The H5N1 strain of influenza - often referred to as bird flu - is first known to have jumped from chickens to humans in 1997. Since 2004 it has ripped through poultry and wild bird populations across Eurasia, and had a 53% mortality rate in the first 147 people it is known to have infected. Health authorities fear this strain, or its descendent, could cause a lethal new flu pandemic in people with the potential to kill billions.
Flu has been a regular scourge of humanity for thousands of years. Flu viruses each possess a mere 10 genes encoded in RNA. All of the 16 known genetic subgroups originate in water birds, and especially in ducks.The virus is well adapted to their immune systems, and does not usually make them sick. This leaves the animals free to move around and spread the virus - just what it needs to persist.
But sometimes a bird flu virus jumps to an animal whose immune system it is not adapted to.
Vietnam's Health Ministry has confirmed that bird flu virus killed a four-year-old child in the country's north, the country's first human case in nearly five months, state-run radio said on Thursday.
Two new human cases found after the death of the young woman announced
only a day ago:
Two Egyptians have tested positive for the deadly H5N1 bird flu virus, a day after an Egyptian woman died of the disease, Egypt's health ministry said on Thursday.
"There are two cases today, one in Damietta and one in Menoufia... Today lab results confirmed that they are infected with bird flu," Amr Kandeel, head of communicable disease control at the health ministry, told Reuters.
testing done by pakistan's national institute of health earlier in december identified nine possible cases, five in the family cluster, a female doctor who had treated members of the family and three poultry cullers who were unrelated to the family. a 10th possible case, another brother in the family, died without having been tested.
some of the testing was done using a pcr or polymerase chain reaction test that looks for traces of virus in sputum samples. in other cases the pakistan lab looked for antibodies in blood using a test which has not been validated as effective in finding h5n1 infections.
retesting by the who collaborating centre for influenza in london and by experts from a u.s. naval laboratory in cairo - known as namru-3 - did not support the initial positive findings on all the suspect cases, hayden admitted.
the negative results may be the product of degradation of specimens due to multiple freezing and thawing of the samples, he said. other factors could have affected the results, including when in the course of infection the specimens were taken or whether suspect cases were started on antiviral drugs before a sample was taken.
"we've got a host of technical issues," he said. "there are practicalities of samples breaking down over time in the transportation."
the above comments are from recent media reports in response to the who situation update on the h5n1 infections in pakistan. as noted above, ten patients tested positive by labs in pakistan, but only one was positive in testing done by who regional reference labs (a mobile unit shipped to pakistan from namru-3, and mill hill testing of samples shipped to london). additional antibody testing will be done on serum samples, and sequencing is in progress. however, the negative data indicate that at most there will only be sequence data from the second fatal case.
the above comments also give a litany of reasons for the false negatives, but these false negatives raise concerns about past and future cases, as well as risk assessments.
the limited release of disease onset dates clearly shows sustained human to human transmission for a month, at a minimum. this chain may have stretched from the index case to a surviving brother to a deceased brother to a second deceased brother, to one or more health care workers. the exact components of the chain are still unclear because all of the onset dates have not been released, and all of the positives have not been supported by clear clinical data. however, not all h5n1 infections produce easily differentiated clinical presentations, which is why lab confirmation is important and why the false negatives increase concerns.
clusters are useful for delineating problems that lead to false negatives. in the cluster from pakistan, factors for negative data include failing to test, or testing after repeated freeze and thaws or after the start of tamiflu treatment.
the test failure for the first fatality remains unclear. although most media reports indicate no sample was collected, at least one report quotes a health care worker as saying samples were collected from both fatal cases. it is unclear if there was no sample, or if it was lost, dropped, improperly tested, or results were lost, withheld, etc.
false negatives could have been due to sample degradation due to a failure to make aliquots of the sample resulting in multiple freeze / thaws. freeze / thaws could also be due to packaging, shipping, or receiving issues.
there are literally hundreds of ways to generate false negatives, which is why such negatives are viewed with caution by the scientific community, but these negatives frequently drive press releases, risk assessment, and policy by others.
unfortunately, false negatives are quite common in h5n1 testing. most surveillance groups have yet to find h5n1 in a live wild bird. on clinical samples, negatives were eventually generated for serum samples collected from patients in northern vietnam in 2005. in turkey, only 12 of the 21 positives were confirmed in england, and sequences were released from only four of the positives. many patients initially test negative, and those on tamiflu continue to test negative because of suppression of viral rna levels.
the assays used for h5n1 are largely confirmatory, and work best when samples are collected at the proper time and handled correctly. virtually all human h5n1 isolates come from samples collected when the patient is near death.
thus, in pakistan viral isolation is limited to two patients, and samples are missing from one of the two. false negatives also lead to false transmission chain terminations, as well as failures to test contacts of the falsely negative patients.
thus, the false negatives in pakistan increase pandemic concerns.
yet another brother, who lives on long island, n.y., travelled to pakistan with his son to attend at least one of the funerals. on their return to the united states they suffered mild respiratory symptoms and consulted their family doctor, who alerted new york state public health authorities.
in early december, the u.s. centers for disease control sent a plane to albany, n.y., to collect samples taken from the two. testing at the cdc's labs in atlanta confirmed the man and his son were not infected with the virus.
the above comments on the son contradict the state of new york report that indicated the family of the brother from new york was not exposed to the cases in pakistan. if both father and son flew to pakistan for the funeral(s), there would have been exposure. the first brother died november 19 and the other fatally infected brother developed symptoms november 21, so if the father and son were in pakistan for the first funeral, they would have been exposed to the second brother, as well as the source that infected the second brother. moreover, if they then stayed for the second funeral, they would have been in pakistan for more than 3 weeks, since they returned to new york on december 5.
pakistan acknowledged that some relatives did not develop pneumonia because they were receiving tamiflu prior to collection of samples. it seems likely that both father and son from the us would also be receiving tamiflu.
another suspected case in pakistan
news out of pakistan has been disturbing and now we have another suspect birdflu case in an 8 yr old child. on the plus side is that not all cases in pakistan have been fatal:
"there is no suggestion that the virus has changed into a form that poses a broader risk," who spokesman john rainford told afp. "if that had been the case, we would have witnessed more cases of human transmission."
the above comments have taken the who spin into the stratosphere. who consultants have already identified receptor binding domain changes in h5n1. in clade 2.2 these receptor binding domain changes have been associated with larger clusters involving human transmission. in pakistan, most of the patients lab confirmed locally were linked to clusters.
the larger cluster appears to be the longest ever reported for h5n1 and was sustained more a month. although who hasn’t provided the details in a who situation update, the disease onset dates were cited in media reports quoting who officials.
receptor binding domain changes can lead to more efficient binding to human receptors. this affinity leads to larger clusters and human to human transmission. in clade 2.2 this change is of concern because another change, pb2 e627k, has become fixed.
prior to the outbreak at qinghai lake in may, 2005, e627k had never been reported in h5n1 from birds. it was found in some h5n1 from patients and was also in the only reported bird flu fatality that wasn’t h5n1, the h7n7 fatality in 2003 in the netherlands. it also increased h5n1 virulence in mice and was associated with an increase in viral replication at lower temperatures (33 c). this temperature corresponds to the temperature of a human nose or throat in the winter. consequently e627k is a human polymorphism and has been found in all human flu isolates dating back to 1918.
therefore, changes in the receptor binding domain are of concern and the association of receptor binding domain changes with clusters and human to human transmission has been clear, especially for the qingahi strain (clade 2.2).
a prediction of such a change, s227n, in qinghai h5n1 flying into the middle east was predicted based on donor sequences in endemic h9n2. the first qinghai h5n1 human case was in turkey, and the isolate from the index case had s227n. although the change wasn’t in the isolate from the sister of the index case, it was in a second isolate from turkey. thus, two of the four turkey isolates had s227n. in turkey 21 cases were lab confirmed and virtually all cases were from lab confirmed clusters.
the outbreak in turkey was followed by an outbreak in iraq. although only three patients were h5n1 confirmed in iraq, two were in a cluster and both had two receptor binding changes, n186s and q196r.
the outbreak in iraq was followed by an outbreak in azerbaijan,and again the vast majority of the cases were in clusters and again another receptor binding domain change, n186k, was found in all human isolates from azerbaijan.
the next human outbreak was in egypt, and the vast majority of cases there were not clusters. however, a year ago egypt did have a cluster of three, the largest reported there to date. h5n1 was isolated from two of the three fatal infections and both had two receptor binding domain changes, v223i and m230i. m230i was another “human” polymorphism found in all human seasonal flu isolates, including influenza b
thus, the association of receptor binding domain changes with larger clusters and increased human to human transmission for clade 2.2 qinghai h5n1 is quite clear, and similar changes in the ha sequence from the sole confirmed positive in pakistan is likely.
in pakistan, 8 of the 9 patients testing positive locally were in clusters. six were in the larger cluster while two were in the smaller cluster. only one positive was not in a cluster.
although a receptor binding domain is not a certainty (isolation procedures can select away from receptor binding domain changes present in the h5n1 from the patient), it is likely that the h5n1 from pakistan will be clade 2.2 (qinghai), which has been true for all h5n1 cases west of china, and the h5n1 in the patient will have a receptor binding domain change.
Fatma Fathi Mohamed died in hospital in the Nile Delta city of Mansoura, three days after she was admitted to a smaller local hospital with a high temperature and difficulty breathing, it said in a statement carried by the state news agency MENA.
The above comments describe the fourth confirmed H5N1 case in Egypt this week. The case is also the second fatality, suggesting a more virulent H5N1 is circulating in Egypt at this time. The other two patients are in critical condition or in the ICU.
H5N1 has been spreading in Egypt this week, indicating more human cases are likely.
...data for last season suggest that much of the difference between the high case fatality rate at the beginning of the season... and the case fatality rate for the remainder of the season...could be explained by newly acquired polymorphisms that were associated with fatal cases.
Egypt offers an ideal situation for rapid evolution. It lies at the intersection of two major flyways and has the large Nile Delta, which offers the opportunity for co-infections. The population of Egypt is concentrated in the Nile Delta and along the Nile River. The backyard poultry can mix with the wild birds, increasing opportunities for additional co-infections in domestic poultry.
One of the markers from northern isolates was M230I, a polymorphism found in all human seasonal flu isolates (H1N1, H3N2, influenza B). This marker was associated with fatal cases. All cases with M230I died...
There appear to be two different large outbreaks in poultry. Interesting that
they are using the military to do the culling:
Via Xinhuanet, a report that appears to deal with a separate outbreak and cull from the one reported here on December 28: Some 2,600 fowls culled in northern Bangladesh. Excerpt:
Bangladesh Army personnel Sunday culled about 2,600 fowls affected by bird flu in Bangladesh's northern Gaibandha district, about 210 km northwest of capital Dhaka.
Politics and bird flu are not a good mix
From Emmy Fitri of the Jakarta Post, read it before it disappears:
the haifa district physician, prof. shmuel rishpon, confirmed thursday that a deadly strain of the bird flu virus has infected chickens at a petting zoo in a binyamina kindergarten.
earlier thursday morning 18 of the 25 chickens in the kindergarten's petting zoo, were found dead.
"the virus was identified as h5n1 bird flue,"
"the kindergarten staff has been given preventive medicines and as far as we know, none of the children or their parents came in contact with the birds.
the above comments confirm h5n1 in haifa, israel. the finding was not unexpected because of the mortality in the chickens and initial positive data for influenza. the lack of contact between children and chickens at a kindergarten petting zoo remains unclear. the staff has been placed on prophylactic oseltamivir (tamiflu).
the confirmation of h5n1 in israel suggests bird flu is widespread in the middle east. the are a reported 38 outbreaks in saudi arabia, and the number of outbreaks in egypt is likely to be higher (see satellite map). birds are migrating through the region at this time, and reported outbreaks upstream have been high this fall in europe and possibly iran..
thus far confirmed human cases have been limited to egypt (see satellite map) and pakistan, although there was a suspect case at the kuwait airport. the similarity between h5n1 in egypt and israel in 2006 raises the possibility of more human cases in the area. in addition to the recent confirmed cases in egypt, a growing number of suspect cases are being reported.
the outbreak in pakistan is likely to be the uva lake strain, which has been widespread in europe and in kuwait early last year. it is likely that the uva lake strain has now migrated into the middle east and may be recombining with resident sequences.
other than germany and russia, all other countries have withheld recent h5n1 sequences.
The WHO has said some of those who died having contracted the H5N1 virus strain showed moderate resistance to Tamiflu, the antiviral drug.
The above comments indicate N294S has re-emerged in the Egypt. Almost exactly one year ago N294S was detected in the Gharbiya cluster. It reduced the efficiency of oseltamivir (Tamiflu) by 15-20 fold and was characterized by WHO as having “moderate” resistance because another resistance marker, H294Y, reduces efficiency by 1000 fold in patients in Vietnam.
However, the effectiveness of Tamiflu in the treatment of H5N1 is borderline, so a twenty fold reduction can lead to fatal infections. Since there have only been four recent confirmed fatalities in Egypt, at least half have H5N1 with moderate resistance, which is almost certainly due to N294S.
Last year H5N1 sequences were generated from two of the three cluster members. Samples had been collected prior to Tamiflu treatment, as well as 48 hours after treatment started (A/Egypt/14724-NAMRU3/2006 and A/Egypt/14724-NAMRU3/2006,). All four NA sequences had N294S, indicating the change was already present in the H5N1, which was probably in birds, based on the epidemiology of the cluster.
The presence of N294S in birds was supported by two prior isolates from China, A/duck/Zhejiang/bj/2002(H5N1) and A/Duck/Hong Kong/380.5/2001(H5N1), which had the same change. The change was not found in a limited survey of bird isolates in Egypt, and was not detected in any of the subsequent human isolates.
However, the new fatal cases this season raise the possibility that N294S has returned, and the comments above suggest that this is the case, since “some” patients have “moderate” resistance.
It is not clear if these infections involve the Gharbiya strain, detected a year ago, or the Uva Lake strain which has appended the N294S onto a new genetic background via recombination.
Release of the human and bird sequences in Egypt would be useful...
From a new blogger, an Egyptian woman, very interesting. Check out her vintage photos of the Egptian royal family. They are gorgeous!
Here is something about Tamiflu resistance in Egypt. Crofsblog gets the credit for finding this link.
He thinks she means sanitation when she writes the word, sanity:
the who has said some of those who died having contracted the h5n1 virus strain showed moderate resistance to tamiflu, the antiviral drug.
the above comments on moderate resistance to tamiflu in some of the fatal cases are cause for concern. since there are only four confirmed fatalities, at least half are tamiflu resistant. the "moderate" resistance is almost certainly due to n294s, the genetic change found in the gharbiya cluster a year ago. n294s was present in samples collected prior to treatment, indicating it was circulating in egypt.
who and the eastern mediterranean branch of who, have issued situation updates on all five confirmed cases. the updates from the mediterranean branch were most detailed, with dates for the start of tamiflu treatment for four of the five cases. the date of sample collection matched the start date for tamiflu, indicating the resistance found in the h5n1 sequences was present prior to treatment. this conclusion is further supported by the dates of death, which were shortly after the treatment date, limiting the time for resistance to develop.
thus, like the cases from last year, the sequence data indicates n294s is circulating in egypt. however, unlike last year, when reported n294s was limited to a single cluster, it appears to be present in multiple patients. the exact number is unclear. tamiflu resistance is not mentioned in any of the updates on the five patients.
last season who issued a comprehensive report that was somewhat delayed because samples were sent for biological confirmation of the sequence data. the biological results matched data generated with h5n1 from asia that also had n294s. the resistance was in the range of 15-20 fold, which was markedly less than h294y, but high enough to raise serious treatment concerns. n294s has been seen previously in h5n1 in ducks from china, but it was only detected in one case in vietnam.
in contrast, n294s was in a cluster in gharbiya last season, and now is in "some" of the recent four fatal cases. since who has acknowledged that the resistance is "moderate", the number of confirmed cases with n294s should be delineated in an update, and sequences should be released.
the five confirmed cases were widely dispersed in egypt, and only two of the five came from the same governorate, menoufia, which also has the largest number of hospitalized suspect cases (see satellite map). all four fatal cases were from separate governorates.
recently, egypt announced a tamiflu dispersal plan in gharbiya and possibly additional governorates in the nile delta. the frequency of h5n1 in patients in egypt is not well understood. although egypt has the highest number of confirmed h5n1 outside of asia, the number of suspect cases is markedly higher than confirmed cases. false negatives are always a possibility, but the failure to get sequence data from the third gharbya cluster member, and 9 of the 10 h5n1 infected patients in pakistan tested negative on confirmatory testing. these data indicate that false negatives are easily generated, so the number of human h5n1 cases may be markedly higher than the number of confirmed cases.
tamiflu is used liberally in egypt, which raises the possibility of selection of n294s. selection may explain while multiple patients with n294s were found in a small number of confirmed fatal cases, raising concerns that n294s is becoming dominant in egypt.
last season n294s was not reported in any of the poultry isolates and was limited to the one cluster. the comments above suggest that n294s is widespread in egypt, and cause for concern.
release of human and poultry sequences would be useful. n294s is well defined, both with regard to biological resistance and coding, as seen in sequences from last season in (a/egypt/14724-namru3/2006 and a/egypt/14725-namru3/2006).
indigo girl
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Vietnam
http://crofsblogs.typepad.com/h5n1/2007/12/vietnam-confirm.html