Natural resources including water, energy, and food have an increase in demand due to the global population increases. The sustainable management of these resources is an urgent global issue. These resources combined in a very vital nexus are called the water–energy–food (WEF) nexus. The field of nanotechnology offers promising solutions to overcome several problems in the WEF nexus. This review is the first report that focuses on the suggested applications of nanofibers in the WEF sectors. An economic value of nanofibers in WEF sectors was confirmed, which was mainly successfully applied for producing clean water, sustainable energy, and safe food. Biotechnological solutions of nanofibers include various activities in water, energy, and food industries. These activities may include the production of fresh water and wastewater treatment, producing, converting, and storing energy, and different activities in the food sector. Furthermore, microbial applications of nanofibers in the biomedicine sector, and the most important biotechnological approaches, mainly plant tissue culture, are the specific focus of the current study. Applying nanofibers in the field of plant tissue culture is a promising approach because these nanofibers can prevent any microbial contamination under in vitro conditions, but the loss of media by evaporation is the main challenge in this application. The main challenges of nanofiber production and application depend on the type of nanofibers and their application. Different sectors are related to almost all activities in our life; however, enormous open questions still need to be answered, especially the green approach that can be used to solve the accumulative problems in those sectors. The need for research on integrated systems is also urgent in the nexus of WEF under the umbrella of environmental sustainability, global climate change, and the concept of one’s health.