Kimberly B. Hummel

Genetic variability of the glycoprotein genes of current wild-type measles isolates

The glycoprotein coding sequences from three wild-type measles viruses isolated in the United States during 1988-1989 were examined by mRNA templated sequencing to determine whether contemporary strains have undergone genetic changes relative to the vaccine strain, Moraten. These studies revealed variation in the hemagglutinin (HA) gene and, to a far lesser degree, the fusion (F) gene. The F protein coding region was highly conserved with only three predicted amino acid changes. Among the predicted amino acid changes identified in the HA was a new potential glycosylation site at residue 416, located toward the carboxy-terminal end of the HA peptide. Eighty percent of the predicted amino acid changes in the HA shared by the three wild-type isolates were clustered near the five previously identified potential glycosylation sites. A linear pattern of evolutionary change was observed after comparing the predicted amino acid HA changes from the 1988-1989 viruses to those predicted in the HA protein from U.S. wild types isolated in 1977 and 1983.

Lack of Association between Measles Virus Vaccine and Autism with Enteropathy: A Case-Control Study

BACKGROUND The presence of measles virus (MV) RNA in bowel tissue from children with autism spectrum disorders (ASD) and gastrointestinal (GI) disturbances was reported in 1998. Subsequent investigations found no associations between MV exposure and ASD but did not test for the presence of MV RNA in bowel or focus on children with ASD and GI disturbances. Failure to replicate the original study design may contribute to continued public concern with respect to the safety of the measles, mumps, and rubella (MMR) vaccine. METHODOLOGY/PRINCIPAL FINDINGS The objective of this case-control study was to determine whether children with GI disturbances and autism are more likely than children with GI disturbances alone to have MV RNA and/or inflammation in bowel tissues and if autism and/or GI episode onset relate temporally to receipt of MMR. The sample was an age-matched group of US children undergoing clinically-indicated ileocolonoscopy. Ileal and cecal tissues from 25 children with autism and GI disturbances and 13 children with GI disturbances alone (controls) were evaluated by real-time reverse transcription (RT)-PCR for presence of MV RNA in three laboratories blinded to diagnosis, including one wherein the original findings suggesting a link between MV and ASD were reported. The temporal order of onset of GI episodes and autism relative to timing of MMR administration was examined. We found no differences between case and control groups in the presence of MV RNA in ileum and cecum. Results were consistent across the three laboratory sites. GI symptom and autism onset were unrelated to MMR timing. Eighty-eight percent of ASD cases had behavioral regression. CONCLUSIONS/SIGNIFICANCE This study provides strong evidence against association of autism with persistent MV RNA in the GI tract or MMR exposure. Autism with GI disturbances is associated with elevated rates of regression in language or other skills and may represent an endophenotype distinct from other ASD.

Characterization of Nipah Virus from Outbreaks in Bangladesh, 2008-2010

New genotyping scheme facilitates classification of virus sequences.

Real-Time Reverse Transcription-PCR Assay for Detection of Mumps Virus RNA in Clinical Specimens

ABSTRACT The mumps virus is a negative-strand RNA virus in the family Paramyxoviridae. Mumps infection results in an acute illness with symptoms including fever, headache, and myalgia, followed by swelling of the salivary glands. Complications of mumps can include meningitis, deafness, pancreatitis, orchitis, and first-trimester abortion. Laboratory confirmation of mumps infection can be made by the detection of immunoglobulin M-specific antibodies to mumps virus in acute-phase serum samples, the isolation of mumps virus in cell culture, or by detection of the RNA of the mumps virus by reverse transcription (RT)-PCR. We developed and validated a multiplex real-time RT-PCR assay for rapid mumps diagnosis in a clinical setting. This assay used oligonucleotide primers and a TaqMan probe targeting the mumps SH gene, as well as primers and a probe that targeted the human RNase P gene to assess the presence of PCR inhibitors and as a measure of specimen quality. The test was specific, since it did not amplify a product from near-neighbor viruses, as well as sensitive and accurate. Real-time RT-PCR results showed 100% correlation with results from viral culture, the gold standard for mumps diagnostic testing. Assay efficiency was over 90% and displayed good precision after performing inter- and intraassay replicates. Thus, we have developed and validated a molecular method for rapidly diagnosing mumps infection that may be used to complement existing techniques.

The C, V and W proteins of Nipah virus inhibit minigenome replication.

Nipah virus (NiV) is a recently emergent, highly pathogenic, zoonotic paramyxovirus of the genus Henipavirus. Like the phosphoprotein (P) gene of other paramyxoviruses, the P gene of NiV is predicted to encode three additional proteins, C, V and W. When the C, V and W proteins of NiV were tested for their ability to inhibit expression of the chloramphenicol acetyltransferase (CAT) reporter gene in plasmid-based, minigenome replication assays, each protein inhibited CAT expression in a dose-dependent manner. The C, V and W proteins of NiV also inhibited expression of CAT from a measles virus (MV) minigenome, but not from a human parainfluenzavirus 3 (hPIV3) minigenome. Interestingly, the C and V proteins of MV, which have previously been shown to inhibit MV minigenome replication, also inhibited NiV minigenome replication; however, they were not able to inhibit hPIV3 minigenome replication. In contrast, the C protein of hPIV3 inhibited minigenome replication of hPIV3, NiV and MV. Although there is very limited amino acid sequence similarity between the C, V and W proteins within the paramyxoviruses, the heterotypic inhibition of replication suggests that these proteins may share functional properties.

Induction of intercellular adhesion molecule 1 gene expression by measles virus in human umbilical vein endothelial cells

The expression of intercellular adhesion molecule 1 (ICAM‐1) by endothelial cells is important for the regulation of adhesion and transendothelial migration of a variety of leukocytes that express the integrins lymphocyte function‐associated antigen 1 (LFA‐1) and/or Mac‐1. Here, we demonstrate strain‐specific differences in the ability of measles virus (MV) to induce ICAM‐1 expression. The vaccine strain Moraten (Mor) rapidly induced high levels of ICAM‐1 mRNA and protein expression, whereas the vaccine strain CAM‐70 and the Edmonston wild type (Ed‐wt) strain were far less effective, even when they were used at very high multiplicities of infection (MOIs). Strain‐specific differences in the induction were not a consequence of differences in the ability to infect ECs. Furthermore, induction of ICAM‐1 by Mor was not dependent on de novo expression of MV or cellular proteins. Dual‐immunofluorescence analysis indicated that there was no association between the expression of either MV nucleocapsid or hemagglutinin protein and the induction of ICAM‐1 expression. Some human umbilical vein endothelial cells (HUVECs) that expressed high nucleocapsid protein in response to either Mor or CAM‐70 failed to express elevated ICAM‐1, whereas some HUVECs that were incubated with Mor expressed high ICAM‐1 prior to expression of MV nucleocapsid protein. Strain‐specific differences in the ability of Mor and CAM‐70 to induce ICAM‐1 correlated with their ability to activate the latent transcription factor NF‐κB. These studies suggest a preexisting component of MV particles that leads to strain‐specific differences in the activation of NF‐κB and the induction of ICAM‐1 gene expression. J. Med. Virol. 57:9–16, 1999.