Whole Genome Assembly And Comparative Genomics Analysis In Swamp Buffalo And River Buffalo

There are two sub-species of domesticated water buffalo: swamp buffalo and river buffalo.Swamp buffaloes are mainly distributed in the East and Southeast Asian countries as a working animal for paddy farming for thousands of years.They have powerful body,long service life,and docile disposition.River buffaloes are mainly distributed in the South Asia and partial countries in Europe and America.They are prized for their distinctive,high-quality milk typified by high fat and dry matter content.The distinct characters between swamp buffalo and river buffalo are resulted from long term breeding selection,while it is uncertain which genes contribute.On the other hand,swamp buffalo and river buffalo can produce fertile offspring despite of differences in the body size,habits and distribution.Phylogenetic researches about divergent time and domesticated location are obsessed in the field,which hinders comprehension of evolution in buffalo.With the advance of agricultural machinery,swamp buffaloes are increasingly neglected in agriculture.Hybrid breeding between two sub-species of buffalo becomes important.It is necessary to build reference sequences and combine researches in genomics to solve issues of breeding and evolution.Although several river buffalo genomes are published in NCBI recently,the quality is limited deep researches in swamp and buffalo.We sequenced the whole genome sequences of two sub-species of buffalo,assembled chromosome-level scaffolds genome,and compared genomes to identify variances.Main results of this study are as follows:1.Assembly of reference genomes for swamp buffalo and river buffalo.We sequenced Pacbio RS2 long reads for swamp buffalo(?50X)and river buffalo(?20x),which yielded total length about 2.6 Gb.The N50 of contigs in swamp buffalo and river buffalo was 8.8/3.1 Mb and the length is longer than the assembly of cattle in UMD?3.1.1 and Btau?5.0.1.Combining the data of Bionano and Hi-C,contigs extended to chromosome-level scaffolds with N50 of 117/116 Mb for swamp buffalo/river buffalo,and the number or size of chromosomes were corresponded to the former researches.BUSCO assessments indicated that the genomes were 96.8%/96.0% complete,underscoring the high quality of the genomes and the gene structure predictions.Synteny among swamp buffalo,river buffalo and cattle indicated that the largest chromosome of swamp buffalo was formed by the fusion between chromosome 4 and chromosome 9 of river buffalo,and chromosome 4 of river buffalo was formed by the fusion between chromosome 5 and chromosome 29 of cattle.2.Annotation of reference genomes for swamp buffalo and river buffalo.Using repeat-masked genomes,we identified 19,279/20,202 gene models in the swamp/river buffalo genomes.The average of gene length,CDS length and exon length was 43,778/39,912 bp,1,662/1,408 bp,and 4,581/4,380 bp respectively.More than 90% of genes had function annotation.The percentage of total repeat sequences of genome was 46% in swamp/river buffalo,and the percentage of short interspersed repeated segments(SINE),long interspersed repeated segments(LINE),and long terminal repeats(LTR)was 12%,28% and 5% respectively.The percentage between Bubalus and Bos remained constant.3.Gene family analysis.We compared orthologous and paralogous genes among swamp buffalo,river buffalo and 8 other mammals.There were 850 genes significantly expanded in Bubalus,and they were enriched related to energetic metabolism,transport,and heat stress in GO term.We focused genes in ABCC family and HSP90 family.Expansion of ABCC family influenced ATP function,muscle development and neuro development in buffaloes.The copy number of HSP90 AA and HSP90 AB were expanded in both sub-species of bufdfaloes,which influenced heat acclimation.There were 229 genes significantly expanded specifically in swamp buffalo,and they were enriched related to biosynthesis of polyamines in GO term.S-adenosylmethionine decarboxylase(AMD1)duplicated in swamp buffalo,which influenced muscle development.There were 373 genes significantly expanded specifically in river buffalo,and they were enriched related to ion channel and Eph receptor family.4.Evolution analysis.Ks distribution indicated that swamp buffalo and river buffalo diverged about 1.13 Ma,which is much longer than the human domestication of history and supported independent domestication.Demographic history using PSMC showed that effective population size between swamp buffalo and river buffalo diverged about 1 Ma during Xixiabangma glacial period.In a typical glacial period,sea level declined and land expanded.Terrestrial channel formed between South Asia and Southeast Asian,which would contributed buffalo ancestors migration and species separation.VPS16 and CGREF1 were identified as positive selection genes,which related to disease resistance and muscle development.