http:∥xuebao.gxnu.edu.cn

表 4 Alpha 多樣性指數(shù)分析結(jié)果

Tab. 4 Alpha diversity index

樣本組別

多樣性指數(shù)值

Ace Chao1 Shannon Simpson

MA 313.14±5.99 319.61±5.47 3.62±0.09

ab

0.06±0.01

ab

MB 314.46±3.37 315.88±3.48 3.72±0.16

ab

0.06±0.01

ab

MC 310.67±4.84 315.27±6.40 3.38±0.14

b

0.10±0.01

ab

MD 307.23±20.01 305.31±25.97 3.26±0.46

ab

0.15±0.08

a

ME 311.10±8.92 318.39±8.20 3.76±0.19

ab

0.15±0.08

a

MF 304.13±8.34 307.57±8.07 3.87±0.07

a

0.05±0.01

b

注:同列數(shù)據(jù)肩標(biāo)小寫(xiě)字母相鄰表示差異顯著(P<0.05),小寫(xiě)字母相間表示差異極顯著(P<0.01)。

2.3.2 腸道菌群 Beta 多樣性分析

腸道菌群 Beta 多樣性的主坐標(biāo)分析(PCoA)和無(wú)度量多維標(biāo)定法(NMDS)分析結(jié)果見(jiàn)圖 4。 將數(shù)據(jù)

以點(diǎn)的形式反映在二維坐標(biāo)圖上,樣本距離越近,表示物種組成結(jié)構(gòu)越相似。 6 個(gè)組樣本極大程度聚集在

一起,微生物群落結(jié)構(gòu)的差異顯著(P>0.05),表明 6 個(gè)組樣本的微生物群落結(jié)構(gòu)具有較高的相似性。

點(diǎn)分別表示各樣品;不同顏色代表不同分組;橫、縱坐標(biāo)為導(dǎo)致樣品間差異最大的 2 個(gè)特征值,以百分?jǐn)?shù)的形式體現(xiàn)主要影響程度

圖 4 PCoA 和 NMDS 分析結(jié)果

Fig. 4 PCoA and NMDS analysis chart

3 討論

3.1 BCAAs 比例對(duì)攸縣麻鴨腸道菌群組成的影響

腸道微生物主要是指腸道細(xì)菌,由有益菌、致病菌及中性菌構(gòu)成,腸道細(xì)菌種群多達(dá) 500 種,但這些種

主要?dú)w屬于少數(shù)的幾個(gè)門(mén)類(lèi)[18]

。 研究發(fā)現(xiàn),厚壁菌門(mén)和擬桿菌門(mén)是單胃大型動(dòng)物與小型動(dòng)物腸道的兩大

優(yōu)勢(shì)門(mén)類(lèi)[18]

。 除去以上兩大優(yōu)勢(shì)門(mén)類(lèi),鼠腸道中的主要菌群還有變形菌門(mén)與脫鐵桿菌門(mén)[19]

,反芻動(dòng)物和

雞腸道中主要菌群還有變形菌門(mén)[20-21]

。 本研究中,兩大優(yōu)勢(shì)門(mén)類(lèi)為厚壁菌門(mén)和擬桿菌門(mén),這與前人研究

結(jié)果[18-21]相一致,第三大優(yōu)勢(shì)門(mén)類(lèi)為變形菌門(mén),與反芻動(dòng)物和雞腸道主要菌群更為一致。 以上結(jié)果表明,

動(dòng)物腸道的優(yōu)勢(shì)門(mén)類(lèi)不會(huì)被輕易改變,但飼糧中添加 BCAAs 會(huì)對(duì)其中部分主要菌門(mén)有一定的影響。 另有

研究指出,15 月齡小鼠腸道中 BCAAs 補(bǔ)充組的擬桿菌門(mén)豐度較對(duì)照組下降3.9%,厚壁菌門(mén)豐度上升

247

廣西師范大學(xué)學(xué)報(bào)(自然科學(xué)版),2022,40(2)

3.62%,變形菌門(mén)作為其第三優(yōu)勢(shì)門(mén)類(lèi)在補(bǔ)充組中的豐度遠(yuǎn)遠(yuǎn)低于對(duì)照組[17]

,而變形菌門(mén)中通常包含較多

的致病菌,因此,補(bǔ)充 BCAAs 可能有助于降低腸道菌群中的致病菌。 結(jié)合本研究,ME 組變形菌門(mén)豐度最

低且顯著低于 MD 組,可以推測(cè) BCAAs 中 Leu 比例在三者中為 37% ~40%時(shí),更有益于攸縣麻鴨的腸道健

康。 本研究還發(fā)現(xiàn) MF 組放線(xiàn)菌門(mén)豐度顯著高于 MA 組,放線(xiàn)菌門(mén)中通常包含較多的共生菌與益生菌,所

以 MF 組可能更有益于攸縣麻鴨腸道菌群。

屬水平上,BCAAs 比例對(duì)豐度前 10 的菌屬均無(wú)顯著影響,然而各組間仍有一定差異。 擬桿菌是腸道

中的共生菌,為身體提供必需的營(yíng)養(yǎng),在 MC 與 MF 組中豐度最高;脫硫弧菌是常見(jiàn)的致病菌,在 MA 組的

豐度最高。 BCAAs 可以被腸腔內(nèi)的細(xì)菌利用[22-24]

,提示它們可能參與調(diào)節(jié)腸道菌群的組成。 在對(duì)小鼠的

研究中,發(fā)現(xiàn) BCAAs 能夠影響其腸道菌群的結(jié)構(gòu),并增加潛在益生菌雙歧桿菌的豐度和降低致病菌腸桿

菌的比例[17]

。 本研究中 MF 組出現(xiàn)較多的雙歧桿菌。 此外,豬的低蛋白飲食降低了腸道乳酸菌水平,但

BCAAs 平衡顯著改善了乳酸菌的增殖[25]

;Ile 促進(jìn)了鯉魚(yú)腸中乳酸桿菌和芽孢桿菌的生長(zhǎng),抑制了氣單胞

菌和大腸桿菌的生長(zhǎng)[26]

。 然而對(duì)仔豬的研究中卻發(fā)現(xiàn) BCAAs 補(bǔ)充組與正常蛋白組的結(jié)腸微生物菌群沒(méi)

有差異[22]

。 以上表明,BCAAs 對(duì)腸道菌群一定是有影響的,得到不一致的結(jié)果可能與動(dòng)物品種、飼料組成

和 BCAAs 添加水平以及比例等問(wèn)題有關(guān)。 因此,需要進(jìn)一步深入研究來(lái)揭示 BCAAs 對(duì)機(jī)體腸道菌群的

影響。

3.2 BCAAs 比例對(duì)攸縣麻鴨腸道菌群結(jié)構(gòu)與多樣性的影響

Yang 等[17]研究指出,補(bǔ)充 BCAAs 使小鼠腸道微生物多樣性更豐富;文獻(xiàn)[22-24]認(rèn)為,BCAAs 通過(guò)

被腸道中細(xì)菌利用進(jìn)而可能參與腸道微生物種類(lèi)和多樣性的調(diào)節(jié)。 但目前沒(méi)有發(fā)現(xiàn) BCAAs 比例對(duì)腸道

菌群影響的研究。 在本研究中,Alpha 多樣性分析結(jié)果表明,6 組 Ace 和 Chao1 指數(shù)無(wú)顯著差異(即 6 組具

有相同的物種豐度),而 MF 組 Shannon 指數(shù)顯著大于 MC 組,Simpson 指數(shù)極顯著小于 MC 組和 MA 組。

在相同物種豐度條件下,Shannon 指數(shù)越大,Simpson 指數(shù)越小,說(shuō)明物種多樣性越高,由此可見(jiàn),在本研究

條件下,MF 組具有最高的微生物多樣性,MC 組腸道微生物多樣性最低。 在 Beta 多樣性分析中,PCoA 與

NMDS 分析都是采取降維的思想對(duì)多樣本數(shù)據(jù)進(jìn)行定位、分析和歸類(lèi),而 NMDS 分析克服了以前排序方法

(包括 PCoA 在內(nèi))的缺點(diǎn),更具有可靠性。 結(jié)合 OTU 聚類(lèi)分析發(fā)現(xiàn),雖然 MD 組具有一特有的 OTU,但 6

組微生物菌落結(jié)構(gòu)仍具有較高的相似性,表明不同 BCAAs 比例在該研究條件下沒(méi)有對(duì)攸縣麻鴨的腸道菌

群結(jié)構(gòu)造成影響。

4 結(jié)論

63 日齡攸縣麻鴨盲腸的核心菌群是擬桿菌門(mén)、厚壁菌門(mén)、變形菌門(mén),BCAAs 比例顯著影響變形菌門(mén)及

放線(xiàn)菌門(mén),當(dāng)亮氨酸、纈氨酸、異亮氨酸的質(zhì)量比 m(Leu) ∶ m(Val) ∶ m(Ile)= 1 ∶ 0.9 ∶ 0.8 時(shí),攸縣麻鴨

盲腸微生物物種多樣性最高。

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Effect of Dietary Branched-chain Amino Acid Ratio on Intestinal

Flora of 28-63 Days Old Youxian Duck

SUN Yue

1,2,3

, DAI Qiuzhong

2,4

, JIANG Guitao

2,4

, HUANG Xuan

2,4

,

LI Chuang

2,4

, DENG Ping

2,4

, SUN Tao

1,3?

(1. College of Life Science, Guangxi Normal University, Guilin Guangxi 541006, China; 2. Hunan Institute of

Animal Science and Veterinary Medicine, Changsha Hunan 410131, China; 3. Key Laboratory of Ecology of Rare and

Endangered Species and Environmental Protection (Guangxi Normal University), Ministry of Education, Guilin

Guangxi 541006, China; 4. Hunan Engineering Research Center of Poultry Production Safety, Changsha Hunan 410128, China)

Abstract: A single factor experiment was adopted to investigate the effects of dietary branched-chain Amino Acid

ratio flora of 28-63 days old Youxian duck. 504 Youxian ducks of 28 day-old and healthy and similar in weight

were randomly allocated to 6 groups with 6 replicates each group and 14 in each replicate. Six diets with leucine,

valine and isoleucine ratios of 1 ∶ 0.4 ∶ 0.3 ( group MA), 1 ∶ 0.5 ∶ 0.4 ( group MB), 1 ∶ 0.6 ∶ 0.5 ( group

MC), 1 ∶ 0.7 ∶ 0.6(group MD), 1 ∶ 0.8 ∶ 0.7(group ME) and 1 ∶ 0.9 ∶ 0.8 (group MF) were avilable. The

experiment lasted for 35 days. At the end of the feeding experiment, one duck was selected from each replicate of

each treatment to collect cecal contents, and the cecal microbial structure and flora diversity were detected and

analyzed by 16S rDNA sequencing technology. The results show that: 1) There are 344 identical OTUs in the 6

groups, and only one unique OTU existed in group MD. 2) At the phylum level, the core flora are Bacteroidetes,

Firmicutes and Proteobacteria. At the genus level, Bacteroides and Desulfovibrio are the dominant genera; 3)

The results of comparion among the 6 groups show that, the abundance of Proteobacteria in group MD is

significantly higher than that in group ME ( P < 0. 05), and the abundance of Actinomycetes in group MF is

significantly higher than that in group MA (P<0.05); 4) There is no significant difference in Ace index and

Chao1 index among the 6 groups, however, the Shannon index in group MF is significantly higher than that in

group MC (P<0.05), and Simpson index in group MC is significantly higher than that in group MF (P<0.01)

and significantly higher than that in group MA (P<0.05). In summary, the core flora of the cecum of 63-day-old

Youxian duck is Bacteroides, Firmicutes, and Proteobacteria. The proportion of branched-chain amino acids in

diets significantly affects the Proteobacteria and Actinomycetes. When the ratio of mleucine, valine and

isoleucine is 1 ∶ 0.9 ∶ 0.8, the microbial species diversity of the cecum in Youxian duck is the highest.

Keywords: branched-chain amino acid; Youxian duck; intestinal flora; species abundance; species diversity

(責(zé)任編輯 馬殷華)

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