Bacterial Typing Methods: Aim, Attributes, Types

Human pathogens of one species can compromise very diverse organisms, typing is a tool to identify those organisms up to their strain level. Typing bacteria is an important part of epidemiological studies. Currently, wide range of bacterial typing techniques are in use. These techniques are different from one another in the aspects of study objectives, costs, reliability and discriminatory power.

Aim of typing:

1. Confirmation of epidemiological relationships in the spread of infection.
2. Providing epidemiological hypotheses about epidemiological relationships between bacteria in the absence of epidemiological data.
3. Describing the distribution of bacterial types and identification of affecting factors.

Desirable attributes of typing method

1. Typing techniques should have excellent typeability to be able to type all the isolates studied.
2. In outbreak investigations, a typing method should have the discriminatory power needed to distinguish all epidemiologically unrelated isolates.  Ideally, such a method should discriminate very closely related isolates to reveal person-to-person strain transmission, which is important to develop strategies to prevent further spread.
3. Typing method must be rapid, inexpensive, highly reproducible, and easy to perform and interpret.
4. A typing method that is going to be used in international networks should produce data that are portable and that can be easily accessed via an open-source web-based database or a client-server database connected via the Internet. Additionally, a typing method used for surveillance should rely on an internationally standardized nomenclature, and it should be applicable for a broad range of bacterial species.
5.  There should also be procedures in place to check and validate, by using quantifiable internal and external controls that the typing data are of high quality.

None of the typing methods has all the desired attributes. A thorough understanding of the advantages and limitations of the available typing methods is of crucial importance for selecting the appropriate method. Each typing method has its advantages and limitations that make it useful in some studies and restrictive in others.  Although a particular typing method may have high discriminatory power and good reproducibility, the complexity of the method and interpretation of results, as well as the costs involved in setting up and using the method, maybe beyond the capabilities of the laboratory.

The choice of a typing method, therefore, will depend upon the skill level, and resources of the laboratory and the aim and scope of the study.

Bacterial strains can be differentiated on the basis of their phenotypic or genotyping differences. Genotyping methods show a better performance than phenotypic characterization.

Commonly used typing methods are as follows:

A: Phenotypic Typing Methods: Phenotyping techniques detect characteristics expressed by the microorganism. They are based on biochemical, antigenic or susceptibility (to phages or antimicrobial agents) properties of the organism.

1. Biotyping: Based on metabolic characteristics expressed by an isolate; referred to as ‘biotypes’
2. Serotyping: Based on antigenic determinants expressed by the microorganism; referred to as ‘serotypes’.
3. Phage typing: Based on the pattern of resistance or susceptibility to a standard set of phages; referred to as ‘phage types’.
4. Resistotyping: Based on the resistance or susceptibility of the isolates against a set of arbitrarily chosen chemical agents 
5. Bacteriocin typing: Based on the susceptibility to a set of bacterial peptides (bacteriocin) produced by certain bacteria.
6. Antibiogram typing: Based on the comparison of susceptibility profiles of an isolate to a set of antibiotics.

B: Molecular Typing Methods: Molecular techniques are based on the analysis of chromosomal or extrachromosomal genetic elements (such as plasmid) of the organism.

In recent years a plethora of molecular-typing methods have appeared based on the analysis of fragments of DNA split by specific restriction enzymes. Their discriminatory powers and complexity vary widely. With the advancement of molecular epidemiology, a single machine is now able to generate a wealth of information needed to detect, monitor and control new threats such as drug resistance and the emergence of new pathogens. With the widespread use of molecular typing methods, phenotyping typing methods are now being obsolete.

Commonly used molecular tying methods are as follows:

1. Amplified fragment length polymorphism (AFLP)
2. Enterobacterial repetitive intergenic consensus (ERIC)-PCR
3. Multilocus sequence typing (MLST)
4. Multilocus variable-number tandem repeat analysis (MLVA)
5. Pulsed-field gel electrophoresis (PFGE)
6. PCR Ribotyping (agarose based or sequence-based)
7. Repetitive element PCR typing
8. Restriction endonuclease analysis (REA)
9. Surface layer protein A gene sequence typing (slpAST)
10. Whole-genome sequencing (WGS)

Uses of Typing methods

1. Epidemiologic usefulness:
   1. To investigate the source of different strains in outbreak situations, to check the possibility of laboratory cross-contamination etc.
   2. To determine whether the second episode of disease is due to a previously isolated strain or to a newly infecting strain, and
   3. To determine whether an infection is caused by more than one strain of the organism
2. As an important infection control tool: to monitor the prevalence of certain strains within a healthcare institution or to investigate if a cluster of infections are unrelated or part of an outbreak

References and further reading

1. Van der Merwe, Ruben & Helden, Paul & Warren, R & Sampson, Samantha & Gey van Pittius, Nico. (2014). Phage-based detection of bacterial pathogens. The Analyst. 139. 10.1039/c4an00208c.
2. Sabat A J, Budimir A, Nashev D, Sá-Leão R, van Dijl J M, Laurent F, Grundmann H, Friedrich A W, (2013). Overview of molecular typing methods for outbreak detection and epidemiological surveillance. Euro Surveill.18(4):pii=20380.