Quantum dots (QDs)-based single particle analysis technique enables real-time tracking of the viral infection in live cells with great sensitivity over a long period of time. The porcine reproductive and respiratory syndrome virus (PRRSV) is a small virus with the virion size of 40–60 nm which causes great economic losses to the swine industry worldwide. A clear understanding of the viral infection mechanism is essential for the development of effective antiviral strategies. In this study, we labeled the PRRSV with QDs using the streptavidin–biotin labeling system and monitored the viral infection process in live cells. Our results indicated that the labeling method had negligible effect on viral infectivity. We also observed that prior to the entry, PRRSV vibrated on the plasma membrane, and entered the cells via endosome mediated cell entry pathway. Viruses moved in a slow–fast–slow oscillatory movement pattern and finally accumulated in a perinuclear region of the cell. Our results also showed that once inside the cell, PRRSV moved along the microtubule, microfilament and vimentin cytoskeletal elements. During the transport process, virus particles also made contacts with non-muscle myosin heavy chain II-A (NMHC II-A), visualized as small spheres in cytoplasm. This study can facilitate the application of QDs in virus infection imaging, especially the smaller-sized viruses and provide some novel and important insights into PRRSV infection mechanism.
This is a preview of subscription content, access via your institution.
Buy single article
Instant access to the full article PDF.
Price excludes VAT (USA)
Tax calculation will be finalised during checkout.
Bretscher A (1991) Microfilament structure and function in the cortical cytoskeleton. Annu Rev Cell Biol 7:337–374
Bulgakov AD, Grebennikova TV, Iuzhakov AG, Aliper TI, Nepoklonov EA (2014) Molecular-genetic analysis of the genomes of porcine reproductive and respiratory syndrome virus and porcine circovirus type 2 circulating in the area of Russian federation. Mol Genet Mikrobiol Virusol 4:29–33
Chandrasekar I, Goeckeler ZM, Turney SG, Wang P, Wysolmerski RB, Adelstein RS, Bridgman PC (2014) Nonmuscle myosin II is a critical regulator of clathrin-mediated endocytosis. Traffic 15:418–432
Conzelmann KK, Visser N, Van Woensel P, Thiel HJ (1993) Molecular characterization of porcine reproductive and respiratory syndrome virus, a member of the arterivirus group. Virology 193:329–339
Dumas A, Le-Bury G, Marie-Anais F, Herit F, Mazzolini J, Guilbert T, Bourdoncle P, Russell DG, Benichou S, Zahraoui A, Niedergang F (2015) The HIV-1 protein Vpr impairs phagosome maturation by controlling microtubule-dependent trafficking. J Cell Biol 211:359–372
Gorp HV, Breedam WV, Doorsselaere JV, Delputte PL, Nauwynck HJ (2010) Identification of the CD163 protein domains involved in infection of the porcine reproductive and respiratory syndrome virus. J Virol 84:3101–3105
Huang BH, Lin Y, Zhang ZL, Zhuan F, Liu AA, Xie M, Tian ZQ, Zhang Z, Wang H, Pang DW (2012) Surface labeling of enveloped viruses assisted by host cells. ACS Chem Biol 7:683–688
Hou W, Li Y, Kang W, Wang X, Wu X, Wang S, Liu F (2019) Real-time analysis of quantum dot labeled single porcine epidemic diarrhea virus moving along the microtubules using single particle tracking. Sci Rep 9:1307
Jiang YF, Xia TQ, Zhou YJ, Yu LX, Yang S, Huang QF, Li LW, Gao F, Qu ZH, Tong W, Tong GZ (2015) Characterization of three porcine reproductive and respiratory syndrome virus isolates from a single swine farm bearing strong homology to a vaccine strain. Vet Microbiol 179:242–249
Joo KI, Fang Y, Liu Y, Xiao L, Gu Z, Tai A, Lee CL, Tang Y, Wang P (2011) Enhanced real-time monitoring of adeno-associated virus trafficking by virus-quantum dot conjugates. ACS Nano 5:3523–3535
Kreutz LC, Ackermann MR (1996) Porcine reproductive and respiratory syndrome virus enters cells through a low pH-dependent endocytic pathway. Virus Res 42:137–147
Lakadamyali M, Rust MJ, Babcock HP, Zhuang X (2003) Visualizing infection of individual influenza viruses. Proc Natl Acad Sci USA 100:9280–9285
Liu SL, Zhang ZL, Tian ZQ, Zhao HS, Liu H, Sun EZ, Xiao GF, Zhang W, Wang HZ, Pang DW (2012) Effectively and efficiently dissecting the infection of influenza virus by quantum-dot-based single-particle tracking. ACS Nano 6:141–150
Liu SL, Zhang LJ, Wang ZG, Zhang ZL, Wu QM, Sun EZ, Shi YB, Pang DW (2014) Globally visualizing the microtubule-dependent transport behaviors of influenza virus in live cells. Anal Chem 86:3902–3908
Liu SL, Wang ZG, Zhang ZL, Pang DW (2016) Tracking single viruses infecting their host cells using quantum dots. Chem Soc Rev 45:1211–1224
Lunney JK, Fang Y, Ladinig A, Chen N, Li Y, Rowland B, Renukaradhya GJ (2016) Porcine reproductive and respiratory syndrome virus (PRRSV): pathogenesis and interaction with the immune system. Annu Rev Anim Biosci 4:129–154
Mu C, Lu X, Duan E, Chen J, Li W, Zhang F, Martin DP, Yang M, Xia P, Cui B (2013) Molecular evolution of porcine reproductive and respiratory syndrome virus isolates from central China. Res Vet Sci 95:908–912
Murakami M, Imabayashi K, Watanabe A, Takeuchi N, Ishizaka R, Iohara K, Yamamoto T, Nakamura H, Nakashima M (2012) Identification of novel function of vimentin for quality standard for regenerated pulp tissue. J Endod 38:920–926
Nathan L, Daniel S (2019) Single virion tracking microscopy for the study of virus entry processes in live cells and biomimetic platforms. Adv Exp Med Biol 1215:13–43
Nelsen CJ, Murtaugh MP, Faaberg KS (1999) Porcine reproductive and respiratory syndrome virus comparison: divergent evolution on two continents. J Virol 73:270–280
Simmonds P, Domingo E (2011) Virus evolution. Curr Opin Virol 1:410–412
Song T, Fang L, Wang D, Zhang R, Zeng S, An K, Chen H, Xiao S (2016) Quantitative interactome reveals that porcine reproductive and respiratory syndrome virus nonstructural protein 2 forms a complex with viral nucleocapsid protein and cellular vimentin. J Proteom 142:70–81
Stankevicius A, Buitkuviene J, Sutkiene V, Spancerniene U, Pampariene I, Pautienius A, Oberauskas V, Zilinskas H, Zymantiene J (2016) Detection and molecular characterization of porcine reproductive and respiratory syndrome virus in lithuanian wild boar populations. Acta Vet Scand 58:51
Su Y, Qiao W, Guo T, Tan J, Li Z, Chen Y, Li X, Li Y, Zhou J, Chen Q (2010) Microtubule-dependent retrograde transport of bovine immunodeficiency virus. Cell Microbiol 12:1098–1107
Sun EZ, Liu AA, Zhang ZL, Liu SL, Tian ZQ, Pang DW (2017) Real-time dissection of distinct dynamin-dependent endocytic routes of influenza a virus by quantum dot-based single-virus tracking. ACS Nano 11:4395–4406
Taylor MP, Burgon TB, Kirkegaard K, Jackson WT (2009) Role of microtubules in extracellular release of poliovirus. J Virol 83:6599–6609
Wang WW, Zhang L, Ma XC, Gao JM, Xiao YH, Zhou EM (2011) The role of vimentin during PRRSV infection of Marc-145 cells. Chin J Virol 27:456–461
Wang F, Qiu H, Zhang Q, Peng Z, Liu B (2012) Association of two porcine reproductive and respiratory syndrome virus (PRRSV) receptor genes, CD163 and SN with immune traits. Mol Biol Rep 39:3971–3976
Wang ZY, Huang X, Liu DH, Lu HL, Kim YC, Xu WX (2015) Involvement of actin microfilament in regulation of pacemaking activity increased by hypotonic stress in cultured ICCs of murine intestine. Physiol Res 64:397–405
Wang T, Zheng Z, Zhang X-E, Wang H (2016) Quantum dot-fluorescence in situ hybridisation for Ectromelia virus detection based on biotin–streptavidin interactions. Talanta 158:179–184
Wells KD, Bardot R, Whitworth KM, Trible BR, Fang Y, Mileham A, Kerrigan MA, Samuel MS, Prather RS, Rowland RR (2017) Replacement of porcine CD163 scavenger receptor cysteine-rich domain 5 with a CD163-like homolog confers resistance of pigs to genotype 1 but not genotype 2 porcine reproductive and respiratory syndrome virus. J Virol 91(2):e01521-16
Wen L, Zheng ZH, Liu AA, Lv C, Zhang LJ, Ao J, Zhang ZL, Wang HZ, Lin Y, Pang DW (2017) Tracking single baculovirus retrograde transportation in host cell via quantum dot-labeling of virus internal component. J Nanobiotechnol 15:37
Zhang Q, Yoo D (2015) PRRS virus receptors and their role for pathogenesis. Vet Microbiol 177:229–241
Zhang F, Zheng Z, Liu SL, Lu W, Zhang Z, Zhang C, Zhou P, Zhang Y, Long G, He Z, Pang DW, Hu Q, Wang H (2013a) Self-biotinylation and site-specific double labeling of baculovirus using quantum dots for single-virus in situ tracking. Biomaterials 34:7506–7518
Zhang Y, Ke X, Zheng Z, Zhang C, Zhang Z, Zhang F, Hu Q, He Z, Wang H (2013b) Encapsulating quantum dots into enveloped virus in living cells for tracking virus infection. ACS Nano 7:3896–3904
Zheng Z, Wang H (2013) Tracking viral infection: will quantum dot encapsulation unveil viral mechanisms? Nanomedicine (Lond) 8:1225–1227
Zheng LL, Li CM, Zhen SJ, Li YF, Huang CZ (2017) A dynamic cell entry pathway of respiratory syncytial virus revealed by tracking the quantum dot-labeled single virus. Nanoscale 9:7880–7887
The authors would like to thank for the support from the National Natural Science Foundation of China (Grant Nos. 31570151 and 31490601), the Program for Science and Technology Innovation Talents in Universities of Henan Province (Grant No. 17HASTIT039), the Key Scientific Research Project of Henan Province Higher Education (16A180044) and the Open Research Fund Program of the State Key Laboratory of Virology of China (Grant No. 2017KF005).
Conflict of interest
The authors declare that they have no conflict of interest.
Animal and Human Rights Statement
This article does not contain any studies with human or animal subjects performed by any of the authors.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Supplementary material 2(mp4 6866 kb)
Supplementary material 3 (mp4 36172 kb)
Supplementary material 4 (mp4 10023 kb)
Supplementary material 5 (mp4 13211 kb)
Supplementary material 6 (mp4 6730 kb)
Supplementary material 7 (mp4 8759 kb)
Rights and permissions
About this article
Cite this article
Liang, Z., Li, P., Wang, C. et al. Visualizing the Transport of Porcine Reproductive and Respiratory Syndrome Virus in Live Cells by Quantum Dots-Based Single Virus Tracking. Virol. Sin. 35, 407–416 (2020). https://doi.org/10.1007/s12250-019-00187-0
- Single virus tracking
- Quantum dots (QDs)
- Porcine reproductive and respiratory syndrome virus (PRRSV)