Noninvasive Detection of Echinococcus multilocularis Tapeworm in Urban Area, Estonia

To the Editor: Alveolar echinococcosis, which is caused by the fox tapeworm Echinococcus multilocularis, is an emerging disease in Europe that shows a high mortality rate (1). Humans can become infected after ingesting parasite eggs (e.g., through direct contact with dogs and red foxes [Vulpes vulpes] or with their contaminated feces). E. multilocularis tapeworm eggs are extremely resistant and can remain viable in the environment for years (2). Numbers of red foxes have increased in many countries in Europe in recent decades, and the E. multilocularis tapeworm has also expanded its range. This tapeworm has recently been reported in 17 countries in Europe, including Lithuania, Latvia, and Estonia (1). Foxes and associated tapeworms are also increasingly found in urban areas, prompting considerable public health concern (1,3). Foxes began to colonize urban areas in Estonia in 2005, and they have since been reported in 33 of 47 towns nationwide (L. Plumer et al. unpub. data). Because ≈30% of foxes are infected with the E. multilocularis tapeworm in natural habitats in Estonia (4), it is essential to monitor parasite spillover into urban areas, where it could become a serious public health risk. Consequently, there is an acute need for methods that can effectively detect the parasite and thereby help prevent human infection. Although immunologic (2) and genetic methods (5–7) are available for identifying Echinococcus spp. parasites, a sensitive molecular diagnostic method that detects tapeworms and identifies their host species from degraded fecal samples would be useful. The purposes of this study were to develop a sensitive, noninvasive, genetic method to identify the host species by discriminating between feces of red foxes and dogs; detect E. multilocularis tapeworms in feces and distinguish them from the related parasite E. granulosus; and collect carnivore feces in an urban area in Estonia to identify this tapeworm. Fecal samples suspected to be from red foxes were collected during January–March 2012 and January–March 2013 from streets and grassy areas of Tartu, Estonia. Tartu is a relatively small city (area 39 km2) with 98,000 human inhabitants. We surveyed 14 transects, each ≈4 km in length, that included all major districts in the city (Figure). Each transect was searched weekly during the study period (total ≈850 km surveyed). A total of 137 fecal samples were collected and stored at –80°C for >1 week to avoid risk of infection from any Echinococcus spp. eggs present (2) because E. multilocularis (4) and E. granulosus (8,9) tapeworms have been found in Estonia. Samples of ≈250 mg were placed into 2-mL tubes, heated at 65°C for 15 min, and stored at –80°C. The heating and cooling procedure helps to break the parasite egg shells, enabling more efficient DNA