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Specific pathogen-free (SPF) animals are bred and managed to exclude pathogens associated with significant morbidity or mortality that may secondarily pose a risk to public health, food safety and food security, and research replicability. Generating and maintaining SPF animals requires detailed biosecurity planning for control of housing, environmental, and husbandry factors and a history of regimented pathogen testing. Successful programs involve comprehensive risk analysis and exclusion protocols that are rooted in a thorough understanding of pathogen lifecycle and modes of transmission. In this manuscript we review the current state of SPF in domestic agriculture (pigs and poultry), aquaculture (salmonids and shrimp), and small laboratory mammals. As the use of laboratory fish, especially zebrafish (Danio rerio), as models of human disease is expanding exponentially, it is prudent to define standards for SPF in this field. We use the guiding principles from other SPF industries and evaluate zebrafish pathogens against criteria to be on an SPF list, to propose recommendations for establishing and maintaining SPF laboratory zebrafish.

Preventing xenograft rejection is one of the greatest challenges of transplantation medicine. Here, we describe a reproducible, long-term survival of cardiac xenografts from alpha 1-3 galactosyltransferase gene knockout pigs, which express human complement regulatory protein CD46 and human thrombomodulin (GTKO.hCD46.hTBM), that were transplanted into baboons. Our immunomodulatory drug regimen includes induction with anti-thymocyte globulin and αCD20 antibody, followed by maintenance with mycophenolate mofetil and an intensively dosed αCD40 (2C10R4) antibody. Median (298 days) and longest (945 days) graft survival in five consecutive recipients using this regimen is significantly prolonged over our recently established survival benchmarks (180 and 500 days, respectively). Remarkably, the reduction of αCD40 antibody dose on day 100 or after 1 year resulted in recrudescence of anti-pig antibody and graft failure. In conclusion, genetic modifications (GTKO.hCD46.hTBM) combined with the treatment regimen tested here consistently prevent humoral rejection and systemic coagulation pathway dysregulation, sustaining long-term cardiac xenograft survival beyond 900 days., Tweaking immune characteristics of donors and recipients could allow for successful cross-species organ transplantation. Here, the authors show that an anti-CD40 antibody therapy of baboons that received heart transplants from genetically modified pigs is key to their long-term survival.

Body condition scoring (BCS) is a simple, rapid, noninvasive tool used to assess body condition in animals. In this study, we developed and validated a diagram-based BCS for adult zebrafish (Danio rerio), a popular research model. After receiving 20 min of hands-on training regarding the scoring system, 5 people each rated 95 adult zebrafish. The fish then were euthanized and measured to establish body condition indices (BMI and the Fulton K factor). Both condition indices were highly correlated with fish width. Using correlation data and observed trends in fish width, we established expected BCS definitions. We validated the BCS definitions in 2 ways. First, we calculated the Pearson correlation coefficient between the average observed BCS and expected BCS; this statistic revealed very strong correlation between observed and expected BCS. In addition, we assessed the predictive power of BCS by using multinomial logistic regression and then applied the fitted model to evaluate the accuracy of the predictions (BCS compared with BMI, 85%; BCS compared with K factor, 61%). Finally, to determine the robustness of BCS to variation among raters, we calculated the intraclass correlation coefficient and demonstrated high interrater reliability. In conclusion, adult zebrafish BCS can be used to quickly identify animals with different body condition indices (thin to obese). In addition, the diagram-based chart is easy to use and implement accurately, with minimal training.