Forum: Final Adaptation of H 5 N 1 to Humans Role of Mammalian Reservoirs 5

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ANON-YYZ – at 10:15 anon_22 – at 10:09

“I think the biggest near-term risk is a new reassortant from south China. This is still ongoing, with signs of increasing virulence towards mammals. “

More details, links? I must have missed some news from China.

The evolution of H5N1 influenza viruses in ducks in southern China

“The transmission of H5N1 ‘‘bird flu’’ to humans in 1997 first established the ability of avian influenza viruses to be transmitted to humans despite their preferential binding to avian sialic acid receptors (i.e., those with a terminal
2,3Gal linkage) (14). Before 1997, there had been isolated reports of human infection with H7N7 influenza virus (usually causing conjunctivitis) (12), but there was no convincing evidence of repeated transmission of avian viruses to humans. How did avian influenza viruses come to acquire a progressively greater capacity to infect mammals? Here, we characterize a series of 21 H5N1 influenza viruses isolated from apparently healthy domestic ducks in coastal provinces and cities of southern China from 1999 through 2002. These isolates were highly lethal to chickens and demonstrated the progressive acquisition of pathogenicity to mice (a mammalian host). They were genomically heterogenous, having acquired multiple gene segments and deletions in their nonstructural (NS) and neuraminidase (NA) genes. We propose a hypothetical mechanism to explain the selection of H5N1 viruses with increasing pathogenicity to mice.”

“Our results demonstrate that while circulating in domestic ducks, H5N1 viruses gradually acquired the characteristics that make them lethal in mice. One possible explanation for this finding is the transmission of duck H5N1 viruses to humans, the selective evolution of the viruses in humans, and their subsequent transmission back to ducks.”

“An alternative possibility is involvement of the pig (the postulated intermediate host) in selection. In the region of China where the H5N1 viruses characterized in this study were isolated, pigs and ducks are housed in close proximity, especially in farming villages, where families typically own a small number of pigs and ducks. Our working hypothesis is that H5N1 viruses have gradually acquired the ability to replicate in mammals by means of selection pressure created by possible transmission between pigs and ducks. To date, there are no reports of the isolation of H5N1 viruses from pigs, although pigs have been experimentally infected with H5N1 viruses (33). We have recently obtained preliminary virological and serological evidence of H5N1 virus infection of pigs in Fujian province.”

H5N1 reassortants 1999 to 2002

H5N1 reassortants show increasing virulence in mice

In the table for virulence in mice, the column to look at is the ‘pathotype’, which shows the trend in increasing virulence.

Another study offers a different but equally disturbing insight.

Are Ducks Contributing to the Endemicity of Highly Pathogenic H5N1 Influenza Virus in Asia?

“In the past, most avian influenza viruses were found to preferentially replicate in the gastrointestinal tract of wild ducks, to be excreted at high levels in the feces, and to be transmitted primarily via the oral-fecal route (15). However, in the current study, virus was excreted at high levels in the trachea (upper respiratory tract), rather than in the cloaca. We observed similar results in experimentally inoculated ducks and ducks infected by contact with birds inoculated with H5N1 viruses isolated in late 2002 from Hong Kong (24). Analysis of the combined data from our studies shows that there is a significant difference in levels of tracheal virus shedding and cloacal virus shedding in ducks, particularly in those infected by recent H5N1 viruses (post-2002). Although route of inoculation may affect the viral shedding pattern or organ tropism in an infected host, the same observation was made among inoculated ducks and ducks infected by contact, confirming that this was not an experimental artifact. Additionally, similar results were observed among Pekin ducks exposed to nine different H5N1 virus isolates from 2003–2004 (see Table S2 in the supplemental material). This is in contrast to previous studies, which reported that experimental infection via the natural route in domestic ducks (Pekin and Sheldrake ducks) with H5N1 viruses isolated in Hong Kong in 1997 and from the coastal province of southern China in 1999 to 2002 resulted in similar titers of virus shedding in the trachea and in the cloaca (4, 22).

“Thus, the increased level of tracheal virus shedding is a particularity of the recent viruses emerging in Asia, from late 2002 onwards, and so far has been observed in both a wild and a domestic duck species. These results indicate that the digestive tract may no longer be the main site of replication in ducks infected by recent H5N1 viruses, and this will most likely influence the natural history of the virus. Indeed, the main path of transmission may have shifted from an oral-fecal route to a more oral-oral route or even an airborne route—or a combination of all of these. This putative change in transmission route could affect the epidemiology of H5N1 viruses and may result in an increase in transmissibility. If this is in fact the case, the implications for H5N1 surveillance and control are important.”

To summarize, we are seeing:

1) frequent reassortment creating new strains

2) these new strains show increasing virulence in mammals

3) possible shift towards airborne transmission

anon_22

Disturbing indeed. The shift of replication site suggests to me a change in the HA cleavage site, a change in the HA bidning domain, or both. Expert comments are welcome.

“They were genomically heterogenous, having acquired multiple gene segments and deletions in their nonstructural (NS) and neuraminidase (NA) genes.”

“One possible explanation for this finding is the transmission of duck H5N1 viruses to humans, the selective evolution of the viruses in humans, and their subsequent transmission back to ducks.”

That one is just sitting dead in the water waiting for a broadside from Niman. Let the games begin…

Airborne between ducks.

Once that is achieved, would H5N1 have learned something that will help it to change to airborne between humans?

ANON-YYZ – at 13:51 “Airborne between ducks.

Once that is achieved, would H5N1 have learned something that will help it to change to airborne between humans?”

First of all, it will definitely increase the chance of b2h transmission.

Secondly, my guess is whether it will enhance airborne h2h will probably depend on what the difference in receptor binding is between respiratory and GI receptors in birds, and whether changes enhancing binding to avian respiratory tract receptors will enhance binding to human respiratory tract receptors.

There’s a March 2006 article that’s been mentioned several times in other threads. I was able to obtain a copy of the full text and will attempt to briefly review the basic results of the study. Title: “The polymerase complex genes contribute to the high virulence of the human H5N1 influenza virus isolate A/Vietnam/1203/04.” The results were rather shocking, at least to me, regarding the large influence of the polymerase complex genes on lethality (PB1, PB2, PA).

Background Information

The research team used two H5N1 isolates: (human) A/Vietnam/1203/04, and (avian) A/Chicken/Vietnam/C58/04. Both are of the Z genotype, both are from Vietnam 2004, and both contain the hemagglutinin multibasic amino acid cleavage sequence PQRERRRKKR|G. Both sequences are lethal to chickens. The human strain is lethal to ferrets and mice, whereas the chicken strain is not lethal to those animals. The experiments used reverse genetic procedures to replace various segments of one isolate into the other isolate, and tested ferrets and mice for illness and lethality. The study also checked other results, but the material is too extensive for this brief review.

Results

• The unaltered chicken isolate caused 0% lethality in both ferrets and mice. However, the unaltered human isolate caused 100% lethality in both ferrets and mice.
• When the HA and NA from the human isolate were knocked out and replaced with HA and NA from the (previously nonlethal) chicken isolate, there was still 100% lethality in the ferrets and mice. This is an important result and proves that focusing only on characteristics of HA and NA in terms of lethality, is probably short-sighted.
• When the HA and NA from the chicken isolate were knocked out and replaced with the HA and NA from the (previously lethal) human isolate, there was still 0% lethality in the ferrets and mice.
• When the three polymerase segments (PB1, PB2 & PA) were knocked out of the human isolate and replaced with the same segments from the (previously nonlethal) chicken isolate, the lethality of the original (lethal) human strain was completely attentuated in all ferrets and mice - there was 0% lethality in the animals.
• Conversely, when the same three polymerase segments were knocked out of the (previously nonlethal) chicken isolate and replaced with the same segments from the (lethal) human isolate, it resulted in 33% lethality in the ferrets and 100% lethality in the mice.
• When the NS segment was knocked out of the (previously lethal) human isolate and replaced with the NS from the chicken isolate, the change resulted in lethality to 16% of ferrets and 100% of mice.
• When the PB2 segment was knocked out of the (previously lethal) human isolate and replaced with the PB2 from the chicken isolate, the change resulted in lethality to 33% of ferrets and 13% of mice.
• When the PB1 segment was knocked out of the (previously lethal) human isolate and replaced with the PB1 from the chicken isolate, the change resulted in lethality to 50% of ferrets and 0% of mice.

It is necessary to read the entire article to get the full impact of the study. The team also assessed culture plaque sizes of the various reassortments that were tested. They cited limitations due to the fact that they could only use a certain number of animals with each test, as the biosafety lab had limited space.

Other brief conclusions quoted from the article

• “the three human P-genes are sufficient to convert a nonlethal virus into a lethal virus in mice but not in ferrets.”
• both the human and the chicken isolates “have preferential binding affinity for alpha-2,3-linked SA [sialic acid] receptors, typical of avian viruses.”
• “Survival of all mice and ferrets inoculated with a RG [reverse genetics] VN1203 virus that had its polymerase genes derived from the chicken isolate showed that these genes are important for virulence.”
• “HA residues outside the cleavage site do not have a critical impact on tropism. Thus, VN1203 (H5N1) influenza’s systemic spread and neurotropism are not the result of changes in receptor binding…Inefficient binding and cell entry of an avian H5N1 virus in a mammalian host can be compensated by intracellular cleavage of the HA and by efficient replication/transcription of the viral genome by PB1 or PB2. Thus, multibasic amino acids in the cleavage site of the HA are necessary but not sufficient for high lethality in mammalian species.” (Please see full article for the authors’ reasoning that supports these conclusions).

What does this mean? Is it saying that it can spread without a change in the receptor binding? Laymens terms please. Thanks in advance.

“HA residues outside the cleavage site do not have a critical impact on tropism. Thus, VN1203 (H5N1) influenza’s systemic spread and neurotropism are not the result of changes in receptor binding…Inefficient binding and cell entry of an avian H5N1 virus in a mammalian host can be compensated by intracellular cleavage of the HA and by efficient replication/transcription of the viral genome by PB1 or PB2. Thus, multibasic amino acids in the cleavage site of the HA are necessary but not sufficient for high lethality in mammalian species.” (Please see full article for the authors’ reasoning that supports these conclusions).”

Commonground at 14:13,

I’m not sure what you mean by “spread” - did you mean spread of the virus within the body, or spread from one host to another?

The authors of the article in this instance, seem to be addressing the issues the cell tropism (what types of cells are vulnerable), and transmission. They stated, “contributions of receptor-binding properties to cell tropism and virus transmission are not known. Our finding that recombinant human virus containing the chicken HA retained its lethality demonstrates that HA residues outside the cleavage site do not have a critical impact on tropism…H5N1 viruses whose HA is selective for alpha-2,3-linked SA can spread efficiently after experimental infection.”

The study did look at pathological effects on the various tissues, including lungs, liver, brain, spleen, and neurons. They did not experimentally assess transmission from one host to another. However, they did look at plaque size of the virus cultures, which relates to replication efficiency of the virus. The largest pattern showed large plaque size correlated with the reassorted viruses that caused 100% lethality, which would imply efficient replication.

My take on this article would thus lead to answering your question with a qualified “yes”. The evidence from it associates higher virus replication capability with the three human isolate polymerase genes, than those from the chicken isolate.

beehiver,

Thanks for that study. Yes, this is a very important point. Interestingly and disturbingly, the findings mirror those from the polymerase genes of the 1918 virus. Characterization of the 1918 influenza virus polymerase genes, Taubenberger et al.

“A total of ten amino acid changes in the polymerase proteins consistently differentiate the 1918 and subsequent human influenza virus sequences from avian virus sequences. Notably, a number of the same changes have been found in recently circulating, highly pathogenic H5N1 viruses that have caused illness and death in humans and are feared to be the precursors of a new influenza pandemic. The sequence changes identified here may be important in the adaptation of influenza viruses to humans.”

The 5 PB2 changes, for example, are almost completely preserved in human sequences, but “Only a small number of avian influenza isolates show any of these five changes, and it is intriguing that almost all of these isolates are from HPAI H5N1 or H7N7 viruses, or from the H9N2 lineage that infected a small number of humans in China in the late 1990s (ref. 29).”

For example, “Only 5 out of 282 available avian PB2 sequences have a Ser residue at position 199, four of these being 1997 H5N1 isolates from Hong Kong.”

Even more interesting is E627K: “This residue has been implicated in host adaptation, and has previously been shown to be crucial for high pathogenicity in mice infected with the 1997 H5N1 virus. Of the avian isolates, 19 out of 345 have a Lys residue at position 627, 18 of which are HPAI H5N1 or H7N7 avian influenza viruses. Sixteen of these were recently characterized H5N1 isolates from a die-off of wild waterfowl around Qinghai Lake in western China in 2005.”

“In human H5N1 isolates, 11 out of 37 have the E627K change: A/Hong Kong/483/1997 and A/Hong Kong/485/1997, four out of six isolates from Vietnam in 2004, and two out of three isolates from Thailand in 2004. The E627K mutation was seen in six out of seven H5N1 isolates from Thai tigers in 2004, and was also present in the H7N7 virus responsible for the single human fatality during the HPAI H7N7 outbreak in the Netherlands in 2003. It was not noted in the contemporaneous chicken isolates.”

A letter (copy attached) published in the August issue of Emerging Infectious Diseases reports the finding of H5N1 HPAI viruses in cats in Iraq. The letter concludes that ¡®death in cats, spatially and temporally associated with unusual deaths in poultry, especially when the cats show positive results of a rapid antigen detection test for influenza A, should be considered to indicate a presumptive diagnosis of HPAI, and appropriate response should ensue.¡¯ The report prompted the attached article in the latest issue of Nature ¡ª raising the possibility of, in some circumstances, using cats as ‘sentinels’ for these viruses.

Nature, Tuesday 25 July 2006 http://www.nature.com/news/2006/060724/full/060724-4.html Published online: 25 July 2006; doi:10.1038/news060724–4

More cats found with bird flu Researchers suggest feline ‘sentinels’ could identify dangerous outbreaks By: Declan Butler

Domestic cats may be widely susceptible to infection with the avian flu H5N1 virus, according to scientists who this week reported the virus in two dead cats in northern Iraq. The latest reports, following recent cat cases in Austria, Germany, Thailand and Indonesia, reinforce the hypothesis that cats may play a role in the spread of the virus, although none of the human victims thus far is thought to have caught the virus from a cat. The findings also suggest that cats might help provide an early-warning system for avian flu by acting as ‘sentinels’, say the scientists, who work at a US Naval Medical Research Unit in Cairo, Egypt. Many remote areas lack the veterinary infrastructure to test quickly for H5N1. So as a proxy, they argue, H5N1 should be immediately suspected and guarded against whenever unusual bird and cat die-offs happen together. Bird flu continues to hit Asia. Thailand has just seen a resurgence in chicken cases this week, after being apparently free of the virus for a year. Cat curiosity H5N1 was first reported in domestic cats in Thailand in 2004, and a later survey showed that some Thai cats carry antibodies to the virus. Further lab work showed that cats can carry the virus in their guts and faeces, and so could contaminate the environment to spread the virus. “In nature we saw exactly what they saw in the lab,” says Samuel Yingst, the lead author of the new work, speaking from Bishkek, Kyrgyzstan. The researchers tested the cats out of curiosity during a two-week field trip last February near Erbil City, in Kurdish northern Iraq, after hearing anecdotal reports of cat deaths associated with H5N1 outbreaks in Turkey and Iraq the month before. Their findings are published in the August issue of Emerging Infectious Diseases. “It’s conceivable that cats could spread the virus,” says Yingst, although he suspects that they may be ‘dead-end’ hosts that die after receiving the disease without passing it on. Baffling virus Dick Thompson, a spokesman for the World Health Organization (WHO), says the latest paper does not change the WHO’s current position: “there is no present evidence that domestic cats play a role in the transmission cycle of H5N1 viruses. To date, no human case has been linked to exposure to a diseased cat.” That said, some cases of H5N1 continue to baffle scientists. There have been reports of cat die-offs in Indonesia in areas where no bird outbreaks have been reported, for example. And one cat virus has been shown to share gene sequences with human cases; gene sequences that have not been reported in poultry samples. Some human cases from Java “have no obvious avian counterparts”, concluded a dozen international experts in animal and human health at the Avian Influenza Expert Consultation meeting in Jakarta from 20¨C23 June. They said there is an “urgent need” to compare human, cat and bird sequences, but that such efforts are being hindered by a lack of data. On the alert In the meantime, the study could help spark an idea for early-alert systems. “Where cats show respiratory infections in areas where avian flu is endemic, H5N1 will probably be one of the causes,” says Magdi Saad, a co-author on the work. Cats could therefore serve “as sentinels in areas which don’t have access to good diagnostics”, adds Yingst. “I’d completely agree that cats can serve as sentinels, they seem very susceptible,” says Albert Osterhaus, whose team at Erasmus University in Rotterdam, the Netherlands, has shown that experimentally infected cats can transmit the virus. Other carnivores are also likely to fall foul of flu, he says, adding that his group is now also looking at ferrets, foxes and seals. Sentinels are important as a first alert, the experts agree, but they are no substitute for detailed investigation.

Reference: Yingst, S.L., Saad, M.D. and Felt, S.A. (2006) Emerging Infectious Diseases, Vol. 12, No. 8, available online at http://www.cdc.gov/ncidod/EID/vol12no08/06-0264.htm

Emerging Infectious Diseases, Vol. 12, No. 8, dated August 2006

http://www.cdc.gov/ncidod/EID/vol12no08/06-0264.htm Posted online ahead of print on Monday 17 July 2006

Qinghai-like H5N1 from Domestic Cats, Northern Iraq Samuel L. Yingst,* Magdi D. Saad,* and Stephen A. Felt*

• US Naval Medical Research Unit No. 3, Cairo, Egypt

To the Editor: Natural infection of several cat species with highly pathogenic avian influenza (HPAI) H5N1 viruses in Thailand (1¨C4) and experimental infection of domestic cats with similar viruses have been reported (5,6). Thus, literature describing HPAI H5N1 infection of cats is limited to descriptions of infections with a subset of clade I viruses. HPAI H5N1 viruses, highly similar to viruses isolated from Qinghai Lake in western People’s Republic of China in spring 2005, are now rapidly disseminating throughout Eurasia and Africa. To our knowledge, this is the first report of a Qinghai-like virus detected in domestic cats. This finding is noteworthy because the host range of influenza viruses is determined by the antigenic characteristics of the hemagglutinin and neuraminidase molecules; clade II viruses are antigenically distinct from clade I viruses, and Qinghai-like viruses are genetically distinct from other clade II viruses.

Personal communications in January 2006 from field veterinarians noted deaths of domestic cats that were associated with suspected (eventually confirmed) H5N1 outbreaks in eastern Turkey (2 villages) and Kurdish northern Iraq (Sarcapcarn in Sulymaniyah Governorate and Grd Jotyar in Erbil Governorate). The clinical conditions of the birds did not suggest HPAI to villagers or consulting veterinarians. In both scenarios in Iraq, results of rapid antigen detection tests with the Anigen kit (Suwon, Republic of Korea), while positive for influenza A, were negative for H5, so the outbreaks were not thought to be caused by HPAI, but concern about the unusual deaths in cats remained. Because the regions are remote and veterinary services limited, such anecdotal reports have rarely been followed up.

After H5N1 influenza was diagnosed in a person in Sarcapcarn, Kurdish northern Iraq, the government of Iraq requested a World Health Organization investigation, which was supported in part by Naval Medical Research Unit No. 3 veterinarians. While investigating the situation in Erbil Governorate, the team was informed of suspicious deaths in cats associated with the death of all 51 chickens in a household in Grd Jotyar (¡Ö10 km north of Erbil City) From February 3 to February 5, 2006, five cats reportedly died; 2 of these were available for examination on February 8. A sick goose from an adjacent household was killed and underwent necropsy with the cats. Gross pathologic changes in cats were similar to those previously reported, except that severe hemorrhagic pancreatitis was observed (5,6). Tissues from these animals and archived tissues from 1 of the 51 chickens were conveyed to Cairo for virologic examination.

An influenza A H5 virus was present in multiple organs in all species from the outbreak site in Grd Jotyar (see Table). cDNA for sequencing was amplified directly from RNA extracts from pathologic materials without virus isolation. On the basis of sequence analysis of the full HA1 gene and 219 amino acids of the HA2 gene, the viruses from the goose and 1 cat from Grd Jotyar and from the person who died from Sarcapcarn (sequence derived from PCR amplification from first-passage egg material) are >99% identical at the nucleotide and amino acid levels (GenBank nos. DQ435200¨C02). Thus, no indication of virus adaptation to cats was found. The viruses from Iraq are most closely related to currently circulating Qinghai-like viruses, but when compared with A/bar-headed goose/Qinghai/65/2005 (H5N1) (GenBank no. DQ095622), they share only 97.4% identity at the nucleic acid level with 3 amino acid substitutions of unknown significance. On the other hand, the virus from the cat is only 93.4% identical to A/tiger/Thailand/CU-T4/2004(H5N1) (GenBank no. AY972539). These results are not surprising, given that these strains are representative of different clades (8,9). Sequencing of 1,349 bp of the N gene from cat 1 and the goose (to be submitted to GenBank) show identity at the amino acid level, and that the N genes of viruses infecting the cat and goose are >99% identical to that of A/bar-headed goose/Qinghai/65/2005(H5N1). These findings support the notion that cats may be broadly susceptible to circulating H5N1 viruses and thus may play a role in reassortment, antigenic drift, and transmission.

The route of infection in these cats cannot be determined definitively. How cats behave when eating birds makes both oral and respiratory infection possible. However, the source of infection seems clear in that an identical H5N1 virus was detected in samples from a goose from the same dwelling, and an H5 virus was detected from archived samples from 1 of 51 chickens that died over the course of a few days. The potential for horizontal spread cannot be ruled out since we detected viral RNA in gut, stool, and trachea; clinical signs developed in all cats, and all died from the acute illness 2¨C4 days after the chicken deaths began; therefore, simultaneous exposure seems likely. Death in cats, spatially and temporally associated with unusual deaths in poultry, especially when the cats show positive results of a rapid antigen detection test for influenza A, should be considered to indicate a presumptive diagnosis of HPAI, and appropriate response should ensue.

Acknowledgments

We thank Elham Botrus Shabo, Saman Najeeb, Faisal Polus, Sura Jabar, Saidawan Omer Yussif, and Burhan Sulaiman for facilitation and technical assistance in sampling and performing necropsies, and Bradford Camp, Odis Kendrick, and Kosar Shaheer for communications and security.

This work was supported by funding through the Naval Medical Research Center work unit GEIS E0018.

 References 

1. Keawcharoen J, Oraveerakul K, Kuiken T, Fouchier RA, Amonsin A, Payungporn S, et al. Avian influenza H5N1 in tigers and leopards. Emerg Infect Dis. 2004;10:2189¨C91.

2. Thanawongnuwech R, Amonsin A, Tantilertcharoen R, Damrongwatanapokin S, Theamboonlers A, Payungporn S, et al. Probable tiger-to-tiger transmission of avian influenza H5N1. Emerg Infect Dis. 2005;5:699¨C701. Erratum in Emerg Infect Dis. 2005;11:976.

3. Songserm T, Amonsin A, Jam-on R, Sae-Heng N, Meemak N, Pariyothorn N, et al. Avian influenza H5N1 in naturally infected domestic cat. Emerg Infect Dis. 2006;12:681¨C3.

4. Amonsin A, Payungporn S, Theamboonlers A, Thanawongnuwech R, Suradhat S, Pariyothorn N, et al. Genetic characterization of H5N1 influenza A viruses isolated from zoo tigers in Thailand. Virology. 2006;344:480¨C91.

5. Rimmelzwaan GF, van Riel D, Baars M, Bestebroer TM, van Amerongen G, Fouchier RA, et al. Influenza A virus (H5N1) infection in cats causes systemic disease with potential novel routes of virus spread within and between hosts. Am J Pathol. 2006;168:176¨C83.

6. Kuiken T, Rimmelzwaan G, van Riel D, van Amerongen G, Baars M, Fouchier R, et al. Avian H5N1 influenza in cats. Science. 2004;306:241.

7. Spackman E, Senne DA, Myers TJ, Bulaga LL, Garber LP, Prdue ML, et al. Development of a real-time reverse transcriptase PCR assay for type A influenza virus and the avian H5 and H7 hemagglutinin subtypes. J Clin Microbiol. 2002;40:3256¨C60.

8. World Health Organization Global Influenza Program Surveillance Network. Evolution of H5N1 avian influenza viruses in Asia. Emerg Infect Dis. 2005;11:1515¨C21.

9. Chen H, Smith GJD. LI KS, Wang J, Fan XH, Rayner JM, et al. Establishment of multiple sublineages of H5N1 influenza virus in Asia: implications for pandemic control. Proc Natl Acad Sci U S A. 2006;103:2845¨C50.

anonymoous,

please chose a handle so we can tell our anonymouses apart.

Melanie apologies. I posted the article from Declan and Emerging diseases above. It was an email sent to me by Mike Nunn.

Melanie I posted the article from Declan and Emerging diseases above. It was an email sent to me by Mike Nunn.

Melanie I posted the article from Declan and Emerging diseases above. It was an email sent to me by Mike Nunn.

Andrew,

Thank you for posting that. I was reading it earlier and wondering what is the difference between cats as hosts vs pigs?

What are possible implications from both the virological and epidemiological points of view?

Thanks again for keeping us informed.

anon_22 at 12:35, thanks so much for your response and posting information from the Nature-Taubenberger-1918 polymerase article. I am picking up a full copy of that article this week for thorough review.

The authors of the March 2006 article (Salomon et al, referenced above at 14:01), also looked at the E627K change in PB2, here is their comment on that issue:

“The generation of a RG [reverse genetics] virus of VN1203 with a point mutation in PB2 changing lysine (K) at position 627 to glutamic acid (E) was not lethal when administered to mice (unpublished data). This demonstrates that K627 of VN1203 PB2 contributes to lethality.”

So yes, it appears that the E627K is a highly suspicious lethality factor, although it may be working in conjunction with other factors such as the multibasic cleavage site in HA, etc.

I will try to do some BLAST work to recheck for E627K in H5N1 isolates. This day job thing, and other responsibilities, keeps me from doing as much as I would like, wish I could do more!

Andrew, thanks so much for the additional update about cats as possible mammalian carrier.

beehiver,

“I will try to do some BLAST work to recheck for E627K in H5N1 isolates.”

I think the Taubenberger study may have that already, at least up to the point of publication. Look in the supplementary material. I don’t have time to do that today cos I’m travelling.

10–4, thanks for letting us know anon-22.

There is another cluster in Indonesia from the Karo region.

Three Indonesians in hospital, bird flu suspected

JAKARTA, Aug 2 (Reuters) - Three Indonesian children have been hospitalised with suspected bird flu in a North Sumatran district where seven members of an extended family died from the disease in May, officials said on Wednesday.

The three — two siblings aged 10 and six and their 18-month-old neighbour — were admitted to the state-run Adam Malik hospital on Tuesday after showing symptoms of bird flu, said hospital director Luhur Suroso.

“We are testing samples taken from them,” Suroso told Reuters. He declined to give further details.

Hariadi Wibisono, the health ministry’s director-general of control of animal-borne diseases, said the patients lived in Karo district in North Sumatra province where bird flu killed as many as seven people in an extended family in May.

The deaths in Karo were the biggest cluster of the disease the country has recorded and sparked fears of a global pandemic of bird flu infections in humans.

The World Health Organisation said in May that two members of the cluster, an Indonesian man and his son, might have caught the virus in a case of direct human-to-human transmission, but the virus did not spread very far if this did happen.

“They are from the same district but a different village,” Wibisono said, referring to the three new suspected cases.

Indonesia has recorded 42 deaths from the H5N1 bird flu virus, equalling Vietnam, where no one is known to have died of the disease this year.

Human cases of bird flu have been rising steadily in Indonesia since its first known outbreak in poultry in late 2003.

Worldwide, the disease has killed at least 134 people since it re-emerged in east Asia in 2003.

Indonesia has been criticised for not doing enough to stamp out H5N1, which still remains essentially an animal disease but experts fear could spark a pandemic if it mutates into a form that can pass easily among people.

The government has so far shied away from mass culling of poultry, citing lack of funds and impracticality in a country with millions of backyard fowl. REUTERS

I smell something cooking here, and it isn’t Kare Ayam.

Andrew,

We were just talking about this on the CDC thread, about the re-emergence of the virus in Karo.

As I understand it, no culling was done at the time of the outbreak. That being the case, we can infer that the origin source is still there.

But can we also infer that there is the much more serious risk of the strain that had passed through patients of that original cluster having then re-infected poultry (or even pigs), so that we have a B-H-H-H-B-H-H situation?

Can you comment on that please? Thank you.

annon 22. I just jumped in here and haven’t read all the threads but reading your post, I think a case could be made for Monotreme’s third wave…in other words this may not be BHHHBHH as you so intelligently have surmised but the source of the reinfection would be an entrenched mutation in a tertiary mammal (mice, rats etc. take your choice).

It would most likely be completely asymptomatic in the tertiary host so that we would not observe abnormal patterns of sudden death.

Tom,

Well, Monotreme’s hypothesis is sound, as I have said already. But this current concern of mine is more of a epidemiological investigation. That is, what is the relationship between the current (suspected) clusters and the biggest cluster so far, which only happened not so long ago and not so far away. (I’m still looking up maps trying to find the distance between Kubu Sembelang, the village of the big cluster, and the current village Sumbul.

I want to know ASAP if there is a chance that this is the same virus that had infected that family, either a) from the same source, or worse, b) from birds that had acquired that virus from the family of 8 patients.

anon_22 – at 19:33

19 chicken died near the new cluster (I think confirmed bird flu, have to double check)

annon 22. I completely agree except for…”b) from birds that had acquired that virus from the family of 8 patients.”

It could very easily be a hidden mammalian carrier (although it could also be birds). It would most likely have to be something new that has not existed in China, Vietnam or Thailand…that’s where the mammal may fit into it.

…are their small mammals in Indonesia that would not be in those other countries?

Sequencing should answer the first part of the question, no matter how closely the two clusters live.

Tom DVM – at 19:43

Don’t know if this will work for you:

Mammals in Indonesia:

http://tinyurl.com/pudy8

Mammals in China:

http://tinyurl.com/mktas

Tom,

I agree.

But I am looking for the link between the clusters as well.

Maybe I should start a different thread for clarity.

:-)

Anon-YYZ Thanks.

I just posted this new thread for the current cluster in karo to discuss whether this might be a continuation of the cluster in May.

This is important. Please check it out and tell me what you guys think.

Closed to maintain Forum speed.