Bacillus licheniformis-fermented products and enramycin differentially modulate microbiota and antibiotic resistome in the cecal digesta of broilers. - Related Documents




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353501.0000Bacillus licheniformis-fermented products and enramycin differentially modulate microbiota and antibiotic resistome in the cecal digesta of broilers. Since antibiotic resistance is a global health issues, the use of antibiotics in animal feed for growth promotion has been restricted in many countries. Bacillus licheniformis probiotic is a potential alternative to antibiotics for increasing poultry performance. Through metagenomic sequencing, this study investigated the effects of B. licheniformis-fermented products (BLFPs) and enramycin on the microbial community composition and antibiotic resistance gene (ARG) distribution in the cecal digesta of broilers at the age of 35 d. In total, 144 one-day-old male broiler chicks (Ross 308) were randomly assigned to 4 dietary treatments as follows: basal diet (control [C] group), basal diet plus 10 mg/kg enramycin (E group), basal diet plus 1 g/kg BLFPs (L group), and basal diet plus 3 g/kg BLFPs (H group), with 6 replicate cages per treatment group and 6 birds per cage. The results indicated that the cecal alpha diversity (richness and evenness) of bacterial species was higher in the H group than in the C group. Principal coordinate analysis of microbiota and the ARG composition indicated clear differences among the cecal samples of the groups. In the cecal digesta, the abundance of active bacteria associated with probiotic properties, such as Lactobacillus crispatus and Akkermansia muciniphila, was higher in the H group than in the other groups. Enramycin treatment promoted the expression of peptide (bcrA), glycopeptide (vanRI), and lincosamide (lsaE) resistance genes but inhibited the expression of aminocoumarin (parY) and pleuromutilin (TaeA) resistance genes. BLFP (1 and 3 g/kg) treatment suppressed the expression of aminoglycoside (ANT(6)-Ib), streptogramin (vatB), and peptide (ugd) resistance genes but enhanced the expression of macrolide (efrA) and aminocoumarin (novA) resistance genes. The abundance of peptide resistance genes in Bacteroides spp. was lower in the H group than in the C group. The abundance of lincosamide resistance genes in Lactobacillus spp. was higher in the E group than in the other groups. These results demonstrated that differential changes in the structure of 3 g/kg BLFPs and enramycin-induced cecal microbial communities accompany changes in the abundance of bacterial hosts carrying specific ARGs in the cecal microbiota of broilers.202235841645
741710.9995Limited impacts of high doses of dietary copper on the gut bacterial metal resistome explain negligible co-selection of antibiotic resistance. High dietary intake of Cu has previously been linked to the selection of Cu resistance and co-selection of antibiotic resistance in specific gut bacteria. Based on a novel HT-qPCR metal resistance gene chip as combined with 16S rRNA gene amplicon sequencing and phenotypic resistance typing of Escherichia coli isolates, we here report the impacts of two contrasting Cu-based feed additives on the swine gut bacterial metal resistome and community assembly. DNA was extracted from fecal samples (n = 80) collected at day 26 and 116 of the experiment from 200 pigs allotted to five dietary treatments: negative control (NC) diet with 20 μg CuSO(4) g(-1) and four diets added 125 or 250 μg CuSO(4) g(-1) feed or 125 or 250 μg Cu(2)O g(-1) feed to the NC diet. Dietary Cu supplementation reduced the relative abundance of Lactobacillus, but it had negligible impacts on bacterial community composition relative to the gut microbiome maturation effect (time). The relative importance of different bacterial community assembly processes was not markedly affected by the dietary Cu treatments, and differences in swine gut metal resistome composition could be explained primarily by differences in bacterial community composition rather than by dietary Cu treatments. High dietary Cu intake (250 μg Cu g(-1)) selected for phenotypic Cu resistance in E. coli isolates, but surprisingly it did not result in increased prevalence of the Cu resistance genes targeted by the HT-qPCR chip. In conclusion, the lacking impacts of dietary Cu on the gut bacterial metal resistome explain results from a previous study showing that even high therapeutic doses of dietary Cu did not cause co-selection of antibiotic resistance genes and mobile genetic elements known to harbor these genes.202337201857
719720.9994The response of copper resistance genes, antibiotic resistance genes, and intl1/2 to copper addition during anaerobic digestion in laboratory. Heavy metal pollution can serve as a selective pressure for antibiotic resistance genes in polluted environments. Anaerobic fermentation, as a recommended wastewater treatment method, is an effective mitigation measure of antibiotic resistance diffusion. To explore the influence of copper on anaerobic fermentation, we exposed the fermentation substrate to copper in a laboratory setup. We found that the relative abundance of 8 genes (pcoD, tetT, tetA, tetB, tetO, qnrS, ermA and ermB) increased at the late stage of fermentation and their abundance was linked to copper content. Corynebacterium and Streptococcus were significantly positively correlated with ermA, ermB, tetA and tetB (P < 0.05). The relative abundance of tetT was significantly positively correlated with Terrisporobacter, Clostridium_sensu_stricto_1 and Turicibacter (P < 0.05). We screened 90 strains of copper resistant bacteria from blank, medium and high copper test groups on days 25, 31 and 37. The number of fragments carried by a single strain increased with time while intl1, ermA and ermB existed in almost all combinations of the multiple fragments we identified. The relative abundance of these three genes were linearly correlated with Corynebacterium and Streptococcus. The antibiotic resistance genes carried by class 1 integrons gradually increased with time in the fermentation system and integrons carrying ermA and ermB most likely contributed to host survival through the late stages of fermentation. The genera Corynebacterium and Streptococcus may be the primary carriers of such integrated mobile gene element and this was most likely the reason for their rebound in relative abundance during the late fermentation stages.202133418156
712530.9994Persistence of resistance to erythromycin and tetracycline in swine manure during simulated composting and lagoon treatments. The use of antimicrobials in food animal production leads to the development of antimicrobial resistance (AMR), and animal manure constitutes the largest reservoir of such AMR. In previous studies, composted swine manure was found to contain substantially lower abundance of AMR genes that encode resistance to tetracyclines (tet genes) and macrolide-lincosamide-streptogramin B (MLS(B)) superfamily (erm genes), than manures that were treated by lagoons or biofilters. In this study, temporal changes in AMR carried by both cultivated and uncultivated bacteria present in swine manure during simulated composting and lagoon storage were analyzed. Treatments were designed to simulate the environmental conditions of composting (55°C with modest aeration) and lagoon storage (ambient temperature with modest aeration). As determined by selective plate counting, over a 48-day period, cultivated aerobic heterotrophic erythromycin-resistant bacteria and tetracycline-resistant bacteria decreased by more than 4 and 7 logs, respectively, in the simulated composting treatment while only 1 to 2 logs for both resistant bacterial groups in the simulated lagoon treatment. Among six classes each of erm and tet genes quantified by class-specific real-time PCR assays, the abundance of erm(A), erm(C), erm(F), erm(T), erm(X), tet(G), tet(M), tet(O), tet(T), and tet(W) declined marginally during the first 17 days, but dramatically thereafter within 31 days of the composting treatment. No appreciable reduction of any of the erm or tet genes analyzed was observed during the simulated lagoon treatment. Correlation analysis showed that most of the AMR gene classes had similar persistence pattern over the course of the treatments, though not all AMR genes were destructed at the same rate during the treatments.201221811793
712840.9994Composting of chicken litter from commercial broiler farms reduces the abundance of viable enteric bacteria, Firmicutes, and selected antibiotic resistance genes. We examined the ability of composting to remove ARGs and enteric bacteria in litter obtained from broiler chickens fed with a diet supplemented with Bacitracin methylene disalicylate (BDM) (conventional chicken litter), or an antibiotic-free diet (raised without antibiotic (RWA) chicken litter). This was done by evaluating the litter before and after composting for the abundance of ten gene targets associated with antibiotic resistance or horizontal gene transfer, the composition of the bacterial communities, and the abundance of viable enteric bacteria. The abundance of gene targets was determined by qPCR and the microbial community composition of chicken litter determined by 16S rRNA gene amplicon sequencing. Enteric bacteria were enumerated by viable plate count. A majority of the gene targets were more abundant in conventional than in RWA litter. In both litter types, the absolute abundance of all of the target genes decreased after composting except sul1, intI1, incW and erm(F) that remained stable. Composting significantly reduced the abundance of enteric bacteria, including those carrying antibiotic resistance. The major difference in bacterial community composition between conventional and RWA litter was due to members affiliated to the genus Pseudomonas, which were 28% more abundant in conventional than in RWA litter. Composting favoured the presence of thermophilic bacteria, such as those affiliated with the genus Truepera, but decreased the abundance of those bacterial genera associated with cold-adapted species, such as Carnobacterium, Psychrobacter and Oceanisphaera. The present study shows that chicken litter from broilers fed with a diet supplemented with antibiotic has an increased abundance of some ARGs, even after composting. However, we can conclude that fertilization with composted litter represents a reduced risk of transmission of antibiotic resistance genes and enteric bacteria of poultry origin to soil and crops than will fertilization with raw litter.202032768779
712450.9994Changes in diversity of cultured bacteria resistant to erythromycin and tetracycline in swine manure during simulated composting and lagoon storage. This study investigated the impact of composting and lagoon storage on survival and change in diversity of tetracycline-resistant (Tc(r) ) and erythromycin-resistant (Em(r) ) bacteria and the resistance genes they carry in swine manure. Treatments were arranged as a 2 × 2 factorial design: composting vs lagoon storage and 0 vs 1% Surround WP Crop Protectant (a clay product) in three replicates. After 48 days of treatments, resistant bacteria were enumerated by selective plating and identified by 16S rRNA gene sequencing. The erm and the tet gene(s) carried by the resistant isolates were screened using class-specific PCR assays. The plate counts of Tc(r) and Em(r) bacteria decreased by 4-7 logs by composting, but only by 1-2 logs by the lagoon treatment. During the treatments, Acinetobacter gave way to Pseudomonas and Providencia as the largest resistant genera. The clay product had little effect on survival or diversity of resistant bacteria. Of six classes of erm and seven classes of tet genes tested, changes in prevalence were also noted. The results indicate that composting can dramatically shift Tc(r) and Em(r) bacterial populations, and composting can be an effective and practical approach to decrease dissemination of antibiotic resistance from swine farms to the environment. SIGNIFICANCE AND IMPACT OF THE STUDY: The presented research provided evidence that composting is much more effective than lagoon storage in dramatically decreasing culturable bacteria resistant to erythromycin and tetracycline in swine manure. Considerable diversity changes of resistant bacteria were also demonstrated during composting or lagoon storage. Overall, Acinetobacter was the major resistant genus in untreated swine manure, but pseudomonads and Providencia became the major resistant genera after the treatments. This is the first study that investigated diversity changes of cultured bacteria resistant to these two antibiotics during composting and lagoon storage of swine manure. New genes encoding resistance to the two antibiotics were also implied in the cultured isolates.201526031793
696760.9994Effects of Pyroligneous Acid on Diversity and Dynamics of Antibiotic Resistance Genes in Alfalfa Silage. Antibiotic resistance genes (ARGs) are recognized as contaminants due to their potential risk for human and environment. The aim of the present study is to investigate the effects of pyroligneous acid (PA), a waste of biochar production, on fermentation characteristics, diversity, and dynamics of ARGs during ensiling of alfalfa using metagenomic analysis. The results indicated that PA decreased (P < 0.05) dry matter loss, pH value, gas production, coliform bacteria count, protease activity, and nonprotein-N, ammonia-N, and butyric acid contents and increased (P < 0.05) lactic acid content during ensiling. During fermentation, Bacteria, Firmicutes, and Lactobacillus were the most abundant at kingdom, phylum, and genus levels, respectively. Pyroligneous acid reduced the relative abundance of Bacteria and Firmicutes and increased that of Lactobacillus. The detected ARGs belonged to 36 drug classes, including mainly macrolides, tetracycline, lincosamides, and phenicol. These types of ARGs decreased during fermentation and were further reduced by PA. These types of ARGs were positively correlated (P < 0.05) with fermentation parameters like pH value and ammonia-N content and with bacterial communities. At the genus level, the top several drug classes, including macrolide, tetracycline, lincosamide, phenicol, oxazolidinone, streptogramin, pleuromutilin, and glycopeptide, were positively correlated with Staphylococcus, Streptococcus, Listeria, Bacillus, Klebsiella, Clostridium, and Enterobacter, the potential hosts of ARGs. Overall, ARGs in alfalfa silage were abundant and were influenced by the fermentation parameters and microbial community composition. Ensiling could be a feasible way to mitigate ARGs in forages. The addition of PA could not only improve fermentation quality but also reduce ARG pollution of alfalfa silage. IMPORTANCE Antibiotic resistance genes (ARGs) are considered environmental pollutants posing a potential human health risk. Silage is an important and traditional feed, mainly for ruminants. ARGs in silages might influence the diversity and distribution of ARGs in animal intestinal and feces and then the manure and the manured soil. However, the diversity and dynamics of ARGs in silage during fermentation are still unknown. We ensiled alfalfa, one of the most widely used forages, with or without pyroligneous acid (PA), which was proved to have the ability to reduce ARGs in soils. The results showed that ARGs in alfalfa silage were abundant and were influenced by the fermentation parameters and microbial community. The majority of ARGs in alfalfa silage reduced during fermentation. The addition of PA could improve silage quality and reduce ARG pollution in alfalfa silage. This study can provide useful information for understanding and controlling ARG pollution in animal production.202235862964
712670.9994Longitudinal characterization of antimicrobial resistance genes in feces shed from cattle fed different subtherapeutic antibiotics. BACKGROUND: Environmental transmission of antimicrobial-resistant bacteria and resistance gene determinants originating from livestock is affected by their persistence in agricultural-related matrices. This study investigated the effects of administering subtherapeutic concentrations of antimicrobials to beef cattle on the abundance and persistence of resistance genes within the microbial community of fecal deposits. Cattle (three pens per treatment, 10 steers per pen) were administered chlortetracycline, chlortetracycline plus sulfamethazine, tylosin, or no antimicrobials (control). Model fecal deposits (n = 3) were prepared by mixing fresh feces from each pen into a single composite sample. Real-time PCR was used to measure concentrations of tet, sul and erm resistance genes in DNA extracted from composites over 175 days of environmental exposure in the field. The microbial communities were analyzed by quantification and denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S-rRNA. RESULTS: The concentrations of 16S-rRNA in feces were similar across treatments and increased by day 56, declining thereafter. DGGE profiles of 16S-rRNA differed amongst treatments and with time, illustrating temporal shifts in microbial communities. All measured resistance gene determinants were quantifiable in feces after 175 days. Antimicrobial treatment differentially affected the abundance of certain resistance genes but generally not their persistence. In the first 56 days, concentrations of tet(B), tet(C), sul1, sul2, erm(A) tended to increase, and decline thereafter, whereas tet(M) and tet(W) gradually declined over 175 days. At day 7, the concentration of erm(X) was greatest in feces from cattle fed tylosin, compared to all other treatments. CONCLUSION: The abundance of genes coding for antimicrobial resistance in bovine feces can be affected by inclusion of antibiotics in the feed. Resistance genes can persist in feces from cattle beyond 175 days with concentrations of some genes increasing with time. Management practices that accelerate DNA degradation such as frequent land application or composting of manure may reduce the extent to which bovine feces serves as a reservoir of antimicrobial resistance.201121261985
719980.9994Minimum influent concentrations of oxytetracycline, streptomycin and spiramycin in selecting antibiotic resistance in biofilm type wastewater treatment systems. It has been demonstrated that antibiotic resistance could be induced and selected under high antibiotic concentrations in biological wastewater treatment systems. However, little is available regarding the minimum concentrations of antibiotics for selecting antibiotic resistance during wastewater treatment. Herein, the minimum influent concentrations of oxytetracycline, streptomycin, and spiramycin in selecting antibiotic resistance in biofilm type wastewater treatment systems were investigated by spiking respective antibiotic into wastewater with an antibiotic dose increasing from 0 to 0.1, 1, 5, 25, 50 mg/L stepwise over a period of 606 days. Significant increase (p < .01) in the total abundance of antibiotic resistance genes was observed for both streptomycin and oxytetracycline at a dose of 0.1 mg/L according to metagenomic sequencing, while the concentration levels leading to significant increases (p < .05) in resistant bacteria ratio were higher: 5 mg/L for streptomycin and 25 mg/L for oxytetracycline. Although resistome abundance increased with the increase of spiramycin dose, neither the corresponding Macrolide-Lincosamide-Streptogramin (MLS) resistance genes nor the resistant bacteria ratio showed perceptible increase. Partial canonical correspondence analysis showed that both bacterial community shift and mobile genetic elements alteration contributed to the enrichment of resistomes under the presence of streptomycin and oxytetracycline. Regarding spiramycin which is mainly targeting on Gram-positive bacteria, the dominance of the intrinsically resisting Gram-negative bacteria in the biofilm microbiota might be responsible for the vague change of MLS resistant determinants under the spiramycin stress. The results demonstrated that it is possible to prevent the development of antibiotic resistance during wastewater treatment by controlling the influent streptomycin and oxytetracyline concentrations below 0.1 mg/L. This work proposed an actionable approach for the management of antibiotic production wastewater.202032325576
741190.9994Thermophilic Composting of Human Feces: Development of Bacterial Community Composition and Antimicrobial Resistance Gene Pool. In times of climate change, practicing sustainable, climate-resilient, and productive agriculture is of primordial importance. Compost from different resources, now treated as wastes, could be one form of sustainable fertilizer creating a resilience of agriculture to the adverse effects of climate change. However, the safety of the produced compost regarding human pathogens, pharmaceuticals, and related resistance genes must be considered. We have assessed the effect of thermophilic composting of dry toilet contents, green cuttings, and straw, with and without biochar, on fecal indicators, the bacterial community, and antibiotic resistance genes (ARGs). Mature compost samples were analyzed regarding fecal indicator organisms, revealing low levels of Escherichia coli that are in line with German regulations for fertilizers. However, one finding of Salmonella spp. exceeded the threshold value. Cultivation of bacteria from the mature compost resulted in 200 isolates with 36.5% of biosafety level 2 (BSL-2) species. The majority is known as opportunistic pathogens that likewise occur in different environments. A quarter of the isolated BSL-2 strains exhibited multiresistance to different classes of antibiotics. Molecular analysis of total DNA before and after composting revealed changes in bacterial community composition and ARGs. 16S rRNA gene amplicon sequencing showed a decline of the two most abundant phyla Proteobacteria (start: 36-48%, end: 27-30%) and Firmicutes (start: 13-33%, end: 12-16%), whereas the abundance of Chloroflexi, Gemmatimonadetes, and Planctomycetes rose. Groups containing many human pathogens decreased during composting, like Pseudomonadales, Bacilli with Bacillus spp., or Staphylococcaceae and Enterococcaceae. Gene-specific PCR showed a decline in the number of detectable ARGs from 15 before to 8 after composting. The results reveal the importance of sufficiently high temperatures lasting for a sufficiently long period during the thermophilic phase of composting for reducing Salmonella to levels matching the criteria for fertilizers. However, most severe human pathogens that were targeted by isolation conditions were not detected. Cultivation-independent analyses also indicated a decline in bacterial orders comprising many pathogenic bacteria, as well as a decrease in ARGs. In summary, thermophilic composting could be a promising approach for producing hygienically safe organic fertilizer from ecological sanitation.202235250940
8037100.9993Dosage effects of lincomycin mycelial residues on lincomycin resistance genes and soil microbial communities. Lincomycin mycelial residues (LMRs) are one kind of byproduct of the pharmaceutical industry. Hydrothermal treatment has been used to dispose of them and land application is an attractive way to reuse the treated LMRs. However, the safe dose for soil amendment remains unclear. In this study, a lab-scale incubation experiment was conducted to investigate the influence of the amendment dosage on lincomycin resistance genes and soil bacterial communities via quantitative PCR and 16S rRNA sequencing. The results showed that introduced lincomycin degraded quickly in soil and became undetectable after 50 days. Degradation rate of the high amendment amount (100 mg kg(-1)) was almost 4 times faster than that of low amendment amount (10 mg kg(-1)). Moreover, the introduced LMRs induced the increase of lincomycin resistance genes after incubation for 8 days, and two genes (lmrA and lnuB) showed a dosage-related increase. For example, the abundance of gene lmrA was 17.78, 74.13 and 128.82 copies g(-1) soil for lincomycin concentration of 10, 50 and 100 mg kg(-1), respectively. However, the abundance of lincomycin resistance genes recovered to the control level as the incubation period extended to 50 days, indicating a low persistence in soil. In addition, LMRs application markedly shifted the bacterial composition and significant difference was found between control soil, 10 mg kg(-1) and 50 mg kg(-1) lincomycin amended soil. Actually, several genera bacteria were significantly related to the elevation of lincomycin resistance genes. These results provided a comprehensive understanding of the effects of lincomycin dosage on the fate of resistance genes and microbial communities in LMRs applied soil.202031662263
8024110.9993High Concentrations of Tilmicosin Promote the Spread of Multidrug Resistance Gene tolC in the Pig Gut Microbiome Through Mobile Genetic Elements. The impact of antibiotic therapy on the spread of antibiotic resistance genes (ARGs) and its relationship to gut microbiota remains unclear. This study investigated changes in ARGs, mobile genetic elements (MGEs), and gut microbial composition following tilmicosin administration in pigs. Thirty pigs were randomly divided into control (CK), low-concentration (0.2 g/kg; L), and high-concentration (0.4 g/kg; H) groups. Tilmicosin concentration in manure peaked on day 16 of dosing and dropped below detectable levels by day 13 of the withdrawal period. While tilmicosin did not significantly affect the total abundance of macrolide resistance genes (MRGs) (p > 0.05), it significantly increased the abundance of the multidrug resistance gene tolC in the H group compared with the L and CK groups during the withdrawal period (p < 0.05). This increase was associated with a coincidental rise in the abundance of MGEs (e.g., int1 and int2) and the growth of potential tolC-hosting bacteria such as Paenalcaligenes and Proteiniclasticum. Redundancy analysis showed gut microbial composition as the primary driver of MRG abundance, with MGEs, tilmicosin concentration, and manure physicochemical properties playing secondary roles. These findings suggest that high-dose tilmicosin may alter the gut microbiota and promote ARG spread via MGE-mediated transfer.202439795013
8031120.9993Anaerobic Digestion of Tetracycline Spiked Livestock Manure and Poultry Litter Increased the Abundances of Antibiotic and Heavy Metal Resistance Genes. Anaerobic digestion is used for the treatment of animal manure by generating biogas. Heavy metals cause environmental pollutions and co-select for antimicrobial resistance. We evaluated the impact of mesophilic anaerobic digestion of cattle manure (CM), swine manure (SM) and poultry litter (PL) on the concentrations of seven tetracycline [tet(A), tet(B), tet(G), tet(M), tet(O), tet(Q), and tet(W)], macrolide [erm(B)], methicillin (mecA and mecC), copper (copB, pcoA, pcoD, and tcrB) and zinc (czrC) resistance genes, and three bacterial species (E. coli, Enterococcus spp. and Staphylococcus aureus). The total bacterial population and total abundance of the seven tet genes significantly increased in the three manure types after digestion. Concentration of tet(M) was strongly correlated with that of erm(B) and enterococci. As concentration of tetracyclines declined during anaerobic digestion, that of four tet genes (A, B, Q, and W) and 16S rRNA increased, that of tet(M) decreased, and that of tet(G) and tet(O) did not change. Concentrations of copB and pcoA did not change; while that of pcoD did not change in the PL, it increased in the SM and CM. While the concentration of enterococci remained unchanged in CM, it significantly increased in the PL and SM. Concentrations of tcrB significantly increased in the three manure types. While concentrations of S. aureus significantly increased in the CM and PL, that of SM was not affected. Concentrations of mecC significantly increased in all manure types after digestion; while mecA concentrations did not change in the SM, they significantly increased in CM and PL. While concentration of czrC remained low in the CM, it increased in the PL but declined in the SM. In conclusion, while mesophilic anaerobic digestion of animal manure decreased concentration of tetracyclines, it increased the concentrations of total bacteria, tet genes, E. coli, enterococci and S. aureus and methicillin resistance genes. It did not have any effect on concentrations of heavy metals; concentrations of heavy metal resistance genes either increased or remained unaffected depending on the animal species. This study showed the need for post-digestion treatments of animal manure to remove bacteria, antibiotic resistance genes, heavy metals and their resistance genes.202033391245
7071130.9993Impacts of multi-year field exposure of agricultural soil to macrolide antibiotics on the abundance of antibiotic resistance genes and selected mobile genetic elements. Exposure of environmental bacteria to antibiotics may be increasing the global resistome. Antibiotic residues are entrained into agricultural soil through the application of animal and human wastes, and irrigation with reclaimed water. The impact of a mixture of three macrolide antibiotics on the abundance of selected genes associated with antibiotic resistance and genetic mobility were determined in a long-term field experiment undertaken in London, Canada. Replicated plots received annual applications of a mixture of erythromycin, clarithromycin and azithromycin every spring since 2010. Each antibiotic was added directly to the soil at a concentration of either 0.1 or 10 mg kg soil(-1) and all plots were cropped to soybeans. By means of qPCR, no gene targets were enriched in soil exposed to the 0.1 mg kg soil(-1) dose compared to untreated control. In contrast, the relative abundance of several gene targets including int1, sul2 and mphE increased significantly with the annual exposure to the 10 mg kg soil(-1) dose. By means of high-throughput qPCR, numerous gene targets associated with resistance to aminoglycosides, sulfonamides, trimethoprim, streptomycin, quaternary ammonium chemicals as well as mobile genetic elements (tnpA, IS26 and IS6100) were detected in soil exposed to 10 mg kg soil(-1), but not the lower dose. Overall, exposure of soil to macrolide antibiotics increased the relative abundance of numerous gene targets associated with resistance to macrolides and other antibiotics, and mobile genetic elements. This occurred at an exposure dose that is unrealistically high, but did not occur at the lower more realistic exposure dose.202032330714
7202140.9993Cyanobacterial extracellular antibacterial substances could promote the spread of antibiotic resistance: impacts and reasons. Many studies have shown that antibiotic resistance genes (ARGs) can be facilitated by a variety of antibacterial substances. Cyanobacteria are photosynthetic bacteria that are widely distributed in the ocean. Some extracellular substances produced by marine cyanobacteria have been found to possess antibacterial activity. However, the impact of these extracellular substances on ARGs is unclear. Therefore, we established groups of seawater microcosms that contained different concentrations (1000, 100, 10, 1, 0.1, 0.01, and 0 μg mL(-1)) of cyanobacterial extracellular substances (CES), and tracked the changes of 17 types of ARGs, the integron gene (intI1), as well as the bacterial community at different time points. The results showed that CES could enrich most ARGs (15/17) in the initial stage, particularly at low concentrations (10 and 100 μg mL(-1)). The correlation analysis showed a positive correlation between several ARGs and intI1. It is suggested that the abundance of intI1 increased with CES may contribute to the changes of these ARGs, and co-resistance of CES may be the underlying reason for the similar variation pattern of some ARGs. Moreover, the results of qPCR and high-throughput sequencing of 16S rRNA showed that CES had an inhibitory impact on the growth of bacterial communities. High concentrations of CES were found to alter the structure of bacterial communities. Co-occurrence networks showed that bacteria elevated in the high concentration group of CES and might serve as the potential hosts for a variety of ARGs. In general, marine cyanobacteria could play an important role in the global dissemination of ARGs and antibiotic-resistant bacteria (ARBs).202337947439
7123150.9993Presence and fate of antibiotic residues, antibiotic resistance genes and zoonotic bacteria during biological swine manure treatment. The presence and dissemination of antibiotic residues, antibiotic resistance genes and zoonotic bacteria in the environment is of growing concern worldwide. Manure management practices, such as biological removal of nitrogen from swine manure, may help to decrease levels of antibiotic residues, antibiotic resistance genes and zoonotic bacteria present in manure before fertilization, thereby reducing environmental contamination. Therefore, the aim of this study was to monitor the presence and fate of seven antibiotic residues (colistin, sulfadiazine, trimethoprim, doxycycline, oxytetracycline, ceftiofur and tylosin A), nine antibiotic resistance genes (tet(B), tet(L), tet(M), tet(O), tet(Q), tet(W), erm(B), erm(F) and sul2) and two zoonotic bacteria (Salmonella Typhimurium and Campylobacter coli) during biological nitrogen removal from swine manure over time. Samples from the raw manure, the solid fraction, the liquid fraction and the storage lagoon were analyzed on two farms at six time points with an interval of two weeks. Only the antibiotics which were used during the three months preceding the first sampling could be detected before and after biological nitrogen removal from swine manure. Of all the antibiotics studied, doxycycline was recovered in all of the samples and sulfadiazine was recovered in most samples on both farms. For both antibiotics, there appears to be a reduction of the amount of residues present in the storage lagoon compared to the liquid fraction, however, this reduction was not statistically significant. A significant reduction of the relative abundances of most of the antibiotic resistance genes studied was observed when comparing the liquid fraction and the storage lagoon. For tet(L), no differences were observed between the fractions sampled and for sul2 and erm(F), a significant increase in relative abundances was observed on the second farm sampled. For the zoonotic bacteria, a reduction of at least 1 log was observed after biological nitrogen removal from swine manure. The results indicate that the concentration of certain antibiotic residues and several antibiotic resistance genes and the amount of zoonotic bacteria present in the manure may be reduced in the end product of the biological nitrogen removal from swine manure.201930878661
7413160.9993Fecal antibiotic resistance genes were transferred through the distribution of soil-lettuce-snail food chain. Massive antibiotic resistance genes (ARG) were detected in the soil modified by manure, which may affect human life safety through the food chain. However, the transmission of ARGs through the soil-plant-animal food chain is still unclear. Therefore, this study used high-throughput quantitative PCR technology to explore the effects of pig manure application on ARGs and bacterial communities in soil, lettuce phyllosphere, and snail excrement. The results showed that a total of 384 ARGs and 48 MEGs were detected in all samples after 75 days of incubation. The diversity of ARGs and MGEs in soil components increased significantly by 87.04% and 40% with the addition of pig manure. The absolute abundance of ARGs in the phyllosphere of lettuce was significantly higher than that of the control group, with a growth rate of 212.5%. Six common ARGs were detected between the three components of the fertilization group, indicating that there was internal transmission of fecal ARGs between the trophic levels of the food chain. Firmicutes and Proteobacteria were identified as the dominant host bacteria in the food chain system, which were more likely to be used as carriers of ARGs to promote the spread of resistance in the food chain. The results were used to assess the potential ecological risks of livestock and poultry manure. It provides theoretical basis and scientific support for the formulation of ARG prevention and control policies.202337434056
7414170.9993Structure of the manure resistome and the associated mobilome for assessing the risk of antimicrobial resistance transmission to crops. In this study, the impact of bovine and poultry manure on the quantitative and qualitative composition of antibiotic resistance genes (ARGs) and the environmental mobilome associated with antimicrobial resistance in soil and crops was determined with the use of next generation sequencing methods. The aim of the study was to perform a metagenomic analysis of manure to estimate the risk of the transmission of ARGs and bacterial drug resistance carriers to fertilized soil and crops. The total copy number of ARGs was nearly four times higher in poultry manure (555 ppm) than in bovine manure (140 ppm), and this relationship was also noted in fertilized soil. Poultry manure induced a much greater increase in the concentrations of ARGs in the soil environment (196.4 ppm) than bovine manure (137.8 ppm) immediately after supplementation. The application of poultry manure led to the highest increase in the abundance of genes encoding resistance to tetracyclines (9%), aminoglycosides (3.5%), sulfonamides (3%), bacitracin (2%), chloramphenicol (2%), and macrolide-lincosamide-streptogramin antibiotics (1%). Heavy metals were stronger promoters of antibiotic resistance in the environment than antibiotics. Antibiotics exerted a greater influence on maintaining the diversity of ARGs than on increasing their abundance in soil. Large quantities of insertion sequences (IS), including those associated with the mobility of ARGs in the population of ESKAPEE pathogens, are introduced to soil with manure. These IS remain stable for up to several months, which indicates that manure, in particular poultry manure, significantly increases the risk of rapid ARG transfer to the environment. Manure also largely contributes to an increase in the diversity of the resistome and mobilome in the metagenome of bacteria isolated from crops. Bacteria of the phylum Proteobacteria appear to play a major role in the transmission of multiple ARGs in crops grown for human and animal consumption.202234864022
3432180.9993Insights into the amplification of bacterial resistance to erythromycin in activated sludge. Wastewater treatment plants are significant reservoirs for antimicrobial resistance. However, little is known about wastewater treatment effects on the variation of antibiotic resistance. The shifts of bacterial resistance to erythromycin, a macrolide widely used in human medicine, on a lab-scale activated sludge system fed with real wastewater was investigated from levels of bacteria, community and genes, in this study. The resistance variation of total heterotrophic bacteria was studied during the biological treatment process, based on culture dependent method. The alterations of bacterial community resistant to erythromycin and nine typical erythromycin resistance genes were explored with molecular approaches, including high-throughput sequencing and quantitative polymerase chain reaction. The results revealed that the total heterotrophs tolerance level to erythromycin concentrations (higher than 32 mg/L) was significantly amplified during the activated sludge treatment, with the prevalence increased from 9.6% to 21.8%. High-throughput sequencing results demonstrated an obvious increase of the total heterotrophic bacterial diversity resistant to erythromycin. Proteobacteria and Bacteroidetes were the two dominant phyla in the influent and effluent of the bioreactor. However, the prevalence of Proteobacteria decreased from 76% to 59% while the total phyla number increased greatly from 18 to 29 through activated sludge treatment. The gene proportions of erm(A), mef(E) and erm(D) were greatly amplified after biological treatment. It is proposed that the transfer of antibiotic resistance genes through the variable mixtures of bacteria in the activated sludge might be the reason for the antibiotic resistance amplification. The amplified risk of antibiotic resistance in wastewater treatment needs to be paid more attention.201525957255
8032190.9993Enrichment of the Antibiotic Resistance Gene tet(L) in an Alkaline Soil Fertilized With Plant Derived Organic Manure. Fifteen antibiotic resistance genes (ARGs) and intI1, a gene involved in horizontal gene transfer (HGT) of ARGs, were quantified in three different soil samples from a 22 year old field experiment that had received inorganic fertilizer (NPK), organic manure (OM; a mixture of wheat straw, soybean oil cake and cotton cake), and control fields that had received no fertilizer and manure (CK). Tet(L) was the most abundant ARG in OM, which also contained considerable levels of intI1. Molecular analysis of yearly collected archived soils over the past 22 years showed that tet(L) and intI1 were higher in OM soils than in NPK soils. The relative abundance of tet(L) was essentially constant during these years, while the level of intI1 in OM soils decreased over time. The main genotype of tet(L) was the same in archived and in fresh soil, OM, and irrigation water. Phylogenetic analysis of the 16S rRNA genes of tetracycline-resistant bacteria (TRB) isolates indicated that the Firmucutes carrying tet(L) in OM were similar to those in the OM soil, suggesting that OM transferred TRB into the OM soils where they survived. Almost all of the TRB isolated from OM carried tet(L) and belonged to the Firmicutes. Survival of bacteria from the organic manure that carried tet(L) may be the cause of the increased level of tet(L) in OM soil.201829904377