BACKGROUND
streptococcus pyogenes  is a common cause of bacterial throat infections, and also causes mild to severe skin and soft tissue infections, including impetigo, erysipelas and necrotizing fasciitis.
gas can also cause systemic infections including septicaemia and meningitis, which can be fatal.
estimated annual incidence for invasive gas  infection in industrialised countries is approximately  <dig> per  <dig>  per year  <cit> .
scarlet fever  is characterised by rash, “strawberry tongue”, and exudative pharyngitis and is thought to be due to infection with gas secreting superantigens  <cit> .
sf typically has a cyclical incidence, with resurgences occurring on average every  <dig> years  <cit> .
in england and wales the incidence of sf has been in decline for several decades, reducing from  <dig> notifications per  <dig>  population per year in  <dig> to < <dig> per  <dig>  per year in the 2000s, until the marked increase in 2013/ <dig> season for which  <dig> notifications per  <dig>  were noted in the 2013/ <dig> season  <cit> .
the 2013/ <dig> routine surveillance of notifiable infectious diseases showed an unprecedented increase in the number of cases of sf despite igas numbers remaining within expected seasonal range  <cit> .
a study in the north-west london indicated and association of sf with emm <dig> lineage isolates and an increase in emm <dig> and emm <dig> infections co-incident with the upsurge  <cit> .
previous studies have shown an over-representation of ssa, spea and spec in sf isolates  <cit> .
similarly, multiple emm <dig> sf isolates were recently noted in hong kong encoding ssa  <cit> .
in england, a novel phage has been described in emm <dig> lineage invasive and non-invasive gas strains encoding spec  <cit> .
previous studies have also shown that in epidemic periods specific emm gene sequence types may predominate within the mixed population of strains present including in taiwan   <cit> , hong kong  <cit>  and also in a study of pharyngeal isolates from patients with and without sf in lisbon 2002– <dig>  <cit> .
due to the unprecedented increase in the incidence of sf in england and wales, a national incident management team was convened by phe which initiated investigation of isolates of gas from patients with sf, with the intention of collating a representative selection of sf isolates from all regions of england for genomic analysis.
this was undertaken to determine whether the increase in sf was due to the introduction of new/emerging strain/s into the population or transmission of a known genetic element through the population of gas by horizontal gene transfer  resulting in infections with an increased likelihood of causing sf.
RESULTS
a total of  <dig> isolates of which  <dig> were from sf patients and  <dig> from igas patients were included in the analysis.
emm gene typing
analysis of  <dig> isolates in total including  <dig> from patients with sf and  <dig> from patients with igas indicated  <dig> emm types  in the study population from patients with sf, with emm  <dig>   <dig>   <dig>   <dig> and  <dig>  each accounting for more than 5% of the population .
isolates of  <dig> emm types .
isolates from patients with sf were of multiple distinct emm sequence types with emm <dig> predominating, accounting for  <dig> % ; this was  <dig> % higher than the proportion seen amongst the igas isolates .
although there appeared to be an association between emm <dig> and sf phenotype, this was not found to be statistically significant following application of bonferoni’s correction, when sampling bias per submitting region  was considered.
this association was also not found to be significant when genomic lineage  were taken into consideration .
two emm types  were consistently found to be over-represented in isolates from patients with sf versus igas  following application of bonferroni’s correction.fig.
<dig>
emm gene type in patients with sf and igas.
emm types detected in > 5% population of patients with sf and igas .
note the total isolate number per emm type and for all isolates and normalised random and genomic selection is included, in addition to percentage, 95% confidence intervals and the p value obtained using fisher’s exact test  to compare number of isolates per emm type per study selection groups versus igas isolates
phylogenetic analysis
the population structure of sf and igas isolates  is depicted in fig.
<dig>  the radial branch lengths within the tree represent approximately  <dig> snps.
the igas and sf isolate sequences were found to be evenly spread throughout the phylogeny with no obvious association within a single or small number of clades.
for comparison the percentage of historic referred isolates per emm type from patients with sf is included from  <dig> to  <dig> .fig.
<dig> maximum likelihood phylogenetic tree.
maximum likelihood phylogeny with emm type.
raxml phylogeny based on snp variants of sf and igas mapped to mgas <dig>  sf isolates are coloured red and igas isolates in blue  the radial branch lengths within the tree represent approximately  <dig> snps
population structure, hierarchical clustering
isolates were grouped into genomic clusters based on hierarchical clustering using the number of snp differences as the distance metric and complete linkage at less than  <dig> snp differences.
<dig> genomic clusters were identified, of which the majority had less than  <dig> isolates.. the cochran-mantel-haenszel test was used  to look for association of genomic clusters with differing phenotypes, with results plotted by region and time .
of the  <dig> with more than  <dig> isolates per genomic cluster using the cochran-mantel-haenszel,  <dig> did not have over-representation of sf vs igas or vice versa,  <dig> were over-represented in sf isolates vs igas and  <dig> were over-represented in igas.
when looking at specific genomic clusters  per region over time  all genetic clusters with more than  <dig> representative strains had isolates found in patients with igas or sf.
in many  of the larger clusters with greater than  <dig> isolates  the same genomic snp clusters were found to have representative isolates from patients with igas and sf distributed throughout the time period studied.
in genetic clusters with smaller numbers , uneven sampling will have had a greater impact on the distribution and this may account for the apparent less even distribution of igas and sf isolates over time in smaller clusters.
not all regions were able to provide sufficient sf isolates, with one not submitting any, and others submitted more than the 5% target .fig.
<dig> genomic cluster  per region and time.
sixteen genomic clades  with more than  <dig> referred isolates are plotted - the number in title boxes represent the arbitrary cluster number followed by the emm type.
those clusters significantly associated with igas and sf are coloured with blue and red headers respectively
age and genomic cluster analysis
all genomic clusters where sf isolates were over-represented also contained isolates from adults with igas, indicating absence of a specific strain lineage associated with sf .fig.
<dig> age and genomic cluster.
strains of the same genomic clades were found in both adult and child populations.
strains within specific genomic clusters overrepresented in sf  were also found in adult patients with igas.
similarly, clusters over-represented in igas  all contained cases of sf
horizontal gene transfer
analysis of the association of genes with isolates from cases of sf versus igas showed that ssa was present in all sf isolates .
however, ssa was also found in isolates from patients with igas .
phage content
of  <dig> phage sequences included in the study   <dig> were previously described within emm types  <dig>   <dig>   <dig>   <dig>   <dig>   <dig>   <dig>   <dig>   <dig> .
examination of the subset of isolates that consisted of only these emm types  revealed that no phage was significantly associated with isolates from patients with sf.
however, previous reports have linked presence of ssa gene with sf, we hypothesised that closely related phage may be carrying ssa gene, but may not be individually associated with the sf samples.
to test this hypothesis, new “meta-ssa” phage was defined to be present if any one of the following phage were present:  <dig> , spsp <dig>  mgas <dig>  or hk360ssa.
these phage have been characterised to contain ssa gene  <cit> .
repeating the phage association analysis with the additional phage showed association of the “meta-ssa” with sf .
association of “meta-ssa” phage, but not indivdual ssa carrying phages points towards a polyclonal population of sf phage.
although not significantly over-represented in emm  <dig>  isolates from patients with sf compared to those with igas, a phage with >95% identity to the hku <dig> ssa phage {davies,  <dig> #18} was found in 43/ <dig>  emm <dig> isolates and 1/ <dig>  of emm <dig> isolates, and were absent from all other isolates in this study.
discussion
due to the unprecedented increase in the incidence of sf in england and wales in  <dig> genomic analysis was undertaken to determine whether the increase in sf was due to the introduction of new/emerging strain/s into the population or transmission of a known genetic element through the population of gas by hgt resulting in infections with an increased likelihood of causing sf.
this study found no evidence to support the hypothesis that the increased number of cases of sf was due to the introduction of, or increase in, a single genetic lineage within the population or to a known virulence gene/phage phenomenon as analysis of the population structure  showed that isolates belonging to multiple diverse genetic lineages were present in patients with sf.
isolates from the same lineages were also present in patients with igas.
further analysis revealed seven defined genomic clusters  with > <dig> isolates; seven over-represented in patients with sf versus igas, and three in igas versus sf.
however, sf and igas isolates were found in all genomic clusters with > <dig> isolates.
it is possible that isolates within these clusters are more commonly carried in the throat or have greater propensity to cause invasive infection in humans than other lineage strains.
however, the prevalence of these lineages in asymptomatic individuals is not known.
nonetheless, all  <dig> clusters were found in differing geographical areas and across the time period studied, indicating widespread presence in the population.
studies have shown that an expansion of an igas clone from patients in the san francisco bay area during 2005– <dig> differed on average by only  <dig> snps within  <dig> isolates  <cit>  the phylogenetic and epidemiologic data suggest that these  <dig> isolates constitute a distinct clone that caused a geographic cluster of invasive infections  <cit> .
therefore, the variation observed within the isolates in < <dig> snp clusters included in this study must have arisen over a long time period and not within the documented period of increase in sf in  <dig>
in  <dig> a rise was noted in sf in vietnam [referenced in  <cit> , and in  <dig> increases of sf were reported in china and hong kong  <cit> .
these increases were attributed to mixed emm lineage clones with multiple clones of emm <dig> predominating, followed by emm <dig> including strains in which an integrative conjugative element encoding resistance to tetracycline , erythromycin  and clindamycin, and bacteriophage hku <dig> vir encoding superantigens ssa and spec were noted  <cit> .
the finding from this study that the ssa gene was significantly associated with isolates from patients with sf is not surprising, as this is consistent with findings reported by other smaller studies  <cit> .
it has been speculated that in a single emm lineage the acquisition of ssa can confer potential changes in disease and pathogenicity  <cit> .
however this has not been demonstrated in multiple lineages, and is unlikely to occur within an entire population of multiple lineages within the limited time period of a few months.
in terms of phage, no single known phage was found in this study, rather several differing phage carrying ssa in isolates from patients with sf were noted.
however, strains were not analysed for as yet undiscovered phage harbouring ssa or additional genetic factors, hence such genetic elements cannot be fully discounted.
in this study several emm gas lineages were found in patients with sf, similar to that seen historically in england .
two emm lineage clusters were found to be over-represented in sf isolates versus igas; emm <dig> and emm <dig>  consistent with findings from north west london of an over-representation of emm <dig>  <cit> .
these two emm lineages were not found within the same phylogenetic major clade .
all published data regarding large increases in wide geographic areas in sf have involved multiple emm lineage strains, albeit with a single predominant emm lineage described for each study, such as emm <dig> in taiwan, china and hong kong in  <dig>  <cit> .
the predominant strain emm lineage in this study was also found to be emm <dig>  however this lineage was not significantly over-represented in isolates from patients with sf versus igas, unlike turner et al.
<cit>  comparison to isolates from the upper respiratory tract.
historical submissions of isolates from  <dig> to  <dig> from patients with sf indicate an annual fluctuation in the predominant emm type strain per annum, including emm <dig>   <dig> and  <dig> and could indicate type specific fluctuation in population immunity .
however these isolates are submitted on an ad hoc basis and are not from a defined sampling scheme.
changes in predominant emm types in children with sf in the beijing population over time and with area specific changes have recently been documented  <cit> .
further study of two genetic elements recently discovered within some emm <dig> lineage strains  may uncover an important role for these elements in conferring the sf phenotype in patients  <cit> .
the hku <dig> vir phage described in some isolates from the  <dig> hong kong sf increase  <cit>  was not present in isolates in this study, whereas the hku <dig> ssa was present in the majority of emm <dig> type isolates but was not found to be over-represented in isolates from patients with sf.
in summary, the large observed increase in the number of sf cases cannot be attributed to a single genetic factor.
without being able to determine the proportion of the different lineages and the genetic factors they carry within the asymptomatic/carriages population it is not possible to determine if the lineages observed in the disease causing isolates mirrors those seen in the population as whole.
if the population structure within these two human host groups was similar it would lend weight to alternative hypotheses such as the increase in sf cases being due to alternative factors.
these could include host population immunity status, concurrent circulation of other predisposing pathogens such as respiratory viruses, potential declining healthcare attendance for patients with gas infection resulting in increased likelihood for infection to progress to sf and reduction in the use of antimicrobials affecting pharyngeal carriage in the population.
geographically representative sampling was limited by isolates available in clinical laboratories and consequently  <dig> of  <dig> regions of england submitted fewer samples than the requested 5% sample and multiple submissions from localised clusters could not be discounted during collection.
isolates from periods outside the study period, including those during the height of the epidemic peak were not available for comparison.
therefore sufficient isolates were not available to undertake a comprehensive genome wide association study to determine additional other/novel genetic factors associated with sf.
CONCLUSIONS
the increase in sf cases was not caused by the introduction of a single or dominant clone but is associated with multiple lineages within the gas population currently circulating in england.
the ssa gene was over-represented in isolates from patients with sf.
although no single ssa carrying phage was found to be over represented in sf vs igas, the “meta-ssa” phage defined by the presence of : <dig> , spsp <dig>  mgas <dig>  or hk360ssa , was found to be over represented in isolates from patients with sf.
the hku <dig> vir phage described in some isolates from the  <dig> hong kong sf increase  <cit>  was not present in any of the  <dig> isolates analysed in this study.
the hku <dig> ssa phage associated with some emm <dig> isolates from the  <dig> hong kong sf study was present in 43/ <dig> emm <dig> isolates, but was not found to be over-represented in isolates from patients with sf.
two emm types,  <dig> and  <dig>  were found to be over-represented in patients with sf versus igas , but were not restricted to sf patients.
the emm <dig> lineage isolates were not found to be over-represented following genomic and regional normalisation indicating a large proportion of emm <dig> isolates were received from a single region.
other factors need to be examined to determine contribution to the national reported increase in sf.

 streptococcus pyogenes is a common cause of bacterial throat infections, and also causes mild to severe skin and soft tissue infections, including impetigo, erysipelas and necrotizing fasciitis. gas can also cause systemic infections including septicaemia and meningitis, which can be fatal. estimated annual incidence for invasive gas infection in industrialised countries is approximately per per year. scarlet fever is characterised by rash, “strawberry tongue”, and exudative pharyngitis and is thought to be due to infection with gas secreting superantigens. sf typically has a cyclical incidence, with resurgences occurring on average every years. in england and wales the incidence of sf has been in decline for several decades, reducing from notifications per population per year in to < per per year in the 2000s, until the marked increase in 2013/ season for which notifications per were noted in the 2013/ season. the 2013/ routine surveillance of notifiable infectious diseases showed an unprecedented increase in the number of cases of sf despite igas numbers remaining within expected seasonal range. a study in the north-west london indicated and association of sf with emm lineage isolates and an increase in emm and emm infections co-incident with the upsurge. previous studies have shown an over-representation of ssa, spea and spec in sf isolates. similarly, multiple emm sf isolates were recently noted in hong kong encoding ssa. in england, a novel phage has been described in emm lineage invasive and non-invasive gas strains encoding spec. previous studies have also shown that in epidemic periods specific emm gene sequence types may predominate within the mixed population of strains present including in taiwan, hong kong and also in a study of pharyngeal isolates from patients with and without sf in lisbon 2002–. due to the unprecedented increase in the incidence of sf in england and wales, a national incident management team was convened by phe which initiated investigation of isolates of gas from patients with sf, with the intention of collating a representative selection of sf isolates from all regions of england for genomic analysis. this was undertaken to determine whether the increase in sf was due to the introduction of new/emerging strain/s into the population or transmission of a known genetic element through the population of gas by horizontal gene transfer resulting in infections with an increased likelihood of causing sf.  a total of isolates of which were from sf patients and from igas patients were included in the analysis. emm gene typing analysis of isolates in total including from patients with sf and from patients with igas indicated emm types in the study population from patients with sf, with emm and each accounting for more than 5% of the population. isolates of emm types. isolates from patients with sf were of multiple distinct emm sequence types with emm predominating, accounting for %; this was % higher than the proportion seen amongst the igas isolates. although there appeared to be an association between emm and sf phenotype, this was not found to be statistically significant following application of bonferoni’s correction, when sampling bias per submitting region was considered. this association was also not found to be significant when genomic lineage were taken into consideration. two emm types were consistently found to be over-represented in isolates from patients with sf versus igas following application of bonferroni’s correction.fig. emm gene type in patients with sf and igas. emm types detected in > 5% population of patients with sf and igas. note the total isolate number per emm type and for all isolates and normalised random and genomic selection is included, in addition to percentage, 95% confidence intervals and the p value obtained using fisher’s exact test to compare number of isolates per emm type per study selection groups versus igas isolates phylogenetic analysis the population structure of sf and igas isolates is depicted in fig.  the radial branch lengths within the tree represent approximately snps. the igas and sf isolate sequences were found to be evenly spread throughout the phylogeny with no obvious association within a single or small number of clades. for comparison the percentage of historic referred isolates per emm type from patients with sf is included from to.fig. maximum likelihood phylogenetic tree. maximum likelihood phylogeny with emm type. raxml phylogeny based on snp variants of sf and igas mapped to mgas sf isolates are coloured red and igas isolates in blue the radial branch lengths within the tree represent approximately snps population structure, hierarchical clustering isolates were grouped into genomic clusters based on hierarchical clustering using the number of snp differences as the distance metric and complete linkage at less than snp differences. genomic clusters were identified, of which the majority had less than isolates.. the cochran-mantel-haenszel test was used to look for association of genomic clusters with differing phenotypes, with results plotted by region and time. of the with more than isolates per genomic cluster using the cochran-mantel-haenszel, did not have over-representation of sf vs igas or vice versa, were over-represented in sf isolates vs igas and were over-represented in igas. when looking at specific genomic clusters per region over time all genetic clusters with more than representative strains had isolates found in patients with igas or sf. in many of the larger clusters with greater than isolates the same genomic snp clusters were found to have representative isolates from patients with igas and sf distributed throughout the time period studied. in genetic clusters with smaller numbers, uneven sampling will have had a greater impact on the distribution and this may account for the apparent less even distribution of igas and sf isolates over time in smaller clusters. not all regions were able to provide sufficient sf isolates, with one not submitting any, and others submitted more than the 5% target.fig. genomic cluster per region and time. sixteen genomic clades with more than referred isolates are plotted - the number in title boxes represent the arbitrary cluster number followed by the emm type. those clusters significantly associated with igas and sf are coloured with blue and red headers respectively age and genomic cluster analysis all genomic clusters where sf isolates were over-represented also contained isolates from adults with igas, indicating absence of a specific strain lineage associated with sf.fig. age and genomic cluster. strains of the same genomic clades were found in both adult and child populations. strains within specific genomic clusters overrepresented in sf were also found in adult patients with igas. similarly, clusters over-represented in igas all contained cases of sf horizontal gene transfer analysis of the association of genes with isolates from cases of sf versus igas showed that ssa was present in all sf isolates. however, ssa was also found in isolates from patients with igas. phage content of phage sequences included in the study were previously described within emm types. examination of the subset of isolates that consisted of only these emm types revealed that no phage was significantly associated with isolates from patients with sf. however, previous reports have linked presence of ssa gene with sf, we hypothesised that closely related phage may be carrying ssa gene, but may not be individually associated with the sf samples. to test this hypothesis, new “meta-ssa” phage was defined to be present if any one of the following phage were present:, spsp mgas or hk360ssa. these phage have been characterised to contain ssa gene. repeating the phage association analysis with the additional phage showed association of the “meta-ssa” with sf. association of “meta-ssa” phage, but not indivdual ssa carrying phages points towards a polyclonal population of sf phage. although not significantly over-represented in emm isolates from patients with sf compared to those with igas, a phage with >95% identity to the hku ssa phage {davies, #18} was found in 43/ emm isolates and 1/ of emm isolates, and were absent from all other isolates in this study. discussion due to the unprecedented increase in the incidence of sf in england and wales in genomic analysis was undertaken to determine whether the increase in sf was due to the introduction of new/emerging strain/s into the population or transmission of a known genetic element through the population of gas by hgt resulting in infections with an increased likelihood of causing sf. this study found no evidence to support the hypothesis that the increased number of cases of sf was due to the introduction of, or increase in, a single genetic lineage within the population or to a known virulence gene/phage phenomenon as analysis of the population structure showed that isolates belonging to multiple diverse genetic lineages were present in patients with sf. isolates from the same lineages were also present in patients with igas. further analysis revealed seven defined genomic clusters with > isolates; seven over-represented in patients with sf versus igas, and three in igas versus sf. however, sf and igas isolates were found in all genomic clusters with > isolates. it is possible that isolates within these clusters are more commonly carried in the throat or have greater propensity to cause invasive infection in humans than other lineage strains. however, the prevalence of these lineages in asymptomatic individuals is not known. nonetheless, all clusters were found in differing geographical areas and across the time period studied, indicating widespread presence in the population. studies have shown that an expansion of an igas clone from patients in the san francisco bay area during 2005– differed on average by only snps within isolates the phylogenetic and epidemiologic data suggest that these isolates constitute a distinct clone that caused a geographic cluster of invasive infections. therefore, the variation observed within the isolates in < snp clusters included in this study must have arisen over a long time period and not within the documented period of increase in sf in in a rise was noted in sf in vietnam [referenced in, and in increases of sf were reported in china and hong kong. these increases were attributed to mixed emm lineage clones with multiple clones of emm predominating, followed by emm including strains in which an integrative conjugative element encoding resistance to tetracycline, erythromycin and clindamycin, and bacteriophage hku vir encoding superantigens ssa and spec were noted. the finding from this study that the ssa gene was significantly associated with isolates from patients with sf is not surprising, as this is consistent with findings reported by other smaller studies. it has been speculated that in a single emm lineage the acquisition of ssa can confer potential changes in disease and pathogenicity. however this has not been demonstrated in multiple lineages, and is unlikely to occur within an entire population of multiple lineages within the limited time period of a few months. in terms of phage, no single known phage was found in this study, rather several differing phage carrying ssa in isolates from patients with sf were noted. however, strains were not analysed for as yet undiscovered phage harbouring ssa or additional genetic factors, hence such genetic elements cannot be fully discounted. in this study several emm gas lineages were found in patients with sf, similar to that seen historically in england. two emm lineage clusters were found to be over-represented in sf isolates versus igas; emm and emm consistent with findings from north west london of an over-representation of emm. these two emm lineages were not found within the same phylogenetic major clade. all published data regarding large increases in wide geographic areas in sf have involved multiple emm lineage strains, albeit with a single predominant emm lineage described for each study, such as emm in taiwan, china and hong kong in. the predominant strain emm lineage in this study was also found to be emm however this lineage was not significantly over-represented in isolates from patients with sf versus igas, unlike turner et al. comparison to isolates from the upper respiratory tract. historical submissions of isolates from to from patients with sf indicate an annual fluctuation in the predominant emm type strain per annum, including emm and and could indicate type specific fluctuation in population immunity. however these isolates are submitted on an ad hoc basis and are not from a defined sampling scheme. changes in predominant emm types in children with sf in the beijing population over time and with area specific changes have recently been documented. further study of two genetic elements recently discovered within some emm lineage strains may uncover an important role for these elements in conferring the sf phenotype in patients. the hku vir phage described in some isolates from the hong kong sf increase was not present in isolates in this study, whereas the hku ssa was present in the majority of emm type isolates but was not found to be over-represented in isolates from patients with sf. in summary, the large observed increase in the number of sf cases cannot be attributed to a single genetic factor. without being able to determine the proportion of the different lineages and the genetic factors they carry within the asymptomatic/carriages population it is not possible to determine if the lineages observed in the disease causing isolates mirrors those seen in the population as whole. if the population structure within these two human host groups was similar it would lend weight to alternative hypotheses such as the increase in sf cases being due to alternative factors. these could include host population immunity status, concurrent circulation of other predisposing pathogens such as respiratory viruses, potential declining healthcare attendance for patients with gas infection resulting in increased likelihood for infection to progress to sf and reduction in the use of antimicrobials affecting pharyngeal carriage in the population. geographically representative sampling was limited by isolates available in clinical laboratories and consequently of regions of england submitted fewer samples than the requested 5% sample and multiple submissions from localised clusters could not be discounted during collection. isolates from periods outside the study period, including those during the height of the epidemic peak were not available for comparison. therefore sufficient isolates were not available to undertake a comprehensive genome wide association study to determine additional other/novel genetic factors associated with sf.  the increase in sf cases was not caused by the introduction of a single or dominant clone but is associated with multiple lineages within the gas population currently circulating in england. the ssa gene was over-represented in isolates from patients with sf. although no single ssa carrying phage was found to be over represented in sf vs igas, the “meta-ssa” phage defined by the presence of:, spsp mgas or hk360ssa, was found to be over represented in isolates from patients with sf. the hku vir phage described in some isolates from the hong kong sf increase was not present in any of the isolates analysed in this study. the hku ssa phage associated with some emm isolates from the hong kong sf study was present in 43/ emm isolates, but was not found to be over-represented in isolates from patients with sf. two emm types, and were found to be over-represented in patients with sf versus igas, but were not restricted to sf patients. the emm lineage isolates were not found to be over-represented following genomic and regional normalisation indicating a large proportion of emm isolates were received from a single region. other factors need to be examined to determine contribution to the national reported increase in sf. 
during a substantial elevation in scarlet fever notifications in a national genomic study was undertaken of streptococcus pyogenes isolates from patients with sf with comparison to isolates from patients with invasive disease to test the hypotheses that the increase in sf was due to either the introduction of one or more new/emerging strains in the population in england or the transmission of a known genetic element through the population of gas by horizontal gene transfer resulting in infections with an increased likelihood of causing sf. there is no evidence that the increased number of sf cases was a strain-specific or known mobile element specific phenomenon, as the increase in sf cases was associated with multiple lineages of gas. post-normalisation emm types, and were found to be over-represented in patients with sf versus igas. no single phage was found to be over represented in sf vs igas. isolates from patients with sf were of multiple distinct emm sequence types and phylogenetic lineages.
 during a substantial elevation in scarlet fever notifications in a national genomic study was undertaken of streptococcus pyogenes isolates from patients with sf with comparison to isolates from patients with invasive disease to test the hypotheses that the increase in sf was due to either the introduction of one or more new/emerging strains in the population in england or the transmission of a known genetic element through the population of gas by horizontal gene transfer resulting in infections with an increased likelihood of causing sf. isolates were collected to provide geographical representation, for approximately 5% sf isolates from each region from 1st april to 18th june contemporaneous igas isolates for which genomic data were available were included for comparison. data were analysed in order to determine emm gene sequence type, phylogenetic lineage and genomic clade representation, the presence of known prophage elements and the presence of genes known to confer pathogenicity and resistance to antibiotics.  isolates were analysed, from patients with sf and from patients with igas. isolates from patients with sf were of multiple distinct emm sequence types and phylogenetic lineages. prior to data normalisation, emm was the predominant type. post-normalisation emm types, and were found to be over-represented in patients with sf versus igas. a single gene, ssa, was over-represented in isolates from patients with sf. no single phage was found to be over represented in sf vs igas. however, a “meta-ssa” phage defined by the presence of:, spsp mgas or hk360ssa, was found to be over represented. the hku vir phage was not detected yet the hku ssa phage was present in 43/ emm isolates but not found to be over-represented in isolates from patients with sf.  there is no evidence that the increased number of sf cases was a strain-specific or known mobile element specific phenomenon, as the increase in sf cases was associated with multiple lineages of gas. electronic supplementary material the online version of this article contains supplementary material, which is available to authorized users. keywords scarlet feverstreptococcus pyogenesgenomicsigasinvasive gashttp://dx.doi.org/ /501100002141public health englandissue-copyright-statement© the author 2017 