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. 2024 Apr 7;16(4):571.
doi: 10.3390/v16040571.

Characterization of an African Swine Fever Virus Field Isolate from Vietnam with Deletions in the Left Variable Multigene Family Region

Aruna Ambagala  1   2   3 , Kalhari Goonewardene  1 , Ian El Kanoa  1 , Thi Tam Than  4 , Van Tam Nguyen  5 , Thi Ngoc Ha Lai  4 , Thi Lan Nguyen  4 , Cassidy N G Erdelyan  1 , Erin Robert  1   2 , Nikesh Tailor  1 , Chukwunonso Onyilagha  1 , Lindsey Lamboo  1 , Katherine Handel  1 , Michelle Nebroski  1 , Oksana Vernygora  1 , Oliver Lung  1 , Van Phan Le  4
Affiliations

Characterization of an African Swine Fever Virus Field Isolate from Vietnam with Deletions in the Left Variable Multigene Family Region

Aruna Ambagala et al. Viruses. .

Abstract

In this paper, we report the characterization of a genetically modified live-attenuated African swine fever virus (ASFV) field strain isolated from Vietnam. The isolate, ASFV-GUS-Vietnam, belongs to p72 genotype II, has six multi-gene family (MGF) genes deleted, and an Escherichia coli GusA gene (GUS) inserted. When six 6-8-week-old pigs were inoculated with ASFV-GUS-Vietnam oro-nasally (2 × 105 TCID50/pig), they developed viremia, mild fever, lethargy, and inappetence, and shed the virus in their oral and nasal secretions and feces. One of the pigs developed severe clinical signs and was euthanized 12 days post-infection, while the remaining five pigs recovered. When ASFV-GUS-Vietnam was inoculated intramuscularly (2 × 103 TCID50/pig) into four 6-8 weeks old pigs, they also developed viremia, mild fever, lethargy, inappetence, and shed the virus in their oral and nasal secretions and feces. Two contact pigs housed together with the four intramuscularly inoculated pigs, started to develop fever, viremia, loss of appetite, and lethargy 12 days post-contact, confirming horizontal transmission of ASFV-GUS-Vietnam. One of the contact pigs died of ASF on day 23 post-contact, while the other one recovered. The pigs that survived the exposure to ASFV-GUS-Vietnam via the mucosal or parenteral route were fully protected against the highly virulent ASFV Georgia 2007/1 challenge. This study showed that ASFV-GUS-Vietnam field isolate is able to induce complete protection in the majority of the pigs against highly virulent homologous ASFV challenge, but has the potential for horizontal transmission, and can be fatal in some animals. This study highlights the need for proper monitoring and surveillance when ASFV live-attenuated virus-based vaccines are used in the field for ASF control in endemic countries.

Keywords: African swine fever; GUS; MGF; horizontal transmission; live-attenuated virus; virulence.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Initial characterization of the ASFV-GUS-Vietnam field isolate. (A) Hemadsorption observed (arrows) in porcine primary alveolar macrophage (PAM) cultures. (B) Confirmation of GUS insertion in ASFV-GUS-Vietnam isolated from a serum sample. The PCR amplicons were resolved by gel electrophoresis (1%). Lane 1 and 7: 100 bp Ladder. Lane 2: ASFV-GUS-Vietnam DNA. Lane 3: ASFV Georgia 2007/1 DNA. Lane 4: extraction control. Lane 5; no template control. Lane 6: E. coli DNA (positive control).
Figure 2
Figure 2
Rectal temperatures of pigs inoculated ON with ASFV-GUS-Vietnam (2 × 105 TCID50/pig) and challenged IM with ASFV Georgia 2007/1 IM (1 × 103 TCID50/pig). dpi = days post-infection.
Figure 3
Figure 3
ASFV genome detection in clinical samples from pigs inoculated (ON) with ASFV-GUS-Vietnam (2 × 105 TCID50/pig) and challenged (IM) with ASFV Georgia 2007/1 (1 × 103 TCID50/pig). (A) Whole blood, (B) oropharyngeal swabs (OPSWs), (C) buccal swabs (BSWs), (D) nasal swabs (NSWs), (E) rectal swabs (RSWs), and (F) oral fluids. dpi = days post-infection.
Figure 3
Figure 3
ASFV genome detection in clinical samples from pigs inoculated (ON) with ASFV-GUS-Vietnam (2 × 105 TCID50/pig) and challenged (IM) with ASFV Georgia 2007/1 (1 × 103 TCID50/pig). (A) Whole blood, (B) oropharyngeal swabs (OPSWs), (C) buccal swabs (BSWs), (D) nasal swabs (NSWs), (E) rectal swabs (RSWs), and (F) oral fluids. dpi = days post-infection.
Figure 4
Figure 4
Detection of antibodies in sera collected from pigs inoculated ON with ASFV-GUS-Vietnam (2 × 105 TCID50/pig) and challenged IM with ASFV Georgia 2007/1 (1 × 103 TCID50/pig). dpi = days post-infection.
Figure 5
Figure 5
Rectal temperatures of pigs inoculated IM with ASFV-GUS-Vietnam (2 × 103 TCID50/pig) and challenged ON with ASFV Georgia 2007/1 (2 × 105 TCID50/pig). dpi = days post-infection.
Figure 6
Figure 6
ASFV genome detection in clinical samples from pigs inoculated IM with ASFV-GUS-Vietnam (2 × 103 TCID50/pig) and challenged ON with ASFV Georgia 2007/1 (2 × 105 TCID50/pig): (A) whole blood, (B) oropharyngeal swabs (OPSWs), (C) buccal swabs (BSWs), (D) nasal swabs (NSWs), (E) rectal swabs (RSWs), and (F) oral fluids. dpi = days post-infection.
Figure 6
Figure 6
ASFV genome detection in clinical samples from pigs inoculated IM with ASFV-GUS-Vietnam (2 × 103 TCID50/pig) and challenged ON with ASFV Georgia 2007/1 (2 × 105 TCID50/pig): (A) whole blood, (B) oropharyngeal swabs (OPSWs), (C) buccal swabs (BSWs), (D) nasal swabs (NSWs), (E) rectal swabs (RSWs), and (F) oral fluids. dpi = days post-infection.
Figure 7
Figure 7
Detection of antibodies in sera collected from pigs inoculated IM with ASFV-GUS-Vietnam (2 × 103 TCID50/pig) and challenged ON with ASFV Georgia 2007/1 (2 × 105 TCID50/pig). dpi = days post-infection.
Figure 8
Figure 8
Gross pathology observed in pigs #1 and 14. (A) Cyanosis in the ear tip (pig #1), (B) ecchymotic hemorrhages in the skin (pig #1), (C) serosanguineous peritoneal fluid (pig #14), (D) enlarged spleen (pig #14), (E) serosanguineous pericardial effusion (pig #1), (F) epicardial hemorrhages (pig #1), (G) consolidated lungs (pig #1), (H) hemorrhagic submandibular lymph nodes (pig #14), (I) hemorrhagic gastro-hepatic lymph nodes (pig #1), (J) hemorrhages in the bladder (pig #1), (K) petechial hemorrhages in the renal cortex (pig #14), and (L) hemorrhagic gall bladder (pig #14).
Figure 9
Figure 9
Detection of GUS (A) and MGF 505-2R (B) in the ASFV-GUS-Vietnam inoculum and in bone marrow (BM) samples collected from pigs #1 and #14 that died of ASF. NTC = no template control. MW = 100 bp ladder. E. coli = E. coli DNA.
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