Evaluating beluga (Delphinapterus leucas) blow samples as a potential diagnostic for immune function gene expression...

Evaluating respiratory health is important in the management of cetaceans, which are vulnerable to respiratory diseases. Quantifying the expression of genes related to immune function within the respiratory tract could be a valuable tool for directly assessing respiratory health. Blow (exhale) samples allow DNA analysis, and we hypothesized that RNA could also be isolated from blow samples for gene expression studies of immune function. We evaluated the potential to extract RNA from beluga blow samples and tested whether transcripts associated with immune function could be detected with endpoint polymerase chain reaction. A total of 54 blow samples were collected from clinically healthy aquarium belugas (n = 3), and 15 were collected from wild belugas temporarily restrained for health assessment in Bristol Bay, Alaska (n = 9). Although RNA yield varied widely (range, 0-265.2 ng; mean = 85.8; SD = 71.3), measurable RNA was extracted from 97% of the samples. Extracted RNA was assessed in 1-6 PCR reactions targeting housekeeping genes (Rpl8, Gapdh or ActB) or genes associated with immune function (TNFα, IL-12p40 or Cox-2). Fifty of the aquarium samples (93%) amplified at least one transcript; overall PCR success for housekeeping genes (96/110, 87%) and genes associated with immune function (90/104, 87%) were similarly high. Both RNA yield and overall PCR success (27%) were lower for wild beluga samples, which is most likely due to the reduced forcefulness of the exhale when compared with trained or free-swimming belugas. Overall, the high detection rate with PCR suggests measuring gene expression in blow samples could provide diagnostic information about immune responses within the respiratory tract. While further study is required to determine if quantitative gene expression data from blow samples is associated with disease states, the non-invasive nature of this approach may prove valuable for belugas, which face increasing anthropogenic disturbances.