Development and maintenance of a specific pathogen-free (SPF) zebrafish research facility for Pseudoloma neurophilia

Pseudoloma neurophilia (Microsporidia) is very common in zebrafish research facilities. A new zebrafish facility was established at the Sinnhuber Aquatic Resource Laboratory (SARL) at Oregon State University, and thus we used this as an opportunity to establish a Specific Pathogen Free (SPF) colony of zebrafish for this microsporidium. Progeny from 10 zebrafish lines (n = 2,203) were initially transferred to the SARL facility in 2007 following PCR screening of broodstock and a subpopulation of progeny (258/1,000 fish from each family). Screening of fish within the facility was conducted as follows: 1) Moribund or dead fish were examined by histology 2) Each line was regenerated on a 4 mo. rotation, and for each of these major propagations, a subsample was screened for P. neurophilia by PCR, in which 60 fry from were collected randomly at 10 days post hatch and screened by PCR for P. neurophilia in pools of 10 3) Adult fish from each line were retired and euthanized at approximately 1 year of age. Twenty of these fish were examined by histology and the brains and spinal cords of 60 fish were combined in pools of five and screened by PCR 4) Sentinel fish were held in 4 tanks receiving effluent water from all tanks in the facility (20 fish/tank). Twenty fish were examined by histology and the brains of another 60 fish (in pools of five) were screened by PCR for P. neurophilia and 5) 760 4 mo old fish from a toxicology study conducted within the laboratory were examined by histology. To date, we have evaluated 2,800 fish by PCR and 1, 222 fish by histology and have not detected P. neurophilia. Thus we have established 9 lines of zebrafish SPF for P. neurophilia. However, 26 fish exhibited mycobacteriosis, diagnosed by the presence of acid fast bacteria visualized in tissue sections. Forty-nine other fish exhibited chronic inflammatory lesions, including egg associated inflammation and hyperplasia, in which acid-fast bacteria were not detected. Eight exhibited hepatic megalocytosis or hepatocellular pleomorphism, and three exhibited neoplasia (cholangiocellular carcinoma, and two with seminoma). One of the seminomas occurred in a female, and was classified as ovo-testes.