Abstract

Four lined seahorses (Hippocampus erectus) of a captive research collection presented with negative buoyancy and bloated upper trunks. Radiographs of an affected individual showed a markedly reduced volume of air in the gas bladder. We collected brown purulent exudate from the gas bladder lumen via pneumocystocentesis. Wet mount microscopy of the exudate revealed many ova of unknown origin, many motile and non-motile bacteria, and many leukocytes. Three smears of the exudate were prepared and stained with Dif-Quik, Gram, and Ziehl-Neelsen stains. Collectively, these smears demonstrated many macrophages and weakly gram-positive, acid-fast positive bacilli.

Diagnostic necropsies were performed. Gross internal examination revealed enlarged, firm, opaque gas bladders, as well as extremely friable and enlarged posterior kidneys. Gas bladders demonstrated thickened walls, and when opened, contained copious brown purulent exudate, unidentifiable necrotic material, and adult trematodes. Trematodes were relaxed in hot saline, fixed in 10% neutral buffered formalin, and identified as Dictysarca sp. (virens?).

Exudate from the gas bladder lumen was cultured on TSA agar with 5% sheep's blood (incubated at 25°C) and two Lowenstein-Jensen (LJ) slants (incubated at 25°C and 37°C). Pasteurella multocida and Staphylococcus sp were identified from three TSA cultures. Three animals demonstrated mustard yellow growth on LJ media by day 10 at 25°C and by 72h at 37°C. LJ subcultures were acid-fast positive, and identified as Mycobacterium poriferae by HPLC and 16S rDNA sequencing.

Histopathology demonstrated a histiocytic response (characterized by cellular infiltration with proliferative fibroblasts) in kidney and gas bladder exudate tissues, severe edema of the gas bladder wall, and severe infection of the gas bladder lumen with acid-fast positive bacilli. A granulomatous response, commonly observed in other Mycobacterium infected fish species, was noticeably absent.

Under dissection microscopy, we examined a sample (n=2616) of harpacticoid copepods from the research system for epidemiological evidence of metacercariae but found none. Alternatively, seahorses may be infected from ingestion of infected live Artemia, acquire infection in the wild, or in the collector's system, which houses many diverse invertebrates.

This is the first report of infection of a syngnathid by M. poriferae. The bacillus was only found occasionally in the presence of the digene, only in the gas bladder, and only in conjunction with purulent exudate. Other, behaviorally normal animals necropsied from this research population demonstrated high incidence of Dictysarca sp. infection (50%), but without purulent exudate or rapid growing Mycobacterium spp. These observations suggest that Dictysarca sp. may harbor M. poriferae in their cecae and transport them to the host, and/or the pathology imposed by the digene creates an environment conducive to its culture. As Mycobacterium spp. are a group of zoonotic pathogens, stringent biosecurity measures (UV sterilization of water, equipment disinfection, skin protection) are recommended for aquarists.

Acknowledgements

Many thanks to J. Holloway and T. Crosby (University of Florida) for assistance with diagnostic procedures, S. Terrell (Wildlife Diagnostics) for histological interpretation, All Florida Veterinary Laboratories and the National Jewish Medical and Research Center for microbiological identification, and A. Dove (Georgia Aquarium) for digene identification. This work was funded by the Fish Health Extension Laboratory of the UF/IFAS Department of Fisheries and Aquatic Sciences.