Alterations in the Gut Microbial Composition and Diversity of Tibetan Sheep Infected With Echinococcus granulosus

Hydatidosis/cystic echinococcosis (CE) caused by Echinococcus granulosus is a parasitic zoonotic disease worldwide, threatening animal health and production and public health safety. However, it is still unclear that whether E. granulosus infection can result in the alteration of gut microbiota in Tibetan sheep. Therefore, a study was designed to investigate the influences of E. granulosus infection on gut microbiota of Tibetan sheep. A total of 10 ovine small intestinal contents (five from healthy and five from infected) were obtained and subjected to high-throughput sequencing by MiSeq platform. A total of 2,395,641 sequences and 585 operational taxonomic units (OTUs) were identified. Firmicutes and Proteobacteria were the most dominant phyla in all samples. Moreover, the proportions of Armatimonadetes and Firmicutes in the infected Tibetan sheep were significantly decreased, whereas Actinobacteria, Chloroflexi, and Acidobacteria had significantly increased. At the genus level, the Christensenellaceae_R-7_group and Ruminococcaceae_NK4A214_group were the predominant bacterial genera in all the samples. Furthermore, the healthy Tibetan sheep exhibited higher abundances of Intestinimonas, Butyrivibrio, Pseudobutyrivibrio, Ruminococcaceae, Eubacterium_coprostanoligenes_group, Oxobacter, Prevotella_1, Ruminiclostridium_6, Coprococcus_1, Ruminococcus, Lachnospiraceae_UCG-002, Olsenella, and Acetitomaculum, whereas Kocuria, Clostridium_sensu_stricto_1, Slackia, Achromobacter, and Stenotrophomonas levels were lower. In conclusion, our results conveyed an information that E. granulosus infection may cause an increase in pathogenic bacteria and a decrease in beneficial bacteria. Additionally, a significant dynamical change in gut microbiota could be associated with E. granulosus infection.

Genome-wide Analysis of CNVs in Three Populations of Tibetan Sheep using Whole-genome Resequencing

Background: Copy number variation (CNV), an important source of genomic structural variation, can disturb genetic structure, dosage, regulation and expression, and is associated with phenotypic diversity and adaptation to local environments in mammals. In the present study, 24 resequencing datasets were used to characterize CNVs in three ecotypic populations of Tibetan sheep and assess CNVs related to domestication and adaptations in the Qinghai-Tibetan Plateau.Results: A total of 87,832 CNV events accounting for 0.3% of the sheep genome were detected in three Tibetan sheep populations. After merging the overlapping CNVs, 2777 CNV regions (CNVRs) were obtained, among which 1098 CNVRs were shared by the three populations. The average length of these CNVRs was more than 3 kb, and duplication events were more frequent than deletions. Functional analysis showed that the shared CNVRs were significantly enriched in 56 GO terms and 18 KEGG pathways that were mainly concerned with ABC transporters, olfactory transduction and oxygen transport. Moreover, 188 CNVRs overlapped with 97 quantitative trait loci (QTLs), such as growth and carcass QTLs, immunoglobulin QTLs, milk yield QTLs and fecal counts QTLs. PCDH15, APP and GRID2 overlapped with body weight QTLs. Furthermore, Vst analysis at each CNVR showed that RUNX1, LOC101104348, LOC105604082 and PAG11 were highly divergent between HTS and VTS, and RUNX1 and LOC101111988 were significantly differentiated between VTS and OTS. Meaningfully, the duplication of RUNX1 may facilitate the hypoxia adaptation of OTS and HTS in QTP, which deserves further research in detail.Conclusions: In this study, we represented the genome-wide distribution characteristics of CNVs in Tibetan sheep and provided a valuable genetic variation resource, which will facilitate the elucidation of the genetic basis underlying the distinct phenotypic traits and local adaptation of Tibetan sheep.

Variations in HIF-1α Contributed to High Altitude Hypoxia Adaptation via Affected Oxygen Metabolism in Tibetan Sheep

The Tibetan sheep is an indigenous species of the Tibetan plateau and has been well adapted to high-altitude hypoxia. In comparison to lowland sheep breeds, the blood gas indicators have changed and the HIFs signaling pathway is activated in Tibetan sheep. These phenotypic and genetic alterations in Tibetan sheep are thought to be an important basis for adaptation to high-altitude hypoxia and variation in genes encoding the subunits that make up HIFs, such as HIF-1α can affect blood gas indicators. In this study, exons 9, 10, 12 of the HIF-1α gene were sequenced to find variations and 3 SNPs were detected, and these 3 SNPs were genotyped by KASP in 341 Hu sheep and 391 Tibetan sheep. In addition, 197 Hu sheep, 160 Tibetan sheep and 12 Gansu alpine merino sheep were used for blood gas indicators analysis. The results showed significant differences between the blood gas indicators of high-altitude breeds (Tibetan sheep and Gansu alpine merino sheep) and low-altitude breeds (Hu sheep), implying that the differences in blood gas indicators are mainly caused by differences in altitude. The haplotype combinations H2H3 and H1H3 were more frequent in the Tibetan sheep population, H2H3 increases O2 carrying capacity by increasing hematocrit and hemoglobin concentrations; H1H3 makes O2 dissociate more readily from oxyhemoglobin by decreasing partial pressure of oxygen and oxygen saturation. These results suggest that variants at the HIF-1α promote the ability of oxygen utilization in Tibetan sheep, which may underpin the survival and reproduction of Tibetan sheep on the Tibetan plateau.

Rumen Fermentation—Microbiota—Host Gene Expression Interactions to Reveal the Adaptability of Tibetan Sheep in Different Periods

As an important ruminant on the Qinghai-Tibet Plateau, Tibetan sheep can maintain their population reproduction rate in the harsh high-altitude environment of low temperature and low oxygen, which relies on their special plateau adaptations mechanism that they have formed for a long time. Microbiomes (known as “second genomes”) are closely related to the nutrient absorption, adaptability, and health of the host. In this study, rumen fermentation characteristics, the microbiota, and rumen epithelial gene expression of Tibetan sheep in various months were analyzed. The results show that the rumen fermentation characteristics of Tibetan sheep differed in different months. The total SCFAs (short-chain fatty acids), acetate, propionate, and butyrate concentrations were highest in October and lowest in June. The CL (cellulase) activity was highest in February, while the ACX (acid xylanase) activity was highest in April. In addition, the diversity and abundance of rumen microbes differed in different months. Bacteroidetes (53.4%) and Firmicutes (27.4%) were the dominant phyla. Prevotella_1 and Rikenellaceae_RC9_gut_group were the dominant genera. The abundance of Prevotella_1 was highest in June (27.8%) and lowest in December (17.8%). In addition, the expression of CLAUDIN4 (Claudin-4) and ZO1 (Zonula occludens 1) was significantly higher in April than in August and December, while the expression of SGLT1 (Sodium glucose linked transporter 1) was highest in August. Correlation analysis showed that there were interactions among rumen fermentation characteristics, the microbiota, and host gene expression, mainly by adjusting the amino acid metabolism pathway and energy metabolism pathway to improve energy utilization. At the same time, we adjusted the balance of the rumen “core microbiota” to promote the development of rumen and maintain the homeostasis of rumen environment, which makes Tibetan sheep better able to adapt to the harsh environment in different periods of the Qinghai-Tibet Plateau.

Molecular detection of Anaplasma spp., Babesia spp. and Theileria spp. in yaks (Bos grunniens) and Tibetan sheep (Ovis aries) on the Qinghai-Tibetan Plateau, China

Background Anaplasma, Babesia and Theileria are tick-borne pathogens (TBPs) that affect livestock worldwide. However, information on these pathogens in yaks (Bos grunniens) and Tibetan sheep (Ovis aries) on the Qinghai-Tibet Plateau (QTP), China, is limited. In this study, Anaplasma spp., Babesia spp. and Theileria spp. infections were assessed in yaks and Tibetan sheep from Qinghai Province. Methods A total of 734 blood samples were collected from 425 yaks and 309 Tibetan sheep at nine sampling sites. Standard or nested polymerase chain reaction was employed to screen all the blood samples using species- or genus-specific primers. Results The results showed that 14.1% (60/425) of yaks and 79.9% (247/309) of Tibetan sheep were infected with at least one pathogen. Anaplasma ovis, Anaplasma bovis, Anaplasma capra, Anaplasma phagocytophilum, Babesia bovis and Theileria spp. were detected in this study, with total infection rates for all the assessed animals of 22.1% (162/734), 16.3% (120/734), 23.6% (173/734), 8.2% (60/734), 2.7% (20/734) and 19.3% (142/734), respectively. For yaks, the infection rate of A. bovis was 6.4% (27/425), that of B. bovis was 4.7% (20/425) and that of Theileria spp. was 3.3% (14/425). Moreover, 52.4% (162/309) of the Tibetan sheep samples were infected with A. ovis, 30.1% (93/309) with A. bovis, 56.0% (173/309) with A. capra, 19.4% (60/309) with A. phagocytophilum and 41.4% (128/309) with Theileria spp. Conclusions This study revealed the prevalence of Anaplasma spp., Babesia spp. and Theileria spp. in yaks and Tibetan sheep in Qinghai Province, China, and provides new data for a better understanding of the epidemiology of TBPs in these animals in this area of the QTP, China. Graphical Abstract

Seasonal diets supersede host species in shaping the distal gut microbiota of Yaks and Tibetan sheep

Yaks and Tibetan sheep are important and renowned livestock of the Qinghai-Tibetan Plateau (QTP). Both host genetics and environmental factors can shape the composition of gut microbiota, however, there is still no consensus on which is the more dominant factor. To investigate the influence of hosts and seasons on the gut microbiome diversity component, we collected fecal samples from yaks and Tibetan sheep across different seasons (summer and winter), during which they consumed different diets. Using 16S rRNA sequencing, principal component analysis (PCoA) data showed that PCo1 explained 57.4% of the observed variance (P = 0.001) and clearly divided winter samples from summer ones, while PCo2 explained 7.1% of observed variance (P = 0.001) and mainly highlighted differences in host species. Cluster analysis data revealed that the gut microbiota composition displayed a convergence caused by season and not by genetics. Further, we profiled the gut microbial community and found that the more dominant genera in yak and Tibetan sheep microbiota were influenced by seasonal diets factors rather than genetics. This study therefore indicated that seasonal diet can trump host genetics even at higher taxonomic levels, thus providing a cautionary note for the breeding and management of these two species.

Astragalus membranaceus Alters Rumen Bacteria to Enhance Fiber Digestion, Improves Antioxidant Capacity and Immunity Indices of Small Intestinal Mucosa, and Enhances Liver Metabolites for Energy Synthesis in Tibetan Sheep

Natural, non-toxic feed additives can potentially replace chemical medications and antibiotics that are offered sheep to improve performance. In the present study, Tibetan sheep were supplemented with the root of Astragalus membranaceus (AMT), a traditional herb used widely in China. Twenty-four male Tibetan sheep (31 ± 1.4 kg; 9-month-old) were assigned randomly to one of four levels of supplementary AMT: 0 g/kg (A0), 20 g/kg (A20), 50 g/kg (A50) and 80 g/kg (A80) dry matter intake (DMI). The A50 and A80 groups increased the diversity of rumen bacteria on d 14 and the relative abundances of fiber decomposing bacteria. Supplementary AMT upregulated the metabolism of vitamins, nucleotides, amino acids and glycan, and downregulated the metabolism of lipids and carbohydrates. In addition, supplementary AMT enriched rumen bacteria for drug resistance, and reduced bacteria incurring cell motility. In general, AMT supplementation increased the concentrations of catalase (CAT), superoxide dismutase (SOD) total antioxidant capacity (T-AOC) and secretory immunoglobulin A (sIgA) in the small intestinal mucosa and CAT and SOD in meat tissue. The liver tissue metabolome response showed that AMT in the A80 lambs compared to the A0 lambs upregulated the metabolites for energy synthesis. It was concluded that supplementary A. membranaceus increased the relative abundances of fiber decomposing bacteria and improved the antioxidant capacities and immunity indices of small intestinal mucosa and meat tissue in Tibetan sheep.