某集约化肉鸡饲养场PM_(2.5)中抗生素抗性基因的分布特征
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摘要
集约化家禽饲养场是抗生素抗性基因(ARGs)的重要来源,而PM_(2.5)作为ARGs可能向人体暴露的重要途径还未得到很好地研究.本文采集了集约化肉鸡饲养场舍内PM_(2.5)和粪便以及舍外PM_(2.5)样品,利用荧光定量PCR(q PCR)进行一类整合子(int I1)、总细菌(16S r DNA)和6类共19种ARGs丰度的检测.结果显示,除blaGES-1和blaSHV-1之外,其余17种ARGs在6类样品中均有检出.磺胺类、四环素类、大环内酯类和氨基糖苷类抗性基因在舍内粪便中丰度较高,达到1. 04×109～3. 27×1010copies·g-1,粪便是饲养场PM_(2.5)中ARGs的主要来源.舍内PM_(2.5)中以磺胺类和大环内酯类抗性基因丰度较高,分别为(8. 9±1. 9)×107copies·m-3和(5. 6±3. 1)×107copies·m-3,且舍内PM_(2.5)中ARGs丰度明显高于舍外. PM_(2.5)质量浓度与16S r DNA、int I1和ARGs丰度呈显著正相关,表明集约化饲养场中PM_(2.5)是空气传播细菌、ARGs和int I1的储存库和传播者. 6类样品中int I1丰度均高于ARGs,同时int I1和ARGs的共存关系表现出了多药耐药性的威胁,易对饲养人员和家禽健康及周边空气环境造成危害.
Concentrated poultry feeding operations are an important source of antibiotic resistance genes( ARGs). Little attention has been given to PM_(2.5) as a mechanism for exposing ARGs to humans. In this study,PM_(2.5) and fecal samples from inside the broiler feeding operation and PM_(2.5) samples from outside the broiler feeding operation were collected. All samples were subjected to the determination of class Ⅰ integrin( int I1),total bacterial gene( 16 S r DNA),and 19 ARGs of six types by quantitative real-time PCR( q PCR). The results indicated that,excluding blaGES-1 and blaSHV-1,the remaining 17 ARGs were detected in all six samples.Sulfonamide resistance genes,tetracycline resistance genes,macrolide resistance genes,and aminoglycoside resistance genes were abundant in the feces,reaching 1. 04 × 109-3. 27 × 1010 copies·g-1,while feces was an important source of antibiotic resistance genes in PM_(2.5) of the broiler feeding operation. There were high abundances of sulfonamide resistance genes and macrolide resistance genes in PM_(2.5) from inside the broiler feeding operation,reaching( 8. 9 ± 1. 9) × 107 copies·m-3 and( 5. 6 ± 3. 1) × 107 copies·m-3,respectively. The abundance of ARGs in the PM_(2.5) samples from inside the broiler feeding operation was significantly higher compared to the outside PM_(2.5) samples. There was a significant positive correlation between PM_(2.5) mass concentration and 16 S r DNA,int I1,and ARGs abundance,indicating that PM_(2.5) was the reservoir and disseminator of airborne bacteria,ARGs,and int I1 in the broiler feeding operation. The abundance of int I1 was higher than ARGs among all samples,and the co-occurring relationship between int I1 and ARGs demonstrates the threat of multi-drug resistance,which is harmful to the surrounding air environment and the health of the breeder and poultry.
Concentrated poultry feeding operations are an important source of antibiotic resistance genes( ARGs). Little attention has been given to PM_(2.5) as a mechanism for exposing ARGs to humans. In this study,PM_(2.5) and fecal samples from inside the broiler feeding operation and PM_(2.5) samples from outside the broiler feeding operation were collected. All samples were subjected to the determination of class Ⅰ integrin( int I1),total bacterial gene( 16 S r DNA),and 19 ARGs of six types by quantitative real-time PCR( q PCR). The results indicated that,excluding blaGES-1 and blaSHV-1,the remaining 17 ARGs were detected in all six samples.Sulfonamide resistance genes,tetracycline resistance genes,macrolide resistance genes,and aminoglycoside resistance genes were abundant in the feces,reaching 1. 04 × 109-3. 27 × 1010 copies·g-1,while feces was an important source of antibiotic resistance genes in PM_(2.5) of the broiler feeding operation. There were high abundances of sulfonamide resistance genes and macrolide resistance genes in PM_(2.5) from inside the broiler feeding operation,reaching( 8. 9 ± 1. 9) × 107 copies·m-3 and( 5. 6 ± 3. 1) × 107 copies·m-3,respectively. The abundance of ARGs in the PM_(2.5) samples from inside the broiler feeding operation was significantly higher compared to the outside PM_(2.5) samples. There was a significant positive correlation between PM_(2.5) mass concentration and 16 S r DNA,int I1,and ARGs abundance,indicating that PM_(2.5) was the reservoir and disseminator of airborne bacteria,ARGs,and int I1 in the broiler feeding operation. The abundance of int I1 was higher than ARGs among all samples,and the co-occurring relationship between int I1 and ARGs demonstrates the threat of multi-drug resistance,which is harmful to the surrounding air environment and the health of the breeder and poultry.
引文
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[11]高敏,仇天雷,秦玉成,等.养鸡场空气中抗性基因和条件致病菌污染特征[J].环境科学,2017,38(2):510-516.Gao M, Qiu T L, Qin Y C, et al. Sources and pollution characteristics of antibiotic resistance genes and conditional pathogenic bacteria in concentrated poultry feeding operations[J]. Environmental Science,2017,38(2):510-516.
[12] Sapkota A R,Ojo K K,Roberts M C,et al. Antibiotic resistance genes in multidrug-resistant Enterococcus spp. and Streptococcus spp. recovered from the indoor air of a large-scale swine-feeding operation[J]. Letters in Applied Microbiology,2006,43(5):534-540.
[13] Liu D J,Chai T J,Xia X Z,et al. Formation and transmission of Staphylococcus aureus(including MRSA)aerosols carrying antibiotic-resistant genes in a poultry farming environment[J].Science of the Total Environment,2012,426:139-145.
[14] Létourneau V,NehméB,Mériaux A,et al. Human pathogens and tetracycline-resistant bacteria in bioaerosols of swine confinement buildings and in nasal flora of hog producers[J].International Journal of Hygiene and Environmental Health,2010,213(6):444-449.
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[16] Xie J W,Jin L,Luo X S,et al. Seasonal disparities in airborne bacteria and associated antibiotic resistance genes in PM2. 5between urban and rural sites[J]. Environmental Science&Technology Letters,2018,5(2):74-79.
[17] Cao C,Jiang W J,Wang B Y,et al. Inhalable microorganisms in Beijing's PM2. 5and PM10pollutants during a severe smog event[J]. Environmental Science&Technology,2014,48(3):1499-1507.
[18] Mao D Q,Yu S,Rysz M,et al. Prevalence and proliferation of antibiotic resistance genes in two municipal wastewater treatment plants[J]. Water Research,2015,85:458-466.
[19] Tan L,Li L Y,Ashbolt N,et al. Arctic antibiotic resistance gene contamination, a result of anthropogenic activities and natural origin[J]. Science of the Total Environment,2018,621:1176-1184.
[20] Xiao K,Wang Y K,Wu G,et al. Spatiotemporal characteristics of air pollutants(PM10,PM2. 5,SO2,NO2,O3,and CO)in the inland basin city of Chengdu, Southwest China[J].Atmosphere,2018,9(2):74.
[21] Just N, Kirychuk S, Gilbert Y, et al. Bacterial diversity characterization of bioaerosols from cage-housed and floor-housed poultry operations[J]. Environmental Research,2011,111(4):492-498.
[22] Ye Z L,Li Q,Ma S S,et al. Summertime day-night differences of PM2. 5components(inorganic ions,OC,EC,WSOC,WSON,HULIS,and PAHs)in Changzhou,China[J]. Atmosphere,2017,8(10):189.
[23] Ling A L,Pace N R,Hernandez M T,et al. Tetracycline resistance and Class 1 integron genes associated with indoor and outdoor aerosols[J]. Environmental Science&Technology,2013,47(9):4046-4052.
[24] He L Y,Ying G G,Liu Y S,et al. Discharge of swine wastes risks water quality and food safety:antibiotics and antibiotic resistance genes from swine sources to the receiving environments[J]. Environment International,2016,92-93:210-219.
[25]靳璐滨,臧增亮,潘晓滨,等. PM2. 5和PM2. 5~10资料同化及在南京青奥会期间的应用试验[J].中国环境科学,2016,36(2):331-341.Jin L B,Zang Z L,Pan X B,et al. Data assimilation and application experiments of PM2. 5and PM2. 5~10during Nanjing Youth Olympic Games[J]. China Environmental Science,2016,36(2):331-341.
[26] Luby E M,Moorman T B,Soupir M L. Fate and transport of tylosin-resistant bacteria and macrolide resistance genes in artificially drained agricultural fields receiving swine manure[J].Science of the Total Environment,2016,550:1126-1133.
[27] Zhang Y J,Li H C,Gu J,et al. Effects of adding different surfactants on antibiotic resistance genes and intI1 during chicken manure composting[J]. Bioresource Technology,2016,219:545-551.
[28] Qian X,Sun W,Gu J,et al. Variable effects of oxytetracycline on antibiotic resistance gene abundance and the bacterial community during aerobic composting of cow manure[J]. Journal of Hazardous Materials,2016,315:61-69.
[29] Cheng W X,Chen H,Su C,et al. Abundance and persistence of antibiotic resistance genes in livestock farms:a comprehensive investigation in eastern China[J]. Environment International,2013,61:1-7.
[30] Gao M,Qiu T L,Sun Y M,et al. The abundance and diversity of antibiotic resistance genes in the atmospheric environment of composting plants[J]. Environment International,2018,116:229-238.
[31] WHO. WHO Director-general delivers lecture at Georgetown university's global futures initiative[EB/OL]. http://www.who. int/dg/speeches/2015/georgetown-university-lecture/en/,2015-09-30.
[2] Wang J,Ben W W,Yang M,et al. Dissemination of veterinary antibiotics and corresponding resistance genes from a concentrated swine feedlot along the waste treatment paths[J]. Environment International,2016,92-93:317-323.
[3]薛银刚,刘菲,王利平,等.气溶胶中抗生素抗性基因研究进展:以养殖场和医院为例[J].生态毒理学报,2017,12(6):27-37.Xue Y G,Liu F,Wang L P,et al. Progress in research on antibiotic resistance genes in aerosols:take farms and hospitals as examples[J]. Asian Journal of Ecotoxicology,2017,12(6):27-37.
[4] Wu N,Qiao M,Zhang B,et al. Abundance and diversity of tetracycline resistance genes in soils adjacent to representative swine feedlots in China[J]. Environmental Science&Technology,2010,44(18):6933-6939.
[5] Pal C,Bengtsson-Palme J,Kristiansson E,et al. The structure and diversity of human,animal and environmental resistomes[J]. Microbiome,2016,4:54.
[6]隋倩雯,张俊亚,魏源送,等.畜禽养殖过程抗生素使用与耐药病原菌及其抗性基因赋存的研究进展[J].生态毒理学报,2015,10(5):20-34.Sui Q W,Zhang J Y,Wei Y S,et al. Veterinary antibiotics use,occurrence of antibiotic resistance pathogen and its antibiotic resistance genes in animal production:an overview[J]. Asian Journal of Ecotoxicology,2015,10(5):20-34.
[7] Hong P Y,Li X Z,Yang X F,et al. Monitoring airborne biotic contaminants in the indoor environment of pig and poultry confinement buildings[J]. Environmental Microbiology,2012,14(6):1420-1431.
[8] The Guardian. Rise of mega farms:how the US model of intensive farming is invading the world[EB/OL]. https://www.theguardian. com/environment/2017/jul/18/rise-of-mega-farmshow-the-us-model-of-intensive-farming-is-invading-the-world?CMP=Share_AndroidApp_Tweet,2017-07-18.
[9] Gao M,Jia R Z,Qiu T L,et al. Size-related bacterial diversity and tetracycline resistance gene abundance in the air of concentrated poultry feeding operations[J]. Environmental Pollution,2017,220:1342-1348.
[10] Mc Eachran A D, Blackwell B R, Hanson J D, et al.Antibiotics, bacteria, and antibiotic resistance genes:aerial transport from cattle feed yards via particulate matter[J].Environmental Health Perspectives, 2015, 123(4):337-343.
[11]高敏,仇天雷,秦玉成,等.养鸡场空气中抗性基因和条件致病菌污染特征[J].环境科学,2017,38(2):510-516.Gao M, Qiu T L, Qin Y C, et al. Sources and pollution characteristics of antibiotic resistance genes and conditional pathogenic bacteria in concentrated poultry feeding operations[J]. Environmental Science,2017,38(2):510-516.
[12] Sapkota A R,Ojo K K,Roberts M C,et al. Antibiotic resistance genes in multidrug-resistant Enterococcus spp. and Streptococcus spp. recovered from the indoor air of a large-scale swine-feeding operation[J]. Letters in Applied Microbiology,2006,43(5):534-540.
[13] Liu D J,Chai T J,Xia X Z,et al. Formation and transmission of Staphylococcus aureus(including MRSA)aerosols carrying antibiotic-resistant genes in a poultry farming environment[J].Science of the Total Environment,2012,426:139-145.
[14] Létourneau V,NehméB,Mériaux A,et al. Human pathogens and tetracycline-resistant bacteria in bioaerosols of swine confinement buildings and in nasal flora of hog producers[J].International Journal of Hygiene and Environmental Health,2010,213(6):444-449.
[15] Just N A,Létourneau V,Kirychuk S P,et al. Potentially pathogenic bacteria and antimicrobial resistance in bioaerosols from cage-housed and floor-housed poultry operations[J]. The Annals of Occupational Hygiene,2012,56(4):440-449.
[16] Xie J W,Jin L,Luo X S,et al. Seasonal disparities in airborne bacteria and associated antibiotic resistance genes in PM2. 5between urban and rural sites[J]. Environmental Science&Technology Letters,2018,5(2):74-79.
[17] Cao C,Jiang W J,Wang B Y,et al. Inhalable microorganisms in Beijing's PM2. 5and PM10pollutants during a severe smog event[J]. Environmental Science&Technology,2014,48(3):1499-1507.
[18] Mao D Q,Yu S,Rysz M,et al. Prevalence and proliferation of antibiotic resistance genes in two municipal wastewater treatment plants[J]. Water Research,2015,85:458-466.
[19] Tan L,Li L Y,Ashbolt N,et al. Arctic antibiotic resistance gene contamination, a result of anthropogenic activities and natural origin[J]. Science of the Total Environment,2018,621:1176-1184.
[20] Xiao K,Wang Y K,Wu G,et al. Spatiotemporal characteristics of air pollutants(PM10,PM2. 5,SO2,NO2,O3,and CO)in the inland basin city of Chengdu, Southwest China[J].Atmosphere,2018,9(2):74.
[21] Just N, Kirychuk S, Gilbert Y, et al. Bacterial diversity characterization of bioaerosols from cage-housed and floor-housed poultry operations[J]. Environmental Research,2011,111(4):492-498.
[22] Ye Z L,Li Q,Ma S S,et al. Summertime day-night differences of PM2. 5components(inorganic ions,OC,EC,WSOC,WSON,HULIS,and PAHs)in Changzhou,China[J]. Atmosphere,2017,8(10):189.
[23] Ling A L,Pace N R,Hernandez M T,et al. Tetracycline resistance and Class 1 integron genes associated with indoor and outdoor aerosols[J]. Environmental Science&Technology,2013,47(9):4046-4052.
[24] He L Y,Ying G G,Liu Y S,et al. Discharge of swine wastes risks water quality and food safety:antibiotics and antibiotic resistance genes from swine sources to the receiving environments[J]. Environment International,2016,92-93:210-219.
[25]靳璐滨,臧增亮,潘晓滨,等. PM2. 5和PM2. 5~10资料同化及在南京青奥会期间的应用试验[J].中国环境科学,2016,36(2):331-341.Jin L B,Zang Z L,Pan X B,et al. Data assimilation and application experiments of PM2. 5and PM2. 5~10during Nanjing Youth Olympic Games[J]. China Environmental Science,2016,36(2):331-341.
[26] Luby E M,Moorman T B,Soupir M L. Fate and transport of tylosin-resistant bacteria and macrolide resistance genes in artificially drained agricultural fields receiving swine manure[J].Science of the Total Environment,2016,550:1126-1133.
[27] Zhang Y J,Li H C,Gu J,et al. Effects of adding different surfactants on antibiotic resistance genes and intI1 during chicken manure composting[J]. Bioresource Technology,2016,219:545-551.
[28] Qian X,Sun W,Gu J,et al. Variable effects of oxytetracycline on antibiotic resistance gene abundance and the bacterial community during aerobic composting of cow manure[J]. Journal of Hazardous Materials,2016,315:61-69.
[29] Cheng W X,Chen H,Su C,et al. Abundance and persistence of antibiotic resistance genes in livestock farms:a comprehensive investigation in eastern China[J]. Environment International,2013,61:1-7.
[30] Gao M,Qiu T L,Sun Y M,et al. The abundance and diversity of antibiotic resistance genes in the atmospheric environment of composting plants[J]. Environment International,2018,116:229-238.
[31] WHO. WHO Director-general delivers lecture at Georgetown university's global futures initiative[EB/OL]. http://www.who. int/dg/speeches/2015/georgetown-university-lecture/en/,2015-09-30.