Antibiotic resistance and genetic diversity of Escherichia coli isolates from traditional and integrated aquaculture in South China

This study investigated antibiotic resistance profiles including antibiotic resistance frequencies, resistance genes and resistance patterns in Escherichia coli strains isolated from traditional and integrated aquaculture systems in South China by using antibiotic susceptibility testing and real time polymerase chain reaction (PCR) technique. The E. coli isolates were found to be resistant to at least one antibiotic among 12 antibiotics. Higher resistance frequencies to ampicillin, sulfamethoxazole, trimethoprime, streptomycin and tetracycline were found compared to the rest antibiotics. Among the 10 tetracycline resistance genes detected in the resistant isolates, the most prevalent tetracycline resistance genes were tetA, tetW and tetB with the frequency of 69.7%, 63.5% and 21.9%, respectively. Three sulfonamide resistance genes were detected in these resistant isolates, with their detection frequencies in the following order: sul2 (55.3%) > sul3 (28.2%) > sul1 (6.2%). Four resistance genes mainly encoding extended-spectrum β-lactamases (ESBLs) were detected in these resistant isolates, with the detection frequencies of blaTEM (28.4%) > blaOXA (9.7%) > blaCTX (9.3%) > blaCARB (5.2%) > blaSHV (0.0%). It was found that the integrated aquaculture system exhibited generally higher prevalence of antibiotic resistance than the traditional aquaculture system. An integrated aquaculture system could facilitate development of bacterial resistance and spread of the antibiotic resistance genes, and consequently become an important reservoir of resistance genes.     

Occurrence and abundance of tetracycline,sulfonamide resistance genes,and class 1 integron in five wastewater treatment plants

To understand the transport and fate of antibiotic resistance genes in wastewater treatment plants, 12 resistance genes (ten tetracycline resistance genes, two sulfonamides genes) and class 1 integron gene (intI1) were studied in five wastewater treatment plants with different treatment processes and different sewage sources. Among these resistance genes, sulfonamides genes (sul1 and sul2) were of the most prevalent genes with detection frequency of 100 %. The effluent water contained fewer types of resistance genes than the influent in most selected plants. The abundance of five quantified resistance genes (tetG, tetW, tetX, sul1, and intI1) decreased in effluent of plants treating domestic or industrial wastewater with anaerobic/aerobic or membrane bioreactor (MBR) technologies, but tetG, tetX, sul1, and intI1 increased along the treatment units of plants treating vitamin C production wastewater by anaerobic/aerobic technology. In plant treating cephalosporins production wastewater by UASB/aerobic process, the quantities of tetG, tetX, and sul1 first decreased in anaerobic effluent water but then increased in aerobic effluent water.     

Detection of antibiotic resistance and tetracycline resistance genes in Enterobacteriaceae isolated from the Pearl rivers in South China

This study investigated antibiotic resistance profiles and tetracycline resistance genes in Enterobacteriaceae family isolates from the Pearl rivers. The Enterobacteriaceae isolates were tested for susceptibility to seven antibiotics ampicillin, chloramphenicol, ciprofloxacin, levofloxacin, sulphamethoxazole/trimethoprim, tetracycline and trimethoprim. In Liuxi reservoir, with an exception to ampicillin resistant strains (11%) no other antibiotic resistance bacterial strains were detected. However, multiple drug resistance in bacterial isolates from the other sites of Pearl rivers was observed which is possibly due to sewage discharge and input from other anthropogenic sources along the rivers. Four tetracycline resistance genes tet A, tet B, tet C and tet D were detected in the isolates from the rivers. The genes tet A and tet B were widely detected with the detection frequencies of 43% and 40% respectively. Ciprofloxacin and levofloxacin resistant enteric bacteria were also isolated from the pig and duck manures which suggest a wider distribution of human specific drugs in the environment. This investigation provided a baseline data on antibiotic resistance profiles and tetracycline resistance genes in the Pearl rivers delta.     

Human health implications of clinically relevant bacteria in wastewater habitats

The objective of this review is to reflect on the multiple roles of bacteria in wastewater habitats with particular emphasis on their harmful potential for human health. Indigenous bacteria promote a series of biochemical and metabolic transformations indispensable to achieve wastewater treatment. Some of these bacteria may be pathogenic or harbour antibiotic resistance or virulence genes harmful for human health. Several chemical contaminants (heavy metals, disinfectants and antibiotics) may select these bacteria or their genes. Worldwide studies show that treated wastewater contain antibiotic resistant bacteria or genes encoding virulence or antimicrobial resistance, evidencing that treatment processes may fail to remove efficiently these bio-pollutants. The contamination of the surrounding environment, such as rivers or lakes receiving such effluents, is also documented in several studies. The current state of the art suggests that only some of antibiotic resistance and virulence potential in wastewater is known. Moreover, wastewater habitats may favour the evolution and dissemination of new resistance and virulence genes and the emergence of new pathogens. For these reasons, additional research is needed in order to obtain a more detailed assessment of the long-term effects of wastewater discharges. In particular, it is important to measure the human and environmental health risks associated with wastewater reuse.     

Exploring antibiotic resistance genes,mobile gene elements,and virulence gene factors in an urban freshwater samples using metagenomic analysis

Environmental Science and Pollution Research - Antibiotic resistance genes (ARGs) and antimicrobial resistance elements (AMR) are novel environmental contaminants that pose a significant risk to...     

Environmental pollution by antibiotics and by antibiotic resistance determinants

Antibiotics are among the most successful drugs used for human therapy. However, since they can challenge microbial populations, they must be considered as important pollutants as well. Besides being used for human therapy, antibiotics are extensively used for animal farming and for agricultural purposes. Residues from human environments and from farms may contain antibiotics and antibiotic resistance genes that can contaminate natural environments. The clearest consequence of antibiotic release in natural environments is the selection of resistant bacteria. The same resistance genes found at clinical settings are currently disseminated among pristine ecosystems without any record of antibiotic contamination. Nevertheless, the effect of antibiotics on the biosphere is wider than this and can impact the structure and activity of environmental microbiota. Along the article, we review the impact that pollution by antibiotics or by antibiotic resistance genes may have for both human health and for the evolution of environmental microbial populations.     

The treatment of wastewater containing pharmaceuticals in microcosm constructed wetlands: the occurrence of integrons (int1–2) and associated resistance genes (sul1–3, qacEΔ1)

Environmental Science and Pollution Research - The aim of this study was to analyze the occurrence of sulfonamide resistance genes (sul1–3) and other genetic elements as antiseptic resistance...     

Heavy metal resistance and genotypic analysis of metal resistance genes in gram-positive and gram-negative bacteria present in Ni-rich serpentine soil and in the rhizosphere of Alyssum murale

Forty-six bacterial cultures, including one culture collection strain, thirty from the rhizosphere of Alyssum murale and fifteen from Ni-rich soil, were tested for their ability to tolerate arsenate, cadmium, chromium, zinc, mercury, lead, cobalt, copper, and nickel in their growth medium. The resistance patterns, expressed as minimum inhibitory concentrations, for all cultures to the nine different metal ions were surveyed by using the agar dilution method. A large number of the cultures were resistant to Ni (100%), Pb (100%), Zn (100%), Cu (98%), and Co (93%). However, 82, 71, 58 and 47% were sensitive to As, Hg, Cd and Cr(VI), respectively. All cultures had multiple metal-resistant, with heptametal resistance as the major pattern (28.8%). Five of the cultures (about of 11.2% of the total), specifically Arthrobacter rhombi AY509239, Clavibacter xyli AY509235, Microbacterium arabinogalactanolyticum AY509226, Rhizobium mongolense AY509209 and Variovorax paradoxus AY512828 were tolerant to nine different metals. The polymerase chain reaction in combination with DNA sequence analysis was used to investigate the genetic mechanism responsible for the metal resistance in some of these gram-positive and gram-negative bacteria that were, highly resistant to Hg, Zn, Cr and Ni. The czc, chr, ncc and mer genes that are responsible for resistance to Zn, Cr, Ni and Hg, respectively, were shown to be present in these bacteria by using PCR. In the case of, M. arabinogalactanolyticum AY509226 these genes were shown to have high homology to the czcD, chrB, nccA, and mer genes of Ralstonia metallidurans CH34. Therefore, Hg, Zn, Cr and Ni resistance genes are widely distributed in both gram-positive and gram-negative isolates obtained from A. murale rhizosphere and Ni-rich soils.     

A comparison of antibiotics,antibiotic resistance genes,and bacterial community in broiler and layer manure following composting

Animal manure is an important source of antibiotics and antibiotic resistance genes (ARGs) in the environment. However, the difference of antibiotic residues and ARG profiles in layer and broiler manure as well as their compost remains unexplored. In this study, we investigated the profiles of twelve antibiotics, seventeen ARGs, and class 1 integrase gene (intI1) in layer and broiler manure, and the corresponding compost at large-scale. Compared with layer manure, broiler manure exhibited approximately six times more residual tetracyclines, especially chlortetracycline. The relative abundances of qnrS and ermA genes in broiler manure were significantly higher than those in layer manure. The concentration of tetracyclines not only had a significantly positive correlation with tetracycline resistance genes (tetA and tetC) but was also positively correlated with quinolone resistance (qepA, qnrB, and qnrS) and macrolide resistance (ermA and ermT). Most ARGs in manure were reduced after composting. However, the relative abundance of sulfonamide resistance gene sul1 increased up to 2.41% after composting, which was significantly higher than that of broiler (0.41%) and layer (0.62%) manure. The associated bacterial community was characterized by high-throughput 16S rRNA gene sequencing. The relative abundances of thermophilic bacteria had significant positive correlations with the abundance of sul1 in compost. The composting has a significant impact on the ARG-associated gut microbes in poultry manure. Variation partitioning analysis indicated that the change of bacterial community compositions and antibiotics contributed partially to the shift in ARG profiles. The results indicate that at industry-scale production broiler manure had more antibiotics and ARGs than layer manure did, and composting decreased most ARG abundances in poultry manure except for sulfonamide resistance genes.

     

Commercially available garden products as important sources of antibiotic resistance genes—a survey

Environmental Science and Pollution Research - The dissemination of antibiotic resistance genes (ARGs) in the environment contributes to the global rise in antibiotic resistant infections....     

Monitoring and evaluation of antibiotic resistance genes in three rivers in northeast China

Environmental Science and Pollution Research - Antibiotic resistance genes (ARGs) have become an important public health problem. In this study, we used metagenomic sequencing to analyze the...     

Increase of antibiotic resistance genes via horizontal transfer in single- and two-chamber microbial electrolysis cells

Environmental Science and Pollution Research - Microbial electrolysis cells (MECs) have been applied for antibiotic degradation but simultaneously induced antibiotic resistance genes (ARGs), thus...     

Diversity of antibiotic resistance genes and encoding ribosomal protection proteins gene in livestock waste polluted environment

The rapid development and increase of antibiotic resistance are global phenomena resulting from the extensive use of antibiotics in human clinics and animal feeding operations. Antibiotics can promote the occurrence of antibiotic resistance genes (ARGs), which can be transferred horizontally to humans and animals through water and the food chain. In this study, the presence and abundance of ARGs in livestock waste was monitored by quantitative PCR. A diverse set of bacteria and tetracycline resistance genes encoding ribosomal protection proteins (RPPs) from three livestock farms and a river were analyzed through denaturing gradient gel electrophoresis (DGGE). The abundance of sul(I) was 10³ to 10⁵ orders of magnitude higher than that of sul(II). Among 11 tet-ARGs, the most abundant was tet(O). The results regarding bacterial diversity indicated that the presence of antibiotics might have an evident impact on bacterial diversity at every site, particularly at the investigated swine producer. The effect of livestock waste on the bacterial diversity of soil was stronger than that of water. Furthermore, a sequencing analysis showed that tet(M) exhibited two genotypes, while the other RPPs-encoding genes exhibited at least three genotypes. This study showed that various ARGs and RPPs-encoding genes are particularly widespread among livestock.     

人工湿地中抗生素抗性大肠杆菌和抗性基因的去除与分布

抗生素的滥用导致抗生素抗性菌和抗性基因随生活污水和养殖废水的排放在环境中肆意散播,其去除及环境行为越来越受到关注。采用K-B纸片法测定了9套不同工艺构型模拟人工湿地中大肠杆菌对7种抗生素的抗性率,并应用多重PCR检测磺胺类sul1、2、3与四环素tetA、B、C、D抗性基因,探究人工湿地对抗性菌的去除效率及抗性菌、抗性基因的分布规律。结果显示,人工湿地能有效去除污水中70%左右的抗性大肠杆菌,有效降低了细菌抗性的传播风险;共计分离出535株大肠肝菌中有378株对一种以上抗生素有抗性性,以四环素、磺胺类和氨苄西林抗性率最高,达到25%以上,其他4种抗性率较低,不足20%;2种抗性基因的检出率都在70%以上;对不同采样点大肠杆菌的抗性性及抗性基因的比较发现,各部分大肠杆菌的抗性水平、多重抗性指数(MRI)以及抗性基因(sul、tet)检出率和组合数表现出:基质≥出水>进水,推测抗性菌被湿地基质截留,在基质生物膜上发生抗性基因的重组,并释放抗性菌,提高了出水中抗性水平和抗性基因检出率。     

Detecting antibiotic resistance genes and human potential pathogenic Bacteria in fishmeal by culture-independent method

Environmental Science and Pollution Research - Fishmeal is a fundamental ingredient of feedstuffs and is used globally in aquaculture. However, there are few data on the antibiotic resistance genes...     

Antibiotic resistance and virulence factors of Escherichia coli from eagles and goshawks

One of the major global problems in medicine is microbial resistance to antibiotics (antimicrobial resistance) and this has become an increasingly frequent research topic. This study focuses on antimicrobial resistance, phylogenetic and genetic characterization of Escherichia coli from wild birds: ten isolates from eagles (Aquila chrysaetos), nine from goshawks (Accipiter gentilis) and 24 from broilers in the Slovak Republic. Twenty-two strains with presence of int1 gene were selected and examined for the presence or absence of transposon gene (tn3), genes of antibiotic resistance and virulence factors. We detected sequence type (ST) in eagles ST 442 with genes iss, papC, iutA, cvaC, tsh, fyuA, iroN, kps, feoB, sitA, irp2, ireA for virulence factors and tetA, sul1, sul2, dfrA, aadA for antibiotic resistance; in goshawks ST 1011 with iss, papC, fyuA, iroN, feoB, sitA and qnrS1, tetA, sul1, sul2, dfrA, aadA, respectively. These ST types have been found in humans too and should be evaluated further for possible zoonotic potential and transfer of resistance genes from the environment.     

Occurrence and distribution of veterinary antibiotics and tetracycline resistance genes in farmland soils around swine feedlots in Fujian Province, China

Six antibiotics, tetracyclines (TCs), and quinolones (QNs) in farmland soils from four coastal cities in Fujian Province of China were investigated. Oxytetracycline was most frequently detected, followed by enrofloxacin, ciprofloxacin, chlorotetracycline, ofloxacin, and tetracycline, with maximum concentrations of 613.2, 637.3, 237.3, 2668.9, 205.7, and 189.8 μg kg^?1, respectively. Samples from Putian City contained the highest maximum concentration of ∑TCs (3,064.2 μg kg^?1), whereas those from Fuzhou City contained the highest maximum concentration of ∑QNs (897.8 μg kg^?1). It is noteworthy that the ∑TCs and ∑QNs in 46.4 and 28.6 % of samples exceeded the ecotoxic effect trigger value (100 μg kg^?1), respectively. The concentrations of these antibiotics and five tetracycline resistance genes in four soil plots at depth profiles were quantified thereafter. In most cases, both antibiotics and resistance genes decreased with the increase of depth. Some antibiotics can be detected at a depth of 60–80 cm where the abundance of tetO, tetM, and tetX reached up to 10⁷ copies g^?1. Additionally, the sum of all tet genes (normalized to 16S rRNA genes) correlated with ∑TCs significantly (r?=?0.676). Our results suggest that resistance determinants can migrate to deeper soil layers and would probably contaminate groundwater by vertical transport.     

Investigation on gene transfer from genetically modified corn (Zea mays L.) plants to soil bacteria

Knowledge about the prevalence and diversity of antibiotic resistance genes in soil bacteria communities is required to evaluate the possibility and ecological consequences of the transfer of these genes carried by genetically modified (GM) plants to soil bacteria. The neomycin phosphotransferase gene (nptII) conferring resistance to kanamycin and neomycin is one of the antibiotic resistance genes commonly present in GM plants. In this study, we investigated kanamycin-resistant (Km(R)) and neomycin-resistant (Nm(R)) soil bacterial populations in a 3-year field trial using a commercial GM corn (Zea mays L.) carrying the nptII gene and its near isogenic line. The results showed that a portion (2.3 - 15.6 %) of cultivable soil bacteria was naturally resistant to kanamycin or neomycin. However, no significant difference in the population level of Km(R) or Nm(R) soil bacteria was observed between the GM and non-GM corn fields. The nptII gene was not detected in any of the total 3000 Km(R) or Nm(R) isolates screened by PCR. Further, total soil bacterial cells were collected through Nycodenz gradient centrifugation and bacterial community DNA was subjected to PCR. Detection limit was about 500 cells per gram of fresh soil. Our study suggests that the nptII gene was relatively rare in the soil bacterial populations and there was no evidence of gene transfer from a GM corn plant to soil bacteria based on the data from total soil bacterial communities.     

Tetracycline resistance in semi-arid agricultural soils under long-term swine effluent application

Annually, millions pounds of antibiotics are released unmetabolized into environment along with animal wastes. Accumulation of antibiotics in soils could potentially induce the persistence of antibiotic resistant bacteria. Antibiotics such as tetracyclines and tetracycline-resistant bacteria have been previously detected in fields fertilized with animal manure. However, little is known about the accumulation of tetracyclines and the development of tetracycline resistance in semi-arid soils. Here we demonstrate that continuous land application with swine effluent, containing trace amounts of chlortetracycline, does not necessarily induce tetracycline resistance in soil bacteria. Based on the testing of more than 3,000 bacteria isolated from the amended soils, we found no significant increase in the occurrence and level of chlortetracycline resistant bacteria in soils after 15 years of continuous swine effluent fertilization. To account for a possible transfer of tetracycline-resistant bacteria originated from the swine effluent to soils, we analyzed two commonly found tetracycline resistant genes, tet(O) and tet(M), in the swine effluent and fertilized soils. Both genes were present in the swine effluent, however, they were not detectable in soils applied with swine effluent. Our data demonstrate that agronomic application of manure from antibiotic treated swine effluent does not necessarily result in the development of antibiotic bacterial resistance in soils. Apparently, concentrations of chlortetracycline present in manure are not significant enough to induce the development of antibiotic bacterial resistance.     

The platelet-activating factor acetylhydrolase gene derived from Trichoderma harzianum induces maize resistance to Curvularia lunata through the jasmonic acid signaling pathway

Platelet-activating factor acetylhydrolase (PAF-AH) derived from Trichoderma harzianum was upregulated by the interaction of T. harzianum with maize roots or the foliar pathogen Curvularia lunata. PAF-AH was associated with chitinase and cellulase expressions, but especially with chitinase, because its activity in the KO40 transformant (PAF-AH disruption transformant) was lower, compared with the wild-type strain T28. The result demonstrated that the colonization of maize roots by T. harzianum induced systemic protection of leaves inoculated with C. lunata. Such protection was associated with the expression of inducible jasmonic acid pathway-related genes. Moreover, the data from liquid chromatography-mass spectrometry confirmed that the concentration of jasmonic acid in maize leaves was associated with the expression level of defense-related genes, suggesting that PAF-AH induced resistance to the foliar pathogen. Our findings showed that PAF-AH had an important function in inducing systemic resistance to maize leaf spot pathogen.