Rabies virus co-localizes with early(Rab5) and late(Rab7) endosomal proteins in neuronal and SH-SY5Y cells

Rabies virus(RABV) is a highly neurotropic virus that follows clathrin-mediated endocytosis and p H-dependent pathway for trafficking and invasion into endothelial cells. Early(Rab5, EEA1) and late(Rab7, LAMP1) endosomal proteins play critical roles in endosomal sorting, maturity and targeting various molecular cargoes, but their precise functions in the early stage of RABV neuronal infection remain elusive. In this study, the relationship between enigmatic entry of RABV with these endosomal proteins into neuronal and SH-SY5 Y cells was investigated.Immunofluorescence, TCID_(50) titers, electron microscopy and western blotting were carried out to determine the molecular interaction of the nucleoprotein(N) of RABV with early or late endosomal proteins in these cell lines. The expression of N was also determined by down-regulating Rab5 and Rab7 in both cell lines through RNA interference. The results were indicative that N proficiently colocalized with Rab5/EEA1 and Rab7/LAMP1 in both cell lines at 24 and 48 h post-infection, while N titers significantly decreased in early infection of RABV. Down-regulation of Rab5 and Rab7 did not inhibit N expression, but it prevented productive infection via blocking the normal trafficking of RABV in a low pH environment. Ultrathin sections of cells studied by electron microscope also verified the close association of RABV with Rab5 and Rab7 in neurons. From the data it was concluded that primary entry of RABV strongly correlates with the kinetics of Rab-proteins present on early and late vesicles, which provides helpful clues to explain the early events of RABV in nerve cells.

Countrywide Survey for MERS-Coronavirus Antibodies in Dromedaries and Humans in Pakistan

Middle East Respiratory Syndrome Coronavirus (MERS-CoV) is a zoonotic pathogen capable of causing severe respiratory disease in humans. Although dromedary camels are considered as a major reservoir host, the MERS-CoV infection dynamics in camels are not fully understood. Through surveillance in Pakistan, nasal (n = 776) and serum (n = 1050)samples were collected from camels between November 2015 and February 2018. Samples were collected from animal markets, free-roaming herds and abattoirs. An in-house ELISA was developed to detect IgG against MERS-CoV. A total of 794 camels were found seropositive for MERS-CoV. Prevalence increased with the age and the highest seroprevalence was recorded in camels aged [ 10 years (81.37%) followed by those aged 3.1–10 years (78.65%) and B 3 years (58.19%).Higher prevalence was observed in female (78.13%) as compared to male (70.70%). Of the camel nasal swabs, 22 were found to be positive by RT-qPCR though with high Ct values. Moreover, 2,409 human serum samples were also collected from four provinces of Pakistan during 2016–2017. Among the sampled population, 840 humans were camel herders.Although we found a high rate of MERS-CoV antibody positive dromedaries (75.62%) in Pakistan, no neutralizing antibodies were detected in humans with and without contact to camels.

狂犬病病毒入胞途径的初步探索

狂犬病病毒（rabies virus,RABV）具有高度嗜神经性,通过识别细胞膜上特异性受体并借助内吞途径侵入细胞。为了探究其入胞途径,本研究在人神经瘤细胞（SH-SY5Y）和非洲绿猴肾上皮细胞（Vero）上,应用氯丙嗪（chlorpromazine）、制霉菌素（nystatin）、巴弗洛霉素a1（bafilomycin a1）、dynasore等抑制剂处理细胞后,进行RABV感染,通过检测病毒N基因拷贝数和RABV效价,对RABV入胞途径进行初步探究。结果显示,RABV通过网格蛋白介导、低pH值和发动蛋白（dynamin）依赖途径侵入SH-SY5Y和Vero细胞,但不通过小窝蛋白依赖途径入胞。这些结果为进一步研究狂犬病病毒感染机制和药物靶点研究提供了新的数据。