Table 1.

Comparison of molecular methods to assess the lung microbiota

Method	Advantages	Disadvantages
DGGE or TGGE	Bands can be isolated for taxonomic identification	Technically difficult; results are variable; highly operator dependent
T-RFLP	High-throughput, inexpensive method for pattern identification	Taxonomy should not be inferred without extensive machine standardization
Clone libraries	Full-length 16S rRNA gene; most accurate taxonomy for 16S rRNA-based methods	Costly; labor intensive; very low throughput; highly operator dependent
454-Pyrosequencing	Long reads span multiple variable regions; high-throughput; very good taxonomic classification; inexpensive with DNA barcoding	Expensive unless samples are barcoded; requires extensive post-run analysis
MiSeq	Long enough read length for good taxonomic classification; dual barcoding for extremely high throughput; very inexpensive per run cost; excellent sequencing depth	Sequencing of amplicons uses nonstandard protocol; extensive post-run analysis

Definition of abbreviations: DGGE = denaturing-gradient gel electrophoresis; TGGE = temperature-gradient gel electrophoresis; T-RFLP = terminal restriction fragment length polymorphisms.