Sequences at Genbank!

[gsgs]

Re: Sequences at Genbank!

I checked all the WRAIR sequences.
It seems that they are less reliable than other sequences.
Additional mutations show up, which are unlikely to be real -
why should WRAIR have more mutations than others ?

wrair2.gif
wraivrdl.gif



one of the low-mutation Canada sequences:

http://www.ncbi.nlm.nih.gov/nuccore/HQ240895

paper: http://www.ncbi.nlm.nih.gov/pmc/article ... 016087.pdf

ahh, identical to this one from May, also in the other segments,
looks like they confused their sequences

http://www.ncbi.nlm.nih.gov/nuccore/HQ240894

--------------------------
and one of the Czech with few mutations:

http://www.ncbi.nlm.nih.gov/nuccore/JQ693489

those 2 Czech viruses are similar to viruses from 2009,
which were completely replaced in 2010/11 by a new strain (from India ?)
So this looks like a date-error by the Czech lab or a contamination
with an old sample from 2009
-------------------


wrair2.gif - earlier pic: May 24 , 2011

http://www.flutrackers.com/forum/showpost.php?p=409421

[gsgs]

Re: Sequences at Genbank!

segment 1, classical H1N1, selected subtable of 10 WRAIR sequences,
nucleotide positions 657-777 in the coding region

seems that this portion was wrongly taken from a H1N1v-sequence
that was processed in the same run, thus giving the impression
of a wrong recombination

                                          0000000000000000000000000
                                          6666666666777777777777777
                                          5666678899001144456667777
                                          7013631439584714733781457

Index                                     GTGATAGCTTAGACTGGGGATACCA
>A/Brisbane/59/2007/07/01(H1N1)           .........................

>A/South Carolina/WRAIR1116P/2009/03/01   ...............AA........
>A/Texas/WRAIR1118P/2009/06/01            .........CC.G.G.AA.......
>A/Medellin/WRAIR1297P/2008/06/28         ..............AAAA..C....
>A/Nevada/WRAIR1257P/2009/02/01           ..............AAAA..C....
>A/Sullana/WRAIR1707P/2009/02/21          .........CCAG.AAAA..C....
>A/Pucallpa/WRAIR1709P/2009/01/20         ..........CAG.AAAA..C....
>A/Piura/WRAIR1708P/2009/03/27            .........CCAG.AAAA..C....
>A/Santa Cruz/WRAIR1706P/2008/12/09       ........GCCAG.AAAA..C....
>A/Cusco/WRAIR1730P/2008/08/20            .........CCAG.AAAA.......
>A/Maracay/WRAIR1729P/2008/09/03          ........GCCAG.AAAA..C....

>A/H1N1v-/index/2009/02/01                AA.CCGTTGCCAG.AAAA..C.TTG

-------------------------------------------------------------------------------------------

http://en.wikipedia.org/wiki/DNA_sequencing#Next-generation_methods

The high demand for low-cost sequencing has driven the development of high-throughput
sequencing (or next-generation sequencing) technologies that parallelize the sequencing
process, producing thousands or millions of sequences at once.[18][19]
In ultra-high-throughput sequencing as many as 500,000 sequencing-by-synthesis
operations may be run in parallel.[21][22][23]

-----------------------------------------------

mutations-over-time picture updated with marks (squares) around the A/WR(number)... sequences,
which are also WRAIR :




-------------------------------------------------------------------
AUTHORS Houng,H.-S.H., Li,T., Verratti,K., Gong,H., Zhou,Y., Lipkin,W.I.,
Street,C., Seriwatana,J., Lyons,A.G., Keyes,J., Yun,T. and
Kuschner,R.A.
TITLE High throughput full genome sequencing of 2009 swine H1N1 isolates
from worldwide origins using Roche 454 FLX system
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 2280)
TITLE Direct Submission
JOURNAL Submitted (10-NOV-2009) Viral Diseases, Walter Reed Army Institute
of Research, 503 Robert Grant Avenue, Silver Spring, MD 20910, USA
-------------------------------------------------------------------

AUTHORS Houng,H.-S., Zhou,Y., Lyons,A. and Kuschner,R.A.
TITLE Full genome sequencing of Influenza by Roche 454 FLX
JOURNAL Unpublished
TITLE Direct Submission
JOURNAL Submitted (10-AUG-2010) Viral Diseases, Walter Reed Army Institute
of Research, 503 Robert Grant Ave., Silver Spring, Maryland 20910,

------------------------------------------------

[gsgs]

Re: Sequences at Genbank!

I found here: (found by (random) googling)

http://www.mdpi.com/2073-4425/1/1/38/pdf

1. Sanger sequencing
>99.9% raw base accuracy
read lengths of up to 1 kb

high-density capillary array electrophoresis
algorithms to deconvolute complex electropherogram tracings

2. Next generation DNA sequencing

2.1. Roche/454 Life Sciences In 2005,

The key advantage of the 454 system when compared to other
platforms is its longer read length and shorter run times.
In eight hours, the second generation GS FLX instrument is capable
of an output of 100 Mb with an average read length of 250 bases
per template. Improvements to the picotiter plate and sequencing
chemistry have increased the read length to an average of 400
bases with a corresponding increase in throughput (400-600 Mb)

=======================================


JCVI:

http://gsc.jcvi.org/projects/msc/influenza/pipeline.shtml

Human influenza samples are currently sequenced using the Sanger sequencing method.
Degenerate primers are designed from aligned reference genomes using a computational
PCR primer design pipeline developed at JCVI to produce tiled amplicons with an optimal
length of 550 bp, with 100 bp overlap in order to provide six-fold sequence coverage of the
influenza genome.
...
Avian influenza samples are subject to a multiplexed Next Gen sequencing strategy to
enable sequencing of 100-200 samples in a single sequencer run. Hybrid Next Generation
sequencing technologies (Roche-454 and Illumina GAIIX )
...
The SISPA method randomly amplifies RNA or DNA and has been used to generate
consensus sequence for 100 samples to > 200X average depth of coverage per 454
or Illumina run.
...
However, owing to biased sequence coverage, a significant number of these viral genomes
require significant finishing by targeted Sanger sequencing of problematic regions

An automated assembly pipeline has been established to assemble hundreds of viral
genomes that have been sequenced using a multiplex bar-coding protocol for 454 and Illumina.
The assembly method involves deconvolution and binning of bar-coded reads and trimming
of barcodes and random hexamers. Binned reads are assembled de novo, and compared
with a database of full length genomic sequences using BLASTN to select the best reference
for mapping assembly. Consensus sequences are generated and variations identified by heuristic
algorithms comparing output of multiple sequencing technologies.

================================================== =================

[gsgs]

Re: Sequences at Genbank!

Canada
--------------

note the strange thing with the lacking mutations in Canada genomes !
late 2009/early 2010

http://magictour.free.fr/panflu/mxai6.GIF

http://www.ncbi.nlm.nih.gov/nuccore/HQ240909

genbank says : 21-Jan.2010, but in the paper they say 10/2009,
(I haven't read the whole paper)
but even that is too late.
They give same accession numbers for multiple viruses


316 ******=H1N1v=A/H1N1pdm genomes from Canada:

[gsgs]

Re: Sequences at Genbank!

one flu-genome (~13300 nucleotides) is assembled from ~200 subsequences of length ~500 nucleotides
as an example I tried this:

http://www.ncbi.nlm.nih.gov/Traces/trace.cgi?cmd=retrieve&size=187&val=species_code%3D%22INFLUENZA+A+VIRUS+%28A%2FCALIFORNIA%2FVRDL116%2F2009%28H1N1%29%29%22&file=trace&gz=on&fasta=on&dopt=fasta&dispmax=500&page=1

pic sang02a.gif

this is available for ~9000 influenza viruses (genomes)

http://www.ncbi.nlm.nih.gov/Traces/trace.cgi?cmd=stat&f=xml_list_species&m=obtain&s=species&letter=I&page=1

avian viruses and viruses samples with new next-generation-sequencing (NGS)
are handled differently

[mixin]

Re: Sequences at Genbank!

Do those little red vertical lines at the bottom of your picture represent the Cancun mutations? Canada has none of them?

What is your opinion on the strange lacking of mutations in the Canadian genomes?

[gsgs]

Re: Sequences at Genbank!

yes, Cancun.
~25% of Canada samples are Cancun. But most are from early 2009,
meanwhile almost 100% should be Cancun, even in Canada.

Canada had extensive pre-Cancun in 2009, unlike USA.

I think they probably got the dates wrong and the low-mutation
genomes are from spring 2009 as well.
Yiming asked them ...

[gsgs]

Re: sequences at genbank !

ahh, what had we been wondering about those Dakota mallards all the years.
Now retracted !

http://www.ncbi.nlm.nih.gov/pubmed/23260489

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3321961/

A/mallard/South Dakota/Sg-00125/2007 (H3N2)
A/northern pintail/South Dakota/Sg-00126/2007 (H3N2)
A/mallard/South Dakota/Sg-00127/2007(H3N2)
A/mallard/South Dakota/Sg-00128/2007(H3N2),




post #7 in this thread from 2009/04/28 :

here the best matches in the 8 segments

this is from my last update, early 2009 (maybe only 2007 in PB1, as I just see)


columns are:

difference in promille
different nucleotides
length of matching part
enumeration
strain


0 0 2280 6904 California/04/2009-04-01/(H1N1)

--------------------------------------------------------------
 304   24  787 2897 Turkey/MO/24093/99(H1N2)
 306   69 2249 3703 mallard Dk/South Dakota/Sg-00127/07(H3N2)
 306   69 2249 3704 mallard Dk/South Dakota/Sg-00128/07
 307   69 2246 3702 pintail Dk/South Dakota/Sg-00126/07(H3N2)
 307   69 2247 3701 mallard Dk/South Dakota/Sg-00125/07(H3N2)
 321   50 1557 2887 Sw/Texas/4199-2/98(H3N2)
 333   52 1557 2888 Sw/Minnesota/9088-2/98(H3N2)
 333   52 1557 2895 Sw/Iowa/8548-1/98
 342   78 2280 3760 Sw/Korea/JNS06/04(H3N2)
 343   54 1573 2894 Wisconsin/10/98(H1N1)
 346   79 2280 3705 Sw/Minnesota/55551/00(H1N2)
-------------------------------------------------------------
 312   71 2271 5644 Wisconsin/10/98(H1N1)
 340   77 2259 5652 mallard Dk/South Dakota/Sg-00125/07(H3N2)
 351   80 2274 5656 Sw/Indiana/9K035/99(H1N2)
 365   83 2271 5638 Sw/Nebraska/209/98(H3N2)
 365   83 2271 5639 Sw/Iowa/533/99(H3N2)
 365   83 2271 5640 Sw/Iowa/569/99(H3N2)
 365   83 2271 5641 Sw/Minnesota/593/99(H3N2)
 369   84 2271 5657 Sw/Korea/CAN04/05(H3N2)
 369   84 2274 5675 Dk/NC/91347/01(H1N2)
 374   43 1149 4066 Sw/Minnesota/9088-2/98(H3N2)
 374   43 1149 4068 Sw/Iowa/8548-1/98
 374   85 2271 5668 Sw/Illinois/100085A/01(H1N2)
 374   85 2271 5670 Sw/Ohio/891/01(H1N2)
 378   86 2271 5654 Sw/Iowa/930/01(H1N2)
 378   86 2271 5682 Sw/Korea/CY04/07(H3N2)
 378   86 2271 5683 Sw/Korea/CY05/07(H3N2)
 382   44 1149 4067 Sw/Texas/4199-2/98(H3N2)
 383   87 2271 5658 Sw/Korea/CAS07/05(H3N2)
 383   87 2271 5659 Sw/Korea/CAS09/06(H3N2)
 383   87 2271 5690 Sw/Indiana/P12439/00(H1N2)
------------------------------------------------------------------
 370   79 2130 4925 mallard duck/South Dakota/Sg-00128/2007
 370   79 2131 4923 pintail duck/South Dakota/Sg-00126/2007(H3N2)
 373   79 2117 4922 mallard duck/South Dakota/Sg-00125/2007(H3N2)
 381   79 2073 4924 mallard duck/South Dakota/Sg-00127/2007(H3N2)
 390   84 2151 2977 Swine/Nebraska/209/98 (H3N2)
 390   84 2151 2978 Swine/Iowa/533/99 (H3N2)
 390   84 2151 2979 Swine/Iowa/569/99 (H3N2)
 390   84 2151 2980 Swine/Minnesota/593/99 (H3N2)
 390   84 2151 3015 Swine/Illinois/100084/01 (H1N2)
 399   86 2151 3011 Swine/Minnesota/55551/00 (H1N2)
----------------------------------------------------------------
 470   80 1700  344 Swine/Indiana/P12439/00 (H1N2)
 482   82 1700  280 Swine/Indiana/9K035/99 (H1N2)
 500   85 1700  504 Turkey/MO/24093/99(H1N2)
 505   86 1700 2468 swine/Minnesota/1192/2001(H1N2)
 511   87 1700  339 Swine/Ohio/891/01(H1N2)
 517   88 1700  549 SW/MN/23124-T/01(H1N2)
 517   88 1700 2115 swine/Guangxi/17/2005(H1N2)
 529   90 1700  550 SW/MN/23124-S/01(H1N2)
 529   90 1700  552 SW/MN/16419/01(H1N2)
 535   91 1700  345 Swine/Illinois/100085A/01 (H1N2)
 542   56 1032  296 Wisconsin/10/98 (H1N1)
------------------------------------------------------------------
 250   13  520 6925 Sw/Oklahoma/18717/99(H3N2)
 251   13  516 6921 Sw/Colorado/23619/99(H3N2)
 262   14  533 6922 Sw/Wisconsin/14094/99(H3N2)
 262   14  533 6924 Sw/Illinois/21587/99(H3N2)
 263   14  531 6923 Sw/Oklahoma/18089/99(H3N2)
 312   16  512 6926 Sw/NorthCarolina/16497/99(H3N2)
 316   31  980 6927 Turkey/MO/24093/99(H1N2)
 329   48 1457 6963 Sw/Iowa/533/99(H3N2)
 336   49 1457 6961 Sw/Indiana/P12439/00(H1N2)
 336   49 1457 6969 Sw/Korea/CY04/07(H3N2)
 336   49 1457 6970 Sw/Korea/CY05/07(H3N2)
 337   33  979 6928 Sw/Texas/4199-2/98(H3N2)
 337   33  979 6929 Sw/Iowa/8548-1/98
 339   19  560 6972 SW/NC/45319/99(H1N2)
 339   19  560 6973 SW/NC/17366/01(H1N2)
 339   19  560 6974 SW/CO/17871/01(H1N2)
 343   50 1457 6971 Sw/Korea/CY09/07(H3N2)
------------------------------------------------------------------
 589   83 1409 1741 Sw/England/195852/92(H1N1)
 652   92 1409 1742 Sw/Germany/Vi5698/95(H1N1)
 712   98 1376 1746 Sw/Belgium/74/85(H1N1)
 751  101 1344 1773 Sw/Ille et Vilaine/1455/99(H1N1)
 753  105 1394 1772 Sw/Cotes d'Armor/1488/99(H1N1)
 754  106 1404 1749 Italy/1424-4/95(H1N1)
 765  108 1410 1737 Sw/Belgium/1/83(H1N1)
 777  104 1338 1777 Sw/Italy/1492/97(H1N1)
 786  103 1310 1778 Sw/Italy/1513-8/98(H1N1)
 787  111 1410 1739 turkey/France/87075/87(H1N1)
 787  111 1410 1788 Sw/Italy/53949/04(H1N1)
 787  111 1410 1789 Sw/Italy/65296/04(H1N1)
 794  112 1410 1738 Sw/Finistere/3616/84(H1N1)
 795  102 1282 1750 Sw/Italy/1424-3/95(H1N1)
 801  113 1410 1736 Sw/Lot/2979/82(H1N1)
 801  113 1410 1790 Sw/Spain/53207/04(H1N1)
 801  113 1410 1791 Sw/Italy/247578/04(H1N1)
 801  113 1410 1793 Sw/Spain/51915/03(H1N1)
 812  106 1305 1774 Cotes d'Armor/1121/00(H1N1)
 815  115 1410 1794 Sw/Virginia/671/1987(H1N1)
--------------------------------------------------------------------
 230   22  955 2735 Sw/HK/5190/99(H3N2)
 230   22  955 2736 Sw/HK/5200/99(H3N2)
 230   22  955 2737 Sw/HK/5212/99(H3N2)
 261   25  955 2739 Sw/HK/1197/02(H3N2)
 261   25  955 2740 Sw/HK/1144/02(H3N2)
 272   26  955 2738 Sw/HK/9745/01(H3N2)
 272   26  955 2743 HK/1774/99(H3N2)
 293   28  955 2741 Sw/HK/7982/00(H3N2)
 314   30  955 2742 Sw/HK/9296/01(H3N2)
 345   33  955 2721 Sw/Leipzig/145/92(H3N2)
 366   35  955 2726 Sw/Cotes d'Armor/1482/99(H1N1)
 397   38  955 2722 wild boar/Germany/WS169/06(H3N2)
 397   38  955 2727 Sw/Italy/1521/98(H1N2)
------------------------------------------------------------
 346   29  838 6427 Sw/Texas/4199-2/98(H3N2)
 346   29  838 6428 Sw/Minnesota/9088-2/98(H3N2)
 357   30  838 6429 Sw/Iowa/8548-1/98
 381   32  838 6444 Sw/Indiana/9K035/99(H1N2)
 395   33  835 6436 Sw/North Carolina/98225/01(H1N2)
 395   33  835 6437 Sw/Minnesota/55551/00(H1N2)
 395   33  835 6445 SW/MN/3327/00(H1N2)
 395   33  835 6453 SW/IN/14810-T/01(H1N2)
 395   33  835 6454 SW/IN/14810-S/01(H1N2)
 395   33  835 6455 Sw/Korea/CAS09/06(H3N2)
 407   34  835 6456 Sw/Korea/CAN04/05(H3N2)
 407   34  835 6457 Sw/Korea/CAS07/05(H3N2)
 407   34  835 6467 Sw/Korea/CAS08/05(H1N1)
 407   34  835 6506 Sw/Iowa/930/01(H1N2)
 417   35  838 6430 Sw/Nebraska/209/98(H3N2)
 417   35  838 6438 Turkey/MO/24093/99(H1N2)
 417   35  838 6483 turkey/OH/313053/04(H3N2)

[mixin]

Re: Sequences at Genbank!

Thanks for finding that.

How well I remember puzzling about those Dakota ducks. I'm glad to know they retracted their paper.

I wonder what prompted them to do it now?

[gsgs]

Re: Sequences at Genbank!

GenBank: CY032878.2 (click to see this obsolete version)
Record removed. This record was removed at the submitter's request because
the sequence may contain contamination.

http://www.ncbi.nlm.nih.gov/nuccore/CY032878

Current status: suppressed, live
I II Version Gi Update Date Action
2 251833724 Jan 8, 2010 02:32 AM
2 251833724 Oct 22, 2009 04:24 PM
2 251833724 Jul 8, 2009 04:47 PM
1 190352826 Jun 17, 2008 12:46 PM
1 190352826 Jun 13, 2008 04:48 PM
Accession CY032878 was first seen at NCBI on Jun 13, 2008 04:48 PM

http://www.ncbi.nlm.nih.gov/nuccore/251833724?sat=4&satkey=37773663

AUTHORS Sreevatsan,S., Ramakrishnan,M.A., Wang,P., Anderson,T.L.,
Goyal,S.M. and Osterholm,M.T.


JOURNAL Submitted (08-JUL-2009) Department of Veterinary Population
Medicine, University of Minnesota, 1354 Eckles Avenue, Saint Paul,
MN 55108, USA

[mixin]

Re: Sequences at Genbank!

The authors didn't admit to contamination in the letter you linked to:

TO THE EDITOR: We would like to retract the letter entitled "Triple Reassortant Swine Influenza A (H3N2) Virus in Waterfowl," which was published the April 2010 issue of Emerging Infectious Diseases (1). The nucleoprotein gene sequences from the viruses reported in that letter are very closely related to those from the earliest detected triple reassortant swine influenza viruses [CY095676 A/sw/Texas/4199-2/1998(H3N2)]. Although these viruses could have acquired a swine-origin segment, the branch lengths are quite short for 9 years of evolution. Therefore, we have withdrawn these 4 isolates from GenBank and subsequently retract this letter.

In the EID publication, they even discussed possible contamination. Did they overlook something?

...
Inasmuch as we identified a swine lineage virus in waterfowl, we first investigated laboratory contamination by using trace back and history of swine virus isolations during the time the surveillance samples were processed. No H3N2 subtype were isolated from swine sources in the Minnesota Veterinary Diagnostic Laboratory during this period. Furthermore, phylogenetic analysis of all HA segment sequences from isolates obtained in that 4-month period confirmed no contamination. We then investigated whether an ecologic niche existed for potential exposure of waterfowl to pigs. We identified a swine herd near the wildlife refuge area where the waterfowl sampling occurred. Pigs were housed outdoors, and the owner of this swine herd reported that geese and ducks inhabit the water ponds/stock dams/slough area to which the pigs had access. Contact with the local veterinarian and the South Dakota Veterinary Diagnostic Laboratory indicated no recent reports of influenza A (H3N2) episodes in the swine herd. In addition, this herd was not vaccinated for swine influenza.
...

[gsgs]

Re: Sequences at Genbank!

lots of sequences from -USA, 2009 and 2010 today (after 4 years ! )

5 new H9N2 genomes from China,2009 the last days ,
H6N1 chicken from Taiwan

catflu and dogflu from Korea, the usual H3N2v swine from USA,
not yet searchable
the most recent searchable sequences are from Apr 9

------------------------
oops, that was yesterday. Today lots of flu-B,USA 2011
and
11 H3N2-HA1s, California,2013 and 2 from Illinois
~20 Corsica H3N2-HAs from early 2012
10 Corsica H3N2-HAs from 2013
12 India H1N1-partial HAs(~400 nucleotides) from Sept.2012

that could be interesting to compare them with Europe H1N1, 2012/3
but my priority is currently on H7N9

------------edit.-----------
now available, 52594 ****** sequences meanwhile (15.Apr.2013)
no H1N1 form Europe 2012-13 yet available
lots of H5N1, Bangladesh 2012 today (148:MNG09,2345:Fuj,7 Z)
5 >A/chicken/Bangladesh/12VIR-7140-13/2012/04/03,H5N1
20: 23, 38, 67, 21, 30, 53, 21, 18 A/avian/Index-MNG/2009(H5N1)
84: 44, 28, 34, 55, 18, 41, 23, 30 Index Fujian,2005
79: 48, 39, 84, 66, 21, 59, 21, 40 A/Ck/HK/YU22/2002,Z
...: 12, 14, 18, --, 07, --, 14, 29 bird-index

[gsgs]

Re: Sequences at Genbank!

13 new genomes from India, 2011-2013
all are the England/2011 strain

some pseudo-recombinations, suggesting sequencing errors, e.g. segment 1 in P121778

http://www.ncbi.nlm.nih.gov/nuccore/KF280700

Submitted (24-JUN-2013) Influenza, National Institute of Virology,
20-A, Dr. Ambedkar Road, Post Box No. 11, Pune, Maharashtra 411001, India


more evidence that seasonal flu comes from India :
HA-mutations A898G,A966G,T1041C were first seen in India

                                                     0000000000000000000000000000000000000000111111111111111111111111111111112222222 0000000000000000000000000000001111111111111111111111111111111111122222222 00000000000000000000000000000000000000000111111111111111111111111111111111111111111222 00000000000000000000000000000000000000000000011111111111111111111111111111 000000000000000000000000000000111111111111111 0000000000000000000000000000000000000000000111111111111 00000000000000000000000000000000 000000000000000000000000000000000000 
                                                     0011122333333444455666677777888888999999000011112223344455555666777799990111111 0000111122222334444456666788990000111112223333344455555778888889900000111 01112222333444555555667777788888999999999000001122234445555556666666777777777889999001 00111222333333444445555566666667778888888999900000011112233444445555666666 111122222233344444556888889999000111222333344 0011111222333334444445555666677777777788999000001112233 00000222233333344455556666777999 000000011111222333333344445666677777 
                                                     2769924112488001248556668889467779245666134645895770624523469689158925599134689 3348456901489664667800367848390349236890350678901201345330119990500077889 94690589347689004688680013511237013456678227888933410272458992455689122334789242578371 29445579013455013673367900458991350114569236802455701573528000360348112255 226923888905504559474002693668588449149256809 3600237349144591223574799277900223445716256245580334915 02337367912245679957780368458247 145668803566066133689912585116800022 
                                                     1516829121847187093476923695013686787369704092892595726213969505956463591520301 0621198549957067124345065645698818011619944613844595387470566895506806140 97584088658228473157377878509213084540388374253406944844217095723578612470621483316735 29144238180017875588909858082075267677278976786136139810466378746361047836 692584258895724092608476470367969137283962913 7604617173602468085925359704402015276094111642957148477 60075870184924342853450259701539 224371888903448523751646398471537902 
-codon-position--------------------------------------  21   2     2    1             22       1 1   21 1 2     1       1   21  21 21                 2    2    1       1  1    21    1          1 2  2221           1 1    2      2  2   2       1     1 1 221   1   1      2 2  1  2                 1   1  11  1      1122 122111 11    11  121        2  1    2 1    1 2 12            1 1           1         1          2 1 12 2  1211   1 2  12 12 1211  1 1  2    1  2 2    1 1  2 1 1     2        1  222  1111     1 1   11  1 2 111      2211121 2 
---Index---------------------------------------------GGGAGATCATATCAATAGGCACATCTTCCAAAGGCGTATCGGCAGGGTCCCGAGAAAAGAAGGACTGAACAGGCAAGGG ATGTCAGGGAATCTACGCTAGAAGTTCCTCCCAAGCGCAGCCTCGACGTCTTCAAGGTCACGTACTAAGCGCC CGCTGCCGCCAGAGGAAAAGGTTCCTAGGTTTGTATATCTAGGGAGAAATTCCCAGTGCGGTCGGCCAGAGCGATCGCGGGGGAGT CATCGTCTAGAGGAGAGTAAAGATGCGTAAAGGGACGATCAACAGGATCTGAGGGAAGAGGCCCCAGTTCCTTC AGAGTCACAGGGTTTAGCGAGAACGCCCAGTGCCGACGGAGACGG GAAGTACAGCGGGGCCAACCCGAGACTGGGATGAAAAGTCCAAAATCCCGCTACG TTCTGGCGTAGAGCTGGTGGAGGAAAGATATG CCCAGGGCATAGCCCCTAAGAGTGTAAAGAGGTATC 
   1 A/******/index/2009/02/01                       ............................................................................... ......................................................................... ...................................................................................... .......................................................................... ............................................. ....................................................... ................................ ....................................        0      0 ,      0      0       1:A/******/index/2009/02/01                       
   5 >A/India/P1114854/2011,2011/09/23,India,H1N1    .........C.A.........T.....T.....A.AC...AA.TA.......G.GT............G...AT..A.. ........A...................C.T..G...T..T.....TA............A...T.......T T...AA.A............AC.....A..CC......A.G.....................A...................A... ......T....A.G........G.C..A............G..G..GC.C....A...GA.T....A....... .........A........................A..A.G...A. .G........A.........T.....C.A..CAC......T.......AA.C... .....AT...A.A...A.....A......... .............T....G...C..G.G...AC...       88     88 ,     88     88       5:>A/India/P1114854/2011,2011/09/23,India,H1N1    
   6 >A/India/P121716/2012,2012/02/24,India,H1N1     .........C.A.........T...C.T.....A.AC...AA.TA.......GTG...A......C..G...AT.GA.. ........A..A................C.T..G...T........TA............A...T.......T T...AA.A............AC.....A..CCA..G..A.G...............C.....A.........A.........A... ......T....A..........G.C.AA.G..........G..G..GC.C....A.G.GA.T....A....... .........A........................A..A.G...A. .G........A.......T.......C.A..CA.G.....T..GGAT.AA.C... CGTG.AT...A.A.C.A.....A......... ........G....T...GG...C..G.G........      107    107 ,    107    107       6:>A/India/P121716/2012,2012/02/24,India,H1N1     
   7 >A/India/P121717/2012,2012/02/24,India,H1N1     .........C.A.........T...C.T.....A.AC...AA.TA.......GTG.......CT.CT.G.T.ATGGA.. ...........A................C.T..G...T........TA............A...T.......T T...AA.A............AC.....A..CCA.....A.G...............C......AA.T..GTA.G........A... ......T....A..........G.C.AA.G..........GG.G..GC.C.T..A.G.GA.T....A....... .........A..................C.....A..A.G...A. .G........A.......T.......C.A..CA.G.....T..G....AA.C... CGT..AT...A.A.C.A.....A......... ........G....T...GG...C..G.G........      113    113 ,    113    113       7:>A/India/P121717/2012,2012/02/24,India,H1N1     
   9 >A/India/P121778/2012,2012/02/24,India,H1N1     .....G............A..T.........GA...C....ATTA.......G.G.......CT...TG...AT..AC. ........A..A..G.A.....G.C....T..G....TG...C......T..........A...T.......T T...AA.A..........G.AC.....A..CCA.....A.G............A..C.....A...................A... ......T....A..........G.C.AA.G..........G.GG..GC.C..A.A...GA.T....A....... .........A........................A..A.G...A. .G.......TA.......T.......CAA..CA..GG...T..G....AA.C... .....AT...A.A.C.A.....A......... ........G....T...GG...C..G.G.......A      107    107 ,    107    107       9:>A/India/P121778/2012,2012/02/24,India,H1N1     
  10 >A/India/P121939/2012,2012/02/29,India,H1N1     .........CGA.........T...C.T.....A.AC...AA.TA.......G.G...A...C.....G...AT..A.. ........A..A...........A....C.T..G...T........TA..CC........A...T.......T T...AA.A............AC.....A..CCA.G...A.G..........T....C.....A...................A.A. ....A.T....AA.........G.C.AA.G..........G..G..G..C....A.G.GA.T....A...AA.. .........A...C.......G............A..A.G...A. .G........A.......T.......C.A..CA.GGG...T..G....AA.C... .....AT...ACAA..A.....A......... ........G....T...GG...C..G.G........      111    111 ,    111    111      10:>A/India/P121939/2012,2012/02/29,India,H1N1     
  12 >A/India/P131845/2013,2013/02/15,India,H1N1     .TA.A.............A.GTG.......TGA...C....ATTAAA.....G.G..G..C.......G...A...A.. ..A..GA.......G.A.C...G...TT.T.T..AT.TG...C.A.......T...ACTCG..G.....T..T T...A..A......A.....AC.....AA..CAC....A.G...GA.C.........ATA..A..T.....T....A.....A... ..C...T....A.....C.C..G.C..AG..A...TA...G..GA..C.C..CAAG.TGA.T.G..A.....C. .A......CAA....GA..TA...C.TT.AC.T.AG.AA..G... ..GA.G.............T....GT......A.........G.....A.T.G.. .....A..........A...G.ATGG.GCG.. .TT....A.C.AT.AT..G.GAC...GG.G...G..      158    158 ,    158    158      12:>A/India/P131845/2013,2013/02/15,India,H1N1     
  13 >A/India/P132194/2013,2013/02/18,India,H1N1     .T.....TGC....GC.....T....CT.....A.AC...AA.TA.......G.G..G..C.......G...A...A.. ...C....A.G.T....A..........C.T..G..AT...T....T.C...........A...T.....AAT TA..A..A.....AA..G..AC.....AA..C......ACG.........C....A..T...A..T..A...........A.A... .....CT..A.A...G.C.C..G.C..A............G..G...C.C...AAG..GA.T....A.....C. G...CT..CAAA..CGA..TA..T..TT.AC.TTA..A...G... AG....T.T.AAA........C....C....CAT...AA.T.......AA.C..A .....A..........A.....A.GG.GC... T....A.......T....GA..CA.G.G..A.....      144    144 ,    144    144      13:>A/India/P132194/2013,2013/02/18,India,H1N1     
   2 >A/India/Nag132467/2013,2013/01/31,India,H1N1   A..G..C.....T...GAAT.T.......G.GA.T.C....ATTA.....A...G......ACTT.......A.G.A.A ..A.T.A..C...CGAA...A.G......T.......TG...CT.........G....ACA..G........T T.......T..AT..G....A...TCT....C....G.A..AAA...........A....A.....TG......C..T.A..A..C ........G.....A...G.T...C.AA.....A.T...T.....A..TCA...A....A.T....A....... .........AA.C......T..G...TT.AC.T.A.TA..A.... ................................A...............AA..... ....AA.A.....T.AA.AT.AA..G.GC... .TTG.........TATC.G...C....G...AC...      133    133 ,    133    133       2:>A/India/Nag132467/2013,2013/01/31,India,H1N1   
   3 >A/India/Nsk12388/2012,2012/01/19,India,H1N1    ..................A..T.........GA...C....ATTA.....A...G......A..T.......A...A.A ..A...A.......G.A.....G.C....T..G....TG...C......T....G......CCG.GCC....T T.....T.TTGA...G....A........G.C......A...AA..T.GC...A.A.....C.AA.T.......C..TA..AAG.C AG.T....G.....A...G.....C.AA..G.A..T.............C....A....AATT..G...T...T ......G..AA.....A..T....ATTT.ACAT.A..A....T.. ....CG......AA..G.......G....A.CA.G............TAAT.... .....AT..G......AC....A..G.GC.C. .TT....A.C..TTAT..G...C....GA...C...      139    139 ,    139    139       3:>A/India/Nsk12388/2012,2012/01/19,India,H1N1    
  14 >A/India/VD122268/2012,2012/03/07,India,H1N1    ..................A..T.........GA...C....ATTAA....A...G......ACTT....A..A...A.A .CA...A.......G.A.....G.C....T..G....TG...C......T..........A..G........T T.....T.TT.A...G...AA..T.....G.C......A...AA..T.....AT.A........A.T.......C..TA..AAG.C A..T....G.....A...G.....C.AA..G....T.............C....A....AATT..G...T...T ...A..G..AA.....A.AT....A.TT.ACAT.A..A....... ....CG..................G....AGCA.G....A........AAT.... .....AT.C.......AC....A..GAGC.C. .TT.T..A.C..TTAT..G...C....GAG......      135    135 ,    135    135      14:>A/India/VD122268/2012,2012/03/07,India,H1N1    
   4 >A/India/P1112874/2011,2011/08/10,India,H1N1    ..................A..T.........GA...C....ATTA....TAA..G......A..T.......A...A.A ..A...A.......G.A..G..G..A...T.......TG...C............A....A..G........T T.......T..A...G....A..........C......A...AA.........T.A..........T.......C..T....A..C ......T...G...A...G..A..C.AA.......T.............C....A....A.TT......T...T ..G....A.AA.C......T......TT.AC.T.A..A......A .....G..................G..........GG...........AAT.... .....A..........A.....A..G.GC... .TT......C...TAT..G...C.C..G.G.AC...      105    105 ,    105    105       4:>A/India/P1112874/2011,2011/08/10,India,H1N1    
  11 >A/India/P12946/2012,2012/02/10,India,H1N1      ................G.A..T.AA...T..GA...CGCG.ATTT...T.A...G......AT.T.......A...A.A ..A.T.A.......GAA....TG......T.......TGA..C..G.....C.G......A..G....A...T T.......T..A...GG...A..........C.....CA...AA.........TCA....A.....T.......CT.T.A..A..C A..T...C......A.A.G.....C.AA......GT.T...........C....A....A.TT.T..CCT...T .........AA.C....T.T..G...TT.AC.T.A.TA......A A....G.G.........G....CAG......CA.G......C......AAT..T. .....A.........AA.AT..A..G.GC..C .TT...A...G..TATC.G...C....G.G......      141    141 ,    141    141      11:>A/India/P12946/2012,2012/02/10,India,H1N1      
   8 >A/India/P121773/2012,2012/02/24,India,H1N1     .............G....A..T.........GA...C....ATTA..G..A...G.G..C....T......AA...A.A T.A...AA......G.A.....G.C....T..G....TG...C......TCC........A..G........T T.TG..T.TTGA...G....A.A......G.C......A...AA..T......T.A........A.T.......C..TA..AAG.C A..T...CG.....A...G....CCTAA..G....T..C...............A....A.TT..G........ ...A..G..AA.....A..T....A.TT.ACAT.A..A......A ....CG....A...TT........G....A.CA.G.............AAT.... .....AT.........AC....A..G..C.C. .TT....A.C..TTAT..G...C....GAG....C.      137    137 ,    137    137       8:>A/India/P121773/2012,2012/02/24,India,H1N1

[gsgs]

Re: Sequences at Genbank!

****** HAs since July 2012
2 out of 150 are non-Cancun
4 out of 150 are non-UK-Europe/2010/11

the new variant (lower part in the pic) was in
Kenya,Casablanca,New York,Brazil,Florida,Beijing,
Ohio-swine,Mississippi,...

[gsgs]

Re: Sequences at Genbank!

119 H1 genomes from 2012/3 from Finland

>~90% are from one main-strain, descending from the England/India/Europe
strain from 2010/11

for the main strain I count these 134=22+19+25+18+17+11+08+14 mutations:

1(22):G161A,G198A,G583A,C666T,A873T,A876G,G878A,T957C,G1030A,
   C1044T,A1060T,G1149A,G1152A,G1188A,C1279A,A1446G,C1467A,
   G1665A,C1719T,G2091A,G2163A,G2191A
2(19):G42A,G168A,A366G,G461A,T474C,A630G,G810A,C885T,C999T,
  C1038T,C1131T,C1186T,A1191G,T1304C,C1896A,A1905G,C2076T,
  G2157A,C2190T
3(25):C99T,C336T,C345T,T435C,G462A,A507G,G663A,T873C,C963A,C1005A,
  G1027A,G1074A,T1170C,A1194C,G1329A,C1428T,G1524A,A1632G,C1687T,
  T1776C,C1824T,A1935G,G1986A,T2115C,G2151A
4(18):T30C,C273T,G340A,A579G,G605C,T658A,G751A,A898G,A966G,
  T1041C,T1056C,G1158A,G1171A,A1230G,A1386G,G1403A,C1408T,G1546A
5(17):A288C,A294G,G298A,G309A,A450G,G459A,A570T,G648A,C930T,
  C963T,G987A,T1059C,C1089T,G1143A,G1248A,A1362G,G2149A
6(11):A131G,A528G,A599G,G615A,C627T,T720C,G721A,A742G,
  C1107A,G1131A,C1134T
7(08):G238A,G459A,G492A,A574G,G600A,A689G,A750G,T781C
8(14):C42T,C54T,C108A,T159C,C204T,C264T,C268A,C315T,A367G,
  T414C,C429A,A614G,C640T,A661G


59,>A/index/******/UK1/2011(H1N1)
    C462A(1),C666T(1),C795T(1),G896A(1),T957C(1),G1030A(1),A1060T(1),G1149A(1),T1266C(1),A1367G(1),A1446G(1),G1797A(1),G2091A(1),G2163A(1)
    T936C(2),A1098G(2),C1186T(2),C1398T(2),C1896A(2),C2190T(2)
    C99T(3),G204A(3),G663A(3),T687C(3),G691A(3),T873C(3),C963A(3),A988G(3),T1548C(3),G1986A(3)
    G340A(4),A579G(4),G605C(4),T658A(4),T1056C(4),G1171A(4),A1386G(4),G1403A(4),C1408T(4)
    G298A(5),C468A(5),G1143A(5),G1248A(5),G1293A(5),A1329G(5)
    T56C(6),G316A(6),T720C(6),G721A(6),A742G(6),C1107A(6),G1131A(6),T1248C(6)
    G238A(7),G492A(7),G600A(7)
    C264T(8),A367G(8),T414C(8)
63,>A/index/******/UK2/2011(H1N1)
    C666T(1),T957C(1),G1030A(1),C1044T(1),A1060T(1),G1149A(1),A1446G(1),G1665A(1),G2091A(1),G2163A(1)
    A366G(2),A630G(2),C999T(2),C1186T(2),A1191G(2),T1304C(2),C1896A(2),A1905G(2),C2190T(2)
    C99T(3),C336T(3),G462A(3),A507G(3),G663A(3),T873C(3),C963A(3),G1027A(3),C1428T(3),G1986A(3),T2115C(3)
    C144T(4),A478G(4),G605C(4),G640A(4),T658A(4),T1056C(4),G1171A(4),G1403A(4),C1408T(4),C1437T(4),C1656T(4)
    G298A(5),C930T(5),C963T(5),G987A(5),T1059C(5),G1143A(5),G1248A(5),A1329G(5)
    A131G(6),G316A(6),T720C(6),G721A(6),A742G(6),C1107A(6),G1131A(6)
    G238A(7),G492A(7),G600A(7)
    C264T(8),C268A(8),A367G(8),T414C(8)

amino-acid mutations:
UK/2011:
1:V344M,I354L
2:I397M,I435T
3:N321K,A343T
4114N,S202T,S220T,E391K,S468N
5:V100I
6:N44S,V106I,V241I,N248D,N369K
7:V80I
8:L90I,I123V

additional amino acid mutations in FIN/2013:
1:R54K,M66I,D195N,R293K,V731I
2:G154D
3:-
4:V251I,K300E,E516K
5:S498N
6:V106I reversed,N200S
7:M192V,K230R,D21G(M2)
8:N205S,T48A(NS2)

with 5 new amino-acid mutations in PB2, they might recheck for the
danger of E627K again


no D239G (D225G in H3-numbering)


segments 1,2,3,5,6,8 are descendent from UK2, while segment 4 is from UK1,
segment 7 can't be decided, it could be from either (but probably also from UK2).

G316A(6), one of the 11 Cancun mutations had been undone here, it mutated again
and now it's again, how it originally was before April 2009
One further of the 61 UK/2011 mutations is missing : A1329G(5),
that one could also have been undone or it happened (in India ?)
after the separation of the UK-strain.
However, 59 of 61 mutations are present, so the common ancester of UK/2011 and Fin/2013
is pretty close to UK/2011,only days or weeks away.
You could even assume it's a direct descendent and pH1N1 survived in Europe or went back to Asia.

All (except 6) the genomes are supposed to be from early 2013 or late 2012,
but the distribution of mutations (see picture hels1.gif) suggests
that some sequences could be from 2011/12 and even 2010/11.
[i.e. this one : http://www.ncbi.nlm.nih.gov/nuccore/KF560221
only 12 mutations --> probably not from 2013 --------- similar to the vax-strain CA/09
contamination or live vax]
[or this one : http://www.ncbi.nlm.nih.gov/nuccore/KF559901
only 32 mutations --> probably not from 2013 ]

Or they survived long periods without mutating in the environment,
but that would be the first such discovery for seasonal flu.
Or there are sequencing errors, the sequences were processed
in the same run and the machine recombined them - we had seen
that before and in fact there are some such indications here too,
I just looked at it briefly so far.

the percentage of nucleotides from {A,T} is up to ~55.70 from 55.28 in 2009
(Cancun:55.36,UK2/2011:55.48,Brevig/1918:54.84,triple-reassortant swine/1998:55.09)