It was observed that KC coatings exhibited a better preservation effect on pork meat after the addition of CEO. The KC–CEO coatings were effective in retarding the growth of total viable count, lactic acid bacteria, and H2S-producing bacteria. In a DPPH test, the level of lipid oxidation in pork meat was also significantly (p < 0.05) reduced by the KC–CEO coatings. Furthermore, these coatings displayed pronounced activity in inhibiting the adverse alterations of pH value and surface color. Practically, KC–CEO-coated samples still exhibited an attractive bright red color at the end of refrigerated storage. Taken together, the developed KC–CEO coatings exerted pronounced antimicrobial and antioxidant activities in pork, thus providing a potential approach to preserving perishable meat.Fresh pork is susceptible to microbial contamination and lipid oxidation, which leads to food safety and quality issues. This study aimed to develop a kappa-carrageenan (KC) coating embedded with cinnamon essential oil (CEO) for antimicrobial and antioxidant purposes in pork meat. The uncoated controls and coated samples were subjected to microbial (total viable count, lactic acid bacteria, and H Its consumption has increased to 55,950,000 tons in 2018, accounting for approximately 64% of total meat consumption in China . Microbibacteria .Cinnamon essential oil (CEO) stands out for its excellent antimicrobial and antioxidant activities ,9,10. ThZataria multiflora Boiss, Mentha pulegium, and Satureja hortensis was carried out in previous studies [In recent years, much attention has been focused on carrageenan as a source of biopolymer for the formulation of edible coatings . Carrage studies ,26,27. HPractically, several KC coatings carrying antimicrobial agents were applied for food preservation. For example, KC coatings loaded with ZnO nanoparticles delayed decay and discoloration, maintained firmness, and reduced the acidity of mango during storage . FurtherThe purpose of the current work was to assess the antimicrobial and antioxidant activities of a KC coating incorporated with CEO in pork meat, which will contribute to knowledge-based utilization of edible coatings during food storage.CEO was provided by Jofont Ltd. , with E-cinnamaldehyde, eugenol, benzaldehyde, and phenylacetaldehyde as the major components as determined in our previous work . This EOFood-grade KC was purchased from Beilian Company . Lean pork meat was procured from a local supermarket and transported to our laboratory in sealed foamed polystyrene boxes containing crushed ice within 1 h. All the media and other chemicals were obtained from Sinoreagent Ltd. .w/v). Glycerol was also added at a ratio of 1.5% (v/v) as a plasticizer. After stirring for 1 h, the mixture was cooled to 60 °C and incorporated with Tween 80 . CEO was then enriched to the desired level of 0% and 2% (v/v), respectively, followed by stirring for 30 min and homogenization for 2 min. The resulting solution was utilized for the subsequent pork meat coating test.KC-based coatings were prepared based on the method of Seol et al.  and Wang2S-producing bacteria), chemical (DPPH and pH), and physical (surface color) analyses.A pork coating test was conducted according to He et al. . Pork mew/v) in a stomacher bag, followed by homogenization for 2 min at room temperature. Decimal dilutions were then prepared with the same solution. Appropriate dilutions of pork homogenates (100 μL) were spread onto plate count agar and incubated at 28 °C for 2 days to determine total viable count (TVC). Appropriate dilutions (1 mL) were also poured onto MRS (de Man Rogosa Sharpe) agar and allowed to solidify, followed by pouring the same medium over the surface to generate another layer. Lactic acid bacteria (LAB) were enumerated on the double-layered plates of MRS medium after incubation at 25 °C for 3 days. In addition, 1 mL inoculum from the appropriate 10-fold dilution was poured onto Iron Agar and incubated at 30 °C for 3 days for H2S-producing bacteria (HSPB) count. All microbiological data were expressed as log CFU/g.Ten grams of pork samples in each group were aseptically added into 90 mL sterile saline solution . The samples were constantly mixed at 25 °C for 0.5 h. The absorbance was then taken at 517 nm as an indicator of antioxidant activity. A higher absorbance value corresponded to a lower antioxidant capacity of KC-based coatings.A DPPH assay was utilized to determine the antioxidant activity of KC-based coatings in pork meat . An aliqA total of 10 g pork meat was blended with sterile distilled water (90 mL), followed by homogenization at 12,000 rpm for 1 min . The resulting homogenate was submitted to pH determination by a PHS-3C digital pH meter .L*, a*, and b* was recorded at different positions of samples. The C* (chroma) and h° (hue angle) values were then calculated by the equations as stated below [A WSC-S colorimeter  was employed to assess the superficial color of pork meat. The parameter of ed below :C* =  from 5.82 log CFU/g on day 0 to 6.39 and 7.44 log CFU/g on day 5 and day 7, respectively. Moreover, treatment with KC coatings did not significantly (p > 0.05) affect the LAB count in pork meat during refrigerated storage, which was in agreement with the observation that KC films did not exert antibacterial activity either via direct contact or by vapor phase [LAB are also critical spoilage microorganisms responsible for the deterioration of food products such as fresh meat and ready-to-eat seafood ,37. As sor phase . On the or phase . It was The rapid growth of HSPB can also result in the deterioration of food products . As depiTaken together, the application of KC and KC–CEO coatings was able to inhibit the growth of some spoilage microorganisms in pork meat, and the latter one with CEO incorporated showed the most pronounced effect. The additive antimicrobial effect of CEO with KC coatings might be closely related to its pronounced capacity to disrupt bacterial cell membranes and cell walls as revealed in our previous work . Thus, Kp < 0.05) lower level of lipid oxidation than the uncoated controls. Moreover, the highest antioxidant activity was achieved by the use of KC–CEO coatings, suggesting the feasibility of the incorporation of CEO into KC coatings to enhance the antioxidative effect in pork samples.Lipid oxidation is another crucial factor affecting meat quality . TherefoSatureja hortensis EO. Interestingly, our current work uncovered that KC coatings exerted a modest antioxidant activity in a real food system, which could be due to the barrier action of these coatings between the oxygen and the samples delaying the myoglobin oxidation process in pork meat [In agreement with the aforementioned observations, Shojaee-Aliabadi et al.  reportedork meat . Moreoveork meat .The pH value is an important indicator of the quality of fresh meat . TherefoIt was noted that pork samples coated with KC and KC–CEO exhibited much lower pH values compared with the uncoated controls , which wL*, C*, and h°) of pork samples were determined in the current work. The L* value of uncoated, KC-coated, and KC–CEO-coated pork samples decreased, remained unchanged and increased during refrigerated storage, respectively  to 100 (white). C* represents color saturation, which varies from dull  to vivid color . h° indicates a color wheel, which ranges from red–purple (0°) to yellow (90°) [L* value, remaining C* value, and increased h° value of the uncoated samples indicated an unattractive brown and dull color, which might result in a decrease in their consumer acceptability. For KC-coated samples, the increased h° value could also lead to discoloration to some extent, despite the remaining L* value and increased C* value. On the contrary, the KC–CEO-coated pork samples exhibited an attractive bright, vivid, red color as suggested by the L*, C*, and h° values, respectively. This desirable color stability could be due to the pronounced antimicrobial and antioxidant activities of the KC–CEO coatings in pork meat [ow (90°) . In the ork meat .Similarly, the incorporation of oregano EO into pectin edible coatings was able to minimize color changes in pork loins . FurtherThe addition of CEO into KC coatings was able to inhibit microbial growth of TVC, LAB, and HSPB in refrigerated pork meat. Moreover, KC coatings enriched with CEO successfully retarded lipid oxidation and adverse alterations in pH and color throughout storage. Hence, the formulated KC–CEO coatings provide a promising approach for the preservation of perishable meat in the food industry. A systematic sensory analysis of pork meat can be considered in future studies to further demonstrate the strengths of these coatings.


The chemical profile of cannabis was determined by high-performance liquid chromatography with ultraviolet detection and gas chromatography with mass spectrometer and flame ionization detector. The extraction methods significantly influenced chemical profile of extracts. The yield of EtOH extracts contained less cannabinoids and terpenes compared to BUT and DME ones. Most of the extracts was predominantly (>60%) composed of various cannabinoids, especially THCA. All of them demonstrated activity against 18 of the 19 microorganisms tested. The minimal inhibitory concentrations (MICs) of the extracts ranged from 4 to 256 μg/mL. In general, the bacteria were more susceptible to the extracts than dermatophytes. Due to the lower content of biologically active substances, the EtOH extracts were less effective against microorganisms. Cannabis extracts may be of value to treat dermatophytosis and other skin diseases caused by various microorganisms. Therefore, they could serve as an alternative or supportive treatment to commonly used antibiotics.Cannabis preparations are gaining popularity among patients with various skin diseases. Due to the lack of scientific evidence, dermatologists remain cautious about their prescriptions. So far, only a few studies have been published about the effects of high-potency cannabis extracts on microorganisms  causing skin problems that affect more than 25% of the worldwide population. Even though, the high-potency cannabis extracts prepared by cold extraction are mostly composed of non-psychoactive tetrahydrocannabinolic acid (THCA) and only low amount of THC, their use in topical treatment can be stigmatized. The  These opportunistic pathogens can cause serious infections, such as endocarditis, sepsis, or streptococcal toxic shock syndrome. However, more frequently they cause painful skin infections such as folliculitis, impetigo, etc.  are predominantly caused by ith them  and due ith them . For exaith them . Althougith them .Trichophyton isolates in India were identified to be resistant to the widely used antibiotic terbinafine (TB) (S. aureus (MRSA)  . The sits (MRSA)  is idents (MRSA) . HoweverCannabis sativa, after its legalization for medicinal purposes, is in the scope of many researches. It has been used in traditional medicine for various purposes including treatment of skin infections , detla-9-tetrahydrocannabinol (THC), cannabigerolic acid (CBGA), and detla-9-tetrahydrocannabiolic acid (THCA)  and alsoUnfortunately, high-potency cannabis is mainly associated with its psychoactive potential caused by THC, which is formed from THCA during the thermal processing of the naïve extract. At the same time, THCA is non-psychoactive cannabinoid with anti-inflammatory, anticancer and neuroprotective effect. For the topical application cannabis is currently prescribed in the form of dried flowers, creams, tinctures, or extracts . Even th2 extraction of cannabis become widely accepted technique in industry , and tetrahydrocannabinolic acid A (THCA-A) were obtained from Cayman Chemicals . Extraction gas butane (BUT), and dimethylether (DME) were provided by RSonic  and Dexso GmbH , respectively. Microbiological growth media Mueller-Hinton Broth (MHB), Sabourad dextrose agar (SDA) were bought from OXOID , horse defibrinated blood was purchased from Thermo Fisher Scientific , antibiotics clotrimazole (CLT), chloramphenicol (CLP), ampicilin (AMP), terbinafine (TB), and RPMI 1640 medium from Sigma-Aldrich.All solvents used for the HPLC and GC analysis were of analytical grade. Acetonitrile (ACN) and formic acid (FA) together with terpene standards  and n-alkane standard were purchased from Sigma-Aldrich . Methanol, n-hexane, dimethyl sulfoxide (DMSO), and ethanol (EtOH) were purchased from VWR Chemicals . Standards of cannabinoids, namely, cannabidivarin (CBDV), cannabidivarinic acid (CBDVA), cannabigerol (CBG), cannabigerolic acid (CBGA), cannabinol (CBN), cannabinolic acid (CBNA), cannabidiol (CBD), cannabidiolic acid (CBDA), cannabichromene (CBC), tetrahydrocannabivarin (THCV), Δ2) equipped with drip irrigation system. Plants were placed in pots containing Euro Pebbles (expanded clay) as growth medium. Air ventilation unit maintained the room temperature and humidity between 22 and 30°C, and 40 and 70%, respectively. The microclimatic conditions were adjusted according to the plant growth phase. Double-ended high-pressure sodium lamps were used to provide a suitable spectrum of light at a power of 1,000 W. The detailed information about plant nutrition were described previously by ® R-100 vacuum evaporator  at 40°C. The Dexso extractor was used for the preparation of butane and dimethyl ether extracts. The extractor was filled with 30 g of plant material to which 500 mL bottle containing compressed BUT or DME was connected. The gas flow was allowed to enter, and the extract was collected at the bottom to the teflon paper. To remove the residues of the gases, the extracts were placed for 2 h under the vacuum at Glass Vac .The dried inflorescences from two cannabis genotypes – Forbidden Fruit (FF)  and Chocolope (CHP)  were used for the preparation of extracts. The plants were cultivated under controlled indoor conditions at the Department of Food Science, Faculty of Agrobiology Food and Natural Resources, Czech University of Life Sciences in Prague in 2020. Each genotype was cultivated on the table , transferred to a HPLC vial, and submitted for analysis.The apparatus consisted of UltiMate 3000 HPLC system  and was equipped with UV detector. Cannabinoids analysis was performed on an Excel SuperC18 column . Compounds were separated using a gradient elution employing water + 0.075% FA (A) and ACN + 0.5% FA (B) as mobile phase under following gradient (A:B): 0′, 30:70; 1′, 30:70; 10′, 0:100; 12′, 0:100; 14′, 30:70. Temperature of the column compartment was set at 40°C. Injection was 10 μL and flow rate was 1 mL/min. Cannabinoids were detected at wavelengths between 190 and 400 nm. Quantification was done under 210 nm. The evaluation of the acquired data was performed using the Chromeleon 7.2 software . Standard calibration curves were prepared in a concentration range of 2–100 μg/mL with six concentration levels . UV peak areas of the standards (at each concentration) were plotted against the corresponding standard concentrations (in μg/mL) using weighted linear regression to generate a standard curve.The extracts were dissolved in methanol at a concentration of 2 mg/mL, transferred to the vial, and submitted to the GC-FID or GC-MS analysis.–1. The oven temperature program began at 60°C with a 3.5 min hold; the temperature was increased at a rate of 3.5°C/min to 155°C and then again at a 30°C min–1 rate to 300°C and held for 10 min . The temperature of the quadrupole was maintained at 230°C and of the ion source at 230°C. The compounds were measured in a scan mode in a range of 55–700 Da. The retention indices of each analyte were calculated from the retention times of n-alkanes by linear interpolation as previously described by Kováts. Identification of the analyzed volatiles was carried out by comparing the spectra with the spectra of available standards and/or by comparing their spectra and retention indices with the NIST Mass Spectral Search Program, version 2.2. The relative quantification of terpenes was performed by 7890A GC coupled with FID detector (Agilent) under the same chromatographic conditions. The detector conditions were set up as follows: t = 300°C; flow of gases - air: 400 mL/min; H2: 30 mL/min; and N2 make up flow: 5 mL/min. All samples were analyzed in triplicate.To identify the terpenic profile, the cannabis extracts were analyzed by GC 7200 coupled with 7890 B qTOF mass detector (Agilent). Separation of individual volatile components was performed on a HP-5MS column . Helium was used as a carrier gas; the flow was set at 1 mL.minStaphylococcus aureus ATCC 25923 and 29213, S. epidermidis CCM 50, S. saprophyticus CCM 2727, S. lugdunensis CCM 4069, S. epidermidis CCM 4418, Streptococcus pyogenes CCM 4425. Their antifungal activity was determined against 12 dermatophytes, namely 3 strains of Nannizzia fulva  two strains of Trichophyton rubrum , Trichophyton tonsurans CCF 4930, Nannizzia gypsea CCF 5215, Epidermophyton floccosum CCM 8339, Microsporum canis CCM 8353, and Trichophyton interdigitale CCM 8377. The strains were obtained from the American Type Culture Collection (ATCC), Czech Collection of Microorganisms (CCM) or Culture Collection of Fungi, Department of Botany, Charles University, Prague (CCF).The antibacterial activity of the extracts was tested against seven bacterial strains, namely, To determine antimicrobial and antifungal activity expressed as minimal inhibitory concentration (MIC), slightly modified broth microdilution methods CLSI , 2009 wec = 51.2 mg/mL). Subsequently, the two-fold serial dilution of the extracts was prepared at concentrations ranging from 4 to 512 μg/mL was prepared to 96 microtiter plates containing MHB as growth medium. In case of S. pyogenes, the MHB was supplemented with 7% horse defibrinated blood. The microplates were inoculated with bacteria at a final density of 0.5 McF . The inocula were prepared from 1 day old bacterial cultures cultivated in MHB at 37°C. The degree of inhibition of bacterial growth was evaluated after 24 h cultivation at 37°C using the BioTek Synergy H1 microplate reader (Agilent) at 512 nm. The lowest concentration that inhibited bacterial growth by 80% compared to the growth control was considered the MIC. Chloramphenicol and ampicilin were used as positive controls.The crude extracts were dissolved in DMSO to prepare the stock solution (c = 51.2 mg/mL) was by two-fold serial dilution dosed to 96 microtiter plates, resulting in concentrations ranging from 8 to 1,024 μg/mL. RPMI 1640 medium with pH 7 was used as growth medium. The microplates were inoculated with dermatophytes at density 3–7 × 105 CFU/mL. Dermatophyte inocula were prepared from 14-day old fungal cultures grown on SDA at 27°C. The degree of fungal growth inhibition was evaluated after 5 days cultivation at 27°C by microplate reader BioTek Synergy H1 at 495 nm. The lowest concentration that inhibited the growth of the dermatophyte by 80% compared to the control was considered MIC. Terbinafine and clotrimazole were used as positive controls.The stock solution of the extracts . As the data did not show normal distribution, the significant differences were evaluated by non-parametric Kruskal-Wallis test.Most of the extracts prepared from two cannabis strains by three different solvents were predominantly (>60%) composed of various cannabinoids, especially THCA. All of them demonstrated activity against 18 of the 19 microorganisms tested. In general, the bacteria were more susceptible to the extracts in comparison to dermatophytes. The MICs of the extracts ranged from 4 to 256 μg/mL. The ethanolic ones were, in general, less potent than the DME and BUT ones. This fact can be attributed to lower content of the active compounds in EtOH extracts.The extraction yield by both gases was on average 4% higher compared to traditional maceration in ethanol . The colThe higher variability in the terpene profile was observed between strains and between extracts of one strain prepared with different solvents . The maiThe solvents had a statistically significant effect on the presence and relative ratio of individual terpenes in the extracts . Both EtAll extracts showed very good to moderate activity against 18 of the 19 microorganisms tested. The MIC of the extracts ranged from 4 to 128 μg/mL and 32 to 256 μg/mL  for bactS. aureus strains (MIC = 4–8 μg/mL). However, almost 10 times higher MIC values (MIC = 64–128 μg/mL) were determined for S. pyogenes. Such a significant decrease in the activity of cannabis extracts could be attributed to the presence of blood in the cultivation medium. As previously reported, in this type of media, cannabinoids become less effective due to their strong affinity to blood proteins   . General2 μg/mL) , therefo2 μg/mL)  has been2 μg/mL) . For exadunensis . While mdermidis  and limodermidis , 2021.T. rubrum CCF 4879 (the most common species causing dermatophytosis in developed countries) and M. canis CCF 6025, while A. insingulare was resistant even to the highest concentrations of the extracts tested. The mode values of the MICs demonstrated that the EtOH extracts were slightly less effective (MIC = 128 μg/mL) compared to the DME and BUT extracts (MIC = 64 μg/mL). Even though some of the cannabis skin preparations are among other disorders also indicated for the treatment of bacterial and fungal skin diseases (Trichophyton mentagrophytes (MIC = 6.25 – 50 μg/mL–1). Also hemp water extracts proved to have moderate activity against T. rubrum and interdigitale (MIC = 500 μg/mL) but not against N. gypsea  was more effective and less laborious. Since DME is successfully used in food industry, its pharmaceutical and commercial potential for plant extraction should be further considered and investigated.It is already known that cannabinoids have a relatively wide spectrum of biological activities for which they are useful in the treatment of skin diseases. However, not much attention has been paid to their effectiveness against bacteria and fungi, which are often accompanied by skin problems. Our research brought new evidence that cannabis extracts may be of value to treat dermatophytosis and other skin diseases caused by various microorganisms and showed that cannabis could serve as an alternative or supportive treatment to commonly used antibiotics.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.TS and ZK: writing – original draft and investigation. JT: investigation, validation, and writing – review and editing. AJ: resources. PK: writing – review and editing. VH: resources and writing – review and editing. AF: conceptualization, data curation, methodology, writing – original draft, review and editing, and supervision. All authors contributed to the article and approved the submitted version.


Gammaproteobacteria increased by FTCs. Moreover, the relative abundance of these taxa was positively correlated with the increment of DOC. Co-occurrence networks confirmed DOM-bacterial interactions, implying that specific microorganisms degrade specific substrates. At class level, Gammaproteobacteria were significantly positively correlated with labile C , whereas other bacterial classes such as Actinobacteria, Alphaproteobacteria, and Thermoleophilia were significantly positively correlated with aromatic compounds in the topsoil. Collectively, FTCs tended to activate DOM and enhance its biodegradability of DOM, potentially hampering DOC accumulation and C sequestration. These findings highlight the potential of DOM molecular mechanisms to regulate the functional states of soil bacterial communities under increased FTCs.Soil dissolved organic matter (DOM) plays a vital role in biogeochemical processes. Global warming leads to increased freeze–thaw cycles (FTCs) in boreal forest soils, which can change DOM production and consumption. However, the interactions between the chemical composition of DOM molecules and the microbial communities that drive C decomposition in the context of freeze–thaw are poorly understood. Here, a FTCs incubation experiment was conducted. Combined with pyrolysis gas chromatography–mass spectrometry and high-throughput sequencing techniques, the relationships between DOM chemodiversity and microbial community structure were assessed. Results indicated that both low-frequency (2FTCs) and high-frequency freeze–thaw cycles (6FTCs) significantly increased soil dissolved organic carbon (DOC) contents in the surface (0–10 cm) and subsurface (50–60 cm) soil layers. In the topsoil, FTCs significantly reduced the relative abundance of aromatic compounds, but increased the relative proportions of alkanes, phenols, fatty acid methyl esters (Me) and polysaccharides in the DOM. In the subsuface soil layer, only the relative abundance of Me in the 6FTCs treatment increased significantly. The response of bacterial communities to FTCs was more sensitive than that of fungi, among which only the relative abundance of  Freeze–thaw cycles (FTCs) increase DOC release and change the chemical structure of DOM in the surface and deep soils in the boreal forest.The improvement of DOM bioavailability is detrimental to DOC accumulation.Gammaproteobacteria play a dominant role in DOC production under the scenarios of intensified freezing–thawing.1.Dissolved organic matter (DOM) is the most active and bioavailable component of organic matter in soil. It is defined as a heterogeneous continuum of organic molecules of various sizes, that are soluble in water and can pass through a 0.45 um pore size filter . Soil DOReduced snow cover at high latitudes leads to an increase in the frequency and severity of soil FTCs . FTCs caLaboratory incubation studies furthermore indicated that 4–93% of soil derived DOM can be decomposed by microorganisms . So, a mcopiotrophic bacteria  tend to favor decomposition of protein components in DOM  FTCs can increase the bioavailability of DOM and can promote the conversion of aromatic compounds into polysaccharides; (2) FTCs increase the activity of dominant species and thus increase mineralization, while eliminating sensitive microorganisms, both of which jointly dominate DOC production.2.2.1.°30′-121°31′ E, 50°49′-50°51′ N, altitude 800–1,000 m). The zonal vegetation is mainly composed of Dahurian larch (Larix gmelinii) mixed with a number of White birches (Betula platyphylla) in the arbor layer, with dense Rhododendron dauricum and Ledum palustre in the herbaceous layer (The Greater Khingan Mountains in Inner Mongolia (Northeast China), are located at the southern edge of the Eurasian permafrost region. This region is characterized by a cold temperate continental monsoon climate with long cold winters and short warm summers, and a fragile permafrost that is sensitivity to global warming. The annual average air temperature is −5.4°C and the mean annual precipitation is 580 mm. The annual average temperature has been increasing 0.32°C/ 10a over the past 60 year . The sample soils from each plot at the same depth were mixed uniformly into a composite sample. Those samples were sieved with a 2-mm mesh to remove roots, gravels, etc. and subsamples were air-dried at 40°C to determine pH, soil moisture, total nitrogen (TN), and total carbon (TC). The remaining subsamples were used to conduct a simulation FTCs experiment.The daily air temperature measured at the Forest Ecosystem Research Station in the Greater Khingan Mountains showed that freeze–thaw mostly occurs from mid-March to mid-April. The average temperature during that period range from −10°C to 5°C, and the number of freeze–thaw cycles in that period varies between 2 and 6 . Accordi2.3.4+-N, NO3−-N) concentrations were measured using a continuous flow autoanalyzer . Water-extractable DOC was determined using a TOC analyzer . Soil pH (soil: water = 1:2.5 w/v) was determined using a portable pH meter .A subsample of each air-dried soil sample was milled with a ball mill  and then analyzed for soil TC and TN concentrations using an elemental analyzer . Fresh soil samples were extracted with 2.0 M KCl (soil: solution = 1:10 w/v) and inorganic nitrogen  with GC injector at 230°C and ion source temperature at 280°C. The carrier gas was helium (1.2 mL min−1). The relative proportion of each compound was equal to the percentage of the peak area of each product to the total peak area.Pyrolysis-gas chromatography–mass spectrometry analysis was conducted for DOM characterization using a multi-shot pyrolyzer  attached to an Agilent 7,890 N gas chromatograph (GC) connected to an Agilent 7000B mass spectrometer (MS). The GC was equipped with an elastic Quartz Capillary Column . DOM-containing extracts were lyophilized using a freeze-dryer . Lyophilized samples (10 mg) were weighted into a small stainless-steel cup and inserted into a pre-heated furnace. The pyrolysis temperature was set as follows: 50°C for 1 min then rose to 600°C, from 50°C to 250°C at a rate of 50°C min2.5.Genomic DNA was extracted from 0.25 g of homogenized soil sample, using a PowerSoil DNA Isolation Kit  according to the manufacturer’s recommendation. The V3-V4 regions of bacterial 16S rRNA were amplified using the primer sets 515F (5′-GTGCCAGCMGCCGCGG-3′) and 907R , following a Monte Carlo permutation test (999 permutations) was conducted to evaluate the influence of soil chemical parameters on bacterial community composition under FTCs treatment. The soil chemical parameters were first examined to reduce the collinearity by eliminating predictors with VIF > 10. A forward selection procedure on environmental variables were performed by the ‘ordiR2step’ function in the vegan package to select the descriptors that affect bacterial community composition. Forward selection is a type of stepwise regression that starts with an empty model and adds variables one by one. In each forward step, it adds the one variable that gives the single best improvement to the regression model and this process is carried out by an automatic procedure.All analyses were conducted in R software (version 4.1.1). Before statistical analysis, the normality of the data was checked by a Shapior–Wilk’s test and the homogeneity of variance was test by a Levene’s test. One-way analysis of variance (ANOVA) combined with a Turkey’s HSD test were used to compare the means in test parameters between FTCs treatments. A Student’s t-test was performed to analyze the differences in abiotic and biotic properties at two soil depths (0–10 cm and 50–60 cm). Principal component analysis (PCA) was performed to evaluate changes in the DOM chemical compositions under different FTCs treatments with the ‘factoextra’ package. After calculating the Bray-Curtis distances, principal coordinates analysis (PCoA) was used to analyze the dissimilarity of soil microbial composition at the OTU level. Differences in the microbial communities between FTC treatments were further tested by analysis of similarities (ANOSIM) and non-parametric multivariate analysis (ADONIS) using the ‘vegan’ package. Mantel tests were implemented to interpret the significance of soil properties on the microbial compositions. The relationships between response variables and explanatory variables under FTCs treatments were tested using ordinary least squares (OLS). Co-occurrence networks were inferred for each soil layer (9 samples per layer) between the bacteria OTUs (abundance >0.05%) and DOM molecules based on the Spearman correlation matrix using the ‘psych’ package. To reduce network complexity, only individual DOM molecules with their abundance >0.01%, a correlation coefficient |3.3.1.4+-N were not significantly different among the FTCs treatments. However, FTCs significantly decreased the subsurface soil NO3—N content . The pyrolytic compounds were grouped according to their chemical similarity and probable origin in the following categories: alkanes, alkenes, aromatics, polyaromatics (PAH), nitrogen compounds (N-comps), phenols, polysaccharides (Polysac), and fatty acid methyl esters Me; . In bothIn the topsoil, 2FTCs significantly increased the relative abundance of long-chain alkanes (C25-C32), while 6FTCs only significantly increased C23 and C25 . In the The PCA of DOM chemical components for each soil layer is presented in 3.3.Proteobacteria (48.60%) and Actinobancteria  and Basidiomycota , followed by Dothideomycetes  correlation with recalcitrant compounds  and alkenes (E7–E25) made up only a small fraction of the pyrolysis products. Long-chain (A25–A33) alkanes and alkenes are typically derived from microbial lipids and plant biopolymers , while mid-chain and short-chain aliphatics can be attribute to microbial polymers . The latAromatic compounds formed during the pyrolysis process mainly originate from proteins, lignin, carbohydrates and charcoal . BenzeneSome pyrolytic compounds were phenol and alkyl phenols, which can be derived from any phenolic precursor such as tannin, lignin, proteinaceous biomass, and carbohydrates . HoweverFTCs increased the relative proportion of polysaccharides in the topsoil layer. Polysaccharides that can be derived from microorganisms include benzofuran, furfural, and methylfuran while residuals from plant material include levoglucosan and levomannosan . Only mi3_enriched conditions  and its derivatives (N6) can be formed by microbial degradation of plant lignin and other phenolic substances under NHnditions . Becausenditions . Peptidenditions . PAH arenditions . Howevernditions . FTCs ha4.3.Betaproteobacteria, Gammaproteobacteria and Flavobacteria has been related to complex DOM profiles , which are not only significantly positively correlated with labile DOM components (particularly polysaccharide compounds), but also with recalcitrant DOM compounds  and Betaproteobacteria  responded quickly to unstable C  is inclined to utilize both labile sucrose and recalcitrant compounds (Microvirga (Alphaproteobacteria) aggregated in heavy metal contaminated and nutrient-deficient soils , could grow slowly in low-nutrient soils and tolerate harsh conditions. The relative proportion of Actinobacteria did not change significantly after FTCs treatment. Classes of Actinobacteria and Thermoleophilia both belonged to the phylum Actinobacteria, and Actinobacteria were not affected by freeze–thaw, while the relative abundance of Thermoleophilia was significantly reduced under FTC treatment. One possible reason is that Thermoleophilia are known to be moderately thermophilic and oil-loving (Actinobacteria and Thermoleophilia correlated to both unstable components (particularly carbohydrates) and recalcitrant compounds via co-metabolism  to decompose rhizodeposits and litter  and long-chain alkanes (A26-A32) in the co-occurrence network , KD4-96 (Chloroflexi), Gemmatimonadetes, and Planctomycetes were not correlated with the DOM chemical compositions, except that KD4-96 was positively correlated with PAH in topsoil. This might be due to the reduction of microbial activity of low-abundance microorganisms under freeze–thaw environments. Another possible reason is that the DOM bioavailability has not changed significantly under the freeze–thaw treatment. Thus, in topsoil, the chemical composition of DOM is one of the major drivers of bacterial community variability under FTCs treatment, while in deeper soil, DOC content is the main factor shaping the bacterial community.Several rant SOM . Acidoba network . AcidobaE9, E12; , B. ShorE9, E12; . Thus, AAcidobacteriae showed negative and positive correlations with alkanes and alkenes, respectively. Alphaproteobacteria presented positive correlations with alkenes and aromatic compounds. A module in the co-occurrence network corresponds to a functional cluster, indicating that microorganisms in that cluster utilize similar DOM molecules  and a decrease in Shannon index. Furthermore, Gammaproteobacteria were dominant in freeze and thaw cycled soils and most likely induced the largest contribution to DOC release. The interactions between DOM molecules and bacterial communities showed that specific microorganisms can degrade specific substrates. In the topsoil, DOM chemical composition shaped bacterial communities, with labile C correlated to Gammaproteobacteria, and more recalcitrant C associated with other bacteria . In comparison, DOC contents were more likely to explain the variation of bacterial communities in the deeper soil layers. This study thus provides new insights into DOC accumulation, transformation, and stability in boreal forest soils under scenarios of intensified freezing–thawing.This study demonstrated that freeze and thaw increased DOC contents in the surface and deep soils of a boreal forest, and changed the chemical composition of DOM. In particular, the decrease of aromatic compounds and the increase of alkanes, phenols, polysaccharides, and Me in the topsoil indicated an improvement of DOM bioavailability. Soil bacteria were more sensitive to FTCs as compared to fungi, manifesting as a decrease in relative abundance of bacterial classes  can be found at: YY, HF, and SC: conceptualization, methodology, data curation, and writing—review and editing. YY: software and writing—original draft preparation. YY, YG, YL, YZ, and FS: formal analysis. HF: investigation and project administration. HF and SC: resources and supervision. KV: visualization. HF and KV: funding acquisition. All authors contributed to the article and approved the submitted version.This research was funded by the Second Tibetan Plateau Scientific Expedition and Research Program (STEP) (No. 2019QZKK1003), the Strategic Priority Research Program of the Chinese Academy of Sciences , National Natural Science Foundation of China (Nos. 41977041 and 31770558), the “Thousand Talents Plan” Project of High-End Innovative Talents of Qinghai Province (No. TTPPHEITQP-2019), and Key research and development projects of Ji’an Science and Technology Bureau (20111ZDF04022). KV received an FWO sabbatical bench fee (number VWH-E1313-SAB/22/016).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.https://www.frontiersin.org/articles/10.3389/fmicb.2022.1012512/full#supplementary-materialThe Supplementary material for this article can be found online at: Click here for additional data file.


Observational studies report inconclusive effects of tea consumption on the risk of Alzheimer’s disease (AD), and the mechanisms are unclear. This study aims to investigate the effects of genetically predicted tea intake (cups of tea consumed per day) on AD, brain volume, and cerebral small vessel disease (CSVD) using the two-sample Mendelian randomization (MR) method.N = 447,485), and AD was from the International Genomics of Alzheimer’s Project . Genetic instruments were retrieved from UK Biobank using brain imaging-derived phenotypes for brain volume outcomes  and genome-wide association studies for CSVD .Summary statistics of tea intake were obtained from UK Biobank  and reached a weak significant level using MR-Egger regression (p < 0.1). The result passed all the sensitivity analyses, including heterogeneity, pleiotropy, and outlier tests. In the secondary MR analysis, per extra cup of tea significantly decreased gray matter  and right hippocampus volume . We found a nonlinear association between tea intake and AD in association analysis, which suggested that over-drinking with more than 13 cups per day might be a risk factor for AD. Association analysis results were consistent with MR results.In the primary MR analysis, tea intake significantly increased the risk of AD using two different methods (ORThis study revealed a potential causal association between per extra cup of tea and an increased risk of AD. Genetically predicted tea intake was associated with a decreased brain volume of gray matter and the right hippocampus, which indicates that over-drinking tea might lead to a decline in language and memory functions. Our results shed light on a novel possible mechanism of tea intake to increase the risk of AD by reducing brain volume.  Alzheimer’s disease (AD), the most cause of dementia, is a neurodegenerative disorder associated with cognitive declines and progressive memory loss that subsequently causes language deficiencies accompanied by behavior disorders , 2. ApprTea, one of the most popular non-alcoholic beverages, has attracted lots of interest for its possible protective effects on body health ; howeverThe results from observational studies are often limited by measurement errors, confounders, and reverse causality. Exposures were identified as risk factors and proposed to be valuable interventions but later proven non-causal. Randomized trials may need decades to produce robust results since some exposures, such as diet, may take years to affect diseases . Meanwhiand related traits. This might give clues on practical prevention strategies for AD patients.This study conducted a Mendelian randomization analysis to assess the causal relationship between tea intake and AD. Cerebral small vessel diseases (CSVD) and brain volume are closely related to both the presence and the severity of the clinical symptoms of AD . In ordeA valid instrumental variable needs to satisfy three assumptions . First, N = 447,485) and tea consumed  was obtained from MRC Integrative Epidemiology Unit (IEU) open genome-wide association study (GWAS) project using GWAS ID ukb-b-6066 and ukb-b-179881 (p < 5 × 10–7). After that, SNPs were clumped with linkage disequilibrium within 1 Mb genomic distance (R-squares > 0.001) using the 1,000 Genome European reference panel and SNPs were excluded if not presented in the outcome GWAS (2 (Tea intake measured by cups of tea consumed per day (UKB Data-field ID: 1488), and tea consumed described as whether consumed tea yesterday (UKB Data-field ID: 100309) were two exposures. Summary-level data on tea intake  . The firReplication analysis was performed using data of a meta-analysis for late-onset AD (LOAD) on 13 cohorts excluded 23andMe conducted by Psychiatric Genomics Consortium based on European and US populations . This meFor secondary outcomes, we performed the MR analysis only using exposure that was significantly associated with our primary outcome.We included total brain volume (gray matter and white matter), gray matter volume, white matter volume, left hippocampus volume, and right hippocampus volume in the study. Summary statistics of brain volume traits were obtained from a GWAS of brain imaging-derived phenotypes in up to 33,224 individuals from the UK Biobank . The braN > 17,663), white matter hyperintensity (WMH) volumes, and three types of brain microbleeds (BMBs), including any BMBs , strictly lobar , and mixed types . Mean diffusivity and fractional anisotropy were obtained from a genetic association study of small vessel disease using MRI markers . Data on BMBs were obtained from a GWAS meta-analysis of 11 population-based cohort studies and three case-control or case-only stroke cohorts  and fractiona anisotropy (FA)  regression, weighted median (WM), and MR-Egger regression. IVW regression was used to derive causal estimates where at least two exposure SNPs were available for analysis. The WM method would have a lower bias than the IVW method, which makes the causal inference more robust when the pleiotropy assumption is not satisfied . The MR-p-values less than 0.005 (0.05/11) were considered significant. MR analyses were performed in R 4.1.0 , and Cochran’s Q test. MR-Egger intercept test and MR-PRESSO global test were used to assess horizontal pleiotropy, and the MR-PRESSO method could also check and correct for outliers. Cochran’s Q test was applied to detect heterogeneity. The Bonferroni correction method was used for multiple testing in the secondary MR analysis, and  R 4.1.0  using Tw R 4.1.0 , Mendeli R 4.1.0 , and MRP R 4.1.0  packages R 4.1.0 .Association analysis between tea intake and Alzheimer’s disease was evaluated using a generalized linear regression model. Participants were divided into eight categories according to tea intake amount, including non-drinkers as well as drinkers consuming less than 1 cup/day, 1–2 cups/day, 3–4 cups/day, 5–6 cups/day, 7–8 cups/day, 9–10 cups/day, 11–12 cups/day, and ≥13 cups/day. Binary AD data and polygenic risk scores (PRS) of AD from UKB were used in the regression model. Binary AD outcome was derived using ICD-10 codes (UKB field-ID: 41202) and self-reported non-cancer illness (UKB field ID: 20002). To better fit the model, we only included participants >65 years old from European ancestry. We used the PRSice2 software  to generThe study design is shown in IVW = 1.48, 95% CI: , p = 3.78 × 10–3; ORWM = 2.00, 95% CI: , p = 3.30 × 10–3). The MR-Egger regression estimate showed close to significance and converging results as IVW estimate and WM estimate . For each extra cup of tea, the odds of AD would be greatly increased by more than 1.48 folds. Along with the insignificant but increasing risk effect of tea consumption, the results showed that per extra cup of tea intake might increase the risk of AD. Sensitivity analysis did not find any pleiotropy effect or heterogeneity, which further indicated the reliability of our results. The analysis had 97% power for detecting the causal association . Secondly, tea intake was associated with decreased gray matter volume  while tea intake was significantly associated with increased white matter volume . The opposite effect directions of tea intake on gray and white matter volumes may elucidate how tea intake affects AD via brain volumes. Meanwhile, we also found that tea intake was strongly associated with decreased right hippocampus volume, which could explain the interaction between tea intake and AD . We did not find any significant association between tea intake and other secondary outcomes.We found significant associations between tea intake and brain volumes . First, p = 1.91 × 10–2), gray matter volume (p = 1.91 × 10–2), and fractional anisotropy (p = 1.91 × 10–2), which suggested potential bias to the MR-Egger estimates. The Cochran’s Q test found heterogeneity in the association with total brain volume (p = 8.87 × 10–5), gray matter volume (p = 4.35 × 10–5), right hippocampus volume (p = 4.71 × 10–3), and fractional anisotropy (p = 1.98 × 10–2). The power analysis showed that the traits with significant results in secondary analysis had enough power of detecting the causal estimates (The MR-Egger intercept test did not find pleiotropy in any of the associations . Howeverstimates .p = 1.14 × 10–2). As the amount of tea intake increases, the protective effects are gradually dismissed and eventually become harmful. As shown, instead of protective effects, people who drink equal to or more than 13 cups of tea per day would have dramatically 67.4% higher odds of AD than non-drinkers . Detailed characteristics of tea intake and AD of 275,285 participants were summarized in The results using binary AD as the response showed a nonlinear association between tea intake and AD . CompareThe two-sample Mendelian randomization analysis found that an extra cup of tea consumed per day was significantly associated with the risk of AD. The association analysis has a consistent finding when the number of cups per day of tea exceeds six. In addition, we found that the number of cups of tea consumed per day was inversely associated with gray matter volume and right hippocampus volume but positively associated with whiter matter volume and total brain volume. Other than that, we did not find a causal association between tea intake and WMH volume, MD, FA, BMBs, and left hippocampus volume.This MR study satisfied the three assumptions of MR analysis. First, we selected genetic instruments using a relative strict cutoff value and the F-statistics of the selected SNPs were greater than 10, which ensures the genetic variants were strongly associated with the exposures. Secondly, after checking the SNPs at the phenoscanner, we rules out the possibility that the genetic variants may be associated with confounders. The MR-Egger test did not find pleiotropy in both primary analysis and secondary analysis, which suggested no violation of pleiotropy. Though, the power of the analysis could be decreased by the small proportion of variance explained, since we have large F-statistics, the power of the analysis was not be greatly influenced.Our MR analysis revealed a risky effect of tea intake on AD. A nominal risk effect of tea intake was found that one extra cup of tea consumed per day would dramatically increase the risk of AD. Similarly, a recent two-sample MR study showed that per extra cup of coffee was associated with an increased risk of AD . A possiIn addition, a nonlinear relationship between tea intake and AD was reported by the association analysis, suggesting daily consumption of 5–6 cups to be the most protective and indicating possible increased risk effects for daily consumption of greater than 13 cups of tea. Similarly, a recent cohort study on UK Biobank also finds a nonlinear relationship between tea intake and risk of dementia and suggested daily consumption of three cups of tea to be the strongest protective . HoweverWe further detected that the amount of tea consumed was associated with brain volumes, which may give clues on how tea intake affects AD. Tea intake was inversely related to gray matter and right hippocampus volume, and positively associated with total brain and white matter volume. Other studies have found that diet or eating behaviors could impact brain volumes –45. For p < 0.05) and is also weak significant in MR-Egger regression (p < 0.1). Also, the primary analysis results passed all the sensitivity analyses, including heterogeneity, pleiotropy, and outliers, demonstrating our results’ robustness and reliability. Meanwhile, when using the conventional genome significance level p < 5 × 10–8 to select genetic instruments, the results still showed that tea intake was significantly associated with increased odds of AD in three methods  and passed all the sensitivity analysis (There are four main strengths of the present study. First, our study is the multiple dimensional investigations of the associations between tea consumption and AD, including association analysis and causal inference using two-sample MR with two phenotypes of tea consumption. Using large-scale GWAS summary statistics from IGAP and UK Biobank rules out potential instrument biases and ensures the credibility and validity of our results. Therefore, our conclusion that tea consumption was associated with an increased risk of AD could be considered comprehensive and robust based on the converging results from different approaches. Second, the main conclusion of causality of tea intake on AD was significant in IVW and WM methods (analysis . A consiThere are some limitations in our analyses. First, the MR analysis of tea consumption and AD was insignificant using an independent AD dataset as replication. However, the overall direction of the replicated MR analyses matched the increasing risk effect of tea consumption. Second, partial sample overlap issues in the exposure and outcome data may bias the results of two-sample MR. The data source of brain volumes, MD, and FA were from UK Biobank . For theIn conclusion, the MR study robustly demonstrated that the genetically predicted tea intake is a causal risk factor for AD and suggested that the risk effects of tea intake on AD may be contributed by decreasing the volume of gray matter and the right hippocampus, which are associated with the decline in language and memory functions. Our results provide more insights into the biological mechanism of tea effect on AD and deliver suggestive guidance of the amount of tea intake per day of tea as an intervention for AD.The original contributions presented in this study are included in the article/YS performed the analysis and wrote the manuscript. RS contributed to the conception, designed the research, and revised the manuscript. All authors approved the final manuscript.


Endoscopic biopsy is recommended for diagnosis of nasopharyngeal carcinoma (NPC). A proportion of lesions are hidden from endoscopic view but detected with magnetic resonance imaging (MRI). This systematic review and meta-analysis investigated the diagnostic performance of MRI for detection of NPC.An electronic search of twelve databases and registries was performed. Studies were included if they compared the diagnostic accuracy of MRI to a reference standard (histopathology) in patients suspected of having NPC. The primary outcome was accuracy for detection of NPC. Random-effects models were used to pool outcomes for sensitivity, specificity, and positive and negative likelihood ratio (LR). Bias and applicability were assessed using the modified QUADAS-2 tool.CI 95.2–99.3%), specificity of 91.7% (95% CI 88.3–94.2%), negative LR of 0.02 (95% CI 0.01–0.05), and positive LR of 11.9 (95% CI 8.35–16.81) for detection of NPC. Most studies were performed in regions where NPC is endemic, and there was a risk of selection bias due to inclusion of retrospective studies and one case–control study. There was limited reporting of study randomization strategy.Nine studies were included involving 1736 patients of whom 337 were diagnosed with NPC. MRI demonstrated a pooled sensitivity of 98.1% (95% This study demonstrates that MRI has a high pooled sensitivity, specificity, and negative predictive value for detection of NPC. MRI may be useful for lesion detection prior to endoscopic biopsy and aid the decision to avoid biopsy in patients with a low post-test probability of disease. Nasopharyngeal carcinoma (NPC) has unique epidemiology, natural history, and treatment response necessitating distinct management paradigms . It encoBiopsy of the primary tumor is required for definitive diagnosis  and is gMRI is established as the preferred modality for locoregional staging of NPC , 10, 12,Unfortunately, the vast majority of patients present with advanced disease, with less than 8% presenting at stage I . Beyond The primary endpoint of this systematic review and meta-analysis is to pool the sensitivity, specificity, likelihood ratios (LRs), and hierarchical summary receiver-operating characteristics (HSROC) of MRI for primary detection of nasopharyngeal carcinoma relative to the reference standard of post-MRI endoscopic biopsy. Secondary endpoints are to identify barriers by way of qualitative or quantitative factors that influence diagnostic accuracy.The search strategy was devised in accordance with the revised PRISMA 2020 statement  and regiWe included prospective and retrospective studies that assessed performance of MRI for diagnosis of NPC in patients with suspected disease, compared to histopathology as the reference standard. Following full-text review, articles were included if they reported sensitivity and specificity values for detection of primary NPC. Papers for which this was not possible were excluded. Articles which assessed accuracy for detection of recurrent tumor were excluded. Studies with risk of overlapping cohorts were scrutinized and the most complete dataset was included to avoid duplication of data. Abstracts, case reports, case series, editorials, and prior systematic reviews or meta-analyses were also excluded due to risk of publication bias and duplication. The selection process was summarized graphically per the PRISMA  guidelinTwo authors (VVG and NNN) independently extracted data including study design, patient demographics, inclusion and exclusion criteria, cohort enrolment and MRI-technique , diagnostic criteria utilized, and diagnostic accuracy data . If multiple diagnostic features were assessed, we included the parameter with best diagnostic performance in the meta-analysis. If data were ambiguous or could not be extracted into 2 × 2 tables, the authors of the paper in question were contacted for clarification where possible. Discrepancies between the 2 reviewers were resolved by discussion and consensus, and results reviewed by the senior investigator (MVC).Included studies were assessed using the revised Quality Assessment of Diagnostic Accuracy of Studies (QUADAS-2) tool . This top-value < 0.05 was considered statistically significant. To pool diagnostic accuracy measures, we used a bivariate mixed-effects regression model that allows correlation between sensitivity and specificity. Results were presented as summary sensitivities, specificities, LRs, and ROC AUC values with 95% CIs. A positive LR > 10 or a negative LR < 0.1 was considered to be strong diagnostic evidence [Statistical analysis was performed using STATA v14.1 . A two-tailed evidence .Interstudy variability was assumed, and an exploration of the causes of variability, including study design (prospective vs. retrospective) differences, was performed using meta-regression. Statistical analyses of variability and publication bias are not included in this report in accordance with updated recommendations by the PRISMA Diagnostic Test Accuracy Group . The aren = 5), post hoc assessment of known nasopharyngeal tumor (n = 7), or for whom data could not be extracted into 2 × 2 tables (n = 2) or a full text could not be obtained (n = 1), there were 9 eligible studies for the meta-analysis. A summary of the study selection process according to the PRISMA format is presented in Fig. A total of 5886 references were identified through searches of 12 electronic databases and registers, of which 23 met criteria for full-text review. Manual search through reference lists yielded 1 additional relevant study. Following exclusion of studies with overlapping cohorts , specificity of 91.7% (95% CI 88.3–94.2%), negative LR of 0.02 (95% CI 0.01–0.05), and positive LR of 11.9 (95% CI 8.35–16.81). The area under the curve (AUC) of the sROC was 0.98 (95% CI 0.97–0.99)  or specificity (p = 0.83) in meta-regression models. There was insufficient data regarding lesion size or tumor stage to facilitate further subgroup analysis.Study design (retrospective vs. prospective) did not demonstrate a significant impact on diagnostic sensitivity (Although MRI was generally performed prior to biopsy, only one study  reportedNasendoscopy and biopsy have long been established as the gold standard for diagnosing NPC. However, NPC may be submucosal or contained within the pharyngeal recess and thus difficult to identify at endoscopy , resultiOur meta-analysis supports the use of MRI as an accurate primary diagnostic tool, with a pooled sensitivity of 98.1%. MRI was also found to have excellent specificity and negative LR. MRI-detected abnormalities could direct biopsy planning whereas a normal MRI examination would establish a low post-test probability of disease. The applicability of these findings is strengthened by the MRI protocols involved, which included T2-weighted and pre- and post-gadolinium T1-weighted sequences on 1, 1.5 and 3 T magnets at slice thicknesses between 1 and 5 mm, reflecting real-world heterogeneity of protocols in clinical practice .The implication for current guidelines is that MRI could be the initial diagnostic test prior to endoscopic biopsy. Pre-operative MRI may increase the yield of endoscopic biopsy by alerting the surgeon to multiple lesions or lesions which may be endoscopically occult, and aid selection of patients who may benefit from intraoperative image-guided stereotactic approaches. The negative LR of MRI implies greatly reduced post-test probability of disease. MRI is a non-invasive technique which does not require anesthesia, yields excellent soft-tissue characterization of the lateral pharyngeal recess, submucosal soft tissues, and retropharyngeal nodes , and alrThe major pitfall for diagnosis highlighted in the included studies is false-positive results due to asymmetric nasopharyngeal hyperplasia , reporteThe results of this systematic review and meta-analysis should be considered in context of several limitations. Only two studies , 34 repoGadolinium-enhanced nasopharynx MRI demonstrates exceptional sensitivity, positive and negative LR for diagnosis of nasopharyngeal carcinoma with good specificity. The results support use of MRI as a primary diagnostic tool for lesion detection, in addition to its established role in locoregional staging. If used prior to endoscopic biopsy, it may increase the yield of biopsy or aid the decision to avoid biopsy in patients with low post-test probability of disease.


Emergency laparotomy (EL) has a high mortality rate. Clinically, frail patients have a poor tolerance for EL. In recent years, sarcopenia has been used as an important indicator of frailty and has received much attention. There have been five different calculation methods of psoas for computed tomography (CT) to measure sarcopenia, but lack of assessment of these calculation methods in Eastern Asian EL patients.We conducted a 2-year retrospective cohort study of patients over 18 years of age who underwent EL in our institution. Five CT measurement values  were calculated to define sarcopenia. Patients with sarcopenia defined by the sex-specific lowest quartile of each measurement were compared with the rest of the cohort. The primary outcome was "ideal outcome", defined as: (1) No postoperative complications of Clavien-Dindo Grade ≥ 4; (2) No mortality within 30 days; (3) When discharged, no need for fluid resuscitation and assisted ventilation, semi-liquid diet tolerated, and able to mobilize independently. The second outcome was mortality at 30-days. Multivariate logistic regression and receiver operating characteristic (ROC) analysis were used.P < 0.001; PMI, P = 0.001; PMD, P = 0.157; TPG, P = 0.006; PBSA, P < 0.001; mortality at 30-days: PML3, P < 0.001; PMI, P = 0.002; PMD, P = 0.088; TPG, P = 0.002; PBSA, P = 0.001). In ROC analysis, the prediction model containing PML3 had the largest area under the curve (AUC) value .Two hundred and twenty-eight patients underwent EL met the inclusion criteria, 192 (84.2%) patients had an ideal outcome after surgery; 32 (14%) patients died within 30 days. Multivariate analysis showed that, except PMD, each calculation method of psoas was independently related to clinical outcome (ideal outcome: PML3, The sarcopenia determined by CT psoas measurements is significantly related to the clinical outcome of EL. The calculation of CT psoas measurement is suitable for application in outcome prediction of EL. In the future, it is necessary to develop a scoring tool that includes sarcopenia to evaluate the risk of EL better.The online version contains supplementary material available at 10.1186/s13017-022-00435-x. Emergency laparotomy has a high mortality rate , and decSarcopenia was first described by Irwin H. Rosenberg in 1988 and used to describe the age-related loss of skeletal muscle quantity and quality . EuropeaThe quantity and quality of muscles should be based on computed tomography (CT) or magnetic resonance imaging (MRI) as the gold standard –9. In prMany studies suggested that sarcopenia was associated with a poor prognosis of emergency laparotomy , 12–17; In the reported studies, there were five different psoas muscle calculation methods. Researchers had compared the prediction of three of them, psoas muscle index (PMI), psoas muscle to L3 vertebral body ratio (PML3) and psoas muscle to body face area ratio (PBSA), in European populations . Lu et aIn this study, we aimed to verify the universality of the conclusion that sarcopenia affected the prognosis of emergency laparotomy in a different population setting. In addition, we compared the ability of five different psoas calculations to predict clinical outcomes, which could be the basis for developing a more reliable risk prediction model.Our institution is a tertiary medical center located in central China. It has a case database, and medical records can be browsed in the local area network. The hospital's institutional review board passed the ethical approval of the study.This study selected adult patients who underwent emergency laparotomy in our prospective database from September 1, 2019, to August 31, 2021. All patients' information was retrieved from our hospital's database, including demographics, comorbidities, preoperative laboratory inspection results, weight, height, body mass index (BMI), American Society of Anesthesiologists (ASA) score, surgical procedures, intraoperative conditions, and prognosis, etc. The Charlson Comorbidity Index (CCI) ; categorical variables were expressed as ong test . P value2. Among the study population, 89 people (39.0%) had previous abdominal surgery; forty-four people (19.3%) were diagnosed with malignant tumors before or after surgery. The median Charlson Comorbidity Index score was 1.0 . Population baseline characteristics are shown in Table A total of 228 patients were enrolled in this study, including 138 (60.5%) men and 90 (39.5%) women. The average age of the population was 57.7 (± 15.8) years, and the average BMI value was 21.7 (± 3.5) kg/mOne hundred and eighteen (51.8%) patients received major surgeries and 110 48.2%) patients received minor surgeries. The commonest surgeries were small bowel resection (24.6%), perforation repair (18.4%), and appendectomy (11.8%). Thirty-three (14.5%) patients underwent more than one of the procedures, with the commonest surgery being small bowel resection combined with abdominal wall hernia repairment (3.5%). The operative procedures are shown in Table % patientThe sex-specific cut-off values for the five psoas muscle measurements are shown in Table We divided the patients into "Sarcopenia" group and "Non-Sarcopenia" group according to each of these five different calculations, respectively.P value = 0.063), the sarcopenia defined by the psoas major measurement values was age-related. In addition, the level of serum albumin was related to the sarcopenia determined by each calculation method.In the baseline characteristics Table , except P value = 0.115) and TPG , sarcopenia was also associated with complications with Clavien-Dindo grade ≥ 3. Besides, the sarcopenia defined by each psoas major measurement value was related to respiratory infection; only the sarcopenia defined by PMD  was related to abdominal infection.Regarding surgical outcome Table , sarcopeThe sarcopenia defined by each psoas major measurement value was closely related to the ideal outcome defined by us and mortality at 30-days. The same result also applied to the length of ICU stay.We performed a univariate regression analysis including the factors related to the prognosis Table .Table 6UEach psoas muscle calculation was related to the ideal outcome, mortality at 30-days, and the occurrence of complications with a Clavien-Dindo score ≥ 2.We performed a multivariate logistic regression analysis on the ideal outcome and mortality at 30-days Table . The incP value = 0.157 and 0.088, respectively) was not an independent risk factor.In all regression models, only PMD . In the study of Tzeng et al., PMD can be used as an independent risk factor for the postoperative hospital stay in In our study, sarcopenia defined by PMD cannot be used as an independent risk factor for clinical outcome in a multivariate analysis  and National Emergency Laparotomy Audit (NELA) models , 36. HowR2 values were larger in the model with the sarcopenia parameter than in the model without , so it was unlikely to evaluate whether the model's predictive ability would be improved by including the psoas muscle measurements as variables. We did not follow up with the patients for a long time, so we cannot evaluate the long-term clinical outcome in this study. We also did not prospectively collect parameters such as nutritional scores or muscle strength measurements to evaluate the patient's skeletal muscle state, so it was impossible to evaluate whether the sarcopenia determined by CT and the set cut-off values were consistent with the clinical diagnosis.The measured values of psoas major muscle determined by CT, except PMD, can be used as an independent risk factor for the prognosis of emergency laparotomy. Large sample research may be needed to accurately determine the CT psoas muscle measurement value as the cut-off value of the diagnostic criteria for sarcopenia. A prognostic model including a sarcopenia parameter should be developed in the future.Additional file 1: The statistical evaluation of the models.


Liver injury (LI) has been frequently observed in patients with dilated cardiomyopathy (DCM), whereas its prognostic value remains blurry. We attempted to appraise the prognostic effect of LI in patients with DCM.This retrospective study included 523 patients with DCM. LI was defined as a threefold increase in aspartate transaminase (≥ 135 U/L) or alanine transaminase (≥ 180 U/L) or a twofold increase in total bilirubin (≥ 41 umol/L) during hospitalization. The population was segmented into non-liver injury (NLI) group and LI group based on liver function test data. To balance differences in covariates at baseline, 1:1 propensity score matching (PSM) was performed.P < 0.001). Similar results were also found in age  and gender stratified analysis . After PSM, the survival rate of patients with LI remained lower than those with NLI . Multivariable Cox regression analysis manifested that LI  showed potent predictive effect on all-cause mortality in patients with DCM, both before and after PSM.Patients with LI had lower survival rate, compared with those with NLI (44.6% vs. 73.8%, The occurrence of LI herald adverse outcomes in patients with DCM and attention to LI may be conducive to risk stratification and management.The online version contains supplementary material available at 10.1186/s40001-022-00876-9. As early as 1930, N. Jolliffe reported a phenomenon of heart failure (HF) accompanied by abnormal liver function (ALF), which predominantly referred to the elevation of bilirubin . A numbeOf note, ALF is not entirely equivalent to liver injury (LI), that is, dysfunction of bilirubin metabolism and damage of hepatocytes , similarThis retrospective study was performed at the First Affiliated Hospital of Guangxi Medical University and included 523 hospitalized DCM patients with HF admitted from August 2012 to May 2020. DCM was defined as left ventricular end-diastolic dimension (LVEDD) > 55 mm for males or > 50 mm for females and left ventricular ejection fraction (LVEF) < 45%, which cannot be explained by hypertension or coronary artery disease, and valvular or congenital heart disease were absent. The diagnosis of dilated cardiomyopathy strictly followed Chinese guidelines. LI was defined as a threefold increase in AST (≥ 135 U/L) or ALT ≥ 180 U/L) or a twofold increase in TB (≥ 41 umol/L) at any time during hospitalization [0 U/L or Exclusion criteria were: (1) chronic liver disease; (2) drug-induced liver injury; (3) acute or chronic biliary tract disease; (4) severe inflammatory or infectious diseases; (5) cardiopulmonary resuscitation; (6) new or old myocardial infarction; (7) coronary revascularization; (8) coronary artery disease complicated with myocardial ischemia; (9) acute myocarditis; (10) chronic kidney disease 5 period or need for hemodialysis; (11) heart transplantation; (12) death in hospital; (13) liver function test and cardiac ultrasound data missing; (14) traumatic patients.All patients underwent physical examinations, blood laboratory tests, 12 lead electrocardiogram and echocardiography during the duration of hospital stay. Medical history collection, physical examinations, 12 lead electrocardiogram and medical records were finished within 8 h of admission. Blood tests of all patients were performed in our central laboratory. Cardiac ultrasonography was implemented by experienced sonographers. Echocardiographic parameters were measured according to the guidelines of the current year , 20, andt test or Wilcoxon rank sum test was used between the two groups, as appropriate. Qualitative data were compared using Chi-square test or Fisher exact test. Ordinal data were compared using Wilcoxon rank sum test. The significance comparison of Kaplan–Meier survival plots was tested by log-rank between the two groups. The initial timepoint for survival analysis was the day of discharge. Schoenfeld residuals were computed to test proportional hazard assumption. Relevant parameters for risk of the primary endpoint were appraised by Univariable Cox regression analysis. Subsequently, parameters with P < 0.05 were included in Multivariable Cox regression analysis, and were screened using the “forward” option. To balance differences in covariates at baseline between groups and enhance the reliability of results, propensity score matching (PSM) was performed. These covariates involved in age, sex, diabetes mellitus, NYHA class, systolic blood pressure (SBP), ALB, blood–urea–nitrogen (BUN), N-terminal pro-B-type natriuretic peptide (NT-proBNP), left atrium diameter (LAD), and renin–angiotensin system (RAS) inhibitor. Considering the ratio of two groups of patients, a 1:1 nearest-neighbor matching was implemented. All statistical analysis was executed by SPSS 23.0 and R 4.2.0. All tests were two-sided. The statistical significance was determined by P < 0.05.Patients were discriminated into non-liver injury (NLI) group and LI group based on liver function test data. Statistical description of quantitative variables was displayed as the mean ± standard deviation or median (interquartile) according to distribution pattern. Classification variables were statistically described in number (percentage). For comparison of quantitative variables, Demographic variables, clinical and behavioral features of the population were exhibited in Table P > 0.05).After PSM, the number of DCM patients in the LI and NLI groups was equivalent 92 vs. 92) (Table  vs. 92 (P < 0.001)     in the LI group and 113 cases  in the NLI group. The Kapan–Meier survival curves testified that patients with LI had lower survival rate than those with NLI . SBP, NT-proBNP, LAD, LVEDD, LVEF, RAS inhibitor and LI played similar prognostic roles in the Univariable Cox regression models after baseline adjustment using PSM .The prediction effect of population characteristics, comorbidities, assessment of cardiac function, hematological indicators, cardiac ultrasound data, and medication and instrument therapy on the primary endpoint are shown in Table P = 0.28) , both before and after PSM  Fig. . In the P < 0.001; HR: 0.973, 95% CI 0.958–0.988, P = 0.001, respectively), NT-proBNP  and LVEDD  all showed strong independent prognostic value, while LVEF  was only independently associated with prognosis after PSM.The risk of death was approximately 1.7 times higher in LI patients than in NLI patients. However, AST, ALT, and TB did not show independent prognostic effects when they were included instead of LI in the multivariable Cox regression model. In addition, after adjusting for confounding factors, SBP  , 9, 13, When AST, ALT and TB were included as a continuous variable in the final model, they did not show an independent predictive value for prognosis in patients with DCM. The reason for this may be that in this study, the changes of liver function indicators within the normal range or slight increase had no potential clinical implication. Only when LI occurred, the prognostic value of liver function indicators was highlighted. Although previous studies , 13 do nThere are no epidemiological studies on AFL or LI in outpatients with cardiovascular disease. A reported ALF occurs in 1–4% of asymptomatic patients . We specWith the early diagnosis and etiological intervention, the development of comprehensive drug therapy and the improvement of device therapy, the survival rate of DCM patients has been considerably improved . During The liver is supplied by both the portal vein (70%) and the hepatic artery 30%), and its blood flow accounts for about one-fifth to one-quarter of the cardiac output . It is i%, and itDCM is characterized by enlarged LVEDD and reduced LVEF, and its underlying molecular mechanism involves activation of the neurohumoral system, which forms the pathophysiological basis of LI occurrence , 36. TheThe protective effect of higher SBP was consistent with previous studies , 43, and2) in univariate COX analysis was not significantly associated with the prognosis of patients with DCM, which may be due to the exclusion of patients with severe chronic kidney disease. Furthermore, the use of RAS inhibitor may improve kidney function, thus affecting its prognostic effect on patients with DCM. More importantly, the vast majority of patients did not have chronic kidney disease and the decrease in eGFR may be a transient phenomenon.Vincenzo Nuzzi et al. found that permanent atrial fibrillation (AF) had a similar effect on the prognosis of DCM patients as LI in this paper, while paroxysmal or persistent AF presented neutral results . HoweverAs far as we know, the reverse action mechanism of cardiogenic LI on the heart may include the following three aspects. First, the development of LI may promote the release of inflammatory factors, activate cell signaling pathways, and cause myocardial dysfunction, similar to those found in cirrhotic cardiomyopathy . Second,We acknowledge that this report had some limitations. The general flaws of retrospective studies were also not immune to this study. First, in addition to liver function indicators, we had no additional liver morphological, pathological, and hemodynamic information to adequately assess the severity of LI and its impact on prognosis. Second, we did not analyze the duration and recovery of LI, and these data may have different effects on the results. Third, the absence of myocardial injury-specific markers prevented us from reliably ruling out the role of myocardial injury in the outcome. Fourth, the lack of data on the assessment of right heart function prevented us from considering all factors that might have influenced the results. Finally, variations in patient medication and device therapy were not recorded during follow-up. This may constitute a latent factor that interfered with the results, even if the baseline levels of drugs were balanced.The results of this study, which incorporated numerous patients with long-term follow-up, revealed that the occurrence of LI predicted a potentially bad prognosis in patients with DCM.Attention to LI is beneficial to identify high-risk DCM patients and guide the next management.Additional file 1: Table S1. Mean dose of drug during hospitalization in the study population. Table S2. Cause of death in the study population. Figure S1. Kaplan–Meier curves of stratified analysis showed the occurrence of the primary outcome in patients with and without liver injury. (A) age ≤ 50 years, (B) age > 50 years, (C) male, (D) female.


Computed tomography angiography (CTA) is a cornerstone in the pre- transcatheter aortic valve replacement (TAVI) assessment. We evaluated the diagnostic performance of CTA and coronary artery calcium score (CACS) for CAD evaluation compared to invasive coronary angiography in a cohort of TAVI patients.In consecutive TAVI patients without prior coronary revascularization and device implants, CAD was assessment by quantitative analysis in CTA. (a) Patients with non-evaluable segments were classified as obstructive CAD. (b) In patients with non-evaluable segments a CACS cut-off of 100 was applied for obstructive CAD. The reference standard was quantitative invasive coronary angiography .100 consecutive patients were retrospectively included, age was 82.3 ± 6.5 years and 30% of patients had CAD. In 16% of the patients, adequate visualization of the entire coronary tree  was possible with CTA, while 84% had at least one segment which was not evaluable for CAD analysis due to impaired image quality. On a per-patient analysis, where patients with low image quality were classified as CAD, CTA showed a sensitivity of 100% (95% CI 88.4–100.0), specificity of 11.4% (95% CI 5.1–21.3), PPV of 32.6% (95% CI 30.8–34.5), NPV of 100% and diagnostic accuracy of 38% (95% CI 28.5–48.3) for obstructive CAD. When applying a combined approach of CTA  and CACS , the sensitivity and NPV remained at 100% and obstructive CAD could be ruled out in 20% of the TAVI patients, versus 8% using CTA alone.In routinely acquired pre-TAVI CTA, the image quality was insufficient in a high proportion of patients for the assessment of the entire coronary artery tree. However, when adding CACS in patients with low image quality to quantitative CTA assessment in patients with good image quality, obstructive CAD could be ruled-out in 1/5 of the patients and may therefore constitute a strategy to streamline pre-procedural workup, and reduce risk, radiation and costs in selected TAVI patients without prior coronary revascularization or device implants.The online version contains supplementary material available at 10.1186/s12872-022-02623-8. Computed tomography angiography (CTA) is an essential component of the pre-interventional transcatheter aortic valve replacement (TAVI) assessment . It provAll patients undergoing TAVI require pre-procedural assessment of coronary anatomy . InvasivAs the indication for TAVI expands to lower risk patients with modest pre-test probabilities for obstructive CAD, the added benefit of an invasive test that is associated with incremental risk, radiation and cost may be of limited value with important implications to both patients and healthcare systems . WhetherThe aim of the current investigation therefore, was to clarify the value of pre-TAVI CTA in delineating coronary anatomy by assessing its diagnostic performance (using both quantitative assessment of CTA and CACS and its combination) as compared to invasive quantitative coronary angiography (QCA) in a prospective cohort of consecutive TAVI patients. We hypothesize that the diagnostic performance of combined quantitative CTA/CACS assessment is such that CAD can be safely and reliably ruled out in select TAVI patients and ultimately obviate the need for additional evaluation with invasive coronary angiography.All patients undergoing TAVI at Bern University Hospital, Bern, Switzerland, are consecutively enrolled into a prospective institutional registry that is a part of the Swiss TAVI registry NCT01368250) . The reg250 . ThePre-TAVI CT was performed as previously published . A standA blinded experienced reader (MH), unaware of the clinical history and results from the invasive measurements, retrospectively performed quantitative CTA data analysis of the coronary arteries on a dedicated software  after selecting the optimal reconstructed phase with regard to image quality. Coronary arteries were assessed according to the AHA segments  and eachThe CACS from each coronary artery was obtained using the Agatston method  and summAs in the case of the CT assessment, an experienced blinded reader (AC) analyzed coronary angiograms without the knowledge of clinical history or results from CT angiography. Coronary arteries were assessed according to the same segmented model used for the CT analysis . Quantitt test or Mann–Whitney U test, as appropriate. Categorical data were analysed with χ2 test or Fisher’s exact test. The p values of all outcomes were two-sided and values less than 0.05 were considered as statistically significant. CI was defined as 95%.Patient information and results are reported for the entire cohort and grouped according to the presence or absence of obstructive CAD. Data are reported as median ± IQR from 25 to 75th percentile, mean ± SD or percentages, as appropriate. Continuous variables were analysed using the Student The pre-specified diagnostic accuracy was performed on a per-patient level with the reference standard of CAD as ≥ 50% and ≥ 70% stenosis on invasive QCA in one of the major epicardial coronary arteries according to the AHA-segment model. Furthermore, using the same cut-off in QCA was used on a per vessel level and on a per segment and patient level. In the first analysis, diagnostic accuracy of CTA was determined on an intention-to-diagnose basis, meaning that no coronary segment was excluded; non-evaluable segments were rated as stenosed, as previously reported . In a seBetween 06/2018 and 02/2019, 100 consecutive patients with severe aortic stenosis undergoing pre-TAVI CT at the Swiss Cardiovascular Center, Bern University Hospital, who fulfill the inclusion and exclusion criteria were analyzed for the present study. In detail, of the 211 patients with severe aortic stenosis who underwent a pre-TAVI CT, 45 patients with prior percutaneous coronary interventions with stent implantation, 15 patients with coronary artery bypass grafting, 7 patients with valve- in valve procedure, 5 patients with pacemakers, and 39 patients with external CTAs were excluded, resulting in 100 patients for the purpose of the present analysis.p < 0.001).The baseline clinical characteristics of the cohort are shown in Table In 16 (16%) out of 100 patients, adequate visualization of the entire coronary tree  was possible with CTA, while 84 (84%) of 100 patients had at least one segment which was not evaluable for CAD analysis due to impaired image quality (e.g. motion artefacts). Out of the 84 patients with non-evaluable segments, a total of 578 segments (578/1533 = 38%) showed a CTA image quality that was insufficient for quantitative CAD evaluation. When focusing only on the left main and proximal segments, 54 out of 100 patients could be assessed, whereas 46% of patients showed at least one segment  which depicted impaired image quality. On a segment analysis, 301 (75%) out of 400 left main and proximal segments showed good image quality.The findings of the performance of CTA in diagnosing CAD using 50% diameter stenosis in invasive QCA as a reference are summarized in Table When assessing only the left main and proximal segments, 15% showed at least a 50% stenosis in invasive QCA. On a per-patient analysis, when non-evaluable segments were rated as possible obstructive CAD, CTA showed a sensitivity of 80% (95% CI 51.9–95.7), specificity of 29.4% (95% CI 20.0–40.03), positive predictive value of 16.7% (95% CI 13.0–21.1), negative predictive value of 89.3% and diagnostic accuracy of 37% (95% CI 27.6–47.2) for obstructive CAD. On a per-patient analysis excluding patients with non-evaluable segments, , CTA had a sensitivity of 57.1% (95% CI 18.4–90.1), specificity of 53.2% (95% CI 38.1–68.9), positive predictive value 15.4% (95% CI 8.2–27.0), negative predictive value of 89.3% and diagnostic accuracy of 53.7% (95% CI 39.6–67.4).The findings on a per segment analysis for 50% diameter stenosis are depicted in Table p < 0.001).The sensitivities and specificities of different CACS thresholds to diagnose a ≥ 50% diameter stenosis on invasive coronary angiography are shown in Fig. To our knowledge, this is the first blinded study assessing the diagnostic performance of quantitative CTA, CACS and combined quantitative CTA/CACS for CAD in a cohort of consecutive patients undergoing TAVI as compared to the gold standard of quantitative invasive coronary angiography. Our analysis shows that (1) a significant proportion of patients (> 80% of patients) have coronary artery segments that are not evaluable by pre-TAVI CTA where per our institutional protocol no medication for rate control or vasodilatation was used; however, when only focusing on the left main and proximal segments, 56% of the patients and 75% of the segments could be assessed (2) in patients with non-evaluable coronary artery segments, a combined approach using CTA and CACS, resulted in high accuracy for ruling-out CAD (i.e. in 20% of the patients), however with poor diagnostic accuracy. These findings provide evidence that pre-procedural diagnostic workup may be streamlined in selected TAVI patients by potentially omitting routine invasive coronary angiography in selected patients. However, as we have only included patients without prior revascularization or device implants, it has to be noted that these findings are only applicable to these selected patient cohorts. Further, as a high number of TAVI patients show evidence of CAD in CTA or may present with impaired CTA image quality, invasive coronary angiography will still be necessary in a large proportion of patients.Previous investigations have evaluated the pre-TAVI CTA as a diagnostic tool to assess CAD , 22, 23.Although a strength of the current study is the inclusion of a prospective cohort of contemporary patients who underwent TAVI with blinded quantitative CTA/CACS and QCA analysis, the results may not be generalizable to all TAVI patients due the inclusion criteria and low patient number. In fact, as there were quite a large patient number excluded due to prior PCI/stent implantation, coronary artery bypass grafting and previous device implants, the results are only applicable for these selected patients. It remains unclear whether different approaches using CTA alone, combined with CACS or CACS alone using different thresholds according to age, gender and clinical CAD pre-test likelihood are required. Moreover, future studies should aim to investigate these research question in a larger cohort, e.g. in a multi-center trial.In routinely acquired pre-TAVI CTA, the image quality was insufficient in a high proportion of patients for the assessment of the entire coronary artery tree. However, when adding CACS in patients with low image quality to quantitative CTA assessment in patients with good image quality, obstructive CAD could be ruled-out in 1/5 of the patients and may therefore constitute a strategy to streamline pre-procedural workup, and reduce risk, radiation and costs in selected TAVI patients without prior coronary revascularization or device implants.Additional file 1: Table S1. CTA/CACS versus QCA as the reference standard (≥ 70% stenosis).