BoLA-DRB3 gene haplotypes show divergence in native Sudanese 2 cattle from Taurine and Zebu breeds

Abstract

. CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint 6 126 from Europe, Asia and the Americas using polymerase chain reaction-sequence 127 based typing (PCR-SBT; [16][17][18][19][20][21][22][23][24][25][26][27][28][29]) and target next generation sequencing (Target-128 NGS; [30]), the most powerful tools used to identify diversity of BoLA-DRB3 alleles in 129 cattle breeds. Until now, private African BoLA-DRB3 alleles have been reported by 130 authors using indirect techniques, such as PCR-RFLP, followed by cloning and 131 sequencing [31][32][33][34]. These studies focused mainly on screening and analysis of a few 132 animals from some African breeds (e.g., Sanga, Kenana, Butana); however, to the 133 best of our knowledge, no study has used SBT or NGS assays to examine genetic 134 diversity of BoLA-DRB3 in large samples of native African cattle breeds. Previous work 135 showed the presence of a high number of private alleles in native breeds.  interspersed among the various clusters (Fig. 3). 192 As shown in Fig. S1, the native Sudanese cattle breeds have an even gene 193 frequency distribution, with a high number of alleles with low frequency. This was . CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint 194 especially extreme in the Baggara breed. Only two, five and seven alleles appeared 195 with frequencies of > 5% in the Baggarar, Kenana and Butana breeds, respectively. BoLA-DRB3 exon 2 and the antigen-binding site (ABS). As expected, the d N /d S ratio 213 was higher when only the ABS was analyzed (  (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020.  (Table 2a) (Table 2a). (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint genetic differentiation (F ST = 0.0076) among native Sudanese breeds, ranging 259 between 0.007 and 0.009 (Table S3). This average value is higher than those  Table S2). When breeds were grouped in terms of the breed's geographical 263 origin, as was done in the Venn diagram, the genetic variance among breed groups 264 and among populations within groups accounted for 1.18% and 3.71% of the total 265 genetic variance. Table S3  (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint 282 distance matrices. Then, dendrograms were constructed from these distance matrices 283 using UPGMA and NJ algorithms. All trees revealed congruent topologies, which were 284 consistent with the historical and geographical origin of the breeds analyzed. As 285 expected, these trees revealed two main clusters which included the Taurine and 286 Zebuine breeds, respectively (Fig. 5a). It is noteworthy that Sudanese breeds were 287 located in a sub-cluster within the Zebuine cluster, with the two dairy breeds located . CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint 427 described above, but also by processes of natural and artificial selection. In Sudan  (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint 477 different genetic backgrounds and that are raised in different environments and 478 production systems, so further association studies are necessary to determine the 479 effect (resistance or susceptibility) of the alleles present in the native cattle breeds of 480 Sudan against different infectious diseases.

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Conclusions and future prospects 482 To the best of our knowledge, this is the first study to document in detail the genetic

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The results demonstrate that the background variation between two cattle groups, Taurine   490 and Zebu, is primarily due to events of origin, selection, and adaptation, which explains the 491 variations found in the diversity of the BoLA-DRB3 genes, not only between the two major 492 groups but also with the Zebu cattle group, as documented in this study. This variation may 493 explain how these cattle from Sudan are well adapted and resistant to various diseases. We 494 presume that this genetic information provides a basis for better design of suitable breeding 495 schemes. Further studies are highly encouraged to link the newly identified variants and their 496 association with specific common disease(s).

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This study has some limitations. Inclusion of more breeds from Sudan may have given 498 a more comprehensive picture. In addition, the unavailability of data for African cattle against 499 which to make comparisons is a shortcoming to this study.

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. CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint

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Sampled populations and genomic DNA extraction 503 The ODK system was used to record the sampling information; breed name, sex,  (Table S1 and Fig. S3)). Seven milliliters of venous 520 blood were collected in EDTA-containing vacutainer tubes. Genomic DNA was 521 extracted using DNeasy® Blood and Tissue Kit, (Qiagen, Germany), following the 522 manufacturer's instructions.

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. CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint 524 PCR amplification and sequencing 525 Exon 2 of the BoLA-DRB3 was amplified by PCR as described by Takeshima  (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020.   (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. . CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint

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. CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint . CC-BY 4.0 International license made available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is The copyright holder for this preprint this version posted August 7, 2020. ; https://doi.org/10.1101/2020.08.07.241133 doi: bioRxiv preprint