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Phylogenetic inference from molecular sequences is becoming an increasingly popular tool to trace the patterns of pathogen dispersal, especially for rapidly evolving RNA viruses [62], [63], [64], [65].
Therefore, maintaining comprehensive phylogenies, based on representative and up-to-date molecular sequences, is challenging.
Useful phylogenetic information retained in molecular sequences is constantly overwritten by more recent substitution events.
One has to consider whether a concatenation of molecular sequences is sensible or whether different segments should be separately analyzed.
Calibration of demographic histories based on molecular sequences is notoriously difficult, particularly when specific rates of molecular evolution are unavailable [ 57, 58].
To study the interaction between a patient and a pathogen from a population, evolutionary and epidemiological perspective, a huge amount of molecular sequences is needed and current molecular techniques provide, with increasing affordability, this kind of data.
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This apparent lack of attention to MSA methods perhaps stems from an era when the study of molecular sequences was limited to what could be successfully amplified, which likely led to biased analyses of closely related sequences.
Perhaps more importantly, our data suggest that inferences drawn from analyses of widely divergent molecular sequences are to be regarded with skepticism, since the outcome heavily depends on the resulting alignment chosen.
For UCLUST and CROP, only molecular sequences are needed as input.
The molecular sequences were aligned using online version of ClustalW2 software [ 38].
Phylogenetic analyses of molecular sequences are an integral part of many modern molecular and evolutionary biology studies.
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