Heavy Metal Resistance Genes Research Articles

Reforestation substantially changed the soil antibiotic resistome and its relationships with metal resistance genes, mobile genetic elements, and pathogens

Revealing the effects of reforestation on soil antibiotic resistome is essential for assessing ecosystem health, yet related studies remain scarce. Here, to determine the responses of the soil antibiotic resistome to reforestation, 30 pairs of cropland and forest soil samples were collected from southwestern China, a region with high environmental heterogeneity. All the forests had been derived from croplands more than one decade ago. The diversity and abundance of soil antibiotic resistance genes (ARGs), metal resistance genes (MRGs), mobile genetic elements (MGEs), and pathogens were determined by metagenomic sequencing and real-time PCR. The results showed that reforestation significantly increased soil microbial abundance and the contents of Cu, total carbon, total nitrogen, total organic carbon, and ammonium nitrogen. Nevertheless, it decreased the contents of soil Zn, Ba, nitrate nitrogen, and available phosphorus. The main soil ARGs identified in this region were vancomycin, multidrug, and bacitracin resistance genes. Reforestation significantly increased the soil ARG abundance by 62.58%, while it decreased the ARG richness by 16.50%. Reforestation exerted no significant effects on the abundance of heavy metal resistance genes and pathogens, but it doubled the abundance of MGEs. Additionally, reforestation substantially decreased the co-occurrence frequencies of ARGs with MRGs and pathogens. In contrast, the correlation between ARGs and MGEs was greatly enhanced by reforestation. Similarly, the correlations between soil ARG abundance and environmental factors were also strengthened by reforestation. These findings suggest that reforestation can substantially affect the soil antibiotic resistome and exerts overall positive effects on soil health by decreasing ARG richness, providing critical information for assessing the effects of “grain for green” project on soil health.

Baseline metagenome-assembled genome (MAG) data of Sikkim hot springs from Indian Himalayan geothermal belt (IHGB) showcasing its potential CAZymes, and sulfur-nitrogen metabolic activity.

Here we present the construction and characterization of metagenome assembled genomes (MAGs) from two hot springs residing in the vicinity of Indian Himalayan Geothermal Belt (IHGB). A total of 78 and 7 taxonomic bins were obtained for Old Yume Samdong (OYS) and New Yume Samdong (NYS) hot springs respectively. After passing all the criteria only 21 and 4 MAGs were further studied based on the successful prediction of their 16S rRNA. Various databases were used such as GTDB, Kaiju, EzTaxon, BLAST XY Plot and NCBI BLAST to get the taxonomic classification of various 16S rRNA predicted MAGs. The bacterial genomes found were from both thermophilic and mesophilic bacteria among which Proteobacteria, Chloroflexi, Bacteroidetes and Firmicutes were the abundant phyla. However, in case of OYS, two genomes belonged to archaeal Methanobacterium and Methanocaldococcus. Functional characterization revealed the richness of CAZymes such as Glycosyl Transferase (GT) (56.7%), Glycoside Hydrolase (GH) (37.4%), Carbohydrate Esterase family (CE) (8.2%), and Polysaccharide Lyase (PL) (1.9%). There were negligible antibiotic resistance genes in the MAGs however, a significant heavy metal tolerance gene was found in the MAGs. Thus, it may be assumed that there is no coexistence of antibiotic and heavy metal resistance genes in these hot spring microbiomes. Since the selected hot springs possess good sulfur content thus, we also checked the presence of genes for sulfur and nitrogen metabolism. It was found that MAGs from both the hot springs possess significant number of genes related to sulfur and nitrogen metabolism.

Characterization and genome analysis of Acinetobacter oleivorans S4 as an efficient hydrocarbon-degrading and plant-growth-promoting rhizobacterium

Plant-growth-promoting rhizobacteria (PGPR) have received increasing attention for assisting phytoremediation. However, the effect of PGPR on total petroleum hydrocarbon (TPH) degradation and plant growth promotion and its underlying mechanism is not well understood. In this study, phenotypic analysis and whole genome sequencing were conducted to comprehensively characterize a newly isolated rhizobacterium strain S4, which was identified as Acinetobacter oleivorans, from a TPH-contaminated soil. The strain degraded 62.5% of initially spiked diesel (1%) in minimal media within six days and utilized n-alkanes with a wide range of chain length (i.e., C12 to C40). In addition, the strain showed phenotypic traits beneficial to plant growth, including siderophore production, indole-3-acetic acid synthesis and phosphate solubilization. Potential metabolic pathways and genes encoding proteins responsible for the phenotypic traits were identified. In a real TPH-contaminated soil, inoculation of Acinetobacter oleivorans S4 significantly enhanced the growth of tall fescue relative to the soil without inoculation. In contrast, inoculation of Bacillus sp. Z7, a hydrocarbon-degrading strain, showed a negligible effect on the growth of tall fescue. The removal efficiency of TPH with inoculation of Acinetobacter oleivorans S4 was significantly higher than those without inoculation or inoculation of Bacillus sp. Z7. These results suggested that traits of PGPR beneficial to plant growth are critical to assist phytoremediation. Furthermore, heavy metal resistance genes and benzoate and phenol degradation genes were found in the genome of Acinetobacter oleivorans S4, suggesting its application potential in broad scenarios.

Effect of Cu addition on sedimentary bacterial community structure and heavy metal resistance gene abundance in mangrove wetlands

IntroductionMangroves are a unique ecosystem, but are often affected by heavy metal pollution. Bacterial communities in mangrove sediments usually respond to heavy-metal pollution stress by expressing specific heavy-metal resistance genes (HMRGs).MethodsMultivariate statistics were performed on Metagenomics as well as environmental data to evaluated the effects of Cu addition on the sedimentary bacterial community structure.Results and discussionUsing metagenomics technology, we found that the Shannon andChao1 indices of natural sediments were significantly higher than in sediments with artificially added Cu. Addition of Cu significantly altered bacterial community structure and diversity at the phylum, class and order levels in sediments. The relative abundance of HMRGs in Cuadded sediments was lower than that in natural sediments. Environmental factors that affect bacterial communities also affect HMRGs. However, the contribution of heavy-metal pollution to the change in HMRG abundance was much lower than its contribution to the change in the bacterial community, indicating that the relationship between HMRG abundance and heavy-metal pollution is complex and needs to be comprehensively studied.

Comparative genomic analysis of Colistin resistant Escherichia coli isolated from pigs, a human and wastewater on colistin withdrawn pig farm

In this study, genomic and plasmid characteristics of Escherichia coli were determined with the aim of deducing how mcr genes may have spread on a colistin withdrawn pig farm. Whole genome hybrid sequencing was applied to six mcr-positive E. coli (MCRPE) strains isolated from pigs, a farmworker and wastewater collected between 2017 and 2019. Among these, mcr-1.1 genes were identified on IncI2 plasmids from a pig and wastewater, and on IncX4 from the human isolate, whereas mcr-3 genes were found on plasmids IncFII and IncHI2 in two porcine strains. The MCRPE isolates exhibited genotypic and phenotypic multidrug resistance (MDR) traits as well as heavy metal and antiseptic resistance genes. The mcr-1.1-IncI2 and IncX4 plasmids carried only colistin resistance genes. Whereas, the mcr-3.5-IncHI2 plasmid presented MDR region, with several mobile genetic elements. Despite the MCRPE strains belonged to different E. coli lineages, mcr-carrying plasmids with high similarities were found in isolates from pigs and wastewater recovered in different years. This study highlighted that several factors, including the resistomic profile of the host bacteria, co-selection via adjunct antibiotic resistance genes, antiseptics, and/or disinfectants, and plasmid-host fitness adaptation may encourage the maintenance of plasmids carrying mcr genes in E. coli.

Transmission mechanisms of antibiotic resistance genes in arsenic-contaminated soil under sulfamethoxazole stress

Numerous studies have revealed the spread mechanism of antibiotic resistance genes (ARGs) in single antibiotic-contaminated soils. However, the comprehensive impacts of heavy metals and antibiotics on ARGs and the underlying mechanisms are still unknown. Here, high-throughput quantitative PCR and high-throughput sequencing were used to investigate changes in ARGs and bacterial communities under various sulfamethoxazole (SMX) regimes (0, 1, 10, 50 mg kg−1) in arsenic (As) contaminated soils. The study found that the abundances of ARGs, mobile genetic elements (MGEs), and heavy metal resistance genes (HMRGs) significantly increased in the soil fortified at 10 and 50 mg kg−1 SMX concentrations. The ARGs abundance increased with the increase in the MGEs abundance. Many significant positive correlations between various ARGs subtypes and HMRGs subtypes were found. These results indicate that the HMRGs and MGEs positively contributed to the enrichment of ARGs in As-contaminated soils under SMX stress. Meanwhile, the abundance of copiotrophic (Actinobacteriota) reduced and oligotrophic (Gemmatimonadota) increased, indicating that the life history strategy of the community changed. In addition, Gemmatimonadota was positively correlated to ARGs, HMRGs, and MGEs, suggesting that Gemmatimonadota, which can cope with As and SMX stress, was the host for resistance genes in the soil. Finally, the study found that MGEs play a determinant role in ARGs proliferation due to the direct utilization of HGT, and the indirect effect for ARGs spread under a co-selection mechanism of ARGs and HMRGs, while the bacterial community showed indirect influences by altering environmental factors to act on MGEs. Collectively, this study revealed new insights into the mechanisms of resistance gene transmission under combined SMX and As contamination in soil ecosystems.

Stingray epidermal microbiomes are species-specific with local adaptations.

Marine host-associated microbiomes are affected by a combination of species-specific (e.g., host ancestry, genotype) and habitat-specific features (e.g., environmental physiochemistry and microbial biogeography). The stingray epidermis provides a gradient of characteristics from high dermal denticles coverage with low mucus to reduce dermal denticles and high levels of mucus. Here we investigate the effects of host phylogeny and habitat by comparing the epidermal microbiomes of Myliobatis californica (bat rays) with a mucus rich epidermis, and Urobatis halleri (round rays) with a mucus reduced epidermis from two locations, Los Angeles and San Diego, California (a 150 km distance). We found that host microbiomes are species-specific and distinct from the water column, however composition of M. californica microbiomes showed more variability between individuals compared to U. halleri. The variability in the microbiome of M. californica caused the microbial taxa to be similar across locations, while U. halleri microbiomes were distinct across locations. Despite taxonomic differences, Shannon diversity is the same across the two locations in U. halleri microbiomes suggesting the taxonomic composition are locally adapted, but diversity is maintained by the host. Myliobatis californica and U. halleri microbiomes maintain functional similarity across Los Angeles and San Diego and each ray showed several unique functional genes. Myliobatis californica has a greater relative abundance of RNA Polymerase III-like genes in the microbiome than U. halleri, suggesting specific adaptations to a heavy mucus environment. Construction of Metagenome Assembled Genomes (MAGs) identified novel microbial species within Rhodobacteraceae, Moraxellaceae, Caulobacteraceae, Alcanivoracaceae and Gammaproteobacteria. All MAGs had a high abundance of active RNA processing genes, heavy metal, and antibiotic resistant genes, suggesting the stingray mucus supports high microbial growth rates, which may drive high levels of competition within the microbiomes increasing the antimicrobial properties of the microbes.

Unexpected enrichment of antibiotic resistance genes and organic remediation genes in high-altitude lakes at Eastern Tibetan Plateau

Elevation has a strong effect on aquatic microbiome. However, we know little about the effects of elevation on functional genes, especially antibiotic resistance genes (ARGs) and organic remediation genes (ORGs) in freshwater ecosystems. In this study, we analyzed five classes of functional genes including ARGs, metal resistance genes (MRGs), ORGs, bacteriophages, and virulence genes between two high-altitude lakes (HALs) and two low-altitude lakes (LALs) in Mountain Siguniang at Eastern Tibetan Plateau by means of GeoChip 5.0. No differences (Student's t-test, p > 0.05) of gene richness including ARGs, MRGs, ORGs, bacteriophages, and virulence genes in HALs and LALs were found. The abundance of most ARGs and ORGs was higher in HALs than in LALs. For MRGs, the abundance of macro metal resistance genes of potassium, calcium, and aluminum was higher in HALs than in LALs (Student's t-test, p < 0.05; all Cohen's d > 0.8). The abundance of some heavy metal resistance genes of lead and mercury was lower in HALs than in LALs (Student's t-test, p < 0.05; all Cohen's d < −0.8). The composition of these functional genes in HALs differed significantly from in LALs. The functional gene network in HALs was also more complex than that in LALs. We speculate that enrichment of ARGs and ORGs in HALs is related to different microbial communities, exogenous ARGs, and enriched persistent organic pollutants through long-range atmospheric transport driven by the Indian monsoon. This study highlights the unexpected enrichment of ARGs, MRGs, and ORGs in remote lakes at high elevations.

Sewage-based surveillance shows presence of Klebsiella pneumoniae resistant against last resort antibiotics in the population in Bergen, Norway.

The aim of this study was to understand the prevalence of antibiotic resistance in Klebsiella pneumoniae present in the population in Bergen city, Norway using city-scale sewage-based surveillance, as well as the potential spread of K. pneumoniae into the marine environment through treated sewage. From a total of 30 sewage samples collected from five different sewage treatment plants (STPs), 563 presumptive K. pneumoniae isolates were obtained on Simmons Citrate Agar with myo-Inositol (SCAI) plates, and 44 presumptive K. pneumoniae isolates on SCAI plates with cefotaxime. Colistin resistance was observed in 35 isolates, while cefotaxime resistance and tigecycline resistance was observed in only five isolates each, out of 563 presumptive K. pneumoniae isolates. All 44 isolates obtained on cefotaxime-containing plates were multidrug-resistant, with 25% (n=11) showing resistance against tigecycline. Clinically important acquired antibiotic resistance genes (ARGs), like blaCTX-M-14, blaCTX-M-15, qnrS1, aac(3)-IIe, tet(A), and sul1, were detected in several sequenced Klebsiella spp. isolates (n=53). All sequenced colistin-resistant isolates (n=13) had a mutation in the mgrB gene with nucleotide substitution at position C88T creating a premature stop codon. All sequenced tigecycline-resistant isolates (n=4) harbored a Tet(A) variant with 22 amino acid (aa) substitutions compared to the reference protein. The sequenced K. pneumoniae isolates (n=44) belonged to 22 different sequence types (STs) with ST730 (29.5%) as most prevalent, followed by pathogenic ST307 (11.4%). Virulence factors, including aerobactin (iutA), enterobactin (entABCDEFS and fepABCDG), salmochelin (iro), and yersiniabactin (ybt) were detected in several sequenced K. pneumoniae isolates, suggesting pathogenicity potential. Heavy metal resistance genes were common in sequenced K. pneumoniae isolates (n=44) with silver (silABCEFPRS) and copper (pcoABDRS) resistance genes present in 79.5% of the isolates. Sewage-based surveillance can be a useful tool for understanding antibiotic resistance in pathogens present within a population and to provide up-to date information on the current resistance situation. Our study presents a framework for population-based surveillance of resistance in K. pneumoniae.

Coal-source acid mine drainage reduced the soil multidrug-dominated antibiotic resistome but increased the heavy metal(loid) resistome and energy production-related metabolism

A recent global scale study found that mining-impacted environments have multi-antibiotic resistance gene (ARG)-dominated resistomes with an abundance similar to urban sewage but much higher than freshwater sediment. These findings raised concern that mining may increase the risk of ARG environmental proliferation. The current study assessed how typical multimetal(loid)-enriched coal-source acid mine drainage (AMD) contamination affects soil resistomes by comparing with background soils unaffected by AMD. Both contaminated and background soils have multidrug-dominated antibiotic resistomes attributed to the acidic environment. AMD-contaminated soils had a lower relative abundance of ARGs (47.45 ± 23.34 ×/Gb) than background soils (85.47 ± 19.71 ×/Gb) but held high-level heavy metal(loid) resistance genes (MRGs, 133.29 ± 29.36 ×/Gb) and transposase- and insertion sequence-dominated mobile genetic elements (MGEs, 188.51 ± 21.81 ×/Gb), which was 56.26 % and 412.12 % higher than background soils, respectively.Procrustes analysis showed that the microbial community and MGEs exerted more influence on driving heavy metal(loid) resistome variation than antibiotic resistome. The microbial community increased energy production-related metabolism to fulfill the increasing energy needs required by acid and heavy metal(loid) resistance. Horizontal gene transfer (HGT) events primarily exchanged energy- and information-related genes to adapt to the harsh AMD environment. These findings provide new insight into the risk of ARG proliferation in mining environments.

Metagenome sequencing to unveil the occurrence and distribution of antibiotic resistome and in a wastewater treatment plant

ABSTRACT The emergence and persistence of antibiotic resistance genes (ARGs) in wastewater treatment plants (WWTPs) has aroused growing public concern for its risk to human health and ecological safety. Moreover, heavy metals concentrated in sewage and sludge could potentially favour co-selection of ARGs and heavy metal resistance genes (HMRGs). In this study, the profile and abundance of antibiotic and metal resistance genes in influent, sludge and effluent were characterized based on the Structured ARG Datebase (SARG) and Antibacterial Biocide and Metal Resistance Gene Datebase (BacMet) by metagenomic analysis. Sequences were aligning against the INTEGRALL, ISFinder, ICEberg and NCBI RefSeq databases to obtain the diversity and abundance of mobile genetic elements (MGEs, e.g.plasmid and transposon). Among them, 20 types of ARGs and 16 types of HMRG were detected in all samples, the influent metagenomes contained many more resistance genes (both ARGs and HMRGs) than the sludge and the influent sample, large reductions in the relatively abundance and diversity of ARG were achieved by biological treatment. ARGs and HMRGs cannot be completely eliminated during the oxidation ditch. A total of 32 species of the potential pathogens were detected, relative abundances of pathogens had no obvious changes. It is suggested that more specific treatments are required to limit their proliferation in the environment. This study can be helpful for further understanding the removal of antibiotic resistance genes in the sewage treatment process via metagenomic sequencing.

Effects of co-selection of antibiotic-resistance and metal-resistance genes on antibiotic-resistance potency of environmental bacteria and related ecological risk factors

The inadequate elimination of micropollutants in wastewater treatment plants (WWTP), cause to increase in the incidence of antibiotic resistant bacterial strains. Growth of microbial pathogens in WWTP is one of the serious public health problems. The widespread and simultaneous emergence of antibiotic resistance genes (ARGs) and heavy metal resistance genes (HMRGs) in the environment with heavy metals create persistent and selective pressure for co-selection of both genes on environmental microorganisms. Co-localization of ARGs and HMRGs on the same horizontal mobile genetic elements (MGEs) allows the spreading of numerous antibiotic-resistant strains of bacteria in aquatic and terrestrial environment. The biofilm formation and colonization potential of environmental bacteria leads to the co-selection of multi-antibiotic resistance and multi-metal tolerance. Horizontal gene transfer (HGT), co-localization of both ARGs and HMRGs on the same MGEs, and the shared resistomes are important bacteria-associated ecological risks factors, which reduce the effectiveness of antibiotics against bacterial infections.

Comparative genomics reveals the acquisition of mobile genetic elements by the plant growth-promoting Pantoea eucrina OB49 in polluted environments

Heavy metal-tolerant plant growth-promoting bacteria (PGPB) have gained popularity in bioremediation in recent years. A genome-assisted study of a heavy metal-tolerant PGPB Pantoea eucrina OB49 isolated from the rhizosphere of wheat grown on a heavy metal-contaminated site is presented. Comparative pan-genome analysis indicated that OB49 acquired heavy metal resistance genes through horizontal gene transfer. On contigs S10 and S12, OB49 has two arsRBCH operons that give arsenic resistance. On the S12 contig, an arsRBCH operon was discovered in conjunction with the merRTPCADE operon, which provides mercury resistance. P. eucrina OB49 may be involved in an ecological alternative for heavy metal remediation and growth promotion of wheat grown in metal-polluted soils. Our results suggested the detection of mobile genetic elements that harbour the ars operon and the fluoride resistance genes adjacent to the mer operon.

Detection of virulence, antimicrobial resistance, and heavy metal resistance properties in Vibrio anguillarum isolated from mullet (Mugil cephalus) cultured in Korea.

In the present study, we identified and characterized 22 strains of V. anguillarum from 145 samples of mullets (Mugill cephallus) cultured in several fish farms in South Korea. They were subjected to pathogenicity tests, antimicrobial susceptibility test, and broth dilution test to detect virulence markers, antimicrobial resistance, and heavy metal resistance properties. All the isolates showed amylase and caseinase activity, followed by gelatinase (90.9%), DNase (45.5%), and hemolysis activities (α = 81.1% and β = 18.2%). The PCR assay revealed that isolates were positive for VAC, ctxAB, AtoxR, tdh, tlh, trh, Vfh, hupO, VPI, and FtoxR virulence genes at different percentages. All the isolates showed multi-drug resistance properties (MAR index ≥ 0.2), while 100% of the isolates were resistant to oxacillin, ticarcillin, streptomycin, and ciprofloxacin. Antimicrobial resistance genes, qnrS (95.5%), qnrB (86.4%), and StrAB (27.3%), were reported. In addition, 40.9% of the isolates were cadmium-tolerant, with the presence of CzcA (86.4%) heavy metal resistance gene. The results revealed potential pathogenicity associated with V. anguillarum in aquaculture and potential health risk associated with consumer health.

Composting reduces the risks of antibiotic resistance genes in maize seeds posed by gentamicin fermentation waste

Using high-throughput quantitative PCR and next generation sequencing, the impact of land application of raw and composted gentamicin fermentation waste (GFW) on antibiotic resistance genes (ARGs) in maize seeds was studied in a three-year field trial. The raw and composted GFW changed both the bacterial community composition and the ARGs diversity in the maize seeds compared to non-amended controls and chemical fertilizer. The abundance of ARGs after raw GFW amendment was significantly higher than other treatments because of a high abundance of aadA1, qacEdeltal and aph(2′)-Id-02; probably induced by gentamicin selection pressure in maize tissues. Meanwhile, the potential host of these three ARGs, pathogenic bacteria Tenacibaculum, also increased significantly in maize seeds after the application of raw GFW. But our result proved that composting could weaken the risk posed by GFW. We further reveal that the key biotic driver for shaping the ARG profiles in maize seeds is bacterial community followed by heavy metal resistance genes, and ARGs are more likely located on bacterial chromosomes. Our findings provide new insight into ARGs dispersal mechanism in maize seeds after long-term GFW application, demonstrate the potential benefits of composting the GFW to reduce risks as well as the potential efficient management method to GFW.

Metagenomic analysis characterizes resistomes of an acidic, multimetal(loid)-enriched coal source mine drainage treatment system

Heavy metal(loid) contaminations caused by mine activities are potential hot spots of antibiotic resistance genes (ARGs) because of heavy metal(loid)-induced co-selection of ARGs and heavy metal(loid) resistance genes (MRGs). This study used high-throughput metagenomic sequencing to analyze the resistome characteristics of a coal source acid mine drainage passive treatment system. The multidrug efflux mechanism dominated the antibiotic resistome, and a highly diverse heavy metal(loid) resistome was dominated by mercury-, iron-, and arsenic--associated resistance. Correlation analysis indicated that mobile gene elements had a greater influence on the dynamic of MRGs than ARGs. Among the metagenome-assembled genomes, six potential pathogens carrying multiple resistance genes resistant to several antibiotics and heavy metal(loid)s were recovered. Pseudomonas spp. contained the highest numbers of resistance genes, with resistance to two types of antibiotics and 12 types of heavy metal(loid)s. Thus, high contents of heavy metal(loid)s drove the co-selection of ARGs and MRGs. The occurrence of potential pathogens containing multiple resistance genes might increase the risk of ARG dissemination in the environment.

Characterization of Avian Pathogenic Escherichia coli Isolated from Broiler Breeders with Colibacillosis in Mississippi

Avian pathogenic Escherichia coli (APEC) causes colibacillosis in poultry, a leading cause of poultry mortality worldwide. It is crucial to control APEC in broiler breeders as it is vertically transferred to progeny via eggs. However, there is only limited knowledge on the current APEC population in breeders. This study characterized 28 APEC strains isolated from broiler breeders with colibacillosis. The genotypic-virulence characteristics as well as antimicrobial and heavy-metal resistance patterns of the isolates were determined. Results showed that O88 is the most prevalent serogroup and B2 is the predominant phylogenetic group. Among virulence genes, genes for iron acquisition (iroN and iutA), protectins (iss and ompT), and toxin production (hlyF) exhibited the highest prevalence. Further, 93% of the isolates carried at least one antimicrobial resistance gene with highest prevalence for tetracycline gene tetA. Among the isolates, 10.71% exhibited multidrug resistance. All isolates carried at least one heavy-metal resistance gene with the highest prevalence for arsenic gene arsC and the highest resistance towards silver. Our findings provide insight into the characteristics of current APEC populations in broiler breeders in Mississippi. This will help future research on the pathogenesis of APEC and the development of effective prevention and control strategies against APEC in broiler breeders.

In silico analyses of diversity and dissemination of antimicrobial resistance genes and mobile genetics elements, for plasmids of enteric pathogens.

The antimicrobial resistance (AMR) mobilome plays a key role in the dissemination of resistance genes encoded by mobile genetics elements (MGEs) including plasmids, transposons (Tns), and insertion sequences (ISs). These MGEs contribute to the dissemination of multidrug resistance (MDR) in enteric bacterial pathogens which have been considered as a global public health risk. To further understand the diversity and distribution of AMR genes and MGEs across different plasmid types, we utilized multiple sequence-based computational approaches to evaluate AMR-associated plasmid genetics. A collection of 1,309 complete plasmid sequences from Gammaproteobacterial species, including 100 plasmids from each of the following 14 incompatibility (Inc) types: A/C, BO, FIA, FIB, FIC, FIIA, HI1, HI2, I1, K, M, N, P except W, where only 9 sequences were available, was extracted from the National Center for Biotechnology Information (NCBI) GenBank database using BLAST tools. The extracted FASTA files were analyzed using the AMRFinderPlus web-based tools to detect antimicrobial, disinfectant, biocide, and heavy metal resistance genes and ISFinder to identify IS/Tn MGEs within the plasmid sequences. In silico prediction based on plasmid replicon types showed that the resistance genes were diverse among plasmids, yet multiple genes were widely distributed across the plasmids from enteric bacterial species. These findings provide insights into the diversity of resistance genes and that MGEs mediate potential transmission of these genes across multiple plasmid replicon types. This notion was supported by the observation that many IS/Tn MGEs and resistance genes known to be associated with them were common across multiple different plasmid types. Our results provide critical insights about how the diverse population of resistance genes that are carried by the different plasmid types can allow for the dissemination of AMR across enteric bacteria. The results also highlight the value of computational-based approaches and in silico analyses for the assessment of AMR and MGEs, which are important elements of molecular epidemiology and public health outcomes.

Isolation of heavy metal-resistant Staphylococcus epidermidis strain TWSL_22 and evaluation of heavy metal bioremediation potential of recombinant E. coli cloned with isolated cadD.

A heavy metal-resistant bacterial strain, TWSL_22 was isolated from an industrial effluent sample and tested for heavy metal tolerance and resistance. The strain was molecularly characterized as Staphylococcus epidermidis based on 16S rDNA gene analysis and the sequence was deposited in the NCBI repository (accession number KT184893.1). Metal removal activity (P<.001) of TWSL_22 was 99.99±0.001%, 74.43±2.51%, and 51.16±4.17% for Cd, Pb, and Cu, respectively. Highest MIC was observed for Cd. Antibiotic susceptibility assays revealed the strain TWSL_22 to be resistant to several antibiotics. The strain was screened for possible heavy metal-resistant genes and presence of cadA, copA, and cadD was confirmed by PCR. A DNA fragment containing complete sequence of cadD (618bp) was isolated and cloned into pET 21a(+), transformed into E. coli BL21 and designated as E. coli/cadDET. E. coli/cadDET showed high metal tolerance capacity and could remove over 82% of heavy metals (Zn2+, Cd2+, Cu2+, and Cr3+) in the industrial effluent.

Decoding whole genome of Anoxybacillus rupiensis TPH1 isolated from tatapani hot spring, India and giving insight into bioremediation ability of TPH1 via heavy metals and azo dyes

A moderately thermophilic, gram-positive genomospecies Anoxybacillus rupiensis TPH1 was isolated from Tatapani hot spring, Chhattisgarh, India. Genome of 3.70 Mb with 42.3% GC subsumed 4131 CDSs, 65 tRNA, 5 rRNA, 35 AMR and 19 drug target genes. Further, comparative genomics of 19 Anoxybacillus spp. exhibited an open pan genome of 13102 genes along with core (10.62%), unique (43.5%) and accessory (45.9%) genes. Moreover, phylogenomic tree displayed clustering of Anoxybacillus spp. into two distinct clades where clade A species harbored larger genomes, more unique genes, CDS and hypothetical proteins than clade B species. Further, distribution of azoreductases showed FMN-binding NADPH azoreductase (AzoRed1) presence in clade A species only and FMN-binding NADH azoreductase (AzoRed2) harboring by species of both clades. Heavy metal resistance genes distribution showed omnipresence of znuA, copZ and arsC in both clades, dispersed presence of cbiM, czcD, merA and feoB over both clades and harboring of nikA and acr3 by few species of clade A only. Additionally, molecular docking of AzoRed1, AzoRed2, ZnuA, CopZ, Acr3, CbiM, CzcD, MerA and NikA with their respective ligands indicated high affinity and stable binding. Conclusively, present study provided insight into gene repertoire of genus Anoxybacillus and a basis for the potential application of this thermophile in bioremediation of azo dyes and heavy metals.