Screening of virus infection in febrile patients with suspected malaria in the Brazilian Amazon

Arthropod-borne viruses (arboviruses) represent a threat to global public health, especially in the tropical and subtropical regions of the world. More than 150 arboviruses can infect humans; they cause mainly febrile illness, although hemorrhagic complications and diseases affecting the central nervous system can also be observed. Arboviruses represent a threat to Brazil and, therefore, a permanent surveillance of these viruses is required to timely reduce the risk of epidemic outbreaks. The Brazilian Amazon region is where the highest number of arboviruses has been detected in the world. Besides, malaria is also endemic in the Amazon region, with a significant predominance of Plasmodium vivax. It is often difficult to differentiate between malaria and arboviral diseases, as they share similar clinical features and laboratory findings, mainly undifferentiated fever. This study aimed to estimate possible viral etiology in patients with febrile syndrome negative for Plasmodium infection, in the Brazilian Amazon. We initially analyzed serum samples of 124 participants with a DNA microarray platform designed for the detection of arboviruses and viruses transmitted by small mammals, but no virus was detected. Then, the serum samples of 76 participants were analyzed with a deep New Generation Sequencing, which showed evidence of the presence of only one arbovirus, the Zika virus in only one pool of 9 serum samples. This result is in contrast with our hypothesis, showing that arboviruses are not frequent in suspected malaria cases in Manaus, Brazil. Other viruses instead of arboviruses were found in this study. Primate erythrovirus 1 was the virus most frequently found virus in the suspected malaria patients, followed by Enterobacteria phage lambda. Besides, we detected, in a lower frequency, the Pegivirus C. In addition to the exogenous viruses, we also detected human endogenous retrovirus in all pools. Due to the high number of viruses that are important in the differential diagnosis of malaria, cost-effective and simple high throughput methods are required, helping molecular surveillance of misdiagnosed viral infections. Further studies with more robust sample sizes in other areas in the Amazon are needed.

虫媒病毒（Arthropod-borne viruses, arboviruses）对全球公共卫生构成威胁，尤以全球热带及亚热带地区为重。目前已有超过150种虫媒病毒可感染人类，此类病毒主要引发发热性疾病，但也可能出现出血并发症及累及中枢神经系统的病症。虫媒病毒对巴西同样构成威胁，因此需对这类病毒开展持续监测，以及时降低疫情暴发风险。巴西亚马逊地区是全球范围内检出虫媒病毒数量最多的区域。此外，亚马逊地区同样存在疟疾流行，且以间日疟原虫（Plasmodium vivax）感染为主。由于疟疾与虫媒病毒病存在相似的临床特征与实验室检测结果，尤以不明原因发热为主要表现，二者往往难以鉴别。本研究旨在探究巴西亚马逊地区疟原虫检测阴性的发热综合征患者的潜在病毒病因。研究初期，我们采用针对虫媒病毒及小型哺乳动物传播病毒设计的DNA微阵列平台，对124名受试者的血清样本进行了检测，但未检出任何病毒。随后，我们采用深度新一代测序技术对76名受试者的血清样本进行分析，仅在1份由9份血清样本组成的混合样本中检出1种虫媒病毒——寨卡病毒（Zika virus）。该结果与本研究的初始假设相悖，表明在巴西马瑙斯市的疑似疟疾病例中，虫媒病毒感染并不常见。本研究还检出了非虫媒类病毒：在疑似疟疾患者中，检出频率最高的为灵长类红细胞病毒1型（Primate erythrovirus 1），其次是肠杆菌噬菌体λ（Enterobacteria phage lambda）。此外，我们还以较低检出频率检测到佩吉病毒C型（Pegivirus C）。除外源性病毒外，我们在所有混合样本中均检出了人类内源性逆转录病毒（human endogenous retrovirus）。鉴于有诸多病毒需纳入疟疾鉴别诊断范畴，亟需开发兼具成本效益与简便性的高通量检测方法，以助力对误诊病毒感染的分子监测。未来需在亚马逊地区其他区域开展更大样本量的后续研究。