Sendai virus intra-host population dynamics and host immunocompetence influence viral virulence during in vivo passage

Peña, José; Chen-Harris, Haiyin; Allen, Jonathan E.; Hwang, Mona; Elsheikh, Maher; Mabery, Shalini; Bielefeldt-Ohmann, Helle; Zemla, Adam T.; Bowen, Richard A.; Borucki, Monica K.

Mice were anesthetized with a mixture of ketamine and xylazine, and inoculated intranasally with 25 ml (1,000 plaque forming unit (PFU)) of stock virus (passage 1) or tracheal lavage fluid from infected mice. Mice were monitored daily for weight loss and visually inspected twice daily for signs of disease such as lethargy, ruffled fur, and hunching. Mice were euthanized 5 days post infection and virus was obtained from infected lung tissue via BAL (Aronson, Herzog, and Jerrells 1995) . The infection protocol of intranasal inoculation and infection of the recipient group with pooled donor BAL (versus homogenized lung tissue) was designed to simulate of viral transmission via the respiratory route. Lavage samples were pooled within each treatment group with equal quantities from each mouse (100 ml) then diluted according to titration results to a concentration of $ 5 Â 10 3 PFU/25 ml for inoculation of the next group. Thus the viral dose was held constant for subsequent in vivo passage for a total of ten serial passages. BAL samples were assayed for viral titer via plaque assay (described below), and lungs were fixed in buffered formalin for histopathology.