It is said “we all live in the past.” It is probably true. If so, how we can use the past to foresee the future could then be our task at present.
While preparing this manuscript, SARS-CoV-2 still continues circulating in human populations and threatening public health. This pandemic has further stressed the importance of closely monitoring the emergence of infectious diseases originating from animal reservoirs and transmitted between animals and people. It is obvious that this pandemics will not be the last; a powerful experimental evolution approach allowing us to closely survey the dynamics of genetic mutations in a population of circulating zoonotic viruses in the short term and possibly predict upcoming variants is therefore pivotal to improve the scientific, medical and societal response to future zoonoses.
Molecular barcoding has a long history in yeast genetics[161]. And to date, it has made great progress in investigating EIDs. Concerning RNA viruses, as mentioned in the article, molecular barcodes have been used as a tracer to follow the consequence of individual viruses encountering evolutionary selection forces in vivo , rendering us better understanding viral transmission fitness and evolutionary dynamics. Molecular barcoding on SARS-CoV-2 are focusing on sample multiplexing thus far; application of molecular barcodes on coronaviruses to gain better understanding of cross-species transmission fitness over time will be extremely important. Whereas molecular barcodes in retroviruses (SIV and HIV)[17,25,61,63]have been applied to identify different polymorphisms on the viral gene, study single provirus transcriptomics and track viral rebound in animal models. Collectively, molecular barcodes provide us a good opportunity for virus studies in a quantitative manner.
The key to using molecular barcodes as universal identifiers for either identifying single molecules or tracing individual viruses is to have a very large initial collection of barcodes. Good complexity of barcodes diminish the probability that two independent molecules harbor the same barcodes. Several approaches used to validate the quality of barcoded libraries, such as calculation of a minimum Levenshtein distance[38] or establishment of a threshold for sorting out trustable barcodes[26]have been mentioned previously. In general, barcode duplications are recommended to be kept at a frequency below 0.1%[60]. It is also important to note that imbalance in the representation of the barcodes in the library has been reported; a thorough investigation into a bias of barcode representation will be required in the future. Eventually, the impact of molecular barcodes on viral infectivity and replication should be always taken into account; fitness of barcoded viruses is recommended to be verified for the usage of every new prepared barcoded library.