>>Chinese government officials, meanwhile, suggested that the patient's illness - if it is indeed SARS - might be marginally different - "a virus variation with (a) new sequence," Xinhua said, quoting Guangdong's Center for Disease Prevention and Control.
The center emphasized that the findings were extremely preliminary, and WHO made no mention of such a discovery.
"The results point to the possibility of the suspect infected by SARS coronavirus, but our sequence test shows ... a new sequence that has not been published," Dr. Chen Qiuxia, a center physician, was quoted as saying.<<
Last year, when SARS emerged on an international scale, it was commonly called the "mysterious illness" by the media because the fatal pneumonia was spreading rapidly and an infectious agent had not yet been isolated. However, after a novel coronavirus was isolated and sequenced, the mystery was not as mysterious. In the ensuing months, a good deal of empirical data was generated which would be used in the design and analysis of laboratory tests designed to confirm the presence of the virus. Due to a number of technical considerations, confirmation could be achieved by a variety of tests that measured the RNA of the virus, antibodies directed against the virus, or the actual isolation of the virus. The limitations of these tests were well known. The RNA levels, as measure by PCR, was generally most accurate on samples collected about 10 days after initial symptoms. Testing too early or too late could give false negative results. The patient's immune response also took time to develop, so negative tests on sera collected less than 28 days after initial symptoms were not considered to be true negatives. The actual isolation of the virus remains as a relatively rare event.
The current Guangzhou case generated significant concern because the patient had classical SARS symptoms, was from Guangzhou, and developed symptoms approximately one year after the initial SARS cases appeared in Guangdong province. Early tests produced a mixed response, in part because samples were collected early, and in part because the SARS cornonavirus detected had a fair number of mutations in all three genes sequenced thus far.
The sequence of the virus cleared up much of the initial lab data, but because the source of the infection has not been identified, the lab data is portrayed in the media and in some official announcements as confusing, perplexing, or complex. However, analysis of the sequence data removes most of this confusion, so it is worthwhile going over some of the data that has been released thus far. The actual sequence will answer additional questions, but the following can be gleaned from yesterday's press release.
Three of the genes of the SARS CoV have been sequenced and compared to other SARS CoVs. The three genes are each about 99% homologous to the sequences of other reported SARS CoVs isolated from patients with probable SARS or animal harboring a virus that is 99.8% homologous to the SARS CoVs from patients. The most important gene for determining host range and tissue tropism is the S gene.
The S gene is composed of 3768 nucleotides and 3768 nucleotides have been sequenced in the Guangzhou isolate. A comparison of the new sequence with those at GenBank indicates that the homology ranges between 98.8% and 99.4%, which would mean that there are between 22 and 44 nucleotide differences between the new sequence and those reported previously. This is a relatively high number and the identification of the changed nucleotides would help considerably in determining the source of the novel SARS coronavirus. A comparison of the complete animal sequence (approximately 30,000 nucleotides) with the human sequences indicates that most of the differences are concentrated in the S gene. 13 changes are found in all human isolates and another 9 changes are found in the vast majority of human isolates. If the new virus has most of these changes, then it probably shared a common recent origin. However, the large number of changes in the S gene suggests that the origin may be slightly different.
The differences in the other genes could address this issue, but the numbers in the press release can be interpreted two ways. The M gene has 666 nucleotides and the press release indicates that the new isolate is 99% homologous in the 658 nucleotides sequenced. This could indicate that sequencing is not complete and the sequence thus far has 6-7 differences. Alternative, the 99% could simply mean that there are a small number of difference and a more precise percent homology will be calculated when all 666 nucleotides are determined. The same situation may exist for the N gene, where 1068 of 1269 nucleotides have been determined. If the 99% however means that thus far 10 or 11 changes have been found in the N gene, then it is likely that the lineage of the new isolate is slightly different than pervious isolates because the N gene is highly conserved. In fact the majority of the human isolates have no differences with the animal sequence (i,e, the homology is 100% and 1269 of 1269 nucleotides are identical).
In any event, the complete sequence of the S gene shows that the latest isolate has drifted quite a bit relative to all of the other known sequences. Recent confirmatory lab tests have focused on neutralizing antibodies in the patient's sera. Since the novel SARS CoV has not been isolated, the only way to measure neutralizing antibodies is to see how well the patient's antibodies react with a previous isolate. The data above indicate that the differences between the novel isolate and the isolate used in the lab test is between 22 and 44 nucleotides. This amount of drift will almost certainly affect the ability of the patient's antibodies (made against the novel agent) to neutralize the lab isolate. Thus, a low neutralizing antibody titer does not indicate that the patient was "slightly exposed" to the novel agent. Instead it means that the novel agent has drifted considerably, which is quite clear from the sequence data.
In summary, there is not confusion on the science or lab confirmation side. The sequence of the novel agent makes it abundantly clear that the Guangzhou patient was recently infected with a SARS CoV which has some significant differences in the S gene when compared to previous isolates. The 1% difference may also apply to two additional genes which have been partially sequenced. The data in the press release lacks sufficient detail to know if the animal reservoir for this virus is similar or identical to previous isolates, and the source of the recent infection has not been determine.
However, the sequence removes confusion that some may have concerning the confirmatory lab data. The patient's SARS symptoms were caused by a novel SARS CoV which has been partially sequenced thus far. |
