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H5N1 HA Cleavage Site Evolution
December 21, 2005
The publication of the H5N1 HA cleavage linked to the first reported human cases in mainland China raises additional concerns about the evolution of H5N1 and its ability to cause human disease.
Cleavage of the HA site is the first step in influenza entry into a cell. Normally cleavage of this site is limited to tissues that express the appropriate protease. However, highly pathogenic avian influenza (HPAI) has been found to have multiple basic amino acids at this site, which allows for cleavage by a variety of protease expressed in various cell types, leading to an expanded tissue tropism. The multi-basic amino acids have been found in H7 and H5 isolates and all such isolates have been HPAI.
This multi-basic cleavage site was first report in H5N1 in Asia in 1996. The cleavage site had a basic amino acid in 7 of the 8 position preceding the cleavage location of wild type influenza. This motif (RERRRKKR) was also detected the following year in Hong Kong patients infected with H5N1. The same sequence was detected in isolates from patients in 2003 in Hong Kong as well as 2004 in Vietnam and Thailand.
Various duck and goose isolates from 2000 and 2001 had variations in the sequence just upstream from the 6 consecutive basic amino acids. These variations included RDRRRKKR, IERRRKKR, and GERRRKKR. In 2003 and 2004 the 6 consecutive basic amino acids were reduced to 5 by changing the first amino acid (REIRRRKKR) or deleting an R (RERRKKR) or K (RERRRKR).
The deletion of the K or R was largely found in China or Japan in 2003 and 2004, but then the deleted K was found in isolates in northern Vietnam this year. This change was associated with an increased transmissibility and decreased fatality rate. Although the change matched the isolates from China or Japan, the cleavage site was on an HA that was in clade 1 although the earlier isolates with the deletion were in clade 2.
In addition 2004 isolates from Thailand changed one of the R’s to a K to produce cleavage sites of REKRRKKR or RERKRKKR.
These evolving cleavage sites continued to be reported this year. The GERRRKKR site was found at Qinghai Lake and has been also reported for isolates from Siberia, Kazakhstan, Tula, Croatia, Romania, and Turkey.
However, a novel HA cleavage site was reported in three isolates from whooping swans in Mongolia (GERRRRKR) and most recently, the cleavage site in the H5N1 associated with two human cases in Hunan was RERRRRR.
The biological or clinical effects of these cleavage site changes remains to be determined. However, the variety of sites is steadily increasing and the effects are uncertain. At least three different cleavage sites (RERRRKKR, RERRKKR, and RERRRR) have been linked to human H5N1 cases and these variations are likely to increase as more sequence data becomes available.
The evolution of the cleavage sites highlights the rapid evolution of H5N1 and suggests the number of different H5N1’s capable of causing human infections may be large and growing larger.