Foot-and-mouth disease (FMD) continues to be a major burden for livestock owners in endemic countries and a continuous threat to FMD-free countries. The epidemiology and control of FMD in Africa is complicated by the presence of five clinically indistinguishable serotypes. Of these the Southern African Territories (SAT) type 3 has received limited attention, likely due to its restricted distribution and it being less frequently detected. We investigated the intratypic genetic variation of the complete P1 capsid-coding region of 22 SAT3 viruses and confirmed the geographical distribution of five of the six SAT3 topotypes. The antigenic cross-reactivity of 12 SAT3 viruses against reference antisera was assessed by performing virus neutralization assays and calculating the r₁-values, which is a ratio of the heterologous neutralizing titer to the homologous neutralizing titer. Interestingly, cross-reactivity between the SAT3 reference antisera and many SAT3 viruses was notably high (r₁-values >0.3). Moreover, some of the SAT3 viruses reacted more strongly to the reference sera compared to the homologous virus (r₁-values >1). An increase in the avidity of the reference antisera to the heterologous viruses could explain some of the higher neutralization titers observed. Subsequently, we used the antigenic variability data and corresponding genetic and structural data to predict naturally occurring amino acid positions that correlate with antigenic changes. We identified four unique residues within the VP1, VP2, and VP3 proteins, associated with a change in cross-reactivity, with two sites that change simultaneously. The analysis of antigenic variation in the context of sequence differences is critical for both surveillance-informed selection of effective vaccines and the rational design of vaccine antigens tailored for specific geographic localities, using reverse genetics.

口蹄疫（FMD）仍然是流行国家牲畜饲养者的主要负担，也是对无口蹄疫国家的持续威胁。由于存在五种临床上无法区分的血清型，非洲口蹄疫的流行病学和控制变得复杂。其中，南部非洲领土 (SAT) 3 型受到的关注有限，可能是因为其分布范围有限且检测频率较低。我们研究了 22 种 SAT3 病毒的完整 P1 衣壳编码区的型内遗传变异，并确认了 6 种 SAT3 拓扑型中的 5 种的地理分布。通过进行病毒中和测定并计算r 1 值来评估12种SAT3病毒针对参考抗血清的抗原交叉反应性，r ₁值是异源中和滴度与同源中和滴度的比率。有趣的是，SAT3 参考抗血清和许多 SAT3 病毒之间的交叉反应性非常高（r ₁值 >0.3）。此外，与同源病毒相比，一些 SAT3 病毒对参考血清的反应更强烈（r ₁ -值> 1）。参考抗血清对异源病毒的亲和力的增加可以解释观察到的一些较高的中和滴度。随后，我们使用抗原变异性数据以及相应的遗传和结构数据来预测与抗原变化相关的自然发生的氨基酸位置。我们在 VP1、VP2 和 VP3 蛋白中鉴定出四个独特的残基，这些残基与交叉反应性的变化相关，其中两个位点同时变化。 在序列差异的背景下分析抗原变异对于根据监测信息选择有效疫苗以及利用反向遗传学合理设计针对特定地理区域的疫苗抗原至关重要。