Cystic echinococcosis (CE), also known as hydatidosis, is a cosmopolitan zoonotic disease caused by the larval stage of the Echinococcus granulosus sensu lato (s.l.) species complex. CE has a worldwide distribution, with a high prevalence in North African countries, especially Tunisia (1–3). This disease is prevalent among humans and animals, particularly in rural areas where close interactions between livestock and dogs facilitate its transmission cycle. Considering its considerably high morbidity rate, CE is among the priority neglected diseases of the World Health Organization which included it in its strategic roadmap for the year 2022 (4). The annual economic loss due to this disease is estimated at approximately USD 3 billion worldwide (5). In addition to its public health impact, CE imposes a significant economic burden on livestock production, including the condemnation of infected viscera, reductions in meat and milk quality and production, and decreased fecundity (6, 7).

The life-cycle of the parasite requires two hosts: canids as definitive hosts and a wide range of herbivorous or omnivorous animals as intermediate hosts. The adult forms are small taeniid worms that develop in the midgut of canids, especially dogs, which release eggs in their feces. After ingestion by an intermediate host, the viable oncosphere transforms into a CE cyst (larval stage). The most commonly involved organs are the liver and lungs but many other anatomical sites may also be affected (8). The importance of traditional livestock farming is responsible for CE transmission due to the frequent close interactions between production animals and dogs. Human infection is caused by accidental ingestion of eggs through contaminated vegetables, water, soil and fomites or by direct contact with infected dogs (9).

Five species are currently recognized as responsible for CE: E. granulosus sensu stricto (s.s) (G1 and G3 genotypes), E. equinus (G4 genotype), E. ortleppi (G5 genotype), E. canadensis (G6–G10 genotypes) and E. felidis (10). E. granulosus s.s. (especially the G1 genotype) is the most common species associated with CE in both animals and humans worldwide, particularly in Tunisia (3, 11).

With a mean annual surgical incidence (ASI) of CE of 12.7 per 100,000 inhabitants and an annual economic loss estimated at USD 10–19 million (12), Tunisia is one of the most endemic countries in the Mediterranean area (13). On the basis of the findings of the latest national survey on CE, the 24 governorates (Tunisian administrative units) were classified as: hyperendemic (ASI > 19), holoendemic (12.7 < ASI < 19), mesoendemic (6.3 < ASI < 12.7) and hypoendemic (ASI < 6.3) regions (14).

While numerous studies have explored human and animal contamination in various parts of the country (3, 15–17), recent data on the prevalence of CE in livestock, especially in the mesoendemic areas of Tunisia, remain scarce.

This study aimed to assess the prevalence of CE in slaughtered animals, analyze the distribution of CE cysts within livestock, explore possible differences related to host species, and identify potential risk factors contributing to CE transmission dynamics which could target control strategies and enhance public health measures.

The prevalence of CE in the slaughtered animals examined in this study highlights significant economic and health challenges in the livestock sector. Specifically, CE-related economic losses in Tunisia have been estimated at US$ 14.7 million, including costs associated with the condemnation of infected organs (US$ 1.76 million), reduced meat and milk production (US$ 16.1 million), decreased fertility in infected animals (US$ 4.02 million), and losses due to reduced livestock productivity in wool (US$ 1.18 million) (12). However, the impact of CE infection varies across host species (ovine, bovine, goat or dromedary). In our study, sheep presented a higher prevalence rate (15.86%) compared to cattle (9.57%), which aligns with the findings of Lahmar et al. (15) in Tunisia, who reported a prevalence of 16.42% (95% CI [15.05–17.87%]) in sheep and 8.56% (95% CI [7.68–9.46%]) in cattle. Comparable differences in CE infection rates have been reported in other Mediterranean countries, such as Italy, Greece, and Algeria, where sheep display higher infection rates than cattle do (23, 24). The elevated prevalence in sheep is largely attributed to their grazing habits in open pastures where they are exposed to E. granulosus s.l. eggs shed by infected dogs. Several studies conducted in Tunisia have highlighted significant environmental contamination with parasite eggs (16, 25, 26). Prolonged exposure increases the likelihood of ingesting parasite eggs, thereby resulting in a higher incidence of CE cyst development. In contrast, cattle, which are typically raised in more controlled environments and fed industrial concentrates, have reduced contact with contaminated environments, explaining their lower infection rate. On the other hand, no cases of CE were observed in dromedaries and goats during this study. These findings are similar to those reported in Tunisia, where CE prevalence in goats ranged from 1.7 to 2.88% and from 0.8 to 5.94% in dromedaries (15, 16). They also align with global studies conducted in Yemen (27), Iran (28), Algeria (29), and China (30). These prevalences are likely attributable to the distinct feeding behaviors of these species as both primarily consume elevated vegetation, such as shrubs and bushes, thereby minimizing their exposure to E. granulosus s.l. eggs. This trend may also be explained by the fact that these animals are often slaughtered at a young age, before the parasite has sufficient time to develop into CE cysts, a process that typically requires several months (31).

A clear correlation was observed between animal age and the CE prevalence, with older animals being significantly more affected. Animals over 4 years of age exhibited the highest infection rates, reaching 83.36% in sheep and 86.42% in cattle. This age-related trend is consistent with the findings of numerous studies, where prolonged exposure to E. granulosus s.l. eggs increases the risk of contamination. Furthermore, an animal’s extended lifespan provides more time for CE cysts to develop and mature within the host (15, 19, 32, 33, 56). In contrast, younger animals, especially those under 2 years of age, show markedly lower prevalence, as they are generally slaughtered before the larvae complete their developmental cycle. Gender also plays a significant role in CE prevalence, with females showing higher infection rates than males. This disparity can be explained by livestock management practices, where females are kept longer for breeding and milk production, unlike males who are typically slaughtered at a younger age, leading to a greater cumulative exposure to the parasite over time.

The liver-lung complex was the most common site of CE cyst localization in both sheep and cattle, followed by CE cysts confined to either the liver or lungs alone. This distribution reflects the known migration pathway of E. granulosus embryos, which first pass through the liver as the primary filter and then through the lungs as the secondary filter (32, 34). The frequent occurrence of hepatic CE cysts is explained by the liver’s proximity to the digestive tract, with larvae migrating through the portal vein as part of their developmental route. However, in some cases, larvae bypass the liver entirely, traveling through the vena cava system to the lungs, where they develop into pulmonary CE cysts (35).

The presence of multiple CE cysts in infected animals is attributed to widespread environmental contamination with E. granulosus eggs, resulting in high levels of exposure (16, 25, 26). Furthermore, a study conducted by M’rad et al. (36), which assessed the genetic diversity of CE cysts using microsatellites, demonstrated successive infection events in host species (ovine and bovine) with E. granulosus eggs. The majority of sheep and cattle harbored between 2 and 10 CE cysts, whereas a smaller percentage of animals carried more than 10 CE cysts. These findings corroborate a study conducted in Sardinia, which demonstrated that most infected animals develop fewer than 10 CE cysts (37). The development of multiple CE cysts may be linked to the host’s immune response, which, despite its efforts to control the infection, allows the parasite to persist through various immune evasion strategies (38). Moreover, the high proportion of aborted CE cysts observed in the present study, especially in older animals, and also reported in Mauritania and Italy (19), indicates a chronic phase of infection. Calcification is part of the immune system’s efforts to contain the parasite, limit CE cyst viability, and reduce the risk of transmission, but does not eliminate the parasite entirely from the host (39).

The fertility of CE cysts plays a crucial role in sustaining the E. granulosus s.l. life cycle, as fertile CE cysts containing viable protoscoleces can develop into adult parasites in definitive hosts. In this study, a high fertility rate was observed in both sheep (98.45%) and cattle (86.81%), with 52.80 ± 24.15% to 67.60 ± 23.63% of the protoscoleces being viable. This high fertility rate was particularly notable in sheep, with no significant difference between sexes, suggesting that both male and female sheep contribute equally to the transmission cycle. In cattle, a striking difference was observed between sexes, with 92.10% of fertile CE cysts found in female cattle, while oxen showed a much lower prevalence of fertile CE cysts. This disparity may be attributed to the longer lifespan of females due to their reproductive role. The prevalence of fertile CE cysts in older animals demonstrates that prolonged longevity correlates with an increased likelihood of developing fertile CE cysts capable of infecting definitive hosts (40). The high fertility and viability rates observed in older breeding females underscore the importance of targeting high-risk animals in control strategies to reduce transmission risks, particularly by ensuring the proper disposal of contaminated offal. Furthermore, the high fertility observed in cattle is characteristic primarily of North African countries (41, 42) and was also noted in the present study. Indeed, previous studies conducted worldwide indicate that E. granulosus s.s. (G1 genotype) cysts, especially in cattle, tend to exhibit low fertility rates (35). National surveillance conducted at French slaughterhouses revealed that CE cysts in cattle are often sterile, with only a minority containing viable protoscoleces (43). This low fertility, with a viability of approximately 5% in some studies, is notably lower than that in other intermediate hosts such as sheep (9). In contrast, in Tunisia, cattle appear to play an active role in maintaining the parasite’s life cycle, posing a risk for dog contamination and, consequently, for human and animal infection. This finding reinforces the importance of targeting cattle in the implementation of control strategies. This observed difference in fertility could be influenced by the regional haplotype of E. granulosus s.s., potentially exhibiting greater compatibility with cattle in North African environments (32, 44–47). For both ovine and bovine hosts, the percentage of fertile CE cysts in the liver was significantly higher (ranging from 72.84% in sheep to 92.10% in cattle) than that in the lungs, which is comparable with the situation in Libya and Iraq (reaching 53.6% in the liver and 14.29% in the lung) (48, 49). However, this variation in fertility between hepatic and pulmonary CE cysts has been shown to differ across studies (50–52). The reasons for these differences remain unclear but may be attributed to several factors, including the developmental stage of the CE cyst, the host’s immune response, and the specific physiological conditions of each organ.

In Tunisia, CE infestation in livestock is due primarily to the G1 genotype of E. granulosus s.s. (16, 42, 44). This is comparable to the epidemiological situation across Mediterranean countries, such as Algeria (29), Italy (19), and Greece (53), emphasizing its significant role in both local and global transmission dynamics. In Tunisian livestock, the high prevalence of this genotype raises concerns not only for its economic impact on animal productivity and veterinary health but also for its implications for public health. Indeed, the G1 genotype of E. granulosus s.s. is, in fact, the primary genotype responsible for human CE cases (54), underlining the zoonotic risk posed by insufficient control of transmission pathways. Phylogeographic analyses, conducted by Kinkar et al. (46), highlight the significant transmission route originating from countries such as Tunisia, which, along with Turkey and Argentina, served as a pathway for G1 genotype dispersion globally. This spread is largely driven by human activities, such as the livestock trade, rather than the natural migration of definitive hosts (46). Factors such as unregulated home slaughter practices, stray dog density, lack of awareness about the CE transmission cycle, and the feeding of raw offal to dogs, facilitate the parasite’s life cycle (2, 24, 25). The World Health Organization recognizes the control and prevention of CE as critical priorities, particularly within a One Health approach, owing to its significant impact on human and animal health, as well as on food supply chains. The practical application of the “One Health” approach involves establishing integrated surveillance networks to monitor CE prevalence in both humans and animals, focusing on identifying high-risk zones, tracking transmission patterns, and evaluating the effectiveness of control measures. Regular community-based deworming programs targeting definitive hosts, such as stray and domestic dogs, are a priority, particularly in high-prevalence areas. These programs should include the provision of cost-effective anthelmintics, community incentives to encourage participation, and collaboration with local authorities and veterinary services to ensure sustainability. Additionally, stricter enforcement of existing slaughterhouse regulations aimed at ensuring the proper disposal of infected offal, particularly from older animals, is crucial for reducing environmental contamination caused by infected dog feces. Public education campaigns must be launched to raise awareness among rural communities about the life cycle of Echinococcus granulosus s.l., the importance of hygiene, and the risks of feeding dogs raw offal. This behavior highlights the significant role that humans play in maintaining the parasite’s life cycle, as feeding dogs raw viscera containing viable CE cysts contributes to environmental contamination with E. granulosus eggs and transmission to livestock. Leveraging local communication channels and involving community leaders can enhance the impact of these campaigns. Furthermore, fostering interdisciplinary collaboration among veterinarians, medical professionals, policymakers, and environmental experts is key to addressing the socioeconomic and ecological factors influencing CE transmission. Potential barriers to implementing the proposed strategies include high densities of stray dogs, which complicate efforts to control definitive hosts through deworming programs. The limited resources for capturing and managing stray dogs exacerbate this issue. Many proposed interventions, such as integrated surveillance systems, regular deworming programs, and enhanced slaughterhouse practices, require significant funding and logistical coordination, which may be challenging in resource-limited settings.

While this study provides valuable insights into the epidemiology of cystic echinococcosis in central Tunisia, several limitations must be acknowledged to contextualize the findings and guide future research efforts. The study is restricted to two central slaughterhouses that process animals from various parts of the country. Although this allows for some extrapolation of the national epidemiological situation, the findings primarily reflect mesoendemic areas in central Tunisia. Expanding the study to include slaughterhouses from other regions with varying endemicity levels would further enhance our knowledge of the spatial distribution of the disease. Another important limitation is that only a subset of the collected cysts was molecularly genotyped, rather than all observed and isolated CE cysts. As a result, we cannot rule out the possibility that additional genotypes were present but not detected in this study. This highlights the importance of expanding genotyping efforts in future studies to further refine our understanding of the genetic diversity of E. granulosus s.l. in Tunisia. Additionally, conducting phylogenetic analyses with another marker, such as the Nad5 gene (55), could improve our understanding of the relationships among the CE cysts sampled in this study and those from different regions. This comparative approach would provide information about the evolutionary dynamics and genetic diversity of the pathogen, offering a more comprehensive view of CE epidemiology in Tunisia and beyond.

In conclusion, this study underscores the significant zoonotic implications of CE in Tunisia, where the G1 genotype of E. granulosus s.s. is prevalent across livestock. The high infection rates observed, particularly in sheep, point to substantial public health risks and ongoing transmission, influenced by local livestock management practices and environmental exposure. The associations between infection rates and factors such as host species, age, and management practices highlight the need for targeted control measures. Addressing CE in Tunisia requires integrated, multisectoral strategies to limit transmission, with a focus on stricter enforcement of existing slaughterhouse regulations, increased public awareness, and targeted deworming programs for definitive hosts, particularly in high-prevalence zones. Further research on the genetic diversity and local adaptations of the G1 genotype will be essential to refine control strategies and reduce the CE burden on both animal health and public health systems. These strategies, combined with enhanced veterinary surveillance, can significantly contribute to breaking the life cycle of the parasite and reducing its burden on both human and animal populations.