Pathogenic bacteria and viruses are the main causes of infectious diseases all over the world. Early diagnosis of such infectious diseases is a critical step in management of their spread and treatment of the infection in its early stages. Therefore, the innovation of smart sensing platforms for point-of-care diagnosis of life-threatening infectious diseases such as COVID-19 is a prerequisite to isolate the patients and provide them with suitable treatment strategies. The developed diagnostic sensors should be highly sensitive, specific, ultrafast, portable, cheap, label-free, and selective. In recent years, different nanosensors have been developed for the detection of bacterial and viral pathogens. We focus here on label-free miniaturized nanosensing platforms that were efficiently applied for pathogenic detection in biological matrices. Such devices include nanopore sensors and nanostructure-integrated lab-on-a-chip sensors that are characterized by portability, simplicity, cost-effectiveness, and ultrafast analysis because they avoid the time-consuming sample preparation steps. Furthermore, nanopore-based sensors could afford single-molecule counting of viruses in biological specimens, yielding high-sensitivity and high-accuracy detection. Moreover, non-invasive nanosensors that are capable of detecting volatile organic compounds emitted from the diseased organ to the skin, urine, or exhaled breath were also reviewed. The merits and applications of all these nanosensors for analysis of pathogenic bacteria and viruses in biological matrices will be discussed in detail, emphasizing the importance of artificial intelligence in advancing specific nanosensors.