
• Diagnostics:
  
  • Gram-negative, nonpigmented, short nonmotile rods
  • Culture
• Treatment:
  
  • Ampicillin, sulfonamides, oxytetracycline

Question 6

What is the etiologic agent of Enteric Septicemia of Catfish?

What is the primarily affected species? Are any others affected?

How is it transmitted?

What are the typical clinical signs?

How is it diagnosed? What samples need to be collected?

What lesions are typically present on necropsy?

How is it treated and prevented?

Answer 6

• Enteric Septicemia of Catfish
  
  • Edwardsiella ictaluri – causative agent of enteric septicemia of catfish
  • Primary pathogen causing significant economic losses in channel catfish – enteric, neurologic and septicemia disease
    
    • Motile, pleomorphic gram-negative bacilli
  • Transmission – ingestion (fecal-oral or scavenging) or through gills
    
    • Fish that resolve signs become carrier and piscivorous birds may act as carriers
  • Epizootic disease seen primarily in channel catfish in aquaculture, but can be seen in other catfish spp too.
    
    • Disease seen in cultured zebrafish and tilapia and wild ayu (different strains)
    • Hybrid catfish more resistant
    • E. ictaluri isolated from wider variety of freshwater fish without morbidity or mortality
  • Permissive water temp – 72-82F
  • Signs
    
    • Most presentations neurologic or enteric/septicemic with neurologic form more chronic.
      
      • Spiral swimming, circling, inappetence, lethargy, low tail position, skin ulcers on the head in fingerlings
    • Enteric/septicemic form more acute – can see erythema, petechiae, ecchymoses, hemorrhages, skin ulcer, exophthalmos, gill pallor, coelomic distention
    • Biphasic – disease in Spring and Fall; mortality > 50% especially in acute disease.
  • Diagnosis
    
    • Multiple consistent findings (signs, cytology, necropsy, histology, culture)
    • Necropsy – petechiae, congestion, coelomic effusion, tissue pallor, splenomegaly, renomegaly with nodules
    • Histo – hemorrhagic enteritis and granulomatous splenitis, nephritis, and encephalitis with intralesional gram negative bacilli
    • PCR offers higher sensitivity.
    • Culture affected tissues (spleen, anterior kidney, liver, brain) or blood
  • Management
    
    • Slowly change temp out of permissive range (72-82F), reduce/withhold food to reduce fecal shedding
    • Antibiotics indicated if water temp remains within permissive range. Florfenicol and sulfadimethoxine/ormetoprim approved for use.
    • Effective commercial immersion vaccines are available

Problem 48 – Enteric Septicemia – Edwardsiella ictaluri

• Host: Ictalurids (Catfish)
  
  • Channel catfish can be asymptomatic carriers
• Clinical Signs/Pathology:
  
  • Enteric or meningeal forms, hole in the head lesions, skin punctuate lesions
  • Hemorrhagic enteritis, liver necrosis, meningitis
• Diagnostics:
  
  • Culture or ELISA of tissue smears
  • Gram negative bacteria
  • This is a fastidious organism that is easily outgrown by other bacteria
• Treatment:
  
  • Oxytetracycline, sulfas
  • Vaccine available
  • Associated with warm water temperatures (seasonal occurrence)
• Edwardsiella.
  
  • Gram negative, family enterobacteriaceae.
    
    • E. ictaluri – enteric septicemia of catfish ESC.
    • E. tarda – Edwwardsiella septicemia, emphysematous putrefactive dz, fish gangrene, red dz of eels).
    • E. piscicida.
  • Edwardsielloses primarily dz of warm water aquaculture.
    
    • Versatile, affect many fish spp.
    • Transmission horizontal from fish or environment, especially E. tarda – resides in intestines of fish and vertebrates.
      
      • Opportunistic, FW and marine spp over broad temp range.
    • E. ictaluri obligate pathogen of FW fish, seasonal at 18-28 deg C water temp.
      
      • Causes ESC.
        
        • Channel catfish farming, SE US and Asian carp.
        • Also tilapia, laboratory zebrafish.
        • Survivors become carriers.
        • Ingestion primary route of infection.
          
          • Loss can be limited by restricted feeding.
      • Chronic form of dz assoc with infection of olfactory sensory epithelium leading to severe meningoencephalitis aka hole in the head.
        
        • Infection erodes through fontanel in dorsal cranium.
      • ESC and hole in the head – both hemorrhagic septicemia in acute phase, multiple organs, necrosis.
      • Ascites prominent feature of ESC in catfish.
        
        • Necrosis of parenchymal organs extensive.
        • Inflammation progresses to granulomas.
        • Lesions in eels – suppurative, liquefactive.
        • Tilapia granuloma formation.
        • ESC also results in ulcerative dermatitis in eels and channel catfish.
          
          • Bacterial embolization to the skin.

Question 7

What is the etiologic agent of edwardsiellosis?

How is this transmitted?

What species are commonly affected?

What are the typical clinical signs?

What lesions are present on necropsy?

How is it diagnosed? What samples need to be collected?

How is it treated and prevented?

Is it zoonotic?

Answer 7

• Edwardsiellosis
  
  • Edwardsiella tarda, Edwardsiella piscicida, and Edwardsiella anguillarum are enteric bacteria that can cause bacterial septicemia known as edwardsiellosis
  • All bony fish susceptible – response may be suppurative and in catfish, disease known as emphysematous putrefactive disease.
  • Differentiation between the species o Edwardsiella requires PCR
  • Small straight gram negative bacilli
  • Transmission – oral
    
    • Also found in invertebrates, amphibians, reptiles, and mammals but role in transmission unknown
  • Disease possible in many marine and freshwater fish
    
    • Epizootics reported in eels, tilapia, red seabream, bass, catfish, flounder, and in marine aquarium teleosts.
    • One report in ribbontail ray
  • Signs
    
    • Clinical course may be acute or chronic, chronic is more common
      
      • Lethargy, spiral swimming, positive buoyancy, inappetence, erythema, petechiae, ecchymoses, skin ulcers, fin erosions.
      • Catfish may show skin abscesses or fistulas
      • Gill pallor (due to anemia), coelomic distention, keratitis, uveitis, hypopyon, exophthalmos may be seen.
      • Mortality low and chronic
  • Diagnosis based on sings, cytology, histology and culture
    
    • Small straight gram negative bacilli on blood smear, effusions, tissue cytology, or histology
    • Necropsy – petechiae, congestion, coelomic effusion, pallor, organomegaly, or nodules in tissues. Malodorous abscesses in viscera or skeletal muscle, esp in catfish
    • Culture of affected tissues (kidney and spleen especially) on routine media
    • PCR or other molecular testing and sequencing in conjunction with other diagnostic
  • Treatment
    
    • Antibiotics based on C/S. Recommend tetracycline, aminoglycoside, fluroquinolone (resistance to macrolides and lincoasmides common)
  • Autogenous vaccines can help with recurrent outbreaks. Commercial vaccines unlikely due to antigenic diversity.
  • Zoonosis
    
    • E. tarda zoonotic but human disease is rare
    • Transmission through oral ingestion raw/undercooked fish. Gastroenteritis most common sequela
    • Inoculation into wounds with local wound infection may occur.
    • Humans can be asymptomatic carriers.

Problem 49 – Emphysematous Putrefactive Disease, Edwardsiellosis – Edwardsiella tarda

• Host: Catfish, Japanese eels
  
  • ZOONOTIC: causes enteric disease
• Clinical Signs/Pathology:
  
  • Malodorous ulcers with draining fistulas, abnormal buoyancy due to gas in tissues
  • Liquefactive necrosis of viscera with fibrinous peritonitis
• Diagnostics:
  
  • Culture, fluorescent antibody
• Treatment: oxytetracycline
• Edwardsiella.
  
  • Gram negative, family enterobacteriaceae.
    
    • E. ictaluri – enteric septicemia of catfish ESC.
    • E. tarda – Edwwardsiella septicemia, emphysematous putrefactive dz, fish gangrene, red dz of eels).
    • E. piscicida.
  • Edwardsielloses primarily dz of warm water aquaculture.
    
    • Versatile, affect many fish spp.
    • Transmission horizontal from fish or environment, especially E. tarda – resides in intestines of fish and vertebrates.
      
      • Opportunistic, FW and marine spp over broad temp range.
      • E. tarda assoc with emphysematous putrefactive dz, large raised cutaneous bullae that rupture to release malodorous gas, cavernous liquefied muscle.
    • Readily culturable on common media.
    • Can ID with molecular testing.
    • E. tarda, E. piscicida cannot be differentiated with ID systems like the API20E substrate panel.

Question 8

Describe the effects of Columnaris and Flexibacteriosis in fish.

What are the etiologic agents? Which affects marine or freshwater fish?

How are they transmitted?

WHat species are commonly affected?

What risk factors are their for infection?

What are the typical clinical signs?

How is this diagnosed? What samples should be collected?

How are these diseases treated and prevented?

Answer 8

• Columnaris and Flexibacteriosis
  
  • Flavobacterium columnare – causative agent of columnaris disease – common in freshwater ponds
    
    • Lesions on the skin, fins, and gills but can be systemic
    • Ubiquitous in freshwater habitats
  • Marine counterpart is Tenacibaculum maritimum (marine flexibacteriosis) – similar lesions but systemic spread less common
    
    • Ubiquitous in saltwater habitats
  • Long motile gram negative bacilli
  • Horizontal transmission common (ingestion, contact, inoculation
    
    • Survival of columnaris increased in warm, hard, alkaline freshwater and high organic loads
  • Signalment – columnaris possible in all freshwater bony fish but common in goldfish, carp, koi, mollies, swordtails and platies. Also common in channel catfish, tilapia, salmon and rainbow trout
    
    • Marine flexibacteriosis possible in all marine teleosts but is common in salmon, barramundi, seabream, amberjack, flounder and turbot. Has also been identified in elasmobranchs (skates and sand tiger shark)
  • Risk factors – highest mortality at 75-90F, high nitrite, alkalinity, hardness and calcium
  • Signs – may see acute mortality without other signs, yellow-white or dark brown discoloration of gills (especially distal aspect of primary lamellae), dyspnea. tachypnea, pruritis,
    
    • Chronic presentation more common – inappetence/reduced feeding, lesions on skin, fins, oral cavity, multifocal pallor develops into ulcers with white or yellow cotton-like mucus plaques. Lesions have a hyperemic yellow or white rim
    • Fin lesions start at the base of the fin and progress distal (unlike typical bacterial fin rot)
  • Diagnosis - direct microscopy of skin scrape/gill biopsy – long bacilli groups in haystacks
    
    • Often seen increased mucus, opportunistic pathogens (scuticocilates), gill hyperplasia and fusion of secondary filaments
    • Histo – necrotizing ulcerative dermatitis or branchitis
    • Culture – contaminants common and organisms are fastidious and can be inhibited by the high-nutrient content of routine media
    • PCR on gill, skin, or cranial kidney
    • LAMP – loop-mediated isothermal amplification used in channel catfish
  • Treatment
    
    • Slowly reduce water temperature to < 75F and if in freshwater, low dose hypersalinity (2-3 g/L) may help.
    • Antibiotics can improve morbidity and mortality but elimination unlikely. Florfenicol, Oxytetracycline, Chloramine-T, and hydrogen peroxide approved in US. Other treatments: enrofloxacin, TMS, Potassium permanganate, copper, oxolinic acid
    • High salinity (> 5 g/L) and low hardness (< 70) reduce columnaris survival
    • USDA approved vaccine available for channel catfish

Problem 37 – Columnaris Infection – Flavobacterium columnaris

• Host/location: Freshwater fish – skin/gills – gill infections are less common but more serious
• Clinical Signs/Pathology:
  
  • Erosive, necrotic skin lesions that become septicemic, fin rot, tail rot
  • Channel catfish may develop septicemia without external lesions
• Life Cycle: This bacteria likely occurs on healthy fish skin
• Diagnosis:
  
  • Culture, typical bacteria on wet mounts of skin or gills
  • Gram negative, gliding rods with haystack appearance
  • Difficult to culture
• Treatment: Oxytetracycline (most resistant to sulfas), copper, potassium permanganate

Flavobacteriacea - ZP

• Group of gram negative bacteria.
  
  • Flavobacterium columnare (columnaris dz aka saddleback).
    
    • FW spp, tropical ornamentals.
  • Flavobacterium psychrophilum (bacterial coldwater dz aka peduncle dz aka rainbow trout fry syndrome).
    
    • Salmonids, non-salmonid FW spp.
  • Flavobacterium branchiophilum (bacterial gill dz).
    
    • Young hatchery-reared salmonids.
  • Tenacibaculum maritimum (tenacibaculosis, SW columnaris, black patch necrosis.
    
    • Marine fish, commercial mariculture.
• Columnaris and coldwater dz in FW, tenacibaculosis in marine environments major dz.
• Columnaris and tenacibaculosis notable for lack of host specificity.
• Morphology – adhere to epithelial surfaces, rods, exhibit gliding motility, produce yellow flexirubin or carotenoid pigments.
  
  • Tissue damage likely promoted by extracellular proteases i.e. chondroitin AC lyase, degrades ground substance of connective tissues.
  • Often persists in environment, outbreaks assoc with stressors i.e. handling, transport, overcrowding, water quality.
  • Diseased and dead fish source of infection.
  • Young fish most susceptible.
• Outbreaks of columnaris and tenacibaculosis above 15 deg C, coldwater dz occurs at 15 deg C and below.
• BGD can occur at temps from 5-20 deg C.
• Internal infection by F. psychrophilum and gill infection by F. columnare often rapidly fatal.
• Columnaris, coldwater dz, tenacibaculosis – ulcerative dermatitis and necrotizing myositis.
  
  • Characteristic lesion – pale yellow discoloration of skin, corresponds to large numbers of long slender bacteria in dermal connective tissues and along epithelial margins.
    
    • Focal depigmentation followed by rapid lesion expansion and necorisis.
  • Inflammatory responses mononuclear, relatively mild.
  • Columnaris – caudal peduncle, mouthparts, fins, dorsum, saddleback lesion. Necrotizing bronchitis.
    
    • Gill lesions – well demarcated yellow-tan areas of necrosis.
      
      • Grossly resembles CyHV3 in koi.
        
        • Consist of mixture of epithelial proliferation and necrosis with bacterial mats.
        • In FW, secondary saprolegnia common.
  • Gross and microscopic lesions of these bacteria virtually pathognomonic.
    
    • Require special media to grow, confirmation via serologic or molecular methodologies.

Question 9

What is the etiologic agent of bacterial cold-water disease? Does it have another name?

What species does it affect?

How is it transmitted?

What are the clinical signs? How does it difffer between adults and fry?

How is it diagnosed? What samples need to be collected?

What lesions are seen on necropsy?

How is this disease treated and managed?

Answer 9

Flavobacterium psychrophilum (CGFM)

• Bacterial cold-water disease – salmonid adults and fingerlings
• Rainbow trout fry syndrome – seen in salmonid fry
• Disease seen when water temperature falls below 59F
• Motile long gram negative bacilli – horizontal and vertical transmission
• Signalment:
  
  • Found in cold freshwater habitats – common problem for freshwater salmonids, especially rainbow trout and coho salmon. Also reported in other teleosts including European eels, common carp, and ayu
• Signs
  
  • For both BCWD and RTFS – lethargy, inappetence, gradual weight loss, coelomic distention, cloacal/anal distention, exophthalmos or gill pallor
  • BCWD: Skin darkening, large cutaneous ulcers/erosions especially around dorsal fin – can be deep enough to reach vertebrae. Mortality can reach 70-90%
  • RTFS: Skin darkening/asymmetrical coloration, neurologic signs (abnormal position in water column and spiral swimming), low mortality and spinal deformities may be seen in survivors
• Diagnosis
  
  • Direct microscopy of skin scrapes or gill biopsies – long gram-negative bacilli (shorter bacilli than F. columnare)
  • Necropsy – splenomegaly, serosanguinous coelomic effusion, tissue pallor;
  • Histo – pyogranulomatous inflammation and necrosis extending in to the muscle, bone and cartilage with intralesional gram negative bacilli (spleen, heart, gills, muscle, bone and cartilage)
  • Confirmed with PCR
  • Culture can be difficult – organisms fastidious and inhibited by nutrient content of routine media at ~60F
  • ELISA and FA developed to screen broodstock with kidney/ovarian tissues
• Management
  
  • Environmental management can resolve morbidity and mortality – slowly increase water temperature, clinical signs resolve as temperature increases
  • Increase cleaning/disinfection – transfer to systems cleaned with sodium hypochlorite
  • Medical management has poorer success due to reduced appetite and immune suppression at lower water temp
  • Antibiotics may be indicated. Florfenicol and Oxytetracycline are approved for use with other treatments (Potassium permanganate, copper sulfate, chloramine T) also reported
• Prevention – screen broodstock with ELISA and disinfect eggs with iodophors to reduce vertical transmission

Problem 38 – Bacterial Cold-Water Disease – Flavobacterium psychrophilum

• Host/location: Salmonids (Coho salmon particularly susceptible) – skin and brain
• Clinical Signs/Pathology:
  
  • Erosions around peduncle, distal part of body, scoliosis, spinal deformities, pigmentation
• Transmission: Isolates from the surface o f normal fish – skin damage initiates infection, also infects eggs
• Diagnosis: Gram-negative filamentous rods with gliding motion
• Treatment: Quaternary ammonium, choramine-T, copper constant flows - oxytetracycline
• Coldwater (ZP)– peduncle, base of skull, kaw, dorsum, perianal area.
  
  • Unlike other members of this group, severe systemic dz.
    
    • Hemorrhagic necrosis in most internal organs and muscle.
    • Can result in complete loss of caudal fin and exposure of vert bone.
    • Predilection for connective tissues.
    • Chronic infection – erratic swimming, blackening of tail and spinal deformities, meningitis, osteitis, periosteal proliferation.
      
      • May resemble whirling dz.

Question 10

What is the etiologic agent of bacterial gill disease?

What species are typically affected?

What are the clinical signs?

How is it transmitted? What are the risk factors for transmission?

How is it diagnosed?

What are the lesions on necropsy? How are the gills affected over time?

How is it treated?

Answer 10

Problem 39 – Bacterial Gill Disease – Flavobacterium branchiphilum

• Host/location: Salmonids - gills
• Clinical Signs/Pathology:
  
  • Lethargy, flared opercula, excess mucus from gills
  • Proliferative branchitis with epithelial hyperplasia, obliteration of interlamellar space, lamellar fusion
• Transmission: Mainly a production management disease – risk factors include low oxygen, high turbidity, high ammonia, overcrowding.
• Diagnosis:
  
  • Role of bacteria may not be exclusive to this organism, but bacteria may be seen,
  • Histopathology
  • Gram negative
• Treatment: Quaternary ammonium, chloramine-T, copper constant flows - oxytetracycline

Bacterial gill disease (BGD) - ZP

• Commercially important, cultured FW fish worldwide.
  
  • Most notably in salmonids.
  • Severe in intensively reared fry and fingerlings.
  • Primarily husbandry and management dz in hatcheries.
  • CS – lethargy, anorexia, dyspnea.
    
    • Flared opercula, crowding water inlets.
  • Explosive morbidity and mortality.
• Causative agent – Flavobacterium branchiophilum.
  
  • Reproduced in RBT – horiz transmission and water exposure.
• Overcrowding, high levels of nitrogenous wastes, low levels od DO factors that may promote BGD.
• Certain spp i.e. brook trout more consistently affected.
• Seasonal pattern suggested – rise in spring and summer.
  
  • Rising temp and higher precipitation possible risk factors.
  • Also higher risk age groups i.e. fry and fingerlings peak in spring.
• Bacterial colonization of gill filaments is preceded by ultrastructural changes to the superficial epithelium of the filament – cytoplasmic blistering and degen of microridges together with irregularities at the tips of filaments.
  
  • Colonization begins at irregular filament tips, proceeds proximally, extends over lamellae.
  • Very high morbidity and mortality.
    
    • Lamellar necrosis and edema (acute first few days of dz).
    • Epithelial cell hyperplasia and mucous cell metaplasia along distal third of filaments in subacute phase (3-5d).
    • Severe epithelial hyperplasia and mucous metaplasia, leading to lamellar and filamental fusion with intralesional filamentous bacteria in clumps of mucus chronic phase (7-14d).
• Dx – examination of whole mount preps or histo of gill.
  
  • Numerous gram-neg filamentous bacteria in interlamellar spaces and along branchial lamellar epithelium.
  • ELISA, IFAT, PCR.
  • Failure to obtain rapid, accurate dx and provide tx can result in high mortality and economic impacts.

Question 11

What is the etiologic agent of Enteric Redmouth Disease in fish?

What are the affected species?

How is it transmitted?

What are the typical clinical signs?

How is it diagnosed? What samples need to be collected?

What lesions are seen on necropsy?

How is it treated?

Answer 11

Yersiniosis

• Yersinia ruckeri – causes hemorrhagic septicemia (known as yersiniosis or enteric red mouth)
• Obligate pathogen of freshwater salmonids – has caused losses in aquaculture
  
  • Serotype 01a cause of most epizootics
  • Short gram negative bacilli, usually motile
• Horizontal transmission – predominantly fecal-oral but waterborne through the gills may also occur. Some evidence of vertical transmission
  
  • Can survive for months outside the hosts and carriers include fathead minnows. Fish that resolve clinical signs continue to shed for up to 2 mo. Also found in aquatic invert, reptiles, birds and mammals.
• Freshwater teleosts most susceptible, particularly salmonids in aquaculture (rainbow trout and Atlantic salmon)
  
  • Also isolated from other freshwater fish – goldfish, common carp, koi, eels, channel catfish
• Permissive water temp 70-80F
• Signs
  
  • Multiple fish usually affected with acute presentation more common in juveniles (chronic seen in adults)
  • Lethargy, swimming near surface, inappetence/reduced feeding, petechia, erythema, hemorrhages (especially around oral cavity and eyes) which often progresses to ulcers, skin darkening, hyphema, keratitis, exophthalmos (may appear blind), gill edema/pallor, coelomic distention
  • Mortality rate usually low, but can have high cumulative loss
• Diagnosis
  
  • Often based on clinical signs and culture results
  • Short gram-negative bacilli seen on blood smear, effusion, tissue cytology or histology
  • Necropsy/histo – petechiae, congestion, yellow to serosanguinous coelomic effusion, tissue pallor, splenomegaly, enteritis (with thick yellow fluid)
  • Culture of kidney (acute infection) or intestine (carriers) often successful with routine media
  • PCR, ELISA, FA also available on blood, fecal or tissue
• Treatment
  
  • Antibiotics usually indicated based on culture and sensitivity – commonly use oxytetracycline, potentiated sulfonamides, florfenicol
  • Restrict access by birds and mammals (may act as vectors)
  • USDA approved vaccine for rainbow trout – effective against biotype 1 but outbreaks of biotype 2 still possible
• Not zoonotic – the yersinia that affect humans have not been reported in fish

Problem 52 – Enteric Redmouth Disease, Slood Spot, Yersiniosis – Yersinia ruckeri

• Host: Salmonid fish (market size fish), other species affected as well
• Clinical Signs/Pathology:
  
  • Darkening of skin, blindness/hyphema, hemorrhage around the mouth, vent, fins, skin
  • Hemorrhagic septicemia, necrosis of hematopoietic tissue (anemia)
• Diagnostics:
  
  • Culture
  • Gram-negative
• Treatment:
  
  • Oxytetracycline, sulfas
  • Stress and temperature are important factors

Question 12

Describe the effects of streptococcal infection in fish.

What are the Strep species that affect them?

How are they transmitted?

What fish are commonly affected?

What are the typical clinical signs?

How is it diagnosed? What samples should be collected?

What lesions are present on necropsy?

How is it treated?

Answer 12

• Streptococcosis
  
  • Emerging pathogens of bony fish – often causes meningoencephalitis, panophthalmitis and septicemia
  • Common cause of epizootics in wild and cultured fish
  • One of the few gram positive infections of fish
    
    • Steptococcus agalactiae and streptococcus iniae most common
  • Transmission typically horizontal through ingestion/contact, and feed can remain a source of infection after > 6 mo of freezing.
  • Disease reported from marine, brackish and freshwater fish, particularly teleosts
    
    • Majority from cultured fish - Tilipia, striped bass, coho salmon, rainbow trout, mullet, sturgeon, channel catfish
    • Less common reports from aquarium fish (rainbow sharks, tetras, cichlids) and rare reported from elasmobranchs (stingrays and shovelnose rays)
    • Common carp and koi may be resistant
  • Signs – can be from individual or multiple fish from one or more species
    
    • Clinical course acute or chronic – spiraling, circling, erratic swimming or loss of buoyancy common as bacteria tend to colonize the CNS. May lose their flight response
    • Reduced appetite, weight loss, darkening of skin, petechiae, erythema (ventrum and fin bases), ulcers, abscess-like swellings along the peduncle, exophthalmos, periocular/periorbital hemorrhage, corneal hypopyon, coelomic distention, spinal deformities, poor reproductive success
    • Large scale epizootics reported for S. iniae
  • Differentials for gram positive cocci in fish – Streptococcus, vagococcus, enterococcus, Lactococcus
  • Diagnosis – combination of cytology, histology and culture (brain or kidney best) .
    
    • Gram positive cocci that form chains may be seen on blood smear, effusions, CSF, tissue cytology or histology
    • Histo – granulocytic, pyogranulomatous or granulomatous inflammation in the CNS, vertebrae and eyes – meningoencephalitis, panophthalmitis and septicemia common
      
      • S. ictalurid in channel catfish unusual that it does not show trophism for the CNS – causes epidermal necrosis, myositis and arthritis
  • Treatment - Antibiotics often indicated – florfenicol approved for us in US
  • Rare zoonotic transmission reported – through inoculation of wounds leading to cellulitis, endocarditis, and arthritis

Problem 53 – Streptococcosis – Streptococcus iniae

• Host: Striped bass, tilapia
  
  • ZOONOTIC disease – nonhealing skin ulcers, arthritis, septicemia, meningitis
• Clinical Signs/Pathology:
  
  • Neurologic disease, can be asymptomatic
  • Exophthalmos with hyphemia is common – sometimes called popeye
  • Bacterial colonies in affected tissues of brain and kidneys
• Diagnostics:
  
  • Gram positive
  • Brain & kidney culture
• Treatment:
  
  • Amoxicillin, erythromycin, oxytetracycline, enrofloxacin
  • Vaccines are underdevelopment
• Gram-positive lactic acid bacteria - ZP
• Lactococcus garvieae, Streptococcus iniae, S. agalactiae, S. parauberis, S. dysgalactiae.
• Several, especially S. iniae are zoonotic.
  
  • S. iniae and S. agalactiae are responsible for major wild fish kills.
• Lactococcus garvieae first described in Japanese yellowtail/Asian mariculture.
• S. iniae major pathogen for loss in cultured and wild fish in FW, brackish and SW.
• S. agalactiae best known for multispecies wild fish kills in brackish water.
• S. parauberis and dysgalactiae most recently recognized.
  
  • S. parauberis in wild striped bass US.
  • S. dysgalactiae originally yellowtail and amberjack, also Nile tilapia.
• Primarily warm water pathogens.
• Influenced by environmental stress.
• L. garvieae and S. iniae, asymptomatic carriers important role in transmission.
• Transmission occurs horizontally.
  
  • Fecal-oral routes in water, cannibalism.
• Predilection for CNS.
  
  • Eyes, coelomic mesothelial surfaces, mild parenchymal involvement.
    
    • Hemolytic and cytolytic toxins damage tissues, avoid phagocytosis.
• Acute to subacute.
  
  • Typical signs of sepsis.
    
    • Some may cause chronic disease and granulomas i.e. strep iniae.
    • Anorexia, lethargy, loss of orientation, erratic swimming, exophthalmia, corneal opacity, skin darkening, hemorrhage.
    • Turbid to hemorrhagic ascites, fibrin, organomegaly with congestion.
    • Cloudy CSF, severe pyogranulomatous meningoencephalitis, hemorrhage panopthalmitis, epicarditis, coelomic polyserositis, cellulitis.
    • Enteritis reported with s. agalactiae and s. parauberis infections, hemoarrhagic enteritis is prominent feature of L. garvieae.
    • Muscle abscesses with S. dysgalactiae, also with S. iniae.
    • The respective diseases cannot be reliably distinguished solely based on histology.
    • Readily cultured, blood agars recommended to observe for hemolysis.

Question 13

What is the etiologic agent of bacterial kidney disease?

What species of fish are affected?

How is this disease transmitted?

What are the typical clinical signs?

How is it diagnosed? What samples need to be collected?

What lesions are present on necropsy?

How is it treated and prevented?

Answer 13

• Renibacterium salmoninarum
  
  • The cause of bacterial kidney disease in salmonids – slowly progressive systemic disease of wild and cultured salmonids
  • Histopath – granulomatous renomegaly with gram positive diplobacilli
  • Horizontal and vertical transmission reported with asymptomatic carriers common. Bacteria can survive for weeks in the environment
  • Affects freshwater and saltwater salmonids – pacific salmonid species are most susceptible (coho, chinook, sockeye, brook trout, artic char)
    
    • Disease most likely if < 1 yr old
  • Permissive water temperature is low with disease seen in fall and winter
  • Signs
    
    • Chronic disease that is slowly progressive – lethargy, poor growth, coelomic distention. May also seen darkening of skin, petechiae, erythema, ulcers, or pustules along the lateral line (spawning rash), exophthalmos
  • Diagnosis – based on necropsy in susceptible species and molecular diagnostics
    
    • Small gram positive diplobacilli on blood smears, effusions, cytology, or histology. Nonmotile. Stain positive with PAS stain.
    • Necropsy – renomegaly with granulomas. Granulomas may also be present in the spleen, liver, and heart. Cavernous lesions in the skeletal muscle adjacent to the kidney
      
      • Can culture from kidney – difficult to grow without kidney disease media (very slow to grow)
      • PCR is common choice, but LAMP may provide faster, simpler, cheaper results.
      • ELISA commonly used to screen broodstock in hatcheries
    • Medical management shows little success. Notifiable to the OIE.
      
      • Antibiotics may reduce morbidity and mortality if early in the disease process – signs often recur – macrolides (Erythromycin commonly used)
    • Prevention – biosecurity (isolation from other farms, all in all out, separation of generations. ELISA on ovarian fluid in broodstock prior to breeding.
      
      • Erythromycin of females pre-spawning can reduce transmission

Problem 54 – Bacterial Kidney Disease, Dee Disease – Renibacterium salmoninarum

• Host: Salmonids
• Clinical Signs/Pathology:
  
  • Lethargy, dark fish, granulomas in kidnesy, exophthalmos, blindness, emaciation
  • Obligate intracellular bacteria, granulomas with bacteria
• Diagnostics:
  
  • Gram positive rods seen in kidney imprints
  • Culture
• Treatment:
  
  • Amoxicillin, erythromycin, oxytetracycline, enrofloxacin
  • Erythromycin injections into female broodstock can reducd infection of eggs

Question 14

Describe mycobacteriosis in fish.

What species of mycobacterium commonly affect fish?

What species are fish are particularly susceptible? Which are more reistant?

How is this transmitted? What risk factors are there for transmission?

What are the typical clinical signs?

How is this diagnosed? What samples, stains, or media need to be used?

How are they treated and managed?

Describe the zoonotic infections associated with these pathogens.

Answer 14

• Mycobacteriosis
  
  • Common chronic disease of teleosts – caused by non tuberculous mycobacteria (NTM)
    
    • Mycobacterium fortuitum (freshwater), Mycobacteria marinum (saltwater and freshwater), and Mycobacterium chelonae (especially in zebrafish)– ubiquitous and common in water and biofilms
  • All fish susceptible – especially Cichlidae (cichlids), Syngnathidae (seahorse, pipefish, seadragons) and Cyprinidae
    
    • Some teleosts (catfish) appear relatively resistant to disease. Rare in elasmobranchs
    • Disease more common in adult and geriatric fish
  • Nonmotile, acid fast positive non-spore forming pleomorphic bacilli
  • Lesions are usually granulomatous (acid fast identified on culture and histo) and medical treatment rarely effective
  • Transmission – typically horizontal (ingestion of live/dead prey, picking at biofilm). Other routes include inoculation and immersion
    
    • NTM common in reptiles/amphibians and these can act as carriers or vectors
  • Risk factors – low pH, high humic and fulvic acids and contaminants (zinc, iron), high water temp
  • Signs
    
    • Often asymptomatic, but when seen is usually chronic and progressive
    • Inappetence, reduced appetite, weight loss, poor growth, lethargy, abnormal swimming, abnormal buoyancy, pigment changes (hyper and hypopigmentation), petechiae, ulcers, cutaneous nodules, firm/soft cutaneous masses, coelomic distention, exophthalmos, spinal deformities, low fecundity/breeding success. Sporadic mortality but epizootics possible
  • Ddx for acid fast granulomatous inflammation – mycobacterium, nocardia, myxozoans, cryptosporidium
  • Diagnosis – presumptive based on granulomas with acid fast bacilli. Definitive requires culture and PCR
    
    • Gram stain and diff quick often take up no stain (ghosting), but acid-fast shows pink to red beaded rods. Sensitivity of cytology higher than culture.
    • Necropsy/histology – organomegaly, granulomas, congestions, necrois (liver, spleen, kidneys, gonads)
      
      • If bacilli only found in GI may be transient
    • Culture possible with routine media but selective media (Lowenstein jenson, Middlebrook, Michison) preferred – growth and ease of culture varies by species
    • PCR recommended for slow growing species
  • Management
    
    • Reduce exposure and stressors – reduce water temp to reduce morbidity and mortality from M. marinum. With virulent strains consider repopulation and disinfection (remove all substrate, media and biofilms)
      
      • Disinfection – heat (140-250F, sunlight, chlorine, phenols, ethyl alcohol, UV
    • Medical treatment often not performed due to poor response and risk of resistance. Reported antibiotics – azithromycin, kanamycin, tobramycin, amikacin with multidrug therapy recommended Avoid drugs used in humans (isoniazid, rifampin, clarithromycin)
    • Prevention – routine monitoring, pasteurization of fish feeds, sentinel programs and disinfection of eggs and larvae in zebrafish colonies. Experimental attenuated live and DNA vaccines can provide some protection
    • Zoonosis – 1/3 of NTM are zoonotic – most common is M marinum in fish, transmitted by inoculation into wounds. Presents as nonhealing skin lesion on extremity (+/- papule, nodule, plaque, exudate)
      
      • Systemic infections possible with NTM species that can tolerate higher temp ( M. fortuitum, M. gordonae) usually in immune-suppressed individuals
        
        • Wear gloved when working with high-risk fish or systems
      • Mycobacteria within the tuberculous complex (M. tuberculosis and M leprae) not identified in fish

Problem 55 – Mycobacteriosis – Mycobacterium marinum

• Host: All fish
  
  • ZOONOTIC – nonhealing ulcers
• Clinical Signs/Pathology:
  
  • Chronic wasting, granulomatous disease
• Diagnostics:
  
  • Gram positive, acid-fast rods
• Treatment:
  
  • Cull, disinfect with phenolic compounds or alcohol
  • Treatment is rarely successful
• Mycobacteriosis and nocardiosis (ZP)
  
  • Chronic, progressive, debilitating, systemic diseases.
  • Gram-positive, acid-fast, or partially AF (nocardia).
  • Short to medium length bacilli
  • Initiate foreign body type host response, evades intraphagocytic lysosomal destruction.
  • Mycobacterium marinum, M. fortuitum, M. chelonae, M. peregrinum, M. abscessus, N. asteroids, N. seriolae, N. crassostreae, and N. salmonicida.
  • Infected water, biofilms, sediment, contaminated fish carcasses.
  • Infections may vary – host spp, virulence of strain, exposure magnitude, robustness of host immune system, length of infection, ambient water temperature.
  • Thin BCS with visceral organomegaly.
  • Parenchyma of spleen and kidney expanded and studded with gray-white nodules.
  • Discrete granulomas, coalescing foci of inflammation.
  • Common in sygnathids.
    
    • Common diagnostic tools – acid fast bacilli in smears, histo, bacterial culture, PCR.
    • Culture requires special agar, long incubation.
    • Persists in biofilms.
    • Chronic, low-level morbidity and mortality, primarily affects older specimens.
  • Atypical acid-fast bacterial infections can be zoonotic, especially for immune-compromised individuals.

Question 15

Describe nocardiosis in fish.

What species of nocardia affect fish?

What species of fish are particularly affected?

What are the typical clinical signs?

How is this diagnosed? What samples need to be collected?

What are the lesions on necropsy?

How is this treated or controlled?

Are these zoonotic bacteria?

Answer 15

• Nocardiosis
  
  • Similar to mycobacteriosis with a chronic presentation, systemic granulomatous inflammation and acid-fast positive rods, but Nocardia are long branching rods that look like fungal hyphae
  • Medical treatment rarely effective
    
    • N. seriolae (marine fish), N. asteroides (freshwater), N salmonicidia, N. crassostreae (fish and oysters)
  • Transmission – typically horizontal, though ingestion reported. Often found in invertebrates that may act as carriers or vectors
  • Disease possible in many freshwater and saltwater fish (ubiquitous in fresh and saltwater habitats) especially neon tetras and seahorses
  • Signs – typically individual fish affected – lethargy, inappetences, weight loss. Later signs include skin ulcers, nodules, or boils (furuncles), coelomic distention and intermittent mortalities
  • Diagnosis – presumptive diagnosis based on cytology and histology, though additional testing required for species identification (PCR, culture)
    
    • Long thin, acid fast +, branching rods on cytology or histology
    • Histo – white to yellow nodules due to granulomatous inflammation within the viscera, especially gills, heart, spleen, liver, kidney and swim bladder
  • Resistant to disinfect, benzalkonium chloride most effective
  • Treatment – rarely recommended due to poor response and risk of resistance. TMS effective in vitro. Long term therapy (several months) often indicated
  • Zoonosis – N. asteroids can cause systemic disease in immunosuppressed people. Nocardiosis typically transmitted by aerosolization of bacteria in soil, standing water, or decaying plants, presenting as slowly progressive pneumonia with cough. Can also be inoculated into wound or surgical site.
• Mycobacteriosis and nocardiosis (ZP)
  
  • Chronic, progressive, debilitating, systemic diseases.
  • Gram-positive, acid-fast, or partially AF (nocardia).
  • Short to medium length bacilli
  • Initiate foreign body type host response, evades intraphagocytic lysosomal destruction.
  • Mycobacterium marinum, M. fortuitum, M. chelonae, M. peregrinum, M. abscessus, N. asteroids, N. seriolae, N. crassostreae, and N. salmonicida.
  • Infected water, biofilms, sediment, contaminated fish carcasses.
  • Infections may vary – host spp, virulence of strain, exposure magnitude, robustness of host immune system, length of infection, ambient water temperature.
  • Thin BCS with visceral organomegaly.
  • Parenchyma of spleen and kidney expanded and studded with gray-white nodules.
  • Discrete granulomas, coalescing foci of inflammation.
  • Common in sygnathids.
    
    • Common diagnostic tools – acid fast bacilli in smears, histo, bacterial culture, PCR.
    • Culture requires special agar, long incubation.
    • Persists in biofilms.
    • Chronic, low-level morbidity and mortality, primarily affects older specimens.
  • Atypical acid-fast bacterial infections can be zoonotic, especially for immune-compromised individuals.

Question 16

What type of organism causes epitheliocystis?

How is this transmitted? What risk factors are there for transmission?

What species are particularly affected?

What are the typical clinical signs?

What differentials should be considered?

How is this diagnosed?

What lesions are seen on necropsy?

How is this treated?

Answer 16

• Epitheliocystis (CGFM)
  
  • Emerging bacterial disease often infecting fish gills – lesions appear as hypertrophied cells that look like cysts on histology
  • Obligate intracellular gram negative bacteria (chlamydia like organism)
  • Transmission – horizontal following rupture of infected cells
  • Chlamydia like organisms reported from multiple species of freshwater and marine teleosts, including aquarium leafy sea dragons and elasmobranchs
    
    • In Atlantic salmon, CLO possible etiology of proliferative gill inflammation seen after transfer to salt water.
  • Signs – fish often asymptomatic, when affected individual fish usually affected
    
    • Small, white spherical nodules on gills, oral cavity and skin often with no other signs. May also see lethargy, reduced appetite/growth, exophthalmos, abnormal swimming, skin ulcers, dyspnea. Morbidity and mortality highest in young fish and disease may be cyclic
  • Differentials for nodules on the gills include herpesviruses, papillomas, granulomatous inflammation, epitheliocystis or other CLOs, Mesomycetozoea, microsporidial xenomas (e.g. Loma spp.), Ichthyophthirius multifiliis, and Cryptocaryon irritans.
  • Diagnosis – tentative diagnosis based on cytology and histology of gills or skin. Definitive diagnosis requires PCR and sequencing
    
    • Histology of epithelial tissue shows large hypertrophied cells that resemble spherical cysts with a single intracytoplasmic, basophilic inclusion or vacuole
  • May be incidental with no treatment needed; CLOS often susceptible to macrolides (azithromycin) and tetracyclines (doxy, oxytet)
  • Not zoonotic
• Epitheliocystis - ZP.
  
  • 50+ spp, FW and SW, warm and cold.
  • Also reported in one chondrostean (white stuegeon) and several chondrichthyans.
  • Affects epithelium of gills and skin.
    
    • Gill infection more clinically important.
    • Gram-negative bacteria that occur intracellularly, in epithelial cells but also in chloride, mucous, and pillar cells, and MP.
    • Majority are in order Chlamydiales (chlamydia-like bacteria).
      
      • Candidatus Piscichlamydia salmonis, Candidatus Clavochlamydia cysticola, Candidatus, Syngnamydia salmonis, Candidatus Parilichlamydia carangidicola.
      • Also several instances of beta proteobacteria i.a. Candidatus Branchiomonas cysticola, BCJ epitheliocystis agent, and gamma proteobacterium Endozoicomonas elysicola have been ID in cases.
  • Can be incidental or cause high mortality.
  • E. elysicola is the only epitheliocystis-assoc bacteria to be isolated in vitro.
  • Koch’s postulates not fulfilled for any of these assoc bacterium.
  • Usually mixed bacterial infections.
  • Risk factors – environmental, management, host factors.
  • Histo – hypertrophied cells, containing intracytoplasmic, membrane-bound, vacuolar inclusions with basophilic, granular contents that displace host cell cytoplasm, often including nucleus, to cell periphery.
  • Gill – proliferative responses vary in severity and extent.
    
    • Hyperplasia, bridging of lamellar epithelium and fusion, hyperplastic epithelium across tips of consecutive lamellae.
    • Varying degrees of necrosis, goblet cell metaplasia, inflammation.
    • Determination of specific agent – PCR, gene analysis, in situ hybridization

Question 17

What organisms cause Francisellosis in fish?

What species are particularly affected?

How is this transmitted?

What are the typical clinical signs?

What differentials shoudl be considered?

How is this diagnosed?

What lesions are present on necropsy?

How is this managed and treated?

Answer 17

• Francisellosis
  
  • Emerging pathogens that can cause septicemia and granulomatous inflammation in multiple fish species
  • Francisella noatunensis subsp noatunensis and Francisella noatunensis subsp chilensis reported from cold water marine teleosts – serious pathogens of cultured Atlantic cod and salmon
    
    • Found in Europe and Chile
  • Francisella orientalis – warm water teleosts (freshwater, brackish and saltwater), serious pathogen in cultured tilapia
    
    • Americas, Asia, UK
  • Transmission – horizontal (fecal-oral) with long environmental stability. Invertebrates (mussels and crabs) may act as vectors.
  • Signs – clinical course can be acute or chronic with lethargy, inappetence, reduced feeding, poor growth, abnormal swimming (spiral/circling) seen. Erythema, petechia, or raised hemorrhagic nodules common. Can also see dark coloration, exophthalmos, white gill nodules, hyperplasia, or pallor
    
    • Acute form has mortality up to 90% with few clinical signs
  • Differentials for bacterial granulomatous lesions in teleosts include Mycobacterium, Nocardia, Edwardsiella, Francisella, Streptococcus, Photobacterium spp., and other piscirickettsial-like organisms
  • Diagnosis – relies on clinical signs and molecular testing
  • Small acid fast negative cocobacilii
    
    • Histo – splenomegaly and renomegaly with white-tan nodules (granulomas and intracellular bacteria) with adjacent muscle and skin often affected.
    • Culture – limited use, organism is fastidious and needs cysteine and iron to grow but specialized media can be used
    • PCR, qPCR, IHC on spleen or kidney available. Sequencing required.
  • Management – for F. orientalis, increase water temp > 82F to reduce mortalities
  • Treatment with antibiotics based on culture and sensitivity. In vitro, florfenicol and oxolinic acid have been recommended.
• Francisella - ZP
  
  • Morphologically similar to P. salmonis.
  • Marine and FW fish and giant abalone.
  • Warm water – Francisella noatunensis subsp orientalis.
  • Cold water – F. noatunensis subsp noatunensis.
  • Transmission horizontal, direct between fish or via contaminated water.
  • Extensive granuloma formation.
    
    • Systemic, spleen, kidney, liver, heart, gill, testes, muscle, brain, eye.
    • Gran negative bacteria observed in cytoplasm of MP in granulomas.
    • Dx – culture ID with gross and microscopic lesions.
    • PCR for identification.

Question 18

What is the etiologic agent of rickettsial septicemia in fish?

What fish are more susceptible?

How is this disease transmitted? Are there any risk factors?

What are the typical clinical signs?

How is this diagnosed?

What are the lesions seen on necropsy?

How is this managed and treated?

Answer 18

• Piscirickettsiosis
  
  • Piscirickettsia salmonis causes rickettsial septicemia- pacific salmonids most susceptible
    
    • P salmonis reported from salmona and brook trout, other piscirickettsial-like organisms from a variety of freshwater and marine fish
    • Risk factors for P. salmonis – transfer to salt water, Fall/Spring, infected stock nearby
  • Gram negative acid fast negative pleomorphic coccobacilli, facultatively intracellular
  • Transmission – horizontal, crustaceans may act as vectors
  • Signs – incubation period is weeks to months and several conspecifics often affected.
    
    • Lethargy, inappetence, dark coloration, gill pallor (anemia), erythema, petechiae, skin ulcers, fin erosions, nodules, coelomic distention
  • Diagnosis – Made by a combination of signs, cytology, histology, molecular testing (PCR)
    
    • Can find organisms on blood smear, effusion, cytology, histology
    • Histo – pathognomonic gray to yellow ring-shaped foci on the liver, serosanguinous coelomic effusion, tissue pallor, hepatomegaly, renomegaly with intracellular bacteria within macrophages
    • Culture limited as organisms are fastidious and need additional cysteine and low temperature
  • Management – response to medical treatment often limited likely due to intracellular nature, antibiotic resistance and reduced feeding
    
    • Commercial vaccines used but efficacy debated
    • Early harvesting (< 6 mo) after transfer to salt water or prior to high temp in summer may reduce risk

Problem 56 – Piscirickettsiosis, Salmonid Rickettsial Septicemia, Huita Disease – Piscirickettsia salmonis

• Host: Salmonids
• Clinical Signs/Pathology:
  
  • Anemic, skin ulcer, swollen abdomen
  • Yellow mucoid material in intestines, spleen and kidney enlargement
  • Vasculitis, hepatocyte necrosis
• Diagnostics:
  
  • Anemia (as low as 2%) is the most consistent sign
  • Tissue smears or sections are best – bacteria seen in hepatocytes and macrophages
  • Gram negative
  • Culture – requires cell culture, highly fastidious
• Treatment:
  
  • Antibiotics not very effective since this is an intracellular pathogen
  • Aminoglycosides, tetracyclines, erythromycin and quinolones have been used
• Piscirickettsiosis - ZP.
  
  • Piscirickettsia salmonis.
  • Gram – negative, nonmotile, aerobic, nonencapsulated, gastidious, typically coccoid intracellular bacteria.
  • SW farmed salmonids. Global. Also ID in seabass.
  • Significant mortality, marine net pens.
    
    • Typically 6-12 weeks after transfer to net pens, mortality 30-90%.
    • Coho salmon considered most susceptible.
    • Atlantic salmon more resistant.
  • Transmission is horizontal, marine reservoir suspected but not identified.
  • Class gammaproteobacteria (also includes Francisella, Coxiella, Legionella).
  • P. salmonis secretes cytotoxic extracellular products in vitro.
  • Characteristic lesion – systemic, septicemic dz with lesions in multiple organs, most severe in liver, kidney, spleen, and intestine.
    
    • Most consistent gross finding – pallor of gills (severe anemia).
    • Hct may be as low as 2% (normal 35-50%).
  • Peripheral smears have high numbers of MP containing bacteria or cellular debris in intracytoplasmic vacuoles.
  • Gross lesions – renomegaly, splenomegaly, ascites, hemorrhages in fat, stomach, SB, skeletal muscle.
  • May have circular cream-colored lesions throughout liver.
  • Necrosis is principal pathologic change followed by granulomatous inflammation in chronic phase.
    
    • Anemia caused by necrosis in renal hematopoietic intesrstitium and spleen.
    • Necrotizing hepatitis with vasculitis and thrombosis.
    • Thrombosis throughout organs.
  • Dx based on recognition of lesions and observation of intracellular gram-negative bacteria.
    
    • Confirmation with IHC, culture isolation and labeling with fluorescent AB, or PCR.
    • Tissues of choice for isolation – kidney, liver, blood.