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he lived in hong kong and took frequent short business trips to mainland china . while in shenzhen in early march 2003 , a fever and productive cough with blood - stained sputum developed .
he was hospitalized in shenzhen and given intravenous antibiotics ( the details of which were not available ) .
he returned to hong kong and sought treatment at prince of wales hospital 8 days after the onset of illness .
his temperature was 37.3c , heart rate 120 beats per minute , blood pressure 110/70 mm hg , and oxygen saturation 93% on room air .
chest radiograph showed patchy consolidations in the right middle zone and mild infiltrates in the left lower zone .
he was admitted to an isolation ward and given supplemental oxygen and cefuroxime,750 mg intravenously every 8 h. blood tests showed leukocytosis ( leukocyte count 14.9x10/l , reference range [ rr ] 4.010.8 x 10/l ) with neutrophilia ( neutrophil count 13.5x10/l , 91% ; rr 41%73% ) and lymphopenia ( lymphocyte count 0.5x10/l , 4% ; rr 19%47% ) . the hemoglobin and platelet count and
arterial blood gas while the patient was receiving 2 l of oxygen showed compensated metabolic acidosis ( ph 7.39 , rr 7.357.45 ; actual bicarbonate 15 mmol / l , rr 2226 mmol / l ; po2 14.2 kpa , rr 10.013.0 kpa ) .
his condition initially improved with normalization of temperature and heart rate , and a satisfactory level of arterial blood oxygen saturation was maintained until early the next morning , when he reported increasing dyspnea and progressed to shock and severe oxygen desaturation .
before he could be transferred to the intensive care unit , he underwent cardiorespiratory arrest , and aggressive resuscitation attempts were not successful .
on admission ( 8 days after the onset of illness ) , sputum and blood samples were taken for routine microscopy , bacterial ( including legionella pneumophilia and mycobacteria ) , and fungal culture .
rapid viral antigen detection by immunofluorescence was performed with commercial assays to identify influenza a and b ; parainfluenza 1 , 2 , and 3 ; respiratory syncytial virus ( rsv ) ; and adenovirus ( dako diagnostics ltd , ely , uk , and chemicon international , inc . , temecula , ca ) .
sars - associated coronavirus ( sars - cov ) was detected by reverse transcription ( rt)-pcr by using primers cor-1 ( sense ) 5 cac cgt ttc tac agg tta gct aac ga 3 , and cor-2 ( antisense ) 5 aaa tgt
tta cgc agg taa gcg taa aa 3 that had been shown to be specific for the novel coronavirus detected from patients with sars ( 6 ) .
virus isolation was performed using rhesus monkey kidney ( llc - mk2 ) , human laryngeal carcinoma ( hep-2 ) , mardin darby canine kidney ( mdck ) , human embryonic lung fibroblast , buffalo green monkey kidney ( bgm ) , and african green monkey kidney ( vero ) monolayers .
after 14 days of incubation , the growth of influenza a and b , parainfluenza 1 , 2 , and 3 , rsv , and adenovirus was examined by commercial monoclonal antibodies with the immunofluorescence technique .
in addition , a hemadsorption test was performed for llc - mk2 and mdck monolayers .
a shell vial culture system , coupled with monoclonal antibody  based detection , was used for the isolation of cytomegalovirus . a serum sample taken on admission ( 8 days after the onset of illness )
was evaluated for hantavirus and atypical pneumonia serology , including legionella pneumophila ; m. pneumoniae ; c. pneumoniae and psittaci , influenza a and b ; parainfluenza 1 , 2 , 3 ; rsv ; and adenovirus .
in addition , an in - house immunofluorescence assay based on sars - cov  infected vero cells was used to detect sars - cov antibodies .
postmortem biopsy specimens were taken from the lungs , heart , kidney , spleen , small bowel , and muscle . pulmonary congestion with edema was noted , but hyaline membranes had not formed ( figure a ) .
interstitial inflammatory cell infiltration was minimal , and intraalveolar organizing lesions were rarely seen , whereas detached atypical pneumocytes were found ( figure b ) .
atypical multinucleated pneumocytes were observed , but definite viral inclusion was not apparent ( figure c ) .
fibrin thrombi were frequently observed in small pulmonary arteries and arterioles ( figure d ) .
the cell culture results were negative , except that a focal refractile rounding cytopathic effect developed 12 days after the left lung tissue sample was injected onto llc - mk2 cells .
this cytopathic effect progressed slowly to cell detachment , and a similar cytopathic effect was observed on other areas of the cell monolayer over the next few days .
the cell culture supernatant was positive by an rt - pcr assay based on primers 5-gag tag gga tca tca agc a-3and 5-gct tag ctg rta tac agt gtt-3 , targeting the f - gene of hmpv .
the nucleotide sequence of the pcr product were identical to the f - gene fragment of hmpv ( genbank accession no .
the cell culture supernatant was passaged to llc - mk2 cells , and the same cytopathic effect was observed after 8 days of incubation .
hmpv particles were shown from the passaged cell culture by electron microscopy . to ascertain the presence of hmpv infection in this patient , the serum sample taken on admission ( 8 days after the onset of illness )
was retrieved for hmpv antibody detection with the immunofluorescence technique , based on llc - mk2 cells infected by the isolated hmpv .
pathologic findings of lung tissue sections . a : pulmonary congestion and edema ( h&e stain , original magnification x100 ) .
d : fibrin thrombi were frequently noted in small pulmonary arteries and arterioles ( h&e stain , original magnification x200 ) .
while the full range of clinical features is still being elucidated , several cases of severe respiratory tract infections have been reported .
boivin et al . ( 2 ) reported that 3 of 12 infected young children < 5 years of age required intensive care , and a child with acute leukemia died subsequently . in that study ,
one of the six infected adult patients ( 1565 years of age ) also required intensive care . in the same study , 2 of 10 infected elderly patients ( ages > 65 years ) died ; 1 also had leukemia and the other had alzheimer s disease . however , none of these patients had an autopsy performed .
this report is the first in which hmpv was the only pathogen identified from postmortem specimens from a patient with a fatal respiratory tract disease .
our patient had a history of good health and no evidence to suggest that his death was a result of exacerbation of underlying cardiac or pulmonary conditions .
coinfection has been found in patients with hmpv - associated respiratory tract disease ( 2 ) and has been suggested to be a factor influencing clinical outcome ( 7 ) . in our patient , no evidence of infection with a copathogen could be demonstrated .
although the patient had died before a late convalescent - phase serum sample could be obtained for a confident exclusion of sars - cov infection by serologic testing , the nasopharyngeal aspirate collected on admission and the postmortem lung tissue were negative for sars - cov by rt - pcr .
the postmortem histologic findings on lung tissue of this patient were different from the classic picture of virus - associated interstitial pneumonitis .
in addition to the minimal inflammatory cell infiltration , multinucleated giant cells that might represent viral cytopathic effect were only occasionally seen .
rather , the predominant histologic finding was the presence of fibrin thrombi in small pulmonary arteries and arterioles .
this thrombotic event was confined to lung tissue and was not found in other tissues .
thrombotic vasculopathy has been observed in infectious diseases caused by hiv , cytomegalovirus , and herpes viruses ( 8,9 ) .
he was negative for hiv antibody and did not fulfill the diagnosis of thrombotic thrombocytopenic purpura .
our patient s signs and symptoms , together with his history of travel to southern china , fulfilled the world health organization criteria for a probable case of sars , but no evidence of coronavirus infection could be found .
the overall pathologic features of this case were distinct from those in the series of sars patients that we had examined and that were reported by others ( 10 ) . in sars patients , coronavirus particles
were seen by electron microscopy in lung cells of most cases , and the lung injuries consistently exhibited features of diffuse alveolar damage .
these sars - associated pathologic features were not observed in this patient , suggesting a different cause of death . in this patient ,
although the possibility of hmpv being a bystander could not be totally excluded , a role of hmpv in severe respiratory tract disease should not be discounted , particularly in sars patients that do not show evidence of coronavirus infection .
hmpv has also been detected in five of the six sars patients living in canada ; four of them were coinfected with coronavirus ( 11 ) .
the index patient of that cluster had stayed in hong kong before the onset of illness .
the patient described in this report was unlikely to have a direct link to the outbreak of sars in our hospital .
this patient was admitted around the same time that an outbreak of sars began in our hospital .
that outbreak mainly involved staff working at the accident and emergency department and the index ward where a sars patient had been treated .
the index ward is located on a different floor from where the patient described in this report was isolated .
in addition , neither fever nor respiratory illness developed in any of the healthcare workers involved in taking care of this patient in the next 2 weeks , and we were not aware of any spread to his family members .
although hmpv and sars - cov might differ in transmission efficiency , their role and clinical outcome in acute respiratory tract disease need to be distinguished .
this distinction is particularly important for analyzing the illness and death associated with the worldwide outbreak of sars .
patients fulfilling the clinical criteria for a probable case of sars might be infected by organisms other than coronavirus .
acute pneumonia developed in a previously healthy man during the outbreak of severe acute respiratory syndrome ( sars ) in southern china in march 2003 . 
 antibiotic treatment was ineffective , and he died 8 days after illness onset . 
 human metapneumovirus was isolated from lung tissue . 
 no other pathogen was found . 
 other etiologic agents should thus be sought in apparent sars cases when coronavirus infection can not be confirmed .