are correctly classified, the efficacy of a drug can be tested and developed against all pathogens within the same taxonomical groups, especially if the groups can be characterized by a molecule, pathway, or trait targeted by the drug. Furthermore, when two pathogens are found to be different in species level, despite the similar phenotypic traits, possibility of different treatments should be considered carefully. [Berman, 2012] 2.5 Culture Dependent WGS Figure 2.6. Culture dependent approach vs direct sequencing approach Several sequencing approaches for routine diagnosis have been proposed, such as culture dependent and culture independent WGS, illustrated in Figure 2.6. In culture dependent WGS, microbiologists cultivate the microbes in patient samples, wait until the microbial colonies emerge, purify the DNA, and proceed to sequencing. Producing sufficient amount of pure organisms in a colony requires days for rapid grower bacteria, or even months for slow grower ones [Köser et al., 2012b]. Figure 2.7 shows the complexity and the timescale of cultivating a bacterial pathogen. Once the whole DNA of the selected colony has been sequenced, computational analysis tools are used to identify the taxonomy of the species. This is done by aligning theirsequences to some defined marker genes, either single loci or multi locus [Larsen et al., 2012]. Once the species is detected, the presence of antimicrobial resistance and virulence factors can also be detected to find out the antibiotic for the patient and to examine the disease progression. Micro dilution and disk diffusion are common for antibiotic susceptibility testing and they take one extra day, or even longer for some bacteria [Köser et al., 2012a]. However, the database of of resistance gene and virulence factor sequences are still incomplete, therefore conventional susceptibility testing is still required in complement to the molecular prediction. Furthermore, WGS also promises a definitive resolution for outbreak investigation, therefore it is essential to identify the strain. Figure 2.7. Workflow of culture-dependent investigation of bacteria according to [Didelot et al., 2012] Compared to traditional phenotypic analysis, the WGS based microbial identification provides more rapid, unambiguous, detailed genotype information of the disease-causing pathogens, and it is very helpful to find targeted treatment for the patient and to prevent the outbreak at earlier stage. The pathogen surveillance
are correctly classified, the efficacy of a drug can be tested and developed against all pathogens within the same taxonomical groups, especially if the groups can be characterized by a molecule, pathway, or trait targeted by the drug. Furthermore, when two pathogens are found to be different in species level, despite the similar phenotypic traits, possibility of different treatments should be considered carefully. [Berman, 2012] 2.5 Culture Dependent WGS Figure 2.6. Culture dependent approach vs direct sequencing approach Several sequencing approaches for routine diagnosis have been proposed, such as culture dependent and culture independent WGS, illustrated in Figure 2.6. In culture dependent WGS, microbiologists cultivate the microbes in patient samples, wait until the microbial colonies emerge, purify the DNA, and proceed to sequencing. Producing sufficient amount of pure organisms in a colony requires days for rapid grower bacteria, or even months for slow grower ones [Köser et al., 2012b]. Figure 2.7 shows the complexity and the timescale of cultivating a bacterial pathogen. Once the whole DNA of the selected colony has been sequenced, computational analysis tools are used to identify the taxonomy of the species. This is done by aligning theirsequences to some defined marker genes, either single loci or multi locus [Larsen et al., 2012]. Once the species is detected, the presence of antimicrobial resistance and virulence factors can also be detected to find out the antibiotic for the patient and to examine the disease progression. Micro dilution and disk diffusion are common for antibiotic susceptibility testing and they take one extra day, or even longer for some bacteria [Köser et al., 2012a]. However, the database of of resistance gene and virulence factor sequences are still incomplete, therefore conventional susceptibility testing is still required in complement to the molecular prediction. Furthermore, WGS also promises a definitive resolution for outbreak investigation, therefore it is essential to identify the strain. Figure 2.7. Workflow of culture-dependent investigation of bacteria according to [Didelot et al., 2012] Compared to traditional phenotypic analysis, the WGS based microbial identification provides more rapid, unambiguous, detailed genotype information of the disease-causing pathogens, and it is very helpful to find targeted treatment for the patient and to prevent the outbreak at earlier stage. The pathogen surveillance
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