Tetracycline-resistant Bacteria Research Articles

Elevated proton motive force is a tetracycline resistance mechanism that leads to the sensitivity to gentamicin in Edwardsiella tarda.

Tetracycline is a commonly used human and veterinary antibiotic that is mostly discharged into environment and thereby tetracycline-resistant bacteria are widely isolated. To combat these resistant bacteria, further understanding for tetracycline resistance mechanisms is needed. Here, GC-MS based untargeted metabolomics with biochemistry and molecular biology techniques was used to explore tetracycline resistance mechanisms of Edwardsiella tarda. Tetracycline-resistant E. tarda (LTB4-RTET ) exhibited a globally repressed metabolism against elevated proton motive force (PMF) as the most characteristic feature. The elevated PMF contributed to the resistance, which was supported by the three results: (i) viability was decreased with increasing PMF inhibitor carbonylcyanide-3-chlorophenylhydrazone; (ii) survival is related to PMF regulated by pH; (iii) LTB4-RTET were sensitive to gentamicin, an antibiotic that is dependent upon PMF to kill bacteria. Meanwhile, gentamicin-resistant E. tarda with low PMF are sensitive to tetracycline is also demonstrated. These results together indicate that the combination of tetracycline with gentamycin will effectively kill both gentamycin and tetracycline resistant bacteria. Therefore, the present study reveals a PMF-enhanced tetracycline resistance mechanism in LTB4-RTET and provides an effective approach to combat resistant bacteria.

An electron transfer tuning strategy for the efficient degradation of tetracycline hydrochloride by Fe-N co-doped carbon materials activated with peroxymonosulfate

In order to address the problems of metal leaching and low electron transfer efficiency, the iron (Fe) nitrogen (N) co-doped MOFs derived carbon catalysts presented a promising strategy to activate peroxymonosulfate (PMS) for water purification. In this work, using the MOF-5 as a template and iron phthalocyanine (FePc) and dicyandiamide (DCD) as a source, Fe, N co-doped carbons were prepared through milling and two-step pyrolysis. The catalytic performance and mechanism of the materials on tetracycline hydrochloride (TC) degradation in the PMS activation system (0.1Fe/N-CPMS) was systematically investigated. The results showed that the stable framework structure of MOFs-derived carbon conduced to encapsulate the Fe nanoparticles within the carbon skeleton, and then the xFe/N-C catalyst with hierarchical pore structure and good electrical conductivity was obtained, which significantly improved the TC catalytic performance. Notably, the 0.1Fe/N-C exhibited the greatest degradation performance achieving a degradation rate of 0.6315 min-1, which was approximately five times higher than that of N-C. Moreover, the degradation performance of the 0.1Fe/N-C kept the stability in various interference conditions, and still remained over 80% after three cycles test. Combine with the characterization and DFT calculation, the mechanism of the TC degradation in 0.1Fe/N-CPMS system was revealed by a non-radical process dominated by electron transfer, and the degradation efficiency was greatly improved by the synergistic effect of Fe/N on the as-prepared material. The received findings provide an electron transfer tuning strategy of Fe/N co-doped carbon nanomaterials synthesis for promoting the non-radical degradation pathways in the application of organic pollutants removal.

Fungus reduces tetracycline-resistant genes in manure treatment by predation of bacteria

New strategies to remove antibiotic resistance genes (ARGs), one of the most pressing threats to public health, are urgently needed. This study showed that the fungus Phanerochaete chrysosporium seeded to a composting reactor (CR) could remarkably reduce tetracycline-resistant genes (TRGs). The reduction efficiencies for the five main TRGs (i.e., tetW, tetO, tetM, tetPA, and tet(32)) increased by 8 to 100 folds compared with the control without P. chrysosporium, and this could be attributed to the decrease in the quantity of bacteria. Enumeration based on green fluorescence protein labeling further showed that P. chrysosporium became dominant in the CR. Meanwhile, the bacteria in the CR invaded the fungal cells via the cell wall defect of chlamydospore or active invasion. Most of the invasive bacteria trapped inside the fungus could not survive, resulting in bacterial death and the degradation of their TRGs by the fungal nucleases. As such, the predation of tetracycline-resistant bacteria by P. chrysosporium was mainly responsible for the enhanced removal of TRGs in the swine manure treatment. This study offers new insights into the microbial control of ARGs.

Effect of veterinary feed directive rule changes on tetracycline-resistant and erythromycin-resistant bacteria (Salmonella, Escherichia, and Campylobacter) in retail meats in the United States.

Antimicrobial-resistant bacteria are a growing public health threat. In 2017 the U.S. Food and Drug Administration implemented Veterinary Feed Directive (VFD) rules changes to limit medically important antimicrobial use in food-producing animals, combating antimicrobial-resistant bacteria. The effect of the VFD rule changes on the occurrence of bacteria resistant to medically-important antimicrobials in retail meats is yet to be investigated in the U.S. This study investigates whether the VFD rule changes affected the occurrence of tetracycline-resistant and erythromycin-resistant bacteria (Salmonella, Escherichia, and Campylobacter) in retail meats in the U.S. Multivariable mixed effect logistic regression models were used to analyze 2002-2019 retail meats surveillance data from the National Antimicrobial Resistance Monitoring System (NARMS) in the U.S. Variables included VFD rule changes, meat type, quarter of year, and raising claims. A potential association between these variables and the occurrence of tetracycline-resistant and erythromycin-resistant bacteria (Salmonella, Escherichia, and Campylobacter) in retail meats was estimated. Analysis included data regarding tetracycline-resistant Salmonella (n = 8,501), Escherichia (n = 20, 283), Campylobacter (n = 9,682), and erythromycin-resistant Campylobacter (n = 10,446) in retail meats. The odds of detecting tetracycline-resistant Escherichia (OR = 0.60), Campylobacter (OR = 0.89), and erythromycin-resistant Campylobacter (OR = 0.43) in chicken breast significantly decreased after the VFD rule changes, compared to the pre-VFD rule change period. The odds of detecting tetracycline-resistant Salmonella (0.66), Escherichia (OR = 0.56), and Campylobacter (OR = 0.33) in ground turkey also significantly decreased. However, the odds of detecting tetracycline-resistant Salmonella (OR = 1.49) in chicken breast and erythromycin-resistant Campylobacter (OR = 4.63) in ground turkey significantly increased. There was no significant change in the odds of detecting tetracycline-resistant Salmonella and Escherichia in ground beef or pork chops. The implementation of VFD rule changes had a beneficial effect by reducing the occurrence of tetracycline-resistant and erythromycin-resistant bacteria in chicken and ground turkey. Ongoing surveillance of antimicrobial resistance and antimicrobial use could complement the implementation of stewardship such as VFD rule in food-producing animals in the U.S.

New insight into the effect of microplastics on antibiotic resistance and bacterial community of biofilm

Microplastics (MPs) could serve as substrates for microbial colonization and biofilm formation. However, research on the effects of different types of microplastics and natural substrates on biofilm formation and community structure in the presence of antibiotic-resistant bacteria (ARB) is limited. In this study, we employed by means of microcosm experiments to analyze the situation of biofilms conditions, bacterial resistance patterns, antibiotic resistance genes (ARGs) distribution, and bacterial community on different substrates using microbial cultivation, high throughtput sequencing and PCR. The result showed that biofilms on different substrates markedly increased with time, with MPs surfaces formed more biofilm than stone. Analyses of antibiotic resistant showed negligible differences in the resistance rate to the same antibiotic at 30 d, but tetB would be selectively enriched on PP and PET. The microbial communities associated with biofilms on MPs and stones exhibited variations during different stages of formation. Notably, phylum WPS-2 and Epsilonbacteraeota were identified as the dominant microbiomes of biofilms on MPs and stones at 30 d, respectively. Correlation analysis suggested that WPS-2 could potentially be a tetracycline-resistant bacterium, while Epsilonbacteraeota did not correlate with any detected ARB. Our results emphasized the potential threat posed by MPs as attachment carriers for bacteria, particularly ARB, in aquatic environments.

Efficient photo-Fenton reaction for tetracycline and antibiotic resistant bacteria removal using hollow Fe-doped In2O3 nanotubes: From theoretical research to practical application

The low exposure of active sites and the slow electron transfer rate still restrict the wide application of the photo-Fenton system of Fe-based photocatalyst in practical water treatment. Herein, we prepared a hollow Fe-doped In2O3 nanotube (h-Fe-In2O3) catalyst for activating hydrogen peroxide (H2O2) to remove tetracycline (TC) and antibiotic resistant bacteria (ARB). Incorporation of Fe could shorten the band gap and increase the absorption capacity of visible light. Meanwhile, the increase of electron density at the Fermi level promotes the interfacial electron transport. The large specific surface area of the tubular structure exposes more Fe active site and the Fe-O-In site reduces the energy barrier of H2O2 activation, resulting in more and faster formation of hydroxyl radicals (•OH). After continuous operation for 600 min, the h-Fe-In2O3 reactor still can remove 85% TC and about 3.5 log ARB in secondary effluent, showing good stability and durability for practical wastewater treatment.

Correlation between Bacterial Cell Density and Abundance of Antibiotic Resistance on Milking Machine Surfaces Assessed by Cultivation and Direct qPCR Methods

The relative abundance of antibiotic-resistant bacteria and antibiotic-resistance genes was surveyed for different parts of a milking machine. A cultivation approach based on swab samples showed a highly diverse microbiota, harboring resistances against cloxacillin, ampicillin, penicillin, and tetracycline. This approach demonstrated a substantial cloxacillin resistance of numerous taxa within milking machine microbiota coming along with regular use of cloxacillin for dry-off therapy of dairy cows. For the less abundant tetracycline-resistant bacteria we found a positive correlation between microbial cell density and relative abundance of tetracycline-resistant microorganisms (R2 = 0.73). This indicated an accelerated dispersion of resistant cells for sampling locations with high cell density. However, the direct quantification of the tetM gene from the swap samples by qPCR showed the reverse relation to bacterial density if normalized against the abundance of 16S rRNA genes (R2 = 0.88). The abundance of 16S rRNA genes was analyzed by qPCR combined with a propidium monoazide treatment, which eliminates 16S rRNA gene signals in negative controls.

Dark repair of sunlight-inactivated tetracycline-resistant bacteria: Mechanisms and important role of bacteria in viable but non-culturable state

The spread of antibiotic resistant bacteria (ARB) in the environment poses a potential threat to human health, and the reactivation of inactivated ARB accelerated the spread of ARB. However, little is known about the reactivation of sunlight-inactivated ARB in natural waters. In this study, the reactivation of sunlight-inactivated ARB in dark conditions was investigated with tetracycline-resistant E. coli (Tc-AR E. coli) as a representative. Results showed that sunlight-inactivated Tc-AR E. coli underwent dark repair to regain tetracycline resistance with dark repair ratios increasing from (0.124 ± 0.012)‱ within 24 h dark treatment to (0.891 ± 0.033)‱ within 48 h. The presence of Suwannee River fulvic acid (SRFA) promoted the reactivation of sunlight-inactivated Tc-AR E. coli and tetracycline inhibited their reactivation. The reactivation of sunlight-inactivated Tc-AR E. coli is mainly attributed to the repair of the tetracycline-specific efflux pump in the cell membrane. Tc-AR E. coli in a viable but non-culturable (VBNC) state was observed and dominated the reactivation as the inactivated ARB remain present in the dark for more than 20 h. These results explained the reason for distribution difference of Tc-ARB at different depths in natural waters, which are of great significance for understanding the environmental behavior of ARB.

Aerosolization behavior of antimicrobial resistance in animal farms: a field study from feces to fine particulate matter.

Antibiotic resistance genes (ARGs) and antibiotic-resistant bacteria (ARB) in animal feces can be released into the atmosphere via aerosolization, posing a high health risk to farm workers. So far, little attention has been paid to the characterization of the aerosolization process. In this study, fecal and fine particulate matter (PM2.5) samples were collected from 20 animal farms involving swine, cattle, layers, and broilers, and the ARGs, ARB, and human pathogenic bacteria (HPB) were loaded in these two media. The results showed that approximately 70% of ARGs, 60% of ARBs, and 43% of HPBs were found to be preferential aerosolization. The bioaerosolization index (BI) of target 30 ARGs varied from 0.04 to 460.07, and the highest value was detected from tetW. The highest BI values of erythromycin- and tetracycline-resistant bacteria were for Kocuria (13119) and Staphylococcus (24746), respectively, and the distribution of BI in the two types of dominant ARB was similar. Regarding the bioaerosolization behavior of HPB, Clostridium saccharolyticum WM1 was the most easily aerosolized pathogen in swine and broiler farms, and Brucella abortus strain CNM 20040339 had the highest value in cattle and layer farms. Notably, the highest BI values for ARGs, ARB, and HPB were universally detected on chicken farms. Most ARGs, ARB, and HPB positively correlated with animal age, stocking density, and breeding area. Temperature and relative humidity have significant effects on the aerosolization behavior of targets, and the effects of these two parameters on the same target are usually opposite. The results of this study provide a basis for a better understanding of the contribution of animal feces to airborne ARGs and HPBs in farms, as well as for controlling the transport of the fecal microbiome to the environment through the aerosolization pathway.

Anaerobic sludge digestion elevates dissemination risks of bacterial antibiotic resistance in effluent supernatant

Anaerobic digestion following a variety of pretreatments is a promising technique for the reduction of excess sludge in municipal wastewater treatment plants (MWWTPs), and eliminations of possible pathogens, viruses, protozoa, and other disease-causing organisms. Notwithstanding a rapidly increasing health concern of antibiotic resistant bacteria (ARB) in MWWTPs, dissemination risks of ARB in anaerobic digestion processes are still poorly understood, especially in the digested supernatant. Taking the representative ARB with respect to the common tetracycline-, sulfamethoxazole-, clindamycin- and ciprofloxacin resistance, we investigated the compositions of ARB in the sludge and supernatant, and quantified their variations along the entire anaerobic sludge digestion process following ultrasonication-, alkali-hydrolysis- and alkali-ultrasonication pretreatments, respectively. Results showed that the abundance of ARB was diminished by up to 90% from the sludge along anaerobic digestion coupling with the pretreatments. Surprisingly, pretreatments clearly boosted the abundance of specific ARB (e.g., 2.3 × 102 CFU/mL of tetracycline-resistant bacteria) in the supernatant that otherwise remained relatively low value of 0.6 × 102 CFU/mL from the direct digestion. Measurements of the soluble-, loosely-bound- and tightly-bound extracellular polymeric substances components revealed a gradually intensified destruction of the sludge aggregates along the entire anaerobic digestion processes, which could be likely responsible to the increase of the ARB abundance in the supernatant. Furthermore, analysis of the bacterial community components showed that the ARB populations were strongly correlated with the occurrence of Bacteroidetes, Patescibacteria, and Tenericutes. Interestingly, intensified conjugal transfer (0.015) of antibiotic resistance genes (ARGs) was observed upon returning of the digested supernatant to the biological treatment system. It implies the likelihood of ARGs spreading and subsequent ecological risks upon anaerobic digestion towards reducing excess sludge, and therefore requires further attentions for the excess sludge treatments especially of supernatant.

Tetracycline resistance potential of heterotrophic bacteria isolated from freshwater fin-fish aquaculture system

This study investigated the tetracycline resistance potential of heterotrophic bacteria isolated from twenty-four freshwater fin-fish culture ponds in Andhra Pradesh, India. A total of 261 tetracycline resistant bacteria (tetR) were recovered from pond water, pond sediment, fish gills, fish intestine, and fish feed. Bacteria with high tetracycline resistance (tetHR) (n=30) that were resistant to tetracycline concentrations above 128 μg mL-1 were predominantly Lactococcus garvieae followed by Enterobacter spp., Lactococcus lactis, Enterobacter hormaechei, Staphylococcus arlettae, Streptococcus lutetiensis, Staphylococcus spp., Brevundimonas faecalis, Exiguobacterium profundum, Lysinibacillus spp., Stutzerimonas stutzeri, Enterobacter cloacae, and Lactococcus taiwanensis. Resistance to 1024μg mL-1 of tetracycline was observed in L. garvieae, S. arlettae, Enterobacter spp., B. faecalis. Tet(A) (67%) was the predominant resistance gene in tetHR followed by tet(L), tet(S), tet(K), and tet(M). At similar concentrations of exposure, tetracycline procured at the farm level (69.5% potency) exhibited lower inhibition against tetHR bacteria compared to pure tetracycline (99% potency). The tetHR bacteria showed higher cross-resistance to furazolidone (100%) followed by co-trimoxazole (47.5%) and enrofloxacin (11%). The maximum threshold of tetracycline resistance at 1024μg mL-1 was observed in S. arlettae, Enterobacter spp., B. faecalis, and L. garvieae and tet(A) was the major determinant found in this study.

Novel iron-carbon micro-electrolysis-anoxic/oxic biofilter system for enhanced treatment of tetracycline wastewater: Significant reduction of antibiotic-resistant and pathogenic bacteria

To mitigate the toxicity of tetracycline (TET) to functional microorganisms and avoid the exceeded discharge of tetracycline-resistant bacteria (TRB), a novel combined iron-carbon micro-electrolysis-anoxic/oxic biofilter (Fe–C-AOBF) system for treatment of pharmaceutical wastewater with high TET concentration was successfully established. Results showed that Fe–C-AOBF had the higher maximum tolerated concentration and the stronger degradation ability for TET than normal biological treatment system. When influent COD, NH4–N and TET were 4000, 200 and 100 mg/L, their removal efficiencies via Fe–C-AOBF could reach 96%, 86% and 90%, respectively. Microbial community analysis revealed micro-electrolysis pretreatment in Fe–C-AOBF system improved the microbial diversity compared to up-flow anaerobic/oxic biological aerated filter (UAF–UBAF). The relative abundance of dominate bacteria with tetracycline resistance genes (TRGs) in the biofilm of Fe–C-AOBF, such as Comamonadaceae, Thauera, Sinobacteracea, Acidaminobacter, and Enterobacteriaceae, was also decreased. Moreover, the absolute concentrations of five normal TRGs in effluent of Fe–C-AOBF were reduced by about 16 orders of magnitude as compared to UAF-UBAF. Due to approximately 70% of original TET has been degraded in micro-electrolysis process and lost its selective pressure to microorganisms, Fe–C micro-electrolysis played a key role in restraining the formation and reproduction of TRB and eliminating the potential risk of pathogenic bacteria (e.g., Arocobacter)in AOBF. Fe–C-AOBF system provided an efficient, stable and green method for antibiotic wastewater treatment with the function of inhibiting the emergence of antibiotic-resistant bacteria.

Tetracycline resistance of Escherichia coli isolated from water, human stool, and fish gills from the Lake Sevan basin.

The ecological state of Lake Sevan, the largest drinking water reservoir for the South Caucasus, formed under the influence of climatic and social changes. This study assesses the bacteriological quality of water in the rivers of the Lake Sevan basin and tetracycline-resistant bacteria isolated from fish and people living near the rivers of the Lake Sevan basin in Armenia in autumn 2019 and spring 2020. No differences have been shown for the tetracycline resistance of the investigated E. coli isolated from the human gut and the Masrik, Argichi, and Gavaraget Rivers. Horizontal gel electrophoresis revealed the same plasmid bands in most of the investigated E. coli with the same tetracycline resistance from the different sources of the Argichi River (obtained from people/fish/water sources where the fish were caught). The results also showed that most of the waters carried Edwardsiella spp., Erwinia spp., Morganella spp., and Proteus spp. in addition to E. coli; the coliform index did not exceed the standard level of 5×104 CFU mL-1 there. These findings highlight the importance of multidisciplinary studies of bacteria from "interacting" ecosystems, which might serve as a basis for the suggestion of microbial antibiotic resistance as another indicator of water pollution.

Ratiometric fluorescence detection of tetracycline for tetracycline adjuvant screening in bacteria

Persistent generation of tetracycline (TC) resistant bacteria poses serious risks for public health. It is of great importance to develop a strategy for rapid screening TC adjuvants for restore TC-susceptibility against resistant bacteria. Herein, a flower-like europium-doped nanoprobe is created by coordinating Eu3+ with adenine (Ade)-graphitic carbon nitride (g-C3N4) conjugate (g-C3N4-Ade-Eu). TC quenches the intrinsic fluorescence of g-C3N4 at 436 nm via internal filter effect (IFE), and meanwhile the antenna effect (AE) of TC significantly promotes the emission of Eu3+ at 618 nm. These fluorescence responsive behaviors of g-C3N4-Ade-Eu ensure ratiometric fluorescence assay of TC within 0.01 − 10 μM, along with a limit of detection (LOD) of 5.0 nM. The nanoflower structure exhibits large surface area, which endows enrichment of TC on the flower surface to ensure a high assay sensitivity. Tetracycline in bacteria is quantified for the first time by this protocol, and it further facilitates the rapid screening (∼3 h) of tetracycline adjuvants in E.coli ER2738.

Insight into the removal of tetracycline-resistant bacteria and resistance genes from mariculture wastewater by ultraviolet/persulfate advanced oxidation process

• UV/PS process completely inactivated tetracycline resistant bacteria. • UV/PS was effective to remove tetA, tetM and tetW in mariculture wastewater. • UV irradiation played an important role in removing tetracycline resistance genes. • Tetracycline dosage inhibited the removal of tetA, tetM and tetW in UV treatment. The antibiotic resistant bacteria and resistance genes are frequently detected recently in mariculture wastewater, thus posing a severe threat to the marine ecological environment. In this study, ultraviolet/peroxysulfate (UV/PS) process was adopted to treat tetracycline resistance pollutants and the influence of the seawater components, especially halogen ions, on UV/PS performance was investigated. The results showed that UV/PS process completely inactivated tetracycline resistant bacteria (TRBs) in five minutes and effectively removed tetracycline resistance genes (TRGs) for 1.18-2.17 log of tetA, tetM and tetW from mariculture wastewater. UV irradiation played an important role in the removal of three TRGs, where SO 4 · − contributed partly to the removal of tetA , while the partial removal of tetM and tetW was ascribed to the matrix composition, especially Cl − in mariculture wastewater. Due to UV competition effect, tetracycline as organic matter impacted heavily on the removal of the three TRGs in UV treatment, but even not in UV/PS system. This study demonstrated that UV/PS is a new alternative process to effectively remove tetracycline resistance pollutants in mariculture wastewater. Graphical Abstract .

Analysis of Antibiotic Resistance and Biofilm-Forming Capacity in Tetracycline-Resistant Bacteria from a Coastal Lagoon

Concerns have been raised regarding co-selection for antibiotic resistance among bacteria exposed to antibiotics used as growth promoters for some livestock and poultry species. Tetracycline had been commonly used for this purpose worldwide, and its residue has been associated with selection of resistant bacteria in aquatic biofilms. This study aimed to determine the resistance profile, the existence of some beta-lactamases genes and the capacity to form biofilm of bacteria isolated from water samples previously exposed to tetracycline (20 mg/L). Thirty-seven tetracycline-resistant bacterial strains were identified as Serratia marcescens, Escherichia coli, Morganella morganii, Pseudomonas aeruginosa, Citrobacter freundii, Providencia alcalifaciens, and Enterococcus faecium. The highest percentage of resistance was for ampicillin (75.75%) and amoxicillin/clavulanic acid (66.66%) in the Gram-negative bacteria and an E. faecium strain showed high resistance to vancomycin (minimum inhibitory concentration 250 μg/mL). Among the strains analyzed, 81.09% had multidrug resistance and eight Gram-negatives carried the blaOXA-48 gene. All strains were able to form biofilm and 43.23% were strong biofilm formers. This study suggests that resistant bacteria can be selected under selection pressure of tetracycline, and that these bacteria could contribute to the maintenance and spread of antimicrobial resistance in this environment.

Effect of tetracycline on nitrogen removal in Moving Bed Biofilm Reactor (MBBR) System.

The effect of tetracycline (TC) on nitrogen removal in wastewater treatment plants has become a new problem. This study investigated the effects of TC on nitrogen removal using a Moving Bed Biofilm Reactor system. The results showed that there was no significant effect on nitrogen removal performance when the concentration of TC was 5 mg/L, and that the total nitrogen (TN) removal efficiency could reach 75–77%. However, when the concentration of TC increased to 10 mg/L, the denitrification performance was affected and the TN removal efficiency decreased to 58%. The abundance of denitrifying bacteria such as those in the genus Thauera decreased, and TC-resistant bacteria gradually became dominant. At a TC concentration of 10 mg/L, there were also increases and decreases, respectively, in the abundance of resistance and denitrification functional genes. The inhibitory effect of TC on denitrification was achieved mainly by the inhibition of nitrite-reducing bacteria.

MALDI-TOF MS for rapid detection and differentiation between Tet(X)-producers and non-Tet(X)-producing tetracycline-resistant Gram-negative bacteria

ABSTRACT The extensive use of tetracycline antibiotics has led to the widespread presence of tetracycline-resistance genes in Gram-negative bacteria and this poses serious threats to human and animal health. In our previous study, we reported a method for rapid detection of Tet(X)-producers using MALDI-TOF MS. However, there have been multiple machineries involved in tetracycline resistance including efflux pump, and ribosomal protection protein. Our previous demonstrated the limitation in probing the non-Tet(X)-producing tetracycline-resistant strains. In this regard, we further developed a MALDI-TOF MS method to detect and differentiate Tet(X)-producers and non-Tet(X)-producing tetracycline-resistant strains. Test strains were incubated with tigecycline and oxytetracycline in separate tubes for 3 h and then analyzed spectral peaks of tigecycline, oxytetracycline, and their metabolite. Strains were distinguished using MS ratio for [metabolite/(metabolite+ tigecycline or oxytetracycline)]. Four control strains and 319 test strains were analyzed and the sensitivity was 98.90% and specificity was 98.34%. This was consistent with the results obtained from LC-MS/MS analysis. Interestingly, we also found that the reactive oxygen species (ROS) produced by tetracycline-susceptible strains were able to promote the degradation of oxytetracycline. Overall, the MALDITet(X)-plus test represents a rapid and reliable method to detect Tet(X)-producers, non-Tet(X)-producing tetracycline-resistant strains, and tetracycline-susceptible strains.

Changes in antibiotic resistance genotypes and phenotypes after two typical sewage disposal processes

Antibiotic resistome is a growing concern around the world. Wastewater treatment plants (WWTPs) have been identified as hotspots for antibiotic resistance gene (ARG) research. However, the distribution of antibiotic resistance genotypes and phenotypes in biofilm wastewater treatment system is poorly understood. In this study, the abundance and fate of antibiotic resistance genotypes and phenotypes in two typical wastewater treatment processes [biological aerated filter (BAF), anaerobic-oxic (A/O)] were quantitatively studied. The average removal rate of total ARGs was greater than 90%. In the biological treatment unit, the abundance of ARGs increased in the A/O unit and decreased in the biofilm unit. In addition, the resistance of tetracycline resistant bacteria changed after sewage disposal, which was closely related to the evolution of bacterial community. In total, the removal rate of resistance bacteria in A/O system was lower than that in BAF system. Genotypes were the basis of determining the phenotypes of microbial resistance. But it is necessary to pay close attention to antibiotic resistance phenotype due to its high variability. More specifically, antibiotic resistance mitigation in WWTPs should focus more on removing bacterial hosts to reduce the release of ARGs into the environment.

Tetracycline resistance in enterococci and Escherichia coli isolated from fresh produce and why it matters

The contamination of fresh produce with antibiotic-resistant bacteria is of particular concern as they are often eaten raw and can be a source for foodborne diseases. Tetracyclines have been largely used in humans, animals and plants which might have accelerated microbial resistance to them. Enterococci and Escherichia coli can be used as indicators to monitor contamination of the fresh produce with tetracycline-resistant bacteria. The investigation related to this issue is very scarce in Oman. This study aimed at identifying tetracycline-resistant enterococci and E. coli in fresh produce at the market place. Thirty-one enterococci and ten E. coli were isolated from local (Oman) and imported fruits and vegetables (N= 105). Using the standard Kirby-Bauer disc diffusion method, resistance to tetracycline was found in 6 (19 %) enterococci, isolated from cucumber, lettuce and radish, and 5 (50 %) E. coli, obtained from cabbage, lettuce and radish. Genetic analysis revealed the presence of tetracycline resistance genes, tet(A) and tet(K), in E. coli and tet(K), tet(L) and tet(M) in enterococci, including Enterococcus sulfureus, Enterococcus mundtii, Enterococcus casseli avus and Enterococcus faecalis. The integron integrase IntI 1 gene, which is known to facilitate the dissemination of antibiotic resistance genes among bacteria, was detected in 2 isolates of E. coli. These results demonstrated the capability of fresh produce to act as a potential source for disseminating tetracycline or possibly other antibiotic-resistant bacteria through the food chain. Thus, control strategies are needed to reduce exposure of the public to such microorganisms.