Spread of Measles Virus D4-Hamburg, Europe, 2008–2011

TOC Summary: More than 24,300 cases were identified in 22 countries.

Switzerland, and Belgium and was repeatedly reimported to Germany. The strain was present in Europe for >27 months and led to >25,000 cases in 12 countries. Spread of the virus was prevalently but not exclusively associated with travel by persons in the Roma ethnic group; because this travel extends beyond the borders of any European country, measures to prevent the spread of measles should be implemented by the region as a whole.
T he 53 member states of the World Health Organization (WHO) European Region (EUR) have set a goal to eliminate measles and rubella virus transmission by 2015 in Europe (1). Elimination targets include 95% vaccination coverage with 2 doses of measles virus-containing vaccine (MVCV), an incidence of <1 measles case per million population, 80% of outbreaks associated with <10 cases, and transmission of indigenous or imported measles virus for no longer than 12 months in a defi ned region (2). Thus, monitoring transmission chains of measles virus is an indispensable tool to assess elimination progress, although the specifi c boundaries of the region have not yet been defi ned for the WHO EUR.
Elimination targets have not yet been met in Germany. Vaccination coverage in Germany, routinely assessed in children 5-6 years of age during an examination before school entry, is still below the required 95% for the second dose of MVCV. Recent outbreaks showed an immunization gap in adolescents and young adults (6). Consequently, outbreaks still occur in Germany every year, although recently they have been more limited in number of cases, length of time, and extent of national transmission (7,8). A total of 915 measles cases were reported in 2008, 571 in 2009, and 780 in 2010; incidence was 7-10 cases/1 million population (www3.rki.de/ SurvStat/QueryFormaspx).
The reasons for Germany's malperformance are complex. Measles virus vaccination is not mandatory, and some groups within the German population do not comply with offi cial vaccination recommendations (9) because of philosophical or religious beliefs or fear of adverse effects (10). As in other countries in Europe, strategies to address hard-to-reach populations and improve access to medical care, preventive measures, and vaccination campaigns have not yet been developed. In this article, we describe exportation of a measles D4 variant from Germany and its subsequent circulation in Europe.

Material and Methods
Serum, urine, and oral fl uid or throat swabs were sent to the WHO Regional Reference Laboratory in Berlin according to the procedures outlined in the WHO LabNet manual (11). Immunoglobulin (Ig) M and IgG serologic testing was performed as described previously (12). Sequencing was performed according to WHO recommendations (13). Sequences were aligned by ClustalW (14) and further analyzed by SeqScape 2.5 and MEGA4 DNA analysis software (15). Phylogenetic trees were constructed by using the neighbor-joining method. Genotype assignment was performed by phylogenetic comparison with the measles virus reference strains as designated by WHO (16). The obtained sequence data, the genotype, the offi cial WHO measles virus sequence name, and relevant epidemiologic data were submitted to the WHO measles sequence data base, MeaNS (www.hpabioinformatics.org.uk/Measles/Public/Web_Front/main. php) or to GenBank. For 69% of the reported cases, the diagnosis was laboratory confi rmed. Complications leading to hospitalization were seen in 40% of the patients (pneumonia or otitis media). The affected age group ranged from a 1-day-old newborn to a 54-year-old adult (median age 13.5 years). Most frequently affected were young children, then young adults. Data with respect to vaccination status were available for 196/216 case-patients. No vaccination was documented for 167 persons (85%); 28 had not yet reached the age of vaccination (>11 months). Twenty-six (13%) previously unvaccinated persons had received MVCV after being exposed to measles virus. Three patients had received 2 doses of MVCV. Several of the 216 cases occurred in the Roma ethnic community.

Results
Seventy-two cases of measles were reported during the same time in Lower Saxony. Fifty-three cases were clearly related to the outbreak in Hamburg. The fi rst cases in Lower Saxony were reported during week 2 (January) and the last case occurred during week 17 (April) of 2009; the peak of the outbreak occurred during week 14 (April). The connection to the Hamburg outbreak was suggested either by the presence of patients in the emergency department of a Hamburg hospital at the time in question, an epidemiologic link, or the result of the sequencing. Many cases occurred in the Roma ethnic group.
Case-patients ranged from 7 months to 42 years of age (median age 15 years); adolescents and younger adults were the main affected age group. Forty-two (79%) casepatients had received no measles vaccination, 10 (19%) had received 1 dose of MVCV, and 1 (2%) had been vaccinated 2 times. In the latter case, primary infection with measles virus was confi rmed by PCR and IgM, but IgG was not detected. Five patients received vaccination after exposure, which did not prevent clinical symptoms. Overall, 47 (89%) of measles cases were confi rmed by laboratory testing. Eleven (21%) case-patients were admitted to a hospital with complications (e.g., pneumonia, otitis media).  (20). The WHO Regional Reference Laboratory in Berlin received 20 specimens at regular intervals from hospitalized persons. Genotype information was obtained for 19/20 case-patients (online Appendix Table, (Figure 1).

Laboratory Investigation of Measles Virus Samples from Bulgaria
Measles virus infection was reconfi rmed for all 20 case-patients by positive test results for IgM, PCR, or both (online Appendix Table). Results were correlated with the clinical data for each case-patient that had been compiled during hospitalization. For 12 case-patients, vaccination status was unknown; a 7-month-old baby was unvaccinated. Seven case-patients (1, 7, 12, 14, 15, 17, and 18) presented vaccination cards that stated the date of 1 or 2 vaccinations with MVCV (online Appendix Table). All had positive IgM and PCR results; 2 had measles virus-specifi c IgG (case-patients 7 and 14). IgG avidity testing showed low avidity and thus a vaccination failure for case-patient 7. The equivocal IgM and the mediocre avidity of IgG in patient 14 did not indicate a primary infection. In summary, lack of immunologic response despite documented vaccination was apparent in 6 of 7 case-patients.

Reimportation of the D4-Hamburg Strain to Germany
In 2010, D4-Hamburg measles virus was reimported to Germany. It appeared fi rst in February in Mannheim, where specimens from 3 case-patients showed a sequence identical to D4-Hamburg (MVs/Mannheim. DEU/07.10[D4]). The virus was introduced by 8 persons from Bulgaria who belonged to a Turkish-speaking minority population, had acquired the infection in Dobrich (Bulgaria), and transmitted the virus to 3 relatives who were living in Mannheim. During June-August 2010, 48 measles cases were reported in Munich; 28 cases occurred among Bulgarian Roma residents in a migrant camp in eastern Munich. Several of these residents worked as cleaning staff at hotels in Munich. From these persons and other hospitalized members of the affected Roma group, the virus spread into the general population.
The age of case-patients in Munich ranged from 9 months to 36 years; 7 case-patients were <7 years of age, and 23 were >18 years of age. One case-patient was hospitalized because of encephalitis. Interviews with the help of an interpreter showed that none of the case-patients had MMR vaccination documents. Therefore, vaccination was offered to all inhabitants of the camp. Twenty-eight cases were investigated at the WHO Regional Reference Laboratory in Berlin. Twenty-three cases were associated with MVs/Muenchen.DEU/22.10[D4] identical with

Discussion
A combination of epidemiologic data and genotyping results enabled us to trace the spread of measles virus D4-Hamburg in Europe. It was imported from London at the end of 2008 to northern Germany (288 cases), then transmitted from Hamburg to Bulgaria, where, after a 7-year absence of measles, an outbreak of 24,379 cases occurred. This was the largest outbreak seen in Europe since an outbreak in the Ukraine in 2006 (22).
Twenty cases from the outbreak in Bulgaria were sampled at different times (April and June 2009, January and June 2010, and March 2011) from persons in distinct districts. The samples were collected initially in northeastern and later in southwestern Bulgaria, thereby following the course of the outbreak. The cases were associated with measles virus sequences such as MVs/ Shumen.BGR/15.09[D4], corresponding to D4-Hamburg. The only exception was MVs/Plovdiv.BGR/23.10/6[D4], which showed 1 mismatch but in all probability developed from MVs/Plovdiv.BGR/23.10/1-5[D4]. Because the samples had been obtained at different times and regions, our analysis provides substantial evidence that D4-Hamburg is responsible for the outbreak in Bulgaria, despite the small number of samples. Samples from 6 of 7 persons showed diagnostic markers of a primary measles infection, although these persons had a certifi cate of prior measles vaccination. Our results therefore demonstrate an urgent need to investigate the vaccination procedures for ethnic minorities.
D4-Hamburg was detected subsequently in Poland (54 cases) (21), Ireland, Northern Ireland, Austria (4 cases), Greece (149 cases) (23), Serbia (14 cases), Belgium (>40 cases), and Macedonia (>400 cases). Sporadic cases were detected in Romania, Turkey, and Switzerland. More than 70 D4-Hamburg-associated cases were detected in Germany after 8 separate reimportations. Taken together, D4-Hamburg was present in Europe from December 2008 to March 2011-that is, at least 27 full months-and caused >25,300 cases. Because sequencing results are not available quickly in most countries, this transmission chain is probably still ongoing. Circulation of imported measles virus for no longer than 12 months (and therefore endemic transmission according to the WHO defi nition) is a marker for successful elimination. We suggest, therefore, that the length of a given transmission chain should not be assessed on a national level but at the level of the all 53 countries within the WHO EUR.
Epidemiologic data showed that the spread of D4-Hamburg across Europe involved predominantly persons from the Roma ethnic group in Bulgaria. Another transmission chain affecting the Roma population in particular was recorded in 2004 in Romania. An outbreak of >8,000 cases associated with MVs/Bucharest. ROU/48.04[D4] commenced in the Roma population. Subsequent spread of D4-Bucharest by traveling Roma persons was observed until 2007 (4). The pronounced sequence deviation of D4-Bucharest and D4-Hamburg indicates the presence of at least 2 distinct and successive transmission chains in the Roma population. Both chains were long lasting and associated with a high number of cases, as well as several fatalities. This and other recent outbreaks in Roma communities (7,24,25) underline the need for the development of strategies to address this ethnic minority at the regional level and to improve their integration into the respective national health services.
The lack of strategies to address reaching the hard-toreach communities in Europe will clearly have an adverse effect on the measles elimination process. In this context, we want to make clear that elimination of measles virus should not be seen exclusively as a Roma-associated problem. Measles virus is a highly infectious agent and will infect any population with low immunity rates. If itinerant groups are underserved by the national health sector, spread of measles virus is highly probable. Because measles outbreaks in western European countries occur mainly in undervaccinated groups (26), reaching the hard-to-reach is not the only important challenge. Thus, closing vaccination gaps in a setting of optional vaccination and vaccine skepticism is another important prerequisite that must be met on Europe's path toward elimination of measles virus.