Epidemiological Markers

Epidemiological markers

The specific protein, polysaccharide antigens, genes in case of microorganisms, and certain threshold value of chemicals or physical factors in case of chemical and physical agents of disease, which are used for typing of microorganisms or health problems are known as epidemiological markers. These marker proteins, polysaccharides, genes are used to identify genera, species, strains and types of microorganisms. Serology, phage typing, biotyping, antibiogram typing are classical typing schemes. The newer methods that focus on the bacterial genome are gold standard. The purpose of epidemiological type identification of species, in a clinical context, is to aid the identification of species which arise from a sourceand to determine the source of the infection either in groups of patients or in individuals. The typing is generally done in the following clinical situations:

1. The clear cut outbreak in which many patients are ill and is attributable to common source.
2. Sporadic infection sin specialized hospital wards.
3. Monitoring of patients in high-risk unit so as to be able to recognize clusters of potentially pathogenic strains.

The typing methods have following characteristics:

1. Discrimination- The typing system makes an accurate distinction between different strain populations of a species.
2. Reproducibility- A typing method should give reproducible results in both the laboratory and clinical context. The stability of a method may be influenced by variation in culture media, incubation time and temperature, inoculum size, structural integrity of the microbial markers.
3. Typability- A typing scheme should allocate the majority of strains to a defined group. Complete typabilityis achieved only by biotyping and electrophoretic methods.
4. Simplicity- Typing methods should be simple, economical to perform. Slide agglutination and other simple serological tests are more widely used than complex ELISA assays or ribotyping.

The typing markers used in the following situations:

1. Commo nsource outbreak
2. Patient to patient spread
3. Sporadic infection
4. Routine monitoring of high-risk patients
5. Antibiotic treatment failure
6. Identification of virulent types

The typing markers are of two types:

Phenotypic markers

The phenotypic markers include cell wall specific proteins, polysaccharides. The phenotyping markers are identified by following methods:

1. Serotyping-Serotyping the use of purified immunoglobulins to detect specific antigens. Common targets for serotyping schemes are surface antigens. Serotyping is done by simple agglutination reaction. The following are the antigens used for developing antibodies for serotyping.

Antigens of Gram-negativebacteria

"O" antigen- cell wall lipopolysaccharide antigen

"R" antigen- core region LPS anitgen

"H" antigen- protein antigen of flagella

"K" antigen- capsular polysaccharide antigen

For example,

Salmonella enterica-1 to 67 serotypes based on "O" antigen

Campylobacter jejuni- 55"O" antigens, 56"H" antigens, 80 "K" antigens

Klebsiella pneumoniae- 75 "K" antigens

Yersinia enterocolitica- 34 "O" antigens

Citrobacter freundii- 32 "O" antigens, 75 "H" antigens

Antigens of Gram-positive bacteria

Capsules, M and T proteins- Streptococcus pyogenes

Coagulase- Staphylococcus aureus

Toxins- Clostridium spp.

Streptococcus pneumoniae has 90 capsular antigens.

2. Bacteriophage typing

Bacteriophagesare bacteria-specific viruses. On the lawn of bacteria on the media, bacteriophages are inoculated. If specifics, plaques are observed. The pattern of susceptibility and resistance to phage lysis differ from one or another and can be used to characterize a strain as belonging to one or another phage type or phagovar. For example, Salmonella typhi has 33 distinct phagovars.

3. Bacteriocin typing

Bacteriocins areproteins produced by bacteria which are lethal for other strains of the same species and ocassionally other species. Bacteriocins inhibit DNA synthesis.

For examples,

Colicin

Macroscopic particles

Microcins, bactericidal peptides

Pyocine typing ofPseudomonasis widely used to characterize it.

4. Biotyping

It is a general biological characterization method, mostly commonly used in clinical microbiology laboratories. The microbial morphology, cultural characteristics, biochemical reactions are used to distinguish isolates. Biochemical tests such as acid production from carbohydrates, carbon source assimilation, nitrate reduction, indole production, H₂S production, MR and VP reactions, and enzymes reactions are used. Physiological tests such s growth at upper and lower temperature, p^H tolerance, salt tolerance are used for biotyping.

5. Antibiogram typing

Antibiogram typing is required as a guide to therapy and also used as an epidemiological marker in tracing hospital cross infections. For example, group Astreptococcus pyogenes(β hemolytic) is bacitracin (0.04 units sensitive and cotrimoxazole-resistant. Strains ofStaphylococcus aureusare often differentiated on the basis of susceptipbility or resistance to a spectrum of antibiotics, e.g. MRSA, VISA.

Genotypic markers

The gene structure of the microorganism is a genotypic marker. For example,pfcrtgene K76T mutation is the molecular marker for chloroquine resistance inPlasmodium falciparum. The followings are the methods of genotyping:

1. G+C ratio

The G+C ratio of each microorganism is fixed and it is useful for the identification of microorganisms.

2. Plasmid profiling

Plasmid, an extrachromosomal DNA is different in different microorganism in size, sequence etc. Therefore, agarose gel electrophoresis along with DNA markers help to identify the microorganisms.

3. Restriction endonuclease pattern

Restriction endonuclease can cleave DNA at specific sites producing DNA fragments. These DNA fragment pattern produced is dependent upon the distribution of the specific DNA sequences recognized by the enzyme (restriction site). Closely related plasmid or DNA will produce the same, or very similar "fingerprints", while unrelated plasmids will produce different fingerprints on electrophoresis.

4. Polymerase chain reaction of specific genes

It is used for artificial replication of DNA. The amplification of certain genes of DNA and its electrophoresis help to identify the microorganisms.

5. Ribotyping.

16S rRNA typing is highly specific. the 16S rRNA is isolated and its sequence is analyzed by electrophoresis and then probed to identify it.

6. DNA hybridization

DNA is isolated, treated with restriction endonuclease and electrophoresis is done. The DNA is then transferred to nitro-cellulose filter and hybridized with radiolabeled DNA probes (P³²). Hybridization is the process in which two single strands of nucleic acid come together to form a stable double standard molecule. The radiolabeled probe helps to identify specific DNA.

References

Gordis, L. Epidemiology. third edition. 2004.

Joshi, Banjara. Fundamentals of Epidemiology. Kathmandu: Quality Printing Press, 2007

Park, K. Park's Text Book of social and prevention Medicine. 18th edition. 2008.