Indiana University-Purdue University Indianapolis (IUPUI) / This study reports the results of the clinical examination of fifty 2 1/2 year old children with congenital defects attributed to infection with prenatal rubella. The results were compared with the results of the clinical examination of fifty normal 2 to 3 year old children. The children with congenital defects attributed to in utero rubella infection had a significantly lower (p<.005) weight at birth and at 2 1/2 years of age. These findings, along with the type and distribution of congenital defects, are in agreement with other reports of children with confirmed rubella embryopathy. The fifty children with rubella embryopathy had a significantly (p<.005) higher incidence of dental defects. Of the rubella children studied, 90 per cent had enamel hypoplasia, 78 per cent had tapered teeth, and 18 per cent had notched anterior teeth. Among the normal children studied, 26 per cent had enamel hypoplasia, 18 per cent had tapered teeth, and none had notched teeth. No other dentofacial abnormalities other than a cleft of the lip and palate in one rubella child were found.

As a model for studying apoptosis associated with pathogenesis of congenital rubella syndrome, bicistronic rubella virus (RUBV) replicons expressing an antibiotic resistance gene in the presence (925-IN) or absence (IN-IN) of RUBV capsid protein (C) were constructed. Apoptosis was assessed by detection of caspase activation, chromatin fragmentation, and flow cytometry. 925-IN cells grew similarly to Vero, but IN-IN cells demonstrated caspase activation, chromatin fragmentation and cell cycle arrest. Whereas Vero cells transfected with P150 exhibited rapid apoptosis not detected in transfected Vero cells stably expressing C, neither exhibited cell cycle alterations, indicating a cell cycle stall not associated with apoptosis. Finally, two human epithelial cells, HEK293 and A549, transfected with P150 failed to exhibit apoptosis, indicating that while replicon-transfected Vero cells are useful for studying apoptosis and cell cycle arrest, the results are not applicable to other cell types.

THESIS Presented to THE FACULTY OF MEDICINE, University of the Witwatergrand. In Partial Fulfillment of the requirements for the Degree DOCTOR OF MEDICINE. / Every morbid condition, occurring in Medicine is based fundamentally on the genetic constitution of the individual, but only diseases whloh are significantly related to hereditary factors are regarded as hereditary diseases. In the thesis presented by the writer, an attempt is made to show how the environment affects the development and expression of factors genetical in origin. The first part of this work is really only a sketchy introduction to the genetic processes involved, and is presented as a background on which to paint the picture of certain conditions regarded as slgnifioantally environmental in origin,. Hence the early chapters cannot be regarded as being vety complete or detailed, but an attempt has been made to present the basic genetic laws, and to Interpret various morbid processes in the foetus and newborn In the light of these laws. Some of these pathological conditions have only recently been understood, and a few are enumerated In the hope of producing an over-all picture. Some are touched upon as being of some practical importance either . at the present date, or possibly in the near future, This brings us to the conception of "prenatal paediatrics." There is a tendency at the present time for a closer relationship to develop between obstetrician and paediatrician Just because of the newer knowledge regarding the reciprocal relationship between maternal and foetal status. Maternal nutri- Importance* Obstetricians, today, in introducing newer and safer procedures, operations, and analgaesic methods for delivering their patients, consider more and more, the management of labour in terms of the effect upon the foetus. Psychological, as well as physiological care of the pregnant woman must be instituted beoause psychological disturbances may significantly affect the actual physiology of pregnancy and parturition. Paediatric attention, initiated early in pregnancy, is therefore not necessarily an impractical procedure. As the genetlo constitution cannot be Varied the aim of “prenatal paediatrics” must be directed towards modification of adverse environmental factors. One of the great aims of Eugenics is to prevent the action of adverse genetio factors, but the scope of eugenics at.the present time Is limited in its application. There appears to be a tendency, as the centuries roll on, for balanced genetical systems to evolve, whioh seem to be resistant to change. Whether the future use of atomic radiation, either in Peace or in War, may change this, remains to be seen. The effects of maternal rubella, as described in the second part of this work, appear to be so diverse in different instances, that the operation of co-existent genetical background factors oannot entirely be excluded. It is essential to obtain better statistical Information regarding gene frequencies. Biochemistry, physiology, and Serology should be allied to Clinical and Social Medicine In an attempt to discover v i i i . and diabetes, and also various types of defedtfe fend malformations, such as congenital morbus cordis * deaf-mutism, mental defioienoy, eto. Art Investigation of the relationship between neonatal stattie fend the development of subsequent disease patterns during later childhood, or even adulthood, will undoubtedly » prove to be of the greatest import. In this work reference has been made to the value of epidemiological Btudies as a means of uncovering mechanisms which produce congenital defects. It should be remembered that, due to differences in diet, medical care, and a thousand other factors of changed environment, maternal status today is different to what it was fifty or one hundred years ago, and any effects for good or ill on the foetal population can. be unearthed by careful, prolonged statistical study. It is obvious that the correlated efforts of many branches of Ollnlcal, Experimental and Social Medicine over long.periods, are essential to produoe useful results. For lnstanoe, mongolism occurring in two slbs may not necessarily represent an hereditary condition, as one of the cases may be incidental, or dependant on environmental factors obtaining for both affected sibs. Similarly, many of the less typloal congenital defects, which in late years have been described as appearing in children born after maternal rubella may be incidental, and either represent the risk to the foetal population at large, or may yet be the means of uncovering the operation of other environmental factors as well. Hence we see the need for a oloser follow up of all morbid or abnormal conditions during pregnancy, particularly/*«,. particularly virus infections, with complete reports on the infants subsequently born, whether normal or otherwise. This need becomes clear, when the latter part of this work is consulted. The investigations carried out on maternal rubella as an aetiological factor in the production of congenital defects is presented as a most important step in the understanding and prevention of unfortunate accidents of foetal development. The present thesis is essentially a review of the literature, but several oases are presented on account.of the paucity of reports in the xrorld literature. In this connection, the writer would like to acknowledge case histories given by Doctors D. G. Melle, 0.N..Javett, M. Chitters, W, Tope, M, Epstein and C-. Faerber all of Johannesburg, and Doctors B. Epstein, P. Oosterhagen and J. Rudolph of Pretoria. Valuable assistance and great courtesy has been offered by the Matron, St. Vinoentfs School for the Deaf, Johannesburg, Mr. Wentworth of.the National Council for the Blind, Pretoria, Dr. le Riche of the Union Health Department, Pretoria, Ma.^or Dreisenstock at Defence Headquarters, Pretoria and those most efficient librarians, Miss A.C. Dick.and Miss Krige of the Witwatersrand Medical Library. My thanks are also due to the doctors at Union Health Department, Pretoria who so graciously plaoed their excellent library material at my disposal* Last but not least, I am indebted to Mrs, T. Mellet, Miss E. Marcus and my wife for clerical assistance. X. E„ F. Pretoria, January, 1951. / IT2017

Objective: To evaluate the impact of rubella vaccination strategies on rubella and CRS incidence and to offer public health suggestions for vaccination strategy in China. Methods: A systematic research of studies was conducted via the United States National Library of Medicine and the National Institutes of Health Medical Database (PubMed) and Google Scholar database. Terms- rubella AND (vaccin* AND coverage AND (reproduct* OR transmiss*) were used as key words in the research via PubMed. “Rubella vaccination strategies” and “rubella and congenital rubella syndrome” and “incidence” were used to search via Google Scholar. A systematic review of literature was conducted. The effectiveness of rubella vaccination strategies on the control and elimination of rubella and CRS incidence was used as primary inclusion criteria. Results: 5 articles were selected according to inclusion criteria. They were two cohort studies, two ecological studies, one mathematical modeling study. From the experience in Canada, around 99% decline of rubella and CRS incidence happened after vaccination strategy with selective approach for children. However, it led to a shift of infection to susceptible groups, and couldn’t interrupt rubella virus circulation. A combined vaccination with adults could control CRS incidence but was not effective for CRS elimination. A vaccination strategy with universal approach for children, males and females in Brazil could prevent child-bearing women from re-infection, thus helped to eliminate CRS cases. Conclusion: Vaccination strategy with universal approach was most effective for CRS control in a short time period with the evidence of a huge reduction of rubella and CRS incidence. As part of national immunization program in China, routine vaccination strategy for rubella should consider both epidemic and demographic factors to assess the effectiveness of children vaccination strategy. / published_or_final_version / Public Health / Master / Master of Public Health

No description available.

Complications of rubella virus infection, including congenital rubella syndrome and the association of rubella virus with joint inflammation, emphasize the need for continued research on rubella virus. The finding that the association of rubella virus infection with joint manifestations is more pronounced with wild strains than with vaccine strains suggested the possibility of strain variation. Several different techniques have been employed in order to compare six rubella virus strains and identify variations in their structural proteins. Differences in biological activities were detected, including the extent of virus production and the ability of various cell types to support replication of rubella virus (tissue tropism). However, the strains were shown to have remarkably similar electrophoretic patterns. Variation appeared to result from alterations in glycosylation. Efforts to isolate the protein components of the two envelope glycoproteins were unsuccessful, and it was therefore not possible to localize variation to either the protein or the carbohydrate components. Future work employing more sensitive methods for examination of fine molecular structure and the correlation of these structures with biological activity will further our understanding of the pathogenesis of rubella virus infection. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

Rubella virus is a small enveloped positive strand RNA virus. Two species of viral RNA are found in infected cells: a full-length genomic RNA and a subgenomic species corresponding to the 3' one third of the genomic RNA molecule. The 24S subgenomic RNA specifies a polyprotein which is cotranslationally processed by endoproteolytic cleavage by host signal peptidase to yield three structural proteins, El, E2 and capsid. El and E2 are membrane glycoproteins forming the virion spikes, and C protein binds to 40S genomic RNA to form a nucleocapsid. El and E2 proteins contain N-linked oligosaccharide as a consequence of their passage through the endoplasmic reticulum (ER) and Golgi apparatus. According to the signal hypothesis, translocation of secretory and membrane proteins into the ER is mediated by a hydrophobic signal peptide. The signal peptides for E2 and El have been localized by in vivo expression of El and E2 cDNAs. Oligonucleotide-directed mutagenesis was used to define the cleavage sites between C, E2, and El, as well as the effect of the cleavages on the transport and processing of E2 and El. The expression of the cleavage site mutants was studied in vitro and in vivo. It was found that uncleaved precursor polypeptides were retained in the ER and very little E2 or El polypeptide was observed at either the Golgi apparatus or the plasma membrane. The E2 and El polypeptides can cross the ER membrane without the cleavage of the signal peptide while the transport of E2 and El beyond the ER requires the cleavage of E2 from C and El from E2. The C-termini of the C and E2 proteins, which were not previously defined, have been partially characterized. Capsid protein does not appear to undergo further proteolytic processing after it is cleaved from E2 by signal peptidase, but E2 may be processed at a second cleavage site at its C-terminus by a trypsin-like enzyme. / Medicine, Faculty of / Medical Genetics, Department of / Graduate

As a model for studying apoptosis associated with pathogenesis of congenital rubella syndrome, bicistronic rubella virus (RUBV) replicons expressing an antibiotic resistance gene in the presence (925-IN) or absence (IN-IN) of RUBV capsid protein (C) were constructed. Apoptosis was assessed by detection of caspase activation, chromatin fragmentation, and flow cytometry. 925-IN cells grew similarly to Vero, but IN-IN cells demonstrated caspase activation, chromatin fragmentation and cell cycle arrest. Whereas Vero cells transfected with P150 exhibited rapid apoptosis not detected in transfected Vero cells stably expressing C, neither exhibited cell cycle alterations, indicating a cell cycle stall not associated with apoptosis. Finally, two human epithelial cells, HEK293 and A549, transfected with P150 failed to exhibit apoptosis, indicating that while replicon-transfected Vero cells are useful for studying apoptosis and cell cycle arrest, the results are not applicable to other cell types.

>Magister Scientiae - MSc / Despite widespread rubella virus (RV) vaccination programs, annually RV still causes severe congenital defects in an estimated 100,000 children globally. A concerted attempt to eradicate RV is currently underway and analytical tools to monitor the global decline of the last remaining RV lineages will be useful for assessing the effectiveness of this endeavour. Importantly, RV evolves rapidly enough that much of its epidemiological information might be inferable from RV genomic sequence data. Using BEASTv1.8.0, I analysed publically available RV sequence data to estimate genome-wide and gene-specific nucleotide substitution rates, to test whether the current estimates of RV substitution rates are representative of the entire RV genome. During these investigations, I specifically accounted for possible confounders of nucleotide substitution rate estimates, such as temporally biased sampling, sporadic recombination, and natural selection favouring either increased or decreased genetic diversity (estimated by the PARRIS and FUBAR methods) at nucleotide sites within RV nucleic acid secondary structures (predicted by the NASP method). I determined that RV nucleotide substitution rates range from 1.19×10-3 substitutions/site/year (in the E1 region) to 7.52×10-4 substitutions/site/year (in the P150 region). I found that these differences between nucleotide substitution rate estimates in various RV gene regions are largely attributable to temporal sampling biases, such that datasets containing a higher proportion of recently sampled sequences will tend to have inflated estimates of mean substitution rates. Although there exists little evidence of positive selection or natural genetic recombination in RV, I revealed that RV genomes possess extensive biologically functional nucleic acid secondary structures and that purifying selection acting to maintain these structures contributes substantially to variations in estimated nucleotide substitution rates across RV genomes. Although both temporal sampling biases and purifying selection favouring the conservation of RV nucleic acid secondary structures have an appreciable impact on substitution rate estimates, I find that these biases do not preclude the use of RV sequence data to date ancestral sequences and evaluate the associated RV phylodynamics. The combination of uniformly high substitution rates across the RV genome and strong temporal signal within the available sequence data enabled me to analyse the epidemiological and demographical dynamics of this virus during these attempts to eradicate it. By implementing a generalized linear model (GLM) and symmetrical model of discretized phylogeographic spread, I was able to identify several predictive variables of geographical RV spread and detect transmission linkages between distinct geographical regions. These results suggest that, in addition to strengthened vaccination strategies, there also needs to be an increased effort to educate people about the effects of vaccination and risks of RV infection.

Rubella virus (RV) is a small, enveloped, positive-stranded RNA virus in the family Togaviridae, and bears striking similarities to the prototype alphaviruses Semliki Forest virus (SFV) and Sindbis virus (SV) in terms of genome organization and structural protein expression strategy. However unlike alphaviruses, RV infection of cultured cells is characterized by relatively long latency periods, slow replication kinetics, limited cytopathology, and the ability to establish a persistent infection in virtually every cell line capable of supporting its growth. RV virions contain three structural proteins C, E2, and El which are derived by post-translational processing of a precursor polyprotein pllO (NF₂-C-E2-El-COOH). Processing and intracellular transport of RV structural proteins has been studied by jn vitro and jn vivo expression of RV cDNAs. It was found that targeting of El and E2 into the endoplasmic reticulum was mediated by two independently functioning signal peptides. Coincident with translocation into the ER, both proteins underwent addition of N-linked glycans and proteolytic processing. C protein did not appear to play a role in the processing of pllO. Expression of the RV structural proteins in COS cells revealed that E2 exited the ER, and was transported through the Golgi to the cell surface in an El-independent manner, although coexpression of El seemed to increase the rate of transport. Conversely, El was retained in a Golgi-like region and was not found on the plasma membrane in the absence of E2. Oligonucleotide-directed mutagenesis of El and E2 cDNAs showed that El andE2 both contain three N-linked glycans respectively. Lack of glycosylation did not appear to affect the intracellular localization of the RV glycoproteins in COS cells. A number of significant differences between RV and SFV/SV structural protein expression strategies were discovered, and their possible relationship to RV virion assembly are discussed. / Medicine, Faculty of / Medical Genetics, Department of / Graduate