Human pathogenic bacteria causing foodborne or zoonotic diseases are a major healthcare concern even in developed countries [1, 2]. Usage of manure as fertilizer has been discussed as a potential source of infection. Moreover, digestates from anaerobic digesters used as fertilizers were also suspected to transfer human pathogenic bacteria onto vegetables or other crops. The recent outbreak of an enterohemorrhagic Escherichia coli O104:H4 strain in Germany in May 2011 is an example for a foodborne disease having vegetables as source of infection. This outbreak led to the infection of about 3,800 patients suffering from acute gastroenteritis or even the hemolytic-uremic syndrome. Epidemiological and surveillance studies were conducted at the same time by German federal institutions to identify the origin of infection. These studies led to the hypothesis that contaminated vegetables like cucumbers or tomatoes might be involved in spreading of the human pathogenic bacterium [3–5]. Press coverage also hypothesized that digestates from agricultural biogas reactors could have been a source causing these infections. Finally, fenugreek sprouts grown from seeds from Egypt were identified as the most likely source of infection .
However, E. coli is not the only relevant potential foodborne pathogen. Examples for other human pathogenic bacteria causing foodborne infections are Listeria monocytogenes, Yersinia enterocolitica or Salmonella species. Moreover, Campylobacter, Vibrio and Clostridium species are also known human pathogens causing foodborne diseases [1, 6]. Particularly the genus Clostridium, which is well known to accomplish the first steps of anaerobic digestion, is widespread in biogas systems. This genus comprises some important pathogens, such as C. botulinum, C. difficile, C. perfringens and C. tetani. For instance, C. botulinum was recently identified in animal feces [7, 8], a potential substrate for agricultural biogas plants. Hence, agricultural biogas plants are also accused to be involved in the spreading of C. botulinum causing chronic botulism [10, 11].
Human pathogenic bacteria are defined as bacteria causing disease in humans  while the term ‘virulence’ describes their degree of pathogenicity. It has been proposed that human pathogenic bacteria can enhance their virulence by acquisition of genes encoding virulence factors [12–14]. These factors may facilitate adhesion to and invasion of (specific) host cells. Moreover virulence factors can promote survival of the pathogen in the host tissue by inhibiting the immune response and increase the pathogenicity by encoding toxins. Resistance against antibiotics can also be seen as a virulence factor as it complicates medical treatment of a human pathogenic bacterial infection [14, 15]. As an example, for the E. coli O104:H4 strain causing the outbreak in Germany it is supposed that it evolved from an enteroaggregative ancestor by acquisition of the shiga toxin encoding Stx-phage and a plasmid encoding aggregative adherent fimbriae and further virulence features [3, 4].
A major substrate component used for biogas production besides agricultural plant material is manure from animals such as pigs, cattle or chicken. It is known that manure can contain potential human pathogenic bacteria such as Salmonella sp., Listeria sp., Campylobacter sp. or E. coli. Thus, spreading of manure might contribute to (zoonotic) bacterial infections [1, 6, 16–18]. However, several studies on lab-scale and agricultural anaerobic digesters showed that a reduction of the overall pathogen load is possible even at low temperatures [16–18]. Reduction of pathogens was shown to be very efficient for bacteria belonging to the family of Enterobacteriaceae, while it was less efficient for Listeria, Clostridia and Enterococci[16–18].
Several metagenomes of experimental and agricultural anaerobic digesters have been published recently [19–23]. These data provided insights into the microbial community involved in anaerobic digestion and methane production and into the underlying metabolic pathways.
To evaluate the risk associated with utilization of digestates from biogas plants as fertilizer on fields, the existing metagenome sequence data from different biogas reactor communities were for the first time analyzed for the presence of sequence tags originating from putative pathogenic bacteria and those representing virulence or resistance determinants.