Abstract

Praziquantel baths are a routine part of marine fish quarantine; however, data to support doses, dosing intervals, or stability of the drug in marine systems are lacking. Over the past 2 yr, New England Aquarium has conducted several modest experiments to assess praziquantel potency, stability, limit of detection, and concentrations during clinical use. In addition, the effects of ozone and activated carbon on praziquantel concentrations were investigated. Results suggest that the residence of time of praziquantel in marine systems is very variable, from less than 24 hr in some systems, to over a week in other systems, even during active attempts to remove it faster.

Introduction

Several reports have documented effective clinical use of praziquantel baths for treatment of external trematode infections of marine fish, and many institutions use praziquantel baths as a routine part of marine fish quarantine.^1-3 In a recent survey of fish quarantine practices at zoos and aquaria, 75% of institutions reported routine use of praziquantel baths, while only 3% tested praziquantel concentrations in treated water.¹ Clinically used protocols vary widely (e.g., 20 ppm for 90 min, 2 ppm for several weeks); however, data to support specific doses, dosing intervals, or stability of the drug in marine systems are lacking. At this time, testing for praziquantel concentrations in sea water is conducted by only three laboratories in the United States, and is moderately expensive ($30–$150 per test). As such, monitoring therapeutic concentrations of praziquantel is not routine.

Methods

Over the past 2 yr, New England Aquarium has conducted several modest experiments to assess praziquantel potency, stability, limit of detection, and concentrations during clinical use. Praziquantel assays were conducted by high performance liquid chromatography at Analytical Research Laboratories, Oklahoma City, OK, USA. Praziquantel was purchased in bulk powdered form from Fishman Chemical, Hobe Sound, FL, USA. Pure samples from three separate lots of praziquantel stored in three separate locations were submitted for potency testing, which was >99% for each sample. For addition of praziquantel to sea water, powder was weighed based on desired concentration and volume of water, and was distributed into the water by squeezing it through a nylon stocking until fully dissipated. Components of life support systems that may remove or denature praziquantel were not used during clinical treatments (i.e., ozone, ultraviolet sterilizers, or activated carbon), but were used experimentally and clinically to reduce concentrations when desired. All praziquantel baths were prepared using natural sea water.

Results

Assays of a serially diluted 6 ppm praziquantel solution in natural sea water (water temp 24°C) demonstrated an approximate limit of detection between 0.015 and 0.04ppm. Detailed accuracy tests were not conducted; however, for four systems in which concentrations were measured 60 min after dosing, concentrations were 73%, 92%, 95%, and 126% of expected values (water temperature 24–26°C). It is very possible that this represented incomplete mixing of the drug rather than test inaccuracy, but specific accuracy testing would be required to assess this possibility.

To assess stability under the influence of ozone and activated carbon, concentrations were monitored in an experimental 3,800 gallon system with no animals (water temperature 24–25°C), with a starting concentration of 3.5ppm. Concentrations were reduced to 65% (24 h), 27% (72 h), 10% (120 h), 4.5% (7 d), and 2.5% (8 d) after starting ozone and carbon.

During routine quarantine treatments, with initial concentrations of 5 ppm or 10 ppm, four systems were surprisingly found to have undetectable praziquantel concentrations within 24 to 96 hr after dosing (water temperatures 24–26°C).

Praziquantel was used clinically for treatment of Benedeniella posterocolpa on a group of newly acquired cownose rays (Rhinoptera bonasus). Rays were housed in a 30,000 gallon closed system (water temperatures 23–24°C) which was treated on Day 1 at a theoretical praziquantel concentration of 3.5 ppm, with desire to maintain this concentration for 1 mo. Supplemental re- doses of 50% were applied on Day 5 and Day 16, and a 100% re-dose was applied on day 22 based on measured concentrations. Measured concentrations were 2.75 ppm (Day 2), 2.55 ppm (Day 4), 4.0 ppm (Day 7), 3.38 ppm (Day 11), 2.44 ppm (Day 14), 1.72 ppm (Day 18), 0.94 ppm (Day 21), and 1.03 ppm (Day 25). Treatment was effective in eradicating the parasites based on examinations over the subsequent 20 mo. For compliance with regulations for the discharge of the treated water, ozone and carbon was applied to the system on Day 29. Concentration on Day 32 was 0.045 ppm, and on Day 40 was undetectable.

Discussion

Collectively, these preliminary observations suggest that the residence of time of praziquantel in marine systems is highly variable, from less than 24 hr in some systems, to over a week in other systems, even during active attempts to remove it faster. Based on these findings, clinicians should consider monitoring therapeutic praziquantel concentrations during treatment. Increasingly strict regulations for discharge water may also increase the need for such testing. Test accuracy, stability during shipping, and variables that influence the persistence or elimination of praziquantel in marine systems are worthy of further study.

Acknowledgments

The author thanks the many members of the Fishes Department and Animal Health Department that contributed to the execution of these experiments and treatments.

Literature Cited

1.  Hadfield, C.A., and L.A. Clayton. 2011. Fish quarantine: current practices in public zoos and aquaria. J. Zoo Wildl. Med. 42:641–650.

2.  Stetter, M., D. Neiffer, A. Stamper, J. Capobianco, I. Burns, and J. Davis. 2005. Medical considerations when exhibiting multiple taxa in large aquarium systems. Proc. Am. Assoc. Zoo. Vet.: 42–44.

3.  Thoney, D.A. 1990. The effects of trichlorphon, praziquantel, and copper sulfate on various stages of the monogenean Benedeniella posterocolpa, a skin parasite of the cownose ray, Rhinoptera bonasus (Mitchill). J. Fish Dis. 13:385–389.