You isolate an infectious substance that is capable of causing disease in plants, but you do not know whether the infectious agent is a bacterium, virus, viroid, or prion. You have four methods at your disposal that you can use to analyze the substance in order to determine the nature of the infectious agent.

I. treating the substance with nucleases that destroy all nucleic acids and then determining whether it is still infectious
II. filtering the substance to remove all elements smaller than what can be easily seen under a light microscope
III. culturing the substance by itself on nutritive medium, away from any plant cells
IV. treating the sample with proteases that digest all proteins and then determining whether it is still infectious

Which treatment could definitively determine whether or not the component is a viroid?
A) I
B) II
C) III
D) IV
E) first II and then III

Analysis of the nature of the specimen like the infectious agents (bacterium, virus, viriod, or prion) is very important to diagnose the diseases caused by them, investigate disease outbreak, to determine the clinical course of the infectious disease etc.

There are many methods like direct detection, cultivation assays, serological assays to detect the pathogen type.

Molecular techniques like polymerase chain reaction (PCR) is done to further analyze or investigate the pathogen, like its microbial genome, the virulence factor etc. These are amplification methods which help to investigate the pathogenesis and disease progression. PCR method involves denaturing the DNA polymerase which is a nuclease type of enzyme. Nucleases are present with exonucleases and endonucleases, both of which acts at specific points on the DNA of the pathogen and will create new templates .