Review of the temporal and geographical distribution of measles virus genotypes in the prevaccine and postvaccine eras

Molecular epidemiological investigation of measles outbreaks can document the interruption of endemic measles transmission and is useful for establishing and clarifying epidemiological links between cases in geographically distinct clusters. To determine the distribution of measles virus genotypes in the prevaccine and postvaccine eras, a literature search of biomedical databases, measles surveillance websites and other electronic sources was conducted for English language reports of measles outbreaks or genetic characterization of measles virus isolates. Genotype assignments based on classification systems other than the currently accepted WHO nomenclature were reassigned using the current criteria. This review gives a comprehensive overview of the distribution of MV genotypes in the prevaccine and postvaccine eras and describes the geographically diverse distribution of some measles virus genotypes and the localized distributions of other genotypes.


Introduction
Although measles virus (MV) is serologically monotypic, the genetic characterization of wild-type viruses has identified eight clades (A -H), which have been divided into 22 genotypes and one proposed genotype. Clades B, C, D, G and H each contain multiple genotypes (B1 -3, C1 -2, D1 -10, G1 -3, H1 -2) while clades A, E and F each contain a single genotype (A, E, F) [1,2]. The sequences of the vaccine strains indicate that the wild type viruses from which they were derived were all members of genotype A. All measles genotypes can be neutralized by serum from vaccinated persons in vitro, although with varying efficiency [3,4]. There are no known biological differences between viruses of different genotypes. Specific measles genotypes are not associated with differences in severity of disease, likelihood of developing severe sequela such as subacute sclerosing panencephalitis or inclusion body encephalitis, or variability in sensitivity of laboratory diagnosis.
Analysis of the variability in the nucleotide sequences of wild-type MVs has enabled the use of molecular epidemiologic techniques for measles surveillance. The molecular data, when used in conjunction with standard case reporting and investigation, can help to identify epidemiological links between geographically distinct cases and outbreaks as well as track importations of MV. [5][6][7]. Also, approximately 5% of vaccine recipients experience mild symptoms (rash and fever) after vaccination and these cases could be misclassified as wild-type measles [8].
Genetic characterization of viral isolates or RT-PCR products is the only laboratory test that can differentiate between vaccine-associated cases and wild-type infection [6,9,10].
In 1998, the World Health Organization (WHO) recommended a standard protocol for the designation of measles genotypes. These recommendations, updated in 2001 and 2003, also included a standard analysis protocol and designation of standard reference strains (see Additional file 2) against which all newly characterized isolates should be compared [2,11,12]. The minimum amount of sequence data required to assign a virus to a genotype are the 450 nucleotides encoding the carboxy terminus of the N protein. The entire sequence of the coding region of the H gene should be obtained from representative isolates [11]. New genotypes are designated if the nucleotide sequence differs from the closest reference sequence by more than 2.5% in N and 2.0% in H [2]. Additionally, phylogenetic analysis should produce similar tree topographies using at least two different analysis methods. Several isolates or clinical specimens should be sequenced and at least one viral isolate should be available as the reference strain. Finally, new genotype classifications should be useful for epidemiological studies, by providing a means to identify the source or transmission pathway of infection and by contributing to our understanding of the global distribution of MV genotypes [2].
The purpose of this summary is to collate all available reports of MV genotypes and to standardize the published genotype nomenclature, according to the current WHO criteria, with the aim of giving a comprehensive overview of the distribution of MV genotypes in the prevaccine and postvaccine eras.

Methods
An examination of the National Library of Medicine "PubMed" [13] search engine using the keyword "measles" combined with "genotypes" and "sequence" was performed to identify English language publications or abstracts describing measles genotyping.
Additional sources included the reference lists of articles identified by "PubMed" and electronic sources such as the CDC and PAHO measles network Internet pages and the NCBI Genbank website [14][15][16]. Measles outbreak alerts were received through the WHO network, which distributes outbreak notifications. In addition, subscription based electronic newsletters such as ProMED mail [17] and Immunization newsbrief [18] were scrutinized for information relating to measles outbreaks. Direct contact was made with the notifying laboratory or health unit requesting genotype information if available.
A table produced by participants at the 1998 WHO meeting listed older classification systems and the comparable genotype classifications under the universally accepted system. This table was used to reclassify genotypes cited in publications prior to 1998 [11]. In some cases, later publications from the same or other groups were used to assign current genotypes to viruses classified before 1998.

Results and Discussion
One hundred and twenty eight studies were identified through the PubMed search, 67 of which described the genotype of MV isolates. Four internet websites were identified (including Genbank) and a further 27 articles were identified from the reference lists of cited publications or from outbreak notification lists such as ProMED [17] and Immunization Newsbrief [18].    Table 1 summarize the temporal and geographical distribution of MV genotypes from the early 1950s to 2004 but do not differentiate between cases of endemic or imported measles virus. Genotype and location specific references are not cited in the following results section but can be found in the relevant genotype specific section in the comprehensive table which accompanies this paper (see Additional file 1, also available from the website of the WHO Western Pacific Regional Reference Laboratory for measles, The Victorian Infectious Diseases Reference Laboratory, Melbourne, Australia http://www.vidrl.org.au/labsandunits/measles/ meas_genotyping.htm).
Routine molecular characterization of wild-type measles viruses was initiated in response to a global resurgence of measles disease in the late 1980s and the concurrent availability of sensitive techniques (e.g. RT-PCR and automated sequencing) for the investigation of viral genomes. Prior to that date, only a few isolates of measles were available for molecular characterization and reliable epidemiologic information was not available for many of these isolates. In the era before the widespread use of measles vaccine, genotypes A, C1, and D1 were detected. Genotype A virus includes the prototype Edmonston strain, the progenitor for most of the current measles vaccines. Analysis of MV sequences obtained from SSPE cases, resulting from initial infections that occurred during the 1950s and 1960s, detected genotypes C1, D1, E and F, providing further evidence that genotype A was not the only genotype detected during the prevaccine era [19][20][21][22][23][24]. However, data from these earlier studies must be interpreted cautiously due to the large number of mutations in SSPE sequences and the lack of standardization. Of course, detection of various genotypes in SSPE cases reflects efforts to study this devastating illness and should not be taken as an indication that one genotype is more likely to cause SSPE than another [25]. Retrospective sequence analysis of viral isolates collected during the 1970s showed continued detection of genotypes C1 and D1 and the first detections of genotypes C2, D2, D4, E and F.
As virologic surveillance expanded in the late 1980s and 1990s, the number of genotypes detected in cases and outbreaks increased substantially to include the 23 genotypes now recognized by the WHO. However, some genotypes (B1, D1, E, F, G1) have not been detected in the last 15 years and are considered inactive.  6 Americas -WHO region of the Americas Genotype A has been detected in acute cases of measles in South and North America, China, Japan, Eastern Europe, Finland and the UK over the last 40 years. Since, it is difficult to distinguish wild-type viruses in genotype A from vaccine strains, these reports must be interpreted with caution since some of the sequences may have been derived from vaccine associated cases or been the result of laboratory contamination [22,26,27]. In the future, detection of genotype A viruses in association with acute cases of measles will need to be thoroughly scrutinized and additional sequence data will need to be obtained from both clinical samples and corresponding viral isolates.
Genotype B2, previously considered inactive [12], has recently been detected in South Africa and Angola [28].  [35]. Genotype D8 appears to be co-circulating with genotype D4 on the Indian subcontinent and Ethiopia, although the first known description of this genotype was in the UK, from where it has been regularly detected. However, investigations have linked UK cases with importations of virus not only from the Indian sub-continent but also from the Balkans and Oman [36]. Genotype D8 has been imported into Australia and the USA from India and Bangladesh. Genotype A few genotype E viruses and related SSPE cases were reported in the early 1970s. Genotype F sequences have been identified on two occasions, both were SSPE cases wherein acute measles infection was documented in 1967 and 1968. Thus both genotypes E and F probably circulated in the pre-vaccine era [19,22].
Clade G, previously consisting of one genotype (G), has recently been expanded to contain three genotypes. The original genotype G (now G1) had not been detected since 1983 and was thought to have been extinct. However, recent investigations have identified two new genotypes (G2 & G3), both of which have been predominantly associated with chains of transmission within and importation from Indonesia and Malaysia [38][39][40].
Clade H viruses originally consisted of a single genotype but recently this clade has been expanded to contain two genotypes (H1 & H2). Both genotypes are predominant in the Asian and South East Asian regions. Genotype H1 has mainly been associated with transmission within or importations from China and was detected during the large measles epidemic in Korea in 2000 -2001 [41]. Genotype H1 may now be dominant in Japan [32,42]. The WHO Western Pacific Regional Reference Laboratory for measles recently confirmed circulation of this genotype in Mongolia. Genotype H2, first described from samples recovered from China, has been more recently associated with importations from Vietnam [43].
Some genotypes of MV are associated with a particular geographical region, while other genotypes are more widely distributed. In particular, clade B is predominant in measles transmission in Sub Saharan and Central Africa, clade G in South East Asia and clade H in South East Asia and China. Clade D viruses, on the other hand, appear to be more widely distributed and are endemic in Eastern Africa, parts of Europe and the Indian sub-continent.

Determination of measles genotypes in countries that
have not yet conducted molecular surveillance can be investigated, by proxy, from cases epidemiologically linked to imported cases. For example, the Philippines have not reported an endemic MV genotype but multiple importations to the USA associated with travel to, or contact with, the Philippines have resulted in the supposition that genotype D3 is the predominant circulating genotype in the Philippines [6,44]. However, caution must be taken when identifying genotypes by proxy as the genotype detected may not be the type that is endemic in the region. In some cases, genotypes have been epidemiologically linked to countries with no history of circulation of that genotype. For example, genotype G2 has been reportedly associated with importations to the UK from Mexico, South Africa and Australia, none of which have reported endemic circulation of genotype G2 [36]. In these cases infection may have occurred while the patient was in transit or at venues frequented by other travellers and might not reflect the circulating genotype.
Simultaneous circulation of multiple genotypes has been reported in several regions. Genotypes D3 and D5 co-circulated in Japan since the mid 1980s and the relative number of isolations changed over time. During the late 1980s genotype D3 was detected more frequently, but by 1990 D5 was more common [42,45]. Genotypes D2 and D4 appear to be co-circulating throughout Eastern and Southern Africa. Genotypes C2 and D6 continue to be detected in some parts of Europe and North Africa [35].
Rima et al described a shift from genotype C2 to genotype D6 in Spain in the early 1990s [24]. Santibanez et al recently demonstrated the shift from detection of mostly genotypes C2 and D6 in Germany to detection of mostly genotype D7 [46]. The shift of genotypes occurs in countries that have sub-optimal measles control programs, resulting in interruption of endemic transmission for short periods. However, failure to maintain high levels of population immunity results in the accumulation of susceptible individuals and creates conditions that favour the rapid transmission of a newly introduced genotype. Therefore, the apparent genotype switching is most likely due to changes in the distribution of susceptible individuals in the region.
Nine new MV genotypes have been identified since 1990 reflecting increased surveillance of measles cases and technological advances, rather than recent evolution. The designation of new genotypes, such as the newly proposed genotype d10, is likely to continue as the molecular analysis of viral isolates becomes routinely integrated into more countries within the global WHO measles laboratory network and more sequence data are added to the database. For example, genotype B3 may eventually be reclassified as two separate genotypes since this genotype contains viruses in two distinct clusters [47][48][49]. Characterization of viruses imported into Australia has detected three previously unrecognised genotypes (D7, D9 & G3) due partly to the frequency of travel between South East Asia and Australia and also to the comprehensive measles surveillance conducted by Australian laboratories [7,38,50].
The mutation rate amongst field isolates of MV is low and appears to be random rather than driven by vaccine pressure or immune responses [3,24,26]. Within a genotype, nucleotide differences (virus lineage) can assist in distinguishing separate episodes of transmission [24,51,52]. In countries or regions with endemic (ongoing and constant MV transmission) measles, many lineages of a single genotype may co-exist; however as countries begin to move from endemic to epidemic measles (MV transmission resulting in a higher number of cases than normally expected, typically against a background of little or no MV transmission) [53], the diversity of sequences within the circulating genotypes decreases [43,[54][55][56][57]. In fact, the genotype D6 virus associated with a large measles outbreak that occurred in several South American countries between 1996 and 1997 had identical N gene sequences suggesting rapid spread of a single lineage [51]. Analysis of measles viruses circulating in Burkina Faso, before and after a mass vaccination campaign, showed that the number of circulating lineages was greatly reduced following the campaign. Sequence analysis of viruses isolated from outbreaks that occurred after the vaccination campaign suggested that virus was introduced from a single source [57].
Many recent measles outbreaks have been reported with no accompanying molecular genotyping investigations, for instance in Afghanistan [58], Niger [59] and the Philippines [60]. These outbreaks highlight the need to extend molecular surveillance capabilities to regions where measles remains endemic. Recent studies have described the recovery of MV RNA by RT-PCR from oral fluid, dried blood and dried oral fluid [61][62][63]. These samples, which are easy to collect, prepare and transport by post to laboratories capable of MV genotyping, have the potential to extend molecular surveillance for measles virus to remote settings and countries with limited infrastructure. However conventional samples such as nasopharyngeal swabs, urine and peripheral blood lymphocytes should continue to be collected, if logistically possible, because of the higher sensitivity of these sample types for detecting MV RNA.
Molecular surveillance undertaken in the early stages of measles control can facilitate identification of endemic genotypes. Over time or after intervention programs continued molecular surveillance, in conjunction with case based epidemiological investigations, can detect the interruption of endemic transmission [1]. Additionally, molecular analysis of specimens from cases facilitates both linkage to, and separation from, contemporaneous cases and clusters, assisting classical epidemiological investigations and the tracking of chains of transmission [7,64].