Cystic echinococcosis (CE) is an endemic disease in different parts of the world and caused by Echinococcus granulosus sensu lato. CE is responsible of having a significant impact on human health and causes economic losses in endemic countries (Budke et al., 2017). World Health Organization has included CE in the list of neglected zoonotic diseases (NZDs) (Kern et al., 2017).

E. granulosus s.l. life cycle is maintained through dogs (definitive hosts), harboring the adult tapeworm, and a range of livestock species like sheep, cattle, goats, pigs and camels as intermediate hosts. Dogs excrete the infective eggs via faeces, in soil and water (Gemmell, 1990), and both intermediate hosts and humans (accidental hosts), can be infected by intake of parasite eggs by contaminated food and water. Following the larval form, called hydatid cyst, develops in internal organs, especially the liver and lungs (Eckert et al., 2000; McManus et al., 2003; McManus and Yang, 2017).

Epidemiological studies conducted in central Asia so far on CE showed that it is a public health challenge. Fifty-eight percent of the total population (270 million people) are at CE risk, from different countries such as Mongolia, Kyrgyzstan, Tajikistan, Kazakhstan, Uzbekistan, Turkmenistan, Iran, Afghanistan, western China, and Pakistan. The majority of residents of these regions are at risk of developing CE, especially herdsman and farmers (Zhang et al., 2015).

Pakistan is an agricultural dependent country; around 8 million families of rural area are affiliated to agriculture, deriving 35–40% income from livestock production (Pakistan Economic Survey, 2020). The agricultural sector contributes 21% to national gross domestic product (GDP) of which 11.9% is contributed by the livestock sector. Thus, livestock plays a crucial role in GDP and the economic stability of Pakistan. According to latest Pakistan Economic Survey (2019–2020), sheep population was estimated as 31.2 million, goat as 78.2 million, cattle as 49.6 million, and buffalo as 41.2 million, and meat production total was estimated almost 4708 tons. Although, various non-infectious and infectious diseases including CE are main hindrance in growth of livestock sector. The economic loss causes by parasitic diseases was reported 26.5 million/annum (Pakistani Rupees), while CE-associated losses were estimated US$ 276.20/100 in sheep and goats and US$ 165.72/100 buffalo, cattle and camels (Latif et al., 2010).

Mitochondrial DNA (mt-DNA) is widely using of understanding the phylogenetic relationships among closely related species because of its rapid sequence evolution (Brown et al., 1979) and large datasets derived from mitochondrial genomes have the potential for resolving problematic issues in Echinococcus taxonomy (Nakao et al., 2010). Therefore, an increasing number of researchers have focused on the parasite's mt-CO1 and NADH dehydrogenase 1 (nad1) regions as genetic markers (Sharbatkhori et al., 2009; Simsek et al., 2010; Nakao et al., 2010; Khan et al., 2020a). Although, Saarma et al. (2009) concluded that the analysis of nuclear DNA data provides a more reliable means of inferring phylogenetic relationships within Echinococcus than mt-DNA, many researchers commonly prefer mitochondrial markers for molecular characterization of E. granulosus s.l. (Kesik et al., 2021, 2022; Khan et al., 2021d). Finally, using different molecular techniques, investigations have reported E. granulosus as a species complex, showing intra-specific variability worldwide. Thus, E. granulosus s.l. is classified currently into E. granulosus sensu stricto (G1, G3), Echinococcus equinus (G4), Echinococcus ortleppi (G5), Echinococcus canadensis (G6/G7, G8 and G10), and Echinococcus felidis (Vuitton et al., 2020).

During the past decades, extensive studies have identified different E. granulosus s.l. genotypes/species responsible for CE globally. However, between 1980 and 2022 only 15 molecular investigations have been conveyed on Echinococcus spp in Pakistan. E. granulosus s.s. (G1, G3) (Khan et al., 2020a; Muqaddas et al., 2020), E. canadensis (G6/G7) (Khan et al., 2020a; Muqaddas et al., 2020), and E. multilocularis (Khan et al., 2020a) were reported from human isolates. Besides, E. granulosus s.s. (G1, G3) (Latif et al., 2010; Ehsan et al., 2017; Alvi et al., 2020; Mehmood et al., 2020; Khan et al., 2021a, 2021b), E. ortleppi (G5) (Alvi et al., 2020; Khan et al., 2020b), and E. equinus (G4) (Khan et al., 2021b) were found in animals in Pakistan. Molecular characterization of E. granulosus s.l. has great impact on epidemiological findings and control strategies. Lack of veterinary supervision in slaughterhouses and proper disposal of offal, significantly contributes to the parasite life cycle and transmission (Ahmed et al., 2017; Khan et al., 2019; 2020c, 2021c).

The current study has been designed to investigate the genetic diversity and haplotypes of E. granulosus s.l. isolates collected from cattle and buffaloes by using PCR and sequence analysis of mt-CO1 gene.