本文選題：節(jié)桿菌 + 篩選鑒定�。� 參考：《河北大學(xué)》2017年碩士論文

【摘要】：在現(xiàn)代集約化高密度養(yǎng)殖模式下,養(yǎng)殖過(guò)程中大量的殘餌,糞便排泄物,動(dòng)植物尸體等含氮有機(jī)物污染嚴(yán)重,因此本實(shí)驗(yàn)主要以氨氮和亞硝態(tài)氮降解能力為主要篩選指標(biāo)進(jìn)行目的菌株的篩選。本實(shí)驗(yàn)獲得一株異養(yǎng)硝化好氧反硝化細(xì)菌,并對(duì)其進(jìn)行了脫氮性能的測(cè)試、菌種的鑒定、發(fā)酵培養(yǎng)基及培養(yǎng)條件的優(yōu)化,分析了菌株對(duì)凡納濱對(duì)蝦(Litopenaeus vannamei)在養(yǎng)殖使用中的安全性以及在凡納濱對(duì)蝦養(yǎng)殖池塘的脫氮作用。研究主要結(jié)果如下:1.異養(yǎng)硝化好氧反硝化細(xì)菌菌株的篩選與鑒定將實(shí)驗(yàn)室保存的10株菌株進(jìn)行異養(yǎng)硝化培養(yǎng)48 h,得到4株氨氮和總氮降解效果較好的菌株,然后將4株菌株分別在反硝化培養(yǎng)基A和反硝化培養(yǎng)基B中進(jìn)行反硝化培養(yǎng),其中菌株7-3在48 h的降解效果最好,48 h硝態(tài)氮去除率為47.37%,亞硝態(tài)氮去除率為100.00%。經(jīng)形態(tài)特征、生理生化、16SrRNA序列分析和系統(tǒng)發(fā)育樹(shù)比較,對(duì)菌株進(jìn)行了菌種鑒定,菌株7-3為邁索爾節(jié)桿菌(Arthrobacter mysorens)。經(jīng)生長(zhǎng)特性研究表明,菌株7-3的0~6 h處于遲緩期,6 h后進(jìn)入對(duì)數(shù)生長(zhǎng)期,33 h后菌體達(dá)到最大生長(zhǎng)量,進(jìn)入穩(wěn)定期,48 h后細(xì)菌進(jìn)入衰亡期。2.節(jié)桿菌7-3液體發(fā)酵培養(yǎng)基配方及條件優(yōu)化以蔗糖蛋白胨培養(yǎng)基作為基礎(chǔ)培養(yǎng)基,采用單因子試驗(yàn)和L9(34)正交試驗(yàn)優(yōu)化了節(jié)桿菌7-3種子培養(yǎng)基的組成,并對(duì)其接種量、發(fā)酵溫度、轉(zhuǎn)速、初始pH值和裝液量等發(fā)酵條件進(jìn)行了優(yōu)化,得到優(yōu)化后的發(fā)酵培養(yǎng)基配方為:玉米粉1%,酵母浸膏1%,K2HPO4 0.2%,KH2PO4 0.2%。確定了最佳培養(yǎng)條件:接種量為7%,發(fā)酵溫度為25℃,轉(zhuǎn)速為220 r/min,初始p H值8.0,裝液量為100 m L。經(jīng)發(fā)酵培養(yǎng)基及發(fā)酵條件的優(yōu)化后,發(fā)酵液活菌數(shù)達(dá)到4.04×1010 CFU/mL。3.菌株對(duì)凡納濱對(duì)蝦的安全性實(shí)驗(yàn)將三株菌的菌液濃度分別設(shè)置為104 CFU/mL、106 CFU/mL、108 CFU/m L三個(gè)濃度梯度添加到5 L凡納濱對(duì)蝦幼蝦養(yǎng)殖容器中,觀察7 d對(duì)蝦的死亡情況,結(jié)果表明,菌株F1508在水體中的含菌量為108 CFU/m L濃度時(shí),對(duì)蝦的累計(jì)死亡率與對(duì)照組無(wú)顯著差異;菌株7-3和菌株683在水體中的含菌量為106 CFU/mL濃度時(shí),對(duì)蝦的累計(jì)死亡率與對(duì)照組無(wú)顯著差異。菌株F1508在108 CFU/mL濃度以下使用時(shí)和菌株7-3和菌株683在含菌量為106 CFU/mL濃度以下在對(duì)蝦養(yǎng)殖使用時(shí)是安全的。4.菌株在對(duì)蝦養(yǎng)殖水體中對(duì)氨氮、亞硝態(tài)氮的脫氮作用將復(fù)合菌制劑應(yīng)用到凡納濱對(duì)蝦海水養(yǎng)殖池塘。試驗(yàn)周期15 d,每天采樣檢測(cè)氨氮、亞硝態(tài)氮的濃度。實(shí)驗(yàn)結(jié)果表明復(fù)合菌劑對(duì)養(yǎng)殖水體中的氨氮和亞硝態(tài)氮均有一定的降解作用,氨氮最大去除率達(dá)到80.31%,亞硝態(tài)氮最大去除率達(dá)到85.58%,對(duì)養(yǎng)殖水體pH的影響均不顯著,p H值波動(dòng)范圍在8.1~8.3。
[Abstract]:In the modern intensive high-density culture model, a large number of residual bait, feces, animal and plant carcasses and other nitrogen-containing organic pollutants are seriously polluted during the breeding process. Therefore, the main screening indexes were ammonia nitrogen and nitrite nitrogen degradation ability. In this experiment, a heterotrophic nitrifying aerobic denitrifying bacteria was obtained, and its denitrification performance, strain identification, fermentation medium and culture conditions were optimized. The safety of the strain to Litopenaeus vannamei) in culture and the denitrification in the pond of Penaeus vannamei) were analyzed. The main results are as follows: 1. Screening and identification of heterotrophic aerobic denitrifying bacteria 4 strains with better degradation of ammonia nitrogen and total nitrogen were obtained by heterotrophic nitrification culture of 10 strains stored in laboratory for 48 h. Then, four strains were cultured in denitrification medium A and denitrification medium B, respectively. The best degradation rate of strain 7-3 was 47.37% and 100.00g% respectively at 48 h. Based on the morphological characteristics, physiological and biochemical analysis of 16s rRNA sequence and comparison of phylogenetic tree, the strain 7-3 was identified as Arthrobacter mysorens. The growth characteristics of strain 7-3 were studied. It was found that the bacteria of strain 7-3 reached the maximum growth rate after entering the logarithmic growth stage for 33 h after 6 h of slow growth and 48 h after entering the stable phase. The formulation and conditions of the liquid fermentation medium of Arthrobacter ganglii 7-3 were optimized by using sucrose peptone medium as the basic medium, and the composition of the medium was optimized by single factor test and L9N 34) orthogonal experiment. The inoculation amount and fermentation temperature of the medium were optimized. The fermentation conditions, such as rotation speed, initial pH value and liquid volume, were optimized. The optimized fermentation medium was obtained as follows: corn powder 1, yeast extract 1 K2HPO 4 0.2 and KH 2PO 4 0.2. The optimum culture conditions were determined as follows: inoculation amount was 7, fermentation temperature was 25 鈩，

本文編號(hào)：2005326