Chikungunya virus (CHIKV) is an alphavirus responsible for numerous epidemics in Africa and Asia. Infection by CHIKV is often characterized by long-lasting, incapacitating arthritis, and some fatal cases have been described among elderly and newborns. Currently, there is no available vaccine or specific treatment against CHIKV. Blood B cells from a donor with history of CHIKV infection were activated, immortalized, amplified, and cloned. Two human mAbs against CHIKV, 5F10 and 8B10, were identified, sequenced, and expressed in recombinant form for characterization. In a plaque reduction neutralization test, 5F10 and 8B10 show mean IC50 of 72 and 46 ng/ml, respectively. Moreover, both mAbs lead to a strong decrease in extracellular spreading of infectious viral particles from infected to uninfected cells. Importantly, the mAbs neutralize different CHIKV isolates from Singapore, Africa, and Indonesia, as well as O’nyong-nyong virus, but do not recognize other alphaviruses tested. Both mAbs are specific for the CHIKV envelope: 5F10 binds to the E2 glycoprotein ectodomain and 8B10 to E1 and/or E2. In conclusion, these two unique human mAbs strongly, broadly, and specifically neutralize CHIKV infection in vitro and might become possible therapeutic tools against CHIKV infection, especially in individuals at risk for severe disease. Importantly, these mAbs will also represent precious tools for future studies on host–pathogen interactions and the rational design of vaccines against CHIKV.

Volume 80, no. 1, p. [85][86][87][88][89][90][91][92][93][94] 2006. Page 86: We have recently realized that a protein sequence alignment comparing the B2 proteins of greasy grouper nervous necrosis virus (GGNNV) and flock house virus (FHV) is erroneous. The FHV sequence used in the article was not that of B2 but rather the frameshifted B1 protein. We have corrected the alignment using the FHV B2 sequence. Our conclusions regarding the low level of sequence similarity between the two proteins remain unchanged. The updated Fig. 1 and legend are shown below.FIG. 1. Amino acid sequences derived from previously published sequences (9, 16, 28) were aligned by using ClustalW, yielding sequence identities of 10.2 and 6.6% and similarities of 13.9 and 16.0% to NoV and FHV, respectively. 4909on May 9, 2018 by guest

Betanodaviruses are small positive-sense bipartite RNA viruses that infect a wide variety of fish species and are notorious for causing lethal outbreaks in juvenile fish hatcheries worldwide. The function of a small nonstructural protein, B2, encoded by the subgenomic RNA3 of betanodaviruses, has remained obscure. Greasy grouper nervous necrosis virus, a betanodavirus model, was used to develop a facile DNA-based reverse genetics system that recapitulated the virus infection cycle, and we used this system to show that B2 is a small nonstructural protein that is essential for high level accumulation of viral RNA1 after RNA transfection of fish, mammalian, and avian cells. The defect in RNA1 accumulation in a B2 mutant was partially complemented by supplying B2 RNA in trans. Confocal analysis of the cellular distribution of B2 indicated that B2 is able to enter the nucleus and accumulates there during the late stages of GGNNV infection. Using human HeLa cells as a cellular RNA interference model, we found that B2 could efficiently antagonize RNA interference, which is a property shared by the distantly related alphanodavirus B2 proteins. This function provides appears to provide an explanation, at least in part, for why B2 mutant RNA1 is severely impaired in its intracellular accumulation. Greasy grouper nervous necrosis virus (GGNNV) is an RNA virus that elicits nervous necrosis upon infection of a variety of fish species (28). GGNNV belongs to the beta subgroup of the virus family Nodaviridae. The nodavirus genome consists of two single-stranded positive-sense RNA molecules, the 3.1-kb RNA1 and the 1.4-kb RNA2. RNA1 encodes an RNA-dependent RNA polymerase (RdRp or protein A), while RNA2 encodes the coat protein, ␣ (25). In addition, a subgenomic RNA transcribed from the 3Ј end of RNA1, termed RNA3 (10, 26), encodes a protein of unknown function called B2.We have previously investigated the subcellular localization of protein A and found that it targets mitochondrial membranes via an N-terminal stretch of approximately 40 amino acids (10), which is also the case with other nodaviruses (23,24). Furthermore, we have characterized the nucleolus localization and apoptotic inducer domains within the ␣ coat protein of the GGNNV (11, 12). However, the function and properties of the remaining GGNNV protein, B2, have remained a mystery.The betanodaviruses of fish studied to date all possess putative B2 proteins with very high sequence homology. The betanodavirus B2 proteins consist of only 75 amino acids and do not possess any characteristic sequence motifs or domains. Several of the alphanodavirus B2 proteins, however, have been characterized and appear to be involved in the suppression of cellular RNA interference, which is now known to act as an intracellular poison against virus infection (7). The B2 protein of Nodamura virus (NoV), which infects both insects and rodents, is required for maximal autonomous replication of RNA1 and RNA3 after transfection of a wide variety of cell lines with RNA1 transcripts synthesized...