The main finding of this study was that jackal-originated worms contained thick-shelled eggs in a significantly lower proportion than those originated from foxes, though the mean intensity and mean egg count per worm did not differ in the two host species. Considering that the investigated specimens originated from naturally infected wildlife, our research presents solely a snapshot of the egg production performance of the parasite in different intestinal ecosystems. By our research design, we could not determine whether thin-shelled eggs originated from younger parasites or resulted from an impediment to egg development. For this reason, a significantly higher proportion of less developed eggs in golden jackals might have more explanations.
On the one hand, more thin-shelled eggs might mean a slower ontogeny of E. multilocularis in jackals. By random sampling, those developmental stages can be overrepresented, which need a longer period to pass through. This explanation accords with experimental findings that the parasite's ontogeny in non-vulpine Canidae proved to be slower than in the red fox [4, 6, 8, 15].
On the other hand, it should not be excluded that some worms cannot complete egg development owing to the suboptimal conditions of their habitat. This phenomenon can be observed in feline definitive hosts, in which both the parasite's intensity and egg production capacity are lower than in canines [4, 6]. In our research, the mean intensity and the proportion of fertile eggs did not differ in the two host species (Table 1).
A third possible cause of numerous thin-shelled eggs is multiple infections that resulted in more distinct classes of worms in the hosts' intestinal tract. However, Al Sabi et al. [15] experienced that in multiple infected hosts, most (74%) worms cannot produce eggs at all. In our study, over 80% of the parasite individuals produced eggs, of which over 80% proved to be fully matured in both hosts.
The suitability of a certain definitive host species depends on the microtopography of the small intestine, the composition of bile, biochemical and nutritive factors in the gut's lumen, and the immune response ability of the local lymphatic tissue [20]. Protoscoleces that enter the small intestine are sensitive to bile acids. Although bile is essential for initiation of larval development, glycine conjugated deoxycholate, which is characteristic for herbivorous bile, possesses a strong lytic effect on the tegument of protoscoleces [17].
Deoxycholate is a secondary bile acid, which is transformed from cholic acid by intestinal bacteria. From the intestinal lumen, deoxycholate is absorbed and transported back to the liver [18, 19]. Hepatocytes sense secondary bile acids as endogenous toxins, thus tolerance to deoxycholate means a better tolerance to also exogenic toxins, such as plant secondary metabolites, which were evolutionary developed in herbs against herbivores. Thus, the interdependent co-evolution of plants, consumers, and the intestinal microbiota were shaping a delicate balance throughout the trophic webs [20, 21]. The bile acid composition of different carnivore mammals is very similar, deoxycholate is conjugated with taurine, and a great proportion of bile acids are cholic acid in the form of sodium cholate or taurocholate, which are less erosive for the worms' tegument [17].
Deoxycholate production depends on the host's microbiota, which is determined by the phylogenetic position of the host species and the diet of the individual [22]. Though both red fox and golden jackal are omnivorous, as they consume less than 70% animal tissue in their diet, deoxycholate producing Firmicutes bacteria are less dominant in golden jackal's microbiota [22, 23] than in members of the Vulpes genus [22, 24]. On the other hand, deoxycholate production in Felidae is very low contrary to the high Firmicutes content in their microbiota [22]. This fact suggests finer interactions between the host, its microbiome and bile acids.
Among the biochemical and nutritional requirements of Echinococcus spp., glucose has a central role. Due to its obligate parasitic lifestyle, E. multilocularis can utilise completely digested nutrients. Carbohydrates are exploited in the form of glucose within an anaerobic environment. Studies on Echinococcus spp. adults revealed that lactate production and lactate dehydrogenase pathway of ATP synthesis characterise their metabolism [3, 25]. Based on these research findings, we hypothesised that access to digested carbohydrates should be a limiting factor for the adult parasites in the intestinal ecosystem.
In wild carnivores, enzymatic breakdown of polysaccharides is limited. The evolution of carnivory led on to the contraction in gene families for metabolism of complex carbohydrates. Amylase enzyme coding AMY2B gene copy number decreased as a result of adaptation to predominant flesh consumption and low carbohydrate content of the diet. Due to the loss of AMY2B copies, the amylase enzyme activity is lower in carnivores than in herbivore mammals. Therefore, the oligosaccharide content achievable for brush border disaccharidases is also limited in a carnivore's small intestine [26, 27].
Although most wild canid species possess only one copy of AMY2B, they can adapt to increased starch content of their food contrary to cats that are hypercarnivores depending on animal originated diet [27, 28]. Golden jackals and red foxes are both opportunistic feeders, carnivorous components of their diet are between 50–70% [12, 29]. Lanszki et al. (2018) pointed out a peculiar distinction between red foxes and golden jackals in utilisation of plant materials. Foxes consume three times more fruits than jackals. Wild fruits' carbohydrates contain 20–60% fructose [30]. Fructose content of the chymus partially inhibits the glucose uptake of the enterocytes [31, 32], which provides accessible glucose for Echinococcus adults located between intestinal villi in intimate connection with the brush border and its disaccharidase enzymes [33]. The positive effect of fructose supplemented diet of the host on Taeniid was studied in Hymenolepis spp. The research demonstrated that fructose alone cannot support the growth and egg production of the parasites but improves the positive effect of a starch-rich diet [34].
Besides fructose, wild berries also contain phenolic compounds [30]. These secondary plant metabolites reduce the gene expression level of sugar transporters of the enterocytes, therefore inhibit glucose uptake by the host [35]. This effect increases the glucose concentration of the intestinal fluid, though little is known about the influence of phenolic compounds on the survival of E. multilocularis. Studies on E. granulosus protoscoleces [36] and Raillietina echinobothrida [37] proved that secondary plant metabolites can damage tegument of cestodes.
Considering that carnivores diverged from their plant eating ancestors, their detoxification ability is limited, thus they hardly tolerate high levels of phenolic compounds [28, 38]. Between different carnivore taxa, detoxification capacity of the liver varies, which is well demonstrated by different levels of contraction and pseudogenization in the UDP-glucuronosyltransferase (UGT) gene family. These genes code the major phenol-detoxification enzymes of the liver [28, 39]. To the best of our knowledge, golden jackal's UGT copy number variation has not been investigated yet. Kondo et al. [40] compared evolutionary features of UGT genes in carnivore taxa. They ascertained gene expansion in domestic dogs and fox species but not in wolves and felids. The authors regarded UGT gene expansion as an adaptation to a degree of herbivory.
Scientific literature on intestinal ecosystems of golden jackal and red fox revealed some variations. Diet analysis demonstrated a preference for fruits in foxes [12]. Studies on components of intestinal microbiota proved that golden jackal harbours less Firmicutes bacteria than vulpine carnivores [22, 23]. These data support the hypothesis that in natural conditions, golden jackal's intestinal fluid contains low concentration of potentially toxic compounds, which can harm E. multilocularis. On the other hand, a small proportion of carbohydrate consumption [12, 29] and presumably low amylase activity [26] in golden jackal provides less accessible glucose for the parasite.
The theory of low carbohydrate level was supported by the finding that in golden jackals, crowding caused conspicuous impact on egg maturation. In golden jackal specimens, mature egg producing individuals were obtained from less crowded (almost third in size) infrapopulations than those of fox origin. This finding suggests that in the golden jackal’s intestinal ecosystem, an intense competition takes place for resources.
Comparing our research findings with literature data, we concluded that the golden jackal's intestinal ecosystem cannot provide a huge amount of nutrients for E. multilocularis. Nevertheless, jackal's gut would carry a lower risk of intoxication for the parasite. In the fox's intestinal ecosystem, the parallel presence of higher amounts of accessible glucose and higher risk of toxic compounds may force the rapid maturation of the worms, which results in shorter longevity and extensive egg production for a short time period. This theory agrees with the experiences gained by experimental infection of different carnivore taxa with E. multilocularis. In these experiments, the researcher determined the shortest, though the most extensive, egg production in foxes [4, 5, 15, 41].
Based on these findings, we supposed that the mutual presence of red fox and golden jackal in a habitat can enhance the public health risk of HAE. Foxes can contribute to the egg load of the environment during the period from late spring to early autumn, which is both the main season for wild berry ripening and breeding of the intermediate hosts of E. multilocularis. During this period, foxes consume a great proportion of fruits [12]. Fruit component increases both sugar and phenolic compound concentration of the intestinal fluid, which might result in maturation and egg production with high peaks but in short phases. The ecosystem in the golden jackal host provides a less turbulent environment for the parasites. In these conditions, the parasites grow and mature less rapidly but can count on a longer lifespan. As a result, egg maturation drags but lasts for long, contributing to steady egg shed almost all year round. This hypothetical egg producing characteristic of jackal's E. multilocularis may mitigate the impact of climate warming by sustaining transmission during the initiation and the termination phase of the rodent's breeding season.
Long-lasting carrying of the parasite might ensure the maintenance of the populations between periods of extensive transmission between definitive and intermediate hosts. This might compensate for the short lifespan of the intermediate hosts of E. multilocularis, thus golden jackals might serve as bridge hosts in multi-host ecosystems. However, their bridge effect is not as remarkable as that of ungulate intermediate hosts of E. granulosus [8]. The survival of adult E. multilocularis in golden jackal is to be determined to assess the potential bridge effect of this host species.
Thompson et al. [5] suggest that short life span of E. multilocularis in foxes increases the possibility of reinfection, thus sexual breeding. This phenomenon contributes to the maintenance of genetic diversity in the E. multilocularis populations [5]. Based on another theory, the combination of continuous inbreeding and rare outcrossing guarantees heterosis in a population [42]. The golden jackal, presumably, is an appropriate reservoir host for inbred genetic lines of E. multilocularis, which can produce heterozygous offspring as a result of outcrossing with another inbred line.
In other aspects of suitability, we could not determine dissimilarities between golden jackal and red fox. Both mean intensity and egg production capacity were statistically similar in the two hosts. Moreover, we experienced that crowding increases the proportion of less matured eggs in E. multilocularis. The phenomenon was identified in both hosts. This finding supports the hypothesis that ongoing infection might reduce the possibility of reinfection [5]. Limitation of egg production in a crowded parasite population might be a symptom of competition for sources, especially for carbohydrates. On the other hand, this self-limitation of larger infrapopulations can contribute to maximal genetic diversity of the whole E. multilocularis population in a certain habitat.
Though it is based on solely deduction, we hypothesise that alternation of sexually and asexually reproduced generations of E. multilocularis forces a continual coevolution with a multi-host system to maintain a constant level of genetic diversity, thus virulence. This might result in a pattern, which is observable in the chase Red Queen dynamic where coevolutionary warfare constantly changes. The variation in selective effect of different types of hosts causes oscillation in the gene frequency, thus relative breeding success of different generations [14, 43]. Previous studies in the same carnivore communities suggested that golden jackal might generate a higher risk for HAE than red fox [11, 44]. Regarding the recent findings, we can conclude that common presence of two, dissimilarly adapted, host species in a multi-host system might stabilise endemic focuses of E. multilocularis even in suboptimal environmental conditions.
By investigation of egg production performance of E. multilocularis adults that originated from different hosts, we proved that the parasite produced more thin-shelled eggs and crowding caused more harm to reproduction in golden jackals than in red foxes. Comparing this finding with the results of other studies dealing with different aspects of intestinal ecosystems of golden jackal and red fox, we supposed that golden jackal might increase the breeding success of E. multilocularis in habitats shared with red foxes. This study investigated naturally infected, hunter-harvested specimens, therefore most of our conclusions are based on a speculative approach, thus our research generated more questions than answers. Our deductive reasoning highlighted potential future research directions, such as determination of E. multilocularis adults’ life-span in the golden jackal, the effect of diet components on the survival of adult parasites, the role of herbivore characteristics in host-parasite coevolution, and the dynamics of E. multilocularis reproduction in other multi-host systems.