We next sought to determine the impact of MHV-1 infection on tissue pathology by examining the lungs and liver of C57BL/6N and C3H/HeJ mice that had been euthanized with more than 20% weight loss or at 8 dpi when the study was terminated (Fig.2A). Lung damage was assessed by identifying and semi-quantitatively scoring inflammation involving the alveolar septa, alveolar space, and surrounding vessels as well as the presence and degree of type II pneumocyte hyperplasia. Liver damage was assessed by identifying and semi-quantitatively scoring hepatocyte vacuolation, hepatocyte necrosis, and overall liver inflammation. The individual parameters were then summed to obtain a composite histopathologic score for each organ. In C57BL/6N mice, no differences were observed between infected lean and obese mice in either tissue (Fig.3A & B). In C3H/HeJ mice, no differences were observed in the lungs of infected obese and lean mice (Fig.3C), whereas the composite liver scores were significantly higher in obese compared to lean mice (Fig.3D, *p=0.0260).

Lean and obese C57BL/6N and C3H/HeJ mice exposed to MHV-1 were euthanized at 8 dpi or when they crossed the threshold of weight loss (20% of the baseline), and tissues were collected for histopathological analysis. H&E-stained images were captured at 20X, scale bar = 100 um.Lung (A) & liver (B) histopathology of C57BL/6N mice.Lungs (top image) and livers (bottom image) from mock and MHV-1 infected lean and obese C57BL/6N mice, showed no significant difference in histopathological lesions between both the groups. Lung (C) & liver (D) histopathology of C3H/HeJ mice.Lungs (top image) and livers (bottom image) from mock and MHV-1 infected lean and obese C3H/HeJ mice were examined. Statistical analysis was carried out by 2way ANOVA. The level of significance is represented as follows: p < 0.05 (*); p < 0.001 (**); p = 0.001 to 0.0001 (***); p < 0.0001 (****), ns = non-significant. The error bars indicate SD of the mean.

In severe cases of COVID-19, lung damage is a driving factor responsible for most patient fatalities⁴⁷ and diffuse alveolar damage (DAD) is a common histological finding in patients with severe COVID-19^48-51. DAD represents injury to type I pneumocytes or endothelial cells in the alveolar septa and is characterized by inflammation in the alveolus and alveolar septa and type II pneumocyte hyperplasia^52-54. Although we observed no differences in composite lung pathology between lean and obese infected groups (Fig.3A & C), we did identify type II pneumocyte hyperplasia and inflammation of alveoli and alveolar septa in C57BL/6N and C3H/HeJ (Fig.4) mice, consistent with DAD, confirming the validity of our MHV-1 infection models for studying COVID-19 pathogenesis.