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Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent <t>IBDV</t> VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.
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Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent <t>IBDV</t> VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.
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Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent <t>IBDV</t> VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.
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Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent <t>IBDV</t> VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.
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Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent <t>IBDV</t> VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.
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Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent <t>IBDV</t> VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.
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Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent IBDV VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.

Journal: Poultry Science

Article Title: Development of a recombinant chimeric Newcastle disease virus-vectored vaccine conferring single-dose, triple protection against genotype VII NDV, IBDV, and H9N2 AIV

doi: 10.1016/j.psj.2026.106993

Figure Lengend Snippet: Generation and characterization of the recombinant rLaS-VIIF/HN-HA-VP2. (A) Schematic representation of the construction strategy for rLaS-VIIF/HN-HA-VP2. The open reading frames (ORFs) of the H9N2 AIV HA gene and the very virulent IBDV VP2 gene, each flanked by the non-coding region (NCR) sequences of the NDV HN gene, were inserted between the P and M genes and the F and HN genes of the pNDFL-VII-F/HN plasmid, respectively, to generate the pNDFL-VII-F/HN-HA-VP2 construct. (B) Western blot analysis of rLaS-VIIF/HN-HA-VP2 expression. Allantoic fluid harvested from embryonated eggs infected with rLaS-VIIF/HN-HA-VP2 was subjected to immunoblotting using specific antisera. Three specific bands corresponding to the uncleaved HA precursor (HA0, ∼62 kDa) and the cleaved subunits HA1 (∼47 kDa) and HA2 (∼25 kDa) were detected. Additionally, a specific band corresponding to the VP2 protein (∼42 kDa) was detected in the rLaS-VIIF/HN-HA-VP2 samples.

Article Snippet: Antibodies against IBDV were measured using the ProFLOK PLUS Infectious Bursal Disease Virus (IBD) Antibody Test Kit (Zoetis, USA) according to the manufacturer's instructions.

Techniques: Recombinant, Plasmid Preparation, Construct, Western Blot, Expressing, Infection

Humoral immune responses in SPF chickens immunized with rLaS-VIIF/HN-HA-VP2 and rLaS-VIIF/HN vaccines. Chickens were immunized via the ocular and nasal routes (100 µL per route) with a total of 200 µL containing 10⁶ EID₅₀ of rLaS-VIIF/HN-HA-VP2 or parental virus rLaS-VIIF/HN. Sera samples were collected before immunization and at day 10 and 20 post-immunization. (A) Antibodies against IBDV were measured using a commercial ELISA kit. (B, C) Antibodies against genotype VII NDV and H9N2 AIV were measured by hemagglutination inhibition (HI) test, respectively. Data are presented as mean ± SD; significance is denoted as * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001).

Journal: Poultry Science

Article Title: Development of a recombinant chimeric Newcastle disease virus-vectored vaccine conferring single-dose, triple protection against genotype VII NDV, IBDV, and H9N2 AIV

doi: 10.1016/j.psj.2026.106993

Figure Lengend Snippet: Humoral immune responses in SPF chickens immunized with rLaS-VIIF/HN-HA-VP2 and rLaS-VIIF/HN vaccines. Chickens were immunized via the ocular and nasal routes (100 µL per route) with a total of 200 µL containing 10⁶ EID₅₀ of rLaS-VIIF/HN-HA-VP2 or parental virus rLaS-VIIF/HN. Sera samples were collected before immunization and at day 10 and 20 post-immunization. (A) Antibodies against IBDV were measured using a commercial ELISA kit. (B, C) Antibodies against genotype VII NDV and H9N2 AIV were measured by hemagglutination inhibition (HI) test, respectively. Data are presented as mean ± SD; significance is denoted as * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001).

Article Snippet: Antibodies against IBDV were measured using the ProFLOK PLUS Infectious Bursal Disease Virus (IBD) Antibody Test Kit (Zoetis, USA) according to the manufacturer's instructions.

Techniques: Vaccines, Virus, Enzyme-linked Immunosorbent Assay, HI Assay

Protective efficacy of the rLaS‑VIIF/HN‑HA-VP2 vaccine against challenge with a very virulent IBDV strain. (A) Survival rates of chickens challenged with IBDV and monitored daily for 7 days post-challenge (dpc; n = 10 per group). (B) Representative gross lesions in the bursa of Fabricius at necropsy (7 dpc). Severe atrophy, accompanied by internal hemorrhage and necrosis, was evident in the bursae of birds in the rLaS‑VIIF/HN control group. In contrast, no notable gross pathological changes were observed in the rLaS‑VIIF/HN‑HA-VP2 immunized group or the uninfected controls. (C) Bursa-to-body weight index (BBIX) in immunized chickens following IBDV challenge. The mean BBIX value was significantly higher in the rLaS‑VIIF/HN‑HA-VP2 immunized group compared to the rLaS‑VIIF/HN control group (****, P < 0.0001). (D) Representative histopathological lesions in the bursa of Fabricius at 7 dpc (H&E stain; original magnification, 400 ×). Bursal tissue from birds in the rLaS‑VIIF/HN control group exhibited severe lymphoid follicle atrophy and lymphoid cell degeneration. No significant histopathological changes were observed in the rLaS‑VIIF/HN‑HA-VP2 immunized group or the uninfected controls.

Journal: Poultry Science

Article Title: Development of a recombinant chimeric Newcastle disease virus-vectored vaccine conferring single-dose, triple protection against genotype VII NDV, IBDV, and H9N2 AIV

doi: 10.1016/j.psj.2026.106993

Figure Lengend Snippet: Protective efficacy of the rLaS‑VIIF/HN‑HA-VP2 vaccine against challenge with a very virulent IBDV strain. (A) Survival rates of chickens challenged with IBDV and monitored daily for 7 days post-challenge (dpc; n = 10 per group). (B) Representative gross lesions in the bursa of Fabricius at necropsy (7 dpc). Severe atrophy, accompanied by internal hemorrhage and necrosis, was evident in the bursae of birds in the rLaS‑VIIF/HN control group. In contrast, no notable gross pathological changes were observed in the rLaS‑VIIF/HN‑HA-VP2 immunized group or the uninfected controls. (C) Bursa-to-body weight index (BBIX) in immunized chickens following IBDV challenge. The mean BBIX value was significantly higher in the rLaS‑VIIF/HN‑HA-VP2 immunized group compared to the rLaS‑VIIF/HN control group (****, P < 0.0001). (D) Representative histopathological lesions in the bursa of Fabricius at 7 dpc (H&E stain; original magnification, 400 ×). Bursal tissue from birds in the rLaS‑VIIF/HN control group exhibited severe lymphoid follicle atrophy and lymphoid cell degeneration. No significant histopathological changes were observed in the rLaS‑VIIF/HN‑HA-VP2 immunized group or the uninfected controls.

Article Snippet: Antibodies against IBDV were measured using the ProFLOK PLUS Infectious Bursal Disease Virus (IBD) Antibody Test Kit (Zoetis, USA) according to the manufacturer's instructions.

Techniques: Control, Staining