新型弱毒活疫苗候选株对B亚型禽偏肺病毒的保护效果研究

doi:10.1016/j.jia.2023.10.025

摘要/Abstract

摘要：

禽偏肺病毒 (Avian metapneumovirus, aMPV)是一种高度传染性的病原体，可引起鸡和火鸡的急性上呼吸道疾病，造成严重的经济损失。目前，B亚型aMPV已成为中国鸡群中主要的优势流行毒株。本研究将B亚型aMPV LN16株在Vero细胞上进行连续传代，传至第30代时，成功获得了一株弱毒候选疫苗株，命名为LN16-A株。将LN16-A以2 × 10⁴ TCID₅₀/只的剂量接种SPF鸡后，接种鸡均未出现任何临床症状和病理损伤，表明LN16-A株已完全致弱，具备良好的安全性。为进一步评价LN16-A是否存在毒力返强风险，将LN16-A株在SPF鸡体内连续进行5次传代，观察每代鸡的临床症状并进行病理组织学检查和病毒载量检测，结果表明，不同传代的鸡均没有出现任何临床症状和病理损伤。此外，随着LN16-A株在鸡体内传代次数的增加，鼻甲骨中的病毒载量逐渐减少，传至第5代时，所有SPF鸡的鼻甲骨中病毒载量均为零。全基因组测序和分析结果表明，与野生型LN16（wtLN16）株的全基因组序列相比，LN16-A株的全基因组序列有9个氨基酸位点的突变，这些突变分别出现在N, P, F, SH, G和L基因上，表明这些基因可能与病毒的毒力有关。为进一步评价LN16-A株对SPF鸡的免疫保护效果，将LN16-A株以800 TCID₅₀/只的剂量免疫SPF鸡，免疫3周后进行攻毒。结果显示，LN16-A株免疫后可以诱导机体产生较高水平的中和抗体，上调CD4+T淋巴细胞亚群的比例并促进Th1（IFN-γ）和Th2（IL-4和IL-6）细胞因子的表达，这表明LN16-A株可以诱导机体产生强烈的体液免疫和细胞免疫。此外，攻毒保护试验结果表明，LN16-A株可对B亚型aMPV强毒攻击提供100%的免疫保护效果，并显著降低攻毒后的排毒水平和鼻甲骨炎症。这些结果表明，LN16-A是一株安全性好、具有良好免疫原性的弱毒活疫苗候选株，可以有效预防B亚型aMPV对鸡的感染。本研究首次以B亚型aMPV鸡源分离株LN16为研究对象，利用传代方法将其成功致弱，系统评价了其作为弱毒疫苗株的安全性和有效性，并分析了可能与病毒毒力有关的关键氨基酸位点，研究结果为进一步研究aMPV的致病机制和安全有效的新型疫苗奠定了基础。

Abstract:

Avian metapneumovirus (aMPV) is a highly contagious pathogen that causes acute upper respiratory tract diseases in chickens and turkeys, resulting in serious economic losses. Subtype B aMPV has recently become the dominant epidemic strain in China. We developed an attenuated aMPV subtype B strain by serial passaging in Vero cells and evaluated its safety and efficacy as a vaccine candidate. The safety test showed that after the 30th passage, the LN16-A strain was fully attenuated, as clinical signs of infection and histological lesions were absent after inoculation. The LN16-A strain did not revert to a virulent strain after five serial passages in chickens. The genomic sequence of LN16-A differed from that of the parent wild-type LN16 (wtLN16) strain and had nine amino acid mutations. In chickens, a single immunization with LN16-A induced robust humoral and cellular immune responses, including the abundant production of neutralizing antibodies, CD4⁺ T lymphocytes, and the Th1 (IFN-γ) and Th2 (IL-4 and IL-6)

cytokines. We also confirmed that LN16-A provided 100% protection against subtype B aMPV and significantly reduced viral shedding and turbinate inflammation. Our findings suggest that the LN16-A strain is a promising live attenuated vaccine candidate that can prevent infection with subtype B aMPV.

Key words: avian metapneumovirus , attenuated , protection , vaccine candidate , chickens

. 新型弱毒活疫苗候选株对B亚型禽偏肺病毒的保护效果研究[J]. Journal of Integrative Agriculture, 2024, 23(5): 1658-1670.

Lingzhai Meng, Mengmeng Yu, Suyan Wang, Yuntong Chen, Yuanling Bao, Peng Liu, Xiaoyan Feng, Tana He, Ru Guo, Tao Zhang, Mingxue Hu, Changjun Liu, Xiaole Qi, Kai Li, Li Gao, Yanping Zhang, Hongyu Cui, Yulong Gao.

A novel live attenuated vaccine candidate protects chickens against subtype B avian metapneumovirus [J]. Journal of Integrative Agriculture, 2024, 23(5): 1658-1670.

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