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Recombination in NA Gene of H5N1 Guangdong Wild Bird
Recombinomics Commentary
March 9, 2006
Recently researchers in China deposit H5N1 sequences from 7 isolates in China collected in 20904. The title of the submissions, "A cohort of AIV H5N1 subtypes isolated from wild aquatic birds and domestic poultry revealed rapid transmission, frequent reassortment, and identifiable recombination", indicated the sequences had examples of recombination. Analysis of the data revealed many examples of recombination. Two of the isolates, a wild bird from Guangdong, A/wild bird/Guangdong/314.2004(H5N1) and a chicken from Henan,
A/chicken/Henan/210/2004(H5N1), have striking examples of recombinations in multiple genes. In most examples, donor sequences were found in a Henan tree sparrow isolate, A/tree sparrow/Henan/1/2004(H5N1) and the recombination took several forms. In one instance the sequence represented approximately one third of the gene. In another the recombined region was short, but contained several polymorphisms. In addition there were single nucleotide changes also found in the same tree sparrow sequence.
As shown below, the wild bird NA was also a recombinant, but the donor sequences were identified in a 2003 H5N1 duck isolate, A/duck/China/E319-2/03(H5N1), from birds smuggled onto Quemoy island. In the series below there are five polymorphism that are spread over almost 600 BP (powitions 284-857) and these changes superficially look like random mutations. However, all five are located in the wild duck and have a limited distribution in the Los Alamos flu database, clearly demonstrating a common source and acquisition of the polymorphisms by recombination.
These data provide additional support for extensive recombination, which in most instances are mistaken for "random mutations." This view is held by WHO and consultants and leads to serious misinterpretations of the sequence data. In WHO's February 20, 2006 announcement on "Avian influenza: significance of mutations in the H5N1virus", the report states that " Influenza viruses are inherently unstable. As these viruses lack a genetic proof-reading mechanism, small errors that occur when the virus copies itself go undetected and uncorrected. Specific mutations and evolution in influenza viruses cannot be predicted……… Mutations and reassortment events are commonly observed in the affected bird populations."
However, as noted above, the "mutations" are not due to recent errors when the virus copies itself. Instead, the polymorphisms are acquired from other virus via homologous recombination. Because of the origin of the genetic material, the polymorphisms can be traced and the tracing shows that the polymorphisms are transmitted and transported by migratory birds and this movement and capture via recombination lead to the frequent genetic changes. Since the changes come from dual infections of existing flu strains, the acquisitions can be predicted. Moreover, these predictions can be used to create vaccine to sequences before they emerge in natures.
These predictions depend on a complete and up to date sequence database. Much of the tracing was les than ideal until recently when the database was updated with sequences from samples that were collected 25-30 years ago. These historical tracings are more predictive when applied to recent sequences. However, the recent sequences are sequestered in a private database, which is not being properly analyzed.
This incomplete analysis is being used to select new vaccines. The latest announcement of a new vaccine based on Indonesian isolates has been done in a vacuum. None of the human H5N1 sequences from Indonesia have been made public. The new vaccine's ability to protect against other human isolates is also unknown because no human sequences from China, Turkey, or Iraq have been released.
These sequences should be released immediately and selection of appropriate vaccines should use evolution via recombination and not the ill conceived and incorrect notion of random mutations.
C183T
AY950249 A/WildDuck/Guangdong/314/2004 2004 H5N1
AY518363 A/duck/China/E319-2/03 2003 H5N1
C284T
DQ321042 A/chicken/Hunan/999/2005 2005 H5N1
DQ321032 A/duck/Guangxi/951/2005 2005 H5N1
DQ095670 A/Duck/Hunan/114/05 2005 H5N1
DQ321043 A/duck/Hunan/1265/2005 2005 H5N1
DQ321034 A/duck/Hunan/127/2005 2005 H5N1
DQ321035 A/duck/Hunan/139/2005 2005 H5N1
DQ321036 A/duck/Hunan/149/2005 2005 H5N1
DQ321037 A/duck/Hunan/152/2005 2005 H5N1
DQ321038 A/duck/Hunan/157/2005 2005 H5N1
DQ321039 A/duck/Hunan/160/2005 2005 H5N1
DQ321044 A/duck/Hunan/1608/2005 2005 H5N1
DQ321045 A/duck/Hunan/1652/2005 2005 H5N1
DQ321040 A/duck/Hunan/166/2005 2005 H5N1
DQ321041 A/duck/Hunan/182/2005 2005 H5N1
DQ095671 A/Duck/Hunan/191/05 2005 H5N1
DQ321070 A/duck/Vietnam/568/2005 2005 H5N1
DQ321028 A/goose/Guangxi/345/2005 2005 H5N1
DQ321029 A/quail/Guangxi/575/2005 2005 H5N1
AY737299 A/chicken/Guangdong/178/04 2004 H5N1
AY737291 A/chicken/Guangdong/191/04 2004 H5N1
DQ321027 A/chicken/Guangxi/2461/2004 2004 H5N1
AY651476 A/Dk/HN/101/2004 2004 H5N1
AY651477 A/Dk/HN/303/2004 2004 H5N1
DQ321009 A/duck/Guangxi/351/2004 2004 H5N1
DQ321010 A/duck/Guangxi/380/2004 2004 H5N1
ISDN48958 A/Duck/Hu nan/15/2004 2004 H5N1
AY651478 A/Ph/ST/44/2004 2004 H5N1
AY950249 A/WildDuck/Guangdong/314/2004 2004 H5N1
AY651475 A/Dk/HN/5806/2003 2003 H5N1
AY518363 A/duck/China/E319-2/03 2003 H5N1
A311T
DQ095669 A/Duck/Fujian/1734/05 2005 H5N1
AY950249 A/WildDuck/Guangdong/314/2004 2004 H5N1
AY518363 A/duck/China/E319-2/03 2003 H5N1
T398C
DQ321063 A/chicken/Bantul/BBVet-I/2005 2005 H5N1
DQ321032 A/duck/Guangxi/951/2005 2005 H5N1
DQ321043 A/duck/Hunan/1265/2005 2005 H5N1
DQ321044 A/duck/Hunan/1608/2005 2005 H5N1
DQ321045 A/duck/Hunan/1652/2005 2005 H5N1
DQ321070 A/duck/Vietnam/568/2005 2005 H5N1
DQ321028 A/goose/Guangxi/345/2005 2005 H5N1
ISDN125875 A/Indonesia/5/05 2005 H5N1
DQ321029 A/quail/Guangxi/575/2005 2005 H5N1
AY737299 A/chicken/Guangdong/178/04 2004 H5N1
DQ321027 A/chicken/Guangxi/2461/2004 2004 H5N1
DQ321061 A/chicken/Yogjakarta/BBVet-IX/2004 2004 H5N1
AY651437 A/Ck/Indonesia/4/2004 2004 H5N1
AY651436 A/Ck/Indonesia/5/2004 2004 H5N1
AY651434 A/Dk/Indonesia/MS/2004 2004 H5N1
DQ321009 A/duck/Guangxi/351/2004 2004 H5N1
DQ321010 A/duck/Guangxi/380/2004 2004 H5N1
ISDN48958 A/Duck/Hu nan/15/2004 2004 H5N1
AY651478 A/Ph/ST/44/2004 2004 H5N1
AY950249 A/WildDuck/Guangdong/314/2004 2004 H5N1
ISDN111354 A/Chicken/Indonesia/11/2003 2003 H5N1
ISDN111353 A/Chicken/Indonesia/7/2003 2003 H5N1
DQ321060 A/chicken/Wonosobo/BPPV4/2003 2003 H5N1
AY651465 A/Ck/HK/YU324/2003 2003 H5N1
ISDN38805 A/Ck/HK/YU357/03 2003 H5N1
AY651435 A/Ck/Indonesia/2A/2003 2003 H5N1
AY651432 A/Ck/Indonesia/BL/2003 2003 H5N1
AY651433 A/Ck/Indonesia/PA/2003 2003 H5N1
AY518363 A/duck/China/E319-2/03 2003 H5N1
A857C
AY950249 A/WildDuck/Guangdong/314/2004 2004 H5N1
AY518363 A/duck/China/E319-2/03 2003 H5N1
Map