RECOMBINOMICS

[dothedd]

All December H1N1 UK Sequences Have S188T
Recombinomics Commentary 23:50
January 18, 2011


The recently released sequences from Iran from mid-December focused attention on the emergence of H1N1 with either S186P or S188T. All four had one of the two changes and three had S188T.

A closer look at the recently released sequences from the UK showed a similar trend.

Ten of the UK sequences were collected in December, and all ten had S188T. All ten had G605C (S188T), T1056C, G1403A (S454N), as had been previously described for the clonal expansion of S188T. However, in the UK the December isolates formed two branches.

The branch which acquired G340A (D100N) via recombination had the two severe cases with D225G (A/England/488038/2010 and A/England/4940476/2010), and a fatal case (A/England/5040499/2010). The other branch had A40G, G207A, and G1629T, which had seven milder cases (A/England/280/2010, A/England/5000135/2010, A/England/5000125/2010, A/England/4980078/2010, A/England/4980074/2010, A/England/4960201/2010, and A/England/4940126/2010).

A similar trend was seen in the November collections. Ten more had S188T, while six had S186P. Only one of the seventeen November collections did not have one of the receptor binding domain change.

These data highlight the emergence of sequences with either S186P or S188T, as well as S188T increasing in frequency in the most recent collections.

A similar trend was seen in the United States and Japan, signaling the emergence and fixing of these changes in 2011.

Release of December H1N1 sequences from additional countries would be useful.

[dothedd]

All Oct/Nov H1N1 US Sequences Have S186P or S188T
Recombinomics Commentary 13:20
January 19, 2011


The recently released HA sequences from Iran and the UK have focused attention on two emerging sub-clades which have S186P or S188T. The most recent sequences are from Iran and were collected in mid-December, 2010. All four have one of the two changes including three with S188T. Three earlier sequences, from October or early December also had the same two markers. Similarly, all December sequences from the UK had S188T, and 16/17 November sequences have one of the markers including ten with S188T.

The CDC has recently released a small series of sequences from the United States and although none are from December, six are from October or November and all six have one of the two markers. The two most recent sequences are from the fatal cases in Lehigh, Pennsylvania. Both sequences (A/Pennsylvania/16/2010 and A/Pennsylvania/17/2010) are identical and have S186P. The other November isolate, A/Kentucky/09/2010 has S188T. Similarly, two of the October isolates, A/Kentucky/08/2010 and A/Wisconsin/10/2010, have S188T, while the third October isolate, A/New Hampshire/03/2010, has S186P.

The emergence of these two sub-clades is supported by the most recent isolates from Russia and Japan, which have S188T. The Russia isolate, A/Moscow/IIV-33/2010, is from October, while the recently release Japan isolate (by NIID at GISAID), A/NIIGATA/1581/2010, is from December.

Thus, there are seventeen December isolates from the UK, Iran, and Japan and fifteen have S188T and the other two have S186P.
These data indicate these two sub-clades will dominate in 2011, and S188T may be fixed.

A recent isolate, A/India/5103/2010 had both markers and was designated a “low reactor” by the CDC, signaling immunological escape, which is a prerequisite for an emerging and dominating sub-clade.

[dothedd]

2011 Mongolia H1N1 Sequence Has S188T
Recombinomics Commentary 15:00
January 20, 2011


The first published 2011 H1N1 sequence has been released at Genbank by the National Center of Communicable Disease in Ulaanbaatar, Mongolia. The HA sequence, A/Ulaanbaatar/190/2011, was collected on January 11, 2011 and has S188T.

This sequence is most closely related to the sequence from Iran, A/Karaj/5327/2010, and supports the fixing of the S188T sub-clade in 2011. It was present in all ten December, 2010 sequences from the United Kingdom, and in four of six December sequences from Iran (the other two had S186P). Similarly, it was in the only December sequence from Japan. This fixing of S188T was also seen in November sequences from the UK, where 10/17 sequences had S188T (6 others had S186P) and the US, where all October and November sequences had S188T or S186P. Moreover, the most recent sequence from Russia (collected in October) also had S188T.

This sub-clade was likely responsible for the pressure on ICU beds in the UK, as well as the over-subscription for ECMO machines in the UK, leading to export ofd patients to neighboring countries for treatment.

The presence of S188T in the most recent sequences from Mongolia, Iran, Japan, United Kingdom, United States, and Russia indicates the current season will produce similar pressure on health care delivery in many countries in the northern hemisphere.

The dominance of this sub-clade provides strong evidence fro immunological escape and raises serious questions about the H1N1 vaccine, which targets A/Califonia/7/2009, an isolate from the spring of 2009.

Release of December 2010 and January, 2011 sequences would be useful.

Mongolia is to be commended for its prompt release of these important sequences.

[dothedd]

H1N1 Now Killing Elderly In UK
Recombinomics Commentary 19:00
January 20, 2011

The HPA week 3 report on influenza in the UK shows a dramatic jump in fatal cases (to 254 from 112), which has more credibility than earlier reports, which were trailing the numbers reported by the media (and the media reports represent a small fraction of the total number of deaths). Agencies are now withholding the number of fatal cases, so the actual number of deaths remains murky. Media reports describe young patients who have flu-like symptoms, which then develops into pneumonia and death, but cases are said to have died from “natural causes”, suggesting the number of flu deaths is markedly higher than the deaths withheld by hospital administrators.

Moreover, the latest HPA report has a dramatic jump in the deaths of the elderly (>64) to 55 from 16 reported through week 2. The number of elderly deaths is now markedly higher than the number of influenza B deaths (16), indicating most of the elderly deaths are now due to H1N1.

In the past, the elderly were generally spared from H1N1 because of seasonal H1N1 infections when they were much younger. Consequently, more than 90% of H1N1 fatalities have been under 65 and initial UK reports indicated that all H1N1 deaths this season were under 65. This recently reported increase in deaths in the elderly population could signal a dramatic change in H1N1 deaths.


Usually deaths in the elderly are not lab confirmed. The patients
are frequently in long term care facilities and die at the facility.
No samples are collected and the number of deaths is extrapolated from death certificates and descriptions of flu-like conditions or pneumonia. Thus, the current number of elderly deaths may also be markedly higher than then 55 lab confirmed cases in the week 3 HPA report. The report includes a graph of deaths in the UK, which is well above expected levels.


The latest sequence data on recent H1N1 isolates in the UK as well as other countries in the northern hemisphere (Mongolia, Iran, Japan, Russia, United States) documents the emergence of a new sub-clade with S188T. The dominance of this sub-clade indicates it is escaping immunological defenses, and this escape may now include responses to earlier seasonal H1N1 infections by the elderly population.

More information on the H1N1 isolated from elderly patients would be useful.

[dothedd]

Henry L Niman, PhD


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Jan20
www.recombinomics.com/whats_new.html

[dothedd]

H1N1 Pennsylvania Deaths Raise Concerns
Recombinomics Commentary 14:30
January 26, 2011

One flu death was reported during week 03, altogether 18 flu deaths have been reported since October of 2010.

The above comments are from the Pennsylvania week 3 report, which confirm a large jump in reported deaths for week 2. No week 2 report was published, but 11 deaths were listed as the season total in the week 1 report, which was leading the country. An alert for Montgomery County had been issued due to three recent deaths, and the 70 hospitalized cases in week 1 represented a dramatic jump from the 23 reported in the prior week. However, the graph of hospitalized cases was not updated with cases in week 2 or week 3, raising concerns that there has been a spike in such cases due to H1N1 in eastern Pennsylvania.

Earlier an alert was issued for Lehigh County due to an H1N1 outbreak that led to two reported H1N1 deaths. The CDC released sequences from these two cases (A/Pennsylvania/17/2010 and A/Pennsylvania/16/2010), 45M and 22M respectively. Both sequences were identical and were closely related to sequences from the UK which had S186P. This week a third sequence, A/Pennsylvania/34/2010, was released at GISAID by the CDC, which was identical to the above two sequence. It was also from a 45M and collected on November 16, 2010, so it is unclear if this third sequence is from a third case, or a second isolate from the earlier index case.

Additional sequences from the CDC on isolates from October and November were almost exclusively from isolates with S186P or S188T, raising concerns that these sub-clades will lead to a spike in hospitalizations and deaths in the upcoming weeks.

An update on the hospitalization in weeks 2 and 3, as well as a clarification on the serotypes and deaths recorded for week 2 would be useful.

[dothedd]

Fixing H1N1 S188T In 2011
Recombinomics Commentary 17:30
January 26, 2011

Recently released European H1N1 sequences by Mill Hill and United States H1N1 sequences by the CDC provide additional support for the fixing of the S188T sub-clade in the northern hemisphere in 2011. Earlier data provided a trend for S188T in the UK, Iran, and Mongolia. The Mill Hill sequences provided more support for Iran and the UK, but also added Luxembourg, La Reunion, and Spain to the list of countries with a dominant or expanding S188T sub-clade in December isolates. Although the CDC has not released December isolates, a recent series collected in mid or late November was heavily populated by the S188T sub-clade.

All of the December UK sequences had S188T, raising concerns that this emerging sub-clade easily escapes from immune recognition generated by prior H1N1 infections or vaccinations. This escape as supported by the low reactor status of an isolate from India that was wild type at positions 156-159, but had S188T and S186P. Similarly, a Kenya sequence, A/Kenya/2708/2010, released by the CDC had a mixed signal at position 158, but also had S188T and was designated a low reactor.

The fixing of S188T raises concerns that increasing frequencies of S188T will lead to an increase in severe and fatal cases as was seen in the UK. Increases in H1N1 infections are being reported in Europe and the United States, as hospitalizations and deaths begin to increase.

It is likely that S188T levels will continue to increase in these regions, leading to significant jumps in severe and fatal H1N1 cases.

[dothedd]

H1N1 S188T Low Reactors In Germany
Recombinomics Commentary 18:00
February 1, 2011


Mill Hill has released another series of recent H1N1 sequences at GISAID. Included were six sequences from three locations in Germany (Nordrhein-Westfalen, Berlin,Baden-Wurttemberg). All six have S188T (see updated list here), but represent different sub-clades with the S188T sub-clade. Remarkably, 5 of the 6 have one or more changes at positions 156, 157, or 158.

This region produces low reactors, which are characterized by a drop in titer of four fold or more. Two of the first designated low reactors in 2009 were from Germany, and both had G158E. One was designated as a low reactor by Mill Hill and the other was designated as a low reactor by the CDC. Although subsequent isolates from the US which had G158E were not designated as low reactors by the CDC, all US low reactors had a change at position 159. Similarly, an isolate from Kenya, which was a mixture at position 158 (G158E plus wild type), also had S188T and was designated as a low reactor, which was true for an isolate from India with S188T and S186P.

Although Mill Hill has not released antigenic characterization data on these recent (all were collected in late December, 2010) isolates, the presence of changes at positions 156, 157, and/or 158, coupled with S188T will almost certainly produce a low reactor designation.

The fixing of S188T (as seen in the above list, S188T is also in recent sequences from Madagascar, Egypt, Latvia) strongly suggests acquisition of S188T facilitates the evasion of prior immune responses to pandemic H1N1 infections or vaccinations.

The presence of these additional changes at positions 156, 157, or 158 in 5 of 6 recent isolates from Germany incates H1N1 is rapidly evolving away from the 2009 immune responses and raises concerns about the continued use of California/07/2009 as an H1N1 vaccine target.

[dothedd]

H1N1 Dominance In Pennsylvania Raises Concerns
Recombinomics Commentary 16:00
February 2, 2011


Of the 16 influenza specimens sutyped (sic) at the state lab, 5 (31%) were the 2009 A/H1N1, 2 (12%) A/H3, 7 (44%) were influenza type A and 2 (13%) were unsutypeable (sic) influenza type A.

The above comments are from the week 4 report from Pennsylvania (PA) and indicate H1N1 has become dominant in the state. The dominance appears to be widespread. Initially, most of the reported H1N1 was in eastern PA. An alert was issued for Lehigh County, where two previously healthy young adults died from H1N1. That was followed by an alert for Montgomery County, where 3 young patients died over a short period at the end of 2010 / beginning of 2011. However, in the latest report, the largest number of reported H1N1 cases is in Allegheny County (Pittsburgh) in western PA.

Moreover, the most affected age group is now 25-49, the prime target for H1N1. The CDC has released three sets of H1N1 sequences from PA and all three were identical. The demographic data matched the outbreak in Lehigh County and the November sequences had S186P. Sequences from other states indicated H1N1 was largely the two sub-clades (with S186P or S188T) dominant in England in November, when a serious outbreak began. The December UK sequences were exclusively the S188T sub-clade, and it is likely that similar dominance is in the United States, although no December / January sequences have been released by the CDC,

The S188T sub-clade in England produced a high rate of severe and fatal cases, which filled ICU beds and put a severe strain on available ECMO machines. The data in PA suggests that a similar scenario will follow in the US. In the UK, H1N1 was the dominant subtype. H3N2 was rare and increases in influenza B have begun recently. In PA, H3N2 was dominant in the fall of 2010, but now H1N1 is on the rise. It is also the dominant serotype in adjacent New Jersey, where the P&I deaths have spiked to 8.53%. A similar trend is likely throughout the US.

In PA, the number of deaths is up to 21, which is almost certainly the highest in the country. Hospitalizations also began to spike in early 2011. The week 1 report had 70 cases, up from 23 reported for week 52 and 11 in week 51. PA did not issue a week 2 report, and hospitalization numbers for week 3 and 4 have not been reported, although they are likely to be markedly higher because the number of reported deaths has almost doubled from 11 in week 1 to 21 by week 4.

Thus, it is likely that the number of severe and fatal H1N1 cases will spike higher in the US in the coming weeks. Release of H1N1 sequences from December and January would be useful.

[dothedd]

H5N1 S227R In 2011 Whooper Swan In Hokkaido Japan
Recombinomics Commentary 13:10
February 3, 2011

The recent series of H5N1 in poultry and wild birds in Japan (see OIE reports here) has raised concerns. Clade 2.3.2 (Fujian strain) had been circulating in wild birds in southern China for several years, but in 2008 it appeared in wild birds in Japan and poultry in South Korea. It was the likely cause of a human cases in South Korea and subsequently appeared in wild birds and poultry in Primorsky.

The presence in wild birds led to the prediction that clade 2.3.2 would migrate to Mongolia and Russia, which was confirmed in 2009 isolates, which led to the prediction that clade 2.3.2 would migrate to Europe, the Middle East, and Africa, as predicted and confirmed for the Qinghai strain (clade 2.2).


A 2011 H5N1 sequence, A/whooper swan/Hokkaido/4/2011, has been released by Hokkaido University at Genbank. The sequence is clade 2.3.2, as expected, but it has the receptor binding domain change S227R.


In 2005, the acquisition of S227N by clade 2.2 was predicted, based recombination between donor sequences in H9N2 in the Middle East and Qinghai H5N1. The ability of H5N1 with S227N to bind to human receptors led to the prediction of human cases in the Middle East, which was confirmed in the sequence from the index cases in Turkey in 2006. S227N was in two of the four human sequences in Turkey and appeared in subsequent human H5N1 sequences in Egypt.

Moreover, additional receptor binding domain changes, V223I and M230I were found in human sequences from the Gharbya cluster.

The presence of V223I and M230I in clade 2.3.2 Fujian sequences raised concerns about human transmission.

The detection of S277R in the recent H5N1 sequence from Japan increases those concerns.

[dothedd]

Love is a mystery - Ludovico Einaudi

[dothedd]

US Pneumonia and Influenza Deaths Spike to 8.45%
Recombinomics Commentary 13:10
February 3, 2011

Tomorrow’s week 4 FluView by the US CDC will show a dramatic spike in the number of pneumonia and influenza (P&I) deaths the United States, based on tomorrow’s MMWR. The rate of 8.45% is a dramatic increase from the 7.49% reported for week 3 and is well above the epidemic threshold.


This spike is likely due to the increases in H1N1, which is now becoming dominant in multiple states. The most recent reports from Pennsylvania and New Jersey indicate H1N1 is now more common than H3N2, and that trend is likely to spread across the US.


The CDC has not released any December or January H1N1 sequences, but the vast majority of sequences from October and November had S188T or S186P, which were dominant in the UK in October and November also. UK experience a large number of severe and fatal H1N1 cases in December and January. Similarly, Canada just released six HA sequences from Ontario collected in late 2010 and all six had S188T or S186P.


Tomorrow’s FluView will also report 6 pediatric deaths including two in New Jersey. This spike is also likely linked to increases in H1N1.

Release of December and January H1N1 sequences in the United States would be useful.

[dothedd]

H1N1 In United States Identical to United Kingdom
Recombinomics Commentary 17:10
February 4, 2011


The CDC has released a series of sequences (at GISAID) from recently collected isolates, which extends the trend generated by the prior sets of sequences from October and November. Most of the latest sequences were collected in December 2010 or January 2011, and all sequences collected after December 10 had either S188T or S186P, with the vast majority carrying S188T. The emergence was similar to the H1N1 seen in the UK, which had an unusually high frequency of severe and fatal cases.

In fact many of the H1N1 sequences in the United States were identical to the sequences from the United Kingdom. Sequences from A/Kansas/08/2010, A/Kentucky/08/2010, A/Delaware/05/2010, A/Indiana/09/2010, and A/Oregon/04/2010 were identical to each other and sequences from A/England/4920303/2010, A/England/119/2010, and A/England/142/2010. Moreover, sequences from A/Connecticut/0/2010, A/Wisconsin/09/2010, A/Maine/03/2010, A/Missouri/01/2010, A/Alaska/02/2011, A/Kentucky/16/2010, and A/Rhode Island/03/2010 were all closely related to the identical sequences above.

These sequences represent infections covering a large portion of the United States, from Alaska to Missouri to Maine. Other sequences had similar relationships with sequences in the United Kingdom, suggesting that the problems in December and January in the UK would soon develop into similar problems in the US.

Recent state reports in the US indicated H1N1 had become the dominant serotype, and the week four pneumonia and influenza deaths for week 4 spike to 8.45% nationally, which is well above the epidemic threshold. Similarly, the number of pediatric deaths jumped to six, even though this is a lagging indicator.

The national week 4 report is likely to extend the above observations and signal increases in serious cases across the country. The dominance of S188T and S186P is likely due to immunological escape and vaccine breakthrough. The WHO regional center in Australia also released recent sequences which were also dominated by S188T and S186P, even though the 2010 flu season has ended for the southern hemisphere. Two of the sequences were from young adults (20M and 21M) who had received the 2010 vaccine and were part of a cluster. Both sequences (which had S186P) from this cluster were identical and provided additional evidence for vaccine escape.

Such escapes are not unexpected. However, the vaccine used in 2010 in the southern and northern hemisphere targeted California/7/2009, which was isolated in April 2009, so escape in 2010 and 2011 was not unexpected. Most remarkably, this outdated target is being used for the 2011 vaccine for the southern hemisphere, which is scandalous.

[dothedd]

Another trH3N2 Case In Pennsylvania Raises Concerns
Recombinomics Commentary 19:10
February 4, 2011


One case of human infection with a novel influenza A virus was reported by the Pennsylvania Department of Health. The patient was infected with a swine origin influenza A (H3N2) virus. The patient reported contact with pigs in the week preceding symptom onset on September 6, 2010,

Initial testing of the specimen indicated a seasonal influenza A (H3N2) virus and the specimen was submitted to CDC as a routine surveillance sample. The delay from onset to detection occurred because attempts to culture the virus were unsuccessful. RT-PCR testing confirmed swine-origin influenza A (H3N2).

The above comments from today’s week 4 FluView describe a second trH3N2 case in Pennsylvania. The prior case was initially announced with a pager alert for two cases, one from Pennsylvania and an earlier case from Illinois. The CDC subsequently indicated the two cases were from Wisconsin and Pennsylvania. The Wisconsin case developed symptoms two after the unannounced case and six weeks prior to the first announced Pennsylvania case. There has been no sequence released, but it is trH3N2 based on PCR data.

Today’s announcement confirms that trH3N2 cases can type as seasonal H3N2, which is not surprising because trH3N2 has human H3 and human N2. However, these sequences are from infections in the mid-1990’s and the virus has been evolving in swine, so the antigenic drift from contemporary seasonal H3N2 is significant. This drift could lead to an “unsubtypable” result of the condition of the sample is less than ideal. The above comments indicate the latest announced case in Pennsylvania did not culture, raising concerns that the large number of “unsubtypable” isolates reported by Pennsylvania in late 2010 and early 2011 may represent more trH3N2 cases that have not been fully characterized. Pennsylvania leads the nation in reported flu deaths, and several have been reported as H3N2.

The CDC has yet to address the number of trH3N2 under investigation. The close sequence related between the human trH3N2 as well as the presence of PB1 E618D in all recent human cases and its absence in all swine trH3N2 isolates indicates the trH3N2 is transmitting human to human.

The latest case is being reported in 2011, but really represents the four trH3N2 case in the fall of 2010, which is an unprecedented frequency. A pager alert was issued when there were two cases. The latest results indicate there were at least three at the time of the alert, and it remains unclear why one of the cases in the pager alert was said to be from Illinois.

More detail on these cases is long overdue.

[dothedd]

trH3N2 Transmission Raises Pandemic Concerns
Recombinomics Commentary 19:20
February 5, 2011

One case of human infection with a novel influenza A virus was reported by the Pennsylvania Department of Health. The patient was infected with a swine origin influenza A (H3N2) virus. The patient reported contact with pigs in the week preceding symptom onset on September 6, 2010, did not require hospitalization, and has since fully recovered. Initial testing of the specimen indicated a seasonal influenza A (H3N2) virus and the specimen was submitted to CDC as a routine surveillance sample.
The delay from onset to detection occurred because attempts to culture the virus were unsuccessful. RT-PCR testing confirmed swine-origin influenza A (H3N2). Six other human infections with swine origin influenza A (H3N2) viruses have been identified in the United States during 2009 through 2010, including one other case from Pennsylvania in week 44 of 2010.

The above comments from the week 4 FluView report raise concerns that trH3N2 is transmitting human to human (H2H) at a level markedly higher that the reported cases mentioned above. In November, 2010 WHO issued a pager alert describing two novel influenza isolates identified in the United States. Both were H3N2 triple reasortants (trH3N2) and were of concern because two such isolates identified over a short time frame was unusual. The first human trH3N2 in the United States was in Kansas in the summer of 2009. Prior to the alert there had been three trH3N2 cases (in Kansas and Iowa in 2009 and in Minnesota in 2010), so two cases in the fall of 2010 raised concerns. The latest report from the CDC indicates there were at least three cases in this time frame, and two of the three were from Pennsylvania.

The above comments confirm that trH3N2 sequences are recognized by current H3 sub-typing reagents, which was expected because the H3 and N2 in swine trH3N2 have a human origin. However, the genes trace back to the mid-1990’s and have been evolving is swine, so there has been significant genetic drift, which could lead to an “unsubtypable” designation if the clinical sample is compromised. Others, like the sample above would simply type as seasonal H3N2, and the swine origin would not be obvious in the absence of more detailed testing, such as antigenic characterization tests, where isolates would register as “low reactors”, or by sequencing, where the relatedness to seasonal H3N2 from the 1990’s would be clear.

Thus, detection of trH3N2 requires more testing than a simple influenza A test or human sub-typing, raising concerns that the number of trH3N2 isolates may be markedly higher than the seven cases reported to date. Moreover, the Kansas sequence, as well as all four 2010 sequences have PB1 E618D. This marker is of concern because it is in virtually all pH1N1 sequences and is not in any swine trH3N2 sequences, suggesting it represents an adaptation to a human host, and signals H2H transmission. Several of the trH3N2 cases do not have a history of contact with swine. Moreover, for cases with swine contact, there has been no demonstration that the swine were H3N2 infected or that H3N2 sequences in the contact swine were closely related to the human trH3N2.

Moreover, as noted, there have been no reports of PB1 E618D in any swine trH3N2, even though it is in 5 of the 6 human trH3N2 sequences.

Therefore more information on additional samples under investigation as well as an explanation for the large number of “unsubtypable” cases in Pennsylvania, which have been noted in weekly reports in 2010 and 2011, would be useful.

[dothedd]

Casual trH3N2 Transmission In Pennsylvania
Recombinomics Commentary 14:20
February 6, 2011


No contact with pigs has been identified in the Pennsylvania case in the week before symptom onset on October 24, 2010; however the case lives in an area close to pig farms.

The above comment is from the CDC week 44 FluView describing one of the new cases associated with the WHO pager altert on novel trH3N2 in the United States. As noted above the Pennsylvania case denied contact with swine prior to development of symptoms on October 24, 2010. The USDA has released two HA sequences from swine isolates in Pennsylvania collected on October 25, 2010. The two swine sequences, A/swine/Pennsylvania/62170-1/2010 and A/swine/Pennsylvania/62170-3/2010 were identical to each other and virtually identical to three sequences (original and virus grown in mammalian cells or eggs), A/Pennsylvania/14/2010, collected from the above patient (45M). These Pennsylvania swine sequences are much closer to the Pennsylvania patient than other trH3N2 sequences from swine or patients, clearly demonstrating a close association between the Pennsylvania swine and human sequences, signaling transmission.

However, the denial of swine contact by the patient leaves open the direction of the transmission, as well as the route. The PB1 from the human Pennsylvania isolate had E618D, which has not been seen in any prior swine H3N2 sequences, but is in the first human trH3N2 sequence reported in the US, Kansas/13/2009, as well as all 2010 human trH3N2 sequences (A/Minnesota/09/2010, A/Wisconsin/12/2010, A/Minnesota/11/2010) indicating it is a recent acquisition and may represent a human adaptation since it is virtually all pH1N1 sequences (human or swine). Its presence or absence in the Pennsylvania swine is not public, but it is likely present, regardless of the direction or route of transmission because once the marker appears in human cases, transmission back to swine is likely, as was seen for pH1N1 which has now been identified in swine worldwide and the recent swine H1 isolates have PB1 E618D.

Thus, the direction of the transmission remains unclear, but the denial of contact with swine suggests transmission is casual and may involve additional human cases linking the reported Pennsylvania case to the Pennsylvania swine, which were presumably in the nearby pig farms described above.

This casual transmission suggests that swine trH3N2 is far more common than the seven cases reported to date, and the additional cases involve human to human transmission, which would not be unexpected sine the H3 and N2 trace back to seasonal H3N2 circulation in the 1990’s. As noted in the week 4 FluView, the second trH3N2 case in Pennsylvania was initially classified as seasonal H3N2 and was changes to swine trH3N2 base on PCR analysis during routine characterization, which was reported 5 months after the fact (the second Pennsylvania case developed symptoms on September 6, 2010, but was not reported until February, 2011 because of a failure to grow the virus).

The number of trH3N2 cases in Pennsylvania is also unclear because of the large number of cases classified as seasonal H3N2 as well as the large number of isolates classified as unsubtypable.

More information on trH3N2 cases under investigation, as well as release of a partial sequence from the second reported case in Pennsylvania as well as the locations of the two Pennsylvania cases would be useful.

[dothedd]

H1N1 S188T Homolgous Recombination In Beijing China
Recombinomics Commentary 19:00
February 7, 2011


Beijing Institute of Microbiology and Epidemiology has released a series of sequences at Genbank from November, 2010 isolates, including five full or partial HA sequences, Each of the three full sequences, A/Beijing/3872/2010, A/Beijing/3856/2010, and A/Beijing/3884/2010 had S188T. Although the two partial sequences, A/Beijing/3848/2010 and A/Beijing/3907/2010, did not cover the region encoding S188T, the vast majority of S188T sub-clade sequences also have T1056C, G1171A (E377K), and G1403A (S454N) and both partial sequences had all three markers. Moreover, the three full sequences had additional markers in this region, C1437T and C1656T which have also been seen in additional S188T sequences and both partial sequences had these two markers also, indicating all five HA sequences were from the S188T sub-clade which had clonally expanded in Beijing in November, 2010. The dominance of this sub-clade in late 2010 / early 2011 was widespread throughout they northern hemisphere as well as in off-season isolates from the southern hemisphere collected in this time frame.

However, one of the five sequences described above, A/Beijing/3872/2010, was missing two consecutive markers, T1056C and G1171A which are found in virtually all S188T sub-clades. This isolate had also gained two markers, C897A and A1215G located in the same region. These two markers were present in other pH1N1 sequences (see list here and here), including A/Beijing/22811/2009, which has C897A and was wild type at positions 1056 and 1171.

Moreover, A/Beijing/3872/2010 has the upstream markers, T72C, C144T, G366A, A478G (S146G), G605C (S188T), G640A (A200T). which are present in the other two November sequences. Thus, Bejing/3872 has the nine common markers flanking the short region (between positions 897 and 1215), were there are four changes (the loss of T1056C and G1171A coupled with the gain of C897A and A1215G).

These concentrated changes are most easily explained by homologous recombination because three of the four changes are present on an earlier Beijing sequence and the rare fourth change is present on two H1N1 sequences which are also wild type at positions 1056 and 1171 and therefore each also has three of the four changes (and the small size of the Beijing sequence database suggests that a sequence with all four changes already exists) could be acquired via a single recombination event producing the acquisition between positions 897 and 1215.

Thus, the five Beijing sequence represent clonal expansion of an evolved S188T sub-type, and A/Beijing/3872/2010 is an example of additional rapid evolution via homologous recombination.

[dothedd]

Pennsylvania Unsubtypables Raise trH3N2 Pandemic Concerns
Recombinomics Commentary 23:55
February 8, 2011


Of the 121 influenza specimens sutyped (sic) at the state lab, 26 (21%) were the 2009 A/H1N1, 24 (28%)(sic) A/H3, 34 (28%) were untyped influenza type A and 8 (7%) influenza type B.

The above comment from the week 5 report from Pennsylvania suggests that the number of unsubtypable samples has spiked higher. The numbers listed above are far from clear because only 92 of the 121 subtyped are categorized, and it is unclear if the term "untyped" refers to those not typed or those unsubtypable. However, the pie chart included in the report list 21 samples as unsutypable (sic), in addition to 176 samples that were influenza A but not typed.


The reason for the large number of unsubtypable samples, as well as the precise definition used in Pennsylvania, remain unclear, but Pennsylvania has also reported two trH3N2 cases, infected in September and October, so the increasing numbers of unsubtypable samples may reflect increased transmission of trH3N2.

The sequences of the 2010 trH3N2 isolates (from MN, WI, PA) clearly supports human to human transmission, and the close sequence similarity between the October isolate, Pennsylvania/14/2010, and two Pennsylvania swine sequences from samples collected a day after disease onset, indicate these sequences are also circulating in swine, which appear to be casually transmitted to humans, since the October PA case has denied contact with swine.

More detail on the increasing numbers of unsubtypable samples, which have been reported in multiple weekly reports in late 2010 and early 2011 in Pennsylvania, as well increasing numbers in other states, is long overdue.

Click LINK to view CHART:
www.recombinomics.com/News/02081101/trH3N2_PA_Unsubtypables.html[/b]

[dothedd]

CDC Silence On Unsubtypables Raises trH3N2 Pandemic Concerns
Recombinomics Commentary 14:40
February 9, 2011


The Pennsylvania week 5 report demonstrates a large spike in unsubtypable cases, which follows earlier 2010 and 2011 reports on smaller numbers of such cases. Unsubtypable cases have also been reported in North Carolina, Connecticut, and California, raising concerns that these cases represent a growing problem which may be linked to the spread of trH3N2. The largest number of unsubtypable cases have been reported in Pennsylvania, which has also reported two trH3N2 cases in the fall of 2010. The most recent case, who developed symptoms on September 6, 2010 was not reported until last week by the CDC in its MMWR and FluView. The five month delay was attributed to a failure to grow the virus and an initial characterization of the sample as seasonal H3N2. Typing trH3N2 as seasonal H3N2 has been a concern, since trH3N2 has human H3 and N2 that was circulating in the 1990’s and as been evolving in swine since it jumped species. Therefore, samples with high levels of virus will sub-type as seasonal H3N2, while samples with low levels may test positive in an influenza A test, but negative on an subtyping test because of the degree of genetic divergence from contemporary seasonal H3N2.

Samples that were influenza A positive but unsubtypable were common in the spring and summer of 2009, when pandemic H1N1 was emerging. Such samples would be positive for influenza A, but negative for seasonal H1n1 because pandemic H1N1 was of swine origin and the swine H1 failed to react with subtyping reagents directed against human sequences. These samples were sent to the CDC for testing, and reagents targeting swine H1 indicated the virus was of swine origin, which was confirmed by sequencing isolated virus. The large number of cases created a backlog and initially states did not have the reagents to subtype the swine isolates, so such samples were classified as unsubtypable samples that were influenza A positive. The CDC subsequently distributed the appropriate test kits for swine H1N1 and subtyping was and is done at the state level, although the test were initially rationed, and states typically only subtype a fraction of the influenza A positive cases.

Cases that are influenza A positive but unsubtypable are not limited to trH3N2. The same result would be generated by other novel influenza such as H7, which has been reported previous in the United States and Canada (as well as Europe), or H5 which has been found in Asia, the Middle East, or Africa. However, all recent novel influenza cases in the US have been trH3N2, and two of the seven cases have been in Pennsylvania.

Although the CDC has reported the seven trH3N2 cases, it has not addressed the unsubtypable cases reported by states, as seen in the graph below, or in the table with the underlying data.



Although there is an unsubtypable column, none of the cases reported by the various states are listed, and the trH3N2 cases reported to date have been typable as seasonal H3N2. However, as the levels of seasonal H3n2 decline, the conditions favoring the emergence of trH3N2 would increase, which might explain the recent increases in unsubtypable cases, especially in Pennsylvania where there have been two confirmed human cases as well as two confirmed swine cases which were closely related to the human case in October. However, the human case in October has denied contact with swine, increasing concerns that the trH3N2 in Pennsylvania is casually transmitting human to human, which is supported by the sequence data from the four cases reported from 2010 (two cases in MN, one in WI, and one in PA – there is no public sequence data from the second case in PA).

The failure of the CDC to acknowledge and address the unsubtypable cases previously reported by multiple states increases pandemic concerns.

Details on these cases are long overdue.

LINK TO CHART:
www.recombinomics.com/News/02091101/trH3N2_CDC_Unsubtypables.html

[dothedd]

H1N1 RBD Changes S188R and D190Y in Costa Rica Swine
Recombinomics Commentary 16:40
February 10, 2011

The USDA has released six HA sequences from Costa Rica swine collected in November, 2010. These sequences are closely related to each other, and quite distinct from the pH1N1 sub-clades dominant in November/December human isolates in the United States and Canada., which have S186P or S188T.

One of the isolates, A/swine/Costa Rica/000125-15/2010, has a cluster of RBD changes (H184D, S188R, D190Y, Q192H, D199Y). Two of these changes (H184D and D190Y) are also in another isolate, A/swine/Costa Rica/000125-3/2010.

Other non-synonymous changes in this series are N34D, K57R, K122N, P162L, S165I, E238K, K286R, A318V, I375V, R549K. Most of these changes are in most of these Costa Rica swine isolates.

This genetic diversity can serve as a reservoir for rapid change in pH1N1 in humans. This virus can readily jump back to humans from swine to introduce these changes into the human population. Most of these changes are already circulating in human isolates (see list here here here).

D190Y has been seen previous in a 2009 sequence in Brazil, A/Brazil/7450/2009, as well as a recent isolate in England, A/England/4500186/2010.

The large number of changes flanking this change is similar to changes associated with the fixing of H274Y in 2009. Virtually all seasonal H1N1 isolates emerging in the fall of 2009 had A193T which was coupled to one or more changes at positions 187, 189, or 196. These changes allowed for immunological escape and rapid spread throughout the northern hemisphere, including countries where the Brisbane/59 sub-clade was circulating in the prior season.

Recently released pH1N1 sequences from the northern hemisphere are sioalted by sub-clades with S186P or S188T, with the latter becoming more dominant. Acquisitions of additional changes via recombination will likely increase spread and virulence.

H1N1 is now becoming dominant in many countries in the northern hemisphere as levels of H3N2 decline. These changes will lead to increases in deaths in pediatric and young adult cases and increase pandemic concerns.

[dothedd]

Pennsylvania trH3N2 Swine Have PB1 E618D
Recombinomics Commentary 15:50
February 11, 2011


The USDA has released sequences from the six internal genes from the Pennsylvania trH3N2 swine, A/swine/Pennsylvania/62170-1/2010 and A/swine/Pennsylvania/62170-3/2010 as well as two additional isolates, A/swine/Pennsylvania/62170-2/2010 and A/swine/Pennsylvania/62170-4/2010, which are closely related to each other and the human trH3N2 from Pennsylvania, A/Pennsylvania/14/2010, as expected from the previously released HA sequences.

Moreover, all four swine PB1 sequences have E618D, representing the first reported swine H3N2 isolates with this change. The change is in 5 of the 6 public trH3N2 human sequences, including the four isolated in 2010. It is a recent change and all swine or human trH3N2 isolates with this change have been isolate after the start of the 2009 pandemic. Virtually all pH1N1 PB1 sequences also have E618D, suggesting it is a change linked to human adaptation of the swine pH1N1.

The presence of this change in the human trH3N2 sequences have raised concerns that it was acquired by recombination, and further acquisitions may increase the transmissibility of trH3N2 in humans.

This concern was increased by the recently released sequences from swine H1N2 sequences in South Dakota which have NA sequences very closely related to the human trH3N2 NA sequence from Pennsylvania (the Pennsylvania swine NA sequences have not been released), and have a pH1N1 MP gene segment. This reassortant demonstrates the exchange of trH1N1 and trH3N2 genes in swine. The presence of PB1 E618D in the South Dakota swine sequences is not known because only the HA, NA, and MP sequences were released.

Although the Pennsylvania swine PB1 sequences have E618D, the sequences are heterogeneous, indicating the trH3N2 has been circulating in the swine population, although the human case denies swine contact.

The presence of E618D as well as relatedness to the NA sequences in the reassortants in South Dakota continue to increase concerns for rapid evolution of tr genes in swine that can re-infect humans and produce more disease with increased severity.

[dothedd]

South Dakota trH1N2 Swine Reassortants Raise Concerns
Recombinomics Commentary 15:50
February 11, 2011


South Dakota State University has released HA, NA, and MP sequences from three recent H1N2 swine reassortants (collected in November and December, 28, 2010). The sequences (A/swine/South Dakota/2/2010, A/swine/South Dakota/3/2010, and A/swine/South Dakota/4/2010) are related to each other, and the NA sequences are closely related to the human trH3N2 from Pennsylvania, A/Pennsylvania/14/2010.

However, the MP sequences are related to pH1N1 sequences indicating the isolates are reassortants and may have the constellation seen in the H1N2 reassortant in Argentina, which had 6 internal genes from pH1N1.

Like the Argentina isolate, all three isolates in South Dakota have human external genes from seasonal H1N2 circulating in 2003. Thus, all known genes can be traced to humans and therefore may be good candidates for human transmission. Indeed the NA genes are almost identical to the NA gene in the human trH3N2 isolate from Pennsylvania, A/Pennsylvania/14/2010, which has PB1 E618D, which is also found in virtually all pH1N1 isolates.

Thus, the three isolates in South Dakota represent additional isolates which may jump to humans, creating swine origin trH1N2, in addition to trH3N2 seen five times in humans in 2010 (and twice in 2009).

Release of sequences from the other five gene segments would be useful. It is likely that all five will also be from pH1N1.

[dothedd]

Recombination Drives Rapid H1N1 Evolution in Costa Rica Swine
Recombinomics Commentary 20:20
February 14, 2011

The USDA recently released six sets of sequences (HA, NA, MP) from swine in Costa Rica. As noted previously, the HA sequences had a large number of synonymous and non-synomous changes, including receptor binding domain changes. However, phylogenetic analysis provided compelling evidence support rapid evolution via homologous recombination between isolates co-circulating in the swine herd. This evolution was most dramatic in isolate A/swine/Costa Rica/000125-3/2010. The HA sequence had a large number of synonymous changes present in all six sequences (G291A, A453T, A708G, G754A, T1023C, A1165A). Similarly, all six sequences had a large series of non-synonymous – A142G (N34D), A212G (K57R), G408T (K122N), G805A (E238K), A899G (K286R), C995T (A318V), C1140T (I375V) which define a rapidly evolving sub-clade that had spread by clonal expansion.

However, these isolates had a number of additional changes shared by a subset of these sequences.

As noted previously, A/swine/Costa Rica/000125-3/2010, has two of the receptor binding domain changes, C592G (H184D) and G610T (D190Y), shared with A/swine/Costa Rica/000125-15/2010, which could be explained by further clonal expansion. However, additional changes which were shared with other isolates from the herd preclude this explanation for this additional evolution. A/swine/Costa Rica/000125-3/2010 shares two other polymorphisms, C528T (P162L) and A1227T, with A/swine/Costa Rica/000125-16/2010. Moreover, another polymorphism, A1222G (T394A) is shared with A/swine/Costa Rica/000125-19/2010, while G1357A (D439N) is shared with A/swine/Costa Rica/000125-20/2010. Thus, four of the five isolates have one or two polymorphisms exclusively shared with A/swine/Costa Rica/000125-3/2010, generating significant discordance. However, discordant combinations are note solely linked to A/swine/Costa Rica/000125-3/2010 polymorphisms. Another change, C647T, is shared by three other isolates, A/swine/Costa Rica/000125-14/2010, A/swine/Costa Rica/000125-16/2010, and A/swine/Costa Rica/000125-19/2010). Thus, all six isolates have polymorphism shared by a small subset (1 or 2) of the other isolates signaling significant recombination between isolates from this herd.

In additional to the recombination in the HA sequences, the NA and MP sequences also had evidence for recombination and discordance. The NA sequence also had multiple synonymous and non-synonymous polymorphisms that were found in all six sequences. However, two additional polymorphisms were present in two different subsets, which also involved A/swine/Costa Rica/000125-3/2010. C531T is shared with A/swine/Costa Rica/000125-19/2010, while C693T (A231V) is shared with A/swine/Costa Rica/000125-14/2010 and A/swine/Costa Rica/000125-16/2010. Moreover, the MP sequence has another polymorphism, C7T, that is not in A/swine/Costa Rica/000125-3/2010, but is shared by a different combination of isolates from the herd (A/swine/Costa Rica/000125-14/2010, A/swine/Costa Rica/000125-19/2010, and A/swine/Costa Rica/000125-20/2010).
Thus, the sequences from this herd clearly demonstrate rapid evolution via recombination, leading a novel sequences in each isolate and in multiple gene segments.

Although the isolates were from swine, these were pH1n1 sequences that jumped from human to swine and such sequences can easily jump back into humans. As noted previously, one of the shared receptor binding changes, D190Y, has already been detected in two human isolates, including a fatal case, A/England/4500186/2010, while another S188R is the position associated with dominance of the S188T sub-clade.

Thus, the recombination between herd members can generate new gen sequences that can seriously impact the human flu gene pool, and such changes to continue to be cause for concern.

[dothedd]

US Pneumonia and Influenza Death Rate Spikes to 8.89%
Recombinomics Commentary 18:30
February 17, 2011


Tomorrow’s MMWR shows a pneumonia and influenza death rate of 8.89% for week 6, which is well above the epidemic threshold. This level is markedly higher than the week 4 spike of 8.45% or the week 5 rate of 8.03%. The week 6 rate is approaching the five year high of 9.1% set in week 11 in 2008. The rate is expected to rise because pandemic H1N1 became the dominant serotype in the US last week, and influenza deaths are a trailing indicator.

Earlier this season H3N2 was dominant, but levels of H3N2 have been declining, while H1N1 has been increasing. A recent report out of the UK indicated the failure rate for the H1N1 vaccine was approximately 50%, which was not a surprise because of the dominance of the sub-clade with S188T in the UK, including sequences which were identical to H1N1 isolates in the United States. This sub-clade has become dominant across the northern hemisphere, including the United States.

Many states are reporting widespread influenza activity, providing additional data supporting immunological escape from the H1N1 circulating last season, as well as the vaccine target for 2009 and the 2010/2011 season. Remarkably, this same outdated target, A/California/07/2009, was recommended by WHO today as the target for the northern hemisphere in 2011/2012.

As seen in the spike in the P&I data and the dominance of the S188T sub-clade, the WHO selection procedure for influenza vaccine targets continues to be hazardous to the world’s health.

[dothedd]

Northern Hemisphere H1N1 Vaccine Failure
Recombinomics Commentary 23:50

February 17, 2011


Unvaccinated 1014/2554 (39.7%)
Vaccinated season 2009/10 only 26/130 (20.0%)
Vaccinated season 2010/11 only 22/100 (22.0%)
Vaccinated both seasons 21/107 (19.6%)


The above data from table 3 in the Eurosurveillance report, "Effectiveness of seasonal 2010/11 and pandemic influenza A(H1N1)2009 vaccines in preventing influenza infection in the United Kingdom: mid-season analysis 2010/11", for samples positive for H1N1 between September 2010 to January 2011, demonstrate a vaccine failure rate of approximately 50%, regardless of vaccination schedule.

This high failure rate was expected based upon the reports of widespread infections, as well as the elevated levels of severe and fatal cases in the UK in this period, primarily in December and January. A vaccine failure was also anticipated due to sequence data which indicated that November cases were almost exclusively due to H1N1 with S186P or S188T, and the presence of S188T in all December sequences from the UK. This domination is largely dependent on immunological escape from the immune response generated in response to infections during the 2009/2010 season or prior vaccinations.

The results in the UK have been extended to most countries in the northern hemisphere, which have reported a similar domination of sub-clades with S186P or S188T. This escape however is poorly represented in antigenic characterization tests, which lack reproducibility and sensitivity. Unfortunately, this outdate procedure is the gold standard for vaccine target selection by the WHO and CDC. Today the WHO announced that the vaccine target for the northern hemisphere in the 2011/2012 season will again be A/California/07/2009. Although the recommendation acknowledged that some isolates produced low reactivity in some antigenic characterization tests using ferret anti-sera, the selection of the 2009 target once again demonstrates the failure of this approach.

The track record of this approach in the targeting of H1N1 has been abysmal, and has been largely driven by the poor data generated in the antigenic characterization test. For the 2007/2008 season, the vaccine target of Solomon Island/03/2006 (clade 2A) was selected. The prior target, New Caledonia/20/1999 had been circulating since 1999 and anti-sera directed against this target (designated clade 1) produced a low reaction against clade 2. However, by the fall of 2007, there was virtually no Solomon Islands (clade 2A) H1N1 circulating. It had been replaced by Brisbane/59/2007 (clade 2B) in Europe and North America, or Hong Kong/2562/2007 (clade 2C) in Asia and western North America. The Brisbane/59 was a concern because of high frequencies of Tamiflu resistance (H274Y).

However, an insensitive ferret anti-sera was used by the CDC. Since the anti-sera could not distinguish between the clade 2A, 2B, and 2C, all were said to be Solomon Island-like and a vaccine match. However, the phylogenetic analysis showed that the three sub-clades were easily distinguished and had multiple amino acid difference. Moreover, the dominance of 2B and 2C indirectly indicated there were antigenic difference which led to the dominance of 2B and 2C at the expense of 2A. These difference were confirmed in a subsequent anti-sera generated against Brisbane/59 grown in mammalian cells. This anti-sera readily distinguished 2A from 2B from 2C and led to the replacement of Solomon Island/3 with Brisbane/59 for the 2008/2009 season. However, Solomon Island was used in the summer of 2008 in the southern hemisphere, when Tamiflu resistance increased to 100%.

Moreover, the dominant sub-clade of Brisbane/59 also had receptor binding domain changes (A193T and additional changes at positions 187 and 189, allowing the virus to escape immunological responses generated against the virus in the prior season. Consequently, the Brisbane/59 that emerged not only was Tamiflu resistant with HA A193T and at least one additional change at positions 187, 189, or 196, leading to vaccine failures when Brisbane was introduced in the 2008/2009 season. Moreover Tamiflu resistance in H1N1 increased to 100% worldwide.

The problems linked to an insensitive and unreliable antigen characterization test continued when pandemic H1N1 emerged in the spring of 2009. The late appearance led to a delay in the fall vaccine, and the pandemic emerged early, so it had already peaked when the vaccine was distributed widely. The drop in activity in November signaled an immunized target population, raising questions about the vaccine effectiveness for the following year, since the target was not changed. Selection of a new target was complicated by an unreliable test.

Initially there was some agreement between testing done by the CDC and Mill Hill. Both had found a low reactor in Germany and both viruses had G158E. This result supported data from two other labs testing H1n1 under different circumstances. MedImmune had tested a clone with G158E which grew well in eggs, but was not selected as a target because anti-sera against wild type produced a dramatically reduced titer (confirming the G158E produced low reactors). Similarly H1N1 grown in the presence of neutralizing monoclonal antibodies generated low reactors which had changes in positions 156-159 including G158E.

However, subsequent isolates from the US which had G158E were not designated low reactors by the CDC, signaling intra-lab variation.

These differences were compounded when a Ukraine isolate A/Lviv/N6/2009 was characterized. Mill Hill designated it as a low reactor, which was a concern because it had D225G. However, the CDC published a sequence showing that an isolate from the same patient had D225G and G158E, yet it was still not called a low reactor by the CDC. An isolate from this patient became a lab standard and the WHO regional lab in Australia and the HPA in the UK published data using the standard and both clearly showed that it was a low reactor. Thus, the CDC data did not match its own data on G158E in Germany or the low reactor designation of the Ukraine isolate generated by three different labs. Moreover, all US isolates designated as low reactors by the CDC had a change at position 159, strongly suggesting that the assay lacked sensitivity for detection of low reactors in the US that did not have a position 159 change.

These discrepancies continued this season. Sub-clades with S188T and S186P emerged in the UK and across the northern hemisphere, indicating these changes fuel immunological escape. These sub-clades became dominant and were tested by the CDC. The testing of isolates collected from patients outside of the US providing some data signaling immunological escape associated with S188T. One isolate from Kenya had a mixed signal at position 158, but was called a low reactor, supporting a contribution by S188T. Similarly, an isolate from India did not have a change in the positions 156-159, but had S188T and S186P and was designated a low reactor. However, a US isolate, A/Kentucky/09/2009 had S188T as well as G158E and D225G, but was still not designated as a low reactor by the CDC. Moreover, the CDC has yet to identify a low reactor in the US this season, even though most isolates have S188T or S186P. This failure of the antigenic characterization test has led to a recommendation of A/California/07/2009 for the 2011/2012 season, even though H1N1 has now become the dominant serotype in the US, influenza is widespread in most of the states, and the pneumonia and influenza death rate for week 6 is 8.9, just shy of the five year high of 9.1 set in week 11 in 2008.

Thus, the CDC and WHO consultants have ignored the emergence of S188T and consistent failures of the antigenic characterization test to generate reliable or predictive data, and selected A/California/07/2009 as the pandemic H1N1 vaccine target for the 2011/2012 season for the northern hemisphere.

[dothedd]

WHO Recommended 2011/2012 Vaccine Targets Unchanged
Recombinomics Commentary 15:10February 18, 2011


Haemagglutination inhibition (HI) tests using postinfection ferret antisera indicated that A(H1N1)pdm09 viruses remained antigenically homogeneous and closely related to the vaccine virus A/California/7/2009. Sequence analysis of the HA genes of A(H1N1)pdm09 viruses indicated that the viruses fell into three genetic subgroups which were antigenically indistinguishable. A small number of viruses showed reductions in reactivity with some ferret antisera in HI assays.

The above comments are from the newly released WHO document on the recommended 2011/2012 vaccine targets, “Recommended composition of influenza virus vaccines for use in the 2011-2012 northern hemisphere influenza season,” which keeps all three targets (pandemic H1N1, seasonal H3N2, influenza B) unchanged from the recommendations for the 2010/2011 season. As indicated above, this recommendation is heavily dependent on antigen characterization tests which use ferret reference sera to characterize new isolates.

However, the track record for this assay is abysmal due to a lack of reproducibility, intra and inter-lab variation, low sensitively, and results which are inconsistent with clinical data. The failures have been glaring for the H1N1 target (both seasonal and pandemic) since the 2007/2008 season, and the above recommendation is clearly at odds with the reports of widespread pandemic H1N1 activity in countries and regions where pandemic H1N1 was widespread last season, as well as published and anecdotal reports on vaccine breakthroughs. Moreover, the latest Pneumonia & Influenza death rate for the latest reporting period in the United States (week 6) is near historic highs and well above the rate recorded last season. These examples of immunological escape this season signal more evolution away from the 2009 isolate, California/7, which is the recommended target for next season.

The shortcomings of the antigen characterization test are detailed here, and discussed in depth here.

The reliance of WHO and the CDC on this fatally flawed test continues to be hazardous to the world’s health and a revised method for the selection of influenza vaccine targets is long overdue.

[dothedd]

WHO Issues H1N1 Warning
Recombinomics Commentary 16:30

February 18, 2011


Experience during the 2010–2011 influenza season in the northern hemisphere has demonstrated that pandemic influenza A(H1N1) 2009 virus is still circulating and, importantly, still causing severe disease in younger people. Continued vigilance is imperative, and the appropriate surveillance, control and treatment practices that allow for the control of both established seasonal influenza strains and pandemic influenza A(H1N1) 2009 virus should be maintained during the post-pandemic period.

The above warning represents the WHO conclusions in the report, “Influenza A(H1N1) 2009 virus: current situation and post-pandemic recommendations” in today’s Weekly Epidemiological record. It follows the August declaration that the H1N1 pandemic phase had ended. At the time H1N1 was widespread in India as well as multiple countries in the southern hemisphere and was killing previously healthy young adults. The recent outbreak in the UK indicated that H1N1 levels were higher than last season, leading to a spike in ICU cases as well as saturation in use of ECMO machines which had increased four fold over levels in the prior season.

Sequence data from UK isolates indicated November isolates were dominated by two sub-clades with S186P or S188T. These sub-clades began to emerge and spread in the southern hemisphere and had become dominant in the UK. Similar sequences were subsequently reported throughout the northern hemisphere, including the United States and Canada in North America, as well as Mongolia, Russia, China, and Japan in Asia. In the US the week 4 P&I death rate spiked to 8.45% and week 6 spiked to 8.89%, just shy of the five year high of 9.1% in week 11 in 2008. This dominance and spread suggests outbreaks similar to the UK will be widespread throughout the northern hemisphere in the upcoming weeks.

The above data supported vaccine escape, which was confirmed by a 50% failure rate in the UK. However, in spite of these dramatic results, the WHO advisory committee recommended the same vaccine targets for the 2011/2012 season, raising serious concerns about the utility of the antigen characterization tests which produced intra and inter-lab variations including results inconsistent with sequence and clinical data (discussed in detail here).

The WHO warning on the emerging H1N1 sub-clades is appropriate, but recommendations for vaccination using targets which produced poor protection against contemporary sub-clades continues to raise serious concerns about the current vaccine target selection and strategy.

[dothedd]

Recombinomics Commentary 19:29
February 18, 2011


Five influenza-associated pediatric deaths were reported. Three of these deaths were associated with 2009 influenza A (H1N1) virus, one was associated with an influenza A virus for which the subtype was not determined, and one was associated with an influenza B virus.

The above comments from the CDC's week 6 FluView indicate that H1N1 is now causing the majority of pediatric deaths in the United States. These data compliment the Pneumonia and Influenza death rate, which has spiked to 8.9%, which is graphically represented below. The graph displays this data historically, and demonstrates its relations to the five year high of 9.1% recorded in week 11 of 2008. As seen in the graph, there is no indication that the week 6 data represents the peak for this season. The H1N1 levels are clearly rising raising concerns that the death rate will continue to rise.

[dothedd]

Unchanged Vaccine Targets Raise Pandemic Concerns
Recombinomics Commentary 14:29
February 19, 2011


This week the WHO recommended leaving the three vaccine targets for the 2011/2012 northern hemisphere flu season unchanged, while the pneumonia and influenza death rate in the US matched the highest level since week 14 of 2008 (8.9%) and approached the five year high of 9.1%, set in week 11 of 2008, as seen in the five year graph below, as published in the CDC’s week 6 FluView.

These record peaks are historically associated with vaccine mismatches. In the 2007/2008 season the three vaccine targets were A/Solomon Islands/3/2006, A/Wisconsin/67/2005, and B/Malaysia/2506/2004. All three targets for the following 2008/2009 season were changed, to A/Brisbane/59/2007, A/Brisbane/10/2007, B/Florida/4/2006.

The decision to leave the targets unchanged for the 2011/2012 was largely based on the antigenic characterization test, which has an extremely poor track record with regard to predictive value, sensitivity, or reproducibility. However, in spite of its abysmal performance, WHO consultants use this assay to recommend targets which are supplied to all major pharmaceutical companies making the flu vaccine, which are then distributed worldwide.

The recent history of the assay for seasonal and pandemic H1N1 has been reviewed recently here and here. The results for the 2007/2008 were particularly instructive. The ferret anti-sera used by the CDC could distinguish clade 1 (represented by New Caledonia/20/1999) from clade 2 (including Solomon Islands/03/2006), but did not distinguish between the three clade 2 sub-clades (Clade 2A - Solomon Islands/3/2006, Clade 2B - Brisbane/59/2007, and Clade 2C- Hong Kong/2562/2006). In the 2007/2008 season, the vaccine target, Solomon Island/3/2006 was no longer circulating. It had been replaced by the other two clade 2 sub-clades.

However, the CDC anti-sera was used to claim that Brisbane/59 and Hong Kong/2562 were a “good match”, which was refuted by the sequence data, the dominance of the clade 2B and clade 2C sub-clades, and a discriminating ferret anti-sera directed against Brisbane/59 grown in mammalian (MDCK) cells. As noted above, this target was then changed in the following northern hemisphere flu season. But the H1N1 mismatch for 2007/2008 season in the northern hemisphere, and 2008 in the southern hemisphere facilitated the emergence of Brisbane/59 like H1N1 which was Tamiflu resistant and had evolved away from the Brisbane/59 target, leading to widespread vaccine failure and the increase in Tamiflu resistance (H274Y) to 100% worldwide.

As noted earlier, the antigen characterization tests on 2009 and 2010 pandemic H1N1 isolates produced another series of irreproducible results including significant intra and inter-lab variations, including the absence of any H1N1 “low reactors” in the United States this season, in spite of reported 50% vaccine failure rate in the UK and record death rates as indicated by the latest P&I data in the graph above.

The latest recommendations by the WHO advisory committee raise serious concerns about the ability of these consultants to integrate clinical and sequence data with an extremely controversial and out dated antigen characterization test, which remains hazardous to the world’s health.

LINK: To view Chart

www.recombinomics.com/News/02191101/US_PI_6_11_Pandemic.html

[dothedd]

Vaccine Failure In Western Pennsylvania
Recombinomics Commentary 23:10
February 21, 2011


Officials said a Vanderbilt man died from the flu over the weekend, while the number of cases in Western Pennsylvania continues to increase.

Health officials said the number of flu cases is increasing even among people who were vaccinated.

"Myself and my colleagues, we observe people that were vaccinated for the flu and are coming down with symptoms. So there seems to be some variant strain circulating." said Dr. Andrew Sahud, chairman of the infection prevention committee at Allegheny General Hospital.

The above comments provide additional anecdotal data on vaccine failures this season across the northern hemisphere. In week 6 the number of H1N1 cases in Allegheny County almost doubled from 23 to 42, representing almost half of the reported cases for the entire state. Thus, it is likely that the above cases are dominated by H1N1, so a vaccine failure would be consistent with data from the UK, where vaccine failure in 50% of vaccinated patients was reported.

Vaccine failure and immunological escape are also supported by the high number of cases being reported throughout the United States. The P&I death rate for week 6 spiked to 8.9%, just shy of the five year peak of 9.1%, providing further evidence for vaccine failure. Last season in the fall of 2009 a serological study indicate almost half of the school age population in Allegheny county had H1N1 antibodies, confirming widespread infections in the greater Pittsburgh area, but H1N1 has made a comeback this season signaling immunological escape. The repeated selection of California/7/2009 for the current season has led to the predicted escape, and the selection of the same targets for the 2011/2012 season provides additional concerns for the use of the antigenic characterization test by vaccine selection committees in the US and worldwide.

The vaccine escape in Western Pennsylvania increases concerns that the procedures used by WHO and CDC to select vaccine targets are hazardous to the world’s health.