水產(chǎn)養(yǎng)殖環(huán)境中磺胺類抗生素抗性基因污染的研究
發(fā)布時間：2018-08-06 19:29
【摘要】：抗生素不僅可以防治水產(chǎn)生物的細菌性疾病,同時還可以促進養(yǎng)殖動物的生長,因此在水產(chǎn)養(yǎng)殖業(yè)中的使用極為廣泛。近年來,隨著抗生素使用的加劇,養(yǎng)殖生態(tài)環(huán)境所遭受的抗生素污染日益嚴重,并由此誘導產(chǎn)生了一種新型的環(huán)境污染物——抗生素抗性基因(ARGs)。抗性基因可以通過基因的水平轉(zhuǎn)移進入人體,影響人類的健康,已經(jīng)引起了國內(nèi)外的廣泛關注。本文試圖以磺胺類抗生素為目標化學物,通過研究太湖周邊養(yǎng)殖區(qū)域(無錫、湖州、常州、蘇州、宜興)不同養(yǎng)殖品種(河蟹、青蝦、鱸魚、青魚、鯽魚、鳊魚、南美白對蝦)養(yǎng)殖池塘的水體和底泥沉積物中抗生素的殘留、抗性菌株的情況以及抗生素抗性基因的污染現(xiàn)狀和變化規(guī)律,并與相近的自然水域相比較,了解養(yǎng)殖水環(huán)境中抗生素抗性基因的污染殘留狀況。研究結(jié)果如下:1、分析了 2015年不同地區(qū)不同品種的養(yǎng)殖池塘在整個養(yǎng)殖周期中水樣和沉積物中磺胺類藥物的分布情況,發(fā)現(xiàn)在整個養(yǎng)殖季節(jié)中,池塘水體和沉積物中磺胺類的含量變化均呈現(xiàn)低-高-低的變化趨勢;2、整個養(yǎng)殖過程中,不同養(yǎng)殖池塘的水樣、沉積物中均可檢出磺胺類的耐藥微生物。隨著磺胺二甲基嘧啶濃度的升高,養(yǎng)殖池塘的水樣、沉積物中微生物的耐藥率均呈現(xiàn)下降趨勢,且濃度越高,耐藥菌的耐藥率越低。同時,相同藥物濃度下,沉積物中細菌的耐藥率要高于水中。3、通過抗性平板分離篩選出具有耐藥性的抗性菌株4株,結(jié)合菌體生理生化特性分析和16SrDNA序列分析,初步確定NHA1401、NHA1402菌株為奇異變形桿菌(Proteus mirabilis),SHA1501、CBA1506 菌株為葡萄球菌屬(Chryseobacterium sp.)和金黃桿菌屬(Staphylococcus sp.)的細菌。4、采用PCR方法對環(huán)太湖不同養(yǎng)殖區(qū)域不同養(yǎng)殖品種的養(yǎng)殖池塘,在同一養(yǎng)殖周期的不同時段(2015年的5-6月、7-8月、9-10月)的水樣和沉積物進行磺胺類抗性基因sulⅠ和sulⅡ的定性分析,結(jié)果顯示,在3個不同的養(yǎng)殖時段中,均可檢出磺胺類抗性基因sulⅠ和sulⅡ。養(yǎng)殖前期(5-6月)2種磺胺類抗性基因的檢出率最低,分別為80%和65%。隨著養(yǎng)殖進程的推進,sulⅠ基因呈現(xiàn)出先升高后下降的趨勢,sulⅡ抗性基因檢出率則在不斷升高。與此同時,磺胺類抗性基因sulⅠ、sulⅡ在太湖自然水體和沉積物中也有檢出。5、通過熒光定量PCR方法分析了環(huán)太湖不同養(yǎng)殖區(qū)域不同養(yǎng)殖品種的養(yǎng)殖池塘中的水樣和沉積物,在同一養(yǎng)殖周期的不同時段的磺胺類抗性基因sulⅠ和sulⅡ的相對豐度,結(jié)果顯示,(1)環(huán)太湖水產(chǎn)養(yǎng)殖區(qū)不同地區(qū)相同品種的養(yǎng)殖池塘環(huán)境中磺胺類抗性基因的相對豐度差異不大,最大不超過一個數(shù)量級;(2)在相同地區(qū)不同品種的水產(chǎn)養(yǎng)殖池塘中,南美白對蝦、青魚、鱸魚養(yǎng)殖池塘中的污染較青蝦、鳊魚為重。除河蟹外,青蝦、南美白對蝦、鱸魚、卿魚、鳊魚和青魚6個品種養(yǎng)殖池塘的水體遭受到的污染程度比沉積物要重,而河蟹養(yǎng)殖池塘則沉積物的污染較重。在水產(chǎn)養(yǎng)殖環(huán)境中,無論是水中還是沉積物中,抗性基因都是存在的,且差異性最大在1～2個數(shù)量級之間;(3)在相同地區(qū)、相同養(yǎng)殖品種的同一個養(yǎng)殖周期中,幾乎有90%的養(yǎng)殖池塘,其磺胺類抗性基因相對豐度的最高值出現(xiàn)在養(yǎng)殖中期的7-8月,只有極少數(shù)的例外。6、在太湖自然水體中,不同地理位置水域的水樣和沉積物中磺胺類抗性基因的相對豐度存在一定的差異,沉積物中磺胺類抗性基因的污染較水體為重。與同期附近的養(yǎng)殖區(qū)域相比,天然水體水樣中的污染程度輕于養(yǎng)殖區(qū),而沉積物中的污染程度與養(yǎng)殖區(qū)相當。研究表明,水產(chǎn)養(yǎng)殖環(huán)境中的抗生素抗性基因并非長期固定累積存在于池塘中,磺胺類抗性基因的產(chǎn)生與養(yǎng)殖周期、養(yǎng)殖中抗生素的投放和使用量存在一定的聯(lián)系。因此,嚴格規(guī)范和控制養(yǎng)殖過程中抗生素的使用,對抗性基因的產(chǎn)生和傳播可起到一定的防范作用,從而使抗性基因的轉(zhuǎn)移得到有效遏制,為人類健康提供保障。
[Abstract]:Antibiotics can not only prevent and cure the bacterial diseases of aquatic organisms, but also promote the growth of aquaculture animals. Therefore, the use of antibiotics is extremely widespread in aquaculture. In recent years, with the intensification of the use of antibiotics, the antibiotic pollution suffered by the ecological environment of aquaculture is increasingly severe, and a new environmental pollution has been induced. The antibiotic resistance gene (ARGs). Resistance genes can be transferred into the human body through the transfer of genes and affect human health. It has aroused widespread concern at home and abroad. This article attempts to use sulfonamides as the target chemicals, through the study of different aquaculture products in Taihu, Wuxi, Huzhou, Changzhou, Suzhou, Yixing). The residue of antibiotics in water and sediment sediments in aquaculture ponds, the situation of resistant strains and the pollution status and change rules of antibiotic resistance genes in aquaculture ponds, such as river crabs, penicus, perch, carp, bream, bream, and Penaeus prawns, and the pollution residue of antibiotic resistance genes in aquaculture water. The results are as follows: 1, the distribution of sulfonamides in water samples and sediments of different breed aquaculture ponds in different regions in 2015 was analyzed. It was found that the variation of sulfonamides in pond water and sediment showed a trend of low high low in the whole culture season, and 2, the whole culture process. In the water samples of different aquaculture ponds, the sulfonamides could be detected in the sediment. With the increase of the concentration of sulfamethazine, the resistance rate of microbes in the aquaculture pond and the sediment showed a decreasing trend, and the higher the concentration, the lower the resistance rate of the drug resistant bacteria. The resistance rate was higher than that of.3 in water. 4 strains resistant to resistance were screened by the resistance plate separation. Combined with the analysis of physiological and biochemical characteristics and 16SrDNA sequence analysis, NHA1401 was preliminarily identified, the strain NHA1402 was Proteus mirabilis, SHA1501, and CBA1506 strain was Staphylococcus (Chryseobacterium sp.) and golden yellow. Bacteriobacterium (Staphylococcus sp.) bacterial.4, the qualitative analysis of sulfonamide resistance genes Sul I and sul II in water samples and sediments at different periods of the same period (5-6 months, 7-8 months, 9-10 months, 2015) by PCR method was used to analyze the qualitative analysis of Sul I and sul II in the water samples and sediments at the same period of the same breeding cycle (5-6 months, 7-8 months, 9-10 months). Sulfonamides resistance genes Sul I and sul II were detected during the breeding period. The detection rates of 2 sulfonamides resistance genes were the lowest in the early stage of culture (5-6 months), respectively, 80% and 65%., respectively, with the advance of the breeding process, the Sul I gene showed a tendency to rise first and then decline, while the detection rate of the Sul II resistance gene was increasing. Gene Sul I, Sul II also detected.5 in Taihu natural water and sediment, and analyzed the water samples and sediments in different aquaculture ponds of different aquaculture regions around Taihu by fluorescence quantitative PCR method. The relative abundance of the sulfonamides resistance based on Sul I and sul II at the same period of the same breeding period showed that (1) The relative abundance of sulfonamides resistance genes in the environment of the same breed aquaculture ponds in different regions of the Taihu aquaculture area is not much different, and the maximum is not more than one order of magnitude. (2) in the aquaculture ponds of different varieties in the same area, the pollution of Penaeus prawns, green fish and perch ponds is more serious than the penaeid shrimp and bream. The water body of the 6 aquaculture ponds of shrimp, white white prawns, perch, Qing fish, bream and green fish in aquaculture ponds are heavier than the sediments, while the river crab aquaculture ponds have heavy pollution. In aquaculture environment, both in water and in sediment, the resistance base is present, and the difference is at the maximum of 1~2 orders of magnitude. (3) in the same area, almost 90% of the aquaculture ponds were found in the same breeding period of the same breed. The highest relative abundance of the sulfonamides resistance gene appeared at 7-8 months in the middle period of the culture, and only a few exceptions.6, in the natural waters of Taihu, the sulfonamides resistant base in the water samples from different geographical locations and the sediments. There is a certain difference in relative abundance, the pollution of the sulfonamides resistance gene in the sediments is heavier than that in the water body. Compared with the aquaculture area near the same period, the pollution degree of the natural water samples is lighter than that in the aquaculture area, and the pollution in the sediments is equivalent to that in the aquaculture area. The non long-term fixed accumulation exists in the pond. The production of the sulfonamides resistance gene is related to the breeding cycle and the dosage of antibiotics in the culture. Therefore, it is necessary to strictly regulate and control the use of antibiotics in the culture process and to prevent the production and transmission of the antagonistic genes, thus making the resistant gene. Transfer is effectively curbed, providing protection for human health.
【學位授予單位】：南京農(nóng)業(yè)大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：S948
本文編號：2168755
[Abstract]:Antibiotics can not only prevent and cure the bacterial diseases of aquatic organisms, but also promote the growth of aquaculture animals. Therefore, the use of antibiotics is extremely widespread in aquaculture. In recent years, with the intensification of the use of antibiotics, the antibiotic pollution suffered by the ecological environment of aquaculture is increasingly severe, and a new environmental pollution has been induced. The antibiotic resistance gene (ARGs). Resistance genes can be transferred into the human body through the transfer of genes and affect human health. It has aroused widespread concern at home and abroad. This article attempts to use sulfonamides as the target chemicals, through the study of different aquaculture products in Taihu, Wuxi, Huzhou, Changzhou, Suzhou, Yixing). The residue of antibiotics in water and sediment sediments in aquaculture ponds, the situation of resistant strains and the pollution status and change rules of antibiotic resistance genes in aquaculture ponds, such as river crabs, penicus, perch, carp, bream, bream, and Penaeus prawns, and the pollution residue of antibiotic resistance genes in aquaculture water. The results are as follows: 1, the distribution of sulfonamides in water samples and sediments of different breed aquaculture ponds in different regions in 2015 was analyzed. It was found that the variation of sulfonamides in pond water and sediment showed a trend of low high low in the whole culture season, and 2, the whole culture process. In the water samples of different aquaculture ponds, the sulfonamides could be detected in the sediment. With the increase of the concentration of sulfamethazine, the resistance rate of microbes in the aquaculture pond and the sediment showed a decreasing trend, and the higher the concentration, the lower the resistance rate of the drug resistant bacteria. The resistance rate was higher than that of.3 in water. 4 strains resistant to resistance were screened by the resistance plate separation. Combined with the analysis of physiological and biochemical characteristics and 16SrDNA sequence analysis, NHA1401 was preliminarily identified, the strain NHA1402 was Proteus mirabilis, SHA1501, and CBA1506 strain was Staphylococcus (Chryseobacterium sp.) and golden yellow. Bacteriobacterium (Staphylococcus sp.) bacterial.4, the qualitative analysis of sulfonamide resistance genes Sul I and sul II in water samples and sediments at different periods of the same period (5-6 months, 7-8 months, 9-10 months, 2015) by PCR method was used to analyze the qualitative analysis of Sul I and sul II in the water samples and sediments at the same period of the same breeding cycle (5-6 months, 7-8 months, 9-10 months). Sulfonamides resistance genes Sul I and sul II were detected during the breeding period. The detection rates of 2 sulfonamides resistance genes were the lowest in the early stage of culture (5-6 months), respectively, 80% and 65%., respectively, with the advance of the breeding process, the Sul I gene showed a tendency to rise first and then decline, while the detection rate of the Sul II resistance gene was increasing. Gene Sul I, Sul II also detected.5 in Taihu natural water and sediment, and analyzed the water samples and sediments in different aquaculture ponds of different aquaculture regions around Taihu by fluorescence quantitative PCR method. The relative abundance of the sulfonamides resistance based on Sul I and sul II at the same period of the same breeding period showed that (1) The relative abundance of sulfonamides resistance genes in the environment of the same breed aquaculture ponds in different regions of the Taihu aquaculture area is not much different, and the maximum is not more than one order of magnitude. (2) in the aquaculture ponds of different varieties in the same area, the pollution of Penaeus prawns, green fish and perch ponds is more serious than the penaeid shrimp and bream. The water body of the 6 aquaculture ponds of shrimp, white white prawns, perch, Qing fish, bream and green fish in aquaculture ponds are heavier than the sediments, while the river crab aquaculture ponds have heavy pollution. In aquaculture environment, both in water and in sediment, the resistance base is present, and the difference is at the maximum of 1~2 orders of magnitude. (3) in the same area, almost 90% of the aquaculture ponds were found in the same breeding period of the same breed. The highest relative abundance of the sulfonamides resistance gene appeared at 7-8 months in the middle period of the culture, and only a few exceptions.6, in the natural waters of Taihu, the sulfonamides resistant base in the water samples from different geographical locations and the sediments. There is a certain difference in relative abundance, the pollution of the sulfonamides resistance gene in the sediments is heavier than that in the water body. Compared with the aquaculture area near the same period, the pollution degree of the natural water samples is lighter than that in the aquaculture area, and the pollution in the sediments is equivalent to that in the aquaculture area. The non long-term fixed accumulation exists in the pond. The production of the sulfonamides resistance gene is related to the breeding cycle and the dosage of antibiotics in the culture. Therefore, it is necessary to strictly regulate and control the use of antibiotics in the culture process and to prevent the production and transmission of the antagonistic genes, thus making the resistant gene. Transfer is effectively curbed, providing protection for human health.
【學位授予單位】：南京農(nóng)業(yè)大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：S948
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