## 2 Do Endophytic Fungi Possess Pathway Genes Encoding Plant Bioactive Metabolites? As previously introduced, endophytic fungi are a polyphyletic group of microorganisms colonizing healthy plant tissues, which developed different symbiotic relationships with their hosts through evolutionary processes [6]. As a consequence of this long-term association implying mutually beneficial relationships, endophytes may play a role in plant metabolism and boost their own biosynthetic activity or may access genetic information to produce bioactive compounds which are analogues to those directly produced by the host plants [5]. However, our knowledge on genetic mechanisms which have eventually allowed endophytic fungi to gain foreign DNA is scanty. One of the hypotheses is horizontal gene transfer (HGT), which is regarded as an important evolutionary mechanism in prokaryotes and is also considered as a possible mechanism for the transmission of genetic material between phylogenetically distant species [37]. Based on evidence of physical clustering of genes for specialized plant metabolic pathways provided by an increasing number of reports [38], HGT is thought to be responsible for the transfer of whole gene clusters from host plants, conferring novel biosynthetic abilities to the associated fungi. According to this hypothesis, many endophytes have developed the capacity to synthesize bioactive products originally known from their hosts, which has triggered the expectation to exploit endophytes as an alternative and sustainable source of important plant metabolites for relieving our dependence on plants. Unfortunately, this expectation remains unfulfilled. One of the main reasons consists in the attenuation or loss of the biosynthetic capacity by the endophytic strains upon subculturing [39; 40]. It has been postulated that it could be consequential to the lack of host stimulation and/or silencing of genes in axenic cultures [41]. However, attempts to reverse the attenuation by adding host tissue extracts to the culturing medium have been unsuccessful [40; 42]. [PERSON] and coworkers considered three alternative hypotheses to explain, firstly, the capacity by endophytic fungi to produce plant secondary metabolites and, secondly, its attenuation in sub-culturing with reference to the observed absence of a key structural gene (here, the strictosidine synthase gene, \(STR\)) in endophytes able to produce terpenoid indole alkaloids [43]. The first hypothesis considers that the function of the missing enzyme might be rendered by another enzyme (encoded by a gene analogue), thus the orthologue gene might be simply absent from the genome of an endophyte. The second hypothesis postulates that endophytic fungi might harbor the missing gene on certain extra-chromosomal elements (ECEs) in the cytoplasm, such as plasmids, which might have been transferred from the host during evolutionary interactions. The third alternative is an extension of the second, saying that the missing gene is included in a plasmid carried by endohyphal bacteria. Like in the former hypothesis, the bacterial plasmid could have acquired the genes through horizontal gene transfer from the host plant. In fact, there are many reports of fungi carrying ECEs, including plasmids [44; 45], and of the presence of endohyphal bacteria in fungi [46; 47; 48]. The two latter hypotheses assume that the attenuation of the secondary metabolite production occurring in cultures results from the steady decay in the number or loss of the endohyphal bacteria or plasmids bearing the genes. In the current post-genomic era, dilemmas concerning the genetic mechanisms enabling production of plant secondary metabolites by endophytic fungi might have a chance to be resolved.