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SCIENCE CHINA Life Sciences, Volume 64 , Issue 5 : 752-765(2021) https://doi.org/10.1007/s11427-021-1904-4

Emergence and prevalence of naturally occurring lower virulent African swine fever viruses in domestic pigs in China in 2020

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  • ReceivedFeb 13, 2021
  • AcceptedFeb 23, 2021
  • PublishedFeb 26, 2021

Abstract


Funded by

the National Key Research and Development Program of China(2018YFC1200601)

Applied Technology Research and Development Project of Heilongjiang Province(GA19B301)

Key-Area Research and Development Program of Guangdong Province(2019B020211004)

the State Key Laboratory of Veterinary Biotechnology Program(SKLVBP201801)

and the National Natural Science Foundation of China(31941002)


Acknowledgment

This work was supported by the National Key Research and Development Program of China (2018YFC1200601), Applied Technology Research and Development Project of Heilongjiang Province (GA19B301), Key-Area Research and Development Program of Guangdong Province (2019B020211004), the State Key Laboratory of Veterinary Biotechnology Program (SKLVBP201801), and the National Natural Science Foundation of China (31941002). We thank Susan Watson for editing the manuscript.


Interest statement

The author(s) declare that they have no conflict of interest.


Supplement

SUPPORTING INFORMATION

The supporting information is available online at https://doi.org/10.1007/s11427-021-1904-4. The supporting materials are published as submitted, without typesetting or editing. The responsibility for scientific accuracy and content remains entirely with the authors.


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  • Figure 1

    Nucleotide insertions, deletions, mutations, and replacement of 22 different African swine fever viruses (ASFVs) isolated in China in 2020. The selected 8 ORFs or regions of 22 ASFVs were compared with the index ASFV isolate in China, HLJ/18 (Pig/HLJ/2018). The names of the ORFs or regions are shown on the top of each panel.

  • Figure 2

    Genetic analysis of the MGF_110-1L gene. A, Phylogenetic tree of the MGF_110-1L gene of 22 ASFVs isolated in 2020 and of HLJ/18. The unrooted tree was constructed by using the neighbor-joining method with a bootstrap value of 1,000 based on the MGF_110-1L gene sequences. B and C, The nucleotide difference and amino acid difference of the MGF_110-1L genes compared with that of HLJ/18.

  • Figure 3

    Characterization of selected ASFVs in vitro. A, PAMs were infected with the selected isolates, HuB/628/20, HLJ/44/20, HLJ/HRB1/20, and HeB/Q3/20 at an MOI of 0.1. The cells were fixed and analyzed by using an IFA at 24 h p.i. The HAD assay was performed for isolates. PAMs were seeded in 6-well plates and inoculated with ASFVs (MOI=0.2). The cell pellets were harvested for morphological assessment by using EM. B, Viral growth curves of isolates in PAMs. Primary PAMs were infected with virus at an MOI of 0.1, supernatants were collected, and viral p72 gene copy numbers were quantified by using qPCR.

  • Figure 4

    Lethality of field ASFV isolates in pigs. A and B, To determine the virulence of ASFVs in pigs, groups of four SPF pigs were inoculated with 102 and 103 HAD of HuB/628/20 and HLJ/44/20, respectively. C and D, Groups of four pigs were inoculated with 103 and 106 TCID50 of HLJ/HRB1/20 and HeB/Q3/20 viruses, respectively. Two naive pigs were cohoused with the pigs inoculated with the 106 TCID50 dose from the first day of inoculation. The pigs were monitored daily for disease signs and survival.

  • Figure 5

    Disease signs and gross lesions in HLJ/HRB1/20-infected pigs. Disease signs include arthroncus (A), paralysis (B), phyma (C), and cutaneous necrosis (D) in surviving pigs. Gross lesions include trichocardia (E), seroperitoneum (F), synarthrophysis (G), tonsil damage (H), and hyperaemia of lymph nodes (submaxillary lymph node (I), inguinal lymph node (J), and mediastinal lymph node (K)) in dead or euthanized HLJ/HRB1/20-infected pigs.

  • Figure 6

    Viral DNA copies in samples collected from HLJ/HRB1/20-infected pigs and contact pigs. Samples, including oral and rectal swabs and blood, were collected at the indicated timepoints from pigs inoculated with HLJ/HRB1/20 and from contact pigs. Viral p72 gene copies in the samples were quantified by using qPCR. The dashed black lines indicate the limit of detection with 1,000 copies per mL. The asterisks indicate that no samples were analyzed at the indicated timepoints because the pigs died.

  • Figure 7

    Viral DNA copies in organs collected from HLJ/HRB1/20-infected pigs and contact pigs. The indicated samples were collected from dead or surviving pigs that were euthanized on day 28 post-inoculation or post-contact. A and B, Pigs were infected with 106 TCID50 or 103 TCID50, respectively. C, The contact pigs were cohoused with the 106 TCID50-inoculated pigs. Viral p72 gene copies were quantified by using qPCR. The dashed black lines indicate the limit of detection. LN1, intestinal lymph node; LN2, inguinal lymph node; LN3, submaxillary lymph node; LN4, bronchial lymph node; LN5, gastrohepatic lymph node; and LN6, mediastinal lymph node.

  • Table 1   African swine fever viruses isolated in China in 2020

    Hemadsorptionphenotype

    Strain name

    Abbreviation

    Isolation time

    Isolation site (Province)

    Source

    Positive

    Pig/Hubei/628/2020

    HuB/628/20

    June 2020

    Hubei

    Lymph nodes

    Pig/Heilongjiang/44/2020

    HLJ/44/20

    July 2020

    Heilongjiang

    Spleen

    Pig/Jilin/1/2020

    JL/1/20

    July 2020

    Jilin

    Spleen

    Pig/Heilongjiang/BY2/2020

    HLJ/BY2/20

    September 2020

    Heilongjiang

    Spleen

    Pig/Liaoning/E24/2020

    LN/E24/20

    September 2020

    Liaoning

    Lymph nodes

    Pig/Hebei/Q1/2020

    HeB/Q1/20

    October 2020

    Hebei

    Blood

    Pig/Hebei/Q4/2020

    HeB/Q4/20

    November 2020

    Hebei

    Serum

    Pig/Liaoning/LA/2020

    LN/LA/20

    November 2020

    Liaoning

    Blood

    Pig/Liaoning/LB/2020

    LN/LB/20

    November 2020

    Liaoning

    Blood

    Pig/Liaoning/LC/2020

    LN/LC/20

    November 2020

    Liaoning

    Blood

    Pig/Liaoning/1068/2020

    LN/1068/20

    December 2020

    Liaoning

    Spleen

    Negative

    Pig/Hubei/1/2020

    HuB/1/20

    June 2020

    Hubei

    Spleen

    Pig/Hubei/4/2020

    HuB/4/20

    June 2020

    Hubei

    Spleen

    Pig/Hubei/SP1/2020

    HuB/SP1/20

    June 2020

    Hubei

    Lymph nodes

    Pig/Heilongjiang/HRB1/2020

    HLJ/HRB1/20

    September 2020

    Heilongjiang

    Spleen

    Pig/Heilongjiang/BLA3/2020

    HLJ/BLA3/20

    November 2020

    Heilongjiang

    Blood

    Pig/Heilongjiang/BLA6/2020

    HLJ/BLA6/20

    November 2020

    Heilongjiang

    Blood

    Pig/Hebei/Q3/2020

    HeB/Q3/20

    November 2020

    Hebei

    Serum

    Pig/Heilongjiang/JMS3/2020

    HLJ/JMS3/20

    December 2020

    Heilongjiang

    Blood

    Pig/Heilongjiang/JMS5/2020

    HLJ/JMS5/20

    December 2020

    Heilongjiang

    Blood

    Pig/Heilongjiang/JMS9/2020

    HLJ/JMS9/20

    December 2020

    Heilongjiang

    Blood

    Pig/Heilongjiang/JMS11/2020

    HLJ/JMS11/20

    December 2020

    Heilongjiang

    Blood

  • Table 2   Disease signs of pigs inoculated with different doses of ASFVs isolated in 2020

    Virus

    Dose

    Pig No.

    The earliest time when disease signs appeared (days post-inoculation, dpi)

    Fever

    Arthroncus

    Phyma

    Cutaneous necrosis

    Depression

    Death

    HuB/628/20

    103 HAD50

    1

    4

    /a)

    /

    /

    5

    7

    2

    4

    /

    /

    /

    5

    8

    3

    4

    /

    /

    /

    5

    6

    4

    3

    /

    /

    /

    5

    8

    102 HAD50

    1

    3

    /

    /

    /

    /

    5

    2

    4

    /

    /

    /

    5

    6

    3

    4

    /

    /

    /

    5

    6

    4

    4

    /

    /

    /

    5

    8

    HLJ/44/20

    103 HAD50

    1

    3

    /

    /

    /

    4

    5

    2

    4

    /

    /

    /

    4

    6

    3

    4

    /

    /

    /

    4

    6

    4

    4

    /

    /

    /

    4

    6

    102 HAD50

    1

    4

    /

    /

    /

    4

    6

    2

    4

    /

    /

    /

    4

    6

    3

    4

    /

    /

    /

    4

    6

    4

    3

    /

    /

    /

    4

    6

    HLJ/HRB1/20

    (non-HAD)

    106 TCID50

    1

    10

    18

    18

    /

    21

    24

    2

    10

    /

    /

    15

    /

    16

    3

    13

    18

    18

    22

    21

    23

    4

    22

    18

    /

    /

    25

    NAb)

    103 TCID50

    1

    17

    18

    20

    27

    27

    NA

    2

    /

    23

    /

    /

    /

    NA

    3

    /

    /

    /

    /

    /

    NA

    4

    9

    /

    /

    /

    /

    NA

    Contact

    1

    /

    /

    /

    /

    /

    NA

    2

    /

    /

    /

    /

    /

    NA

    HeB/Q3/20

    (non-HAD)

    106 TCID50

    1

    9

    10

    /

    /

    9

    NA

    2

    8

    13

    /

    /

    8

    14

    3

    8

    13

    10

    /

    8

    14

    4

    9

    9

    /

    /

    9

    NA

    103 TCID50

    1

    /

    /

    /

    /

    /

    NA

    2

    /

    /

    /

    /

    /

    NA

    3

    8

    /

    /

    /

    19

    NA

    4

    8

    /

    /

    /

    /

    NA

    Contact

    1

    16

    22

    20

    /

    18

    24

    2

    19

    /

    /

    /

    /

    NA

    No disease sign observed. b) The pig survived from the infection.

  • Table 3   Gross lesion of the tissues in the pigs inoculated with different doses of non-HAD HLJ/HRB1/20 virus

    Dose

    Pig No.

    Gross lesions of the tissues observed during necropsy

    Heart

    Seroperitoneum

    Lymph node

    Tonsil

    Spleen

    Lung

    Liver

    106 TCID50

    1

    Trichocardia, hydropericardium, 800 mL, primrose yellow, muddy

    500 mL,primrose yellow, clear

    Hyperaemia, swell

    Hyperaemia

    Hyperaemia

    Edema,extravasated blood

    Hyperaemia

    2

    /a)

    /

    Hyperaemia, swell

    Flush, fester

    Swell, petechia

    Extravasated blood

    /

    3

    Trichocardia, hydropericardium, 100 mL, crimson, muddy

    200 mL, puce, muddy

    Hyperaemia, swell

    Extravasated blood

    Hyperplasia of interstitia

    Edema,hyperaemia

    White spot on the surface

    4

    Trichocardia, hydropericardium, 500 mL, red, muddy

    /

    Hyperaemia, swell

    /

    /

    Carnification

    /

    103 TCID50

    1

    Hydropericardium, 20 mL, primrose yellow, clear

    /

    Hyperaemia, swell

    /

    /

    Carnification, hyperaemia

    /

    2

    Hydropericardium, 50 mL, damask, clear

    /

    Hyperaemia, swell

    /

    /

    /

    /

    3

    Hydropericardium, 100 mL, primrose yellow, muddy

    /

    Hyperaemia, swell

    /

    /

    Carnification,

    /

    4

    Hydropericardium, 50 mL, primrose yellow, muddy

    /

    Hyperaemia, swell

    /

    /

    Carnification, hemorrhagic spots

    /

    Contact

    1

    /

    /

    Hyperaemia, swell

    /

    /

    Atrophy,hemorrhagic spots

    /

    2

    /

    /

    Hyperaemia, swell

    /

    /

    Carnification, atrophy

    /

    No gross lesion observed.

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