Biocide Resistance Research Articles

A computational model for microbial colonization of an antifouling surface.

Biofouling of marine surfaces such as ship hulls is a major industrial problem. Antifouling (AF) paints delay the onset of biofouling by releasing biocidal chemicals. We present a computational model for microbial colonization of a biocide-releasing AF surface. Our model accounts for random arrival from the ocean of microorganisms with different biocide resistance levels, biocide-dependent proliferation or killing, and a transition to a biofilm state. Our computer simulations support a picture in which biocide-resistant microorganisms initially form a loosely attached layer that eventually transitions to a growing biofilm. Once the growing biofilm is established, immigrating microorganisms are shielded from the biocide, allowing more biocide-susceptible strains to proliferate. In our model, colonization of the AF surface is highly stochastic. The waiting time before the biofilm establishes is exponentially distributed, suggesting a Poisson process. The waiting time depends exponentially on both the concentration of biocide at the surface and the rate of arrival of resistant microorganisms from the ocean. Taken together our results suggest that biofouling of AF surfaces may be intrinsically stochastic and hence unpredictable, but immigration of more biocide-resistant species, as well as the biological transition to biofilm physiology, may be important factors controlling the time to biofilm establishment.

Combining Topography and Chemistry to Produce Antibiofouling Surfaces: A Review.

Despite decades of research on the reduction of surface fouling from biomolecules or micro-organisms, the ultimate antibiofouling surface remains undiscovered. The recent covid-19 pandemic strengthened the crucial need for such treatments. Among the numerous approaches that are able to provide surfaces with antibiofouling properties, chemical, biological, and topographical strategies have been implemented for instance in the marine, medical, or food industries. However, many of these methods have a biocidal effect and, with antibioresistance and biocide resistance a growing threat on humanity, strategies based on reducing adsorption of biomolecules and micro-organism are necessary for long-term solutions. Bioinspired strategies, combining both surface chemistry and topography, are currently at the heart of the best innovative and sustainable solutions. The synergistic effect of micro/nanostructuration, together with engineered chemical or biological functionalization is believed to contribute to the development of antibiofouling surfaces. This review aims to present approaches combining hydrophobic or hydrophilic chemistries with a specific topography to avoid biofouling in various industrial environments and healthcare facilities.

Impacts of soybean agriculture on the resistome of the Amazonian soil.

The soils of the Amazon are complex environments with different organisms cohabiting in continuous adaptation processes; this changes significantly when these environments are modified for the development of agricultural activities that alter the chemical, macro, and microbiological compositions. The metagenomic variations and the levels of the environmental impact of four different soil samples from the Amazon region were evaluated, emphasizing the resistome. Soil samples from the organic phase from the different forest, pasture, and transgenic soybean monocultures of 2–14 years old were collected in triplicate at each site. The samples were divided into two groups, and one group was pre-treated to obtain genetic material to perform sequencing for metagenomic analysis; another group carried out the chemical characterization of the soil, determining the pH, the content of cations, and heavy metals; these were carried out in addition to identifying with different databases the components of the microbiological communities, functional genes, antibiotic and biocide resistance genes. A greater diversity of antibiotic resistance genes was observed in the forest soil. In contrast, in monoculture soils, a large number of biocide resistance genes were evidenced, highlighting the diversity and abundance of crop soils, which showed better resistance to heavy metals than other compounds, with a possible dominance of resistance to iron due to the presence of the acn gene. For up to 600 different genes for resistance to antibiotics and 256 genes for biocides were identified, most of which were for heavy metals. The most prevalent was resistance to tetracycline, cephalosporin, penam, fluoroquinolone, chloramphenicol, carbapenem, macrolide, and aminoglycoside, providing evidence for the co-selection of these resistance genes in different soils. Furthermore, the influence of vegetation cover on the forest floor was notable as a protective factor against the impact of human contamination. Regarding chemical characterization, the presence of heavy metals, different stress response mechanisms in monoculture soils, and the abundance of mobile genetic elements in crop and pasture soils stand out. The elimination of the forest increases the diversity of genes for resistance to biocides, favoring the selection of genes for resistance to antibiotics in soils.

Controlling AMR in the Pig Industry: Is It Enough to Restrict Heavy Metals?

Heavy metals have the potential to influence the transmission of antimicrobial resistance (AMR). However, the effect on AMR caused by heavy metals has not been clearly revealed. In this study, we used a microcosm experiment and metagenomics to examine whether common levels of Cu and Zn in pig manure influence AMR transmission in manured soil. We found that the abundance of 204 ARGs significantly increased after manure application, even though the manure did not contain antibiotic residuals. However, the combined addition of low Cu and Zn (500 and 1000 mg/kg, respectively) only caused 14 ARGs to significantly increase, and high Cu and Zn (1000 and 3000 mg/kg, respectively) caused 27 ARGs to significantly increase. The disparity of these numbers suggested that factors within the manure were the primary driving reasons for AMR transmission, rather than metal amendments. A similar trend was found for biocide and metal resistance genes (BMRGs) and mobile genetic elements (MGEs). This study offers deeper insights into AMR transmission in relation to the effects of manure application and heavy metals at commonly reported levels. Our findings recommend that more comprehensive measures in controlling AMR in the pig industry are needed apart from restricting heavy metal additions.

Exudates from Miscanthus x giganteus change the response of a root-associated Pseudomonas putida strain towards heavy metals

The composition of root exudates is modulated by several environmental factors, and it remains unclear how that affects beneficial rhizosphere or inoculated microorganisms under heavy metal (HM) contamination. Therefore, we evaluated the transcriptional response of Pseudomonas putida E36 (a Miscanthus x giganteus isolate with plant growth promotion-related properties) to Cd, Pb and Zn in an in vitro study implementing root exudates from M. x giganteus. To collect root exudates and analyse their composition plants were grown in a pot experiment under HM and control conditions.Our results indicated higher exudation rate for plants challenged with HM. Further, out of 29 organic acids identified and quantified in the root exudates, 8 of them were significantly influenced by HM (e.g., salicylic and terephthalic acid). The transcriptional response of P. putida E36 was significantly affected by the HM addition to the growth medium, increasing the expression of several efflux pumps and stress response-related functional units. The additional supplementation of the growth medium with root exudates from HM-challenged plants resulted in a downregulation of 29% of the functional units upregulated in P. putida E36 as a result of HM addition to the growth medium. Surprisingly, root exudates + HM downregulated the expression of P. putida E36 functional units related to plant colonization (e.g., chemotaxis, motility, biofilm formation) but upregulated its antibiotic and biocide resistance compared to the control treatment without HM.Our findings suggest that HM-induced changes in root exudation pattern may attract beneficial bacteria that are in turn awarded with organic nutrients, helping them cope with HM stress. However, it might affect the ability of these bacteria to colonize plants growing in HM polluted areas. Those findings may offer an insight for future in vivo studies contributing to improvements in phytoremediation measures.

Suitability of Methods to Determine Resistance to Biocidal Active Substances and Disinfectants—A Systematic Review

Biocide resistance is an increasing concern. However, it is currently unknown if an elevated MIC value for an isolate correlates with a lower log10 reduction in suspension tests or carrier tests. The aim of this review was therefore to evaluate if isolates with a suspected tolerance to a biocidal active substance reveal an elevated MIC value and an impaired efficacy in suspension tests and carrier tests. A Medline search was done on 6 July 2022 using the following terms: “resistance biocidal MIC suspension” (16 hits), “resistance biocidal MIC carrier” (22 hits), “resistance biocidal suspension carrier” (41 hits), “tolerance biocidal MIC suspension” (1 hit), “tolerance biocidal MIC carrier” (4 hits) and “tolerance biocidal suspension carrier” (3 hits). Studies were included when a tolerance or resistance to the biocidal active substance or disinfectant was suspected and at least two of the three endpoints were evaluated in parallel in comparison to the control isolates. In three out of five studies, the elevated MIC values did not correlate with an impaired bactericidal efficacy against the biocide-tolerant isolates. In three out of five studies, an impaired activity in the suspension tests was described that correlated with an impaired efficacy in the carrier tests (peracetic acid-tolerant K. pneumoniae and glutaraldehyde-tolerant M. chelonae; the two other studies did not allow a comparison. Overall, the results from the suspension tests and tests under practical conditions allowed to determine a clinically relevant resistance.

Substrate complexity affects the prevalence and interconnections of antibiotic, metal and biocide resistance genes, integron-integrase genes, human pathogens and virulence factors in anaerobic digestion

Anaerobic digestion (AD) is widely used to treat livestock manure that harbors diverse pollutants (resistance genes (ARGs), metal/biocide resistance genes (MBRGs), integron-integrase genes, human pathogens and pathogen virulence factors (VFs)). However, the interplays of these pollutants and the effects of substrate complexity on pollutants in AD are elusive. This study investigated the dynamics of these pollutants and bacterial communities during AD of swine manure, by metatranscriptomic sequencing and amplicon sequencing of 16 S rRNA and 16 S rRNA gene. The pollutant profiles and bacterial communities differed across AD processes, nevertheless with consistent dominance of ARGs of multi-drugs, tetracycline, aminoglycoside and rifamycin, MBRGs of multi-biocides, multi-metals, copper and arsenic, the integron-integrase gene intI1, potential pathogens of Escherichia coli, Streptococcus gallolyticus and Clostridium perfringens, VFs involved in pathogen adherence, and bacterial phyla of Firmicutes, Bacteroidetes and Proteobacteria. Reduced substrate complexity (replacing a part of swine manure, a complex substrate, with a simple substrate, apple waste or fructose) decreased the prevalence and stochastic turnover of ARGs and MBRGs. Network analyses revealed decreased interplays among pollutants under reduced substrate complexity. Our findings provide a mechanical understanding of diverse pollutants dynamics during AD, and reveal the importance of substrate complexity in controlling prevalence and interplays of pollutants.

Higher spreading risk of antibacterial biocide and heavy metal resistance genes than antibiotic resistance genes in aerobic granular sludge

Metagenomic approach was applied to simultaneously reveal the antibiotic resistance genes (ARGs) and antibacterial biocide & metal resistance genes (BMRGs), and the corresponding microbial hosts with high mobility during aerobic granular sludge (AGS) formation process. The results showed that the relative abundance of BMRGs was 88–123 times that of ARGs. AGS process was easier to enrich BMRGs, leading to a greater risk of drug resistance caused by BMRGs than that by ARGs. The enrichments of ARGs and BMRGs in AGS were closely related to several enhanced microbial metabolisms (i.e., cell motility, transposase and ATP-binding cassette transporters) and their corresponding regulatory genes. Several enhanced KEGG Orthologs (KO) functions, such as K01995, K01996, K01997 and K02002, might cause a positive impact on the spread of ARGs and BMRGs, and the main contributors were the largely enriched glycogens accumulating organisms. The first dominant ARGs (adeF) was carried by lots of microbial hosts, which might be enriched and propagated mainly through horizontal gene transfer. Candidatus Competibacter denitrificans simultaneously harbored ARG (cmx) and Cu related RGs (corR). Many enriched bacteria contained simultaneously multiple BMRGs (copR and corR) and mobile genetic elements (integrons and plasmids), granting them high mobility capabilities and contributing to the spread of BMRGs. This study might provide deeper understandings of the proliferation and mobility of ARGs and BMRGs, importantly, highlighted the status of BMRGs, which laid the foundation for the controlling widespread of resistance genes in AGS.

Metagenomic insights into the microbial community structure and resistomes of a tropical agricultural soil persistently inundated with pesticide and animal manure use.

Persistent use of pesticides and animal manure in agricultural soils inadvertently introduced heavy metals and antibiotic/antibiotic resistance genes (ARGs) into the soil with deleterious consequences. The microbiome and heavy metal and antibiotic resistome of a pesticide and animal manure inundated agricultural soil (SL6) obtained from a vegetable farm at Otte, Eiyenkorin, Kwara State, Nigeria, was deciphered via shotgun metagenomics and functional annotation of putative ORFs (open reading frames). Structural metagenomics of SL6 microbiome revealed 29 phyla, 49 classes, 94 orders, 183 families, 366 genera, 424 species, and 260 strains with the preponderance of the phyla Proteobacteria (40%) and Actinobacteria (36%), classes Actinobacteria (36%), Alphaproteobacteria (18%), and Gammaproteobacteria (17%), and genera Kocuria (16%), Sphingobacterium (11%), and Brevundimonas (10%), respectively. Heavy metal resistance genes annotation conducted using Biocide and Metal Resistance Gene Database (BacMet) revealed the detection of genes responsible for the uptake, transport, detoxification, efflux, and regulation of copper, cadmium, zinc, nickel, chromium, cobalt, selenium, tungsten, mercury, and several others. ARG annotation using the Antibiotic Resistance Gene-annotation (ARG-ANNOT) revealed ARGs for 11 antibiotic classes with the preponderance of β-lactamases, mobilized colistin resistance determinant (mcr-1), macrolide-lincosamide-streptogramin (MLS), glycopeptide, and aminoglycoside resistance genes, among others. The persistent use of pesticide and animal manure is strongly believed to play a major role in the proliferation of heavy metal and antibiotic resistance genes in the soil. This study revealed that agricultural soils inundated with pesticide and animal manure use are potential hotspots for ARG spread and may accentuate the spread of multidrug resistant clinical pathogens.

Comparing resistome profiles from anthropogenically impacted and non-impacted areas of two South Shetland Islands – Maritime Antarctica

Whalers Bay, in Deception Island, has one of the most anthropogenically impacted areas in Maritime Antarctica. However, considering the volcanic nature (high concentrations of heavy metals) of Deception Island's soils, this putative anthropogenic impact should be carefully investigated. In this context, the objective of this study was to compare resistome profiles of impacted and non-impacted areas in Deception Island (Whalers Bay, Crater Lake, and Fumarole Bay) and Livingston Island (Hannah Point) in order to investigate the microbiome tolerance/resistance mechanisms selected as a function of environmental drivers. Metagenomics was used to search for genes conferring resistance/tolerance to antibiotics, biocides, and heavy metals. Whalers Bay has a greater diversity of antibiotic, biocide, and heavy metal resistance classes found in its microbiomes. However, Hannah Point, at Livingston Island, has a greater abundance of antibiotic and biocide resistance/tolerance genes. The microbiome of Deception Island's non-impacted areas (Crater Lake and Fumarole Bay) showed resistance/tolerance genes almost entirely to heavy metals. Pb was found in higher concentrations in Whalers Bay soil in comparison to the other areas, indicating human contamination. The non-metric multidimensional scaling (NMDS) analysis revealed that Pb concentrations influenced resistome profiles in Whalers Bay soil. Despite the effect of Pb on the microbial communities of Whalers Bay, most heavy metal concentrations did not have a significant impact on resistome genes, suggesting that the volcanic soil heavy metal concentration of Deception Island has little biological influence.

Virulome and genome analyses identify associations between antimicrobial resistance genes and virulence factors in highly drug-resistant Escherichia coli isolated from veal calves.

Food animals are known reservoirs of multidrug-resistant (MDR) Escherichia coli, but information regarding the factors influencing colonization by these organisms is lacking. Here we report the genomic analysis of 66 MDR E. coli isolates from non-redundant veal calf fecal samples. Genes conferring resistance to aminoglycosides, β-lactams, sulfonamides, and tetracyclines were the most frequent antimicrobial resistance genes (ARGs) detected and included those that confer resistance to clinically significant antibiotics (blaCMY-2, blaCTX-M, mph(A), erm(B), aac(6’)Ib-cr, and qnrS1). Co-occurrence analyses indicated that multiple ARGs significantly co-occurred with each other, and with metal and biocide resistance genes (MRGs and BRGs). Genomic analysis also indicated that the MDR E. coli isolated from veal calves were highly diverse. The most frequently detected genotype was phylogroup A-ST Cplx 10. A high percentage of isolates (50%) were identified as sequence types that are the causative agents of extra-intestinal infections (ExPECs), such as ST69, ST410, ST117, ST88, ST617, ST648, ST10, ST58, and ST167, and an appreciable number of these isolates encoded virulence factors involved in the colonization and infection of the human urinary tract. There was a significant difference in the presence of multiple accessory virulence factors (VFs) between MDR and susceptible strains. VFs associated with enterohemorrhagic infections, such as stx, tir, and eae, were more likely to be harbored by antimicrobial-susceptible strains, while factors associated with extraintestinal infections such as the sit system, aerobactin, and pap fimbriae genes were more likely to be encoded in resistant strains. A comparative analysis of SNPs between strains indicated that several closely related strains were recovered from animals on different farms indicating the potential for resistant strains to circulate among farms. These results indicate that veal calves are a reservoir for a diverse group of MDR E. coli that harbor various resistance genes and virulence factors associated with human infections. Evidence of co-occurrence of ARGs with MRGs, BRGs, and iron-scavenging genes (sit and aerobactin) may lead to management strategies for reducing colonization of resistant bacteria in the calf gut.

Draft Genome Sequencing of Three Glutaraldehyde-Tolerant Bacteria from Produced Water from Hydraulic Fracturing.

ABSTRACTHere, we report the draft genome sequence of three glutaraldehyde-resistant isolates from produced water from hydraulic fracturing operations. The three strains were identified as Marinobacter sp. strain G11, Halomonas sp. strain G15, and Bacillus sp. strain G16. The genome sequences of these isolates will provide insights into biocide resistance in hydraulic fracturing operations.

Antimicrobial and Biocide Resistance among Canine and Feline Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Pseudomonas aeruginosa, and Acinetobacter baumannii Isolates from Diagnostic Submissions.

A total of 215 isolates from infections of dogs and cats, including 49 Enterococcus faecalis, 37 Enterococcus faecium, 59 Escherichia coli, 56 Pseudomonas aeruginosa, and 14 Acinetobacter baumannii, were investigated for their susceptibility to 27 (Gram-positive bacteria) or 20 (Gram-negative bacteria) antimicrobial agents/combinations of antimicrobial agents by broth microdilution according to the recommendations of the Clinical and Laboratory Standards Institute. Moreover, all isolates were analysed for their susceptibility to the biocides benzalkonium chloride, chlorhexidine, polyhexanide, and octenidine by a recently published broth microdilution biocide susceptibility testing method. While the E. faecalis isolates did not show expanded resistances, considerable numbers of the E. faecium isolates were resistant to penicillins, macrolides, tetracyclines, and fluoroquinolones. Even a single vancomycin-resistant isolate that carried the vanA gene cluster was detected. Expanded multiresistance phenotypes were also detected among the E. coli isolates, including a single carbapenem-resistant, blaOXA-48-positive isolate. In addition, multiresistant A. baumannii isolates were detected. The minimal inhibitory concentrations of the biocides showed unimodal distributions but differed with respect to the biocide and the bacterial species investigated. Although there were no indications of a development of biocide resistance, some P. aeruginosa isolates exhibited benzalkonium MICs higher than the highest test concentration.

Antimicrobial and Biocide Resistance among Feline and Canine Staphylococcus aureus and Staphylococcus pseudintermedius Isolates from Diagnostic Submissions.

A total of 114 Staphylococcus isolates from various infections of companion animals, including 43 feline Staphylococcus aureus, 19 canine S. aureus, 11 feline Staphylococcus pseudintermedius and 41 canine S. pseudintermedius were investigated for (i) their susceptibility to 24 antimicrobial agents and three combinations of antimicrobial agents by broth microdilution following CLSI recommendations and (ii) the corresponding resistance genes. In addition, the isolates were tested for their susceptibility to the four biocides benzalkonium chloride, chlorhexidine, polyhexanide and octenidine by a recently developed biocide susceptibility testing protocol. Penicillin resistance via blaZ was the dominant resistance property in all four groups of isolates ranging between 76.7 and 90.9%. About one quarter of the isolates (25.4%) proved to be methicillin-resistant and carried the genes mecA or mecC. Macrolide resistance was the second most prevalent resistance property (27.2%) and all isolates harbored the resistance genes erm(A), erm(B), erm(C), erm(T) or msr(A), alone or in combinations. Fluoroquinolone resistance was detected in 21.1% of all isolates tested, whereas tetracycline resistance via tet(K) and/or tet(M) occurred in 19.3% of the isolates. Resistance to last resort antimicrobial agents in human medicine was seen only in single isolates, if at all. The minimal inhibitory concentrations (MICs) of the four biocides showed unimodal distributions and were very similar for the four groups of staphylococci. Because of the large number of (multi)resistant isolates, antimicrobial susceptibility testing of feline and canine S. aureus and S. pseudintermedius isolates is highly recommended before the start of an antimicrobial chemotherapy. Moreover, no hints towards the development of biocide resistance were detected.

Molecular detection of quaternary ammonium compound resistance genes of Staphylococcus aureus from udder surface and mastitis milk of bovines

Quaternary ammonium compounds (QAC) are widely used biocides in the field of medical and veterinary practice. Resistance to QACs is an emerging problem due to this widespread use. The present study was undertaken to detect the presence of genes that contribute to resistance to QACs in Staphylococcus aureus isolates obtained from udder surface and mastitis milk. A total 22 isolates of S. aureus were obtained from udder washings and milk of bovine mastitis cases. Among these, seventeen isolates were from udder wash samples and five isolates from clinical and subclinical cases of mastitis. Broth microdilution assay was performed to assess the minimum inhibitory concentration (MIC) of the isolates against Cetyltrimethylammonium bromide (CTAB), a QAC. Polymerase chain reaction (PCR) was done targeting qac A/B gene which codes for efflux pump which targets QACs. In the present study qac A/B gene was detected in seven out of 22 isolates of S. aureus. Three of these isolates were obtained from udder washings before milking and four, after milking. None of the S. aureus isolates from mastitis milk possessed the gene. The average minimum inhibitory concentration of qac A/B positive isolates against CTAB was 0.63 ± 0.55 μg/ml. The average MIC between qac A/B positive and negative isolates were statistically insignificant. The study points to the fact that multiple factors could be contributing to biocide resistance in S. aureus.

Exploring the Co-Occurrence of Antibiotic, Metal, and Biocide Resistance Genes in the Urban Agricultural Environment

Exploring the Co-Occurrence of Antibiotic, Metal, and Biocide Resistance Genes in the Urban Agricultural Environment

Genomic characterization of multidrug-resistant extraintestinal pathogenic Escherichia coli isolated from grain culture soils

Multidrug-resistant (MDR) Escherichia coli , mainly extraintestinal pathogenic E. coli (ExPEC), has been widely reported in infections worldwide. In agricultural soils, manure is a hotspot for the dissemination of antimicrobial resistance genes (ARGs) and pathogenic bacteria; however, MDR bacteria have also been reported in soils with no history of manure use. In addition, cross-resistance and co-resistance have been described as responsible for the metal-driven selection of bacteria resistant to antimicrobials. Therefore, the aim of this study was to analyze three MDR E. coli isolates obtained from Brazilian grain culture soil samples with no history of manure use by whole-genome sequencing. The MDR E. coli isolates were recovered from soils from corn and coffee fields, and presented resistance to β-lactams, quinolones, aminoglycosides, tetracyclines, sulphonamides, and dihydrofolate reductase inhibitor. Resistome analysis showed ARGs to several antimicrobials ( i.e. , β-lactams, tetracyclines, aminoglycosides, sulphonamides, trimethoprim, phenicols, fosfomycin, and macrolides) as well as several metal resistance genes and antibacterial biocide resistance genes. In addition, known mutations in quinolone-resistance-determining regions of GyrA (Ser83Leu and Asp87Asn), ParE (Ser458Thr), and ParC (Ser80Ile) were also detected. Virulome analysis showed the presence of virulence genes ( lpfA , mcmA , gad , mchF , iroN , cma , and iss ) associated with ExPEC. Multidrug-resistant ExPEC isolates were assigned to phylogenetic group B1. The presence of MDR B1-ExPEC in soil samples shows the ability of these isolates to survive in soils. This study reports for the first time some sequence types ( i.e. , ST345, ST448, and ST1146) of MDR E. coli in Brazilian soils. Therefore, these findings contribute to the monitoring of antimicrobial resistance and surveillance studies based on whole-genome sequencing worldwide.

Metagenomic profiles of the resistome in subtropical estuaries: Co-occurrence patterns, indicative genes, and driving factors

Estuaries are resistome hotspots owing to resistome accumulation and propagation at these locations from surrounding rivers, yet the large-scale biogeographic pattern of resistome, especially biocide and metal resistance genes (BMRGs) and its driving mechanisms in estuarine waters remains to be elucidated. Here, a metagenomics-based approach was firstly used to investigate resistome and mobilome profiles in waters from 30 subtropical estuaries, South China. The Pearl River estuaries had a higher diversity and abundance of antibiotic resistance genes (ARGs), BMRGs, and mobile genetic elements (MGEs) when compared with estuaries from east and west regions. Genes resistant to multiple antibiotics, metals, and biocides were the most abundant gene types in the resistome. The abundance of MGEs (e.g., intI1, IS91, and tnpA) was highly associated with the total abundance of resistance genes, suggesting their utility as potential indicators for quantitative estimations of the resistome contamination. Further, MGEs contributed more than bacterial communities in shaping the resistome in subtropical estuaries. Physicochemical factors (e.g., pH) regulated MGE composition and stochastic assembly, which mediated the co-selection of ARGs and BMRGs via horizontal gene transfer. Our findings have important implications and provide a reference on the management of ARGs and BMRGs in subtropical estuarine ecosystems.

Cropping system exerts stronger influence on antibiotic resistance gene assemblages in greenhouse soils than reclaimed wastewater irrigation

The effects of reclaimed wastewater (RW) irrigation on the spread of antibiotic resistance genes (ARGs) in soil is modulated by a myriad of biotic and abiotic factors and their relative significance remains vague. We compared microbial communities, assemblages of genes associated with microbial resistance to antibiotics, biocides and metals, and insertion sequences (ISs) in soils following 16 years of irrigation with groundwater (GW), RW or alternately with GW and RW in two greenhouses with different cropping systems, using shotgun metagenome sequencing. The results showed that cropping system exerted greater influence than irrigation on the profile of ISs and resistance genes. This influence was most strongly associated with concentrations of copper, mercury and perfloxacin in the soils. There was no significant difference in soil ARG profiles between continuous RW irrigation and alternating GW and RW irrigation. Proteobacteria, Actinobacteria and Firmicutes and a limited number of ISs were closely associated with the detected ARGs. Most ARGs were found to co-occur with metal and biocide resistance genes through the mechanism of efflux pumps. These findings highlight the significance of understanding and improving crop management in mitigating the dissemination of ARGs in soils irrigated with RW.

Prevalence of resistance genes to biocides in antibiotic-resistant Pseudomonas aeruginosa clinical isolates

Biocides are frequently used as preservative, disinfectant and sterilizer against many microorganisms in hospitals, industry and home. However, the reduced susceptibility rate of Pseudomonas aeruginosa (P. aeruginosa) strains to biocides is increasing. The aim of this study was to evaluate the antimicrobial activity of four frequently used biocides against P. aeruginosa and to determine the prevalence of genes involved in biocide resistance. A total of 76 clinical isolates of P. aeruginosa strains were used in the present study. The minimum inhibitory concentrations (MICs) of four biocides, i.e. chlorhexidine digluconate, benzalkonium chloride, triclosan and formaldehyde, against P. aeruginosa strains were determined using agar dilution method. In addition, the prevalence of biocide resistance genes was determined using the polymerase chain reaction (PCR) method. In the present study, the highest MIC90 and MIC95 (epidemiological cut-off) values were observed for benzalkonium chloride (1024μg/mL), followed by formaldehyde (512μg/mL), triclosan (512μg/mL) and chlorhexidine digluconate (64μg/mL). Furthermore, the prevalence of qacEΔ1, qacE, qacG, fabV, cepA and fabI genes were 73.7% (n = 56), 26.3% (n = 20), 11.8% (n = 9), 84.2% (n = 64), 81.5% (n = 62) and 0% (n = 0), respectively. A significant association was observed between the presence of biocide resistance genes and MICs (p < 0.05). Furthermore, there was no significant association between the presence of biocide resistance genes and antibiotic resistance (p > 0.05), except for levofloxacin and norfloxacin antibiotics and qacE and qacG genes (p < 0.05). Our results revealed that chlorhexidine digluconate is the most effective biocide against P. aeruginosa isolates in Ardabil hospitals. However, we recommend continuous monitoring of the antimicrobial activity of biocides and the prevalence of biocide-associated resistance genes for a better prevention of microorganism dissemination and infection control in hospitals.