Animal viruses with separately packaged RNA segments

dose-response-plaque-assayThere are many examples of viruses with segmented genomes – like influenza viruses – but these genomes segments are packaged in  one virus particle. Sometimes the genome segments are separately packaged in virus particles. Such multicomponent viruses are commonly found to infect plants and fungi, but only recently have examples of such viruses that infect animals been discovered (paper link).
Sequence analysis of viruses isolated from Culex mosquitoes in Central and South American Countries revealed six new viruses with segmented RNA genomes, which was confirmed by gel electrophoresis of RNA extracted from virus particles.
Some of the virus isolates appear to lack the fifth RNA segment, and the results of RNA transfection experiments indicated that this RNA is not needed for viral infectivity.
RNA viruses with segmented genomes are common, but in this case, the surprise came when it was found that the dose-response curve of infection for these viruses was not linear. In other words, one virus particle was not sufficient to infect a cell (illustrated). In this case, between 3 and 4 particles were needed to establish an infection. These findings indicate that the viral RNA segments are separately packaged, and must enter a cell together to initiation infection.
These novel viruses, called Guaico Culex virus (GCXV) are distantly related to flaviviruses, a family of non-segmented, + strand RNA viruses. They are part of a clade of RNA viruses with segmented genomes called the Jingmenvirus, which includes a novel tick-borne virus isolated in China (previously discussed on this blog), and a variant isolated from a red colobus monkey in Uganda. These viruses are also likely to have genomes that are separately packaged.
An interesting question is to identify the selection that lead to the emergence of  multicomponent viruses that require multiple particles to initiate an infection. Perhaps transmission of these types of viruses by insect vectors facilitates the introduction of multiple virus particles into a cell. How such viral genomes emerged and persisted remains a mystery that might be solved by the analysis of other viruses with similar genome architectures.

TWiV 398: Permission to be intuitive

Vincent speaks with Sandy Weller about her career and her work on the mechanisms of synthesis, maturation, cleavage and packaging of viral DNA genomes.

You can find TWiV 398 at, or listen/watch below.

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TWiV 353: STING and the antiviral police

On episode #353 of the science show This Week in Virology, the TWiVniacs discuss twenty-eight years of poliovirus shedding by an immunodeficient patient, and packaging of the innate cytoplasmic signaling molecule cyclic GMP-AMP in virus particles.

You can find TWiV #353 at

Packaging of the segmented influenza RNA genome

The RNA genome of influenza viruses is segmented . The virions of influenza A and B viruses contain 8 different RNAs, while those of influenza C viruses contain 7. How is the correct number of RNA segments inserted into newly synthesized virus particles?

During influenza virus assembly, viral RNAs and viral proteins – called a ribonucleoprotein complex or RNP –  travels to the plasma membrane. There the virion forms by a process called budding, during which the membrane bulges from the cell and is eventually pinched off to form a free particle.


Production of an infectious virus particle requires incorporation of at least one copy of each of the eight RNA segments. Two different mechanisms – random and selective packaging – have been proposed to explain how each virion receives a full complement of genomic RNA.

If the 8 influenza viral RNA segments were randomly packaged into new particles, we would expect to observe 1 infectious particle for every 400 particles assembled (8!/88). This ratio falls within the range of infectious to noninfectious particles that occur in virus stocks. If more than 8 RNA segments could be packaged into each virion, then the fraction of infectious particles would be significantly increased. For example, if 12 RNA molecules could fit into each virion, then 10% of the particles would have the complete viral genome. In support of this mechanism, influenza viruses with more than 8 RNA segments have been observed.

In the selective packaging mechanism, each of the eight genomic RNAs has a different signal that allows incorporation into virus particles. These signals are believed to be within the noncoding and coding sequences at the 5′- and 3′-ends of the viral RNAs. The sequences interact and form structures that are unique to each segment, and which have been shown to be essential for incorporation of each segment into virions. Consistent with this hypothesis, electron microscopy reveals that during budding, the viral RNPs are organized in a distinct pattern, as shown in the image.


This observation argues that RNPs are not randomly incorporated into virions, and is consistent with the presence of specific signals in each RNA segment that enable the RNPs to be packaged as a complete set. The mechanisms by which these signals are recognized, and how they ensure incorporation of one copy of each RNA segment into the particle, are not known.

There is clear evidence for a selective mechanism during the packaging of the bacteriophage ψ6 genome. Viral particles contain one copy each of a S, M, and L dsRNA segment. All particles contain a complete complement of genome segments, as indicated by the fact that every virus particle is infectious. Only the S RNA segment can enter newly formed particles; once that segment is packaged, then the M RNA can enter. The L RNA can only enter particles that contain both the S and M segments. Precise packaging is therefore the result of a serial dependence of packaging of the RNA segments.

Muramoto, Y., Takada, A., Fujii, K., Noda, T., Iwatsuki-Horimoto, K., Watanabe, S., Horimoto, T., Kida, H., & Kawaoka, Y. (2006). Hierarchy among Viral RNA (vRNA) Segments in Their Role in vRNA Incorporation into Influenza A Virions Journal of Virology, 80 (5), 2318-2325 DOI: 10.1128/JVI.80.5.2318-2325.2006

Noda, T., Sagara, H., Yen, A., Takada, A., Kida, H., Cheng, R., & Kawaoka, Y. (2006). Architecture of ribonucleoprotein complexes in influenza A virus particles Nature, 439 (7075), 490-492 DOI: 10.1038/nature04378

Frilander, M. (1995). In Vitro Packaging of the Single-stranded RNA Genomic Precursors of the Segmented Double-stranded RNA Bacteriophage ψ6: The Three Segments Modulate Each Other’s Packaging Efficiency Journal of Molecular Biology, 246 (3), 418-428 DOI: 10.1006/jmbi.1994.0096