Despite only 8% of cattle being found in Europe, European breeds dominate current genetic resources. This adversely impacts cattle research in other important global cattle breeds, especially those from Africa for which genomic resources are particularly limited, despite their disproportionate importance to the continent’s economies. To mitigate this issue, we have generated assemblies of African breeds, which have been integrated with genomic data for 294 diverse cattle into a graph genome that incorporates global cattle diversity. We illustrate how this more representative reference assembly contains an extra 116.1 Mb (4.2%) of sequence absent from the current Hereford sequence and consequently inaccessible to current studies. We further demonstrate how using this graph genome increases read mapping rates, reduces allelic biases and improves the agreement of structural variant calling with independent optical mapping data. Consequently, we present an improved, more representative, reference assembly that will improve global cattle research.

Control of tropical theileriosis, caused by the apicomplexan Theileria annulata, depends on the use of a single drug, buparvaquone, the efficacy of which is compromised by the emergence of resistance. The present study was undertaken to improve understanding of the role of mutations conferring buparvaquone resistance in T. annulata, and the effects of selection pressures on their emergence and spread. First, we investigated genetic characteristics of the cytochrome b locus associated with buparvaquone resistance in 10 susceptible and 7 resistant T. annulata isolates. The 129G (GGC) mutation was found in the Q₀₁ binding pocket and 253S (TCT) and 262S (TCA) mutations were identified within the Q₀₂ binding pocket. Next, we examined field isolates and identified cytochrome b mutations 129G (GGC), 253S (TCT) and 262S (TCA) in 21/75 buffalo-derived and 19/119 cattle-derived T. annulata isolates, providing evidence of positive selection pressure. Both hard and soft selective sweeps were identified, with striking differences between isolates. For example, 19 buffalo-derived and 7 cattle-derived isolates contained 129G (GGC) and 253S (TCT) resistance haplotypes at a high frequency, implying the emergence of resistance by a single mutation. Two buffalo-derived and 12 cattle-derived isolates contained equally high frequencies of 129G (GGC), 253S (TCT), 129G (GGC)/253S (TCT) and 262S (TCA) resistance haplotypes, implying the emergence of resistance by pre-existing or recurrent mutations. Phylogenetic analysis further revealed that 9 and 21 unique haplotypes in buffalo and cattle-derived isolates were present in a single lineage, suggesting a single origin. We propose that animal migration between farms is an important factor in the spread of buparvaquone resistance in endemic regions of Pakistan. The overall outcomes will be useful in understanding how drug resistance emerges and spreads, and this information will help design strategies to optimise the use and lifespan of the single most drug use to control tropical theileriosis.

Trypanosomes are parasitic protozoa that cause severe diseases in humans and animals. The most important species of Trypanosmes include Trypanosoma evansi and Trypanosoma brucei gambiense. The most well-known human diseases are sleeping sickness in Africa and Chagas disease in South America. The most identified animal diseases include Nagana in the African tsetse fly belt and Surra in South Asia, North Africa, and the Middle East. Surra is caused by Trypanosoma evansi. Diminazene resistance is an emerging threat caused by T. evansi infecting animals. The underlying mechanism of diminazene resistance is poorly understood. Trypanosoma brucei gambiense causes African sleeping sickness. The development of diminazene resistance in Trypanosoma brucei gambiense is associated with the alterations in the corresponding P2 adenosine transporter-1 (AT-1) gene. In the present study, by extrapolating the findings from Trypanosoma brucei gambiense, we analyzed genetic diversity in the P2 adenosine transporter-1 gene (AT-1) from T. evansi to explore a potential link between the presence of mutations in this locus and diminazene treatment in ruminants. We examined T. evansi-infected blood samples collected from goats, sheep, camels, buffalo, and cattle in seven known endemic regions of the Punjab province of Pakistan. Heterozygosity (H_e) indices indicated a high level of genetic diversity between seven T. evansi field isolates that had resistance-type mutations at codons 178E/S, 239Y/A/E, and 286S/H/I/D/T of the P2 adenosine transporter-1 (AT-1) locus. A low level of genetic diversity was observed in 19 T. evansi field isolates with susceptible-type mutations at codons A178, G181, D239, and N286 of the P2 adenosine transporter-1 (AT-1) locus. Our results on T. evansi warrant further functional studies to explore the relationship between diminazene resistance and the mutations in AT-1.

Fasciola gigantica liver flukes are responsible for over 3 billion US dollars of production loss annually in farmed livestock and cause widespread zoonotic disease. Nevertheless, the understating of the emergence and spread of the trematode species is poor. The multiplicity of F. gigantica infection and its spread is potentially influenced by multiple factors, including the abundance of suitable intermediate hosts, climatic conditions favoring the completion of the parasite’s lifecycle, and translocation of infected animals or free-living parasite stages between regions. Here we describe the development of a ‘tremabiome’ metabarcoding sequencing method to explore the numbers of F. gigantica genotypes per infection and patterns of parasite spread, based on genetic characteristics of the mitochondrial NADH dehydrogenase 1 (mt-ND-1) locus. We collected F. gigantica from three abattoirs in the Punjab and Balochistan provinces of Pakistan, and our results show a high level of genetic diversity in 20 F. gigantica populations derived from small and large ruminants consigned to slaughter in both provinces. This implies that F. gigantica can reproduce in its definitive hosts through meiosis involving cross- and self-breeding, as described in the closely related species, Fasciola hepatica . The genetic diversity between the 20 populations derived from different locations also illustrates the impact of animal movements on gene flow. Our results demonstrate the predominance of single haplotypes, consistent with a single introduction of F. gigantica infection in 85% of the hosts from which the parasite populations were derived. This is consistent with clonal reproduction in the intermediate snail hosts.