Comparative analysis of the rectal\nand caecal microbial community composition and function\nin adult Erhualian and Sushan pigs

Abstract

Intestinal microbiota plays an important role in nutrition, metabolism and immunity in all mammals. It is comprised of diverse populations of bacteria and other microorganisms whose abundances are impacted by both environmental and host genetic factors. However, the understandings of the intestinal microbiota in different pig breeds remain largely undefined. To examine the differences in intestinal microflora between two pig breeds with different genetic backgrounds under the same environment, 16S rRNA gene amplification and sequencing were performed to investigate the structural composition and potential functions of microbial communities in rectum and caecum of Erhualian and Sushan pigs. The results revealed that the diversity of intestinal microflora in two pig breeds was similar, but the abundance of specific intestinal microflora was different. At the phylum level, the dominant bacteria in caecum and rectum of Erhualian and Sushan pigs were Firmicutes, Acidobacteria and Bacteroides, but their expression abundance was different. Firmicutes and Bacteroidetes in Erhualian pigs were higher than those in Sushan pigs. At the genus level, Lactobacillus was the most abundant in caecum of Sushan pigs (6.83%) and rectum of Erhualian pigs (9.61%), while Ruminococcaceae UCG-005 were dominant in caecum of Erhualian pigs (10.89%) and Streptococcus in rectum of Sushan pigs (24.89%). This study further confirmed the existence of specific microbial community diversity and abundance in different pig breeds. The microbial community diversity and abundance in Erhualian and Sushan pigs were closely related to pig fat deposition and nutrient absorption. 1 Corresponding author: e-mail: [email protected]; [email protected] 2 Intestinal microorganisms in pigs of Landrace, Yorkshire and Duroc pigs. However, Bama mini, Erhualian and Xiaomeishan pigs from Chinese local breeds have high similarity when it comes to microorganisms presence and type. Diao et al. (2016) showed that intestinal microbial abundances in Rongchang, Tibetan and Landrace pigs were different. Tibetan and Rongchang pigs had a higher proportion of Firmicutes and Spirochaetes and a lower proportion of Bacteroidetes than Landrace pigs, and the proportion of Spirochaetes in Tibetan pigs was significantly higher than that in Rongchang pigs. Compared with Tibetan pigs, Landrace and Rongchang pigs contained a higher proportion of Tenericutes and a lower proportion of Fibrobacteres and Elusimicrobia. The above results indicated that the composition of intestinal microorganisms in pigs was probably related to the host’s metabolic type, feeding characteristics and immune function. Erhualian pig is a well-known local pig breed in China, with strong lactation, good motherhood, a high feeding rate, early sexual maturity, especially high resistance to rough feeding and a docile temperament. Sushan pig, as a new hybrid breed of Erhualian and Yorkshire pigs, has the advantages of delicious meat and a certain level of resistance to rough feeding. In pig production, the tolerance to rough feeding and crude fibre of Sushan pigs was found to be lower than that of Erhualian pigs, and the demand of Sushan pigs for nutrients in feed is higher than that of Erhualian pigs. In order to study the difference of tolerance to rough feeding and crude fibre between Erhualian and Sushan pigs, characterization and comparative analysis were performed to investigate the structural composition and potential functions of microbial communities in these two breeds. This study provides a powerful theoretical basis on the potential roles of intestinal microbial communities in resistance to rough feeding, nutrition metabolism and crude fibre digestion for safe and healthy pork production. Material and methods This experiment was reviewed and approved by the Institutional Animal Ethics Committee from the Research Integrity and Ethics Administration of Jiangsu Academy of Agricultural Sciences, China. Animals and sampling Erhualian and Sushan pigs (five barrows each) were fed at the Sushan Pig Breeding Farm (Nanjing, China) under the same conditions and provided with standard diets in accordance with the feeding standard of swine (NY/T 65-2004) issued by the Ministry of Agriculture of the People’s Republic of China (Table 1). At the rapid growth stage (175th day of age), the adult Erhualian (75 kg) and Sushan (90 kg) pigs were slaughtered according to standard procedures. The pigs were dissected for collecting the intestinal contents of the rectum and caecum. The samples were kept at −80 °C for 16S rRNA gene analysis. DNA extraction and PCR amplification Microbial DNA was extracted from rectal and caecal samples using an E.Z.N.A.® soil DNA Kit (Omega Bio-tek, Norcross, GA, USA) according to the manufacturer’s protocols. The final DNA concentration and purification were determined by a NanoDrop 2000 UV-V is spectrophotometer (Thermo Scientific, Waltham, MA, USA), and the DNA quality was checked by 1% agarose gel electrophoresis. The V3/V4 hypervariable regions of the bacterial 16S rRNA gene were amplified using primers 341F (5’-CCTAYGGGRBGCASCAG-3’) and 806R (5’-GGACTACNNGGGTATCTAAT-3’) based on a thermocycler PCR system (GeneAmp PCR System 9700, Applied Biosystem, Foster City, CA, USA). The PCRs were conducted with the following program: initial denaturation at 94 °C for 4 min; 94 °C denaturation for 30 s, 50 °C annealing for 45 s and 72 °C extension for 30 s, repeated for 25 cycles; and Table 1. Composition and nutrient levels of basal diets (air-dry basis), % Indices Content Ingredients maize 61.00 soyabean meal 9.00 wheat bran 27.50 CaHPO4 0.50 limestone 0.50 NaCl 0.50 premix1 1.00 Nutrient levels2 DM 85.43 ash 6.24 CP 15.41 EE 4.31 CF 3.02 DE, MJ /kg 18.32 Ca 0.54 P 0.47 DM – dry matter, CP – crude protein, EE – ether exract, CF – crude fibre, DE – digestible energy; 1 the premix provided the following per kg of diets: mg: Fe 100, Zn 100, Mn 30, Cu 10, Se 0.3, I 0.5, vit. K 3.0, vit. B1 2.0, vit. B2 6.0, vit. B6 3.0, nicotinic acid 30, pantothenic acid 30, folic acid 1.0, biotin 0.2, choline 300; IU: vit. A 8000, vit. D 31000, vit. E 20; μg: vit. B12 30; 2 DE was calculated, while the other values were calculated