William E. Rawls

Human Papillomavirus Infection and Cervical Cancer in Latin America

To evaluate a possible association between infection with human papillomavirus (HPV) and cervical cancer, we performed a multicenter case-control study in Latin America of 759 cases of invasive cervical cancer and 1467 randomly selected age-matched controls. Demographic, sexual, behavioral, and other clinical data were obtained by interview, and HPV DNA was assayed in cervical-swab specimens with use of filter in situ hybridization. Cervical infection with HPV 16 or 18 or both was strongly associated with cervical cancer. HPV DNA was detected in 62 percent of the cases but only 32 percent of the controls, and the relative risk of cancer increased from 2.1 (95 percent confidence interval, 1.6 to 2.8) to 9.1 (6.1 to 13.6) with hybridization reactions of increasing strength. Although the number of sexual partners, age at first intercourse, number of live births, and Pap-smear history were also significant risk factors, the strong associations between infection with HPV 16 or 18 or both and cervical cancer persisted after we adjusted for these variables. These observations are consistent with the hypothesis that genital infection with HPV 16 or 18 may have a role in the pathogenesis of cervical cancer. Other well-known risk factors were also identified in the study, but they did not affect the association between HPV and cervical cancer.

Sexual Behavior, Venereal Diseases, Hygiene Practices, and Invasive Cervical Cancer in a High-Risk Population

A case‐control study of 759 women with invasive cervical cancer and 1430 controls in four Latin American countries evaluated risk in relation to sexual behavior, histories of specific venereal diseases, and hygiene practices. Early age at first sexual intercourse and increasing number of sexual partners were associated with significantly increased risks even after adjustment for their mutual effects. Risk increased to a twofold excess among women reporting first intercourse at 14 to 15 years of age compared with 20+ years. The number of steady sexual partners was a more important predictor of risk than the number of nonsteady partners, particularly before age 30, possibly reflecting the need for prolonged or repeated exposures to a transmissible agent, or different methods of protection against sexually transmitted diseases or pregnancy. Reported frequency of intercourse was not generally associated with risk, except among women reporting increased frequencies before 20 years of age. Histories of gonorrhea or crab lice were associated with increased risk, but histories of other venereal diseases were not significant predictors. No consistently increased risks were detected for women reporting specific hygiene or douching habits, except the practice of washing the genitalia infrequently during menstruation. These results provide support for a period of increased susceptibility to carcinogens during adolescence, and suggest that this may be an important determinant of the high incidence of cervical cancer in Latin America.

Relation of Herpes simplex viruses to human malignancies.

We have been asked to provide our evaluation of the available data concerning the role of herpes simplex viruses in human malignancies. Perhaps we can begin by stating a set of criteria which, if fulfilled, would satisfy us that a particular virus was the causative agent of a particular malignancy. These requirements will form the framework around which the topic will be developed. Of course, we do not wish to imply that these criteria are original or that the form in which we have stated them is unique. These criteria include the following points: (1) There should be some epidemiologic or biological association or link between the malignancy and the suspected virus; (2) Purified preparations of the virus should induce tumors in its natural host or a similar species, and the resulting tumors should be similar to naturally occurring ones; (3) Ideally, one might hope to find similar naturallyoccurring malignancies in other animals caused by similar viruses; (4) The virus should transform cells in culture, and transformed cells should induce tumors similar to naturally occurring ones, and conversely, tumor cells cultured in vitro should be similar to transformed cells; (5) It should be possible to prevent the malignancy by preventing infection with the suspected virus. Needless to say, the different criteria carry different weight, and fulfillment of the second or fifth criteria stated above would provide very strong evidence for an etiologic role of the virus.

Virion-associated ribosomes are not required for the replication of Pichinde virus.

Abstract Ribosomes of host cell origin have been isolated from Pichinde virus. A cell mutant with a temperature-sensitive defect in protein synthesis and 60 S ribosomal subunits was employed to study the role of virion-associated ribosomes in the replication of Pichinde virus. Virions carrying the temperature-sensitive ribosomes replicated as well at the nonpermissive temperature as at the permissive temperature. The results suggest that the virion-associated ribosomes are not required for the replication of Pichinde virus.

Analyses of the genomes of prototype Pichinde arenavirus and a virulent derivative of Pichinde Munchique: Evidence for sequence conservation at the 3′ termini of their viral RNA species

Abstract Sequence analyses of 3′-[ 32 P]pCp end-labeled large (L) and small (S) viral RNA species of prototype Pichinde (PIC) arenavirus and a virulent derivative of Pichinde Munchique (MUC) virus have shown that the two viral RNA species of each virus have similar 3′-terminal RNA sequences. The 3′-terminal sequence of the first 19 nucleotides of the S RNA of either virus is: 5′ … GCCUAGGAUCCACUGUGCG OH 3′. By comparison, the sequence of the first 19 nucleotides of the L RNA species of either virus is identical, except for a position 6 G residue (from the 3′ end), and a position 8 U residue. The first UAC triplet on S RNA occurs at nucleotide position 84 from the 3′ end; for the L RNA it occurs at nucleotide position 31. Whether these UAC triplets represent the initiation points of translation on viral complementary mRNA species remains to be determined.

Inhibition of Pichinde Virus Replication by Actinomycin D

The yields of Pichinde virus, a member of the arenavirus group, were markedly inhibited when infected BHK 21 cells were incubated in the presence of 0.4 to 4 mug/ml of actinomycin D. Maximal inhibition was observed when actinomycin D was added after the adsorption of virus to cultures; however, addition of drug as late as 12 h after infection reduced the 24 h yield by 50%. Virus antigen synthesis, as measured by complement fixation and immunodiffusion, was not dramatically reduced by actinomycin D. The expression of virus antigens on the surface of infected cells was greater on cells treated with actinomycin D than on untreated cells. Putative defective particles with a density of Pichinde virus were not detected in fluids of cultures incubated with actinomycin D and 3H-amino acids. Actinomycin D appears to inhibit Pichinde virus late in the replicative cycle. The observations raise the possibility that the drug inhibits the synthesis of proteins of the host cell membrane which are required for virus maturation.

Requirement of cell nucleus for the replication of an arenavirus.

Baby hamster kidney (BHK21) cells enucleated with cytochalasin B were infected with Pichinde virus or Sindbis virus. Viral replication was measured by plaque assay, and the synthesis of viral antigens was determined by immunofluorescence. Pichinde virus replication was completely inhibited in cells enucleated prior to infection as measured by both techniques while the replication of Sindbis virus was unaffected. Enucleation of cells at different times after infection with Pichinde virus indicated that intact nuclei were required for at least 8 h after infection.

DESTRUCTION OF VIRUS-INFECTED CELLS BY ANTIBODY AND COMPLEMENT

Publisher Summary This chapter explains the destruction of virus-infected cells by antibody and complement. The interaction of antibody with antigen on the surface of infected cells may produce cytolysis in one of several ways. The immunopathological effects are mediated by various components of the complement system, which is activated by union of IgG or IgM antibodies with antigen on the surface of target cells. Complement activation results in the sequential interaction of nine distinct components designated numerically as C 1 , C 4 , C 2 , C 3 , C 5 , C 6 , C 7 , C 8 , and C 9 , in their order of participation in the sequence. While classical antibody cytolysis requires the sequential action of all nine complement components, there are several cleavage products released in the early sequence of the complement cascade that appear to be significant effector molecules in promoting an inflammatory response. However, the role of complement-dependent antibody-mediated cell lysis in the immunopathology and control of virus infections in vivo is yet to be known. Immunological injury to cells may be influenced by a number of factors, some of which pertain to the nature of expression of antigens at the cell surface. Thus, a virus infection that produces a few antigenic sites on the cell surface might escape immunologic injury as the widely separated antigens would be incapable of doublet formation with IgG antibodies, which is necessary for activation of the complement sequence.

Size Estimation of Pichinde Virus RNA by Gel Electrophoresis under Denaturing Conditions

RNA extracted from purified Pichinde virion preparations yields four major species, two of which appear to represent the viral genome while the other two appear to be cellular ribosomal 28S and 18S RNA. The molecular weights of these RNA species have been estimated by gel electrophoresis following denaturation by either methylmercury or glyoxal/dimethyl sulfoxide. The values determined by the two methods are in close agreement. The average molecular weights of L and S Pichinde viral RNA are 2.63 x 10(6) to 2.83 x 10(6) to 1.31 x 10(6), respectively.

Pseudotypes of vesicular stomatitis virus and Pichinde virus.

Super-infection of Pichinde virus-infected cells with vesicular stomatitis virus (VSV) resulted in the production of pseudotype virus which was not neutralized by antiserum to VSV but which was neutralized by antiserum to Pichinde virus. Analysis of pseudotype virus production in relation to the kinetics of replication of Pichinde virus demonstrated that pseudotype virus production occurred when super-infection with VSV was initiated 8 h or more after infecting the cells with Pichinde virus. The quantities of pseudotype virus produced correlated with the quantities of Pichinde virus antigen detected on the surface of the cells both during acute infection and in cells chronically infected with Pichinde virus. The observations indicate that pseudotype of VSV and Pichinde virus are readily formed and that the formation of pseudotype virus may be used to examine the Pichinde virus antigens expressed on the surface of infected cells.