It is found that the same KOH activation procedure applied to reduced graphene oxide (rGO) and pine cone derived biochars results in carbon materials with a similar surface area, pore size distribution and performance in supercapacitor (SC) electrodes. It can be argued that “activated graphene” and activated carbon are essentially the same kind of material with a porous 3D structure. It is demonstrated that the pine cone derived activated carbon (PC-AC) can be used as a main part of aqueous dispersions stabilized by graphene oxide for spray deposition of electrodes. The PC-AC based electrodes prepared using a semi-industrial spray gun machine and laboratory scale blade deposition of these dispersions were compared to pellet electrodes.High surface area carbons are so far the best materials for industrial manufacturing of supercapacitor electrodes. Here we demonstrate that pine cones, an abundant bio-precursor currently considered as a waste in the wood industry, can be used to prepare activated carbons with a BET surface area exceeding 3000 m 2 g−1) was prepared starting from pine cones as a precursor and used as a main part of aqueous dispersions for spray and blade deposition of supercapacitor electrodes.Activated carbon (AC) with extremely high surface area (∼3000 m The energy density of EDLCs is lower compared to batteries.2 g−1) are the key elements of modern commercial SCs. A high specific surface area (SSA) translates, up to a certain extent, to more electrostatic charge storage. The most common high surface area carbon material is microporous activated carbon (AC). There have been many reports focusing on the preparation of low-cost AC from various bio-materials including various kinds of waste  and agricultural wastes . Converting waste into useful materials contributes also to resolving environmental issues related to waste recycling. A variety of activation methods were proposed to convert biochars into AC, each with own advantages and drawbacks. Possibly the most common in industrial applications is steam activation typically providing AC with a surface area of about 2000 m2 g−1.7,9,10 However, a much higher surface area (>3000 m2 g−1) was reported for AC carbons produced by KOH activation.11Porous and electrically conductive materials are required to prepare supercapacitor (SC) electrodes with high energy storage parameters. High surface area carbon materials (2000 m12–15 Activated reduced graphene oxide (a-rGO) or “activated graphene” is a promising material for the preparation of SC electrodes, due to a combination of a high SSA (∼3000 m2 g−1), high conductivity and mechanical stability.16,17 Gravimetric capacitance values in the range of 90–300 F g−1 have been reported for supercapacitors prepared using a-rGO16–19 with some variation of precursors, modification of the structure and pore size distribution of these materials.20–22Many kinds of nanostructured carbon materials and composites have been tested as electrode materials on the laboratory scale in recent years.e.g. due to influence of added components and specifics of deposition. Spray-gun deposition of electrodes was demonstrated in recent years as a feasible method to prepare large area electrodes for SCs on the semi-industrial scale.23–26 Recently, we have demonstrated that stable aqueous dispersions can be prepared using micrometer sized a-rGO grains and graphene oxide (GO) as a stabilizing agent.27 The dispersion also included fumed silica as a rheology modifying component and carbon nanotubes (CNTs) in order to improve electrical contact between a-rGO grains in electrodes. Thermal annealing was demonstrated to further improve performance of SC devices due to conversion of electrically insulating GO into conductive rGO. The dispersions have been prepared in liter amounts demonstrated to be sufficiently stable for deposition using industrial spray deposition.28It should be noted that the performance of SCs depends not only on the properties of porous materials but also on the methods of electrode preparation, 27,28 to prepare “activated graphene” but using natural biochar instead of rGO as a precursor. We also evaluated the performance of these ACs in SC devices.Activated graphene is a promising material for SCs, but it is relatively expensive due to the high cost of the precursor rGO. It should be noted that a-rGO is prepared by the KOH activation method, which is also commonly used to prepare AC. Both a-rGO and AC are carbon materials with a 3D porous structure. The main difference between “activated graphene” and AC is the type of precursor used for the activation. Reduced GO is used to produce “activated graphene”, and most commonly, various biochars are used to prepare AC. It is not yet clear if using rGO as a precursor provides significant advantages in terms of supercapacitor performance since the 2D sheets are anyway connected into a rigid 3D structure in the process of activation. Preparation of rGO is also a relatively complex process. It starts with oxidation of graphite into graphite oxide, which involves using strong acids, slow adding of an oxidant and thermally controlled conditions. The graphite oxide is then subjected to explosive thermal exfoliation to produce rGO. Biochars are a lot easier to prepare starting from variety of organic precursors using simple thermal annealing procedures. Therefore, we decided to prepare AC by exactly the same KOH activation procedure as the one previously used in our earlier studies2 g−1) obtained using KOH activation of pine-cone based biochars.29–33 Activated carbon from pine cones has been studied previously as a bio-sorbent for metals, CO2, dye waste, nitrate, and removal of lead(ii) ions from aqueous solutions by adsorption.8,29,31,33–36 There are relatively few reports on the use of pine cone derived carbons for the preparation of SC electrodes.32,37–41 However, most of these studies were performed using AC with a SSA (1000–2000 m2 g−1) significantly lower than that of a-rGO and prepared by somewhat different procedures32,38,41 with a single report on the material with ∼2500 m2 g−1 which is still below the surface area of a-rGO (up to ∼3000 m2 g−1).37Pine cones were selected as a starting material for preparation of AC due to its high abundance in Scandinavia  and reports of extremely high surface area materials  of pine cone derived AC (PCAC) was not always evaluated using verified procedures specific for microporous materials designed by J. Rouquerol et al.42 and requires independent verification. The value of the BET SSA of microporous materials is particularly dependent on the selection of the relative pressure (P/Po) range for the BET plot and can easily be extracted erroneously with rather different values from the same N2 isotherm. The correct procedure, which allows reading reproducible values of the SSA in microporous materials, includes a special method to select the right interval of relative pressures for a BET plot.42 The SSA value is also more reliably determined using suitable DFT models . Moreover, evaluation of pore size distribution in microporous materials requires rather detailed isotherms with many points at a low relative pressure interval which was not always the case in published studies.Moreover, an extremely high BET SSA  in dispersions used for spray deposition of SC electrodes.In this study we produced AC with an exceptionally high BET surface area of ∼3000 mThe scheme of this study is presented in −1 and keeping at 500 °C for 2 h under the flow of argon. This carbonization temperature program was chosen based on a previously reported procedure.29 The carbonized pine cones (biochar) were crushed into fine powder using an agate mortar before the activation procedure.The pine cones were collected on a sunny day in a forest outside of Umeå, Sweden. The site was far from the city and industry to minimize the risk of inorganic contaminants in the pine cones. Pine cones were washed several times in DI water and dried at 70° for a few days. The dry pine cones were carbonized in a tube furnace by heating from room temperature to 500 °C with a rate of 5 K min17,43 The biochar powder was mixed with KOH in a 1 : 8 proportion by weight and placed in a water/ethanol mixture (3 : 7 by volume). Typically, we used 50 ml of the water/ethanol solution for every 1 g of biochar. The mixture was magnetically stirred overnight and dried in a vacuum oven. The dry powder was then annealed under argon flow using the following temperature profile:The biochar produced from pine cones was activated using the KOH procedure optimized in our earlier publications for the rGO precursor to produce the highest BET surface area.−1).- Heating from 25 °C to 200 °C in 20 min .- Heating from 200 °C to 850 °C in 2 h 20 min .The reaction product was washed with 10% aqueous solution of acetic acid, filtered using a PTFE filter, then washed with water (until the pH was in the range 4–6) and dried.The carbonization step from dry pine cones to biochar has a yield of 32.2 wt%, and the activation step has a yield of 32.5 wt%. The amount of dry pine cones used was 16.2 g, which yielded 1.3 g of activated material, which corresponds to 10.5 wt%.et al.29 which was reported to result in AC with the highest SSA (3931 m2 g−1). KOH was grinded in a mortar and mixed with dry biochar (2 : 1 by weight). The mixture was then placed in a tube furnace and heated from room temperature to 800 °C with a heating rate of 3 K min−1, followed by 1 h annealing at 800 °C before cooling down to room temperature. The reaction product was washed with 10% HCl solution with stirring for 5 min and then rinsed with water before drying at 105 °C for 3 h. However, analysis of AC produced using this procedure showed a BET SSA value of ∼1850 m2 g−1, much smaller than expected from the report and our own procedure described above. For this reason, the procedure described by K. Li et al.29 was not used in our experiments with electrode preparation.A separate batch of activated pine cones was prepared according to the procedure described by K. Li −1 under nitrogen or air flow (40 mL min−1). SEM images were collected using a Zeiss Merlin FEG-SEM microscope. A Panalytical X'pert diffractometer was used to record XRD patterns in the reflection mode with CuKα radiation. Raman spectra were recorded using a Renishaw Invia Raman spectrometer equipped with a 514 nm laser. Fourier transform infrared (FTIR) spectra were recorded using powder samples using a Bruker IFS 66v spectrometer.The as-synthesized AC was characterized using X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD), Raman spectroscopy, thermogravimetric analysis (TGA), SEM imaging, BET analysis and Fourier-transform infrared (FTIR) spectroscopy. XPS spectra were recorded with a Kratos Axis Ultra electron spectrometer equipped with a delay line detector. A monochromatic Al Kα source operated at 150 W, a hybrid lens system with a magnetic lens, providing an analysis area of 0.3 × 0.7 mm, and a charge neutralizer were used for the measurements. The binding energy scale was adjusted with respect to the C 1s line of aliphatic carbon, set at 285.0 eV. All spectra were processed with the Kratos software. TGA was performed by using a Mettler Toledo TGA/DSC1 STARe System. Experiments were performed from room temperature up to 700 °C at a heating rate of 3 K minP/P0 for the BET plot was selected using a procedure optimized for microporous materials. The procedure takes into account Rouquerol parameters and is provided as a part of standard Quantachrome ASiQWin software package.42 The value of the BET SSA was also verified using the BETSI software44 which provided nearly identical values  equipped with a turbo pump at liquid nitrogen temperature. The relative pressure interval 2 : CNT).27 In some samples (for blade deposition), an increased amount of GO was also tested in the proportion (10 : 2 : 1 : 1). Electrodes were obtained by simply drying the dispersion on suitable current collectors . As was shown in our previous work, the electrodes prepared from this dispersion maintain a high SSA of the precursor material, which is useful for SC applications.27 To further improve the mechanical stability and conductivity of the electrodes, we subjected them to annealing at 200 °C for 1 h. Typically, the concentration of the dispersion used in spray deposition was 2.5 mg ml−1.Pine cone activated carbon (PC-AC) powder was dispersed in water with additives ) according to the optimized component ratio 10 : 1 : 1 : 1 (activated pine cone : GO : SiO−1).For blade deposition, we prepared a dispersion with a higher concentration of PC-AC  before and after spraying the graphene based dispersion.47 The dispersions stored for several days before deposition were mildly sonicated to improve stability and uniformity of the concentration between the top and the bottom of the bottle. We also used magnetic stirring to keep the suspension more stable during the spray deposition process.The dispersions were gun-sprayed on carbon coated aluminium foil (En’Safe Primed by ARMOR) collectors with a thickness of 20 μm. The spray gun machine has a moving spray-nozzle and maintains a constant distance of ∼15 cm from the surface. The collectors are placed on a hotplate heated up to a temperature above the boiling point of water (>100 °C) to ensure the instantaneous evaporation of the solvent droplets.48 AC-PC powder was mixed with poly(tetrafluoroethylene) in a weight ratio of 8 : 1, and some ethanol was added. The ethanol-wetted paste was compressed in a piston cylinder cell at 50 MPa to make 12 mm diameter pellets with a typical thickness of 130–200 μm. To ensure a homogeneous thickness of the pellets, the pressure loading–release cycle was applied nine times. Then the electrodes were dried in a vacuum oven for 1 h at 200–250 °C. The annealing results in conversion of GO into rGO which improves the electrical conductivity of electrodes.A separate set of electrodes were prepared as pellets, according to the procedure described in our previous work.2 for electrodes measured in aqueous electrolyte and 0.5–1 cm2 for electrodes measured in organic electrolyte. The typical electrode mass loading was 2–3 mg cm−2 for the electrodes made from drying dispersion and 1.4–1.8 mg cm−2 for spray deposition. The typical weight of pellet electrodes was 5–7 mg. Glass fiber membranes (Whatman GF/A) were used as separators for all measurements. CV curves were measured in the voltage range 0–1 V for aqueous electrolyte and 0–2.7 V for organic electrolyte using scan rates of 50, 100, 200 and 500 mV s−1. Charge–discharge curves were measured in the voltage range 0–1 V for aqueous electrolyte and 0–2.7 V for organic electrolyte at specific current values ranging from 0.5 A g−1 up to 100 A g−1.Electrochemical characterization of electrodes was performed using a potentiostat (Autolab PGSTAT204 or Iviumstat XR) and a standard two-electrode cell. For each measurement, two square electrodes were cut from a larger sheet (except for pellet electrodes) and weighed. Mass of the coating per unit mass of the current collector was measured by scraping off and weighing a reference sample with and without the coating (not necessary for pellet electrodes). Typical electrode sizes were 1.3–1.5 cm2 g−1, 3047 m2 g−1 and 3048 m2 g−1). An example of the BET plot composed of 7 points in the range of p/po of 0.124–0.274 selected using the Rouquerol procedure is shown in Fig. 1S  was reproduced well in three synthesis batches  impurities. Considering that a natural source was used for the preparation of PC-AC, the samples showed a rather small amount of impurities as confirmed by XPS, which revealed C/O ratios of 10.1 (batch 1) and 18.6 (batch 3).Chemical analysis of PC-AC was performed using XPS  which sh2O3 in this sample. Excluding oxygen from the Al2O3 impurity, the C/O ratio increases up to a value of 67.Higher oxygen content (C/O = 10.1) correlates with the increased amount of AlThe C 1s part of the XPS spectra shows the standard for AC components at 284.4 eV (not oxidized carbons), 285.3 eV and 287.7 eV from carbons connected to oxygen functional groups  and the satellite 290.9 eV component.2 isotherms. Once again, the PC-AC material shows a strong similarity with earlier reported a-rGO in microscopic texture revealed by SEM. The only difference which was observed in the SEM images is the absence of clear layered textures typically found in the a-rGO grains.The PC-AC samples consist of grains with size in the range ∼1–100 μm, which are visibly porous only in the high resolution images , as expeID/IG of about 5.64 .27Electrodes for testing in supercapacitors were prepared using three methods. The first two methods involved preparation of aqueous dispersions with PC-AC as a main component. The dispersions included graphene oxide as the surfactant, fumed silica as a rheology modified agent, CNTs for better mechanical stability and improved electrical contact between with PC-AC as a main component which contributes to a high surface area of the electrode. The SSA of electrodes prepared from AC is about 20% smaller compared to the SSA of precursor material due to added components.4/acetonitrile electrolyte. These results were compared to tests with electrodes deposited by blade deposition and tests with pressurized pellet electrodes.The dispersions were spray-coated on carbon plated aluminum foil and evaluated as electrodes in SC devices using TE-BF28 The parameters which had to be optimized include the nozzle speed, dispersion concentration, flow speed and temperature of the substrate. Visibly uniform deposition of electrodes with a mass loading of 3.22 mg cm−2 was achieved  compared to 2580 m2 g−1 for Kuraray AC. However, it is still somewhat lower compared to a-rGO which showed 130 F g−1 in 1 M TEA-BF4/Ac electrolyte.The maximal capacitance of PC-AC is clearly improved compared to standard commercial Kuraray AC, which was used in our previous studies (84 F g−1 ref. ). The imThe advantage of spray deposition is that the thickness of electrodes can be increased using multiple runs over the surface of a current collector. Adding an additional layer by blade deposition typically destroys the sample. It should be emphasized that spray deposition was performed using the semi-industrial scale over a relatively large area. This method can easily be scaled up for industrial production as compared to laboratory scale handmade blade deposition.The performance of PC-AC was also verified using pellet electrodes prepared by high pressure compaction. In this case, the electrodes do not include graphene oxide and fumed silica as a part of formulation but require a binder (PTFE in this case). As a result, the weight fraction of PC-AC is almost the same as in dispersions, but the thickness of electrodes can be made significantly larger.−1 at 0.5 A g−1.The gravimetric capacitance of pellet electrodes was found to be in good agreement (considering the significantly larger thickness of pellets) with values obtained using electrodes prepared from sprayed and bladed dispersions providing the highest value of 90 F gThe Nyquist plots  show tha49τ) separating the domination of capacitance and resistance. In agreement with Impedance data can also be treated in terms of the frequency-dependent complex capacitance that can be used to know the efficiency of a capacitive rate response.The sprayed sample is efficiently capacitive only in the lower frequency domain (the higher characteristic response time).As can be seen, annealing and preparation conditions together with the mass loading are all determining factors for the efficiency of a capacitive rate response. The bladed and pelletized electrodes provide a similar cell response in our samples. The increase in the overall resistance for the sprayed electrodes  slows do28The energy storage parameters of the SC with spray deposited electrodes are similar to those made with pellets as confirmed by the corresponding Ragone plots . As expeIt is an interesting observation considering that PC-AC and a-rGO materials showed a very similar surface area and essentially a microporous nature. This means that some other parameters are important in the overall performance of electrodes such as the nature of the remaining oxygen groups and differences in defects derived from different precursors after essentially similar activation procedures.The performance of blade deposited and pellet electrodes prepared using PC-AC was also verified in 6 M KOH electrolyte  using st−1 at 0.5 A g−1 , Fig. S6, S7.†4). Ionic liquids allow higher energy density due to the higher operating voltage. However, ionic liquids are low in electrolyte transport properties (viscosity and conductivity), which intrinsically limits the high-power performance. EMI-BF4 was chosen since it has one of the highest viscosities and conductivities known to date.Electrodes prepared using PC-AC were also tested using an ionic liquid, 1-ethyl-3-methylimidazolium tetrafluoroborate (EMI-BFInsufficient electrode wetting can be explained by the components used in the electrode preparation (such as rGO and fumed silica used in dispersions) and the electrode compaction method, which results in a poor compatibility with viscous IL electrolytes. At the same time, the pellets lead to a standard impedance response with a higher contact resistance due to more IL taking part in the interface, but also an almost ideal vertical-line capacitive response in the low-frequency range.The real capacitance component  shows th−1. At the higher rate, typically required from supercapacitors, only does the low rGO-related SSA provide some capacitance, which is also accordingly low (F g−1). The overall rate performance is reflected in a Ragone plot  precursor. The PC-AC material was evaluated for preparation of supercapacitor electrodes using aqueous dispersions (spray and blade deposition on current collectors) and as free-standing pellets. It is demonstrated that aqueous dispersions based on PC-AC are compatible with spray gun deposition using large area current collectors. The energy storage parameters of electrodes deposited using the semi-industrial spray gun machine were comparable to electrodes prepared by laboratory scale methods.In summary, pine cones were used as a naturally abundant precursor biomaterial for the preparation of activated carbon with an extremely high surface area of ∼3050 mThe energy storage parameters of supercapacitors prepared using PC-AC electrodes appeared to be almost on the same level as “activated graphene” with a similar surface area. Therefore, using “green” and inexpensive pine cone derived AC can be considered as a feasible alternative to “activated graphene.” The biochar is obviously a lot easier to prepare by simple thermal carbonization whereas the cost of pine cones is negligible. In contrast, rGO is prepared starting from graphite using oxidation in concentrated acids to prepare GO and explosive thermal exfoliation of GO to prepare rGO with a high surface area. Pine cones are currently considered as a waste in the forest/wood industry. It is also an abundant precursor available in large amounts in industrial regions such as Europe , Canada and the USA.There are no conflicts to declare.NA-004-D2NA00362G-s001


Klebsiella species are ubiquitous in nature and can be found in the natural environment and on mucosal surfaces of mammals and it is an important multidrug-resistant pathogen affecting humans and is a major source for hospital acquired infections. The aim of this study is to investigate the prevalence of ESBL enzyme among Klebsiella pneumoniae isolates by phenotypic methods from different hospital wards and detection of ESBL resistance genes such as TEM and SHV in Sulaimani city/ Kurdistan–Iraq.Klebsiella pneumoniae isolates were collected from different clinical samples from different hospitals, the isolates were identified by standard technique. Screening of ESBLs was undertaken by using double disk diffusion and standard disk diffusion methods. Real-time PCR was used for genotypic detection of TEM and SHV genes according to the standard protocol.Klebsiella pneumoniae isolates; 28 were ESBL positive, The pattern of antimicrobial susceptibility testing showed that the most resistant antibiotic are AMP (100%), AMC (100%) followed by CAZ (83.33%), CTX (75.9%), CPM (74%), ATM (70.37%). Both TEM and SHV genes were detected among 28 (51.85%) ESBL positive by using Real-time PCR method.Out of 54 Klebsiella pneumoniae.SHV gene was detected in most of the isolates of ESBL producers of  Enterobacteriaceae, plasmid-mediated enzymes capable of hydrolyzing a β-lactam bond in commonly prescribed β-lactam antibiotics, such as penicillins, broad spectrum cephalosporins and aztreonam Reviewers' comments:Reviewer's Responses to Questions  Comments to the Author1. Is the manuscript technically sound, and do the data support the conclusions?  The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: YesReviewer #2: Partly**********  2. Has the statistical analysis been performed appropriately and rigorously?   Reviewer #1: NoReviewer #2: Yes**********  3. Have the authors made all data underlying the findings in their manuscript fully available?PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.  The Reviewer #1: YesReviewer #2: Yes**********  4. Is the manuscript presented in an intelligible fashion and written in standard English?  PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.Reviewer #1: NoReviewer #2: No**********  5. Review Comments to the Author  Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. Reviewer #1: N0 Line number Reviewer’s Comments/Suggestions1 Abstract Most results of antibiotic not clarify clearly in abstract sectionKeywords Need to arrange alphbetically2 The manuscript contains many grammatical errors that must be corrected especialy the discussion and introduction section need too much correction3 highlighted in Green Correct the word4 highlighted in Red Delete word or character5 Many Word not writed in fully form like” EBLS,ICU,U.S.A,MICs,OPD)and others, also the term EBLS sometimes writed “ESBLS” other palces writed “ESBLs” it must write in unique and scinetific style6 Highlighted in Yellow Give a space7 Results - İn this sectio the concentration of each antibiotiv must mentioned inside the (table 4).-  this wrong value .8 Discussion The authors mentioned in this section this wrong reference lined with yellow” The other drug such as amoxicillin-clavulanic acid also shows high resistance rate (96.29%), the same result was reported in Erbil , Najaf and Duhok / Iraq , , (Qadir (2019). Respectively. So Gupta and his collegous study founded in U.S.A9 References All the refernce not in one unique style; the year of publication sometime after the name of authors and sometimes after the title of article, sometiems after the year of publication come comma others semicolon10 Others important notice - Many word wtrited wrongly like „standardise,Fifty four,30 µg must become30µg (without any space),TEM and SHV writed with different format like balSHV,blasCTM- Table 1,2 need to re-type in word format not in picture foramt- Chi-sequare and correlation analysis test mentioned in statistical anlalysis section but actualy not calculate for the data (for comparision), the authors must delete these characters , also the authors must mentioned which version of M.soft excel they used- Table 3, 4,5 ,6 and 7 must re-type again with word format adn without coluration .- The manuscript not contanin conclusion and any Acknowledgement.Reviewer #2: After reading the manuscript titled: I suggest the need for major revisions.I believe that the presented research is very relevant and worth publishing, however the authors need to pay attention to a few issues detailed below. First of all, spelling inconsistencies, such as VITEK / Vitek, are noticeable. The authors do not pay attention to the correct spelling of the name of the microorganisms , the names of the authors cited are often written in lower case. Punctuation errors are also noticeable.Introduction:1) “Extended-spectrum beta-lactamase (ESBL)-producing bacteria is plasmid-mediated enzymes…” – should be corrected, bacteria aren’t enzymes!2) “…evolutionary families based on their amino acid sequence TEM and sulphydryl variable SHV were the major types…” is this one sentence?3) ” The fact that patients most likely to become infected with ESBL-producing Enterobacteriaceae are those…” – Enterobacteriaceae should be in italics4) “It is estimated that Klebsiella species cause 8% of all hospital-acquired infections...” – Klebsiella should be in italics5) “Antimicrobial resistance in Klebsiella Pneumoniae…” – pneumoniae should be lowercase6) “Combinations of all types of bla genes were reported in this species, this may be due to either carriage of an antibiotic-resistant plasmid encoding an array of antibiotic resistance genes or due to acquisition of transposons containing different bla genes on the same plasmid” – bla should be in italics7) “Many different techniques exist for confirming ESBL production , Phenotypic tests for ESBL detection” – this sentences should be separated by a period, not a comma. Do not put spaces in front of the punctuation marks8) “” – too much brackets9) “.” – too much brackets, write surnames with capital letters10) “Another confirming method involves at least an eight-fold reduction in the MICs of ceftazidime or cefotaxime in the presence of clavulanic acid, The isolates then reported as resistant to all penicillins, cephalosporins and aztreonam (muhamed & Abas 2019).” - Separate sentences with a period and not with a comma, and write surnames with a capital letter11) “.” – surnames with a capital letterMethods1) shon etal.(2013). – surnames with a capital letter2) biomeireux – company name should be write: bioMérieux3) “Vitek® 2 biomeireux machine was used for the identification of all isolated Klebsiella according to the manufacturer’s recommendations” – which card was used?4) tryptic soya broth - who is the medium’s producer?5) Eppendoruf – correct to Eppendorf6) “Antibiotic susceptibility pattern of Klebsiella pneumoniae isolates were performed according to the Kirby–Bauer disk diffusion method  by using 15 commonly used antibiotic agents and all of the inoculated plates were aerobically incubated at 37°C for 18–24 hours, and the results were interpreted based on the instruction provided by the CLSI (2018 ).” - please mention here the antibiotics and their concentration7) “disks” – correct to “discs” in each place8) “Mueller-Hinton Agar plate” – no information about producer9) VITEK or Vitek – please standardize10) “Then let it to dissolve well and leave it for 15 minute at room temperature and mixed by vortexing, stock primer diluted 1:10 (10 μL of stock primer with 90 μL of DNA and RNA free distil water) to prepare working primer in concentration 10 pmol/μL (Ali& Ismael (2017).” - please correct the syntax so that it doesn't look like an instructionDiscussion1) “this indicate that UTI is the most common infection in hospitals and Klebsiella is the second most common isolated bacteria” – Klebsiella should be in italics2) “ICU samples . this indicates” – remove space before period and start sentence with a capital letter**********  6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.If you choose “no”, your identity will remain anonymous but your review may still be made public.Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.  Reviewer #1: NoReviewer #2: Nohttps://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, Attachmentmanuscript-26-10-2021.docxSubmitted filename: Click here for additional data file.Attachmentrenamed_36ab1.docxSubmitted filename: Click here for additional data file. 14 Dec 20211/ all Klebsiella pneumoniae were checked.2/ tables written and putted in word format 3/ figure uploaded in APACE style 4/ Reference checked according to submission guide line 5/ all reviewer comment checked step by step 6/ 2 file uploaded one is manuscript ( re new one after correction ) , other revised one as follow : green word : corrected and newly added according to reviewer comment , red words : deleted word 7/ cover page checked according to file template of plos one \\AttachmentRespons to reviewer.docxSubmitted filename: Click here for additional data file. 5 Apr 2022Phenotypic and genotypic detection of extended spectrum beta lactamase enzyme in Klebsiella PneumoniaePONE-D-21-32646R1Dear Dr. Anoar,We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.Within one week, you’ll receive an e-mail detailing the required amendments. 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The most predominant complexes of both guests with hosts were the 1:1 guest: host complexes. Stability constants of 2322 ± 547 M−1, 7281 ± 689 M−1, 3566 ± 473 M−1 were obtained for the complexes of BaP with CB[6], CB[7], and CB[8], respectively. On the other hand, stability constants of 5900.270 ± 326 M−1, 726.87 ± 78 M−1, 3327.059 ± 153 M−1 were obtained for the complexes of FLT with CB[6], CB[7], and CB[8], respectively. Molecular dynamic (MD) simulations were used to study the mode and mechanism of the inclusion process and to monitor the stability of these complexes in aqueous media at an atomistic level. Analysis of MD trajectories has shown that both BaP and FLT form stable inclusion complexes with CB[7] and CB[8] in aqueous media throughout the simulation time, subsequently corroborating the experimental results. Nevertheless, the small size of CB[6] prohibited the encapsulation of the two PAHs inside the cavity, but stable exclusion complex was observed between them. The main driving forces for the stability of these complexes are the hydrophobic forces, van der Waals interactions, electrostatic effect, the π····π and C–H···π interaction. These results suggest that BaP and FLT can form stable complexes with CB[n] (n = 6–8) in solution.The inclusion complexes of cucurbit[n]uril, CB[n] (n = 6–8), with poly aromatic hydrocarbon (PAH) Benzo(a)Pyrene (BaP), and fluoranthene (FLT) were investigated carefully in aqueous media. Fluorescence and  Polycyclic aromatic hydrocarbons (PAHs) are a large group of over 200 different compounds only made of carbon and hydrogen atoms containing two or more fused aromatic rings present in a mixture ,2. PAHs Benzo(a)Pyrene (BaP) is among the most carcinogenic PAHs. It is a molecule with five fused, hexagonal rings  Figure d. BaP exCucurbiturils CB[n]s are a family of macrocyclic compounds self-assembled from a condensation reaction of formaldehyde and glycoluril catalyzed by acid. These pumpkin-shaped containers have a characteristic rigid hydrophobic cavity accessible through two polar portals rimmed with carbonyl groups . The cavRecently, considerable literature has grown up around using CB[n] as a host molecule; however, no single study exists on its complexation with an unsubstituted BaP and FLT. PAH detection procedures suffer from several major drawbacks; therefore, to validate a new more sensitive, selective, cheap, and easy analytical procedure, this paper shines new light on the influence of this macrocyclic molecule in the spectroscopic behavior of BaP and FLT. The complexes between the guests and these cavitands are developed in an attempt to enhance the electronic properties of the guests such as the fluorescence. This will allow the establishment of a platform for the development of simple direct analytical methods for their quantification in complex matrixes such as food sample.1H NMR spectroscopy, and fluorescence spectroscopy. Furthermore, we utilized MD simulations in aqueous media to have a better insight into the major interactions between the guest and host and to identify the main factors stabilizing the complexes formed.The complexes in the solution were characterized by All chemicals used in this study were purchased from Sigma Aldrich  and were used without further purification. The host’s CB[n] were synthesized based on literature methods , and theThe fluorescence spectra were obtained using a PerkinElmer LS55 fluorescence spectrophotometer  equipped with a xenon lamp using a quartz cell of 1.0 cm path length. All measurements were recorded at room temperature. The stock solutions of BaP and FLT were prepared in methanol, ultrapure water was used to prepare the working solutions, and the concentration of this solution was kept at 1.0 μM, while the concentration of hosts was varied from 0.1 to 0.6 mM for CB[n].1H NMR data were processed with MestReNova v6.0.2-5475 software .The nuclear magnetic resonance (NMR) spectroscopy experiments were carried out using a Bruker Avance Ⅲ HD 700 MHz spectrometer  equipped with a 5 mm TCI H/C/N cryoprobe. The proton NMR experiment was run using the zg30 pulse program operating at 700.13 MHz. The acquisition parameters were as follows: 90° proton pulse width of 8.00 µs, relaxation delay of 1 s, and 128 scans. The spectrum was recorded at 298 K and processed using TOPSPIN 3.2 software . All host–guest complexes were prepared with DMSO-d6. All chemical shifts  are referenced to the solvent signal (DMSO-d6). The initial structure of BaP and FLT were built using Chemdraw and their geometries optimized using the DFT-B3LYP method using a 6-31G* basis set . On the Molecular docking experiments were performed using the Autodock Iprogram (version 4.2) . In thisThe MD simulations were carried out using the Desmond molecular simulations package, as distributed in the Schrodinger 2015 suite of programs. The OPLS_2005 all-atom force field with explicit solvent (TIP3P water model) was used throughout the calculations. Simulations were run with periodic boundary conditions, in an orthorhombic box with the solute, viz., placed in the middle at 20 Å distance from each of the box’s edges. The SHAKE algorithm was used to constrain covalent bonds between hydrogen and heavy atoms. Long-range electrostatic interactions were dealt with using the Ewald smooth particle mesh (PME) method . The sol−7 M) and upon complexation. According to the literature, some organic molecules tend to form aggregates in aqueous media through π-π type interactions between the aromatic rings  (n = 6–8) hosts was studied carefully by the spectrofluorimetric technique. The fluorescence emission spectrum of BaP shows two intense characteristic vibronic structure emission bands at 405 and 420 nm and a broader less intense one at 540 nm when excited at 385 nm . At highic rings ; specifi−7 M) displays a distinct enhancement on the 405 and 420 nm bands upon the addition of the three hosts, whereas the peak at 540 shows a monotonous increase in intensity with the increase in the hosts’ concentration (The fluorescence emission spectrum of the polyaromatic compound BaP solution (5 × 10ntration . These r3) to encapsulate the BaP molecule comfortably and form a stable inclusion complex with high emission intensity; The hydrophobic cavity of a cucurbituril encapsulates water molecules, which have high energy in accordance with their nonpolar cavities, and due to the weak dispersion interactions with weakly polarizable CB[n] cavity walls. Hence, the environment inside the cavity is not comfortable for these water molecules, which have stabilized themselves by hydrogen bonding interaction outside the cavity . In return for that, the addition of hydrophobic molecules or moiety to the cavity and expulsion of high-energy water molecules is a highly exothermic process . Moreove−6 M FLT solution, as shown in −4 M, indicating an equilibrium state  can provide further explanation for this variation. PC is defined as the ratio of guest volume to the volume of the host cavity. It is considered one of the most important factors determining the stability of the host–guest complex. It works as an estimator of the steric goodness of fit of host–guest inclusion complexes. In general, stable inclusion complexes result when PC values are in the range from 45 to 65% . Therefore, it was used here to predict the possibility of formation of BaP and FLT inclusion complexes with these hosts and to investigate their stability. MD provides valuable information pertinent to the fluctuations and conformational changes in atoms and molecules in materials . The congyr can be used to characterize the typical distance traveled by the body while the RMSD is used as a quantitative measure of the similarity between two or more complexes or molecular structures. BaP and FLT with CB[6] produce unstable trajectories, as is evident from the RMSD plots in The rThe π∙∙∙∙π interaction is between the carbonyl group of CB[n] molecule and the PAH molecules, whereas C–H∙∙∙∙π interactions are between the methylene group on the outer surface of CB[n] molecules with the PAH molecules . The RMSFLT enters the cavity of CB[7] by pushing the fused two benzene rings aside , as shown in gyr) of the hosts, guests, and their corresponding inclusion complexes during the simulation time are summarized in gyr of all complexes are comparable to those of the guests and are generally less than the sum of rgyr of the individual host and guest. This indicates the existence of a compact association between the guest and the host, except in the case of CB[6].The radius of gyration Pyrene (BaP) and Fluranthene (FLT) were obtained with CB[n] (n = 6–8). The existence of the complexes was supported by fluorescence spectrophotometry, and


An increase in tyre and road wear associated with winter road maintenance operations is also understood to contribute to the temporal increase of several metals including Cu, Zn, Co, Cr and V in the < 63-µm solids fraction in the road catchment gully pots. The concentrations of As and Pb decrease in all size fractions in both catchments, with the implementation of unleaded fuels (for Pb in housing catchment only), End-of-Life Vehicle Directive (Directive 2000/53/EC) (for Pb in both catchments), and strengthened industrial emission reduction measures suggested as possible drivers. The high contamination load for Zn, Cu, Cd and Pb in < 63-µm sediments from low-traffic housing catchment also emphasised the necessity of tracing and restricting non-traffic-related metal sources. Further seasonal monitoring of gully pot sediments is recommended to fully follow up the development of metals loading in both catchments.Diffuse pollution is recognised as a major challenge in achieving EU Water Framework Directive compliance, with urban runoff being a key pathway connecting various sources to receiving waters. Gully pots, as one of the ubiquitous urban drainage infrastructures, are placed at the inlets of piped drainage pipe network and actively drain runoff from urban catchment with suspended solids proportionally retained. The physiochemical properties of these retained solids reflect the activities within the catchment during the accumulation period. In this work, seven gully pots in two catchment types (highway and housing) in Luleå, Sweden were fully emptied and sediments analysed for total mass, particle size distribution and selected metal concentrations by six size fractions. The results of this sampling campaign are compared with the results of a 2005 study of the same gully pots to identify changes in the physicochemical properties of sediments over time and examine whether changes identified can be linked to changes in wider catchment management practices. The results highlight the potential impacts of winter road maintenance operations (e.g. up to a 15-fold higher solids loading rate in road catchment gully pots), reaching a normalised solids accumulation rate of 0.176–0.819 kg mThe online version contains supplementary material available at 10.1007/s11356-022-20341-8. Mitigating diffuse pollution is identified in the EU Water Framework Directive EU WFD  as a meaApproaches to mitigating urban diffuse pollution can be broadly categorised into two types: structural measures  and non-structural policies and practices which target pollutants at sources . Stormwater quality-control facilities are often regarded as an end-of-pipe solution and can be applied at different (sub-) catchment scales. Regional-scale solutions include larger storage systems, e.g. stormwater ponds that receive the runoff from a whole catchment of multiple land-use types. Local-scale solutions include swales, rain-gardens, green roofs and gully pots (GPs) which drain from a relatively smaller area. Whilst end-of-pipe solutions have shown their efficacy in alleviating the adverse impacts of urban runoff quality on receiving waters, increasing attention is focusing on opportunities to mitigate the release of diffuse pollutants at sources as upstream, cost-effective policy-based control measures Urbona . HoweverExamples of directives addressing both point and diffuse pollutions include the European Union Water Framework Directive EU WFD ; identifThe eight GPs are located in two types of catchments in Luleå, Sweden. Four GPs (R1–R4) are located along a highway with an annual average daily traffic (AADT) of over 10,000 vehicles/day (v/d), of which 8% are heavy goods vehicles. A further four GPs are located in a housing area (H1–H4) consisting of detached residences with chimneys (for wood-burning heating), private garages and gardens, with an AADT of less than 500 v/d . Full information on the dimensions, accumulation periods and contributing drainage areas of the seven GPs sampled is provided in Table The current sampling programme took place 21The size of the directly connected area to each GP was determined by implementing a catchment delineation process in ArcGIS software-based on aerial photography-based digital elevation data (resolution of 0.25 m) for Luleå, provided by Läntmateriet . The catGPs were emptied by an eductor truck that employed hydraulic dredging to loosen the compacted sediment bed in combination with a vacuum pump to collect the sediment–water mixture. After each GP was emptied, the collected mixture  was placed in a container lined with a layer of new high-density polyethylene (HDPE) to avoid contact between sample and container. The eductor truck was thoroughly cleaned between GPs to avoid cross-contamination between sites. The sediment–water mixture in each container was allowed to stand for 1 h before supernatant water samples were collected from each container and stored at 4 °C prior to the analyses. The remaining water volume was decanted using a peristaltic pump, and the total collected water volume was recorded. The container and wet slurry were then weighed before sediment samples were taken using the conning and quartering procedures described in ASTM .3 for every 100 ml sample and then analysed with sector field inductively coupled plasma mass spectrometry (ICP-SFMS) following SS-EN ISO 17294–2:2016 and US EPA Method 200.8:1994. For particulate fractions, samples were dried at 50 °C and digested before analysis with ICP-SFMS following modified SS EN ISO 17294–1,2 and US EPA Method 200.8.Water samples were analysed for suspended solids (SS) and selected metals  in both particulate and dissolved fractions . SS was determined using the multiple filter method (MFM) proposed by Nordqvist et al.  where thd) and selected metals. Mass-based PSD analyses followed ISO 11277:2009, where sediment samples were first wet-sieved to 63 µm with the < 63-µm size fraction further analysed using laser diffraction. Dry mass fractions are reported as the ratio of sample dry mass  by the sample wet weight. Selected metals contents were determined for the total fraction (≤ 2 mm) and for six size fractions . These size fractions were derived by first drying sediments at 50 °C, followed by their sieving through woven stainless steel wire cloth sieves of corresponding pore sizes. Metals were determined by ICP-SFMS following modified SS-EN ISO 17294–1,2 and US EPA Method 200.8:1994, after being digested with 5 ml 7 M nitric acid in a modified microwave oven. A lower reporting limit for metals , dry mass fraction , Material B (branded aggregated sizes of 4–8 mm) and Material C (branded aggregated sizes of 0–6 mm), Material A and B are used on residential streets, pedestrian and bike paths (including the H2-H4 study area), whilst Material C is used on high trafficked roads (including the road catchment in this study where R1–R4 are located).CFcatchment-year = contamination factor; B = local background concentration in local native soils (< 63 µm).Contamination factors (CF) for selected metals  were calculated using local background soil concentration data (available for the < 63-µm fraction) reported by Andersson et al. . CFs werI (kg/(year·m2)) = normalised solids accumulation rate in GP; SS (mg/L) = suspended solids; V (L) = volume of discharged water; fd = dry fraction; Mwet (kg) = mass of wet solids; T (year) = length of accumulation period; A (m2) = total effective connected area.The GPs investigated in this work and other studies do not share the same accumulation time period or receive runoff from the same sized catchment areas. As both parameters impact on the total mass of solid accumulated in GPs, the use of absolute values hinders the direct comparison of total solid dry mass between sites and studies. To address this, the total dry mass of solids in each investigated GP was normalised by the accumulation period and drainage catchment area, expressed as per Eq. :2\documeρ) > 0.6 and significant if p < 0.05.Anderson–Darling normality tests indicated a non-normal distribution of the dataset, and therefore non-parametric methods were used. Mood’s median tests (Minitab 19) were applied to examine the level of significance associated with (1) temporal changes of selected metals determined within the six sediment size fractions for each catchment type; and (2) impacts of catchment types on selected metal contents in sediments by size fractions for respective sampling occasion. Spearman’s rank-order correlation tests (Minitab 19) were also applied to examine the significance of relationships between selected metals in sediments by six size fractions for the respective sampling occasion. The correlation is considered strong if correlation coefficient  of < 63-µm particles than those from H2–H4 where particles in this size range account for less than 10% (by mass) of total sediments. For both catchments, particles within the size range of 8–63 µm dominate the < 63-µm sediments  from the road catchment (except 125–250 µm and 500–1000 µm) and only the < 63 µm and 1–2-mm fractions in the housing catchment  was only identified for the < 63-µm fraction from the road catchment and the 63–125-µm sediments from the housing catchment. Cd also exhibits an overall increase for both catchments  are used to quantify the magnitude of soil/sediment contamination relative to values reported for local uncontaminated soil. Figure CF for both Cu and Zn showed a clear increase in both catchments between the campaigns. For example, in the road catchment, the Zn CF more than doubled from a mean of 4.01 for the 2005 campaign to 9.39 in the latest sampling occasion (resulting in a classification of very high) and Cu increased from a mean CF of 3.92 in the 2005 campaign to 5.46 in the 2020 sampling occasion. Though the CF of Zn and Cu did not exhibit an equivalent level of increase in housing catchment sediments, an increase in CF was detected and sediments were classified as (or close to) a very high contamination level.In terms of Cd, whilst the road catchment sediments increased from a mean value of 2.16 to 3.79 (a shift from a moderate to a considerable contamination level) between sampling campaigns, the trend in the housing catchment is less clear. This is exemplified with the Cd CFs of housing catchment sediments in the 2005 campaign demonstrating significant levels of variation from moderate to very high levels. A similar pattern in Cd CFs is also shown in the 2020 sampling campaign, suggesting that activities at an individual housing (or plot level) have the potential to generate localised pollutant hotpots. A similar range in CF is also shown for Pb in the same 2005 housing catchment GP sediments, supporting the potential for residential activities to impact at a localised scale. However, a similar range in Cd and Pb levels is not seen in the housing catchment 2020 data set or observed in the road catchment data set where, overall, Pb concentrations showed a decreasing CF over time.The presented physicochemical properties were not reported for sediments from a specific year/time point but represent the physicochemical characteristics of a time-composite sediment sample of the entire sediment bed during the whole accumulation period prior to the sampling occasion. The physicochemical composition of GP sediments is a function of the activities which take place in the catchment between GP emptying campaigns. The following sections consider GP sediment composition data in relation to catchment activities with a focus on traffic and related winter road maintenance operations, regulatory changes and the influence of non-traffic sources.Compared with the PSD of GP sediments derived from the 2020 sampling campaign Fig. , those cComparing the PSD curves from both sampling campaigns, a threefold higher mass percentage of < 63-µm sediments fraction was reported in road catchment GPs in comparison with the previous sampling occasion Figs.  and 6. TBoth the 2005 and 2020 sampling campaigns indicate higher normalised solid accumulation rates in the road catchment in comparison to the housing catchment Fig. . Such a Besides the catchment differences related to the availability of solids being supplied to the investigated GPs, the differences in the GP designs may also contribute to the varied solids accumulation rates between sites. GPs in the road catchment have generally deeper sumps compared with those of housing catchment GPs Table . Such a As described in Sect. 2.4.2, the normalised solids accumulation rates presented in this work do not assume a constant sediments accumulation behaviour over the monitored time-frames, but instead account for differences in accumulation time periods and catchment sizes. The achievement of a hypothetically constant sediments accumulation rate would require a steady solids supply to the drainage catchment, stationary transport of solids to GPs in both wet and dry weathers and uniform solids retention efficiency, all of which are thought unlikely given the inherent variability of processes within am urban catchment. This is demonstrated by the results presented by Post et al. , where sBased on the findings of Post et al. , it is aρ = 1, p < 0.0005) indicating they derive from a common source. The impacts of winter road maintenance operations are also indicated by the strong correlations for Zn–V , suspended solids; ρ = 0.964, p < 0.0005 for < 63-µm fraction) and Zn-Co  in samples from the 2020 sampling, which are attributed to both the wear of V–Co-alloyed steel studs and/or the abrasion of V-containing asphalt  used in the housing catchment are of a larger size fraction than those analysed (i.e. < 2 mm) for the determination of total metal concentrations. However, the presence of metals in road grits at concentrations higher than those reported in GPs sediments suggests that abraded grit particles may contribute to concentrations of Cr, Co, V and Ni Fig. . With thThe concentrations of Zn for < 63-µm sediments of the road catchment GPs in this study are at the same magnitude as the mean concentration of 395 mg/kg reported in Pitt  for GP sWhilst AADT remained relatively constant between sampling periods, a range of regulatory measures directly affecting catchment activities were implemented. For example, legislation requiring the removal of Pb additives from gasoline (Ordinance in SFS1985:838) was implemented in January 1995. This regulatory measure is particularly relevant to the decreases in Pb CFs in the housing catchment sediments between the two sampling occasions Fig.  had an an = 2) to 0.1895 mg/kg (n = 4).Though the implementation of the EU ELV Directive  also limIn addition to the direct traffic-related sources, atmospheric deposition also contributes metals to the urban drainage system  was identified for suspended solids of the 2020 sampling occasion, suggesting a potential common source for both metals. Though commonly reported as a traffic-related pollutants released through the fossil fuel combustion process  with Pb in suspended solids and sediment samples supporting earlier studies of roadside soil samples  and inorganic  pollutants to the urban drainage system in the catchment (Hedenmark and Alkeblad −2 year−1) in road catchment gully pots of the latest sampling occasion than the previous one. These measures are also likely to have contributed to the higher mass fraction of < 63-µm particles in road catchment (mean of 19%) in comparison to the housing catchment (mean of 8.5%).Winter road maintenance operations e.g. applying traction grits and the compulsory use of studded tyres may contribute up to a 15-times higher solids loading rate  (for Pb only) and strengthened industrial emission reduction measures (both As and Pb) implemented by local industries over the evaluated period suggested as possible factors in this change. Likewise, the relative increase in local industrial Cd emissions could contribute to the increase in Cd sediment concentrations in both catchments.A temporal increase of Cu, Zn, Co, Cr and V is observed, especially fine solids in both catchments. The increase in the road catchment concentrations is considered a function of winter road maintenance operations whilst the source of the temporal increase in the housing catchment remains unclear.CFs estimated for < 63-µm fraction indicate that the impact of anthropogenic activities can be identified at a GP level. The fact that there are high CFs for Zn, Cu and Cd in both catchments and Pb (housing catchment only) highlights that traffic is not the only source of metals.This study evaluated sediments collected from the gully pots during two sampling campaigns with a time interval of 15 years. The sediments derived from these two sampling occasions represented varying accumulation periods with data used to evaluate changes in normalised solids accumulation rates, particle size distributions and selected metals concentrations. Results are discussed in relation to changes within the catchment structural and non-structural activities. Results suggest that:By design, gully pot sediments reflect activities at a local catchment scale. However, the results of this study also suggest that GP pollutant concentrations may also be influenced by long-range transport processes. Hence, changes in GP sediment pollutant concentrations may also be used to evaluate the impact of wider catchment management practices. Further monitoring with an extension of the pollutants analysis list is therefore recommended not only to follow up the metals loading changes in both catchments but also to provide a basis for future identification of temporal changes to other pollutants of emerging concerns.Supplementary file1 (DOCX 183 kb)Below is the link to the electronic supplementary material.


The structure, elemental compositions and morphology of the products were characterized by XRD, XPS, EDX, SEM and TEM, respectively. In addition, microwave absorption performance and the intrinsic mechanism are investigated in-depth. The paraffin-based composites with 20 wt.% filling contents exhibited superior microwave absorption capacities in view of both maximum reflection loss value  and effective absorption bandwidth , which can be ascribed to unique hollow structure and good impedance matching. With these considerations in mind, this study provides a reference for the construction of high-performance microwave absorbers with unique hollow structure.Hollow nanoboxes structure have raised great attention as microwave absorption materials on account of their ultralow density and large specific area. By introducing an adjustable interior cavity structure, the dielectric loss and microwave absorption performance were affected by the tunable complex permittivity and impedance matching was improved. In our study, hollow CoS The electromagnetic radiation generated by the extensive use of new electronic devices such as household electrical appliances, wireless communication and advanced military radars has led to the deterioration of the electromagnetic environment, posing a serious threat to human health and safety, normal operation of equipment as well as national defense security, which has aroused worldwide concern ,2,3,4,5.2) nanoparticles, because of their high thermal stability and good electronic conductivity, show special advantages in microwave absorption fields [2 has a half-metallic nature and unique magnetic properties, which could enhance the absorption performance. Recently, hollow or porous nanostructure materials have aroused widespread research interest in the microwave absorption field, due to their multi-reflection and scattering as well as the improved impedance matching. Therefore, hollow carbon spheres [1−xS hollow spheres [9S8 nanotubes [2 nanoboxes [2@MoS2 core-shell composites [Recently, transition metal sulfides (TMSs), due to their excellent electric and magnetic properties, have been used in a wide range of applications, such as in lithium-ion batteries ,10, supen fields . Further spheres , hollow  spheres , yolk-sh spheres , tubular spheres  and 3D g spheres ,22 are b spheres , porous anotubes , hierarcanoboxes  and paramposites . However2 to successfully fabricate CoS2 hollow nanoboxes with tunable interiors and shell thickness. As absorbents, the paraffin-based composites with 20 wt.% filler loading displayed excellent microwave absorbing ability, which is considered by the modification of electronic and geometric effects. Moreover, the current studies provided extensive impetus for the rational design and controllable synthesis of well-defined hollow nanoboxes with excellent absorption performance.In our work, we first adopt the surfactant-assisted solution method and in situ ion-exchange process in the presence of CS3)2·6H2O), 2-Methylimidazole and Hexadecyl trimethyl ammonium Bromide (CTAB) was purchased from Aladdin Corporation. Anhydrous ethanol was purchased from Sinopharm Chemical Reagent Co., Ltd. .All reagents were at analytic reagent grade and used as received without additional purification. Cobalt nitrate (Co(NO3)2·6H2O (0.292 g) and CTAB (0.01 g) were dissolved into 10 mL deionized water under ultrasonic stirring to form clear solution (defined as A solution). 2-methylimidazole  were dissolved into 70 mL deionized water under stirring to form solution B. After that, solution A was injected into solution B rapidly and stirred continuously for 2 h. After the reaction, the purple products were centrifuged, washed three times with water and redispersed in 60 mL anhydrous ethanol containing 0.3 mL CS2. Finally, the mixture was sealed into a 100 mL stainless steel autoclave and kept at 140 °C for 8 h. The autoclave was cooled down to room temperature naturally and the black precipitate was washed with ethanol several times. The final products were collected by centrifugation and dried in a hot oven at 70 °C for 12 h. The synthetic procedure is depicted in 2 as the sulfur source. The solid ZIF-67 precursor was successfully transformed into CoS2 nanoboxes. Due to the different diffusion rate of inward diffusion of vulcanizing agent and outward diffusion of metal ions, the vulcanizing agent reacted with metal ions on the surface of the solid ZIF-67 precursor and began to produce a core-shell structure. With the extension of vulcanization time, the ion exchange reaction was completed and the hollow CoS2 nanoboxes with unique composition and structure were obtained.In this typical synthesis process, Co using Cu Kα radiation (λ = 0.15418 nm). The morphology and inner structure information were investigated by field-emission scanning electron microscope , transmission electron microscope , high resolution TEM as well as scanning TEM (STEM) images and elemental mapping analyses. The chemical states and elemental composition of the samples were determined through X-ray photoelectron spectroscopy (XPS). The electromagnetic parameters of the products were measured by a network analyzer  in the frequency range of 1–18 GHz via a coaxial ring method.2 hollow nanoboxes were examined by powder X-ray diffraction (XRD) spectra, field emission scanning electron microscopy (FESEM) and transmission electron microscopy (TEM). As shown in 2 (PDF JCPDS No. 41-1471) [2 hollow nanoboxes during in situ sulfidation.The crystalline structure and morphology of as-obtained solid ZIF-67 precursor and CoS41-1471) ,29. Rema3/2 and Co 2p1/2 of Co2+ state, respectively, implying the existence of Co-S bond in the CoS2 nanoboxes. Furthermore, the other two peaks located at 802.1 eV and 784.3 eV suggested the existence of N-Co bond due to the insufficient sulfurization of ZIF-67 precursor. In 22−, respectively.The surface chemical composition of the samples were identified by X-ray photoelectron spectroscopy (XPS) . In the 2 hollow nanoboxes. As shown in 2, the amounts of CS2 and the reaction temperatures have great influence on the morphology: high dosage and high reaction temperature will destroy its structure  images further corroborated the well-defined hollow structure , effective absorption bandwidth (EAB) and absorbent matching thickness (dm) are the main factors influencing the microwave absorption performance of absorbing materials. The effective absorption bandwidth refers to the reflection loss value below −10 dB, which is equal to 90% of the incident electromagnetic wave be attenuated. According to transmission line theory, the reflection loss values of absorbing materials can be calculated by the following formula [Zin, Z0, rμ, rε, f, d and c are the input impedance of the absorber, the impedance of free space, relative complex permeability and relative complex, the microwave frequency, the thickness of the absorber and the velocity of light, respectively. Taking the practical applications into consideration, the filling contents ratio is an important index to evaluate the microwave absorption performance of wax-based absorbents. To investigate the microwave absorption performance of CoS2 hollow nanoboxes, various filling contents of the products were mixed with paraffin to form composites by a simple blending method. RLs and RL curves of the as-obtained hollow CoS2/paraffin composites with different thicknesses (1.5–5 mm) in 1–18 GHz at various filler loadings. When the filling content of CoS2 hollow nanoboxes was 10 wt%, the minimum RL value was −12.65 dB at 13.84 GHz with an optimum matching thickness of 3.1 mm  and complex permeability (μr = μ′ − jμ″), where the real parts (ε′ and μ′) stand for the storage capacity of electrical and magnetic energy and the imaginary parts (ε″ and μ″) represent the decay of the electric and magnetic energy of the material, respectively [ε″ and μ″ values could endow the better microwave absorption properties. In order to better explore the associated microwave absorbing mechanism of CoS2 hollow nanoboxes, the frequency dependence relative permittivity for CoS2/paraffin with various filling contents is depicted in ε′ and ε″ increase with the increase in the filling contents and the composites containing 30 wt% CoS2 hollow nanoboxes have much higher ε′ and ε″ values than those of the other two samples, which could endow them with better microwave absorption performance. Nevertheless, their absorbing performance is weaker than that of samples with 20 wt% filling contents, which may be due to their poor impedance matching performance. The ε′ and ε″ values display a downward trend in the frequency range of 2–18 GHz, though a vigorous fluctuation occurred at 7–18 GHz in ε″ curve of all composite samples. Such a phenomenon existed in many pristine absorbing materials and most composites can be explained by frequency dispersion behavior. The multiple resonance peaks founded in the ε″ curves of all samples maybe correlated with the interfaces, which would cause the displacement current lag. According to the Debye theory and free electron theory, it is deduced that dipolar polarization triggered by nitrogen atoms and defects in materials, interfacial polarization originating in various interfaces, and metal sulfide materials’ inherent conductivity loss together give rise to dielectric loss. Commonly, the relaxation process has a significant influence on the dielectric loss of microwave absorption materials, which can be explained by a Cole–Cole semicircle. According to the Debye dipolar relaxation, the relative complex permittivity (εr) can be described by the following equation [εs is the static permittivity; τ and ε∞ are the polarization relaxation time, and relative dielectric permittivity at the high-frequency limit, respectively. From Equation (4), it can be deduced as shown belowIt is well known that the microwave absorption properties of absorbing materials mainly depend on the complex permittivity εrε′ and ε″ can be deducedAccording to Equations (5) and (6), the final equation indicating the relationship between ε′ versus ε″ for 20 wt% samples are portrayed in ε′′ curve. However, without ignorance, the Cole–Cole semicircles are distorted, indicating there might be other mechanisms.Thus, the curve of ε′ versus ε″ is a single semicircle, which is named a Cole–Cole semicircle, and one relaxation process usually corresponds to one semicircle. Curves of 2 hollow nanoboxes could produce weak magnetic loss, which contributed to the improvement of the microwave absorption performance to some degree. Available literature sources show that the magnetic loss comes from hysteresis loss, domain-wall resonance, eddy current resonance, natural resonance and exchange resonance [μ0 is the permeability in vacuum, σ is the conductivity of microspheres and d is the thickness. Based on the Equation (7), if C0 is a constant, the existence of eddy current effect would be identified. From the curves of μ″(μ′)−2 f −1 against frequency shown in fr, r, Hα, |K1| and Ms are defined as the frequency of natural resonance, gyromagnetic ratio, anisotropic energy, anisotropic coefficient and saturation magnetization, respectively. The natural resonance can be influenced by two aspects according to the natural-resonance equation. On the one hand, the smaller size of magnetic cores gives rise to the increase in anisotropic energy of the composite due to the surface anisotropic field by the reduced size effect. On the other hand, the lower Ms value of the weak magnetic CoS2 will increase with higher anisotropic energy. As is recognized, the higher anisotropic energy is beneficial to enhance microwave absorption properties. Furthermore, the peaks at high frequency can be ascribed to the exchange resonance.In addition to dielectric loss, the presence of CoSesonance . In consesonance :(8)C0=μ″Z) and attenuation constant (α) of absorbing materials can be calculated by the following formulas, respectively [Electromagnetic wave reflection usually includes surface reflection and multiple reflection, in which multiple reflections are usually caused by scattering effects due to inhomogeneity within the material internal. As we all know, adjusting and designing porous and hollow structures can realize multiple reflections, thus effectively extending the electromagnetic wave propagation path. Furthermore, the presence of hollow structures can regulate electromagnetic parameters to achieve impedance matching characteristics, improving the electromagnetic wave absorption of materials’ performance. The impedance matching ratio As presented in 2 hollow nanoboxes have been successfully fabricated by the in situ ion-exchange process. The amounts of CS2 and the reaction temperature as well as reaction time play an important role in hollow nanoboxes’ structure during the ion-exchange process. The microwave absorption performance of different contents of paraffin-based composites was investigated in detailed. The results demonstrate that paraffin-based CoS2 composites with 20 wt.% filling contents show significantly enhanced microwave absorption performance with minRL of up to −54.48 dB at 17.4 GHz and the best effective absorption bandwidth of 6.0 GHz (RL ≤ −10 dB) due to their special hollow nanoboxes structure and good impedance matching. The unique hollow structure contributes to the multiple reflection of electromagnetic wave and could regulate the electromagnetic parameters.In summary, the CoS