Molecular epidemiology of respiratory syncytial virus: rapid identification of subgroup A lineages

doi:10.1016/0166-0934(92)90088-U

Journal of Virological Methods

Volume 40, Issue 3, 1 December 1992, Pages 297-306

https://doi.org/10.1016/0166-0934(92)90088-U Get rights and content

Abstract

Methods for the rapid analysis of samples of respiratory syncytial (RS) virus are described using the polymerase chain reaction (PCR) followed by restriction mapping. Isolates (either clinical samples or tissue culture grown virus) can readily be divided into subgroups and then further classified into lineages. These methods enable examination of large numbers of isolates by molecular techniques, thereby facilitating research into the molecular epidemiology of the virus.

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Comparison of virological profiles of respiratory syncytial virus and rhinovirus in acute lower tract respiratory infections in very young Chilean infants, according to their clinical outcome
2014, Journal of Clinical Virology
Citation Excerpt :
The reaction was carried out for 5 min at 94 °C and for 35 cycles at 94 °C for 1 min, 55 °C for 1 min and 72 °C for 1 min and followed by 10 min of extension at 72 °C. Amplified products of 653 bp of RSV A and 656 bp of RSV B were visualized using ethidium bromide under UV light on a 1.5% agarose gel [16]. When required, semi nested PCR was performed with GAB and F1 primers [18] resulting in 489 bp and 492 bp products, depending on genotype.
Respiratory syncytial virus (RSV) and rhinovirus (HRV) are the main cause of acute lower respiratory tract infections (ALRTIs) in infants. Viral and host-related risk factors for severe disease have also not been clearly established.
To assess whether certain viral features of RSV and, or HRV are associated with severe ALRTI.
RSV and HRV were studied in nasopharyngeal samples of infants by immunofluorescence, Luminex^® and/or real-time RT-PCR assays. Quantitation and genotyping of RSV and HRV by PCR were done.
Of 124 virus positive specimens, 74 (59.7%) had RSV; 22 (17.7%) HRV and 28 (22.6%) RSV-HRV co-infection. Hospitalization was required in 57/74 RSV infants (77.0%); in 10/22 HRV cases (45.5%) (p = 0.006) and in 15/28 co-infected by both viruses (53.6%) (p = 0.003). Severe cases were 33/74 (44.6%) RSV infections, 2/22 HRV cases (9.1%), (p < 0.002) and 6/28 (21.4%) patients co-infected by RSV–HRV (p < 0.026). Three genotypes (NA1, B7, B9) of RSV circulated during the study. In 33 severe infants, NA1 was detected in 19 cases (57.6%); B7 in 13 (39.4%) and B9 in 1 (3.0%) (p < 0.01; OR = 10.0). RSV loads were similar between outpatients and hospitalized infants (p = 0.7) and among different severities (p = 0.7). NA1 loads were higher than other strains (p = 0.049). Three geno-groups of HRV circulated homogeneously.
In very young infants, RSV cause more severe disease than HRV. Co-infection does not increase the severity of illness. NA1 RSV genotype was associated with major frequency of hospitalization, severe respiratory disease and higher viral load.
Respiratory syncytial virus infection and recurrent wheezing in Chilean infants: A genetic background?
2013, Infection, Genetics and Evolution
Citation Excerpt :
Reverse transcription in a Perkin Elmer Gene Amp® PCR System 2400 was performed using a F gene primer (5′ TGTCTAACTATTTGAACA 3′, nucleotides 844–861 of F gene of Long RSV strain) as published (López et al., 1998). A fragment of the N gene with specific primers (Cane and Pringle, 1992) was amplified by real time PCR in a Light Cycler 1.5 instrument (Roche®). A clinical score system, previously published by authors (Larrañaga et al., 2009), was used in order to characterize the severity of the RSV infection.
Respiratory syncytial virus (RSV) infection has been associated to recurrent wheezing, but pathogenic mechanisms are unclear. Interleukin-4/Interleukin-13 (IL-4/IL-13) pathway is involved in both conditions. A common host genetic susceptibility may exist in patients whom RSV will trigger severe illness and those who develop recurrent wheezing.
To assess, by a candidate-gene approach, whether genetic polymorphisms in IL-4/IL-13 pathway are associated with RSV infection severity and its outcome in Chilean children.
A cohort of 118 RSV-infected infants was analyzed and followed for one year. Severity of acute infection and later recurrent wheezing were characterized. Alleles and genotypes frequencies were determined for two SNP in each of the genes IL-4, IL-13 and IL-4Rα. Association tests and interaction analyses were performed.
Enrollment included 60 moderate and 58 severe cases. Two SNP were found associated to severity during acute infection in IL-4Rα gene (Gln551Arg, Ile50Val). The follow up was completed in 71% of patients (84/118). Later recurrent wheezing was 54% in severe group, versus 31% in moderate cases (p = 0.035). In relation to outcome, allele Ile50 in IL-4Rα was more frequent in patients with moderate disease and no wheezing outcome. A common protector genotype is proposed for Chilean children: IL-4Rα Ile/Ile.
Genetic variations in the host are associated to infection severity and outcome. A common genetic background might be influencing both pathologies.
SP-A1, SP-A2 and SP-D gene polymorphisms in severe acute respiratory syncytial infection in Chilean infants
2011, Infection, Genetics and Evolution
Citation Excerpt :
Reverse transcription was performed with F gene primer (5′ tgtctaactatttgaaca 3′, based in F gene of Long strain) in a Perkin Elmer Gene Amp® PCR System 2400. A fragment of the N gene with specific primers (Cane and Pringle, 1992) was amplified by real time PCR using a Light Cycler® Fast Start DNA Master SybR Green I kit (Light Cycler 1.5 Roche®). RSV cases were confirmed as positive for any infant, if their first NPA sample was positive by IFA at the emergency room and their second NPA sample also identified RSV by either real time PCR, IFA or viral isolation.
Respiratory syncytial virus (RSV) is the principal pathogen that causes acute lower respiratory tract infection (ALRI) in infants. Severe RSV-ALRI has been associated with the host genetic susceptibility. To assess whether severe RSV disease in infants is associated with certain single nucleotide polymorphism (SNP) into the gene of SP-A1, SP-A2 and SP-D, a prospective study was performed among blood donors and RSV-infected infants aged < or = 6 months, considering their severity, according to a strict scoring system. Allele and genotype frequencies were compared using χ²-test. Association studies and haplotype analysis were tested by using Armitagës trend test and Unphased 3.0 program.
A total of 118 RSV-infected infants and 104 blood donors were enrolled into the study; 59 infants had a severe respiratory disease, 34 children developed a moderate illness and 25 had a mild disease. There was no difference in the allelic and genotypic frequencies of SP-A1, but intragenic haplotypes showed significant differences among infected infants and blood donors (p = 0.0021). 1A⁰ variant of SP-A2 was the most frequent allele in all groups. Thr₁₁ allele of SP-D is significantly higher in RSV infants (p = 0.028), as given by its higher frequency in severe disease (p = 0.046). Heterozygous Thr₁₁/Met₁₁ was significantly more common in infected infants (p = 0.037), because it has higher frequency in critically ill children (p = 0.017). Thr₁₆₀ allele was significantly higher in severe infants compared with blood donors (p = 0.046) and infants with mild disease (p = 0.018). Thr₁₁-Thr₁₆₀-Ser₂₇₀ haplotype was significantly more common in RSV-infants, due to severe (p = 0.00000034) and moderate disease (p = 0.000009). Differences were also found among severe and mild disease (p = 0.026). Differences found with other authors, indicate the need for local studies to identify genetic biomarkers of severity.
Human respiratory syncytial virus genomic and antigenic variants isolated in two hospitals during one epidemic, in Santiago, Chile
2008, Journal of Clinical Virology
Citation Excerpt :
The mixture was incubated for 60 min at 37 °C, followed by 5 min at 95 °C in a Gene Amp PCR System 2400 thermocycler (PerkinElmer®). Fragments of the genes N (nucleotides 858–1135) and G (nucleotides 1–584) were amplified as described previously (Cane and Pringle, 1992). PCR products were purified by columns (Qiagen®) and their genetic variability was analyzed by restriction fragments length polymorphism (RFLP) as previously described (Cane and Pringle, 1992).
Human respiratory syncytial virus (HRSV) is a major cause of severe lower respiratory tract infection (LRI) in children. Distinct variants of the viruses have been described.
The objective was to compare the antigenic and genetic variability of HRSV strains recovered from infants admitted to two hospitals during one epidemic in a big city.
We analyzed nasopharyngeal aspirates from 201 infants admitted for LRI to two hospitals during 2002 in Santiago, Chile. The analyses were carried out using a panel of monoclonal antibodies against G glycoprotein epitopes (EIA) and RFLP for N and G genes.
No differences in HRSV groups A/B and in N patterns distribution were observed among both hospitals. On the contrary, antigenic and genetic G patterns displayed a wide diversity of strains circulating during one epidemic, in one big city.
RSV variability assessment depended rather on the tool used for analysis than on the geographical location.
Understanding the transmission dynamics of respiratory syncytial virus using multiple time series and nested models
2007, Mathematical Biosciences
The nature and role of re-infection and partial immunity are likely to be important determinants of the transmission dynamics of human respiratory syncytial virus (hRSV). We propose a single model structure that captures four possible host responses to infection and subsequent reinfection: partial susceptibility, altered infection duration, reduced infectiousness and temporary immunity (which might be partial). The magnitude of these responses is determined by four homotopy parameters, and by setting some of these parameters to extreme values we generate a set of eight nested, deterministic transmission models. In order to investigate hRSV transmission dynamics, we applied these models to incidence data from eight international locations. Seasonality is included as cyclic variation in transmission. Parameters associated with the natural history of the infection were assumed to be independent of geographic location, while others, such as those associated with seasonality, were assumed location specific. Models incorporating either of the two extreme assumptions for immunity (none or solid and lifelong) were unable to reproduce the observed dynamics. Model fits with either waning or partial immunity to disease or both were visually comparable. The best fitting structure was a lifelong partial immunity to both disease and infection. Observed patterns were reproduced by stochastic simulations using the parameter values estimated from the deterministic models.
Respiratory Syncytial Virus Group A and B Genotypes and Disease Severity among Cuban Children
2006, Archives of Medical Research
Citation Excerpt :
The mixture was incubated for 2 h at the recommended temperature according to the manufacturer's instructions for each enzyme. The digests were subjected to electrophoresis in 4% agarose gels (3:1) in a mixture of three parts of LE agarose (analytical grade, Promega) and 1 part LM agarose (Low Melting, Promega) containing 5 μg/mL of ethidium bromide in 1X Tris-borate buffer and then photographed (14,15). Medical records from the 40 selected patients who had positive results of IFA and PCR for RSV were abstracted, and the following parameters were recorded: age, breastfeeding, birthweight and prematurity.
Respiratory syncytial virus (RSV) is the leading cause of serious lower tract infections in infants. Comorbid conditions such as chronic diseases and prematurity have been associated with greater severity illness, but virus genotypes and disease severity is still unknown.
Forty selected strains of RSV group A and B from Cuban infants with acute respiratory disease (ARD) over five seasons were studied. Viral RNA was extracted and polymerase chain reaction (PCR) was carried out using direct primers directed to parts of the nucleoprotein (N) and fusion (F) genes, respectively. Amplicons were digested using restriction fragment length polymorphism (RFLP) to define the association between virus and disease severity. Disease severity was assessed as very mild, mild, moderate, and severe.
Three of six known N genotypes were detected. NP4 and NP3 were found more frequently; moreover, it was difficult to establish a relationship between N genotypes and disease severity. Five genotypes in F gene were found: F1, F2, F5, F9 and F11; F9 and F11 were associated with very mild disease, but F1 genotype appears to be associated with moderate to severe disease.
At least five combinations of N and F genotypes circulated in the studied infants in Cuba. Patients with F1NP4 genotype showed moderate to severe disease. Relationship between genotypes and disease severity was established.

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Molecular epidemiology of respiratory syncytial virus: rapid identification of subgroup A lineages

Abstract

Virus Res.

Virus Res.

Virology

Antigenic characterization of respiratory syncytial virus strains with monoclonal antibodies

J. Infec. Dis.

Sequence of the major nucleocapsid protein gene of pneumonia virus of mice: sequence comparisons suggest structural homology between nucleocapsid proteins of pneumoviruses, paramyxoviruses, rhabdoviruses and filoviruses

J. Gen. Virol.

Respiratory syncytial virus heterogeneity during an epidemic: analysis by limited nucleotide sequencing (SH gene) and restriction mapping (N gene)

J. Gen. Virol.

Identification of variable domains of the attachment (G) protein of subgroup A respiratory syncytial viruses

J. Gen. Virol.