Lİnk between deadly d225g mutatİon and tamİflu resİstance h274 mutatİon

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December 2, 2009

1,937,292 Influenza/ARI

116,982 Hospitalized

446 Dead

The above numbers represent the latest update from the Ukraine Ministry of Health. As the number of cases approach 2 million, the rate of increase has slowed and shifted to the east. The largest daily update was in Donetsk, which rose by 5892 cases to 127,584 (see map). However. Increases in deaths are being report across the eastern border in Russia, were there have been 30 deaths reported in Saratov. Similarly, almost 200 deaths have been reported in Turkey, raising concerns of D225G spread.

The initial data showed strong association of D225G with fatal cases. Of the 9 samples from western Ukraine, all four fatal cases had D225G. In contrast nasal washes from surviving cases were positive for the same sub-clade, but were wild type at position 225, suggesting that infections were mixtures and virus with D225G targeted the lung, while wild type remained in the upper respiratory tract.

This tissue tropism poses surveillance problems, because most sequences come from nasal pharyngeal swabs, which may not reflect D228G in the lung. Currently, only 1% of HA sequences have D225G, although it was present in April in the vaccine target, A/California/7/2009, as a mixture. The version with D225G was selected for the target in the attenuated carrier distributed as a flu mist. In contrast, the killed infected vaccine has the HA version of California/7 with wild type D at position 225, raising concerns that killed vaccine or natural immunity will select for D225G. This concern was increased when Mill Hill labeled Ukraine H1N1 with D225G a low reactor.

The reporting of D225G in all fatal cases in Ukraine should be followed up with additional sequence data on more of the 400+ fatalities. Collections from the upper and lower respiratory tract in fatal cases would help define the significance of D225G in fatal lung cases, and also determine the ratio of components of mixtures with and without D225G.

An increase in the D225G ratio would be cause for concern.

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Re: Lİnk between deadly d225g mutatİon and tamİflu resİstance h274 mutatİon

To further characterize the observed reduction in zanamivir
sensitivity exhibited by some of the isolates on the PRA, we also
performed sequence analysis of the HA gene in all of the isolates.
Mutations at one of two positions in the HA gene, N163 and D225,
were observed only in the viral isolates with reduced zanamivir
sensitivity (Table 3, full alignment Figure S5). Non-conserved
mutations at these two positions, N163K/H and D225G, thus may
be sufficient to cause reduced sensitivity to NAIs in MDCK cells.
Although a zanamivir-sensitive isolate, A/St.Louis/758/09, carries
a N163T mutation, this isolate also has a unique, nearby V189A
mutation that might negate the effects of the N163T mutation such
that the virus remains sensitive to zanamivir (Table 3). Sequence
alignment with a published IFV A H1 crystal structure reveals that
N163, D225, as well as V189 lie near the sialic acid binding pocket
of HA (Figure 2). Therefore, the observed mutations might affect
receptor binding affinity of the HA protein in MDCK cells.


In the 2007 and 2009 clinical isolates that appear to have
decreased sensitivity to zanamivir based on PRA, we have
identified mutations involving two amino acid residues in the
HA gene that are proximate to the sialic acid binding site, N163
and D225. Mutation of N163 may be highly relevant for NAIresistance
because glycosylation of asparagine (N) near the sialic
binding site directly impacts the HA receptor binding affinity
[29,30]. Mishin et al. reported that introduction of an N163G
mutation made a recombinant laboratory IFV strain more
sensitive to NAIs in MDCK cells [31].



Similarly in our study,
the 2007 and 2009 isolates that are resistant to oseltamivir and have
reduced sensitivity to zanamivir carry mutations in both NA
(H274Y) and HA (N163K/T/H and D225G), but the observed HA
mutations have not been reported previously in drug-resistant IFV
strains.

http://www.plosone.org/article/related/info:doi%2F10.1371%2Fjournal.pone.0007838;jsessionid=F54E293ACE4E0D8ACF439DAB5CC17032

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Re: Lİnk between deadly d225g mutatİon and tamİflu resİstance h274 mutatİon

Tamiflu resistance of influenza H1N1 strains

by Vincent Racaniello on 9 January 2009

Yesterday the New York Times ran an article on the resistance to Tamiflu of current influenza H1N1 strains circulating in the US. I wrote a post about this issue on 22 December 2008, so I’m happy to see the Times following my lead. But there is an issue with the Times article that I’d like to address here. According to the article:

The single mutation that creates Tamiflu resistance appears to be spontaneous, and not a reaction to overuse of the drug.

Drug-resistant viruses are not ‘reactions’ to overuse of the drug. The drug selects, from the diverse viral population in an individual, those viruses that can multiply in its presence. Usually the drug-resistant mutants are already in the host, and outpace other drug-sensitive viruses. Is that what the writer means by ’spontaneous’? Not in this case. What apparently happened is that the mutation that causes drug-resistance, a change from histidine to tyrosine at position 274 of the viral NA protein, emerged in parts of the world were little Tamiflu is used. There was some other reason why this change was selected for in those populations. The article implies that the his->tyr change accompanied a second amino acid change at position 193 of the HA protein which improved the ability of the virus to infect people. This change did not affect resistance to Tamiflu, but apparently it only persisted when the change at 274 was also present. It so happened that the 274 change also conferred resistance to Tamiflu. Thus, when this virus arrived in parts of the world where Tamiflu is used, the resistance was noted. None of this is made particularly clear from the article.

I also have an issue with the author describing the amino acid changes in the ‘N’ and ‘H’ genes. The correct nomenclature is NA and HA. The author might have been mislead by the strain designation which uses only ‘H’, e.g. H1N1. It’s a small point but I believe that the devil is in the details.

What about the two other anti-influenza drugs? And the other strain currently circulating, H3N2?

Most of the flu in the US now is caused by H1N1 strains. So although the H3N2 strains are sensitive to Tamiflu, it’s not much help.

The Tamiflu-resistant H1N1 strains are sensitive to another drug, Relenza (zanamivir). But that drug must be inhaled and is not appropriate for everyone. However, these H1N1 strains are sensitive to Rimantadine, so its use is a good alternative. Most H3N2 strains are resistant to Rimantadine, which is why it has not been used much in recent years.

Nevertheless, our anti-influenza drug arsenal is much too small. It’s worth recalling the following information from Principles of Virology (ASM Press):

With about 1016 human immunodeficiency virus (HIV) genomes on the planet today, it is highly probable that somewhere there exist HIV genomes that are resistant to every one of the antiviral drugs that we have now or are likely to have in the future.

AIDS is no longer a death sentence because we have a deep arsenal of antiviral drugs that can control the infection. Patients are treated with a combination of three anti-HIV-1 drugs at a time. When resistance inevitably emerges, the patient is switched to another combination of three. The high levels of HIV-1 replication in many hosts, coupled with the large numbers of viral mutants that are produced, ensure that resistance will emerge.

Influenza virus shares similar features as HIV-1: high replication rates in many hosts, and the generation of large numbers of viral mutants. Therefore any antiviral strategy that employs only three drugs is bound to fail. The difference with influenza, of course, is that an excellent vaccine is available, and should be used whenever possible. The antiviral compounds should only be used in the face of an outbreak when immunization has not been sufficiently comprehensive. However, I suspect that the use of Tamiflu and Relenza is far more prevalent than desired. How many people rush for a prescription at the first signs of a respiratory infection? And how many of those have already been immunized? This was not the intended use for these antiviral compounds.

If we want to seriously use antiviral to treat influenza (which I don’t think is a good idea except in certain cases), we need to have a far deeper arsenal of antiviral drugs.

http://www.nytimes.com/2009/01/09/health/09flu.html

http://www.virology.ws/2009/01/09/tamiflu-resistance-of-influenza-h1n1-strains/

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Re: Lİnk between deadly d225g mutatİon and tamİflu resİstance h274 mutatİon

The interaction between the 2009 H1N1 influenza A hemagglutinin and neuraminidase: mutations, co-mutations, and the NA stalk motifs

http://www.scirp.org/journal/jbise/