Dengue virus requires apoptosis linked gene-2-interacting protein X (ALIX) for viral propagation.

The endosomal sorting complexes required for transport (ESCRT) pathway accessory protein apoptosis linked gene-2-interacting protein X (ALIX) has been shown to be upregulated during dengue virus (DENV) replication. Yeast-two-hybrid screens have additionally shown that ALIX interacts with DENV NS3 protein, but evaluation of the interaction through a replicon assay failed to show a functional significance to the interaction. In this study the interaction between DENV NS3 and ALIX was investigated by co-immunoprecipitation, and functional significance assessed by investigation of DENV production in ALIX expression regulated cells. The results showed that ALIX both interacted and co-localized with DENV NS3 protein and that upregulation of ALIX resulted in a significantly increased viral titer, while either siRNA or CRISPR-Cas9 mediated down regulation of ALIX significantly reduced viral production, without affecting relative DENV genome levels. These results are consistent with ALIX playing a significant role in the DENV replication cycle either during late infection or at viral egress.

Actin Interacts with Dengue Virus 2 and 4 Envelope Proteins.

Dengue virus (DENV) remains a significant public health problem in many tropical and sub-tropical countries worldwide. The DENV envelope (E) protein is the major antigenic determinant and the protein that mediates receptor binding and endosomal fusion. In contrast to some other DENV proteins, relatively few cellular interacting proteins have been identified. To address this issue a co-immuoprecipitation strategy was employed. The predominant co-immunoprecipitating proteins identified were actin and actin related proteins, however the results suggested that actin was the only bona fide interacting partner. Actin was shown to interact with the E protein of DENV 2 and 4, and the interaction between actin and DENV E protein was shown to occur in a truncated DENV consisting of only domains I and II. Actin was shown to decrease during infection, but this was not associated with a decrease in gene transcription. Actin-related proteins also showed a decrease in expression during infection that was not transcriptionally regulated. Cytoskeletal reorganization was not observed during infection, suggesting that the interaction between actin and E protein has a cell type specific component.

Delayed antibody dependent enhancement of low passage dengue virus 4 isolates.

BACKGROUND: The concept of antibody dependent enhancement (ADE) of dengue virus (DENV) infection is a cornerstone of our current understanding of dengue pathogenesis, although some questions as to the mechanism remain, particularly in regards to the behavior of low and high passage virus isolates. This study utilized two low passage DENV 4 isolates and a laboratory adapted DENV 4 isolate to investigate the potential of low passage isolates to undergo ADE. RESULTS: Little or no ADE of infection was observed on day 2 post infection with low passage isolates, while high enhancement of infection was seen with the laboratory adapted virus. However, both of the low passage isolates showed high levels of infection (60-100%) by day 5 post infection. CONCLUSIONS: These results show that low passage DENV 4 viruses undergo ADE mediated infection, but that the process is significantly temporally delayed as compared to laboratory adapted DENV 4.

Retrospective screening of acute undifferentiated fever serum samples with universal flavivirus primers.

INTRODUCTION: Fever is a common symptom of many tropical diseases and in many cases the etiologic agent remains unidentified as a consequence of either the etiologic agent not being part of routine diagnostic screening or as a consequence of false negatives on standard diagnostic tests. METHODOLOGY: This study screened a well characterized panel of 274 serum samples collected on day of admission from adult patients with acute undifferentiated fever admitted to a hospital in Nakhon Ratchasima, Thailand by RT-PCR using pan-flavivirus degenerate primers. RESULTS: Subsequent clinical diagnosis was achieved for 38 of the patients, and included 19 cases of dengue fever. RT-PCR screening identified seven positive samples (2.5%) which were revealed by sequence analysis to be dengue virus 1 (2 cases), dengue virus 2 (2 cases) and dengue virus 3 (3 cases). Only 5 out of 19 (26%) serum samples from patients subsequently diagnosed with dengue were positive, but 2 samples which clinically remained undiagnosed were shown to be positive for dengue virus. Sequence analysis suggested that the dengue virus 3 cases occurred as a result of importation of a strain of dengue from India or China. No other flaviviruses were identified. CONCLUSIONS: No evidence was found of other flaviviruses besides dengue circulating in this population. Despite improved diagnostic tests, cases of dengue are still evading correct diagnosis.