Tracking down a pathogen

TLDR

If you have owned goldfish for any amount of time you have experienced weird mysterious diseases.  The forums are full of people asking for a diagnosis and a few angelic people willing to help them.  I consider myself an advanced hobbyist, I do my water changes, I test when necessary, I cycle my tanks, I know my stuff.  Avoid my mistakes, do not reuse filter media, it is cheap just replace it.  There is good information and bad information online, there are experts and people who think they are experts, it is hard to separate the wheat from the chaff.  Seek out professional assistance when necessary.

http://www.cahfs.ucdavis.edu/test_fees/index.cfm

The Beginning

I was burnt out on planted tanks and decided to turn my 200 gallon planted tank into a goldfish “pond”, less heating, less lighting, less maintenance, less cost, less energy expenditure all around.  I ripped the tank apart, kept my filter cycled and started the hunt for goldfish.  I quickly found and settled on some very nice Jikins, they looked like regular fish with fancy tails, I was not into the real oddballs yet, perfect.

My first problem.  The waste output from my tiny planted tank fish was nothing compared to goldfish and my precycled filter really needed to cycle again, damnit, should have seen that one coming.  I dealt with the cycling issues and onto the next problem.  My Jikins had what appeared to be bruising, odd but maybe some latent shipping damage.  A few of them were also kind of lethargic, this was no good.  So into the quarantine tank we go because I don’t want to medicate 200 gallons of water, that would be crazy.  After lots of clean water and I believe some API General cure that I had on hand (Metro and Prazi), they came around and everyone was better for a while.  Sick goldfish are par for the course, I thought the worst of it was over.

Living with it

My goldfish fish addiction flourished and I started ordering fish from all over the place, this is not advisable but, I quarantine, no problem.  It is all sort of a blur now but I had Ryukin, Ranchu, Oranda, Tamasaba, imports, domestic, all kinds of fish.  My super cool brand new broad tail tiger Ryukin made it through quarantine and into the main tank then a month later he got sick and had to be removed and treated.  I dosed the main tank with prazi because that is what everyone says to even though I don’t think I was dealing with parasites.

A month later after a heatwave I had ich.  I treated the main display with high heat and high salinity and that knocked it out.
Then my favorite Jikin got sick again, treated in QT but he did not make it.

A month or so later one of my Ranchu started acting up, off to QT for treatment, all fixed up, back to the main tank.
Then a different Ranchu look slightly more pineconey than he should, off to QT for a round of metro and kana.  After not seeing any improvement with these meds or Epsom salts I assumed this was normal for him and it was just my eyes playing tricks on me.  As long as he was acting normal he was cleared for return back to the display tank.

Another Jikin got sick, floaty problems this time, QT’d and treated with Epsom salt, fasting, followed by a duckweed diet, all fixed up and back to the main tank.

Then a different Jikin fell ill, this time I thought, no meds just good ol’ fashion clean water and salt.  Maybe I was killing my fish with all the meds? Nope that did not help and he died.

Everyone was fine for a while then a different Ranchu fell ill.

This was all becoming too much, something was going on and about every month or two someone would get sick.  They would get better in QT, then back to the main tank.  I decided to treat the main tank with Paraguard, this did not help.  Then I treated with Metronidazole and Kanamycin, this is the combo that had been working best for me in QT.  I even tried mixing it into their food to target any internal bacteria or parasites.  I had already treated the tank with salt for ich so that did not help whatever mystery bug I had on my hands

A quick aside.  Kanamycin is a broad-spectrum antibiotic and antifungal.  Metronidazole treats parasites and anaerobic bacteria.  I was really trying to cover a lot of bases because I did not know what I was dealing with.  I did a lot of online research and reached out for help on the forums.   At some point I stopped asking for help because I realized the people helping could not diagnose my problem online and I got tired of being asked for my water parameters.

Just to add insult to injury, during one course of treatment I killed off the beneficial bacteria in my biological filter and had to cycle my tank again.

I’ve had enough

So now what?  Is this just a string of bad luck and multiple different problems from mixing fish from different sources?  Is this mystery pathogen a figment of my imagination?  How to get to the bottom of this? 

I did some digging and calling around and found that UC Davis has a fish vet and a lab.  I was not interested in driving my fish up to Davis and paying for a vet visit.  I got in touch with the lab and for a very reasonable price I could send them 3 fish for necropsy.  The process was straight forward.  I euthanized 3 sick fish with clove oil, packed them with ice packs, and overnighted them to them the lab.  They provided some initial findings within a few days then further information when they received the results back from their tests.  Finally, I had some hard facts, tests, labs, stains, people with PhDs who did not treat me like an idiot.

The Diagnosis.

Disseminated mycobacteriosis with:
Red Ranchu – A: 1. Kidney, mesentery, liver, pancreas, heart, intestinal serosa, and ovary: Granulomas, multiple with necrotic and cellular debris and /or mineral, and acid-fast bacilli. Other Findings 2. Gill: Branchitis, proliferative and lymphohistiocytic, marked, diffuse 3. Coelomic cavity: Serositis / coleomitis, pleocellular, moderate and luminal serous fluid. 4. Integument and skeletal muscle: Myonecrosis, multifocal, moderate with interstitial lymphohistiocytic myositis and focal epidermal ulceration

Calico Ranchu – B 1. Ovary, pancreas, mesentery and liver: Granulomas, multiple with rare numbers of acid-fast bacilli. Other Findings 2. Liver: Hepatocellular lipid-type vacuolar change, diffuse. 3. Gill: Branchitis, proliferative and lymphohistiocytic, moderate, diffuse 4. Coelomic cavity: Serositis / coleomitis, pleocellular, moderate and luminal serous fluid.

Butterfly telescope – C 1. Kidney: Granulomas, few with rare acid-fast bacil

Other Finding

 2. Gill: Branchitis, lymphohistiocytic, multifocal, mild. Ancillary tests

Negative for koi herpes virus [gill] – PCR.

Rare to moderate numbers of Aeromonas hydrophila, Aeromonas sp., and Shewanella putrifaciens isolated [liver, kidney and gill] - A, B and C.

Large numbers of Aeromonas sp and Vibrio sp. isolated [liver] – C

Case Summary

Findings are similar in the red and calico Ranchu and consist of granulomas in multiple organs. No etiologic agents were seen in tissue sections on routine H & E stains; a likely differential is Mycobacterium sp. infection and special stains are pending. In these fish there was also significant proliferative and lymphohistiocytic branchitis suggestive of alterations in the water column, and moderate serositis / coelomitis. Lesions in the butterfly telescope were mild and consist of few granulomas in the kidney, and branchitis. Tissues were negative for koi herpesvirus. Bacteriology and additional histology to follow. Submitted water samples were held in case further testing is required. 3/12/18. Acid-fast bacilli were seen in granulomas in multiple organs in both Ranchu and in the renal granulomas in the butterfly telescope, consistent with Mycobacterium sp. infection. Bacterial isolates are likely normal flora in aquatic environment and may have contributed to the branchitis noted. All testing complete.

So uh… dumb it down for me doc.

I am not a scientist, I am not a doctor, I do not even pretend to know what all the above diagnosis and case summary mean.  I had a couple phone calls with the doctor and what I have settled upon is that I have a strain of mycobacterium in my tank that is particularly nasty and there is no treating it.  According to wikipedia there are over 190 recognized species of mycobacterium*.  Fish TB is caused by mycobacterium.

Wait whut?

Fish TB?  At one point a long time ago when I was working with wild caught Sawbwa resplendens I thought I was dealing with internal parasites or a bacterial infection that was responsive to API General Cure (metro), and at some point I thought maybe I was dealing with fish TB.  Sawbwa resplendens are wild caught in Burma and do horrible in captivity which is why I was breeding them.  They always seemed to die off in my tank, I had some success breeding them but eventually gave up.  Of course I did not sterilize my tank before switching to goldfish, hell I even used the same filter media.  Damnit!  Mystery solved as far as I am concerned.

Now what?

I reached out to some of the people that I really respect in the goldfish community to get their opinions before I made my decision.  If you are a normal person with a fish tank and some pet fish and you have mycobacteriosis you keep your water clean, feed a healthy diet, and when your fish start to get really sick you euthanize them.  I am not a normal person, I am breeding goldfish and I do not want to risk transferring this strain of mycobacteria to my breeding stock so I decided to euthanize all my fish from that tank.  It was a really bad day to say the least but I could not risk spreading this disease around.  I threw away all the plants, all the filter media, basically everything that could not be sterilized or was easy to replace.  The tank is dry, the filter is dry, the parts of the filtration and tank that I am keeping have been through a bleach treatment and are awaiting a treatment with Virkon**.  After that it will sit dry for a few more weeks.  Once it is up and running I will run another round of Virkon through the water column.  Big water change, then cycle, then fish.

Closing

This whole ordeal lasted about a year, it was stressful and cost a ridiculous amount of money.  Learn from it and move on, it is all you can do.

 

* https://en.wikipedia.org/wiki/Mycobacterium

King HC, Khera-Butler T, James P, Oakley BB, Erenso G, Aseffa A, Knight R, Wellington EM, Courtenay O (2017) Environmental reservoirs of pathogenic mycobacteria across the Ethiopian biogeographical landscape. PLoS One 12(3):e0173811. doi: 10.1371/journal.pone.0173811

** https://pentairaes.com/aquatic-disinfectant.html?gclid=Cj0KCQjwkKPVBRDtARIsAA2CG6HJeNfXlWX2cKO6rCOQwyf7fp4LkPpmI9GYK1KL-p0SB-f9iRU-VNwaAthgEALw_wcB

 

 

FINAL REPORT

Specimens Received:      3 Carcass;This report supersedes all previous reports for this case

Date Collected: 03/01/2018     Date Received: 03/02/2018

Comments:    UPS - Owner paid $120.  Chk:#  391

 

C a s e   C o n t a c t s

 

 

Submitter Xxxxxxxxxx, Xxxxxxxxxx

                                     

Xxxxxxxxxx

CA   xxxxxxxxxx

 

S p e c i m e n   D e t a i l s

 

 

Animal/Source

ID Type

Taxonomy

Gender

Age

XXXXXXXXXX-01

CAHFS Internal ID

Fish

Unknown

 

 

L a b o r a t o r y   F i n d i n g s / D i a g n o s i s

 

 

Three female fish submitted with history of lethargy and lack of appetite.

Disseminated mycobacteriosis with:

Red ranchu – A:

1.  Kidney, mesentery, liver, pancreas, heart, intestinal serosa, and ovary: Granulomas, multiple with necrotic and cellular debris and /or mineral, and acid-fast bacilli.

Other Findings

2.  Gill: Branchitis, proliferative and lymphohistiocytic, marked, diffuse

3.  Coelomic cavity: Serositis / coleomitis, pleocellular, moderate and luminal serous fluid.

4.  Integument and skeletal muscle: Myonecrosis, multifocal, moderate with interstitial lymphohistiocytic myositis and focal epidermal ulceration.

Calico ranchu – B

1.  Ovary, pancreas, mesentery and liver: Granulomas, multiple with rare numbers of acid-fast bacilli.

Other Findings

2.  Liver: Hepatocellular lipid-type vacuolar change, diffuse.

3.  Gill: Branchitis, proliferative and lymphohistiocytic, moderate, diffuse

4.  Coelomic cavity: Serositis / coleomitis, pleocellular, moderate and luminal serous fluid.

Butterfly telescope – C

1.  Kidney: Granulomas, few with rare acid-fast bacilli.

Other Finding

2.  Gill: Branchitis, lymphohistiocytic, multifocal, mild.

Ancillary tests

* Negative for koi herpes virus [gill] – PCR.

* Rare to moderate numbers of Aeromonas hydrophila, Aeromonas sp., and Shewanella putrifaciens isolated [liver, kidney and gill] - A, B and C.

* Large numbers of Aeromonas sp and Vibrio sp. isolated [liver] - C.

C a s e   S u m m a r y

Findings are similar in the red and calico ranchu and consist of granulomas in multiple organs. No etiologic agents were seen in tissue sections on routine H & E stains; a likely differential is Mycobacterium sp. infection and special stains are pending. In these fish there was also significant proliferative and lymphohistiocytic branchitis suggestive of alterations in the water column, and moderate serositis / coelomitis.

Lesions in the butterfly telescope were mild and consist of few granulomas in the kidney, and branchitis. Tissues were negative for koi herpesvirus.

Bacteriology and additional histology to follow.

Submitted water samples were held in case further testing is required.

3/12/18. Acid-fast bacilli were seen in granulomas in multiple organs in both ranchu and in the renal granulomas in the butterfly telescope, consistent with Mycobacterium sp. infection.

Bacterial isolates are likely normal flora in aquatic environment and may have contributed to the branchitis noted.

All testing complete.

C l i n i c a l  H i s t o r y

All 3 fish have been in my care for at least a year, the 2 Ranchu are domestically bred and the Butterfly is an import.  My tanks hold a mix of imported and domestic fish.  I have 2 tanks with seemingly related problems.  Tank 1 200gal, weekly 50% water change, 14 fish all ~3-6".  Tank 2 90 gallons, weekly 50% water changes, 7 fish all under 3-6".  Both tanks 68F currently the gold ranchu is the most ill.  Symptoms:  Lethargy, lack of appetite.  The calico ranchu and brown butterfly telescope are eating but are also lethargic and sitting on the bottom of the tank. 

Duration of illness:  Chronic

Disease suspected:  Subacute mycobacterium

Treatments to date have been:  Ich-raise temp to 86 and maintain .2% salt for 7 days, kanaplex (kanamycin) - 1 tsp/20gal 5 days wc and redose every 48 hours, metronidazole-both in the water and in the food on different occasions , 1/4tsp/20 gal every 24hr treat 10 days 25% wc between each dose, food-1/2c food 1c water, 2tsp metro, PraziPro-1tsp/20gal <5% praziquantel by weight. I have a few theories:  1) reused filter media from wild caught sawbwa resplendens has given me a nasty internal parasite or resistant bacteria.  2) Imported Goldfish are immune to and carrying nasty bugs that my domestic fish have not been exposed to do not have immunity to.  3) Poor genetics and coincidence appearing to be chronic illness. 4) Bad advice on treatments to date and I have something totally curable.

G r o s s  O b s e r v a t i o n s

Three [3] female fish were submitted for necropsy which commenced at 1200 hours on 2nd March, 2018.

A: 68g, red ranchu, total length 11cm and fork length 9cm

B: 63.1g, calico ranchu, total length 11.5cm and fork length 10cm.

C: 42.7g, butterfly telescope, total length 12cm and fork length 10cm.

All three fish were in good post-mortem condition, and fish A and B had pendulous abdomens with increased amounts of serous fluid. No parasites were seen on microscopic examination of gill clip, and skin and intestinal scrappings. Fish A had multiple, variably sized white nodules in mesentery, along intestinal serosa, in liver and kidney; similar but fewer nodules were present in liver and ovary of fish B. The butterfly telescope had large amounts of fine sand in the intestinal tract. No gross lesions were seen in other viscera and brain.

B a c t e r i o l o g y

BACTERIAL AEROBIC CULTURE

Animal/Source             Specimen

Specimen Type

Results

 

XXXXXXXXXX-01

A

Liver Tissue

Aeromonas hydrophila Rare#

XXXXXXXXXX-01

A

Kidney Tissue

Aeromonas hydrophila Rare#

Aeromonas sp. Rare#

XXXXXXXXXX-01

B

Kidney Tissue

Aeromonas hydrophila Sm#

XXXXXXXXXX-01

A

gill

Aeromonas sp. Sm#

XXXXXXXXXX-01

B

gill

Aeromonas hydrophila Mod#

XXXXXXXXXX-01

C

gill

Aeromonas hydrophila Mod#

Shewanella putrifaciens Mod#

XXXXXXXXXX-01

C

Kidney Tissue

Aeromonas hydrophila Sm#

XXXXXXXXXX-01

B

Liver Tissue

Aeromonas hydrophila Mod#

XXXXXXXXXX-01

C

Liver Tissue

Aeromonas sp. Lg#

Vibrio sp. Lg#

 

 

B i o t e c h n o l o g y

Koi Herpes Virus, DNA, qtPCR

Animal/Source             Specimen

Specimen Type

Results

 

XXXXXXXXXX-01         A

gill

Negative

XXXXXXXXXX-01         B

gill

Negative

XXXXXXXXXX-01         C

gill

Negative

 

 

H i s t o l o g y

           

Tissues examined include brain, gill, heart, liver, spleen, kidney, pancreas, esophagus, stomach, intestine, mesentery, ovary, and swim bladder. Findings are described below:

Fish A – red ranchu

Kidney: There are multiple, variably sized granulomas in melanomacrophage centers and scattered randomly throughout the parenchyma. Individual granulomas consist of nodular aggregates of macrophages, or necrotic and cellular debris and cholesterol clefts, or debris and mineral, surrounded by a collar of flattened macrophages several layers thick. Similar granulomas are present in the pancreas, liver and intestinal serosa.

Mesentery: There are large numbers of granulomas throughout the mesentery and moderate pleocellular infiltrates in interstitium between and around granulomas. 

Heart: Solitary necrotic granuloma in myocardium.

Ovary: Moderate, numbers of discrete to coalescent necrotic granulomas.

Gill: There is lamellar epithelial hyperplasia with bridging along the tops of consecutive lamellae and, moderate interlamellar infiltrates of lymphocytes and macrophages. There are similar infiltrates in connective tissue and musculature of the gill arch.

3/12/18. Additional tissues examined include integument, skeletal muscle, vertebrae, spinal cord, eye [globe and lens], semicircular canal, and cranial vault.

Skeletal muscle and fascia: Multiple foci of myofiber degeneration and necrosis accompanied by mild to moderate interstitial infiltrates of lymphocytes and macrophages that extend into surrounding fascia.

Integument: Focally extensive epidermal ulceration in skin overlying the head with moderate lymphohistiocytic infiltrates in exposed dermis and extending into musculature.

Ziehl-Nelsen stain: Small numbers of acid-fast bacilli among cellular debris in some of the granulomas.

Gram stain: Small numbers of gram-positive short rods among cellular debris in granulomas. PAS stain: No fungal hyphae seen in tissue sections.

 

B:

Ovary: Multiple, variable sized, discrete to coalescent granulomas, with moderate pleocellular infiltrates and melanomacrophages in interstitium between granuloma.

Pancreas and mesentery: Interstitium is markedly expanded by large numbers of discrete to coalescent granulomas with extensive pleocellular infiltrates in interstitium between and around the granulomas. 

Kidney: Small numbers of granulomas primarily in melanomacrophage centers.

Liver: Diffuse lipid-type vacuolar change, and small numbers of granulomas randomly throughout the parenchyma. 

Gill: Multifocal lamellar epithelial hyperplasia with bridging, and moderate lymphohistiocytic interlamellar infiltrates. There are extensive infiltrates at the base of the lamellae.

3/12/18. Additional tissues examined include integument, skeletal muscle, spinal cord, eye [globe and lens], semicircular canal, and cranial vault. No lesions identified.

Ziehl-Nelsen stain: Rare numbers of acid-fast bacilli among cellular debris in some of the granulomas.

Gram stain: Few gram-positive short rods among cellular debris in granulomas. PAS stain: No fungal hyphae seen in tissue sections.

C:

Kidney: There are few small granulomas in melanomacrophage centers. Gill: Mild, multifocal, lymphohistiocytic infiltrates in lamellae.

3/12/18. Additional tissues examined include integument, skeletal muscle, spinal cord, thymus, pseudobranch, eye [globe and lens], semicircular canal, and cranial vault. No lesions identified.

Ziehl-Nelsen stain: Rare numbers of acid-fast bacilli among cellular debris in few granulomas.

Gram stain: Few gram-positive short rods among cellular debris in granulomas. PAS stain: No fungal hyphae seen in tissue sections.


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