Applications of Molecular Epidemiology to Surveillance

Introduction

Molecular epidemiology is the study of genetic and environmental factors, identified at the molecular level, contributing to the etiology, distribution and prevention of disease in human populations. It is the conjunction of molecular biology and epidemiology. The use of molecular biology techniques in epidemiological applications, such as disease surveillance, outbreak investigation and the identification of modes of transmission and risk factors, significantly improves data quality by providing more sensitive and specific measurements [1].

Regarding the contribution of molecular epidemiology to surveillance, i.e. the systematic and ongoing collection, analysis and interpretation of data regarding public health, the continuing development of molecular methods provides novel tools for enhanced surveillance and outbreak investigations of communicable diseases. The selection of the appropriate molecular method depends significantly on the nature of the epidemiological ‘problem’ to be solved, as well as the time and geographical context in which the method is going to be used. For example, in outbreak investigations the method must have the discriminatory power needed to distinguish cases that are epidemiologically unrelated and, ideally, discriminate closely related findings that reveal person-to-person transmission. At the same time, the method must have high reproducibility and be low cost, and be rapid and easy to perform and interpret. When the method is used in the context of epidemiological surveillance, it must yield results consistent over time to allow implementation of efficient communicable disease control measures [5].

Molecular epidemiology has applications in the surveillance of several communicable diseases, such as measles, meningitis and West Nile Virus (WNV) infection.

 

Measles

While measles remains a significant cause of morbidity and mortality among children in developing countries, the number of deaths attributed to measles world-wide decreased by 70% between 2000 and 2011, mainly because of vaccine efficiency [8]. The World Health Organization (WHO) has included measles in the list of infectious diseases to be eliminated by the year 2020. Specifically, in the region of Europe the target date for interrupting the indigenous measles virus transmission has been set as 2015.

In view of the goal of measles elimination, it is very important to assess the circulation of wild-type measles virus [2]. Genetic analysis is necessary for detection of the virus and its different genotypes, and is essential for understanding the epidemiology of the disease and its surveillance.  Laboratory-based surveillance in conjunction with the mandatory notifiable disease surveillance system is fundamental for detecting cases of measles, because laboratory diagnosis is necessary for case confirmation, particularly when measles incidence is low and most cases of rash and fever are actually caused by other infections and syndromes [3]. Furthermore, the findings from genetic analyses help classify a case as endemic or imported, which is valuable information not only for the purposes of surveillance but for case management as well. After sufficient epidemiological information has been collected and the virus genotype of laboratory-confirmed cases is known, the case can be classified as endemic or imported depending on whether the genotype detected belongs to the endemic genotypes or not. The use of genetic analyses in surveillance allows the monitoring of virus genotypes circulating over time in a specific country or region, and allows the verification or not of any interruption in the transmission of endemic measles. Evidence of the absence of endemic genotypes is one of the criteria for verifying measles elimination in a country or region, and this evidence comes exclusively from genotyping [4].

Additionally, data from molecular analyses can help confirm the source of a virus or indicate the potential source of disease in cases where the source is unknown. Furthermore, genetic analyses can help define cases or outbreaks as related or unrelated. This information is valuable for disease control programs, and the contribution of molecular methods is essential in evaluating the efficiency of measles control measures.

 

Meningitis

The management and prevention of the spread of meningitis, which is a very serious communicable disease, requires accurate and rapid diagnosis, achieved by the application of molecular techniques. Bacterial meningitis is responsible for the greatest burden of morbidity, and meningococcus (Neisseria meningitidis), pneumococcus (Streptococcus pneumoniae) and hemophilus influenza type b (Haemophilus influenzae type b) account for more than 75% of all bacterial meningitis cases.

By using molecular techniques, such as the polymerase chain reaction (PCR), detection and typing of the infectious agent can be achieved in less than 2 hours, significantly improving case confirmation and disease surveillance [7].

Several techniques are used for the molecular typing of every meningitis infectious agent. Results such as the sequencing of the gene porA to determine genotypic characteristics, and the detection of the gene gna1870 encoding the lipoprotein that binds human complement factor H (factor H binding protein, fHbp), contribute significantly to the development of vaccines against meningococcal meningitis. Other techniques, such as multilocus sequence typing (MLST) and variable number tandem repeat analysis (VNTR), contribute to the epidemiological monitoring of the evolution of the meningococcus bacterium and outbreak investigation, respectively.

Molecular typing and serotyping of clinical specimens for the detection of the most frequent serotypes of S. pneumoniae, which accounts for a high percentage of meningitis morbidity and mortality in children, significantly improves disease surveillance by providing useful information on the circulating serotypes that are not included in the 7-valent conjugated vaccine.

With regard to the use of molecular techniques for the detection of H. influenza type b, which causes serious invasive disease in children under the age of 5 years, identification of the bacterium and serotyping has contributed greatly to a) estimating the disease rate through the surveillance of confirmed cases, and b) evaluating the vaccine efficiency [7].

 

West Nile Virus infection

WNV infection occurs after a bite from a WNV-infected mosquito. The virus is mainly found in birds, which are the primary hosts; humans and mammals are considered to be dead-end hosts because during viremia the virus titer in their bloodstream is low and insufficient for mosquito infection. The virus causes mild symptoms of influenza-like illness in about 20% of people who have been infected with WNV, while the majority of infected individuals are asymptomatic. More severe signs and symptoms from the central nervous system develop in less than 1% of the infected population.

For the successful restriction of infection transmission, it is necessary to carry out surveillance, which is significantly facilitated by detection of the virus in biological specimens from patients. The molecular tests used for WNV detection vary depending on the clinical phase of the infection. In asymptomatic patients with low viremia, the molecular method used (e.g. during monitoring of infections through active surveillance of people who live or work in a region where WNV is known to be circulating) must be characterized by high sensitivity in order to detect the virus even at low levels. The viral load in biological specimens from patients with suspected WNV infection is hypothesized to be higher than in asymptomatic infected individuals and, consequently, the molecular methods used for diagnosis do not have to be characterized by the same high sensitivity that is required for the methods used for asymptomatic individual screening [6].

WNV strains have been grouped phylogenetically into seven different lineages. Lineage 1 dominates in Europe but in some European areas lineages 1 and 2 overlap. PCR, reverse transcription PCR (RT-PCR), real-time RT-PCR and nucleic acid amplification tests (ΝΑΤ) are some of the molecular methods used for WNV detection, depending on the requirements; for example, in order to distinguish between lineages 1 and 2, a novel real-time quantitative RT-PCR test has been developed [6]. Phylogenetic separation provides important information regarding virus origin and transmission in specific geographical areas. Circulation of the virus in a given area usually precedes the onset of an outbreak by a year or more, and the detection of a specific viral strain in space and time helps the instigation of an efficient WNV infection surveillance system [9].

 

Conclusions

Without doubt, molecular epidemiology not only improves our understanding of disease pathogenesis by providing clues regarding specific mechanisms, molecules and genes that influence the risk of developing a disease [1], but also assists epidemiological disease surveillance by providing reliable data with high sensitivity and specificity. Additionally, with the results of molecular techniques it is possible to identify the origin of a disease, the route of transmission/spread, and to assess the effectiveness of preventative measures.

References
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  4. Rota PABrown KMankertz A, et al. 2011. Global distribution of measles genotypes and measles molecular epidemiology. JID 204[EHF1] :S514-S523.
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  8. World Health Organization. 2013. Weekly Epidemiological Record 88(3):29–36. Available at: http://www.who.int/wer [Accessed 25 September 2013].
  9. Zehender GEbranati EBernini F, et al. 2011. Phylogeography and epidemiological history of West Nile virus genotype 1a in Europe and the Mediterranean basin. Infect Genet Evol 11(3):646-653.

 Lambrou Angeliki, Head of the Laboratory-based Epidemiological Surveillance Office, HCDCP