The patient was from Guangzhou, the capital of the southern province of Guangdong. A fever (39°C) developed on February 22, 2006. He was hospitalized on February 26 and died on March 2. Diagnosis of influenza virus infection was made on March 3. Throat swab specimens obtained on March 1 and 2 tested positive for HPAI H5N1 virus by reverse transcription (RT)–PCR. Virus was isolated and named A/Guangzhou/1/2006 (H5N1).
Epidemiologic studies showed that the patient did not slaughter, process, or cook birds. However, while looking for work before his illness, he visited 9 food markets that had live birds. All 9 markets were located in the central part of the city (). He visited food market F twice a day from January 23 to 27 and food market G on February 17 for 30 minutes. Before his illness, he and his girlfriend (whom he lived with) shopped at markets B and F on February 20–22. He also visited food market I from February 10 to February 20. The dates he visited the other food markets could not be determined. Onset of fever occurred on February 22.
Serum and swab sample results from live birds and animal cages sampled at markets in Guangzhou, People's Republic of China*
The food markets were typically large, clean, and well managed and had vendors selling vegetables, fruits, raw and cooked meats, food flavorings, beverages, and other goods. They are typical of larger food markets in cities in the People's Republic of China. The only difference between markets in Guanzhou in southern China and those in cities in northern China is that more (2–9
) booths are used to sell live birds in Guanzhou. Wire cages are stacked next to each other with ≈5–10 birds in each cage (chickens, geese, ducks, and pigeons). Each species of bird is placed in separate cages; chickens are the most common species. All cages are located in a closed room separated by a glass window from customers, who choose the bird they prefer. When a live bird is selected, it is slaughtered in view of the customer. Sanitation inspections are routinely performed by municipal authorities. No diseased or dead birds were observed during this investigation.
Animal cages were swabbed and anal swabs of live birds were obtained at the food markets () on March 3 and 4 and tested for HPAI by using RT-PCR (3
) for the hemagglutinin (H5), neuraminidase (N1), and membrane (M) genes. Positive PCR results were confirmed by sequencing. None of 94 anal swabs from live birds tested positive for HPAI H5N1. However, 1 of 79 animal cage swabs tested positive for HPAI H5N1 (). The positive swab was from a goose cage at market I (), the market that the patient visited from February 10 to February 20. The nucleotide sequences of H and M genes from specimens from this patient were compared with those from the animal cage swab and submitted to GenBank (accession nos. DQ842487–90). Forty-eight variations were found in the NA gene and 15 were found in the HA gene, which resulted in 17 HA amino acid and 3 NA amino acid changes, respectively. Phylogenetic analysis with the neighbor-joining method using the ClustalX program (4
) suggested that the 2 strains are related to each other and to duck isolates ().
Figure 1 Detection of avian influenza virus H5N1 from an animal cage for geese by reverse transcription–PCR. Viral RNA was extracted from the sample and amplified by using 3 pairs of primers specific for membrane (M), hemagglutinin (H5), and neuraminidase (more ...)
Figure 2 Phylogenetic relationships of representative H5N1 influenza virus strains and patient and animal cage isolates (indicated by asterisks) used in this study. A) Hemagglutinin gene (nt positions 29–1650). B) Neuramidase gene (nt positions 28–1323). (more ...)
Serum samples were obtained from 110 of 121 poultry purveyors working at the live bird food markets and screened for antibody to H5N1 to determine if subclinical infections occurred. One of 110 serum samples was positive (titer 320) by hemagglutination-inhibition assay with turkey erythrocytes (Lampire Biologic Laboratories, Pipersville, PA, USA) and H5N1 virus strains A/Hong Kong/486/97 and A/Vietnam/1194/04/H5N1 (5
). Neutralizing antibody titers against the 2 strains of virus were 1,280 and 640, respectively. The positive serum sample was from a 44-year-old man who slaughtered birds for 5 years. He slaughtered ≈100 chickens/day and did not report any recent respiratory diseases. He denied any contact with ill birds.