Epidemiological and molecular analysis of avian influenza A(H7N9) virus in Shanghai, China, 2013–2017

Figures & data

Figure 1 Confirmed cases of avian influenza A(H7N9) virus infections in humans, birds, and the environment in Shanghai reported to the WHO and Food and Agriculture Organization (n=85), by month of illness onset, 2013–2017.

Notes: Origin: Publicly released infections in the Disease Outbreak News (http://www.who.int/csr/don/en/) or avian influenza report issued by the Department of Health, Government of the Hong Kong Special Administrative Region (http://www.chp.gov.hk/en/index.html).

Table 1 Confirmed human cases of avian influenza A(H7N9) virus in Shanghai as of 31 December 2017

Case no.	Date of onset*	Gender	Age (years)	Situation	Notes
1	2013-02-19	M	87	Died
2	2013-02-19	M	27	Died
3	2013-03-17	M	2
4	2013-03-22	F	67	Died
5	2013-03-25	M	58	Died
6	2013-03-27	F	52	Died
7	2013-03-28	M	73	Died
8	2013-03-28	M	48	Died
9	2013-03-29	M	65
10	2013-03-29	M	67
11	2013-03-31	M	4
12	2013-03-31	M	74	Died
13	2013-04-01	M	64	Died
14	2013-04-01	M	62
15	2013-04-01	F	76
16	2013-04-01	M	56	Died
17	2013-04-02	F	83	Died
18	2013-04-03	M	77	Died
19	2013-04-03	M	53
20	2013-04-03	M	86	Died
21	2013-04-04	F	81
22	2013-04-04	M	68
23	2013-04-04	M	78
24	2013-04-05	M	78
25	2013-04-05	M	73
26	2013-04-08	F	80	Died
27	2013-04-08	M	54
28	2013-04-10	M	47
29	2013-04-12	M	89	Died
30	2013-04-13	F	75
31	2013-04-13	M	68
32	2013-04-13	M	75
33	2013-04	N/A	N/A	Died
34	2013-12-26	M	86
35	2014-01-01	F	58	Critical
36	2014-01-01	M	35	Died
37	2014-01-03	M	54	Died
38	2014-01-06	F	61
39	2014-01-06	M	56	Critical
40	2014-01-10	M	77	Died
41	2014-01-11	M	31	Died
42	2014-11-28	M	38
43	2015-01-18	M	69
44	2015-01-22	M	73
45	2015-01-23	F	75
46	2015-02-12	M	53
47	2015-04-05	M	52
48	2015-04-07	F	37
49	2015-06-10	M	61
50	2015-12-21	M	59
51	2016-01-01	F	58
52	2016-01-30 (estimated)	M	66		2016-02-08 (WHO reported date)
53	2016-03 (estimated)	F	79		2016-03 (WHO reported date)
54	2016-12-12 (estimated)	M	45		2016-12-22 (WHO reported date)
55	2016-12-19 (estimated)	M	34		2016-12-29 (WHO reported date)
56	2016-12-29 (estimated)	M	58		2017-01-07 (WHO reported date)
57	2017-01-10 (estimated)	M	60	-	2017-01-11 (WHO reported date)
58	2017-01-31 (estimated)	M	58	-	2017-02-10 (WHO reported date)
59	2016-02-21 (estimated)	M	43		2017-03-03 (WHO reported date)

Notes: *Dates are presented as year-month-day. For eight infections whose date of onset was missing, the estimated onset date was calculated by subtracting 10 days from the WHO reported date.

Abbreviations: F, female; M, male; N/A, not available.

Figure 2 Age, gender, and death distributions of confirmed cases of avian influenza A(H7N9) virus infection during the five epidemics in Shanghai, 2013–2017.

Notes: (A) Gender distribution, (B) death distribution, (C) host distribution, and (D) age distribution in all epidemics (1st, February 2013–September 2013; 2nd, October 2013–September 2014; 3rd, October 2014–September 2015; 4th, October 2015–September 2016; 5th, October 2016–September 2017; and All, 2013–2017). One case (case no. 33) was excluded from A and D because the data were not available.

Table 2 Amino acid homology (%) of the eight genes of avian influenza A(H7N9) virus strains in Shanghai, China, 2013–2017

	HA	NA	MP	PB2	PB1	PA	NS	NP
Amino acid homology between strains	95.4-100	94.5-100	92.1-100	97.5-100	97.6-100	96.4-100	86.7-100	95.1-100

Abbreviations: HA, hemagglutinin; MP, matrix protein; NA, neuraminidase; NP, nucleoprotein; NS, non-structural protein; PA, polymerase acidic protein; PB1, polymerase basic protein 1; PB2, polymerase basic protein 2.

Table 3 Amino acid homology (%) of the surface HA and NA proteins between the avian influenza A(H7N9) Shanghai strains and WHO-recommended vaccine candidates

	A/Anhui/1/2013	A/Shanghai/02/2013	A/Guangdong/17SF003/2016	A/HongKong/125/2017
HA	96.7-100	96.7-100	94.9-97.8	95.1-98.9
NA	96.5-100	96.3-100	94.7-97.6	96.1-98.9

Abbreviations: HA, hemagglutinin; NA, neuraminidase.

Figure 3 Phylogenetic trees deduced using (A) the maximum-likelihood method and (B) the neighbor-joining method of HA, displaying the genetic divergences of the H7N9 viruses, Shanghai, 2013–2017.

Notes: The current study includes a total of 59 HA full sequences that were downloaded. The trees are rooted with A/turkey/Minnesota/1988 as the outgroup. The WHO-recommended vaccine candidates are indicated by a black circle.
Abbreviation: HA, hemagglutinin.

Figure 4 Phylogenetic trees deduced using (A) the maximum-likelihood method and (B) the neighbor-joining method of NA, displaying the genetic divergences of the H7N9 viruses, Shanghai, 2013–2017.

Notes: The current study includes a total of 77 NA full sequences that were downloaded. The trees are rooted with A/turkey/Minnesota/1988 as the outgroup. The WHO-recommended vaccine candidates are indicated by a black circle.
Abbreviation: NA, neuraminidase.

Table 4 Characterization of key molecular markers of avian influenza A(H7N9) viruses in Shanghai, 2013–2017

Gene	Function	Mutation	Amino acid	Human	Non-human	Total
HA*	Increased binding to humantype influenza receptor	G186V	G	3	2	5
			V	25	25	50
		Q226L/1	Q	1	3	4
			L/1	24	27	51
	Enhanced virulence in poultry	Cleavage site with insertion of multiple basic amino acids	PK^GR	28	27	55
NA*	Oseltamivir resistance	R292K	R	25	45	70
			K	3	0	3
	Increased virulence	69-73 deletion	QISNT	1	2	3
PB2	Increased virulence in mice	E627K	E	7	25	32
			K	19	1	20
	Enhanced transmission in guinea pigs	D701N	D	25	26	51
			N	1	0	1
PB1	Increased transmission in ferrets	1368V	1	2	0	2
			V	19	24	43
PA	Species-associated signature positions	V100A	V	2	8	10
			A	23	19	42
		K356R	K	0	2	2
			R	25	25	50
NS	Altered antiviral response in the host	N205s	N	0	0	0
			S	34	28	62
	Increased pathogenesis in mice	P42s	P	0	0	0
			S	34	28	62
	Altered virulence in mice	D92E	D	33	28	61
			e	1	0	1

Notes: *The HA gene was under the H3 numbering system and the NA gene was under the N2 numbering system. Other internal genes were numbered from the start codon.

Abbreviations: HA, hemagglutinin; NA, neuraminidase; NS, non-structural protein; PA, polymerase acidic protein; PB1, polymerase basic protein 1; PB2, polymerase basic protein 2; ↓, cleavage site.

Table 5 Signature amino acid mutations of avian influenza A(H7N9) viruses in Shanghai with WHO-recommended vaccine candidates, 2013–2017

Strain	Collection date*	HA			NA		PB2			PBI	PA		NS		D92E
		G 186V	Q226L/1	Cleavage site	R292K	69-73 deletion	E627K	D701N	K702R	1368V	V100A	K356R	N205S	P42S
A/turkey/Minnesota/1 /1988	1988-07-13			PENPKTRG		QISNT							N205S	P42S
A/Anhui/1/2013 (VC)	2013-01-03	G186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/Minhang/S01/2013	2013-02-27		Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/Shanghai/5190T/2013	2013	G186V	Q226L	PEIPKGR-1-GLF	R292K		E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/1/2013	2013-02-26		Q226L	PEIPKGR-1-GLF	R292K		E627K				V100A	K356R	N205S	P42S
A/Shanghai/3/2013	2013-02-27	G186V	Q226L	PEIPKGR-1-GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/02/2013	2013-03-05	G 186V	Q226L	PEIPKGR-1-GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/2/2013	2013-03-05	G 186V	Q226L	PEIPKGR-1-GLF						1368V			N205S	P42S
A/Shanghai/4664T/2013	2013-03-05	G 186V	Q226L	PEIPKGR-1-GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/4/2013	2013-03-09	G 186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/Shanghai/07/2013	2013-03-18	G 186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/environment/S hanghai/RLO 1 /2013	2013-03-30	G 186V	Q226L	PEIPKGR-1-GLF							V100A	K356R	N205S	P42S
A/chicken/Shanghai/017/2013	2013-04	G 186 V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/019/2013	2013-04	G 186 V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/pigeon/Shanghai/S 1069/2013	2013-04-02	G 186 V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/Shanghai/05/2013	2013-04-02			PEIPKGR-1-GLF		QISNT				1368V		K356R	N205S	P42S	D92E
A/chicken/Shanghai/S 1053/2013	2013-04-03	G 186 V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S 1055/2013	2013-04-03	G186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S 1076/2013	2013-04-03	G186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S 1077/2013	2013-04-03	G186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S 1078/2013	2013-04-03	G186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S 1079/2013	2013-04-03	G186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S 1080/2013	2013-04-03	G 186V	Q226L	PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S 1358/2013	2013-04-03	G 186V	Q226L	PEIPKGR-1-GLF						1368V		K356R	N205S	P42S
A/chicken/Shanghai/S 1410/2013	2013-04-03	G 186V		PEIPKGR-1-GLF						1368V	V100A	K356R	N205S	P42S
A/chicken/Shanghai/S1413/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/environment/Shanghai/S1088/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/environment/Shanghai/S1436/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V		K356R	N205S	P42S
A/environment/Shanghai/S1437/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V		K356R	N205S	P42S
A/environment/Shanghai/S1438/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V		K356R	N205S	P42S
A/environment/Shanghai/S1439/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/pigeon/Shanghai/1421/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/pigeon/Shanghai/S1423/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/Shanghai/17/2013	2013-04-03	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/ION/2013	2013-04-04	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/8/2013	2013-04-07	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/9/2013	2013-04-08	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/Shanghai/10/2013	2013-04-09	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/Shanghai/15/2013	2013-04-09	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/06-A/2013	2013-04-10	G186V	Q226L	PEIPKGR GL	R292K		E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/11/2013	2013-04-10	G186V	Q226L	PEIPKGR GLF			E627K				V100A	K356R	N205S	P42S
A/Shanghai/12/2013	2013-04-10	G186V	Q226L	PEIPKGR GLF			E627K			1368V			N205S	P42S
A/Shanghai/13/2013	2013-04-10	G186V	Q226I	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/14/2013	2013-04-10	G186V	Q226L	PEIPKGR GL			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/16/2013	2013-04-10	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/MH01/2013	2013-04-17	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/tree sparrow/Shanghai/01/2013	2013-05-09	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/01/2014	2014-01-03	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/02619/2014-HA	2014-01-03	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/PD-01/2014	2014-01-17	G186V	Q226L	PEIPKGR GLF			E627K			1368V	V100A	K356R	N205S	P42S
A/Shanghai/PD-02/2014	2014-01-17	G186V	Q226L	PEIPKGR GLF			E627K			1368V		K356R	N205S	P42S
A/chicken/Shanghai/PD-CN-02/2014	2014-01-21	G186V	Q226L	PEIPKGR GLF						1368V	V100A	K356R	N205S	P42S
A/environment/Shanghai/PD-JZ-01/2014	2014-01-21	G186V	Q226L	PEIPKGR GLF						1368V		K356R	N205S	P42S
A/duck/Shanghai/SD015/2015	2015-01-17			PEVPKGRG		QISNT				1368V			N205S	P42S
A/duck/Shanghai/SD016/2015	2015-01-17			PETPKGRG		QISNT				1368V			N205S	P42S
A/chicken/Shanghai/S4100/2015	2015-11-11	G186V	Q226L	PEIPKGR GLF						1368V		K356R	N205S	P42S
A/Guangdong/17SF003/2016-HA (VC)	2017-01-03	G186V		PEVPKRKRTARG	R292K				K702R	1368V		K356R	N205S	P42S
A/HongKong/125/2017 (VC)	2017-01-05	G186V	Q226L	PEIPKGRG					K702R	1368V		K356R	N205S	P42S

Note: *Dates are presented as year-month-day.

Abbreviations: HA, hemagglutinin; NA, neuraminidase; NS, non-structural protein; PA, polymerase acidic protein; PB1, polymerase basic protein 1; PB2, polymerase basic protein 2; ↓, cleavage site.