Fig. 8: Type I and type III IFNs protect bat organoids from zoonotic virus infection.
From: Bat organoids reveal antiviral responses at epithelial surfaces
a, Experimental workflow showing the administration of universal IFNα2, IFNλ1-like or IFNλ3-like 12 h before, during or 8 h after treatment with VSV infection of bat nasalORG or SIORG. Intracellular VSV viral RNA was measured 24 h after infection. b,c, RT–qPCR analysis of intracellular VSV-NP viral RNA in bat SIORG (n = 3) (b) or nasalORG (n = 3) (c), normalized to EEF1A1 (2−ΔCt) and expressed as a percentage relative to no IFN control. A two-sided Student’s t-test was used to compare VSV viral RNA levels across before, during and after treatment, and no IFN conditions, comparing each IFN treatment to no IFN control. d, Experimental workflow showing bat nasalORG or SIORG treated with universal IFNα2 or bat IFNλ1-like for 3 h, followed by washout and incubation in IFN-free medium for 24 h and RNA collection at 0, 3 and 24 h after IFN washout. e,f, RT–qPCR analysis of ISGs (normalized to EEF1A1) in bat SIORG (n = 3) (e) or nasalORG (n = 3) (f) treated with universal IFNα2 or bat IFNλ1-like at 3 h (left) or 24 h (right) after IFN removal. A two-sided Student’s t-test was used to compare ISG mRNA levels between treatments. g, RT–qPCR analysis of intracellular SINV-eGFPnsP2-P726G nsP1 RNA (normalized to EEF1A1 (2−ΔCt) and expressed as fold change relative to the mean of the sgScrambled control) in bat SIORG engineered with Cas9 and targeted guide RNA (sgIRF9, sgIFNAR2, sgIFNLR1) or control (sgScrambled), infected with SINV-eGFPnsP2-P726G. A two-sided Student’s t-test was used to compare SINV nsP1 viral RNA levels between treatment groups. nsP1, nonstructural protein 1. h, RT–qPCR analysis of intracellular MARV-L viral RNA (normalized to EEF1A1) in bat SIORG, nasalORG or alvORG (n = 4) treated with or without IFNs during a 3-day MARV infection. A two-sided Student’s t-test was used to compare MARV viral RNA levels between IFN-treated and untreated controls. i, RT–qPCR analysis of MARV-L viral RNA in bat alvORG treated with or without 5 µM ruxolitinib during a 3-day infection. A two-sided Student’s t-test was used to compare MARV viral RNA levels between ruxolitinib and dimethylsulfoxide (DMSO) mock-treated alvORG and MARV-infected alvORG. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.
