Fig. 1

Experimental design and identification of coronavirus-derived RNA and proteins in cat flea. Representative images of a laboratory-reared domestic cat flea and tissues used for extraction of RNA and plasma membrane proteins for RT-PCR and proteomics analysis. Schematic representation of the coronavirus genome organization and virion structure based on SARS-CoV-2 [15]. Coronavirus proteins identified by proteomics analysis included ORF1a, ORF1b, protein M and protein 7b. Genomic RNA (gRNA) serves as mRNA for ORF1a and ORF1b. Other major subgenomic RNAs (sgRNAs) are produced to encode for envelope (e.g. protein M) and accessory (e.g. protein 7b) proteins in addition to the gRNA. Real-time RT-PCR targeting ORF1b identified in coronavirus-derived RNA in laboratory-reared unfed cat flea (sample 2) and feral cat flea (sample 3). MW, molecular weight O’GeneRuler 1 kb Plus DNA Ladder (Thermo Fisher Scientific); sample 1, laboratory-reared unfed cat flea, samples 4 and 5, positive controls (Table 1); samples 6 and 7, nuclease-free water negative controls