INVESTIGATIONS IN FISH CONTROL

99. Evaluation of 215 Candidate Fungicides for Use in Fish Culture

OCH 3

UNITED STATES DEPARTMENT OF THE INTERIOR 

FISH AND WILDLIFE SERVICE
Investigations in Fish Control, published by the Fish and Wildlife Service, include reports on the results of work 

at the National Fisheries Research Center at LaCrosse, Wisconsin, and reports of other studies related to fish control 

and acute toxicity of piscicides. Though each report is regarded as a separate publication, several may be issued 

under a single cover, for economy. See inside back cover for list of current issues.

Copies of this publication may be obtained from the Publications Unit, U.S. Fish and Wildlife Service, 18th 

and C Streets, NW., Mail Stop 1111, Room 130, Arlington Square Building, Washington, DC 20240, or may 

be purchased from the National Technical Information Service (NTIS), 5285 Port Royal Road, Springfield, 

VA 22161.

ISSN 0565-0704
INVESTIGATIONS IN FISH CONTROL

99. Evaluation of 215 Candidate Fungicides for Use in Fish Culture

By Tom A. Bailey and Susan M. Jeffrey

U.S. Fish and Wildlife Service 

National Fisheries Research Center LaCrosse

P.O. Box 818 

LaCrosse, Wisconsin 54602

1989

Evaluation of 215 Candidate Fungicides 

for Use in Fish Culture

by

Tom A. Bailey and Susan M. Jeffrey

U.S. Fish and Wildlife Service 

National Fisheries Research Center LaCrosse

P.O. Box 818 

LaCrosse, Wisconsin 54602

ABSTRACT. The fungicidal activity of 215 compounds was evaluated by comparing their effectiveness with that of malachite green. More than half were found to be unsuitable as aquatic fungicides 

in preliminary screening tests because of their lack of activity against fungi, toxicity to fish or eggs, 

insolubility in water, or potential carcinogenicity. After further testing, 30 compounds were selected 

for evaluation of their activity against fungi on eggs of rainbow trout (Oncorhynchus mykiss). Four 

compounds 8-quinolinol, 8-quinolinol sulfate, dichlorophen, and formalin inhibited fungal growths 

on dead and live fish eggs and demonstrated the greatest potential as replacements for malachite 

green. The quinolinols cannot be considered for use as aquatic antifungal agents, however, because 

they were toxic to eggs at efficacious concentrations (>1.0 mg/L). Dichlorophen, at concentrations 

of 5.0 mg/L, was effective for eggs of channel catfish (Ictaluruspunctatus) that had been separated, 

but was toxic for eggs clustered in the original egg mass; it showed potential for use on channel 

catfish eggs at 2.5 mg/L for 15 min, but was only marginally effective against fungi on rainbow 

trout eggs. Formalin controlled fungal infections on rainbow trout eggs at a concentration of 250 mg/L 

for 60 min a concentration substantially lower than has been previously reported.

Fishes are commonly parasitized by various species of 

aquatic fungi that cause fungal infections or saprolegniasis; 

several species of the pathogenic fungi may occur together 

on a single infected fish. Saprolegniasis is usually considered to be secondary to bacterial or viral infections 

(Richards 1977; Richards and Pickering 1978), but there 

is evidence that some of the Saprolegniales may act as 

primary pathogens (Neish 1977; Richards and Pickering 

1978). Fungal infections often follow physical damage 

to the surface of fish (Roberts and Shepherd 1974; 

Richards and Pickering 1978) caused by handling and 

intensive culture conditions. In the wild, fungal infections 

develop in abrasions received during territorial defense, 

redd digging, or spawning (White 1975; Richards and 

Pickering 1978).

Fungal infections of hatchery-reared fish can usually 

be controlled with formalin (Neish and Hughes 1980; 

Bailey 1984) or malachite green (Meyer and Hoffman 

1976; Bailey 1983a). Formalin is the only registered

aquatic fungicide, but it is generally not completely effective on adult fish. The antifungal activity of malachite 

green is extremely high, but it is unlikely to be approved 

(registered) by the U.S. Food and Drug Administration 

(Schnick and Meyer 1978) because of its potential 

teratogenicity (Meyer and Jorgenson 1983). Its use is now 

limited to the treatment of nonfood fish (e.g., eggs or adult 

salmon held for spawning) under an Investigational New 

Animal Drug Application held by the U.S. Fish and 

Wildlife Service.

Because of the status of these two fungicides and 

the substantial monetary losses that result from fungal 

infections in fish hatcheries, fish culturists need an 

effective registered fungicide to replace malachite 

green. Our purpose here is to report on the efficacies 

of a number of potential fungicides against three 

species of aquatic fungi and to evaluate their potential 

for approval and future use on fish and incubating 

fish eggs.

1
INVESTIGATIONS IN FISH CONTROL 99

Materials and Methods

Pure strains of aquatic fungi were obtained from the 

American Type Culture Collection (ATCC). Saprolegnia 

hypogyna (ATCC 28275) and Achlya flagellata (ATCC 

14566) were used for in vitro testing (preliminary screening and determination of minimum inhibitory concentration) and Saprolegnia ferax (ATCC 36146) was used to 

infect eggs of rainbow trout (Onchorhynchus mykiss) for 

confirmatory and in vivo tests.

Test procedures used were those developed by Bailey 

(1983a,b; 1984). The method involves an in vitro screening technique modified from that of Golden and Oster 

(1947) and a minimum inhibitory concentration determination based on the percent inhibition of growth in diameter 

of colonies. Cultures of fungi on agar were exposed to 

five concentrations of each candidate compound for 

15 and 60 min.

An egg to agar transfer test was conducted to establish 

the concentrations that would control fungal growth on 

a natural substrate (dead rainbow trout eggs) for 48 h 

(Bailey 1984). Dead (cold shocked) rainbow trout eggs 

were exposed to chemicals at concentrations of 10 to 

100 times their minimum inhibitory concentrations. The 

egg to agar transfer provides a confirmatory test and is 

representative of the in vivo activity of a compound. 

Levels of inhibition are the concentrations that prevent 

growth of fungi on either the egg surface or on agar.

The most promising candidate compounds were tested 

on incubating (green) rainbow trout eggs received from 

Trout Lodge, McMillin, Washington, and Erwin National 

Fish Hatchery, Tennessee. Duplicate groups of 500 eggs 

were artificially infected with S. ferax and exposed to 

candidate fungicides in Heath incubator trays for 15 or 

60 min. Treatments were administered three times weekly 

for 2 weeks. Two control groups of eggs were used for 

each study: one was uninfected with fungi and untreated 

with fungicide; the other was infected with fungi but untreated. Upon completion of the testing, the degree of 

fungal infection and the number of eggs that hatched were 

analyzed to determine the efficacy of each chemical. The 

data were then compared to those for malachite green, 

the reference compound. A chi-square test (a = 0.05, 

df = 1) was used to determine whether treatments with 

the candidate fungicides caused significant differences in 

rates of infection and hatching.

Results

A total of 215 compounds were tested to determine 

antifungal activity against the three pure-strain species of

pathogenic aquatic fungi. Of these, 120 were considered 

unsuitable as aquatic fungicides because they were inactive against the fungi, toxic to fish or eggs, insoluble in 

water, or potentially carcinogenic (Table 1; Tables 1-6 

follow the text). Although four copper-8-quinolinolate, 

crystal violet, Du-Ter, and Phaltan were highly effective against the fungi, all were toxic to fish and fish eggs 

at the effective levels or were potentially teratogenic (e.g., 

crystal violet, a chemical homologue of malachite green). 

The minimum inhibitory concentrations of candidate compounds were based on fungal growth (colony diameter) 

and categorized in three levels of activity (Tables 2-4): 

high (<10 mg/L), moderate (>10 to 100 mg/L), or low 

(>100 mg/L).

Before in vivo testing, the most promising compounds 

were evaluated further in confirmatory tests (Table 5). 

Only four of the candidate compounds adequately inhibited fungal growth on incubating rainbow trout eggs 

in in vivo tests (Table 6): dichlorophen (2.0 mg/L), formalin (250 mg/L), 8-quinolinol (30.0 mg/L), and 

8-quinolinol sulfate (35.0 mg/L).

The proportions of rainbow trout eggs infected were 

all significantly lower than in the untreated controls after 

15-min exposures to dichlorophen (45%, % 2 = 13.3), 

formalin (37%, x 2 = 30.4), 8-quinolinol (83%, x 2 = 

146.2), and 8-quinolinol sulfate (20%, x 2 = 40-8). 

Likewise, inhibition after 60-min exposures was significant (dichlorophen: 48%, x 2 = 7.8; formalin: 15%, x 2 

= 261.9; 8-quinolinol: 30%, x 2 = 24.8; 8-quinolinol 

sulfate: 20%, x 2 - 28.3). Hatching success was highest 

after treatments with dichlorophen and 8-quinolinol 

sulfate (Figure).

100-

90-

o

z 80-

T~

o 70-

^

x 60-

Q

z 50-

<

5 ^0-

LJ

^ 20-

10- 

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i

V

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-

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ES 56 HATCH- 15MIN 

ESJ % HATCH-60MIN

1

1

I , MG FORM DICH 8-Q-S 

CANDIDATE FUNGICIDES

8-Q

Figure. Relative percent fungal infection of rainbow trout eggs 

after treatment with candidate fungicides and resulting hatching success. MG = malachite green; FORM = formalin; 

DICH = dichlorophen; 8-Q-S = 8-quinolinol sulfate; and 8-Q 

= 8-quinolinol.
EVALUATION OF CANDIDATE FUNGICIDES

Channel catfish eggs that had been separated and exposed to 5.0 mg/L dichlorophen for 15 min showed only 

2% fungal infection, and hatching success was about 91 %. 

When eggs were exposed to 2.5 mg/L dichlorophen, the 

fungal infection rate rose to 5.0% but hatching rate for 

the uninfected eggs improved to 96%.

Eggs of channel catfish (Ictalurus punctatus), which 

were allowed to remain in the egg mass, were exposed 

to 5.0 mg/L and 2.5 mg/L dichlorophen. Treatment with 

5.0 mg/L dichlorophen killed 98% of the eggs; however, 

treatment with 2.5 mg/L dichlorophen resulted in 20% 

infection and 88% hatching success.

Of the candidate fungicides used in 60-min exposures, 

formalin at 250 mg/L provided the best control of fungal 

growth on rainbow trout eggs (only 15% infected) and 

yielded an acceptable level of hatching (>65%) after a 

15- or 60-min exposure (Figure). In order of decreasing 

efficacy, the compounds ranked as follows: formalin 

> 8-quinolinol sulfate > dichlorophen > 8-quinolinol.

Discussion

In laboratory tests, the powerful antifungal agent malachite green was 5 to 30 times more effective than all 

except 5 of the 215 candidate compounds: copper-8- 

quinolinolate, crystal violet, Du-Ter (47%), Phaltan, and 

8-quinolinol. Malachite green thus provides an excellent 

reference standard with which candidate fungicides can 

be compared. The registration of malachite green is 

unlikely, although limited use of the compound might be 

acceptable if suitable filtration units are used (Marking 

et al. 1989).

The 8-quinolinol compounds controlled fungi in situ; 

however, the parent compound and the water soluble salts 

had detrimental side effects: At efficacious concentrations, 

8-quinolinol reduced the hatching to unacceptable levels 

and 8-quinolinol sulfate caused posthatching mortality.

Dichlorophen, an anthelmintic (Shah et al. 1984), has 

shown some potential for controlling pathogenic fungi 

(Mussa and Russell 1977; Alderman and Polglase 1984) 

and protozoans (Takeuchi et al. 1985; Griffin 1989). 

Alderman and Polglase (1984) reported that the sodium 

salt of dichlorophen is effective in vitro against fungi at 

1.0 mg/L. We did not test the sodium salt of dichlorophen, 

but the minimum inhibitory concentration for the phenol 

form of the compound was 7.5 mg/L. Perhaps water 

solubility enhances the toxicity.

In an unpublished report on preliminary tests with channel catfish eggs, B. R. Griffin 1 noted that 15-min immersions in 5 or 10 mg/L of dichlorophen for 4 consecutive

days arrested the spread of fungal growth, and resulted 

in no evidence of pre- or posthatching toxicity. The results 

of our tests of separated channel catfish eggs exposed to 

dichlorophen generally agreed with these observations. 

In contrast, dichlorophen was extremely toxic when we 

used it on eggs clustered in the egg mass.

In in vivo tests with fungused rainbow trout eggs, 

dichlorophen was only marginally effective. In preliminary toxicity tests, groups of rainbow trout eggs were 

subjected to 15-min dips in solutions of dichlorophen for 

4 consecutive days. Within 48 h after treatments, mortality was high. Dichlorophen is also toxic to fish. The 

LC50 values for channel catfish and rainbow trout were 

8.2 and 3.0 mg/L in 1-h exposures and 0.9 and 0.5 mg/L 

in 3-h exposures.

It is clear from data shown here that formalin was 

superior to any of the candidate fungicides screened. Unfortunately, hatchery managers have not fully exploited 

its effectiveness. Formalin continues to show excellent 

antifungal activity against natural fungal infections caused 

by the Saprolegniales. The safe, controlled use of formalin 

should help control fungal problems until better fungicides 

can be developed.

Our data showed that dichlorophen may have potential 

as an antifungal for use on channel catfish eggs; however, 

its usefulness on fungused rainbow trout eggs is still questionable. The toxicity of quinolinols to fish and to fish 

eggs will probably eliminate them from further consideration as replacements for malachite green.

References

Alderman, D. J., and J. S. Polglase. 1984. A comparative

investigation of the effects of fungicides on Saprolegnia

parasitica and Aphanomyces astaci. Trans. Br. Mycol. Soc.

83:313-318. 

Bailey, T. A. 1983a. Method for in vitro screening of aquatic

fungicides. J. Fish Dis. 6:91-100. 

Bailey, T. A. 1983b. Screening fungicides for use in fish culture:

evaluation of the agar plug transfer, cellophane transfer, and

agar dilution methods. Prog. Fish-Cult. 45:24-27. 

Bailey, T. A. 1984. Effects of twenty-five compounds on four

species of aquatic fungi (Saprolegniales) pathogenic to fish.

Aquaculture 38:97-104. 

Golden, J. J., and K. A. Oster. 1947. Evaluation of a fungicidal laboratory test method. J. Am. Pharm. Assoc.

36:350-362.

Griffin, B. R. 1987. Dichlorophen has potential for use as a fishery 

chemical. U.S. Fish and Wildlife Service Research Information Bulletin 

87-119 (an internal Service document).
INVESTIGATIONS IN FISH CONTROL 99

Griffin, B. R. 1989. Screening of chemicals for controlling protozoan parasites of fish. Prog. Fish-Cult. In press.

Marking, L. L., D. Leith, and J. Davis. 1989. Development 

of a carbon filter system for removing malachite green from 

hatchery effluents. Prog. Fish-Cult. In Press.

Meyer, F. P., and G. L. Hoffman. 1976. Parasites and diseases 

of warmwater fishes. U.S. Fish Wildl. Serv., Resour. Publ. 

127. 20 pp.

Meyer, F. P., and T. A. Jorgenson. 1983. Teratological and 

other effects of malachite green on development of rainbow 

trout and rabbits. Trans. Am. Fish. Soc. 112:818-824.

Mussa, A. G. A., and P. E. Russell. 1977. Seed treatment with 

dichlorphen to control Fusarium solani form species phaseoli 

on Phaseolus vulgaris. Plant Physiol. 26:195-198.

Neish, G. A. 1977. Observations on saprolegniasis of adult 

sockeye salmon, Oncorhynchus nerka (Walbaum). J. Fish 

Biol. 10:513-522.

Neish, G. A., and G. C. Hughes. 1980. Fungal diseases offish. 

Pages 47-49 in S. F. Snieszko and H. R. Axelrod, eds. 

Diseases of Fish, Book 6. T.F.H. Publications, Inc., Neptune City, N.J.

Richards, R. H. 1977. Mycology of fishes. Pages 205-215 in

R. J. Roberts, ed. Fish Pathology. Bailiere Tindall, London.

Richards, R. H., and A. D. Pickering. 1978. Frequency and 

distribution patterns of Saprolegnia infection in wild and 

hatchery-reared brown trout Salmo trutta L. and char Salvelinus alpinus L. J. Fish Dis. 1:69-82.

Roberts, R. J., and C. J. Shepherd. 1974. Handbook of trout 

and salmon diseases. Fishing News (Books), West Byfleet, 

Surrey, England.

Schnick, R. A., and F. P. Meyer. 1978. Registration of thirtythree fishery chemicals: Status of research and estimated costs 

of required contract studies. U.S. Fish Wildl. Serv., Invest. 

Fish Control 86. 19 pp.

Shah, K. A., S. A. Samuels, M. K. Michaels, and J. W. Warren. 1984. Liquid chromatographic analysis of dichlorophen 

and its major impurity. J. Pharm. Sci. 76:822-823.

Takeuchi, I., S. Kobayashi, M. Tanabe, and T. Fujiwara. 1985. 

In vitro inhibition of Giardia lamblia and Trichomonas vaginalis growth by bithionol, dichlorophen, and hexachlorophene. 

Antimicrob. Agents Chemother. 27:65-70.

White, D. A. 1975. Ecology of an annual Saprolegnia sp. 

(Phycomycete) outbreak in wild brown trout. Int. Ver. Theor. 

Angew. Limnol. Verh. 19:2456-2460.

Table 1. Chemicals that were ineffective against Saprolegniales, toxic to nontarget organisms, or otherwise considered unsuitable for further testing.

A-23 (Abbott Laboratories)

A-48 (Abbott Laboratories)

A-49 (Abbott Laboratories)

A-56 (Abbott Laboratories)

A-150A (Lilly Laboratories)

A56620 (Abbott Laboratories)

Acriflavin

7-Amino-4-methylcoumarin

7-Amino-4-trifluoromethylcoumarin

2-Aminopyridine

Ascorbic acid

Baycor

Bayleton

Baytan

BayVP2674

Benomyl

Benzalkonium chloride3

Boric acid

Busan 85

Busan 881

Captanb

Captan 50Wb

Carbenicillin disodium salt

Catch and Release3

Catechol

Cellulase (pH 7.69)

Citcop-4E

Clotrimazole

CO (tech) (Derse & Schroeder)

Copper carbonate

Copper sulfate (basic) 3

Copper sulfate (tri-basic)a

m-Coumaric acid

o-Coumaric acid

P-Coumaric acid

Coumarin

Coumarin 35

Coumarin 138

Coumarin 152

Coumarin 153

Coumarin 311

Cuprimyxin (0.5%)a

Cutrine-plus

Disulfiram

Dithane M-45 (Mancozeb)

Dithane Z-78 (Zineb)

EDTA (disodium)

Fast green

5-Fluorocytosine

Formalin3

Fosfomycin disodium salt

Fumagillin

Funginex

Griseofulvin

Halox E-1003

2-Hydroxy-5-nitropyridine3

Ipronidazole3

Ipropran

Kanamycin sulfate3

Kasumin (50%)

Metalaxyl

Methyl orange

Methylene blue

Morestan

Nalidixic acid (Na + salt)

OAC-3289 3

O-Phenylphenol (Dowcide A)

O-Phenylphenol (Dowcide 1)

Oxolinic acid (20 %) 3

Phaltan 50%

2-Phenoxyethanol

Piperalin

Plantvax

Polyoxin AL (filtered)

Polyoxin AL (suspension)

Polyoxin Z (filtered)

Polyoxin Z (suspension)

Potable aqua

Potassium chloride

Potassium iodide

Potassium sorbate

Previcur-N 3

Quinoline hydrochloride

Quinoline QL

Quinolinol HC1

2-Quinolinol

6-Quinolinol
EVALUATION OF CANDIDATE FUNGICIDES

Table 1. Continued.

7-Quinolinol

8-Quinolinol benzoate 

8-Quinolinol (free base) 

4-Quinolinol-2-carboxylic acid 

8-Quinolinol-5-sulfonic acid 

8-Quinolinol potassium sulfate 

4-Quinolinol-trihydrate 

8-Quinolyl acetate 

R07-4488/006b 

Ridomil 

Roccal II (50%)

Roccal (tech)

Ronilan

Salicylanilide I

Small Fish Savera

Sodium chloride

Sodium iodide

Sodium sulfate

Sodium sulfite

Streptomycin 804

T204C (Tavolek Company)

Tamed iodine

Tannic acid 

Terrazole

Thuringiensin (3%) 

Topsin M 

1-2-4-Triazole 

Trifluralin

Ultramarine blue CP-18 

Ultramarine blue FP-64 

Vancide 51 

Vangard 10W 

Vitavax

a Antifungal activity less than desirable, but screened in further tests because of water solubility or current registrations. 

Rejected because of toxicity or carcinogenicity.

Table 2. Minimum inhibitory concentrations (mg/L) of candidate aquatic fungicides that showed high antifungal 

activity against Achlya flagellata and Saprolegnia hypogyna in 15- and 60-min exposures.

Chemical

Achlya flagellata

15 min 60 min

Saprolegnia hypogyna

15 min 60 min

AL-BB-001

Alpha-terthienyl

Amical 48

P-Benzoquinone

Busan 30

Busan 30L

Busan 30WB

Busan 1030

Copper oxychloride sulfate

Copper-8-quinolinolate

Crystal violet

Dichlorophen

Du-Ter (47.5%)

Du-Ter (tech)

Dyrene

Herbisan 5

5-Hydroxy-l ,4-naphthoquinone

8-Hydroxy-5-nitroquinoline

Malachite green

Malachite green (carbinol)

Malachite green (neutralized)

Phaltan (tech)

Polyphase 17WD

Polyphase AF-1

Polyphase P-100

8-Quinolinol

8-Quinolinol copper salt

8-Quinolinol HC1

V-101 (Abbott Laboratories)

V-102 (Abbott Laboratories)

5.0

5.0

30.0

5.0

10.0

10.0

10.0

>10<100

1.0

1.0

10.0

1.0

3.0

30.0

10.0

10.0

30.0

1.0

<10

<10

1.0

5.0

3.0

5.0

1.0

3.0

5.0

10.0

30.0

8.0

1.0

3.0

10.0

3.0

10.0

10.0

10.0

10.0

1.0

0.3

7.5

1.0

3.0

10.0

10.0

6.0

9.0

1.0

<10

<10

1.0

3.0

3.0

5.0

7.5

3.0

>15<30

5.0

10.0

5.0

3.0

10.0

5.0

10.0

10.0

10.0

10.0

1.0

0.75

7.5

1.0

1.0

10.0

10.0

6.0

3.0

0.5

<10

<10

1.0

3.0

3.0

3.0

1.0

3.0

5.0

3.0

5.0

5.0

1.0

0.75

10.0

3.0

10.0

10.0

10.0

10.0

1.0

0.5

7.5

1.0

1.0

10.0

10.0

6.0

9.0

0.5

<10

<10

1.0

1.0

1.0

1.0

1.0

1.0

5.0

1.0

3.0
INVESTIGATIONS IN FISH CONTROL 99

Table 3. Minimum inhibitory concentrations (mg/L) of candidate aquatic fungicides that showed moderate antifungal

activity in 15- and 60-min exposures.

Achlya flagellata

Chemical

Actidione

AO (tech) (Derse & Schroeder)

Bayer 73

Black algaetrine

Blasticidin-S

Botran

Cellulase (pH 5.02)

Chlorazene (powder)

Chlorazene (tablets)

Copper sulfate (tri-basic)

Defungit

5,7-Dichloro-8-quinolinol

Dithane M-22 (Maneb)

Dithianon

Dodine

Ethyl violet

Hyamine 3500

2-Hydroxy- 1 ,4-naphthoquinone

Irumamycin

Karathane

Lesan (technical)

Miconazole

5 -Nitro- 8-quinolinol

5-Nitrosalicylic acid

Polyalkylene glycol-iodine

Potassium permanganate

8-Quinolinol sulfate

TFM (36.3%)

TFM (85.6%)

TFM (tech)

2-(4-Thiazolyl) benzimidazole

Thiram

Thiram 42S

15 min

30.0

15.0

25.0

MOO

500

75.0

MOO

50.0

MOO

100

125

MOO

50.0

700

30.0

15.0

500

75.0

MOO

100

60.0

300

24.0

112.5

100

210

100

75.0

30.0

30.0

30.0

100

MOO

60 min

30.0

3.0

15.0

75.0

500

75.0

MO

30.0

>10<100

100

100

>10<100

75.0

75.0

30.0

15.0

100

45.0

>10<100

100

60.0

75.0

8.0

75.0

100

80.0

100

50.0

30.0

30.0

30.0

75.0

MOO

Saprolegnia hypogyna

15 min

>30

40.0

30.0

50.0

30

75.0

MOO

75.0

>10<100

100

125

M0<100

50.0

300

75.0

15.0

500

75.0

>10<100

100

60.0

75.0

8.0

112.5

150

210

300

75.0

30.0

30.0

30.0

30.0

>10<100

60 min

>30

15.0

30.0

10.0

3.0

50.0

MO

30.0

>10<100

100

100

>10<100

50.0

100

75.0

15.0

100

45.0

>10<100

30.0

36.0

30.0

24.0

45.0

75.0

80.0

100

10.0

30.0

30.0

30.0

30.0

10.0
EVALUATION OF CANDIDATE FUNGICIDES

Table 4. Minimum inhibitory concentrations (mg/L) of candidate aquatic fungicides that showed low antifungal activity

in 15- and 60-min exposures.

Achlya flagellata

Chemical

Amobam

Amphotericin B-Type I

BAS-389-01F

Benzalkonium chloride

Catch and Release

CGA-64251

Copper sulfate (basic)

Copper sulfate (crystals)

Cuprimyxin (0.5%)

Diquat dibromide

Enilconazole

Formalin

Halox E-100

2-Hydroxy-5-nitropyridine

Ipronidazole

Kanamycin sulfate

Ketoconazole

Lesan 70% WP

Leuco-malachite green

Naftifine

Nalidixic acid (free acid)

OAC-3289

Oxolinic acid (20%)

Poly ram 80% (filtered)

Polyram 80% (suspension)

Previcur-N

8-Quinolinol citrate

Small Fish Saver

Sodium omadine

Solricin 135

Surflan

Thiabendazole

15 min

MOO

>1,000

500

500

> 1,000

1,000

300

300

2,500

5,000

750

700

> 1,000

> 1,000

> 1,000

> 1,000

300

> 1,000

800

MOO

500

>1,000

1,000

500

> 1,000

MOO

1,000

MOO

250

200

1,000

60 min

MOO

> 1,000

500

300

>1,000

500

100

300

750

2,500

300

200

750

> 1,000

> 1,000

> 1,000

300

> 1,000

800

MOO

500

> 1,000

750

500

>1,000

MOO

750

MOO

250

150

1,000

Saprolegnia hypogyna

15 min

MOO

> 1,000

300

500

> 1,000

500

100

300

875

30.0

300

700

> 1,000

> 1,000

> 1,000

1,000

300

100

>800

MOO

500

> 1,000

> 1,000

750

500

>1,000

MOO

750

MOO

250

200

300

60 min

MOO

>1,000

300

300

> 1,000

300

100

300

750

30.0

100

200

> 1,000

> 1,000

> 1,000

1,000

100

100

>800

MOO

750

> 1,000

> 1,000

500

500

> 1,000

MOO

500

MOO

250

150

300
INVESTIGATIONS IN FISH CONTROL 99

Table 5. Concentrations (mg/L) of fungicides that were effective for the control of aquatic fungi on dead rainbow

trout eggs in egg to agar transfer tests.

Incubation time

Chemical

Actidione

AL-BB-001 (40%)

Amical 48

p -Benzoquinone

Black algaetrine

Busan 30

Busan SOL

Busan 1030

Busan 30WB

Chloramine-T

Copper-8-quinolinolate

Defungit

Dichlorophen

Dithianon

Dodine

Du-Ter (47.5%)

Du-Ter (tech)

Formalin

5-Hydroxy-l ,4-naphthoquinone

LD (Alcide Corp.)

Lesan (70%)

Malachite green

5-Nitro-8-quinolinol

Phaltan (technial)

Polyphase 17WD

Polyphase P-100

Potassium permanganate

8-Quinolinol

8-Quinolinol copper salt

8-Quinolinol sulfate

TFM (36.3%)

TFM (85.6%)

TFM (tech)

Thiabendazole

Thiram 42S

V-101 (Abbott Laboratories)

V-102 (Abbott Laboratories)

15 min

100

_ a

>250

>100

75

10

75

100

>10

1,000

10.0

>300

188

75

300

8.0

MOO

30

100

50

150

100

100

> 1,000

24 h

60 min

100

25

MOO

50

10

75

75

MO

10.0

>300

188

75

100

8.0

10

10

100

50

250

100

100

> 1,000

15 min

100

>250

MOO

75

MOO

MOO

100

MO

3,000

MOO

>300

188

>2,000

M,000

>80

MOO

50

300

>50

375

300

100

>1,000

168 h

60 min

100

25

MOO

50

100

100

100

MO

3,000

100

>300

188

>2,000

750

>80

10

10

100

>50

250

300

100

> 1,000

'Dashes indicate chemicals tested in in vivo tests, but not in egg to agar transfer tests.
EVALUATION OF CANDIDATE FUNGICIDES

Table 6. Evaluation ofcandidate fungicides selected for in vivo tests with artificially infected live rainbow trout eggs.

Chemical

Actidione

AL-BB-001 (40%)

Alcide

Amical 48

p-Benzoquinone

Black algaetrine

Busan 30WB

Chloramine-T

Copper-8-quinolinolate

Defungit

Dichlorophen

Dithianon

Dodine

Du-Ter (tech)

Du-Ter (47.5%)

Formalin

5-Hydroxy- 1 ,4-naphthoquinone

LD (Alcide Corp.)

Malachite green

5-Nitro-8-quinolinol

Phaltan (technial)

Polyphase 17WD

Polyphase P-100

Potassium permanganate

8-Quinolinol

8-Quinolinol copper salt

8-Quinolinol sulfate

TFM (36.3%)

Thiabendazole

Thiram 42 S

V-101 (Abbott Laboratories)

V-102 (Abbott Laboratories)

Concentration 

(mg/L)

25

10

1:10:1

25

10

30

1

75

20

50

2

100

75

1

1

250

10

1:10:1

5

15

5

10

10

50

30

1

35

15

30

30

15

5

Effect on eggs

Toxic

Toxic

Toxic

Toxic

Toxic

Toxic

Toxic

Toxic

Toxic

Ineffective

Marginally effective

Toxic

Toxic

Toxic

Toxic

Effective

Highly toxic

Toxic

Effective3

Toxic

Toxic

Toxic

Toxic

Marginally effective

Effective

Toxic

Effective13

Toxic

Toxic

Toxic

Toxic

Toxic

a Potential risk to human health. 

b Post-hatch mortality.

Bailey, Tom A., and Susan M. Jeffrey. 1989. Evaluation of 215 Candidate 

Fungicides for Use in Fish Culture. U.S. Fish. Wildl. Serv., Invest. Fish 

Control 99. 9 pp.

The fungicidal activity of 215 compounds was evaluated by comparing their 

effectiveness with that of malachite green. More than half were found to be unsuitable as aquatic fungicides in preliminary screening tests because of their lack 

of activity against fungi, toxicity to fish or eggs, insolubility in water, or potential carcinogenicity. Four compounds 8-quinolinol, 8-quinolinol sulfate, 

dichlorophen, and formalin inhibited fungal growths on dead and live fish eggs 

and demonstrated the greatest potential as replacements for malachite green. The 

quinolinols are questionable for use as aquatic fungicides because of their toxicity. Dichlorophen holds some promise for use against fungus on eggs of channel catfish (Ictalurus punctatus), but formalin was effective at a concentration 

of 250 mg/L substantially lower than has been previously reported.

Key words: Fungicides, dichlorophen, formalin, 8-quinolinol, fish eggs.

HBailey, Tom A., and Susan M. Jeffrey. 198

Fungicides for Use in Fish Culture. U.S

Control 99. 9 pp.

The fungicidal activity of 215 compounds 

effectiveness with that of malachite green. Mo

suitable as aquatic fungicides in preliminary sc

of activity against fungi, toxicity to fish or egg

tial carcinogenicity. Four compounds 8-q

dichlorophen, and formalin inhibited fungal 

and demonstrated the greatest potential as repl

quinolinols are questionable for use as aquati

icity. Dichlorophen holds some promise for u

nel catfish (Ictalurus punctatus), but formalin 

of 250 mg/L substantially lower than has b

Key words: Fungicides, dichlorophen, form

Bailey, Tom A., and Susan M. Jeffrey. 1989. Evaluation of 215 Candidate Fungicides for Use in 

Fish Culture. U.S. Fish. Wildl. Serv., Invest. Fish Control 99. 9 pp.

The fungicidal activity of 215 compounds was evaluated by comparing their effectiveness with that 

of malachite green. More than half were found to be unsuitable as aquatic fungicides in preliminary 

screening tests because of their lack of activity against fungi, toxicity to fish or eggs, insolubility in 

water, or potential carcinogenicity. Four compounds 8-quinolinol, 8-quinolinol sulfate, dichlorophen, 

and formalin inhibited fungal growths on dead and live fish eggs and demonstrated the greatest potential 

as replacements for malachite green. The quinolinols are questionable for use as aquatic fungicides 

because of their toxicity. Dichlorophen holds some promise for use against fungus on eggs of channel 

catfish (Ictaluruspunctatus), but formalin was effective at a concentration of 250 mg/L substantially 

lower than has been previously reported.

Key words: Fungicides, dichlorophen, formalin, 8-quinolinol, fish eggs.

Bailey, Tom A., and Susan M. Jeffrey. 1989. Evaluation 

Fish Culture. U.S. Fish. Wildl. Serv., Invest. Fish C

The fungicidal activity of 215 compounds was evaluated 

of malachite green. More than half were found to be unsu

screening tests because of their lack of activity against fu

water, or potential carcinogenicity. Four compounds 8-qui

and formalin inhibited fungal growths on dead and live fish 

as replacements for malachite green. The quinolinols are 

because of their toxicity. Dichlorophen holds some promis

catfish (Ictaluruspunctatus), but formalin was effective at 

lower than has been previously reported.

Key words: Fungicides, dichlorophen, formalin, 8-quino



(Reports 87 through 89 are in one cover.)

87. Ethyl-p-aminobenzoate (Benzocaine): Efficacy as an Anesthetic for Five Species of Freshwater Fish, by V. K. Dawson 

and P. A. Gilderhus. 1979. 5 pp.

88. Influences of Selected Environmental Factors on the Activity of a Prospective Fish Toxicant, 2-(Digeranyl-amino)-ethanol, 

in Laboratory Tests, by C. A. Launer and T. D. Bills. 1979. 4 pp.

89. Toxicities of the Lampricides 3-Trifluoromethyl-4-nitrophenol (TFM) and the 2-Aminoethanol Salt of 2'.5-Dichloro-4'- 

nitrosalicylanilide (Bayer 73) to Four Bird Species, by R. H. Hudson. 1979. 5 pp.

(Reports 90 and 91 are in one cover.)

90. Accumulation and Loss of 2',5-Dichloro-4'-nitrosalicylanilide (Bayer 73) by Fish: Laboratory Studies, by V. K. Dawson, 

J. B. Sills, and Charles W. Luhning. 1982. 5 pp.

91. Effects of Synergized Rotenone on Nontarget Organisms in Ponds, by R. M. Burress. 1982. 7 pp.

(Reports 92 through 94 are in one cover.)

92. Acute and Chronic Toxicity of Rotenone to Daphnia magna, by J. J. Rach, T. D. Bills, and L. L. Marking. 1988. 5 pp.

93. Toxicity of Rotenone to Developing Rainbow Trout, by T. D. Bills, J. J. Rach, and L. L. Marking. 1988. 3 pp.

94. Oral Toxicity of Rotenone to Mammals, by L. L. Marking. 1988. 5 pp.

95. Deposition and Persistence of Rotenone in Shallow Ponds During Cold and Warm Seasons, by P. A. Gilderhus. V. K. 

Dawson. and J. L. Alien. 1988. 7 pp.

(Reports 96 and 97 are in one cover.)

96. Effects of Environmental Factors on the Toxicity of Chloramine-T to Fish, by T. D. Bills, L. L. Marking, V. K. Dawson, 

and J. J. Rach. 1988. 6 pp.

97. Effects of Organic Matter and Loading Rates of Fish on the Toxicity of Chloramine-T, by T. D. Bills, L. L. Marking, 

V. K. Dawson, and G. E. Howe. 1988. 4 pp.

98. History of Acute Toxicity Tests with Fish, 1863-1987, by J. B. Hunn. 1989. 10 pp.

NOTE: Use of trade names does not imply U.S. Government endorsement of commercial products.
TAKE PRIDE

in America

U.S. DEPARTMENT OF THE INTERIOR

FISH AND WILDLIFE SERVICE

As the Nation's principal conservation agency, the Department of the Interior has responsibility for most of our nationally owned public lands and natural resources. This includes 

fostering the wisest use of our land and water resources, protecting our fish and wildlife, 

preserving the environmental and cultural values of our national parks and historical places, 

and providing for the enjoyment of life through outdoor recreation. The Department assesses 

our energy and mineral resources and works to assure that their development is in the best 

interests of all our people. The Department also has a major responsibility for American 

Indian reservation communities and for people who live in island territories under U.S. 

administration.