Review of Cryptosporidium and Giardia in the eastern part of Europe, 2016

Introduction This paper reviews the current knowledge and understanding of Cryptosporidium spp. and Giardia spp. in humans, animals and the environment in 10 countries in the eastern part of Europe: Bosnia and Herzegovina, Croatia, Czech Republic, Estonia, Hungary, Latvia, Poland, Romania, Serbia and Slovenia. Methods: Published scientific papers and conference proceedings from the international and local literature, official national health service reports, national databases and doctoral theses in local languages were reviewed to provide an extensive overview on the epidemiology, diagnostics and research on these pathogens, as well as analyse knowledge gaps and areas for further research. Results: Cryptosporidium spp. and Giardia spp. were found to be common in eastern Europe, but the results from different countries are difficult to compare because of variations in reporting practices and detection methodologies used. Conclusion: Upgrading and making the diagnosis/detection procedures more uniform is recommended throughout the region. Public health authorities should actively work towards increasing reporting and standardising reporting practices as these prerequisites for the reported data to be valid and therefore necessary for appropriate control plans.


Introduction
Cryptosporidium spp. and Giardia spp. have been ranked as the sixth and 11th most important foodborne parasites globally, respectively [1]. Both parasites are shed in the faeces of infected hosts and can infect new hosts via faecal-contaminated soil, water, feed and food [2]. Several Cryptosporidium species are clearly zoonotic, including C. parvum, while human giardiasis is caused by two genetically different groups of G. intestinalis, referred to as assemblages A and B, which can infect other mammalian hosts and thus have a zoonotic potential [3]. Control of pathogens that can be transmitted among humans, animals and the environment is best achieved with the One Health approach.
Among food-borne diseases, cryptosporidiosis and giardiasis cause a considerable burden at the global level [4], but the burden at regional and national levels is largely unknown [1,5]. Moreover, the current estimates of the burden caused by zoonotic pathogens only include a part of the potential impacts and true costs. In a One Health context, the estimates of disease burden would address that in humans and that in animals, including reduced human and animal health, economic losses, environmental contamination and the impact on biodiversity.
The most common clinical presentation of human cryptosporidiosis is profuse watery diarrhoea with abdominal pain, low-grade fever, nausea, vomiting and weight loss. It is often asymptomatic, mild or selflimiting in immunocompetent individuals and serious, even fatal in immunosuppressed individuals, such as HIV-infected persons [6,7]. A Cryptosporidium spp. infection can also be fatal in several mammalian animals and chronic in reptiles [8].
The clinical features of acute giardiasis in humans are similar to cryptosporidiosis, and include severe diarrhoea, abdominal cramps, nausea and weight loss. These symptoms may persist for a few weeks or evolve   [11]. c Regional data, does not represent the whole country. Croatia: data from Istria region; Slovenia: data from various regions; Bosnia and Herzegovina: data from Canton of Sarajevo; Serbia: data from region of Nis.
into a chronic reoccurring disease. The infection may be asymptomatic or a subclinical course [9]. Giardia spp. infection in cattle, goats and sheep can cause nutrient malabsorption that can consequently result in a reduction of weight gain. Although mortality due to giardiasis is uncommon, fatal giardiasis has been reported in chinchillas and birds [10].
Microscopic examination of stool specimens remains the cornerstone of diagnostic testing for these parasites, although molecular methods and immunological assays can effectively replace microscopic approaches. Microscopy is cheap, but requires a skilled parasitologist and the diagnostic yield is dependent on proper stool collection. The treatment options for both include antiparasitic drugs and fluid therapy.  [11]. These countries make up 20% of the EU population [12]. Considering that Cryptosporidium spp. and Giardia spp. are transmitted via similar pathways, and that one fourth of all giardiasis cases notified in the EU were from these 10 countries, the low proportion of cryptosporidiosis cases suggests underreporting. In general, relatively little is known about the presence of Cryptosporidium spp. and Giardia spp. in the eastern part of Europe despite their public health relevance. This review aimed to assess the significance of Cryptosporidium spp. and Giardia spp. infections in humans and animals, as well as their occurrence in the environment based on (locally) available data. While the data are challenging to compare, they provide an overall picture of the situation and main knowledge gaps.

Methods
For the purpose of this analysis, we considered the following 19 (Figure).
The contacted experts from each country gathered data from sources including national official health service reports, national databases, and international and national publications. These experts also conducted a PubMed (Medline) literature search between April and October 2016 to identify internationally published data while Google databases, using defined qualifiers for G iardia, Cryptosporidium and geographic location (e.g. Hungary), were used to identify data from grey literature. In addition, searches in local databases identified doctoral theses, journals and other publications available in the main local languages (Bosnian, Croatian, Czech, Estonian, Hungarian, Latvian, Polish, Romanian, Serbian, Slovenian) in the participating countries. Data on epidemiology, diagnostics and research of the two parasites in humans, in animals, and in the environment were extracted.
Based on the extracted data, the 95% confidence intervals (CI) of prevalence and the two-tailed p values of two-by-two table comparisons were calculated using the mid-P exact method with the OpenEpi v.3.01 programme [13]. If detailed data were not given, we report the count, percentage and CI as presented in the original publication. Data in this paper are presented on a country-by-country basis in alphabethical order.

Croatia Humans
There has been an obligation for clinicians to report both parasites since 2012. However, the only data available for this review for the years 2006 to 2015 were those obtained from the Department of Microbiology, Public Health Institute of the Istrian Region, which serves an area of ca 200,000 inhabitants. Of the stool samples examined for Giardia cysts, 245,321 came from the obligatory occupational health checks of healthy people working with food and beverages while 17,183 were sent by clinicians for diagnostic purposes. Routine methods are used, including merthiolateiodine-formaldehyde concentration (MIFC), to concentrate protozoa and worm eggs from faecal samples. For patients with bloody diarrhoea, Ziehl-Neelsen staining and microscopy of bloody stool are used to determine the presence of Cryptosporidium spp. The presence of G. intestinalis and Cryptosporidium spp. in human stool samples in the region of Istria is presented in Table 1.

Czech Republic Humans
In the years 1975 to 1982, a total of 1,750 immunocompetent persons, mostly employed by agricultural enterprises, were examined for the presence of gastrointestinal parasites [18]. Of these, none were positive for Cryptosporidiumspp., but 0.80% (14/1,750; 95% CI: 0.48-1.38) were positive for G. intestinalis using Breza's, MIFC and Army Medical Service III concentration techniques and direct microscopy [19,20]. The first human cryptosporidiosis case in the Czech Republic was recorded by Ditrich et al. in an immunodeficient patient in 1991 [21]. The authors identified the Cryptosporidium isolate from that case as C. baileyi, but the identification was made without molecular analysis and therefore cannot be considered accurate. Based on the data from National Reference  In a study mapping the occurrence of various diarrhoeal pathogens in children hospitalised with diarrhoea between 1992 and 1996, 11.32% (12/106; 95% CI: 6.28-18.45) were positive for Cryptosporidium based on aniline-methyl-violet staining of stool smears [22,23]. Nine of 106 Cryptosporidium-positive samples originated from immunocompetent children 5  [29].
A few studies have reported on the presence of Giardia in dogs, domestic animals and wild ungulates (Table 3). Unfortunately, all studies were based on microscopic examination of samples using native preparation, flotation in Sheather's sugar or Breza's solution, or staining methods. As no genotyping tools were used and information on genetic assemblages is lacking.

Estonia Humans
Cryptosporidiosis and giardiasis are notifiable diseases in Estonia. From 1991 until 2016, a total of 134 cases of cryptosporidiosis have been reported by the Health Board, of which only a few have been in the recent years (Table 1) [59,60]. For 1991 to 1992, the official Estonian reports mention 33 cryptosporidiosis cases (personal communication, J Epštein, February 2014). During the same years, stool samples of patients with intestinal diseases (n = 1,518) were examined at one hospital using an unspecified microscopy method and Cryptosporidium oocysts were found in 3.34% (49/1,469; 95% CI: 2.51-4.35) of the stools from patients with acute intestinal disease who were 0-14 years of age [61]. Since 1999, reports on cryptosporidiosis have originated from two of 15 counties, Harjumaa and Raplamaa, and since 2010, all the individuals diagnosed with cryptosporidiosis were children [59]. The official data thus do not appear to include known outbreaks occurring among veterinary students [62]. One such case was caused by the C. parvumsubtype IIaA16G1R1, and there was evidence of the infection having originated from calf faeces [62].
According to the number of cases reported to ECDC from 2007 to 2016, Estonia has the second highest rate of laboratory-confirmed giardiasis cases with a   (Table 1), which is three times higher than the EU mean for the same time period [11]. In particular, the reported incidence rate among children 0-4 years of age in Estonia between 2007 to 2016 (152.22 individuals per 100,000 inhabitants) was 10 times higher than the rate in all reporting countries (15.45 individuals per 100,000 inhabitants) [11]. In a national health report of the Health Board, 46.84% (549/1,172; 95% CI: 44.00-49.71) of individuals with reported giardiasis in 2010 to 2014 were children less than 5 years of age [59]. The same report reported that 5.12-17.51% of all patients with giardiasis were hospitalised and that 70.20-80.71% of the annually reported cases in 2010 to 2014 originated from one county, Harjumaa, where the capital Tallinn is located [59].

Table 5b
Prevalence of Cryptosporidium spp. and Giardia spp. in domestic animals including pets and wild animals using different methods Poland, 1997-2014 oocysts at the time of the study. C. parvum and C. andersoni have been described in cattle less than 12 months of age [64]. The prevalence of shedding Cryptosporidium spp. oocysts was higher (p < 0.001) in animals older than 12 months of age compared with younger animals. However, evaluated with a semiquantitative scale, the younger animals appeared to shed in higher numbers [64]. Management practices that appeared to increase the magnitude of oocysts shedding included early removal of a calf from its mother [65]. Cryptosporidium spp. oocysts were detected with IFT in ovine faeces collected from 60.87% (56/92; 95% CI: 50.63-70.43) of sheep herds on the islands of Hiiumaa, Vormsi and Saaremaa [66].

Hungary Humans
Stool samples for both Cryptosporidium spp. oocysts and Giardia spp. cysts are routinely tested at the Department of Parasitology, National Center for Epidemiology and Regional Parasitological Laboratories in Budapest, Hungary using microscopic examination of the wet mount (saline and iodine) preparation, MIFC technique for concentration of the protozoan cysts, ELISA/immunochromatographic test (ICT) antigen detection and/or Kinyoun staining. The data are shown in Table 1.
Based on an epidemiological survey, the seroprevalence for a positive response to the 27-kDa Cryptosporidium antigen was significantly higher in communities where the drinking water originated from surface water than in the control city where riverbank filtration was used (p < 0.001). A logistic regression analysis of risk factors showed that bathing in outdoor pools was also associated with a positive response to the 15/17-kDa Cryptosporidium antigen complex (p = 0.0197) [67].
The association between the consumption of Giardiapositive drinking water and asymptomatic giardiasis was investigated in 2007. Despite this being a field investigation where only a single stool sample was examined from each participant, G. intestinalis infections were found in 4.00% (4/100; 95% CI: 1.28-9.36) of asymptomatic individuals. In both water samples and asymptomatic persons, G. intestinalis assemblage B was detected [68]. . All sequenced SSU rRNA samples belonged to dog-specific assemblages C and D. Although canine giardiasis is highly prevalent in the studied geographical areas, it did not present zoonotic potential and the infection rate declined with increasing age of the dogs [71].

Environment
The presence of Cryptosporidium oocysts and Giardia cysts in different water sources (surface water, wastewater, raw water and drinking water) was investigated during the period 2000 to 2007 by microscopy using Method 1623 of the United States Environmental Protection Agency (US EPA). Up to three Cryptosporidium oocysts/100 L and up to 63.6 Giardia cysts/100 L were detected in drinking water [72]. The highest concentration in raw water was 50 Cryptosporidium oocysts/100 L and  [75]. All positive samples were from children up to 4 years of age, and isolates belonged to species C. parvum and C. hominis. In 2008, Bajer et al. reported Cryptosporidium infectio ns in persons with immunodeficiencies; C. hominis, C. meleagridis and C. parvum were found in children with primary immunodeficiencies (PID), but only C. parvum was found in children and adults with a secondary immunosuppression (i.e. after cancer treatment) [76].
A 2010 study including 232 people from the west-central region of Poland found G. intestinalis in 1.29% (3/232; 95% CI: 0.33-3.48) of the collected faecal samples by direct microscopy. Three subgenotypes of Giardia were detected: a cosmopolitan subgenotype AII and two new subgenotypes A and B [77]. Examination of the faeces of 31,504 children 7 years of age from 15 Polish provinces in 2002 to 2003 found G. intestinalis in the faeces of 0.69% (217/31,504; 95% CI: 0.60-0.78) of the children using direct microscopy and Lugol's iodine staining method [78]. In another study from 2008 to 2009, of 120 children with watery diarrhoea resembling a parasite infection, 12.50% (15/120; 95% CI: 7.44-19.35) tested positive for Giardia antigens in the faeces using an immunochromatographic test [79].

Animals
Several prevalence studies have been performed on animals in Poland for both parasites using a wide range of detection techniques. The results are summarised in Table 5.

Environment
Cryptosporidium spp. contamination of tap water has been confirmed by microscopy, IFA and PCR in one of twelve examined samples from the city of Poznan [80].

Examination of surface waters
The presence of G. intestinalis assemblages A and B, and Cryptosporidium oocysts has been found in 45 [81]. The Vistula River (n = 21) and the Zegrzyński Lake (n = 8) were tested for the presence of Cryptosporidium oocysts and Giardia cysts using a Filta-Max filtration capsules and xpress automatic station (IDEXX Laboratories, Inc., Westbrook, US) for filter elution, immunomagnetic separation (IMS) and IFT [82]. Giardia cysts were found in all samples from the Zegrzynski Lake (range: 10-45/100 L) and in all samples from the Vistula River (range: 10-389/100 L). Cryptosporidium oocysts were present in 50.00% (4/8; 95% CI: 18.41-81.59) of samples from the Zegrzyński Lake and in 47.62% (10/21; 95% CI: 27.29-68.57) of samples from the Vistula River. Their number in both cases was similar and ranged from 5 to 25 oocyst/100 L. Cryptosporidium oocysts were also detected in 50 of 68 surface water samples collected monthly from intakes (n = 13) and recreational waters (n = 4) in the Krakow area during June to September 2012. Giardia cysts were only detected in samples taken from three sampling locations [83].

Using animals as indicators of contamination
Rotifers taken from three lakes located near the city of Poznań were used as an indicator of recreational water contamination [85]. Cryptosporidium oocysts were detected in rotifers and water from the lakes using the fluorescence in situ hybridisation (FISH) method. Mussels collected from Poznań's municipal reservoir, Lake Malta, have been examined by direct microscopy (wet smear and smears stained with Ziehl-Neelsen and iron haematoxylin) and MERIFLUOR IFT Cryptosporidium/Giardia kit (Meridian Bioscience Inc., Cincinnati, US) [86]. Cryptosporidium oocysts were detected in 15.38% (12/78; 95% CI: 8.61-24.69) of the mussels.

Contamination of food products
Fresh vegetables and soft fruit have been investigated using IMS and molecular methods [87]. Cryptosporidiumoocysts were found on 6 of 128 vegetables, and C. parvum was identified by subtyping (gp60) from celery. The authors speculated that the presence of Cryptosporidium on vegetables could be associated with products originating from regions with considerable livestock production [87].

Romania Humans
Between 2008 and 2012, a total of 16 Cryptosporidium spp. infections were reported by the Romanian National Public Health Institute (Table 1). In a study using ELISA, a Cryptosporidium prevalence of 4.04% (17/421; 95% CI: 2.45-6.26) was reported from western Romania [88]. Molecular characterisation of five isolates indicated the presence of species C. parvum (n = 3) and C. ubiquitum (n = 2) [88]. Vieira et al. has also reported the presence of the C. parvum subtype IIdA22G1 in faecal samples of four children under 12 years of age from Timiş County in this area of Romania [89].

Animals
Over the last decade, epidemiological surveys were carried out with the aim of finding Cryptosporidium oocysts and Giardia cysts in livestock, pets and wildlife stool samples. Research focusing on livestock is limited and mostly involves the western [89,[92][93][94], central and north-western [95] regions of the country. The methods applied included non-molecular (conventional acid-fast staining and classical microscopic examination, coproantigen detection immunoassays) and molecular tools (PCR-restriction fragment length polymorphism (RFLP), DNA sequencing). Results are summarised in Table 6.

Environment
Investigations on the occurrence of Cryptosporidium spp. oocysts and Giardia spp. cysts in the main rivers of western Romania using the US EPA's Method 1623 showed their presence in 7.54% (4/53; 95% CI: 2.44-17.21) and 41.50% (22/53; 95% CI: 28.87-55.06) of raw surface water samples, respectively. Genetic characterisation of the isolates demonstrated the presence of domestic/wild canid origin C. canis (n = 1) and the human/animal origin C. parvumIIaA16G1R1 subtype (n = 1), as well as G. intestinalis assemblages AII (n = 12) and E, the ruminant origin assemblage (n = 1) [96]. In another study, conducted in the same region, 27.27% (3/11; 95% CI: 7.45-57.81) of the tested wastewater samples were positive for the zoonotic C. parvum, with IIaA15G2R1 (n = 2) and IIdA18G1 subtypes. Also, the occurrence of Giardia spp. were recorded in different surface water types with a detection rate of 90.91% (10/11 53-57.59) in ponds. The registered and successfully sequenced G. intestinalis assemblages were: assemblage E (n = 12) in all tested water bodies, assemblage AII (n = 9) in all tested water bodies except for ponds, and the domestic/wild canid specific assemblage D in a pond [97].

Serbia Humans
In Serbia, giardiasis is a notifiable disease, while cryptosporidiosis is not. Not only that cryptosporidiosis is not reportable, it has also seldom been the subject of research. The only description of cryptosporidiosis in immunocompetent individuals is a report of a family outbreak in 2010 [98]. Conversely, a long-term analysis in immunocompromised individuals carried out between 1985 and 2008 found cryptosporidiosis in 10.50% (50/476; 95% CI: 7.98-13.50) of HIV-infected patients with gastrointestinal symptoms. This finding placed cryptosporidiosis as the second most common cause of gastrointestinal disorders, following oesophageal candidiasis, among all opportunistic diseases in this patient category [99].
On the other hand, giardiasis apparently occurs much more frequently. From 2005 to 2014, a total of 1,996 cases of giardiasis (Table 1) were reported by the Institute of Public Health of Serbia [100]. However, the number of examinations carried out, the clinical reasons for testing and the methods used in particular laboratories are not reported. Analysis of the reports showed that the number of reported cases of giardiasis decreased from 4.6 per 100,000 inhabitants in 2005 to 1.1 per 100,000 inhabitants in 2014. There was no difference (p = 0.255) in the distribution of cases between females and males (48.5% of cases were female and 51.5% were male). Infections were most often diagnosed in people aged 20-40 (45.6%), while 11.9% of all cases were reported in children up to 10 years of age. Giardiasis occurrence was associated with seasonality (p < 0.0001), with one third of the cases being diagnosed between August and October. The incidence peak coincided with increased outdoor activities and increased water consumption during hot weather periods. Giardiasis is widespread throughout Serbia, but the data seem to indicate that it is more common in northern than in central Serbia (10-year mean of 4.5 cases/100,000 inhabitants vs 2.1 cases/100,000 inhabitants). Whether the observed fluctuations reflect a real change in the infection dynamics or are merely the result of differences in the detection of cases or reporting of these remains to be explored. Official reports do not differentiate between cases and do not describe whether reported cases were symptomatic or accidental findings of possibly asymptomatic individuals, for example, during routine examinations of cooks, bakers, restaurant staff, etc. for obligatory occupational health checks. Regional investigations conducted by the Department for Parasitology at the Public Health Centre of Niš (southern Serbia) between 2004 and 2008, did report the number of investigations making it possible to estimate the prevalence of Giardia, which was 0.28% (Table 1). Miladinovic-Tasic and colleagues carried out several studies on giardiasis in different populations; the results from ones that examined healthy adults as a part of obligatory occupational health checks showed a decrease in the prevalence of giardiasis from 0.43% (64/14,833; 95% CI: 0.33-0.55) in 2002 to 0.16% (53/32,814; 95% CI: 0.12-0.21) in 2008 [101][102][103]. High infection rates were registered in establishments where people were in close contact, such as individuals in psychiatric institutions (6/100; 6.00%, 95% CI: 2.47-12.06) [101], specialised institutions for children with disabilities (7/106; 6.60%, 95% CI: 2.93-12.62) [101] and refugee camps (7/122; 5.74%, 95% CI: 2.54-11.02) [102]. In patients with diarrhoea, the prevalence of giardiasis was as high as 10% in adults and 4% in children under 14 years of age [101,102]. The prevalence of giardiasis has also been studied in schoolchildren. Nikolić et al conducted an extensive long-term study throughout central Serbia between 1985 and 2005 that involved a total of 6,645 asymptomatic children 7-11 years of age, representing approximately 10% of the total agematched population (n = 69,232) [104]. The methods used included microscopy after conventional concentration techniques. Despite this being a field investigation where only a single stool sample was examined from each participant, the results showed the presence of Giardia infection in all examined regions, with infection rates ranging from 3.2 to 14.2%, and an overall prevalence of 6.10% (405/6,645; 95% CI: 5.54-6.69). This is significantly higher than the figures in the official reports. Interestingly, the prevalence of Giardia was similar in urban (7.0%) and rural (6.5%) areas. Another study had previously shown a similarly high prevalence of 8.00% (14/175; 95% CI: 4. 63-12.76) in the highly urban area of the city of Belgrade [105]. Finally, a study carried out in 2004 in south-western Serbia estimated a giardiasis prevalence of 5.62% (45/800; 95% CI: 4.18-7.39) in asymptomatic schoolchildren [106].

Animals
Neither Cryptosporidium nor Giardia infections are notifiable in animals in Serbia. However, several studies have investigated such infections in cattle, swine, lambs and goats (Table 7).
In an examination of 160 cattle from the Belgrade area, Cryptosporidium oocysts were detected in 34 analysis showed that the successfully subtyped assemblage A isolates belonged to the sub-assemblage AII while the assemblage B isolates belonged to the sub-assemblage BIV [115].

Animals
According to genotyping studies, the transmission of Cryptosporidium between cattle and humans is of epidemiological relevance in Slovenia. The most common C. parvum subtypes in cattle were also found in humans [116,117]. The Cryptosporidium species and subtypes detected in cattle in Slovenia are presented in Table 8.  86) of goats were found to be Giardia-positive, while no cysts were found in horses and deer. In terms of cattle, only the non-zoonotic assemblage E of G. intestinalis has been found in 36 faecal samples from livestock using a real-time PCR assay [118]. Although the sample size is limited, the results of this study suggest a less important role of livestock in the transmission of Giardiato humans in Slovenia.

Discussion
Analysis of the data obtained from a total of 10 countries showed that both Cryptosporidium spp. and Giardia spp. are commonly found in animals and in the environment when investigated, while giardiasis is more commonly reported in humans than cryptosporidiosis. Based on the number of reported cases in the ECDC Surveillance Atlas of Infectious Diseases, the difference between western Europe and eastern Europe appears more striking for cryptosporidiosis than for giardiasis [11].
Both parasites are prevalent in eastern Europe, but the number of reported cases varies greatly between the investigated countries; the causes of this variation include true differences in exposure and susceptibility, variable provision and access to healthcare systems, and differences in case definition, laboratory diagnosis, recording of cases and reporting. The national health systems of the countries covered here operate differently. Eight countries are members of the EU, and in these, both cryptosporidiosis and giardiasis are notifiable. In Bosnia and Herzegovina, neither disease is notifiable, and in Serbia, only giardiasis is notifiable. The different reporting standards may lead to varied levels of underreporting and varied recognition of the diseases as public health issue. Making a disease mandatorily notifiable is an important step for obtaining accurate data, however, the quality and representativeness of the data obtained depends strongly on which patients are tested and which diagnostic tests are used. In many countries, neither the number of samples investigated nor the methods used for testing are reported. In our opinion, more transparency and uniformity in the collection of surveillance data are needed to further improve its quality. Currently, data available from the ECDC Surveillance Atlas of Infectious Diseases does not allow for reliable inter-country comparisons as demonstrated by the discrepancy in the reported occurrence of both diseases in humans when comparing surveillance data available via in the ECDC Surveillance Atlas of Infectious Diseases with the data provided by the public health laboratories (Table 1). Some countries provided lower or higher notification rates than that reported by public health laboratories. For example, no evidence of human infections of G. intestinalis was recorded for Romania in the ECDC Surveillance Atlas of Infectious Diseases (Table 1). Primary care doctors or physicians frequently treat patients with diarrhoeal disease symptomatically, without testing faecal samples for pathogens.
Another striking observation of our analysis is the discrepancy in the number of human cases between official reports of public health authorities (Table 1) and research-derived data. Although routine investigations and research studies are never directly comparable, the studies indicate more human infections than what is reflected in the routine investigations, therefore suggesting under-reporting throughout eastern Europe. One reason why research studies report more cases than public health authorities may be the ability to use more sophisticated methodology than that available for routine purposes. Under-reporting, which leads to underestimation of the burden of infection, is further anticipated because not all infected individuals exhibit clinical symptoms and some symptomatic persons do not seek medical care.
Data on the occurrence of Cryptosporidium spp. and Giardia spp. in animals in eastern Europe differ broadly in terms of targeted animal species and depth of analysis. This review showed that both Cryptosporidium spp. and Giardia spp. are common parasites of domestic animals, including pets, in eastern Europe, and importantly, genotypes pathogenic to humans, including C. parvum and G. intestinalis assemblage A and B, are prevalent. C.parvum subtype IIaA16G1R1 is a common subtype in the region, found in both cattle and humans in the Czech Republic, Estonia, Hungary, Romania and Slovenia [62,69,89,117,119]. It has also been suggested that birds may be carriers of human pathogenic species and genotypes of Giardia and Cryptosporidium [70].
Reports on presence of Cryptosporidium spp. and Giardia spp. in food were scarce from this region. Waterborne and food-borne outbreaks are clearly important to establish the burden of disease, but it is likely that many smaller outbreaks are currently missed [120,121].
Baseline data as well as improved understanding of the epidemiology, infection sources, reservoirs and transmission of cryptosporidiosis and giardiasis in eastern Europe are needed. Surveillance studies and outbreak investigations using molecular tools at the subtype level are warranted. In addition, consensus and updated methods that are harmonised across countries are required to make the data more comparable. Reducing public health risks from zoonoses and other threats at the human-animal-ecosystem interface must consider the complexity of interactions among humans, animals and the various environments in which they live. This requires communication and collaboration among the sectors responsible for human health, animal health and the environment in a One Health approach. Although the presented results may be important for public health specialists, epidemiologists, drinking and wastewater managers, veterinarians, farmers and the public in general, further addressing the knowledge gaps in a timely manner would greatly contribute to understanding the complex picture of cryptosporidiosis and giardiasis epidemiology and thus set the stage for appropriate future control plans. The article is partly based upon collaboration within the framework of COST Action FA1408 (A European Network for Foodborne Parasites (Euro-FBP)), supported by COST (European Cooperation in Science and Technology) We also acknowledge the kind help of the following individuals: Pál Szakál for editorial work on this manuscript, Age Kärssin for helping to summarise the veterinary diagnostic data from Estonia, Eszter Mezei for retrieving the epidemiological data from Hungary, Antra Bormane and Rita Korotinska from the Latvian Disease and Control Centre for providing the epidemiological data from Latvia.