登革热由登革病毒（DENV）引起，主要由埃及伊蚊、白纹伊蚊叮咬吸血传播，是近几十年在全球扩散最快的重要蚊媒传染病。世界半数以上人口生活在登革热感染风险中。持续的城市化、全球化和气候变化深刻影响传染病的流行传播，并逐步重塑病毒种群的分子系统地理结构，频发的重大传染病疫情给人类生存和全球公共卫生带来前所未有的挑战。近年来，以系统地理学和系统动力学为理论基础研发的多种分析方法和工具，帮助我们在病毒物种的分子进化水平更为精准地明晰和理解病毒传播扩散机制。当前针对SARS-CoV-2的高分辨率基因分型系列研究，在传染病流行传播中活跃谱系及其传播链的快速追踪溯源方面呈现出强大的解析能力。尽管基于媒介蚊虫生态数据和登革热流行数据进行疾病负担评估和预测已在全球范围内开展，目前的DENV常规监测乃至未来的实时监测预警，仍亟需构建具有高分辨率且整合时空流行病学背景信息的DENV全球统一基因分型框架，对于自全球视野探究DENV的种群遗传结构及其传播动力学规律，并建立全球DENV协同监测体系，至关重要。
DENV血清1型（DENV-1）广泛流行于亚洲、美洲和大洋洲，近年来由DENV-1引发的登革热疫情在中国、意大利等多国报道呈现向高纬度地区不断扩散趋势。本研究聚焦于DENV-1, 采集1944 - 2018年78个流行国家/地区5003株符合纳入条件的DENV-1完整E基因序列，整合相关时空流行病学背景信息，基于系统发育学、种群遗传学、系统地理学和系统动力学建立了统一的DENV-1全球高分辨率基因分型框架，其包含5个基因型（Genotype）、41个基因亚型（Subgenotype）和210个基因分支（Clade），三级基因分型的平均遗传距离（MPDs）分别为2-6%，0.8-2%和≦0.8%。
一方面，本研究揭示了DENV-1种群分布呈现显著的地理限制性特征，表征为洲际-基因型（Continent-Genotype）、区域-基因亚型（Region-Subgenotype）和国家-分支（Nation-Clade）的分层流行配对（stratified spatial-genetic epidemic pairs）规律，从而确定和划分了全球12个可促进DENV-1区域性协同监测控制的流行区域（Regions），包括: 西非地区（WAFR）、红海地区（RSR）、西南印度洋地区（SWIO）、南亚次大陆（SASC）、湄公河-中国地区（GMS-China）、东南亚（SEA）、菲律宾群岛（PHI）、西太平洋地区（TWP）、太平洋地区（OCE）、中北美地区（CNA）、加勒比海地区（CAR）和南美地区（SA）。流行区域内近邻国家/地区之间病毒亚群相似，92.3%的基因亚型中来自同一地区的毒株比例≧50%。
另一方面，虽然区域内DENV-1的基因亚型在总体趋势上呈现明显的空间传播限制，但仍有如5K（在SEA、SASC、SA、CAR、CNA、OCE、SWIO和WAFR地区）、5C（GMS-China, SEA和SASC）、5M与5N（SA、CAR和CNA）等多个基因亚型出现显著的跨区域扩张。相比DENV-1的跨洲际传播，洲际内部区域及其近邻区域传播则更为频繁，尤其是GMS-China、SEA和SA等区域的交互传播活跃度高。这在一定程度上诠释了DENV-1近年来在全球的快速播散，并提醒我们需要重点关注跨区域甚至跨洲际扩散的可能引发大规模流行的DENV-1亚群。因而，基于本研究所建立的DENV-1统一分型框架，及其所呈示的病毒遗传种群具体分布情况，可迅速展开特定区域内/间的国际协作，追踪溯源并实时更新DENV-1基因亚群的新发、再现、传播与漂移，以形成具有高成本效益的登革热区域乃至全球监测控制战略。
进一步基于此框架展开对DENV-1全球传播模式的探究，发现泰国、越南、印度尼西亚、菲律宾、印度和巴西等传统的登革热本土流行国家（endemic countries）仍是持续的DENV-1传染源输出中心，而以输入性疫情为主的中国、澳大利亚和美国等登革热疫情新发流行国家（emerging epidemic countries）则表现出日趋增强的DENV-1扩散趋势。同时，孤支现象（singleton）发生在12个DENV-1基因亚型仅有单一进化支，63个基因分支仅有单条毒株序列，提示其原因之一可能是由于监测不足，尤其是在非洲和印度次大陆，DENV-1毒株序列报道早，却分别在1970s和1990s出现跨越十年的零毒株序列报道，提示因监测缺失而存在的隐匿疫情。因而，利用该全球统一基因分型框架，基于在一个或多个国家/地区共同传播的DENV-1基因亚型和分支建立精准的日常监测控制协同体系，对控制DENV-1的规模化暴发流行、阻遏本土化进程以及解决监测不足的问题均具有积极的推进作用。
总之，本研究揭示了基于该全球统一DENV-1基因分型框架建立分层协同的全球监测控制策略与体系的可行性、必要性和迫切性，以精准阻遏登革热在全球的快速扩散。同时该分型框架也有望将经典的DENV E基因分型与基因组流行病学以及风险建模相关联，展现出在流行定量评估等方面的应用前景。研究团队同时表示将保持对该分型框架的定期更新与发布。
Urbanization, climate change, and globalization including rising global tourism are re-shaping the phylogeographic structure of virus populations, possibly contributing to a succession of severe viral pandemics as an unprecedented challenge to global public health. To investigate viral dispersion mechanisms in phylogeography and phylodynamics, various analytical methods and general genotyping methods have been developed for more accurate pathogen identification beyond viral species. Recently, the high-resolution genotyping scheme for surveillance manifested itself to be powerful in rapidly tracking the active lineages and transmission chains of SARS-CoV-2. Even though disease burden assessment and prediction using mosquito ecological data and dengue epidemic data have been investigated globally, there is still lack of a systematic scheme for understanding the DENV population structure, dynamics and mechanisms of cross-national, cross-regional, and even cross-continental transmission in a global profile.
Targeting DENV-1 spreading prominently in recent decades, by reconciling all qualified complete E gene sequences of 5003 DENV-1 strains, with the relative epidemiological information confirmed by literature research, from 78 epidemic countries/areas ranging from 1944 to 2018, the present study has established a unified global high-resolution genotyping framework of DENV-1 designating three hierarchical layers of clade, subgenotype and genotype with respective mean pairwise distances (MPDs)≦0.8%, 0.8-2%, and 2-6%.
On the one hand, the global epidemic patterns of DENV-1 showed strong geographic constraints representing stratified spatial-genetic epidemic pairs of Continent-Genotype, Region-Subgenotype and Nation-Clade. Thereby 12 epidemic regions have been identified including: GMS-China, Great Mekong Subregion-China; SEA, Southeast Asia; PHI, Philippines; TWP, Tropical Western Pacific; OCE, Oceania; WAFR, West African Region; RSR, Red Sea Region; SWIO, Southwest Indian Ocean; SASC, South Asia Subcontinent; CNA, Central North America; CAR, Caribbean; and SA, South America, which prospectively facilitates the region-based coordination. As for the pair of the Region-Subgenotype, 92.3% and 71.8% of the 39 designated subgenotypes, with ≥50% and ≥80% isolates respectively, mainly circulated in a single region.
On the other hand, the present study also reminds us to be concerned about the cross-regional and even cross-continental diffusion of DENV-1, which potentially arouse large epidemics, in reference to the stratified spatial-genetic epidemic pairs. For example, several subgenotypes including 5K, 5B, 4F, 5C, 5M and 5N were found spreading across several regions. The cross-regional transmissions were mostly observed in adjacent regions, especially in the regions of Asian or American Continents, but only a few cross-continental transmissions occurred. Collectively, the transmissions of subgenotypes cross countries/regions, on the rise in recent decades, can explain its continuous global expansion to some extent. Therefore, international collaborations based on the framework in promptly pinpointing and periodically updating the hotspots of emerging, transmission, drifting, and replacement of DENV-1 will give rise to cost-effective surveillance strategy for regional and global control of dengue.
In addition, through in-depth explorations of the global transmission pattern of DENV-1 using this framework, we observed that the traditional endemic countries such as Thailand, Vietnam, Indonesia, the Philippines, India, and Brazil apparently displayed as the persisting dominant source centers, while the emerging epidemic countries such as China, Australia, and the USA, where dengue outbreaks were frequently triggered by importation, showed a growing trend of DENV-1 diffusion. Meanwhile, the singletons in subgenotypes (12/41) and clades (63/208) recognized in our study suggest that they may be partially caused by widely deficient surveillance. The probably hidden epidemics were found especially in Africa and India. Then, our framework can be utilized in an accurate daily coordinated surveillance based on the population compositions of its clades and subgenotypes, which is prospectively valuable for hampering the ongoing transition process of epidemic to endemics and addressing the issue of inadequate monitoring.
The genotyping framework and its utilization in quantitatively assessing DENV-1 epidemics has laid a foundation and re-unveiled the urgency for establishing a stratified coordinated surveillance platform for blocking global spreading of dengue. This framework is moreover expected to bridge classical DENV-1 genotyping with genomic epidemiology and even pathogen-mosquito-disease-associated risk modelling. The authors will promote the public availability to the framework and keep periodic updating of it as well.
