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PubMed1995

Interspecific reassortment of genomic segments in the evolution of cucumoviruses.

Segmented genomes of RNA viruses are thought to evolve and be maintained in analogy to sexual recombination and reassortment in eukaryotic systems. If reassortment among genomes is an important event in cucumoviral evolution, then such events should be detectable among extant viruses. In this study, phylogenetic analyses of cucumoviruses were performed using aligned amino acid sequences. The results reveal different relationships among species when the three genomic segments are compared, suggesting that reassortment events have given rise to extant forms. In addition, we describe a cucumovirus isolate that is composed of genomic segments from two distinct viral species. These results indicate that reassortment events may provide a mechanism for speciation in cucumoviruses.

Amino Acid Sequence↗

PubMed1998

Tomato spotted wilt Tospovirus genome reassortment and genome segment-specific adaptation.

A system to associate specific genome segments with viral phenotypes and to study factors influencing genome reassortment was developed for tomato spotted wilt Tospovirus (TSWV). Reassortant isolates were generated by co-inoculating a TSWV isolate, TSWV-D, with TSWV-10 or TSWV-MD. The parental origin of each genome segment in putative reassortant isolates was determined by segment-specific restriction fragment length polymorphisms. The TSWV isolates readily exchanged genome segments in a nonrandom fashion. The S RNA from TSWV-D was dominant over the S RNA from TSWV-10. The intergenic region (IGR) of the S RNA was correlated with competitiveness of this genome segment in reassortant isolates. The less competitive S RNA contained a net increase of 62 nt, including a 33-nt duplication in the IGR. This duplicate sequence was highly conserved among isolates from the southeastern United States and an isolate from Bulgaria. The evidence supports the hypothesis that the IGR of the S RNA with an ambisense coding strategy serves a regulatory function which influences the occurrence of this segment in the viral population. In addition, it was demonstrated that stable parental phenotypes can be mapped to specific genome segments as well as generating novel phenotypes not associated with either parent.

Adaptation, Biological↗

PubMed2001

Reassortant analysis of guinea pig virulence of pichinde virus variants.

The new world arenavirus Pichinde (PIC) is the basis of an accepted small animal model for human Lassa fever. PIC (Munchique strain) variant P2 is attenuated in guinea pigs, whereas variant P18 is extremely virulent. Previous sequence analysis of the S segments of these two viruses indicated a small number of possible virulence markers in the glycoprotein precursor (GPC) and nucleoprotein (NP) genes. In order to determine the role of these S segment genes in guinea pig virulence in this system, we have generated reassortant viruses. When tested in outbred guinea pigs, the reassortant containing the S segment from the virulent parent P18 (S18L2) caused significantly higher morbidity than the reciprocal reassortant. This increased morbidity was associated with higher viral titers in serum and spleen. However, the S18L2 reassortant was not as fully virulent in this system as the P18 parent, indicating a role for L segment genes in virulence.

Animals↗

PubMed1996

WC3 reassortant vaccines in children.

Bovine rotavirus strain WC3 (P7[5], G6) administered at the 12th passage level was well tolerated clinically in infants and efficiently induced serum virus neutralizing antibody (VNA) with bovine rotavirus G6 specificity. The protective efficacy of WC3 vaccine against all rotavirus disease was inconsistent, varying in four separate trials from 76% to 0%; some selective protection against severe disease was seen in all trials. WC3 reassortants containing the gene for an individual human rotavirus VP7 (G) or VP4 (P) surface antigen were also well tolerated, but preferentially induced VNA to the WC3 parent. Efficacy trials of human G1 VP7 reassortant WI79-9 (P7[5], G1) consistently led to > 60% protection against all rotavirus disease. A quadrivalent WC3 reassortant vaccine was developed to contain four separate monovalent reassortants expressing human rotaviruses surface proteins G1, G2, G3, and P1A [8] respectively. In a multicenter trial including 439 infants, this vaccine induced 67.1% protection against all rotavirus disease (defined as positive for rotavirus antigen by ELISA only [p = < 0.001]) and 72.6% protection when the standard for rotavirus diagnosis was a positive test of stool for both rotavirus antigen by ELISA and rotavirus RNA by electropherotype analysis (p = < 0.001). In this trial, episodes of the most severe rotavirus disease (clinical severity score > 16.0 eight cases) occurred only in placebo recipients.

Animals↗

PubMed2000

Cross-protection and reassortment studies with avian H2 influenza viruses.

In order to assess the degree of immune cross-protection among avian H2 influenza virus strains, mice were immunised with beta-propiolactone-inactivated virus preparations and infected intranasally with mouse-adapted variant of A/Black Duck/New Jersey/1580/78 (H2N3) strain. The experiments with 11 avian H2 strains revealed that both Eurasian and American H2 avian influenza viruses exhibit either high or moderate degree of cross-protection. The grouping of the strains in accordance with their cross-protection efficiency does not coincide with H2 phylogenetic branches. Several reassortant clones were obtained with the use of A/Pintail Duck/Primorie/695/76 (H2N3) strain and high-yield X-67 reassortant as parent viruses, among them a high-yield H2N3 reassortant. Taking into account the data on cross-protection among avian H2 strains, the high-yield H2N3 reassortant may be regarded as a prototype strain to be used for the preparation of killed vaccines in the case of a new appearance of avian H2 haemagglutinin in circulation in humans.

Animals↗

PubMedDate not supplied

Non-random selection of gene segment 3 and random selection of gene segment 5 observed in reassortants generated in vitro between rotavirus SA11 and RRV.

A total of 154 clones of reassortants between rotavirus RRV and SA11 were isolated in vitro in order to compare the selection modes of each RNA segment in reassortant formation with those in the in vivo experiment (carried out by Gombold and Ramig) using the same parent strains in which the occurrence of non-random selection of gene segments 3 and 5 was observed. In reassortants isolated from a cross RRV x SA11-L2 (66 isolates) and a cross RRV x SA11-S1 (88 isolates), 36 and 57 different genotypes were identified, respectively. Segregation rates of SA11 RNA segments 1, 5, 8, 9 and 11 were significantly different as compared with those previously reported in the in vivo experiment. While RNA segment 3 was preferentially selected from an SA11 parent, the segregation rate of SA11 gene 5 was relatively low in vitro. Thus, the selection mode of rotavirus RNA segments was found to be different between in vivo and in vitro reassortments.

Animals↗

PubMed1997

Use of oligonucleotide probes for selecting potential high-yielding influenza reassortants.

A new method for rapid screening of high-yielding reassortants of influenza virus, as candidates for vaccine production, is described. Oligonucleotide probes specific for all the parent genes of A/PR/8/34 (PR8), except the HA and the NA were designed based on database information available for different influenza strains. Digoxigenin labelled probes were tested by slot-blot hybridizations to purified RNA from a panel of A/PR/8/34 wild type and A/PR/8/34 reassortant viruses. The results show that the vast majority of reassortants selected for their high growth yield had acquired the non-structural (NS), matrix (M) and RNA polymerase 2 (PB2) genes from the PR8 parent. It is proposed that probes for these genes provide the potential for a simple and rapid procedure for selection of candidate high-yield reassortants for vaccine production.

Genes, Viral↗

PubMed2001

Hypertrophy, hyperplasia, and infectious virus in gut-associated lymphoid tissue of mice after oral inoculation with simian-human or bovine-human reassortant rotaviruses.

Oral inoculation of infants with a vaccine that contains simian-human reassortant rotaviruses has been found to be a rare cause of intussusception. Because intussusception can be associated with enlargement of gut-associated lymphoid tissue, we studied the capacity of simian-human and bovine-human reassortant rotaviruses to cause lymphoid hypertrophy and hyperplasia of Peyer's patches (PP) of adult BALB/c mice. Neither hypertrophy nor hyperplasia was detected in PP after oral inoculation with simian-human or bovine-human reassortant rotaviruses. However, infectious virus was detected in PP and mesenteric lymph nodes after oral inoculation with simian, but not bovine, reassortant rotaviruses. Implications of these findings on the pathogenesis of intussusception are discussed.

Administration, Oral↗

PubMed1996

The development of multivalent bovine rotavirus (strain WC3) reassortant vaccine for infants.

Laboratory and clinical studies have been directed toward development of a vaccine against rotavirus gastroenteritis in infants. First, bovine rotavirus strain WC3, which did not induce neutralizing antibodies to predominant human rotavirus (HRV) serotypes, was determined to be safe and immunogenic; however, it was not protective in all efficacy trials. HRVs adapted to cell culture retained some virulence for infants, but when further attenuated by cold adaptation, they were poorly immunogenic. Reassortant rotaviruses were designed to express HRV surface proteins VP7 (G) or VP4 (P) while retaining a bovine WC3 genome background. Reassortants containing either HRV surface protein and as few as four bovine rotavirus genes were safe in infants. A monovalent WC3 reassortant of serotype G1 specificity was 64%-100% protective in placebo-controlled trials. A quadrivalent WC3 reassortant vaccine with components of HRV G1, G2, G3, and P[8] specificity induced 67% protection against all rotavirus disease in a multicenter efficacy trial.

Animals↗

PubMed2004

A comparison of the effects of oral inoculation with Rotashield and pentavalent reassortant rotavirus vaccine (WC3-PV) on suckling CB17scid mice.

The effects of oral inoculation into infant CB17(scid) mice of two reassortant rotavirus vaccines were compared. The vaccines were Rotashield and WC3-PV, a mixture of five reassortants (G1, G2, G3, G4 and P1; pentavalent reassortant vaccine). Control mice were inoculated with a placebo. At 6 days post-inoculation (p.i.), 8 of 13 (62 %; P<0.005) Rotashield-inoculated mice developed hepatitis and/or bile-duct obstruction compared with none of 11 mice given WC3-PV and none of 14 given placebo. In the Rotashield-inoculated mice, only serotype G3 rhesus rotavirus (RRV) was isolated from multiple sites, including intestine, liver, pancreas, spleen, blood and mesenteric lymph nodes. Recovery of RRV from Rotashield-inoculated mice followed a biphasic pattern. The two peaks of RRV recovery appeared to coincide firstly with replication in the intestine during days 1-3 p.i., and secondly with virus infection of the liver from days 10 to 15 p.i. WC3 reassortants of four different serotypes were detected only at day 1 p.i. in the intestine, liver, pancreas and blood cells from three WC3-PV-inoculated mouse pups. However, WC3-PV did not produce any hepatopathology. Rotashield and WC3-PV appeared to exhibit different biological activity in infant CB17(scid) mouse pups.

Administration, Oral↗

PubMed2006

Increasing appearance of reassortant influenza B virus in Taiwan from 2002 to 2005.

Genetic and antigenic analyses of influenza B virus field strains isolated in Taiwan from 1998 to 2005 were performed. To investigate the molecular evolution of influenza B viruses, sequence analysis of the hemagglutinin (HA1 subunit) and neuraminidase genes was performed. All influenza B viruses isolated between 1998 and 2000 belonged to the B/Yamagata/16/88 lineage. The B/Victoria/2/87 lineage, which was cocirculating with the Yamagata lineage, was identified in Taiwan in March 2001. Concurrently, there was an increasing prevalence of this lineage in many parts of the world, including North America and Europe, during the 2001-2002 season. Since 2002, genetic reassortants of influenza B virus with the Victoria lineage of hemagglutinin and the Yamagata lineage of neuraminidase have been found at a rate of 46%. Therefore, in 2002, at least three sublineages of influenza B virus strains, the B/Shanghai/361/2002-like strain (Yamagata lineage), the B/Hong Kong/330/01-like strain (Victoria lineage), and the B/Hong Kong/1351/02-like strain (B reassortant lineage), were identified in Taiwan. The results showed that genetically distinct lineages can cocirculate in the population and that the reassortment among these strains plays a role in generating the genetic diversity of influenza B viruses. Interestingly, from January to April 2005, B reassortant viruses became dominant (73%) in Taiwan, which indicated that a mismatch had occurred between the influenza B vaccine strain recommended for the 2004-2005 season in the Northern hemisphere by the World Health Organization and the epidemic strain.

Antigens, Viral↗

PubMed2004

Identification of host-specificity determinants in betanodaviruses by using reassortants between striped jack nervous necrosis virus and sevenband grouper nervous necrosis virus.

Betanodaviruses, the causal agents of viral nervous necrosis in marine fish, have bipartite positive-sense RNAs as genomes. The larger genomic segment, RNA1 (3.1 kb), encodes an RNA-dependent RNA polymerase, and the smaller genomic segment, RNA2 (1.4 kb), codes for the coat protein. Betanodaviruses have marked host specificity, although the primary structures of the viral RNAs and encoded proteins are similar among betanodaviruses. However, no mechanism underlying the host specificity has yet been reported. To evaluate viral factors that control host specificity, we first constructed a cDNA-mediated infectious RNA transcription system for sevenband grouper nervous necrosis virus (SGNNV) in addition to that for striped jack nervous necrosis virus (SJNNV), which was previously established by us. We then tested two reassortants between SJNNV and SGNNV for infectivity in the host fish from which they originated. When striped jack and sevenband grouper larvae were bath challenged with the reassortant virus comprising SJNNV RNA1 and SGNNV RNA2, sevenband groupers were killed exclusively, similar to inoculation with SGNNV. Conversely, inoculations with the reassortant virus comprising SGNNV RNA1 and SJNNV RNA2 killed striped jacks but did not affect sevenband groupers. Immunofluorescence microscopic studies using anti-SJNNV polyclonal antibodies revealed that both of the reassortants multiplied in the brains, spinal cords, and retinas of infected fish, similar to infections with parental virus inoculations. These results indicate that viral RNA2 and/or encoded coat protein controls host specificity in SJNNV and SGNNV.

Animals↗

PubMed2006

Isolation and genetic characterization of new reassortant H3N1 swine influenza virus from pigs in the midwestern United States.

Since the introduction of H3N2 swine influenza viruses (SIVs) into U.S. swine in 1998, H1N2 and H1N1 reassortant viruses have emerged from reassortment between classical H1N1 and H3N2 viruses. In 2004, a new reassortant H3N1 virus (A/Swine/Minnesota/00395/2004) was identified from coughing pigs. Phylogenetic analyses revealed a hemagglutinin segment similar to those of contemporary cluster III H3N2 SIVs and a neuraminidase sequence of contemporary H1N1 origin. The internal genes were of swine, human, and avian influenza virus origin, similar to those of contemporary U.S. cluster III H3N2 SIVs. The recovery of H3N1 is further evidence of reassortment among SIVs and justifies continuous surveillance.

Animals↗

PubMed2026

Genomic and Clinicopathological Characterization of a Reassortant HPAI H5N1 (Clade 2.3.4.4b) in an Endangered Cinereous Vulture (Aegypius monachus) in South Korea, 2026.

Clade 2.3.4.4b highly pathogenic avian influenza viruses (HPAIVs) continue to circulate widely in East Asia and undergo frequent reassortment in wild birds. Raptors are regarded as spillover hosts that may be exposed through predation or scavenging, yet integrated clinicopathologic and genomic investigations in cinereous vultures remain limited. Here, we describe a fatal H5N1 HPAIV infection in a cinereous vulture (Aegypius monachus) found in South Korea on January 17, 2026. On presentation, the cinereous vulture showed severe neurologic dysfunction, including inability to stand, right-sided head tilt with pathologic nystagmus, reduced oculocephalic and palpebral reflexes, and intermittent bilateral leg tremors. The cinereous vulture died within 2 days after rescue, and a complete necropsy was performed. Hematologic and biochemical testing revealed marked heterophil predominance, severe lymphopenia, mild monocytosis, and globulin values near the upper end of the reference interval. An oropharyngeal swab tested positive for influenza A virus, and a virus isolate, designated A/Cinereous_Vulture/Korea/26-JBN47/2026(H5N1), was recovered in embryonated chicken eggs. Histopathology showed nonsuppurative encephalitis and necrotizing myocarditis, and influenza A nucleoprotein was detected immunohistochemically in neurons and cardiomyocytes. Tissue real-time RT-PCR showed the lowest cycle threshold value in the brain. Whole-genome sequencing demonstrated that 26-JBN47 belonged to clade 2.3.4.4b and contained a polybasic HA cleavage site (PLREKRRKR/GLF). Segment-level phylogenetic analysis revealed a reassortant genome constellation comprising a maintained H5N1 backbone in HA, NA, and M; low PAIV (LPAIV)-associated but H5N1-incorporated PA and NP segments; flyway-associated PB2 and NS segments; and a PB1 segment phylogenetically linked to regional LPAIV lineages. Molecular marker analysis identified multiple substitutions previously reported to be associated with receptor-binding properties, polymerase-related fitness, virulence, and host-response modulation, whereas canonical PB2 mammalian-adaptive markers were absent. These findings show that 26-JBN47 was a reassortant clade 2.3.4.4b H5N1 HPAIV associated with systemic disease and clinicopathological findings consistent with neurotropic and cardiotropic infection in a cinereous vulture. They also support the potential value of scavenging raptors as sentinels of local or regional HPAIV circulation involving reassortant viruses in East Asia.

Animals↗

PubMed2002

[A preliminary genetic reassortment between Hantaan virus and Seoul virus strains].

OBJECTIVE: To determine the frequency and characteristics of reassortment among Hantaan and Seoul viruses causing hemorrhagic fever with renal syndrome (HFRS). METHODS: Mixed infections were initiated in tissue culture, using Hantaan virus strain 76 - 118 and Seoul virus strain SR-11. Potential reassortant virus plaques were picked out by multiplex RT-PCR, using primers specific for individual genome segments (L, M, S) of each strain. RESULTS: Most of the progeny virus plaques (68.19% of 44) had parental genotype of 76 - 118 strain or SR-11 strain while 2 of 44 plaques had mixed genotypes that yielded RT-PCR bands for the same segment of both parental strains. Reassortant viruses were detected in 68.19% of 44 progeny plaques tested, involving the M and S segments. In addition, approximately 4.55% of the progeny virus plaques appeared to contain S or M segments originating from both parental virus strains, showing that they were diploid. CONCLUSION: Genetic reassortment can occur between Hantaan virus and Seoul virus strains.

Animals↗

PubMedDate not supplied

[B-cell and cytotoxic lymphocyte immune response to virulent, attenuated and reassortant influenza viruses].

The quantitative estimation of the production of the virus specific IgA-, IgM-, IgG1, IgG2a, IgG2b and IgG3-antibody secreting B-cells (ASC) and extent of the immune response of the cytotoxic T-lymphocytes was carried out in mice after primary and secondary immunization with the parental cold adapted (ca) virulent epidemic wild-type (wt) master strain viruses and their reassortant variant (RV) with the incorporated in their genom different genes from ca master strain. The chick embryo derived ca master strain virus A/Len/134/47/57 (H2N2) reduced or eliminated the viral potency to induce the primary B-cellular response. Reassortant incorporation of wt derived HA and NA genes into the ca virus genome restored the virus immunogenic activity concerning the ASC production, but at the lower level than parental virulent virus. Reassortance of the viruses according to generally accepted genom formula 6/2 was associated with decrement of the functional activity of the cytotoxic T-lymphocytes memory and of the primary immune response especially. The data obtain demonstrate the necessity for the control of the immunogenicity of the reassortant viruses at the cloning stage.

Animals↗

PubMed1998

Characterization of a rotavirus rearranged gene 11 by gene reassortment.

The effect of replacement of gene 11 of rotavirus SA-11 by a gene carrying a head to tail duplication obtained from a swine rotavirus strain was studied. The swine rotavirus strain with a duplicated gene (CC86) exhibits both a phenotype that allows to overgrow other viral strains when coinfected and an increased plaque size when plated in both CV-1 and MA-104 monkey kidney cells. Using reassortment methods the duplicated gene of the swine rotavirus was introduced into the SA-11 virus, replacing the regular gene 11. The reassorted strain was characterized to find out the origin of each of the other viral gene segments. Based on electrophoretic mobilities segments 1, 2, 3, 5, 7, 8 and 10 were identified as of SA-11. The SA-11 origin of the segments 4, 6 and 9 was confirmed by neutralization with polyclonal and monoclonal antibodies and by ELISA. The results suggest that the new reassortant virus was a monoreassortant carrying SA-11 genes except the duplicated gene originated from the swine virus CC86. The ability to in vivo replicate and to synthesize viral proteins was identical in the reassorted virus and the parental strains. Sequence analysis indicates that the new phenotype does not originate in the duplication of gene 11 but possibly from mutations in the coding region of NSP5 gene that may result in different phosphorylation patterns of the protein.

Amino Acid Sequence↗

PubMed1983

Phenotypic variation in influenza virus reassortants with identical gene constellations.

To determine which gene segments of influenza A viruses are responsible for the property of tissue tropism, reassortants were produced between the avian influenza strain, A/Mal/NY/6750/78 [H2N2] (Mal/NY), and a human strain, A/Udorn/307/72 [H3N2] (Udorn). The avian strain replicates in the intestinal tract of ducks and the human strain does not. Eight reassortants were shown by hybridization analysis to have the same gene constellation, having received hemagglutinin gene segment 4 from Udorn and the remaining seven gene segments from Mal/NY. With one exception, all reassortants containing the Udorn HA were restricted in their ability to traverse the digestive tract of ducks and replicate therein. The exception, reassortant R2, replicated to high titers in the intestinal tract. The R2 virus was shown to possess a hemagglutinin molecule that was antigenically distinguishable from the Udorn parent with polyclonal and monoclonal antibodies. The virus was "nonavid" in reaction with antihemagglutinin antibodies and was phenotypically similar to avian influenza viruses. The results suggest that the R2 hemagglutinin has undergone mutation(s) altering tissue tropism and antigenic properties of the virus. These studies illustrate the importance of the hemagglutinin gene in determining tissue tropism and present an example of phenotypic variation in a virus population with the same gene constellation but do not exclude a requirement for other gene products.

Animals↗