Human Parasitic Diseases in Bulgaria in Between 2013-2014

Background: In Bulgaria, more than 20 autochthonous human parasitic infections have been described and some of them are widespread. Over 50 imported protozoan and helminthic infections represent diagnostic and therapeutic challenges and pose epidemiological risks due to the possibility of local transmission. Aims: To establish the distribution of autochthonous and imported parasitic diseases among the population of the country over a 2-year period (2013-2014) and to evaluate their significance in the public health system. Study Design: Cross sectional study. Methods: We used the annual reports by regional health inspectorates and data from the National Reference Laboratory at the National Centre of Infectious and Parasitic Diseases on all individuals infected with parasitic diseases in the country. Prevalence was calculated for parasitic diseases with few or absent clinical manifestations (oligosymptomatic or asymptomatic infections). Incidence per 100.000 was calculated for diseases with an overt clinical picture or those that required hospitalisation and specialised medical interventions (e.g. surgery). Results: During the research period, parasitological studies were conducted on 1441.244 persons, and parasitic infections were diagnosed in 22.039 individuals. Distribution of various parasitic pathogens among the population displayed statistically significant differences in prevalence for some intestinal parasites (enterobiasis 0.81%, giardiasis 0.34% and blastocystosis 0.22%). For certain zoonotic diseases such as cystic echinococcosis (average incidence of 3.99 per 100.000) and trichinellosis (average incidence of 0.8 per 100.000), the incidence exceeds several times the annual incidence recorded in the European Union. Conclusion: Parasitic diseases still pose a substantial problem with social and medical impacts on the residents of our country. Improved efficiency regarding autochthonous and imported parasitic diseases is essential in providing the public health system the tools it needs to combat these diseases. Attention should be focused on the various imported vector-borne parasitic diseases (e.g. malaria and cutaneous leishmaniasis) for which the country is potentially endemic.

According to the World Health Organization (1), over 3 billion people around the world suffer from one or more parasitic diseases, and in most tropical or subtropical countries, they are widespread and a leading cause for morbidity and mortality in the population. In Bulgaria, more than 20 human parasitic infections with autochthonous distribution have been described and some of them are widespread: enterobiasis, hydatid disease, and trichinellosis (2). Growing relationships with tropical and subtropical countries increase the risk of inadvertently importing tropical parasitic diseases. Over 50 imported protozoan and helminthic infections represent diagnostic and therapeutic challenges and pose epidemiological risk due to the possibility of local transmission. More than 2 million Bulgarians travel annually across all regions of the world (3). The geopolitical changes in recent years have led to strong migratory pressure on European countries (including Bulgaria) due to migrants coming from the Middle East and North and sub-Saharan Africa. This mass movement of people increases the risk of the introduction and spread of parasitic pathology that currently has no local distribution. The aim of this study was to conduct a retrospective epidemiological analysis on the prevalence of autochthonous and imported human parasitic diseases in Bulgaria for a period of 2 years (2013-2014) and to evaluate their significance on the public health system in the country.

MATERIALS AND METHODS
The use of these data was consistent with the regulations of the National Centre of Infectious and Parasitic Diseases (NCIPD), Sofia, Bulgaria (Approval number: 1000-107/20.02.17)

Surveillance system
Under current regulations, 11 parasitic diseases are subject to mandatory reporting and registration: ascariasis, cryptosporidiosis, congenital toxoplasmosis, echinococcosis, giardiasis, hymenolepiasis, malaria, taeniasis, trichinellosis, trichuriasis and visceral leishmaniasis (4). Surveillance and control of all parasitic diseases is carried out in accordance with the Ministry of Health (MoH) regulation regarding 'the diagnosis, prevention and control of local parasitic diseases' (5). Data on diagnosed cases are provided to the respective administrative district by the Departments of Medical Parasitology and Epidemiology at the Regional Health Inspectorates (RHI). In the Department of Parasitology and Tropical Medicine at the NCIPD, the information is summarised, analysed and presented to the MoH.

Patients and samples
According to the country's legislation, the following groups of people undergo an annual mandatory check-up for intestinal parasites: children attending nurseries and kindergartens and the personnel working in them, the staff in public food services, persons working in the field of production and trade of food products, and migrants and refugees who arrived in Bulgaria. Our survey includes all Bulgarian and foreign nationals residing in the country who were examined for parasitic diseases from January 2013 to December 2014.

Parasitological diagnosis
In Bulgaria, parasitological diagnosis is carried out in 89 laboratories: 28 laboratories at RHIs and 61 independent medical diagnostic laboratories or laboratories in hospitals based in the 28 districts of the country. Routine and confirmatory diagnostic activities are carried out at the National Reference Laboratory (NRL) at NCIPD. Stool specimens were examined for intestinal helminths and protozoa by various methods: direct wet smear, Lugol's iodine, formalin-ether and other concentration techniques and culture of larval-stage nematodes. For detection of Enterobius vermicularis eggs, the transparent tape test was used. For intestinal protozoa, staining techniques, culture methods and rapid immunochromatographic tests were used. For blood and tissue parasites, the following techniques were used: staining for examination of blood and bone marrow smears, culture methods, detection of specific antibodies by ELISA, Western blot, rapid immunochromatographic tests and molecular analysis by polymerase chain reaction (PCR). Immunological diagnosis and PCR were performed with certified, CE-marked commercial kits (Table 1).

Data collection
We used the annual reports by RHIs on all individuals infected with parasitic diseases in the country and data from the NRL at NCIPD. Data from the National Statistical Institute on the country's population size was also collected and these data were used to calculate parameters such as prevalence and incidence. Prevalence was calculated for parasitic diseases with few or absent clinical manifestations (oligosymptomatic or asymptomatic infections), and incidence per 100.000 was calculated for diseases with overt clinical picture or requiring hospitalisation and highly specialised medical care (e.g. surgery).  Table 2). Diagnosis was confirmed by microscopic methods in 68% of the cases and in 32% by immunological or molecular methods.

Diseases with local transmission
Cystic echinococcosis: A total of 639 cases of hydatid disease were recorded. The relative share of primary cases was 90.85%

Imported parasitic diseases
During the 2-year study period, 8.970 persons were examined, including 611 Bulgarian citizens (

Parasitic infections with local transmission
During the research period, around 10% of the country's population was examined for parasitic infections. Two of these, including cystic echinococcosis (CE) and trichinellosis, have important medical and public health implications. A disturbing fact is the high proportion of echinococcosis among children and adolescents (17.7%, n=113), which indicates a high transmission rate in the country (7-9). The incidence of cystic hydatid disease in the European Union for the reviewed period was 0.18 per 100.000 (10,11), but in Bulgaria the incidence is 30-40 times higher. Possible reasons for the high incidence of human CE in Bulgaria is the large number of free-ranging stray dogs and the breeding of yard and shepherd dogs in some rural areas that are not regularly dewormed which, in turn, leads to a high infection rate among domestic animals (7). In a study of the prevalence of echinococcosis among livestock at slaughter meat production houses, animals that were most affected were the sheep (6.4%), followed by cattle (4.3%), goats (0.9%) and pigs (less than 0.1%) (7,12). After the political and economic changes, human trichinellosis became a re-emerging zoonosis in Bulgaria since 1991. Between 1990 and 2006, 145 trichinellosis outbreaks and 238 sporadic cases were recorded (13). In 2013-2014, 536 cases of human trichinellosis (10,11) were recorded in the EU, of which 130 (24.3%) were registered in Bulgaria. Accordingly, the incidence was 0.05 (2013) and 0.07 (2014) per 100.000 for the EU, and 0.82 and 0.83 per 100.000 for Bulgaria (6). Regarding the Trichinella spp. causing outbreaks in the country, some previous (14) and recent studies (6) confirmed the invariable presence of 2 species, T. britovi and T. spiralis. In Bulgaria, toxocariasis is not a notifiable disease, but according to previous studies of patients with allergy and/or eosinophilia, its seroprevalence ranges between 12.25% and 14.01%, (15)(16)(17)(18) (20). According to some authors, approximately 25%-30% of the world human population is infected by Toxoplasma spp. (21). Actually, the prevalence varies wildly between countries (from 10% to 80%) and often within a given country or between different communities in the same region (22 (27). During this study, most of the autochthonous cases of visceral leishmaniasis were recorded in 2 municipalities (Petrich and Parvomay) located in southern Bulgaria, and 2 cases were imported from Greece and Portugal.

Imported parasitic diseases
Over the course of our study, we observed imported parasitic diseases such as malaria, cutaneous leishmaniasis, urogenital schistosomiasis and loiasis, none of which had local transmission in the country. The rest of the imported cases were intestinal parasitic diseases such as soil-transmitted helminth infections (ascariasis and trichuriasis), communicable parasitic diseases (enterobiasis, hymenolepiasis, giardiasis, and blastocystosis) and visceral leishmaniasis, all of which have local distribution. The gathering of large groups of migrants with such parasitic infections in endemic areas may lead to formation of new foci of transmission or to an increase in the intensity of the pre-existing ones. It seems that the import of parasitic diseases from endemic regions will continue in the coming years. This influx is associated with expanding trade, political and economic relations, tourism and, unfortunately, mass migration of people. The country has favourable climate and fauna conditions for local distribution of a number of parasitic diseases. In 1995 a local outbreak of imported malaria that reported illegal immigrants from Afghanistan as a source was recorded (28). Similar cases have been reported in Greece between 2009 and 2013 with registered cases of tertian malaria with local transmission imported by migrants (29). The local transmission of parasitic diseases directly depends on the effectiveness of the system for surveillance and control.
Regardless of the improvement of living standards and tools for health education, parasitic diseases are a significant part of the overall pathology registered in Bulgaria. It is essential to constantly improve the efficiency of the public health system in order to deal with autochthonous parasitic diseases. It is also imperative to maintain a state of increased preparedness for action against tropical parasitic diseases that may be imported more often into the country in the coming years.