| H5N1 in Cats in Indonesia Recombinomics Commentary October 7, 2006 A study conducted by the Indonesian Environment Information Center (PILI) in Yogyakarta found that stray cats had caught the H5N1 virus through contact with infected poultry at traditional markets. The above comments provide additional evidence implicating an alternative source of H5N1 human infections in Indonesia. Recently, Indonesia summarized the results of poultry tests by a WHO affiliated lab in Australia (http://www.recombinomics.com/News/10050602/H5N1_Indo_Match_Failures.html). Concerns had been voiced in an H5N1 meeting in Jakarta in June because the sequences from the vast majority of human cases on Java did not match the poultry H5N1 collected between 2003 and 2005. Consequently, 91 poultry samples (http://www.recombinomics.com/News/08120601/H5N1_Match_Unlikely.html) were sent to Australia for virus isolation and sequencing. The samples were from collections between September 2005 and March 2006 and failed to match (http://www.recombinomics.com/News/08110601/H5N1_Match_Failure.html) Java poultry isolates with most of the sequences from patients. All but one of the human isolates from patients on Java had a novel cleavage site, RESRRKKR, as well as a number of associated changes in all 8 gene segments that readily distinguished the human sequences from the poultry sequence (see green sequences in lower branch (http://www.recombinomics.com/phylo/Indo_Karo_HA1.html)). One duck from Indramayu (http://www.recombinomics.com/News/08310601/H5N1_Indo_Duck_Cleavage.html) had the novel cleavage site, but that duck isolates had additional changes, including a silent change in the cleavage site, which was found in only a small subset of human isolates. The vast majority of human isolates, including those from Indramayu, failed to match the duck sequences. Two matching sequences were found in chickens in central Sumatra (http://www.recombinomics.com/News/09140601/H5N1_Sumatra_Chickens.html), but the match failure on Java, coupled with the almost universal matching of human sequences, including the first sequence isolate in July 2005 as well as isolates from a wide range of geographical locations, including East Java, suggested human infections were largely due to H5N1 in a reservoir other than poultry. The only match on Java of the human sequences was from a throat swab of a cat. A/feline/Indonesia/CDC1 (http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=nucleotide&list_uids=113495213&dopt=genbank)/2006(H5N1), from Jakarta on January 22, 2006. H5N1 has also been detected in swine in Indonesia, but the swine sequences, including the HA cleavage site, did not match the human sequences. The discovery of more H5N1 in cats in Indonesia raises the possibility that the cats are also in contact with an alternate reservoir. Sequence data on H5N1 from cats, and “infected poultry in traditional markets”, including those in and around Jakarta, where most of the reported human cases have been located, would help resolve the role of cats in H5N1 transmission. Media Sources (http://www.flutrackers.com/forum/showthread.php?t=11352) Phylogenetic Trees (http://www.recombinomics.com/phylo.html) http://www.recombinomics.com/News/10070601/H5N1_Indonesia_Cats.html | |||||||||
Recombinomics Commentary
October 19, 2006
In the past few weeks, the Beijing Genome Institute has released a series of complete H5N1 sequences from poultry, wild birds, and swine in China. Most of the sequences are from isolates collected between 1997 and 2005 in northeastern China. Many sequences were initially deposited at GenBank on February 28, 2005 under the title, “A cohort of AIV H5N1 subtypes isolated from wild aquatic birds and domestic poultry revealed rapid transmission, frequent reassortment, and identifiable recombination events.” Those sequences were made public on February 2, 2006 and had many clear cut examples of recombination (http://www.recombinomics.com/News/03090602/H5N1_Recombination_Guangdong_NA.html).
The current series confirms (http://www.recombinomics.com/News/10050603/H5N1_Recombination_Confirmation.html) the earlier sequences and provide a roadmap of recombination events (http://www.recombinomics.com/News/10120601/H5N1_China_Evolution.html) between 1997 and 2005. These sequences define the evolution of H5N1 in China and highlight the critical role of recombination and migratory birds (http://www.recombinomics.com/News/03060605/H5N1_Recombination_Henan_PB2.html). The sequences trace back to low path isolates in Hong Kong in the late 1970’s and show how H5N1 is evolving (http://www.recombinomics.com/News/10060601/H5N1_H7N1_PB1_Recombination.html) since the first reported sequence from a 1996 goose in Guangdong.
Of interest are two H5N1 swine sequences from Guangxi and Henan provinces. These sequence demonstrate the transmission and transport of H5N1 genetic information into the Qinghai strain, which has now been transmitted and transported to most countries in Europe, the Middle East, and Africa. This strain is recombining with wild birds and poultry (http://www.recombinomics.com/News/10060601/H5N1_H7N1_PB1_Recombination.html) in China and swine appear to add additional selection and acquisition of mammalian polymorphisms.
The swine sequences can be found in dog, cat, and wild cat isolates from Thailand, as well as human isolates in Thailand, Vietnam, China, and Indonesia. The movement of pieces of genes into Indonesian patients, including those on Java as well North Sumatra (the Karo cluster) is striking, and provides additional insight into the evolution and adaptation of H5N1 in mammals. Some of the swine data is discussed here (http://www.flutrackers.com/forum/showthread.php?t=11718) and here (http://www.flutrackers.com/forum/showthread.php?t=11774).
The data demonstrates the value in a database of full influenza sequences. These data add considerably to H5N1 evolution in China and the spread to southeast Asia and Indonesia. The data also demonstrate movement of sequences to and from North America.
The data clearly show that H5N1 is evolving vioa recombination in a predictable manner that involves migratory birds, dual infections, and exchange of small and large bits of genetic information via recombination.
The database however, still has a large number of holes. Much of the prior sequences of H5N1 in Hong Kong, China, and southeast Asia are partial sequences, or sequences for one or two of the eight gene segments. The filling in of those holes would create a more robust database. The NIAID has a flu sequencing project (http://www.niaid.nih.gov/dmid/genomes/mscs/influenza.htm) which will generate full sequences on all eight gene segments at no charge. The first H5n1 sequences from that project were also released for the first time in the past several weeks. In addition, full sequences from human isolates in Indonesia have been released ad the number of poultry sequences for one of the eight gene segments has also been increased. However, sequences collected in Europe beginning in October of 2005 have been withheld as have H5 sequences collected in Canada. These large sets of withheld data also create large holes in the sequence database.
In addition to withheld data, there are also significant limits on collections. The data from Indonesia indicate the vast majority of poultry H5N1 isolates do not match the human isolates. However, there are no swine or wild bird sequences from Indonesia. There is one cat sequence, which does match the human sequences, and media reports indicate additional cat positives have been found, but none of the sequences have been released.
Similarly, Canada and the United States have now isolated H5 from wild birds in 2005 and 2006, but thus far only 4 of 8 sequences from one H5N2 sample, collected over a year ago in August, 2005 (http://www.recombinomics.com/News/11190504/H5_LPAI_Quebec_British_Columbia.html) in British Columbia has been released. It has evidence of recombination and acquisition of North American swine H1N1 (http://www.recombinomics.com/News/07190601/H5_BC_Mallard_Swine.html) sequences and Asian H5N1 Qinghai sequences, demonstrating rapid evolution of low path H5 in North America. Full sequences of the recent H5N1 isolates in Canada and the United States would provide information on nthe source and rate of this evolution and provide more information on the false negatives recently announced by the USDA. (http://www.recombinomics.com/News/10180601/H5N1_USDA_Failures.html)
The predictable evolution of H5N1 is now abundantly clear, and the release of full sequence data is long overdue.
Media Sources (http://www.flutrackers.com/forum/showthread.php?t=11718)
Phylogenetic Trees (http://www.recombinomics.com/phylo.html)
http://www.recombinomics.com/News/10190601/H5N1_China_Swine_Evolution.html
| Mammalian H5N1 Evolution in Indonesia Recombinomics Commentary October 7, 2006 The recent announcements of H5N1 bird flu in cats in Indonesia (http://www.recombinomics.com/News/10070601/H5N1_Indonesia_Cats.html), coupled with results from expanded sequencing of poultry (http://www.recombinomics.com/News/10050602/H5N1_Indo_Match_Failures.html) strongly signal the existence of a separate mammalian H5N1 reservoir in Indonesia that is responsible for the vast majority of human cases. This separate reservoir creates significant problems, because most of the attention has focused on infected poultry, and the mammalian reservoir has been significantly under investigated and under reported. Therefore, a review of the evolution of this mammalian reservoir is useful. Although Indonesia did not acknowledge H5N1 infections until 2004, the first isolates were from birds in 2003. Sequence analysis of the 2003 and 2004 isolates indicated the H5N1 was Clade 2 and had a number of genetic markers that were specific for Indonesia. The first human case was reported in July of 2005, and the sequence of the isolate, A/Indonesia/5 (http://flu.lanl.gov/search/view_record.html?accession=ISDN125873&database=fluA)/2005 had the Indonesia specific markers, but also had a number of unique polymorphisms, including one that created a novel HA cleavage site (http://www.recombinomics.com/News/03260601/H5N1_Vaccine_Indo_Sequence.html), RESRRKKR. However, the second human isolate, A/Indonesia/6/2005 had the more common HA cleavage site, RERRRKKR, and was similar to bird isolates. Subsequent human isolates in 2005 had the novel cleavage site, but several had an additional silent change, so although the cleavage site matched the first sequence at the protein level, there were a number of changes at the nucleotide level that divided the sequences with the novel cleavage site into two groups. However, as the number of sequences increased in early 2006, it became increasingly clear that the human sequences were separating from the poultry sequences, all of which had the common bird cleavage site. The first match of the human sequences was from H5N1 from a throat swab of a cat (http://effectmeasure.blogspot.com/2006/02/indonesia-and-questions-about-its.html) in Indramayu near a residence were two siblings died from H5N1 infections. The sequence of the cat isolate not only matched the more recent human sequences, but was very close to the sequences of the isolates from the two siblings, A/Indonesia/283H/2006 and A/Indonesia/286H/2006, as well as other human isolates from Indramayu, A/Indonesia/292H/2006 and A/Indonesia/304H/2006. New isolates in 2006 collected from patients in East and West Java were sequenced and all were matches of more recent sequences such as the four human and one cat isolate from Indramayu. In May of 2006 however, there was a new cluster in the Karo regency in north Sumatra. This outbreak was the largest to date and involve secondary and tertiary transmissions of H5N1. Consequently a meeting was call by WHO and consultants in Jakarta in June. Included in the presentation was a phylogenetic tree (http://www.recombinomics.com/phylo/Indo_Karo_HA1.html) that summarized the H5N1 Indonesian isolates as of June 12, 2006. The tree, which had the human sequences in green and the Karo cluster shaded in pink, clearly showed the match problem. All of the sequences with the novel cleavage site were on the lower portion of the tree and there were no poultry sequences on these two lower branches. Moreover, all of the recent human sequences from Java were on the lowest branch, which was even further from the poultry isolates. Thus, the human isolates were evolving away from the poultry isolates, suggesting the existence of a separate mammalian reservoir. However, all of the human isolates were from July, 2005 or later, while most of the bird isolates were from earlier dates. Therefore 91 samples (http://www.recombinomics.com/News/08120601/H5N1_Match_Unlikely.html) were schedule for shipment to Australia for virus isolation and sequencing. The samples were from infections between September, 2005 and March, 2006. As sequences from these more recent and geographically dispersed isolates began to be published, it was becoming increasingly clear that the vast majority of the human infections on Java were not from domestic poultry (http://www.recombinomics.com/News/08120601/H5N1_Match_Unlikely.html). Each human sequence mapped to the lower portion of the tree and which was more distinct from the bird sequences. The second set of new bird sequences included an isolate with the novel cleavage site. It was from a duck on Indramayu (http://www.recombinomics.com/News/08310601/H5N1_Indo_Duck_Cleavage.html) isolated in 2006. However, that isolate match the upper branch of the human sequences, which were composed of six isolates from three patients in 2005. Thus, although every human isolate in 2006 was matching the lower branch, the one duck sequence matched the upper branch. The third set of poultry sequence had two matched with the lower branch. However, the two matches were from chickens in central Sumatra from 2005. Thus, none of the poultry isolates matched the lower human branch, while all human isolates, as well as the cat isolate, matched the lower human branch. These data again supported a separate reservoir for the human sequences, and the only matches on Java were from the one cat, and all human isolates. The recent announcement indicates more H5N1 has been detected in cats, but the sequences of those isolates have not been released. Swine H5N1 sequences have been reported, but none match (http://www.recombinomics.com/phylo/Nidom_swine_HA.html) the human sequences. The match failures pose a major problem because testing of humans is largely limited to patients how have been near dead or dying poultry. However, the poultry association has not been linked to the human infections, so an expanded testing of patients with symptoms is warranted. Similarly, more sequencing of H5N1 from other reservoirs is warranted by the match failures between mammalian and avian sequences. Media Sources (http://www.flutrackers.com/forum/showthread.php?t=11352) Phylogenetic Trees (http://www.recombinomics.com/phylo.html) http://www.recombinomics.com/News/10070602/H5N1_Indonesia_Mammalian.html | |||||||||
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