Höller, Christiane

Kemper, Nicole

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I Mitteilunaen zur Kieler Polarforschuna Heh 20-2004 Nicole Kemper 1 and Christiane Höller 1 ·2 Institute of Hygiene and Environmental Medicine. University Kiel 2 present address: Bavarian Agency for Health and Food Safety, Ober­schle1ßheim, Germany Semi-domesticated Reindeer in Fennoscandia - a Risk to Human Health? (Workpackage 9.2) lntroduction As one main point of the REN MAN project was the investigation of the environ­ment, the objective of this workpackage was to examine and evaluate the relati­ons between reindeer, the environment and human health regarding microorga­nisms that might cause serious diseases in both, animal and man. Free-ranging animals may serve as sentinels or reservoirs for diseases in livestock and man. Transmission of infectious agents to man may occur through direct contacts to free-ranging animals including cervids (Fanning & Edwards, 1991), by contami­nation of the environment through faecal shedding (Aavitsland & Hofshagen, 1999) or by consumption of venison (Keene et al., 1997). In contrast to dome­stic animals, however, the epidemiological situation in free-ranging animals and in their habitat is difficult to assess. There is a lack of information regarding the human health risk due to faecal shedding of pathogens by reindeer. Bacteria, such as Campylobacter spp., Entero­coccus spp., Escherichia coli, Salmonella spp., Yersinia (Y.) spp. and the parasites Cryptosporidium spp. are among the most important agents in causing zoonosis like enteric and other diseases. They were isolated before from healthy and di­seased domestic ruminants (De Rycke et al., 1986; Munoz et al., 1996; Busato et al., 1998 and 1999, Tham et al., 1999). In E. coli, the ability to cause severe ill­ness in humans and animals is associated to the occurrence of several virulence factors like the production of shigatoxins. Therefore, the presence of shigatoxin genes can be an allusion for the virulence of certain strains, also known as shiga­toxin producing E. coli (STEC). Besides STEC, Campylobacter spp., Salmonella spp. and Yersinia spp. are a pro­blem for meat production with a high infection risk for humans consuming con­taminated products. In addition, these bacteria are known to cause illness espe­cially in young animals. Thus, a certain importance of these pathogens as well in reindeer production may not be excluded. Salmonella spp. (Kuronen et al., 1998) and Yersinia spp. (Rehbinder & Nikander, 1999) have been found associated to mortality in reindeer in Finland and Sweden. Campylobacter spp., that have not been associated to mortality in reindeer yet, may also occur (Kobayashi et al., 1999; Lahti et al., 2001). 39 1 1 40 i ,, .' Mitteilunaen zur Kieler Polarforschuna Heft 20-2004 1 The impact of Cryptosporidium spp. on human health has become more impor­tant during the recent years as waterborne cryptosporidiosis outbreaks cannot be excluded. As free-ranging reindeer can contaminate surface waters by faecal shedding, the occurrence of these pathogens should also be determined. Regar­ding Cryptosporidium spp. in northern European reindeer, no data is available. Ruminants, such as reindeer, may be a reservoir for all these pathogens and more knowledge is required to better protect man, reindeer, wild and farm animals against outbreaks of diseases caused by these infectious organisms. This is of special importance as corralling of reindeer for winter feeding is increasing, eventually raising the prevalence of infectious microorganisms and the inci­dence of i nfectious diseases in rei ndeer. Materialsand Methods During the RENMAN project, 2 243 faeces samples from healthy reindeer, adults and calves, of both genders were examined for the occurrence of Campylobaeter spp., Enterococcus spp., E. coli, Salmonella spp., Yersinia spp., and in addition, for the occurrence of the parasites Cryptosporidium spp .. The samples were taken in the course of one year (June 2001 - April 2002) from Finnish and Norwegian free-ranging and corralled reindeer herds, considering parameters such as degree of intensification of husbandry, location and season. 5amples were taken off the ground or per rectum from slaughter animals, sent to Kiel, Germany, directly after collection and conserved frozen at - 4 °C until further processed . The examination of the occurrence of the different bacteria species was perfor­med according to standard procedures. PCR was used to detect the occurrence of shigatoxin1 and 2 genes (stx1, stx2), the intimin gene (eae) and EHEC-hemoly­sin gene (hlyEHEc) in E. coli. To detect various Yersinia-genes, PCR was performed using primers for the virulence gens 165 rRNA, yadA, V-antigen and 165 rRNA. For the detection of Cryptosporidium oocysts, immunomagnetic separation and immunofluorescence microscopy were applied. Furthermore, 392 soil samples and 50 water samples were examined for these pathogens. For statistical ana­lyses, the data was evaluated with the 5tatistica 5.0 software (5tatSoft GmbH, Hamburg, Germany), following the instructions of Trampisch & Windeler (1997). For all analyses, differences were considered significant at ps0.05. Results In 2 224 (99.2%) out of the total number of 2 243 faecal samples one or more of the examined bacteria species were isolated. Campylobacter sp, identified as C. hyointestinalis, was detected in one sample only (0.04%). Enterococcus spp.were isolated in 2 084 (92.9%) samples. Escherichia coli were isolated in 2 123(94. 7%) samples. There was no evidence of the occurrence of Salmonella spp.nor Cryptosporidium spp. Table 1 shows the results for the detection of the f. colitoxin genes by PCR. One hundred and eight (4,8%) strains of Yersinia spp. wereI Mitteilunqen zur Kieler Polarforschung Heft 20-2004isolated, consisting of Y. enterocolitica Biogroup lA (n=29), Y. intermedia (n=2), Y. kristensenii (n=72), Y. mollaretii (n=3) and Y .  rhodei (n=2).Regarding the degree of intensification of reindeer husbandry, the season or the stxl stx2 /!, 14%)nErol,=2123 0 -eae -11 (0.52%) h[yEHEC eae+hlyEHrc 21 [ 2 (0.99%) {0.09%}geographic origin, no significant differences were found for Enterococcus spp. and E. coli, whereas the prevalences of Yersinia spp. differed significantly: pre­valences for Yersinia spp. in free-ranging reindeer in summer and autumn were significantly higher than in fenced reindeer during winter. Figure 1 illustrates the prevalences of Yersinia spp. and the other isolated bacteria in free-ranging and corralled reindeer. Prevalences of Yersinia spp. in Norwegian reindeer {10.24%) were significantly higher than in Finnish reindeer {3.60%). E. coli was isolated from 14 {3.57%) of the soil samples. Enterococcus spp. was detected in 6 samp­les {1.53.%) and Yersinia spp. in 3 samples (0.77%). Nine {18%) of the water samples were positive for E. coli and 2 {4%) for Enterococcus spp .. 100 -90 � 80 70 : 60 j 50 ,e "° C1> .... a) Sl C1> 30 20 10 :g O o o Campylobacter Enterococcu• •PP spp. Discussion � ..... "' ..... "' C1> ..... E. coli -·-0 .... <D Yerslnla •PP Faecal samples of reindeer were examined for the occurrence of important en­teric pathogens in order to get information about the human and animal health risk. Human infections may be accomplished through direct animal contacts or through contamination of the environment due to faecal shedding in reindeer husbandry areas, or by consumption of contaminated reindeer meat. All bacteria investigated in this study may be found in Northern Europe in the environment in aquatic, terrestrial and animal reservoirs (Kapperud, 1981) and were isolated from the intestinal tract of healthy or diseased ruminants worldwide {Adesiyun Table 1 Occurrence of E. co/i d toxin genes in 2 123 iso/ote stroins (prevalences in paren theses) Figure 1 Preva/ences of ana-lysed pathogens in faeces of free-rangmg (n•1 579} and corralled reindeer ( n·664) 41 1 1 42 Mitteilunaen zur Kieler Polarforschuna Heft 20-2004 1 et al. 1998; Busato et al., 1998). Even though most of the isolated bacteria strains do not have the potential to cause severe human or animal health pro­blems, certain strains might be a risk, especially for immunosupressed, old or very young persons and animals. Therefore, one has to regard the epidemiologi­cal impact of transmission of these infectious agents from the environment to reindeer and man and vice versa. In reindeer, Enterococcus spp. and E. coli occurred in very high prevalences, sho­wing the affiliation of these two species to the normal intestinal flora of healthy reindeer. Concerning f. coli, there are only few reports on diseases caused by shigatoxin-producing bacteria in ruminants (Sherwood, 1985; Mainil, 1999), ho­wever these bacteria are of extreme importance in causing severe diseases in humans (Griffin & Tauxe, 1991). As the genes encoding stx1, eae and hlyEHEc were detected only in very low numbers of the isolated f. coli-strains, the human health risk due to f. coli excreted by reindeer can be considered very low at the moment. These results comply with another study detecting no E. coli 0157: H7 in 1 387 faecal and 421 meat samples from reindeer (Lahti et al., 2001). But this situation might change very quickly, as it is known that STEC virulence factors are mobile within bacterial populations (Yamamoto et al., 1984; Pupo et al.,1997). Yersinia spp. was isolated in 108 samples. The identified species Y. intermedia, Y. kristensenii, Y. mollaretii and Y. rhodei have been isolated before from va­rious environmental samples (fresh water, soil, etc.), food, healthy animals andhealthy and diseased humans (Baier & Puppel, 1981; Sulakvelidze, 2000). Eventhough these species are widely distributed in nature, their actual impact onhuman health is a matter of controversy. But as they are isolated from personsassociated to gastrointestinal disorders, the role of these species should not bedisregarded (Sulakvelidze, 2000). The isolated Y. enterocolitica strains belongedto Biogroup lA, which embraces the nonpathogenic European Y. enterocoliticastrains, often isolated from environmental samples, foods, animal and humanfaeces (Bottone, 1997).Campylobacter hyointestinalis was isolated from one sample only. As the cultiva­tion of Campylobacter spp. is exceedingly difficult, the real prevalence might be higher. Hitherto C. hyointestinalis has been associated only sporadically with hu­man gastrointestinal disorders (Edmonds et al., 1987, Gorciewicz et al., 2002). Even though the prevalence for Campylobacter spp. in this study was very low, it shows that reindeer can be carriers of C. hyointestinalis. This is approved by ano­ther study detecting C. hyointestinalis in a prevalence of 6% in Finnish reindeer faeces (Hänninen et al., 2002). Regarding the soil and water samples, there was no hint that the detected bacteria were reindeer-originated. All positive water samples were taken next to human settlements and human origin is the most probable reason for these positive results. f Mitteilunaen zur Kieler Polarforschuna Heft 20- 2004Summarizing it can be stated that the examined enteropathogens were eit­her not detected at all (Salmonella spp. and Cryptosporidium spp.), in very small numbers (Campylobacter spp.) or if detected, their virulence and pa­thogenicity was very low (E. coli and Yersinia spp.). In the present situation in northern Europe the potential human and animal health risk by reindeer, excreting various important enteropathogenic bacteria and Cryptosporidium spp., has to be estimated as very low. These results are very important, especially regarding the status of reindeer meat as a natural product for the consumer, as for the production no antibiotic treatment is required so far, in contrast to the situation of ordinary farm animals. lt may be expected that free-ranging ruminants develop some of the same production limiting diseases that affect domestic ruminants, if brought into an intensive far­ming environment (Mackintosh, 1998), that may have an impact on human health as well. References Adesiyun AA, Seepersadsingh N, Inder L, Caesar K (1998). Some bacterial enteropathogens in wildlife and racing pigeons from Trinidad. J Wildl Dis 34: 73-80. Aavitsland P, Hofshagen T (1999). Salmonella utbrudd i Her0y kommune. Nytt fra miij0 og samfunnsmedisin 3: 12. Baier R, Puppel H (1981). Enteritis durch „atypische" Yersinien. Dtsch Med Wschr 106: 208-210. Bottone EJ (1997). Yersinia enterocolitica: the charisma continues. Clin Mi­crobiol Rev 10: 257-276. 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Semi-domesticated Reindeer in Fennoscandia - a Risk to Human Health? (Workpackage 9.2)

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