Isolation and characterization of recombinants between attenuated and virulent aphthovirus strains.

A guanidine-resistant mutant of the attenuated strain of aphthovirus type 01 strain Campos and the original wild-type strain were crossed to generate recombinant viruses. Two independently derived recombinant viruses were isolated. One isolate (RI) contained the P1 (structural proteins) gene region of attenuated strain and P3 (polymerase precursor) gene region of the wild-type strain. The other isolate (RII) had a genomic structure complementary to that of RI, this is, P1 of the wild-type strain and P3 of the attenuated virus. Recombinant RII inherited some in vitro phenotypic markers that were characteristic of the attenuated strain, whereas the RI recombinant had in vitro behavior that was similar to that of the wild-type strain. The data obtained suggest that the polymerase precursor (P3) of the attenuated strain (01 Campos) could be involved in the determination of the attenuated phenotype for fetal bovine kidney cells and, eventually, for cattle.     

Recombination in RNA

The aphthovirus genome consists of a single molecule of single-stranded RNA that encodes all the virus-induced proteins. We isolated recombinant aphthoviruses from cells simultaneously infected with temperature-sensitive mutants of two different subtype strains. Analysis of the proteins induced by 16 independently generated recombinants revealed two types of protein pattern, which were consistent with single genetic crossovers on the 5' side and 3' side, respectively, of the central P34-coding region. Recombinants invariably inherited all four coat proteins from the same parent, and novel recombinant proteins were not observed. RNAase T1 fingerprints of virus RNA, prepared from representatives of each recombinant type, confirmed the approximate crossover sites that had been deduced from the inheritance of proteins. These fingerprints provide molecular evidence of recombination at the level of RNA and demonstrate the potential of RNA recombination for producing genetic diversity among picornaviruses.     

Mapping of attenuating sequences of an avirulent poliovirus type 2 strain.

A mouse model for poliomyelitis was used to identify genomic sequences that attenuate neurovirulence of poliovirus strain P2/P712. This type 2 strain is avirulent in primates and mice yet grows as well as virulent strains in cell culture. The approach used was to exchange portions of the genome of the mouse-virulent P2/Lansing strain with the corresponding region from P2/P712 to identify sequences that could attenuate Lansing neurovirulence in mice. A full-length infectious cDNA of P2/P712 was assembled and used to construct recombinants between P2/P712 and P2/Lansing. The results of neurovirulence testing of 11 recombinants indicated that strong attenuating determinants are located in the 5' noncoding region of P2/P712 and a region encoding capsid protein VP1 and 2Apro, 2B, and part of 2C. An attenuating determinant was further localized to between nucleotides 456 and 628 of P2/P712. A third sequence from P2/P712, nucleotides 752 to 2268, encoding VP4, VP2, and part of VP3, was weakly attenuating. The sequence from nucleotide 4454, approximately halfway through the 2C-coding region, to the end of the P2/P712 genome did not contain attenuating determinants. Nucleotide sequence analysis revealed that P2/P712 differs from the type 2 Sabin vaccine strain by only 22 nucleotides. Six differences lead to amino acid changes in the coding region, and four differences are in the 5' noncoding region. These studies show that, like the type 1 and type 3 Sabin vaccine strains, the attenuated type 2 strain P712 contains multiple attenuating sequences, including strongly attenuating sequences in the 5' noncoding region of the genome.     

Physical mapping of drug resistance mutations defines an active center of the herpes simplex virus DNA polymerase enzyme.

The genome structures of herpes simplex virus type 1 (HSV-1)/HSV-2 intertypic recombinants have been previously determined by restriction endonuclease analysis, and these recombinants and their parental strains have been employed to demonstrate that mutations within the HSV DNA polymerase locus induce an altered HSV DNA polymerase activity, exhibiting resistance to three inhibitors of DNA polymerase. The viral DNA polymerases induced by two recombinants and their parental strains were purified and shown to possess similar molecular weights (142,000 to 144,000) and similar sensitivity to compounds which distinguish viral and cellular DNA polymerases. The HSV DNA polymerases induced by the resistant recombinant and the resistant parental strain were resistant to inhibition by phosphonoacetic acid, acycloguanosine triphosphate, and the 2',3'-dideoxynucleoside triphosphates. The resistant recombinant (R6-34) induced as much acycloguanosine triphosphate as did the sensitive recombinant (R6-26), but viral DNA synthesis in infected cells and the viral DNA polymerase activity were not inhibited. The 2',3'-dideoxynucleoside-triphosphates were effective competitive inhibitors for the HSV DNA polymerase, and the Ki values for the four 2',3'-dideoxynucleoside triphosphates were determined for the four viral DNA polymerases. The polymerases of the resistant recombinant and the resistant parent possessed a much higher Ki for the 2',3'-dideoxynucleoside triphosphates and for phosphonoacetic acid than did the sensitive strains. A 1.3-kilobase-pair region of HSV-1 DNA within the HSV DNA polymerase locus contained mutations which conferred resistance to three DNA polymerase inhibitors. This region of DNA sequences encoded for an amino acid sequence of 42,000 molecular weight and defined an active center of the HSV DNA polymerase enzyme.     

Genetic basis of the neurovirulence of pseudorabies virus.

Lomniczi et al. (J. Virol. 49:970-979, 1984) have shown previously that two attenuated vaccine strains of pseudorabies virus have a similar deletion in the short unique (US) region of the genome. The region which is deleted normally codes for several translationally competent mRNAs. As expected, these mRNAs are not formed in the cells infected with the vaccine strains. The function specified by these mRNAs is thus not necessary for growth in cell culture. Using intracerebral inoculation of 1-day-old chicks as a test system, we have attempted to determine whether a gene within the region that is missing from the attenuated strains specifies functions that are required for the expression of virulence. An analysis of recombinants between the Bartha vaccine strain and a virulent pseudorabies virus strain (having or lacking a thymidine kinase gene [TK+ or TK-]) revealed the following. None of the recombinant plaque isolates that were either TK- or which had a deletion in the US was virulent. Not all recombinant plaque isolates which were both TK+ and had an intact US were virulent. These results indicate that both thymidine kinase activity and an intact US were necessary but not sufficient for the expression of virulence. Marker rescue experiments involving cotransfection of the Bartha strain DNA and a restriction fragment spanning the region of the genome that was missing from the Bartha strain resulted in the isolation of virions to which an intact US had been restored. These virions were not virulent but had an improved ability to replicate in the brains of chicks compared with that of the parental nonrescued Bartha strain. Our results show that genes in the US region, which are missing from the Bartha strain, were necessary for virulence but that this strain was also defective in other genes required for the expression of virulence. Thus, the virulence of pseudorabies virus, as measured by intracerebral inoculation into chicks, appears to be controlled multigenically.     

Identification of two determinants that attenuate vaccine-related type 2 poliovirus.

The poliovirus P2/P712 strain is an attenuated virus that is closely related to the type 2 Sabin vaccine strain. By using a mouse model for poliomyelitis, sequences responsible for attenuation of the P2/P712 strain were previously mapped to the 5' noncoding region of the genome and a central region encoding VP1, 2Apro, 2B, and part of 2C. To identify specific determinants that attenuate the P2/P712 strain, recombinants between this virus and the mouse-adapted P2/Lansing were constructed and their neurovirulence in mice was determined. By using this approach, the attenuation determinant in the central region was mapped to capsid protein VP1. Candidate attenuating sequences in VP1 and the 5' noncoding region were identified by comparing the P2/P712 sequence with that of vaccine-associated isolate P2/P117, and the P2/117 sequences were introduced into the P2/Lansing-P2/P712 recombinants by site-directed mutagenesis. Results of neurovirulence assays in mice indicate that an A at nucleotide 481 in the 5' noncoding region and isoleucine (Ile) at position 143 of capsid protein VP1 are the major determinants of attenuation of P2/P712. These determinants also attenuated neurovirulence in transgenic mice expressing human poliovirus receptors, a new model for poliomyelitis in which virulent viruses are not host restricted. These results demonstrate that A-481 and Ile-143 are general determinants of attenuation.     

Multigenic control of virus virulence in intertypic recombinants of herpes simplex virus

The virulence of herpes simplex virus (HSV) type 1 x type 2 intertypic recombinants was determined following infection of corneas of outbred New Zealand White rabbits. None of the four recombinants was as virulent for rabbits as type 1 parent. All the four recombinants having an insert of type 2 virus genome between 0.35 and 0.576 map units (m.u.) and/or 0.82 and 1.00 m.u. exhibited intermediate virulence between their type 1 and type 2 parents. The results indicate that intertypic recombinants are moderated in their virulence independent of their parental virulence and therefore there exists a multigenic control of HSV virulence.     

Recombinants between herpes simplex virus types 1 and 2: analyses of genome structures and expression of immediate early polypeptides.

Recombinants between temperature-sensitive mutants of herpes simplex virus types 1 (HSV-1) and 2 (HSV-2) were constructed. Using restriction endonucleases, we analyzed the genome composition of 17 intertypic recombinants and detected crossovers in every region of the genome. The virion DNA of one recombinant appeared to be largely "frozen" in two of the four possible genome arrangements of HSV. Knowledge of the genome structures of recombinants enabled us to physically map immediate early polypeptides. We present evidence that the immediate early polypeptide Vmw IE 110 of HSV-1 and its functionally equivalent polypeptide, Vmw IE 118, of HSV-2 may map in the repetitive sequences bounding the long unique region of HSV.     

A group B coxsackievirus/poliovirus 5' nontranslated region chimera can act as an attenuated vaccine strain in mice

The linear, single-stranded enterovirus RNA genome is flanked at either end with a nontranslated region (NTR). By replacing the entire 5' NTR of coxsackievirus B3 (CVB3) with that from type 1 poliovirus, a progeny virus was obtained following transfection of HeLa cells. The chimeric virus, CPV/49, replicates like the parental CVB3 strain in HeLa cells but is attenuated for replication and yield in primary human coronary artery endothelial cell cultures, in a human pancreas tumor cell line, and in primary murine heart fibroblast cultures. Western blotting analyses of CPV/49 replication in murine heart fibroblast cultures demonstrate that synthesis of CPV/49 proteins is significantly slower than that of the parental CVB3 strain. CPV/49 replicates in murine hearts and pancreata, causing no disease in hearts and a minor pancreatic inflammation in some mice that resolves by 28 days postinoculation. A single inoculation with CPV/49 induces protective anti-CVB3 neutralizing antibody titers that completely protect mice from both heart and pancreatic disease when mice are challenged 28 days p.i. with genetically diverse virulent strains of CVB3. That a chimeric CVB3 strain, created from sequences of two virulent viruses, is sufficiently attenuated to act as an avirulent, protective vaccine strain in mice suggests that chimeric genome technology merits further evaluation for the development of new nonpoliovirus enteroviral vectors.     

Recombinants between endogenous and exogenous avian tumor viruses: role of the C region and other portions of the genome in the control of replication and transformation.

Endogenous retroviruses of chickens are closely related to exogenous viruses isolated from spontaneous tumors in the same species, yet differ in a number of important characteristics, including the ability to transform cells in culture, ability to cause sarcomas or leukemias, host range, and growth rate in cell culture. To correlate these differences with specific sequence differences between the two viral genomes, the genome RNA of transforming subgroup E recombinants between the Prague strain of Rous sarcoma virus, subgroup B (Pr-RSV-B), and the endogenous Rous-associated virus-0 (RAV-0), Subgroup E, and seven nontransforming subgroup E recombinants between the transformation-defective mutant of Pr-RSV-B and RAV-0 was examined by oligonucleotide fingerprinting. The pattern of inheritance among the recombinant viruses of regions of the genome in which Pr-RSV-B and RAV-0 differ allowed us to draw the following conclusions. (i) Nonselected parts of the genome were, with a few exceptions, inherited by the recombinant virus progeny randomly from either parent, with no obvious linkage between neighboring sequences. (ii) A small region in the Pr-RSV-B genome which maps in the 5' region was found in all transforming but only some of the nontransforming recombinants, suggesting that it plays a role in the control of the expression of transformation. (iii) A region of the Pr-RSV-B genome which maps between env and src was similarly linked to the src gene and may be either part of the structural gene for src or a control sequence regulating the expression of src. (iv) The C region at the extreme 3' end of the virus genome which is closely related in all the exogenous avian retroviruses but distinctly different in the endogenous viruses is the major determinant responsible for the differences in growth rate between RAV-0 and Pr-RSV-B. This latter observation allowed us to redefine the C region as a genetic locus, c, with two alleles cn (in RAV-0) and cx (in exogenous viruses).     

Natural genetic exchanges between vaccine and wild poliovirus strains in humans

In a previous study of poliovirus vaccine-derived strains isolated from patients with vaccine-associated paralytic poliomyelitis (VAPP) (9, 11), we reported that a high proportion (over 50%) of viruses had a recombinant genome. Most were intertypic vaccine/vaccine recombinants. However, some had restriction fragment length polymorphism (RFLP) profiles different from those of poliovirus vaccine strains. We demonstrate here that five such recombinants, of 88 VAPP strains examined, carried sequences of wild (nonvaccine) origin. To identify the parental wild donor of these sequences, we used RFLP profiles and nucleotide sequencing to look for similarity in the 3D polymerase-coding region of 61 wild, cocirculating poliovirus isolates (43 type 1, 16 type 2, and 2 type 3 isolates). In only one case was the donor identified, and it was a wild type 1 poliovirus. For the other four vaccine/wild recombinants, the wild parent could not be identified. The possibility that the wild sequences were of a non-poliovirus-enterovirus origin could not be excluded. Another vaccine/wild recombinant, isolated in Belarus from a VAPP case, indicated that the poliovirus vaccine/wild recombination is not an isolated phenomenon. We also found wild polioviruses (2 of 15) carrying vaccine-derived sequences in the 3' moiety of their genome. All these results suggest that genetic exchanges with wild poliovirus and perhaps with nonpoliovirus enteroviruses, are also a natural means of evolution for poliovirus vaccine strains.     

Isolation of intertypic recombinants of Epstein-Barr virus from T-cell-immunocompromised individuals.

All wild-type isolates of Epstein-Barr virus (EBV) analyzed to date for allelic polymorphisms of the nuclear antigen EBNA2 gene (in the BamHI YH region of the genome) and of the EBNA3A,-3B, -3C genes (tandemly arranged in the BamHI E region) have proved either uniformly type 1 or uniformly type 2 at all four loci. The absence of detectable intertypic recombination in the wild probably reflects the rarity with which individual carriers, and certainly individual target cells, become coinfected with both virus types. Studying a group of human immunodeficiency virus-positive T-cell-immunocompromised patients known to be at enhanced risk of multiple EBV infections, we have isolated intertypic EBV recombinants from 2 of 40 patients analyzed. These recombinants, whose in vitro transforming capacity appeared at least equal to that of type 1 strains, carried a type 1 EBNA2 allele and type 2 EBNA3A,-3B, and -3C alleles. This was clearly demonstrable at the DNA level by PCR amplification using type-specific primer-probe combinations and was confirmed at the protein level (for EBNA2 and EBNA3C) by immunoblotting with type-specific antibodies. In one patient, the recombinant appeared to be the predominant strain, being the virus most commonly rescued by in vitro transformation both from the blood and from the throat washings on two separate occasions 20 months apart. A regular type 1 virus strain was also present in this individual, but this was not related to the recombinant since the two viruses carried type 1 EBNA2 genes with different patterns of variance from the B95.8 prototype sequence. In the other patient, recombinants were isolated on one occasion from the blood and on a separate occasion, 21 months later, from the throat; these recombinants were almost certainly related, being identical at several genomic polymorphisms and differing only in one facet of the "EBNAprint," the size of the EBNA1 protein. Three different type 1 viruses were also isolated from this patient, two of which carried EBNA2 genes with the same pattern of sequence variation from B95.8 as the recombinant; however, since this is a fairly common pattern of variance, the relationship of these viruses to the recombinant remains an open question. We infer that intertypic recombinants of EBV are not uncommon in HIV-positive T-cell-immunocompromised patients, that they arise in such individuals as a consequence of their increased frequency of mixed-type infections, and that they will prove capable of efficient transmission in the human population.     

Complete genome sequence of a recombinant coxsackievirus b4 from a patient with a fatal case of hand, foot, and mouth disease in guangxi, china

The coxsackievirus B4 (CVB4) belongs to human enterovirus B species within the family Picornaviridae. Here we report a novel complete genome sequence of a recombinant CVB4 strain, CVB4/GX/10, which was isolated from a patient with a fatal case of hand, foot, and mouth disease in China. The complete genome consists of 7,293 nucleotides, excluding the 3' poly(A) tail, and has an open reading frame that maps between nucleotide positions 742 and 7293 and encodes a 2,183-amino-acid polyprotein. Phylogenetic analysis based on different genome regions reveals that CVB4/GX/10 is closest to a CVB4 strain, EPIHFMD-CLOSE CONTACT-16, in the 5' half (VP4～2B) of the genome, although it is closer to a Chinese CVB5 strain, CVB5/Henan/2010, in the 3' half (2C～3D) of the genome. Furthermore, similar bootscan analysis based on the whole genomes demonstrates that recombination has possibly occurred within the 2C domain and that CVB4/GX/10 is a possible progeny of intertypic recombination of the CVB4 strain EPIHFMD-CLOSE CONTACT-16 and CVB5/Henan/2010 that occurred during their cocirculation and evolution, which is a relatively common phenomenon in enteroviruses.     

Differences in replication of attenuated and neurovirulent polioviruses in human neuroblastoma cell line SH-SY5Y.

A base change from C to U at position 472 of the 5' noncoding region of the poliovirus genome is known to be a major determinant of attenuation in the P3/Sabin vaccine strain. To determine the biochemical basis for the attenuated phenotype imparted by this mutation, a cell line in which replication of neurovirulent and attenuated viruses could be distinguished was identified. A pair of P3/Sabin-P2/Lansing viral recombinants that differ only at position 472 was used; the viruses replicated equally well in HeLa cells, but the virus with a U at base 472 was attenuated in mice. In the human neuroblastoma cell line SH-SY5Y, recombinants with a U at base 472 replicated to approximately 10-fold-lower titers than did neurovirulent viruses with a C at this position. Analysis of viral RNA and protein synthesis indicated that translation of the attenuated viral RNA was specifically reduced in SH-SY5Y cells.     

A recombinant virus between the Sabin 1 and Sabin 3 vaccine strains of poliovirus as a possible candidate for a new type 3 poliovirus live vaccine strain.

Biological tests including the monkey neurovirulence test performed on recombinants between the virulent Mahoney and attenuated Sabin 1 strains of type 1 poliovirus indicated that the genome region encoding mainly the viral capsid proteins had little correlation with the neurovirulence or attenuation phenotype of the virus. The results suggested that new vaccine strains of type 2 and type 3 polioviruses may be constructed in vitro by replacing the sequence encoding the antigenic determinants in viral capsid proteins of the Sabin 1 genome by the corresponding sequences of the type 2 and type 3 genome, respectively. Accordingly, we constructed recombinants between the Sabin 1 and Sabin 3 strains of poliovirus in which genome sequences of the Sabin 1 strain encoding most or all capsid proteins were replaced by the corresponding genome sequences of the Sabin 3 strain. One of the recombinant viruses thus constructed was fully viable and showed antigenicity and immunogenicity identical to those of type 3 poliovirus. The monkey neurovirulence tests and in vitro phenotypic marker tests (temperature sensitivity of growth, sodium bicarbonate concentration dependency of growth under agar overlay, and size of plaque) were performed on the recombinant virus. The stability of the virus in regard to the temperature sensitivity phenotype was also tested. The results suggested that the recombinant virus is a possible candidate for a new type 3 poliovirus vaccine strain.     

Mapping of sequences required for mouse neurovirulence of poliovirus type 2 Lansing.

Intracerebral inoculation of mice with poliovirus type 2 Lansing induces a fatal paralysis, while most other poliovirus strains are unable to cause disease in the mouse. To determine the molecular basis for Lansing virus neurovirulence, we determined the complete nucleotide sequence of the Lansing viral genome from cloned cDNA. The deduced amino acid sequence was compared with that of two mouse-avirulent strains. There are 83 amino acid differences between the Lansing and Sabin type 2 strain and 179 differences between the Lansing and Mahoney type 1 strain scattered throughout the genome. To further localize Lansing sequences important for mouse neurovirulence, four intertypic recombinants were isolated by exchanging DNA restriction fragments between the Lansing 2 and Mahoney 1 infectious poliovirus cDNA clones. Plasmids were transfected into HeLa cells, and infectious recombinant viruses were recovered. All four recombinant viruses, which contained the Lansing capsid region and different amounts of the Mahoney genome, were neurovirulent for 18- to 21-day-old Swiss-Webster mice by the intracerebral route. The genome of neurovirulent recombinant PRV5.1 contained only nucleotides 631 to 3413 from Lansing, encoding primarily the viral capsid proteins. Therefore, the ability of Lansing virus to cause paralysis in mice is due to the viral capsid. The Lansing capsid sequence differs from that of the mouse avirulent Sabin 2 strain at 32 of 879 amino acid positions: 1 in VP4, 5 in VP2, 4 in VP3, and 22 in VP1.     

In vitro binding of cattle PstI SINE with a 33-kDa nuclear protein.

A PstI family of SINEs (short interspersed elements) has been identified in some of the members of the family Bovidae, for example, cattle, buffalo and goat. In vitro DNA-protein interactions were studied to provide a better understanding of the function of these SINEs in the genome. Use of one such cattle PstI interspersed repeat sequence, as a probe in gel retardation assays, has lead to the identification of a repeat DNA-binding factor PIRBP (PstI interspersed repeat binding protein) from cattle liver nuclear extract. Southwestern analysis with liver nuclear extracts from cattle, goat, and buffalo revealed the presence of a PIRBP-like nuclear factor in all three species belonging to the family Bovidae. Deletion analysis localized the PIRBP binding site to an 80-bp (337-417 bp) region within the cattle PstI sequence. UV crosslinking and Southwestern analyses clearly indicated that PIRBP is a singular, small polypeptide of 33-kDa molecular mass. Homology search of the nucleic acids database revealed that the cattle PstI sequence was associated with many different genes of the family Bovidae, either in the 5' flanking region, 5' locus activating region, 3' UTR or in intervening sequences. The binding of the cattle PstI SINE by PIRBP and its association with the regulatory regions of the genes suggests that it plays an important role in the bovine genome.     

Recovery of virulent and RNase-negative attenuated type 2 bovine viral diarrhea viruses from infectious cDNA clones

Cloned cDNA derived from the genome of the virulent type 2 bovine viral diarrhea virus (BVDV) strain NY'93/C was sequenced and served for establishment of the infectious cDNA clone pKANE40A. Virus recovered from pKANE40A exhibited growth characteristics similar to those of wild-type BVDV NY'93/C and proved to be clinically indistinguishable from the wild-type virus in animal experiments. A virus mutant in which the RNase residing in the viral glycoprotein E(rns) was inactivated, revealed an attenuated phenotype. The plasmid pKANE40A represents the first infectious cDNA clone established for a type 2 BVDV and offers a variety of new approaches to analyze the mechanisms of BVDV-induced disease in cattle.     

Complete genome sequences of a Korean virulent porcine epidemic diarrhea virus and its attenuated counterpart

A virulent porcine epidemic diarrhea virus (PEDV) strain, DR13, was obtained from suckling pigs suspected of having porcine epidemic diarrhea in 1999 in Korea, and its attenuated counterpart was derived from virulent strain DR13 by serial propagation in Vero cells. This report describes the first complete genome sequences of virulent PEDV and its attenuated counterpart, which will provide important insights into the molecular basis of the attenuation of PEDV.     

RNA recombination of murine coronaviruses: recombination between fusion-positive mouse hepatitis virus A59 and fusion-negative mouse hepatitis virus 2.

It has previously been shown that the murine coronavirus mouse hepatitis virus (MHV) undergoes RNA recombination at a relatively high frequency in both tissue culture and infected animals. Thus far, all of the recombination sites had been localized at the 5' half of the RNA genome. We have now performed a cross between MHV-2, a fusion-negative murine coronavirus, and a temperature-sensitive mutant of the A59 strain of MHV, which is fusion positive at the permissive temperature. By selecting fusion-positive viruses at the nonpermissive temperature, we isolated several recombinants containing multiple crossovers in a single genome. Some of the recombinants became fusion negative during the plaque purification. The fusion ability of the recombinants parallels the presence or absence of the A59 genomic sequences encoding peplomers. Several of the recombinants have crossovers within 3' end genes which encode viral structural proteins, N and E1. These recombination sites were not specifically selected with the selection markers used. This finding, together with results of previous recombination studies, indicates that RNA recombination can occur almost anywhere from the 5' end to the 3' end along the entire genome. The data also show that the replacement of A59 genetic sequences at the 5' end of gene C, which encodes the peplomer protein, with the fusion-negative MHV-2 sequences do not affect the fusion ability of the recombinant viruses. Thus, the crucial determinant for the fusion-inducing capability appears to reside in the more carboxyl portion of the peplomer protein.