The Effect of In Vitro Cultivation on the Transcriptome of Adult Brugia malayi.
The Effect of In Vitro Cultivation on the Transcriptome of Adult Brugia malayi.
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DOI：
10.1371/journal.pntd.0004311
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发表时间：
2016-01
影响因子：
3.8
通讯作者：
Geary TG
中科院分区：
文献类型：
--
作者：
Ballesteros C;Tritten L;O'Neill M;Burkman E;Zaky WI;Xia J;Moorhead A;Williams SA;Geary TG
丝状线虫在热带国家的人口中造成严重和使人衰弱的感染，助长了根深蒂固的贫困循环。在实验室环境中，啮齿类动物体内只能维持一种人类丝虫病，马来布鲁氏菌。在采用培养系统的实验中，它是一种广泛使用的模式生物，对蠕虫的影响尚不清楚。利用Illumina RNA测序技术，我们在四个时间点对马来芽孢杆菌雌性成虫体外维持的基因表达变化进行了表征：从宿主移除后立即，在运输后立即收到，在液体培养基中48小时和5天后。剧烈的环境变化和从宿主到培养之间24小时的时间间隔导致了基因表达谱的全球失调。通过对时间点的两两比较，我们发现最多有562个差异表达基因。在离开宿主和运输后的最初冲击后，在培养48小时后，直到实验停止，仍有一些应力指纹。编码角质层胶原蛋白和蛇形蛋白的几个基因强烈而持续的上调就是最好的例证。这些发现表明，马来芽孢杆菌在离开宿主并适应新环境后，至少可以在培养物中维持数天，作为药理学和生物学研究的有效系统。然而，编码几种应激指标的基因在实验停止前仍处于失调状态。丝虫病感染会造成严重的身体损伤，影响到热带和亚热带国家数千万人。为了更好地了解这些寄生虫的生物学和药理学，马来布鲁贾经常被用作模型。这种寄生虫可以在蒙古鸟类的实验室中维持，使研究人员能够在体内和体外测试药物，以及其他研究。将蠕虫从宿主体内移除并进行培养可能会影响其生理的许多方面，包括对药物的反应，但蠕虫在培养过程中发生的变化程度尚不清楚。利用深度RNA测序和生物信息学工具，我们在培养的5天内检测了马来芽孢杆菌雌性在四个不同时间点的全球转录组学特征。关注在不同时间点差异表达的基因，我们观察到在宿主解剖和装运后接收之间表达谱的普遍扰动。实验结束时，在控制条件下5天后，几个基因的表达仍有变化；特别是，编码角质层胶原蛋白的基因被显著地表达和强烈地过表达。
Filarial nematodes cause serious and debilitating infections in human populations of tropical countries, contributing to an entrenched cycle of poverty. Only one human filarial parasite, Brugia malayi, can be maintained in rodents in the laboratory setting. It has been a widely used model organism in experiments that employ culture systems, the impact of which on the worms is unknown. Using Illumina RNA sequencing, we characterized changes in gene expression upon in vitro maintenance of adult B. malayi female worms at four time points: immediately upon removal from the host, immediately after receipt following shipment, and after 48 h and 5 days in liquid culture media. The dramatic environmental change and the 24 h time lapse between removal from the host and establishment in culture caused a globally dysregulated gene expression profile. We found a maximum of 562 differentially expressed genes based on pairwise comparison between time points. After an initial shock upon removal from the host and shipping, a few stress fingerprints remained after 48 h in culture and until the experiment was stopped. This was best illustrated by a strong and persistent up-regulation of several genes encoding cuticle collagens, as well as serpins. These findings suggest that B. malayi can be maintained in culture as a valid system for pharmacological and biological studies, at least for several days after removal from the host and adaptation to the new environment. However, genes encoding several stress indicators remained dysregulated until the experiment was stopped. Infections with filarial worms cause serious physical impairment and affect tens of millions of people in tropical and subtropical countries. To better understand the biology and pharmacology of these parasites, Brugia malayi is often used as a model. This parasite can be maintained in the laboratory in Mongolian jirds, enabling researchers to test drugs in vivo and in vitro, among other studies. The effects of removing worms from their hosts and culturing them may affect many aspects of their physiology, including response to drugs, but the extent to which the worms undergo changes during culture has remained unknown. Using deep RNA sequencing and bioinformatics tools, we examined the global transcriptomic profile of B. malayi females at four different time points over 5 days in culture. Focusing on genes that are differentially expressed at various time points, we observed a general perturbation of the expression profile between dissection from the host and receipt after shipment. The expression of several genes remained changed at the end of the experiment, after 5 days under controlled conditions; in particular, genes encoding cuticle collagens were prominently represented and strongly overexpressed.