入侵紅火蟻防治用藥影響土壤生態環境之研究

The effect of red imported fire ant controlling insecticides on soil ecosystem

張靜淑(Ching-Shu Chang)

指導教授 ： 王一雄

共同指導教授 ： 顏瑞泓

國立臺灣大學/生物資源暨農學院/農業化學研究所/博士(2011年)

https://doi.org/10.6342/NTU.2011.00500

摘要

本研究旨在探討入侵紅火蟻（Red Imported Fire Ant, RIFA; Solenopsis invicta）防治用藥三種餌劑中的有效成分，包含百利普芬、芬普尼與賜諾殺施用於土壤環境中，經過長天數後之生物性降解與對土壤細菌族群結構之影響。
在添加不同濃度之藥劑的生物性降解結果顯示，對三種藥劑百利普芬、芬普尼與賜諾殺而言，當添加濃度為1 mg Kg-1時降解速率最快，其次為5 mg Kg-1添加，10 mg Kg-1添加時降解速率最慢；即藥劑濃度愈高，降解速率愈慢。
在不同溫度處理之生物性降解結果顯示，以添加10 mg Kg-1藥劑之處理為例，百利普芬在實驗初期，當溫度為40℃時之降解速率最快；實驗之第7天起，30℃處理之降解速度最快。芬普尼在實驗初期，不同溫度處理有相似之降解速率；實驗之第21天起，40℃處理之降解速度最快。賜諾殺在實驗全程中當溫度為30∼40℃時有較快之降解速率，特別是在實驗的第0∼21天。
在細菌族群結構的變遷方面，以聚合酶鏈鎖反應放大土壤細菌特定片段之DNA，結合變性梯度凝膠電泳（PCR-DGGE）的技術，再輔以Bio-Rad所發展之Quantity one軟體，並選用UPGMA法進行聚類分析，以進ㄧ步探討藥劑對土壤細菌族群結構之影響。添加10 mg Kg-1與1 mg Kg-1百利普芬在實驗早期菌相的相似度分別有59%與6 %。經過91天培養，10 mg Kg-1處理不同時期之細菌族群結構之相似度僅餘32%，（未添加藥劑之對照組仍有53%之相似度），在實驗過程中優勢的細菌族群包括Massilia albidiflava，與一些文獻上顯示無法在培養基中所培養的細菌（uncultured bacterium）。1 mg Kg-1處理經過91天培養與初期之相似度為57%，且細菌的族群數量較10 mg Kg-1處理豐富，顯示即便是低濃度百利普芬添加亦對土壤細菌族群造成影響。實驗全程中優勢的細菌族群包括Clostridium jejuense與尚有一些文獻上顯示無法在培養基中所培養的細菌；實驗晚期的優勢族群為Duganella sp. HF15。
添加10 mg Kg-1芬普尼於實驗第7天，細菌族群結構有明顯之改變，與其他時期相較僅有29%之相似度，顯示芬普尼對細菌族群的影響在實驗的早期即顯現。實驗全程中的優勢族群包括Acidobacteriacear bacterium Gsoil 1619, uncultured Acidobacterium sp., Methylobacterium sp. nac_9等；Rubrivivax gelatinosus是實驗第7天開始明顯存在之細菌族群；Nevskia soli是實驗後期之優勢細菌族群。添加1 mg Kg-1芬普尼經過91天培養，菌群間有55%之相似度。實驗全程中穩定的細菌族群包括uncultured gamma proteobacterium與一些uncultured bacterium；實驗初期的優勢族群有uncultured Chloroflexi bacterium；實驗第21天開始明顯存在之細菌族群有uncultured Methylobacillus sp., Acidobacteriaceae bacterium Gsoil 1619與uncultured Sinorhizobium sp.; 實驗初期與後期皆顯著的細菌族群有uncultured Pelobacter sp.。
添加10 mg Kg-1賜諾殺於實驗第7天起發現菌群結構發生變化，與其他培養天數相較相似度為46%；比較實驗第91天與第0天培養，相似度為71%，顯示經過長天數培養，實驗後期之菌群結構有恢復與初期相似之傾向。添加1 mg Kg-1 賜諾殺於實驗第0∼14天，菌相結構之相似度為66%。經過91天培養，細菌族群結構與實驗第0∼14天相較，仍有61%之相似度；Acidobacteriacear bacterium Gsoil 1619, uncultured Acidobacterium sp與uncultured Pseudomomadales bacterium，皆為實驗過程中穩定存在之細菌族群。添加50 mg Kg-1賜諾殺於實驗77∼91天與實驗0∼49天的相似度僅有44%。研究結果顯示：儘管賜諾殺是天然的發酵產物，但高濃度仍造成土壤細菌族群的影響。
結果顯示，添加百利普芬、芬普尼、賜諾殺於土壤環境中確實造成土壤細菌族群結構之改變，添加的濃度愈高所造成的影響愈劇，且其衝擊在實驗進行的早期即顯現。

關鍵字

百利普芬 ； 芬普尼 ； 賜諾殺 ； 聚合酶鏈鎖反應 ； 變性梯度凝膠電泳 ； 細菌族群結構

並列摘要

The objects of the study were to examine the dissipation rate of three kinds of ingredient in the baits for prevention and cure of the Red Imported Fire Ant (RIFA, Solenopsis invicta）as well as the change in the soil bacterial community. Residues of chemicals were measured using high performance liquid chromatography (HPLC) and the changes in bacterial community were determined by comparing the 16S rDNA bands on patterns by denaturing gradient gel electrophoresis (DGGE). The dissipation of three kinds of chemicals was affected by both the concentration applied and incubation temperature. Lower concentrations (1 mg Kg-1) and higher incubation temperatures (30 and 40ºC) showed more rapid dissipation rates.
The population of microbial community decreased rapidly after incubation with 10 mg Kg-1 of pyriproxyfen for 91 days, indicating the toxicity of pyriproxyfen toward bacterial communities in a closed soil ecosystem. Lower concentrations of chemicals showed less toxicity toward the microbial community. From cluster analysis, the structure of the bacterial community showed roughly a 53% similarity throughout the experiment period in the control experiment, indicating the relative stability within soil microbiota without chemical agitation. However, the similarity was lower than 38% both in the one and 10 mg Kg-1 of insecticide pyriproxyfen spiked experiment, indicating the soil bacterial community changed after the insecticide pyriproxyfen was applied. The analysis of the 16S rDNA sequence from soil total DNA according to the BCBI database showed the superiority of bacteria were including Massilia albidiflava and some kinds of uncultured bacteria in 10 mg Kg-1 treatment.
The effects of fipronil showed that the similarity of the bacterial community throughout the experiment period was 29% and 55% in 10 mg Kg-1 and 1 mg Kg-1of insecticide fipronil spiked experiment respectively. The analysis of the 16S rDNA sequence from soil total DNA according to the BCBI database showed the superiority of bacteria were including Acidobacteriacear bacterium Gsoil 1619, uncultured Acidobacterium sp., Methylobacterium sp.nac_9, Rubrivivax gelatinosus and Nevskia soli in 10 mg Kg-1 treatment.
The effects of spinosad showed that the similarity of the bacterial community throughout the experiment period was 46% and 44% in 10 mg Kg-1 and 50 mg Kg-1of insecticide spinosad spiked experiment respectively. The analysis of the 16S rDNA sequence according to the BCBI database showed the superiority of bacteria were including Ramlibacter sp. BXN5-27, Acidobacteriacear bacterium Gsoil 1619, Bacterium Ellin5078, uncultured gamma proteobacterium and some kinds of uncultured bacteria in 10 mg Kg-1 treatment.

並列關鍵字

Pyriproxyfen ； Fipronil ； Spinosad ； PCR ； DGGE ； bacterial community structure

參考文獻

Amann, R. I., W.Ludwig and K.-H. Schleifer. 1995. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbial. Rev. 59(1): 143-169.

Bobe, A., Coste C. M. and J. Copper. 1997. Factors Influencing the Adsorption of Fipronil on Soils. J. Agric. Food Chem. 45: 4861-4865.

Boivin, M. Y., B. Massieux, A. M. Breure, F. P. van den Ende, G. D. Greve, M. Rutgers and W. Admiraal. 2005. Effects of copper and temperature on aquatic bacterial communities. Aquat. Toxico. 71: 345-356.

Chiu. T. C., J. H. Yen, Y. N. Hsieh and Y. S. Wang. 2005. Reductive transformation of dieldrin under anaerobic sediment culture. Chemosphere 60: 1182-1189.

Cowan, D. A. 2000. Microbial genomes- the untapped resource. Trends in biotechnology. 18:14-16.