Molecular Biology and Epidemiology 
for Control of Rotavirus Diarrhea 
Carlos F. Arias, Ph.D. — International Research Scholar 
Dr. Arias is Investigador Titular B, Department of Molecular Biology, at the Biotechnology Institute, 
National Autonomous University of Mexico, Cuernavaca. He received his undergraduate degree in 
biochemistry and his M.S. and Ph.D. degrees in biomedical research from the National University of 
Mexico, Mexico City. He recently conducted sabbatical research with James Strauss at the California 
Institute of Technology. 
ACUTE, infectious diarrhea is the commonest 
Lcause of morbidity and mortality among 
young children living in developing countries, 
accounting for as many as one billion illnesses 
and between four and five million deaths an- 
nually. Rotaviruses are the leading cause of se- 
vere diarrheal disease in children under three 
years of age, and it is estimated that an effective 
vaccine would save about 800,000 children's 
lives per year. 
Because rotavirus plays such an important role 
in severe infantile gastroenteritis, and because 
even advanced levels of hygiene seem unable to 
control the spread of rotavirus infections, there 
has been considerable interest in developing ef- 
fective vaccination strategies. 
Our laboratory is interested in studying how 
rotaviruses attach and enter their host cell and 
how they replicate there to produce the viral 
progeny. Also among our interests are the host 
immune response to rotavirus infection and the 
epidemiology of these viruses in Mexico. 
As in other infectious agents, proteins located 
at the surface of the rotavirus particles are in- 
volved in the early interactions (attachment and 
penetration) of the virions with the host cell. 
These proteins contain antigenic determinants 
that represent major immunological targets. The 
structural and functional characterization of the 
viral surface proteins, as well as the study of im- 
mune response in the infected host, should con- 
tribute to the development of rationally designed 
therapeutic agents and new prevention strate- 
gies. Moreover, the success of a vaccine may de- 
pend on knowledge of the epidemiology of the 
pathogen in the particular geographic area where 
the vaccine is to be used. 
The surface of the rotavirus contains two pro- 
teins, VP4 and VP7. VP4 forms spikes that extend 
from the viral surface. This protein has been asso- 
ciated with a variety of viral functions, including 
the determination of virulence in vivo and the 
ability to agglutinate red blood cells (hemagglu- 
tination). VP4 is also important in the penetra- 
tion of the virion into the cell. On the other hand. 
the protein responsible for the initial attachment 
of rotavirus to its host cell remains controversial; 
both VP4 and VP7 have been proposed. 
The attachment of animal rotaviruses to cells in 
culture is mediated by compounds containing 
sialic acid, since treatment of cells with sialidases 
greatly reduces the binding of virus particles to 
the cell surface. In addition, the hemagglutinat- 
ing activity of rotaviruses and their binding to 
cultured epithelial cells can also be inhibited by 
incubation of the virus with sialoglycoproteins 
such as glycophorin. 
We have isolated rotavirus mutants whose bind- 
ing is no longer inhibited by treatment of cells 
with sialidases or incubation with glycophorin. 
The detailed analysis of these mutants should 
give us information about the protein (s) in- 
volved in the attachment of the virions to the cell. 
Furthermore, we are constructing deletion mu- 
tants and chimeras between VP4 genes of hemag- 
glutinating and nonhemagglutinating rotavi- 
ruses, to define further the protein domains 
involved in the binding of the virus to epithelial 
and red blood cells. 
After the virus's initial attachment to the cell 
surface, the next step in its infection cycle is to 
enter the cell. The entry of the virus particle can 
be augmented by treating it with trypsin, and 
probably is trypsin dependent. Experiments in 
progress have shown that this proteolytic treat- 
ment induces three specific cleavages of VP4. We 
are interested in identifying the cleavage (s) di- 
rectly associated with enhancement of viral infec- 
tivity and in learning about the mechanism 
through which the cleaved VP4 protein mediates 
the virion's penetration of the cell. 
Both surface proteins have also been character- 
ized immunologically. It has been found that anti- 
bodies to either protein neutralize the virus in 
vitro and are capable of passively protecting ani- 
mals from rotavirus challenge. In addition, oral 
infection with live rotavirus stimulates protec- 
tive immunity, which can be mediated by VP4 
and/or VP7. These observations make the two 
proteins attractive candidates for recombinant 
subunit vaccines. 
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