Human trH3N2 / trH1N1 Matches Raise Pandemic Concerns
Recombinomics Commentary 21:10
November 15, 2010

Previously, three of these reports had been swine origin A (H3N2) viruses. The Pennsylvania and Wisconsin cases bring the number of reports swine origin A (H3N2) infections in humans in the United States to five. The viruses identified in Pennsylvania and Wisconsin are similar to viruses that infected a patient in Iowa in September 2009, a patient in Kansas in August 2009 and a patient in Minnesota in May 2010.

The above comments from the CDC update confirms that all five human triple reassortant H3N2 (trH3N2) are “similar” to each other.  Although triple reassortants (TRs) in swine were first described in the late 1990’s, concerns have increased because of human cases such as the five cases described above, as well as the H1 TRs found in humans prior to the 2009 pandemic, which also involved TRs.  The constellations of internal genes are similar.  The polymerase complex is composed of one human flu gene (PB1), and two avian flu genes (PB2 and PA), which give rise to a wide variety of TRs.  The complex has been found in a variety of serotypes, which are easily distinguished by the H and N surface genes, which are frequently swapped.  trH3N2 have a human H3 and N2, and the timing of these introductions can be determined by the sequence of the H and N genes.  Similarly, the internal genes have different evolutionary paths, which form groupings which can be determined through phylogenetic analysis. Similarly, H1 TRs can have H and N represented by swine of human flu genes.  The pandemic H1N1 (pH1N1) had swine H1 and N1.  However, other TRs have human H1 and human N1 or N2.  Thus, there are many combinations of TRs in swine, but only one TR has caused a pandemic characterized by sustained transmission in humans.  However, the recent pager alert on H3N2 TRs has raised concerns that an H3N2 swine pandemic could arise, since H3N2 TRs have human H and N, which the constellation of internals genes, including the polymerase complex is found in pH1N1.  This potential can be evaluated through phylogenetic analysis.

This analysis, however, is limited by the withholding of sequences, or the generation of partial sequences.  However, relationships can be identified through targeted analysis.  Thus, although the CDC has not released the sequences from the more recent trH3N2 cases, one full set of sequences, A/Kansas/13/2009, was released from a patient (22M) associated with the outbreak at the Riley County fair in the summer of 2009.  Similarly, full sets of sequences from swine at the Huron County fair in Ohio in the summer of 2007. 

These sequences, A/swine/Ohio/24366/07 and A/swine/OH/511445/2007, were of interest because the HA sequences from two of the swine isolates matched each other and were identical to two exhibitors (10F and 36M) at the fair, A/Ohio/01/2007 and A/Ohio/02/2007.  These two cases were included the NEJM report of the 11 TRs that preceded the 2009 pandemic.  The cases were patients #4 and #5 in the associated table.  The authors of the paper on the swine sequences noted that the full set of swine sequence matched the full set of human sequences generated by the CDC.  Thus, although the human sequences were not made public, the public swine sequences matched the full sets of sequences from the two human cases.

Although the constellations of the internal genes in TRs are similar, there are distinctions that can be identified through phylogenetic analysis as well as anti-viral resistance due to S31N in M2, which is found invirtually all pH1N1, but M2 is sensitive in most of the H1 and H3 TRs from humans and swine.  Another difference of interest is PB1 E618D, which is in all pH1N1 as well as the human trH3N2 from Kansas.  This new acquisition may signal further adaptation to humans.

However, the full set of sequences in internal genes in the Huron County fair are of interest because the two exhibitors represented the only examples of two confirmed cases from the same outbreak, as described in the NEJM report.  Moreover, about two dozen fair attendees had flu-like symptoms, which are unusual in August in Ohio.

The concern over these internal gene sequences increased when phylogenetic analysis was carried out for the internal genes of the human trH3N2 sequences from the Riley County fair, which indicated that the best match for the 2009 sequences were the 2007 sequences from Ohio.
This match raises concerns that the minor changes in the Ohio TR genes are important for adaptation to humans, which pruced the large number of cases in Ohio, as well as the increasing number of trH3N2 cases, including the two that generated the pager alert last week.

This match suggests that most or all H3N2 sequence withheld by the CDC also are closely related to the published sequences from Ohio and Kansas, and the similarity signals human to human transmission in multiple states (Kansas, Iowa, Minnesota, Wisconsin, and Pennsylvania).
Detection of such sequences is limited because the H3 links to seasonal H3N2 circulating in the mid-1990’s while the N2 links to seasonal H3N2 circulating in 2003.  Thus, the current H3 and N2 sero-typing reagents would not easily distinguish trH3N2 from seasonal H3N2.

Thus, the full set of trH3N2 sequences from human cases should be released, and sequencing efforts of recent H3N2 cases should be expanded.  Although the H3 and N2 sequences have a human lineage, they link to older human sequences which have evolved further in swine and would not be well covered by current H3N2 vaccines.

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