Next-generation sequencing in clinical virology: Discovery of new viruses

Table 1 A comparative evaluation of the different virus discovery approaches showing advantages and disadvantages associated with them

	Classical approaches(Cell culture and infection based)	Nucleic acid sequence-dependent amplification approaches	Nucleic acid sequence-independent amplification approaches	Next-generation sequencers-based metagenomic approaches
Requirement of cell culture systems	Yes, required for virus particle enrichment	Not required	Not required	Not required
Information about the cytopathic effects of the virus	Yes, could be achieved through cell changes	No information could be achieved	No information could be achieved	No information could be achieved
Requirement of special equipments for purification	Yes, Ultracentrifuge/high speed centrifuges, density gradient is required for preparing pure virus	Not necessary, semi pure preparations obtained through low speed centrifuges are suitable	Not necessary, semi pure preparations obtained through low speed centrifuges are suitable	Not necessary, semi pure preparations obtained through low speed centrifuges are suitable
Information about detailed morphological/structural features of the virus	Yes, could be achieved through Electron/Atomic Force microscopy	No information on virus morphology/structure could be achieved directly	No information on virus morphology/structure could be achieved directly	No information on virus morphology/structure could be achieved directly
Time required for virus identification	Long time is required for identification, ranging from days to weeks	Comparatively faster, days required if cloning and sequencing is involved. Faster with microarray based approaches	Comparatively faster, virus could be identified within few days	Fastest available approach, identification could be done within days and even some times within hours
Requirement of prior knowledge about the virus	Not required	Some information is required regarding genus/family to design primers/probes	Being sequence independent technique, no information is required	Being sequence independent technique, no information is required
Dynamic detection range	Very narrow	Narrow	Wide	Extremely wide
Tolerance to non-viral materials	Vulnerable to other pathogens capable of infecting cell	Being sequence dependent, less vulnerable to other sequences from host and other pathogens	Being sequence in-dependent, more vulnerable to other sequences from host and other pathogens. Virus enrichment techniques required before analysis	Being sequence in-dependent, more vulnerable to other sequences from host and other pathogens. Virus enrichment techniques required before analysis
Suitability for discovery of new viruses	Yes	Less suitable, good at discovery of genotypes/variants of known viruses	Yes	Yes
Suitability during outbreaks	Not suitable due to requirement of long time	Not suitable due to requirement of prior sequence information	Yes, but still considerable time is required during outbreaks	Being fast, very much suitable in detecting pathogens in an outbreak scenario