Differential age susceptibility to influenza B / Victoria lineage viruses in the 2015 Australian influenza season

Influenza B viruses make up an important part of the burden from seasonal influenza globally. The 2015 season in Australia saw an unusual predominance of influenza B with a distinctive switch during the season from B/Yamagata/16/88 lineage viruses to B/Victoria/2/87 lineage viruses. We also noted significant differences in the age groups infected by the different B lineages, with B/Victoria infecting a younger population than B/Yamagata, that could not be explained by potential prior exposure.


The 2015 influenza season in Australia
Lineage data was available for 816 influenza B viruses from 2008 and 1,648 from 2015 that were received by the Centre from all over Australia.The formal representativeness of these samples is unknown.Generally there is a bias towards sampling from children and this is seen in most years in most general and sentinel surveillance systems and was also seen in the 2008 [2] and 2015 [1] Australian influenza seasons.It is unlikely that any bias would exist in selecting patients with a particular B lineage, and given the size and the geographical diversity of the samples tested, it is likely these data will provide an accurate estimate of the overall situation with influenza B in Australia during these two years.During the 2015 pre-season period (January-April) and the early part of the influenza season (May-June), B/Yamagata/16/88 lineage (B/Yam) viruses predominated.However, from July to November, B/Victoria/2/87 lineage (B/Vic) viruses increased rapidly and were dominant from August (Figure 1).Notably, this same switch was seen during the 2008 season with similar timing although almost no B/Vic lineage viruses were detected in Australia before June (Figure 1).The distributions of lineages during 2015 were similar when individual Australian states were examined, with the exception of the Northern Territory, which has a small population largely situated in the tropics, that had an almost total B/Vic year (36/39 B viruses).Australia's most populous state, New South Wales, experienced an increase in the proportion of B/ Vic viruses from low levels early in 2015 to 28% during the period from 15 June to 12 July 2015 [3] which according to our study continued to increase over the rest of the influenza season, and B/Vic viruses predominated from July onwards.Children and young adults carry a higher burden of influenza B disease than older adults and the elderly.According to the Australian Paediatric Surveillance Unit 2015 saw 88 children 15 years and younger (median: 3.3 years) hospitalised with severe complications of influenza between 1 July 2015 and 30 September 2015.Roughly two thirds (n = 59) were influenza B cases (lineage unknown) [1].Overall, the average duration of hospitalisation was four days, 20 required an ICU admission, and there were three influenza-associated deaths, all associated with influenza B infections [1].

Antigenic and genetic drift
The move in dominance by the B/Vic viruses in 2015 was not accompanied by any major antigenic changes from the B/Vic viruses that circulated in smaller numbers in 2014.All Australian B/Vic viruses analysed by the Centre were antigenically B/Brisbane/60/2008like as they were in 2014 (data not shown).Equally, the 2015 B/Yam viruses that were analysed remained antigenically B/Phuket3073/2013-like, similar to B/ Yam viruses that circulated in Australia from mid-2014 (data not shown).However, subtle phylogenetic differences in the haemagglutinin (HA) genes of the two B lineages were apparent (Figure 2).The HA gene phylogeny revealed a greater diversity for B/Yam viruses isolated in Australia during 2015 (n = 56) than during 2014 (n = 42) (Figure 2, top panel).The mean time to most recent common ancestor (mTMRCA) extended beyond 2012 and the isolates belonged to three antigenic types (B/Wisconsin/1/2010, B/Massachusetts/2/2012 and B/Phuket/3073/2013), although the majority from 2015 (52 of the 56 viruses from 2015) belonged to the B/Phuket/3073/2013 clade (also known as group 3) with a mTMRCA in mid-2014.Reconstruction of nonsynonymous changes along this phylogeny revealed an amino acid substitution (M267V) in the subclade of B/Phuket/3073/2013-like viruses that were dominant during the 2015 season, suggesting that this mutation may have contributed to increased viral fitness.All Australian B/Vic viruses from 2015 (n = 54) were phylogenetically B/Brisbane/60/2008-like (also referred to as clade 1A) with an mTMRCA in 2013.This clade was made up of three distinct subclades (Figure 2, bottom panel) the largest of which had the non-synonymous amino acid substitutions V161I and I132V that may have also enhanced the fitness of these viruses.

Age distribution of influenza B infections
The age distribution of patients with confirmed B/ Vic infections in 2015 was positively skewed, with a greater number of infections among the younger age groups (mean: 26.4 years, median: 19.9 years).For B/ Yam infections, the age distribution was more even (mean: 42.4 years, median: 43.8 years; p < 0.001 for Wilcoxon rank sum test).This age differential was less evident in 2008 where, despite the high proportion of viruses obtained from children younger than five years, the interquartile range indicated that B/Yam viruses affected a broader age range than the B/Vic viruses (Figure 3).This age difference between lineages has previously been reported from a household study in Hong Kong [4].There, children younger than 15 years had a 13-fold increased risk of secondary influenza infection with a B/Vic virus than with a B/Yam virus, during a period when both lineages were co-circulating.Similar findings were reported in population studies in southern China during the 2009 and 2010 seasons [5], over three seasons in Slovenia (2010-13 [6]) and in our earlier studies from eastern Australia and New Zealand, where major differences in lineage distribution were observed in subjects older than five years [5,7].

Discussion
A predominance of influenza B viruses in an influenza season occurs infrequently, usually in the order of once every 10 years.Prior to 2015, it last occurred in Australia in the 2008 season, where 54% of typed viruses were influenza B, 43% were influenza A and 3% untyped [2].Similarly in Europe for the seasons from 2001/02 to 2010/11, influenza B was the majority influenza type (59.1%) in only one season (2005/06).In the United States (US) over the same period, 2002/03 was the season with the highest proportion of influenza B (42.6%) among of all typed viruses [8].The two antigenic and genetically distinct lineages of influenza B viruses (B/Yam and B/Vic) have co-circulated in various proportions since 2002 in most countries.Trivalent influenza vaccines (containing only one B virus lineage) used over this time have tried to match these changing lineage circulation patterns.Because of a number of poor matches during the 2000s, quadrivalent vaccines (containing viruses from both B lineages) were developed and have recently been introduced in order to improve vaccine effectiveness.The 2015 influenza vaccines licensed in Australia were all traditional inactivated virus vaccines (live attenuated influenza vaccines and recombinant vaccines were not available) with mostly trivalent vaccine containing only the B/ Yam component being used along with low levels of quadrivalent vaccine.
The reasons for the apparent differential age susceptibility between the two B lineages described in this   In addition, it is unlikely given the low levels of childhood vaccination in Australia that this this would have significantly altered the circulation patterns of the influenza B lineages.Vaccination uptake is generally below 10% among Australian children [12].Childhood influenza vaccination is only recommended for children of aboriginal descent five years and younger and for children six months and older with comorbidities [13].
We have suggested previously that there may be some fundamental differences in the receptor specificity of the different influenza B lineages and that the distribution or density of receptors for influenza B viruses in the respiratory tract of humans may differ with age [7].Others have shown differential responses of children to B/Yam and B/Vic antigens contained in influenza vaccines that might also contribute to differential susceptibility to these two lineages 14].Further work is needed to fully understand the basis of these observations and to determine if the differences are due to receptor variation or density during ageing or prior exposure history or a mixture of both.If indeed young children are at an elevated risk of infection with B/Vic viruses, then it may be prudent to prioritise distribution of quadrivalent vaccines (containing viruses from both B lineages) to this age group.This is relevant to the current northern hemisphere influenza season where the trivalent vaccine contains B/Yam lineage virus, but B/Vic lineage viruses are in our view likely to increase substantially during the current season.Use of the quadrivalent vaccine for this subgroup (or preferably for the whole population) would be potentially advantageous in improving influenza vaccine effectiveness.

Figure 1
Figure 1Relative frequency of influenza B subtypes received by month of specimen collection, Australia, 2008 (n = 816) and 2015 (n = 1,648)

Figure 2
Figure 2 Maximum clade credibility trees showing the evolution of haemagglutinin genes of sequenced Australian influenza B viruses from 2014 and 2015 (n = 168)

Table a
Australian influenza B viruses and haemagglutinin gene sequences used to construct Figure2, obtained from The Global Initiative on Sharing All Influenza Data (GISAID) a IMVS: Institute of Medical and Veterinary Science; Pathwest: Pathwest QE II Medical Centre; QHSS: Queensland Health Scientific Services; VIDRL: Victoria Infectious Diseases Laboratory. a All samples were sequenced and submitted by WHO Collaborating Centre for Reference and Research on Influenza, Melbourne, Australia, with the exception of B/Brisbane/47/2015 that was submitted by US Centers for Disease Control and Prevention.b Authors: A: Deng Y-M, Iannello P, Spirason N, Jelley L, Lau H, Komadina N; B: Deng Y-M, Iannello P, Spirason N, Lau H, Komadina N; C: Tilmanis D, Hurt A, Komadina N.

Table b
Australian influenza B viruses and haemagglutinin gene sequences used to construct Figure2, obtained from The Global Initiative on Sharing All Influenza Data (GISAID) a IMVS: Institute of Medical and Veterinary Science; Pathwest: Pathwest QE II Medical Centre; QHSS: Queensland Health Scientific Services; VIDRL: Victoria Infectious Diseases Laboratory. a All samples were sequenced and submitted by WHO Collaborating Centre for Reference and Research on Influenza, Melbourne, Australia, with the exception of B/Brisbane/47/2015 that was submitted by US Centers for Disease Control and Prevention.b Authors: A: Deng Y-M, Iannello P, Spirason N, Jelley L, Lau H, Komadina N; B: Deng Y-M, Iannello P, Spirason N, Lau H, Komadina N; C: Tilmanis D, Hurt A, Komadina N.

Table c
Australian influenza B viruses and haemagglutinin gene sequences used to construct Figure2, obtained from The Global Initiative on Sharing All Influenza Data (GISAID) a IMVS: Institute of Medical and Veterinary Science; Pathwest: Pathwest QE II Medical Centre; QHSS: Queensland Health Scientific Services; VIDRL: Victoria Infectious Diseases Laboratory. a All samples were sequenced and submitted by WHO Collaborating Centre for Reference and Research on Influenza, Melbourne, Australia, with the exception of B/Brisbane/47/2015 that was submitted by US Centers for Disease Control and Prevention.b Authors: A: Deng Y-M, Iannello P, Spirason N, Jelley L, Lau H, Komadina N; B: Deng Y-M, Iannello P, Spirason N, Lau H, Komadina N; C: Tilmanis D, Hurt A, Komadina N. www.eurosurveillance.org

Table d
Australian influenza B viruses and haemagglutinin gene sequences used to construct Figure 2, obtained from The Global Initiative on Sharing All Influenza Data (GISAID) a IMVS: Institute of Medical and Veterinary Science; Pathwest: Pathwest QE II Medical Centre; QHSS: Queensland Health Scientific Services; VIDRL: Victoria Infectious Diseases Laboratory.All samples were sequenced and submitted by WHO Collaborating Centre for Reference and Research on Influenza, Melbourne, Australia, with the exception of B/Brisbane/47/2015 that was submitted by US Centers for Disease Control and Prevention.For example, five-year-old Australian children in 2015 were likely to have been exposed to a mixture of B/Yam viruses, which circulated in 2013 and 2014, and B/Vic viruses, which circulated from 2009 to 2012, as was the case in for five-year-olds in 2008.Possible exposure therefore fails to explain the elevated proportion of five year-old children infected with B/Yam viruses in 2008 or with B/Vic in 2015.
aIt is probable that the prior exposure history of the different age groups has influenced our observations.However, this is difficult to deduce from the present data.