Low-replication influenza virus mediates high pathogenicity through an inflammation-driven lung–heart–brain axis in mice

Figures

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  Figure 1.. Morbidity, mortality, and replication of GX19 viruses. (A) Five 8-week-old C57BL/6 mice per group received intranasal inoculation of either 105 or 106 TCID50 of the indicated viruses. Daily body weight checks were conducted over a 14 dpi. Euthanasia was performed humanely on mice that experienced a ≥ 25% loss of their original body weight. (B) Depicts the survival rate percentages of C57BL/6 mice after exposure to the indicated viruses. (C and D) Six C57BL/6 mice were assigned to groups and given 106 TCID50 of the designated viruses via intranasal administration. At 1 and 4 dpi, three mice from each group were euthanized. Respiratory tract tissues such as the nasal turbinate (NT), trachea (Tr), and lungs (Lu) were harvested individually for TCID50 testing (C) or RT-qPCR (D), respectively. (E) The growth characteristics of the GX19 and CS were compared in MDCK cells maintained at 37°C. The supernatants were collected, and the virus titres were determined using TCID50 in MDCK cells at the indicated time points. Results are expressed as the mean value plus or minus standard deviation (SD), with a sample size of 3 (n = 3). GX19, A/Environment/Guangxi/44461/2019 (H3N2). GX19-6 and…

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  Figure 2.. Pathological changes in the lung, heart, and brain of mice after viral infections. (A)-(C) refer to the images of mice's lungs, hearts, and brains, respectively, by H&E staining (10×, Scale bar = 1 mm for low magnification; 100×, Scale bar = 100 μm for high magnification). (D)–(F) refer to quantitative measures of injuries in mice's lungs, hearts, and brains, respectively (see Materials and Methods). P-values were shown for comparison between CS-6, GX19-5, and GX19-6 using the t-test. n = 3 mice per group except for the brain of the GX19-6 group, where n = 2 due to severe degradation of a mouse brain sample in the H&E experiment. (G)-(I) refer to GSVA results of acute diseases in the lungs, hearts, and brains, respectively.

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  Figure 3.. Transcription analysis of GX19 infection. (A) Number of DEGs identified in the lung, heart, and brain. (B)∼(E) refer to the enriched biological processes of up - and down-regulated DEGs in the lung and heart after infection with GX19-6. The GO terms were clustered, and each subpanel showed the number of GO terms in each cluster. (F) Top 10 enriched KEGG pathways in the mice brain by GSEA analysis.

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  Figure 4.. GX19-6 specific genes and functions relative to GX19-5 and CS-6. (A) and (B) refer to the Venn diagram of DEGs identified in the lung and heart, respectively, that were infected by GX19-6, GX19-5, and CS-6. (C)∼(E) refer to the enriched biological processes of GX19-6-specific DEGs in the mice's lung and heart compared to GX19-5 and CS-6. The GO terms were clustered, and each subpanel showed the number of GO terms in each cluster. (F) and (G) refer to the enriched biological processes of the top 100 GX19-6-exclusive up-- and down-regulated genes, respectively. (H) Comparison of fold changes of inflammation response genes in the mice's lung, heart, and brain between different viruses using the paired Wilcoxon rank-sum test. ns, no significant; *, p-value <0.05; **, p-value <0.01; ***, p-value <0.001. (I) Fold change of inflammation response genes in the mice's lung, heart, and brain after virus infections. (J) Diagram of a hypothesis about the rapid death of mice caused by the GX19 virus.