Among the starch concentrations  investigated, 15% starch concentration provided the best control over the shape of the beads due to its high storage modulus (8947 Pa), indicating higher gel strength. At this condition, the starch beads revealed an average size of ~650 µm, which was significantly smaller than the beads produced with other starch concentrations (10 and 12.5%), and had a density of 0.23 g/cm3. However, at lower starch concentrations (10%), the beads were not able to retain their spherical shape, resulting in larger beads (812–3501 µm). Starch crystallinity decreased by gelatinization, and the starch beads exhibited a porous structure, as observed from their SEM images. Overall, 3D food printing can be an alternative approach to preparing porous beads for the delivery of bioactive compounds with high precision.This study introduces a 3D food printing approach to fabricate spherical starch beads with small sizes and high porosity for the first time. The results illustrated that 3D food printing could generate starch beads in different sizes depending on the nozzle diameter, printing pressure, and ink viscosity. The 3D-printed beads were characterized for their morphology, crystallinity, and textural properties, while the starch-based ink was analyzed for its rheological properties. A suitable printing was attained when viscosity was in the range of 1000–1200 Pa.s at a low shear rate (˂0.1 s Advanced materials with unique properties such as nanoporous structures have received great attention due to their potential applications in modern high-tech fields, including biotechnology and food industries . PotentiAmong food-grade biopolymers, starch has received great attention due to its ability to create a 3D open porous structure by simply gelatinization upon heating. In addition, starch can be easily incorporated into various food formulations since it is abundant, non-toxic, inexpensive, biocompatible, and part of many food systems ,5. StarcStarch hydrogels were successfully shown to increase the bioaccessibility of bioactive compounds such as curcumin  and polyAdditive manufacturing (AM) or 3D printing has recently been adapted to manufacture food products . AM techThus far, 3D food printing has been employed to create 3D geometric shapes of basic food formulations such as potato puree, chocolate, meat paste, cheese, and dough ,13. CompIn addition to creating macroscopic geometric shapes, 3D food printing can be used to precisely control the microstructure of foods. When the technologies of 3D food printing and functional porous materials  are combined, their performance, structure, and application scope can be broadened, especially where intricate structures and shapes are desired. Direct printing of an object with a porous structure is critically challenging, which was recently addressed using polymers with inherent porosity ,14,15. TThe sol–gel method, the main procedure typically applied to make polymer-based hydrogels, can be simply adapted to formulate ink suitable for 3D printing technologies. However, given the demand for both biological function and physical characteristics, the utilization of biopolymers for 3D printing is highly challenging ,14,15. RHigh amylose corn starch (HACS) with 72% amylose content and 11% moisture content (Hylon VII) was kindly obtained from Ingredion  and used as the raw material to generate beads.w/w) were prepared. These starch concentrations were selected based on the preliminary 3D food printing experiments. The samples were then heated up to 95 °C under high shear (4260 rpm) using a Thermomix  and maintained at this temperature for 25 min, and then immediately used as an ink for 3D food printing.First, aqueous starch suspensions of three concentrations  that was equipped with two 10 mL extruders. The ink extrusion was empowered by pneumatic pressure .Next, the developed starch paste was loaded into a 10 mL cartridge, which was then transferred to one of the extruders of the 3D printer previously warmed up to 95 °C. The starch hydrogel was then extruded through nozzles with different diameters  . The nozDuring printing, the cartridge temperature was kept constant at 95 °C. The extrusion pressure was optimized for each printing condition  in the range between 0.027 and 0.17 MPa. The printing speed was kept constant at 6 mm/s, and the Bioprint Pro software was used to print spherical beads. The starch-based ink was deposited at room temperature (23 °C). Retrogradation was performed by keeping the gel beads at 4 °C, and then the beads were transferred to a freezer −80 °C, and finally dried for 24 h at −108 °C using a freeze dryer . High amylose corn starch and the 3D-printed starch beads with starch concentrations of 10, 12.5, and 15% were thereafter referred to as HACS, SP-10, SP-12.5, and SP-15, respectively. All 3D food printing experiments were conducted in triplicates.The bead size and sphericity of the 3D-printed starch beads were determined from their images using ImageJ software . Randomly selected 50 beads were used to determine the mean bead size ± standard deviation and sphericity ± standard deviation.FEI NovaNanolab200  Dual-Beam system equipped with a 30 kV SEM FEG column and a 30 kV FIB column was utilized to visualize the structure of the starch beads. The specimens were prepared by cutting cross-sections from the starch beads, and then they were sputter-coated with a gold layer  to prevent electrical charging. Finally, the SEM images were taken at an acceleration voltage of 10 kV and a current of 10 mA.Furthermore, the macropore size distribution was determined from the SEM images by measuring the size of randomly selected 50 open pores using ImageJ software. The thickness of the beads’ surface was also measured using ImageJ software.b). The volume of the beads was determined via measuring the beads’ dimensions using ImageJ software . The true density was measured using a helium pycnometer . Finally, the porosity (ε) was then determined using the bulk and true density [The weight of a certain amount of the starch beads was divided by their volume to calculate the bulk density  unit  equipped with a Quest attenuated total reflectance (ATR) accessory . The FTIR spectrum was acquired in the range of 4000 to 400 cm64 scans .The crystalline structure of the 3D-printed starch beads was evaluated through an X-ray diffraction (XRD) analysis. The XRD patterns were recorded using a PW3040X’PertMRDθ) with a step size of 0.02 at 45 kV and 40 mA [High-Resolution X-ray diffractometer . The powdered starch beads were packed into the sample holder, and then they were scanned from 5° to 40°  measurements were performed using a PerkinElmer DSC 4000 calorimeter . Briefly, 8 μL of deionized water was added to 3.5 mg (dry basis) of powdered starch beads and sealed in a 50 μL aluminum pan. The DSC measurements were then conducted in a heating cycle from 25 °C to 150 °C at a rate of 5 °C/min under a nitrogen atmosphere. The DSC data were used to determine the gelatinization temperature as well as the melting point of amylose–lipid complexes.−1 at 95 °C.A controlled-stress rheometer  equipped with a Peltier Plate temperature-controlled system and a 40 mm parallel-plate geometry was used to measure the rheological properties. Rheological measurements were performed according to the method reported by Chen et al.  with minw/w) and loading it between the parallel plate geometry with a 40 mm diameter and 1 mm gap. The moisture evaporation from the samples was prevented by covering the edge of the samples with silicon oil. Afterward, the temperature sweeps were carried out from 50 °C to 95 °C at a rate of 2°/min with 0.01% strain and 10 rad/s frequency.The viscoelastic properties of starch hydrogels were determined by using storage modulus (G′), loss modulus (G″), and loss tangent  as the dynamic mechanical parameters. Temperature sweeps were performed by preparing a starch suspension with a certain concentration  was first determined by conducting strain sweeps at a frequency of 10 rad/s. The oscillatory stress sweeps were then performed at the same frequency (10 rad/s) in order to measure yield stress (τy) and flow stress (τf). The stress value related to a 10% drop in G′ value was reported as the yield stress (Pa). Moreover, the crossover point of G′ and G″ was considered as the flow stress. The angular frequency sweep test was carried out by increasing the frequency from 0.1 to 100 rad/sat 95 °C. The average of three replications was reported for each rheological parameter.p ≤ 0.05), was performed using SPSS Statistics software. The average of three replications along with their standard deviations was reported for the measured parameters.The analysis of variance (ANOVA), followed by the LSD comparisons of means test  decreased with decreasing the nozzle diameter and increasing the starch concentration. Regarding the size and sphericity of the starch beads, the best results were obtained with a 15% starch concentration and nozzle 34G (The bead size of starch beads significantly (zzle 34G . As depip < 0.05). At the same starch concentration, nozzle size and printing pressure played important roles in achieving the desired bead size. Smaller nozzle diameters resulted in smaller bead sizes; however, higher printing pressure was required to create beads  at room temperature allowed self-supporting of the starch paste and, in turn, retained the spherical shape without further curing process . TherefoThe morphology of the 3D-printed starch beads is depicted in The granule remnants can be a particular concern when developing small porous spheres. It is important to fabricate homogeneous structures without any granule residues for future applications such as bioactive compound delivery. The heating of starch granules dispersed in an excess amount of water causes irreversible swelling of the granules and amylose dissolution. Complete dissolution of granules occurs only if high enough temperature and shear are used . As repow/w) (p < 0.05). This may provide advantages in protecting and delivering sensitive bioactive compounds. The dense surface of the beads may function as a coating layer leading to a controlled release as it covers the internal open-pore structure. Moreover, the release of the loaded compounds can vary based on the thickness of the surface. Previously, Schroeter et al. [The surfaces of the 3D-printed starch beads are also shown in r et al.  reported2 drying or lyophilization. They concluded that the main parameter greatly affecting the aerogel properties was the amylose/amylopectin ratio. Furthermore, it was reported that during retrogradation, semi-crystalline clusters are formed through the association of amylose into parallel double helices linked via amorphous regions [The starch beads prepared in this study showed porous structure similar to the results reported by Zou and Budtova , who pre regions . At the  regions ,24, whicIn this study, freeze-dried starch beads indicated an intricate network with a porous structure . This stFurthermore, the macropore size distribution, ranging between 0.5 and 5 µm, is presented in 2 drying, where beads with finer networks can be generated, granting them more potential in being applied in delivery systems. The effect of the drying method and starch type is investigated in the next step of this project.As mentioned above, the macroporous structure observed in this study was created through freeze-drying, which can be improved by using supercritical COThe density and porosity of the starch beads are represented in 3 (p < 0.05) (No shrinkage was observed during the preparation of starch beads. The high amylose content 72%) resulted in a faster retrogradation and higher gel strength, increasing the resistance to collapse during the drying process. The starch beads revealed low densities ranging between 0.14 and 0.23 g/cm3 a, simila2% result3 b. As exp < 0.05) ; this wa < 0.05) . A similThe chemical structure of starch beads was elucidated by an FTIR spectroscopy. The FTIR spectra demonstrated identical characteristic peaks; however, their intensities differed slightly .−1 was ascribed to the OH stretching mode. The width of this peak implies the hydrogen bonding content. For starch beads, SP-10, SP-12.5, and SP-15, OH stretching peak shifted slightly to a higher wavenumber , which can be related to the reduction in crystalline structure [−1 was attributed to the –CH2 stretching vibration. The C–H bending of CH2 revealed IR peaks at 1433 and 1409 cm−1. The other bands at 1150, 1105, and 990 cm−1 were assigned to C–O, C–C, and C–O–H stretching vibrations, respectively [−1 was associated with C–O–C at the β-glycosidic linkage [A strong absorption peak centered at 3282 cmtructure ,32. The  linkage .−1 revealed two shoulders at 1022 cm−1 and 1054 cm−1, which exhibited the amorphous and crystalline bands, respectively. The higher intensity of the shoulder appeared at 1022 cm−1, which suggests the greater extent of short-range double helices [−1 was also quantified to compare the crystallinity of the starch beads. This ratio decreased in starch beads compared to HACS, confirming the loss of crystalline structure during gelatinization. For HACS, SP-10, SP-12.5, and SP-15, the 1054/1022 cm−1 was calculated as 1.48, 1.23, 1.27, and 1.34, respectively.In addition, the starch crystallinity was determined by FTIR spectra by characterizing the changes in the amorphous and crystalline domains. As seen in  helices . The int−1 was attributed to the water tightly bound through hydrogen bonding as part of the crystalline structure [The single peak that appeared at1646 cmtructure . This peθ = 15.02°, 17.03°, 19.75°, 22.08°, and 23.90°, representing a B-type XRD pattern. The previous studies reported that high amylose starches typically show a B-type XRD pattern, which is identified by five XRD peaks at 2θ = 15.26°, 17.21°, 19.75°, 22.32°, and 24.08° [The diffraction profiles of HACS and 3D-printed starch beads are presented in d 24.08° .As expected, the intensity of the sharp peaks observed in the XRD pattern of HACS became significantly weaker and broader, indicating the decrease in crystallinity after the gelatinization for all three concentrations ,35. ThisTwo endothermic peaks were observed in the DSC thermogram of HACS, with the main peak appearing at 70 °C and the other smaller peak at 95 °C . AccordiAs depicted in Generally, starches have a lipid content between 0.5 and 1.5% ,41. TherThe rheological behavior of the bio-ink is critical for extrusion-based 3D printing. The desired properties, including shape stability and resolution, highly depend on the rheological properties of the printing material. Specifically, the ink should show shear-thinning behavior with appropriate flow stress to be printable using an extrusion-based 3D printer, especially with a small nozzle size ,43,44,45−1), and the power-law index, respectively. The power-law parameters are presented in The steady shear rheological parameters were measured to investigate the viscoelasticity of starch suspensions as the ink for extrusion-based 3D printing. At a constant shear rate, the viscosity increased as the starch concentration increased a. The doStorage modulus (G′) and tan δ are two parameters reflecting the viscoelastic characteristics of starch paste. The gelatinized starch obtained by increasing the temperature of a starch solution was able to respond to the elastic deformation and support the printed object ,19. The Higher starch concentrations resulted in higher G′ values . Based oThe highest starch concentration (15%) revealed the highest G′ d. StarchThe frequency sweeps of the storage (G′) and loss (G″) moduli at 95 °C are depicted in w/w) and nozzle size  were investigated and optimized for the smallest bead size. The smallest bead size (~650 µm) was achieved using a 15% starch concentration with a nozzle size of 0.08 mm. Moreover, the sphericity of the beads increased with increasing the starch concentration; as starch pastes’ viscosity and storage modulus increased with starch concentration, resulting in a better shape fidelity  at 15% starch concentration. However, higher starch concentration led to higher density (0.23 g/cm3) and lower porosity (86%). All the 3D-printed starch beads showed an open porous structure. However, a nonporous layer covering the 3D-printed beads was observed, and its thickness increased as the starch concentration increased, which may provide extra protection in encapsulation applications. The crystallinity of starch decreased mainly due to the gelatinization process involved prior to and during 3D printing. DSC thermograms indicated that the amylopectin crystalline structure was irreversibly melted due to gelatinization. Overall, the proposed 3D food printing approach provides precise control over the shape, size, and composition of the 3D-printed beads while eliminating the need for surfactants, organic solvents, and extra extraction steps to generate starch beads. The porous structure of the 3D-printed starch beads enables them to be used for the delivery of bioactive compounds/drugs in the food and pharmaceutical industries.In the present study, extrusion-based 3D food printing was used to fabricate spherical porous starch beads. Three-dimensional food printing parameters, namely starch concentration (10, 12.5, and 15%,


Background: The anatomic variation of hepatic vein in the left lateral segment (LLS) increases the risk of outflow complication in pediatric living liver transplantation (LDLT). Here, we share a modified method for dual hepatic vein reconstruction in pediatric LDLT using LLS with two wide orifices.Methods: From Sep 2018 to Dec 2019, 434 pediatric LDLTs using LLS were performed in our center. Hepatic veins of grafts were classified into three types with emphasis on the number, size, and location of orifices at the cut surface: a single opening ; two adjacent orifices ; two wide orifices with orifices distances <20 mm ; and two wide orifices with orifices distances >20 mm . Rv was defined as the ratio of diameter of V2 and V3 (refer to hepatic vein drained segments II and III). We developed a modified dual hepatic vein anastomosis to reconstruct outflow for type IIIb grafts with Rv ≤1. Briefly, the hepatic vein of segment II was anastomosed to the common stump of middle hepatic vein (MHV) and left hepatic vein (LHV), followed by unification of V3 and the longitudinal incision orifice in inferior venous cave (IVC).Results: During median follow-up of 15.6 months (7.5–22.9 months), no hepatic vein complications occurred.Conclusion: This novel modified dual hepatic vein anastomosis could serve as a feasible surgical option for type IIIb LLS grafts with Rv ≤1 in pediatric LDLT. Liver transplantation is the standard choice for children with end-stage liver diseases . Due to Four hundred fifty-three LDLT procedures were performed at the Department of Liver Surgery, Renji Hospital, School of Medicine, Shanghai Jiaotong University from September 2018 to December 2019. All LDLT procedures were approved by the ethics committee of Renji Hospital and were performed in accordance with the relevant regulations. A retrospective analysis of left hepatic vein (LHV) variations in LLS grafts was performed with emphasis on the number, size, and location of orifices at the cut surface. Computer tomography image data for 434 donors using LLS were analyzed. The anatomical variations of LHV were classified into three types. Very small hepatic vein branches (<5 mm in diameter) draining into the middle hepatic vein (MHV) or inferior vena cava (IVC) were not considered because they could be sacrificed without the risk of significant hepatic vein congestion. Eight type IIIb LLS grafts were considered fitter for modified dual hepatic vein anastomosis.in situ through the left portal vein, first with 4°C lactated Ringer's solution (200 ml), and then with 4°C UW solution .LLS graft harvest strategies were formulated according to computational simulation of virtual LLS graft with commercial three-dimensional reconstruction software . Graft/recipient's body weight ratio (GRWR) was calculated, and all 434 donors received left lateral segmentectomy. The hepatic parenchyma was transected using an ultrasonic aspirator  and bipolar electric cautery. The triangular and hepatogastric ligaments were dissected, and the hepatic veins were isolated. The left portal vein and left hepatic artery were transected and the sheath around the left bile duct was left undisturbed to maintain the blood flow of the biliary system. The isolated graft was then perfused Hepatic artery and portal vein were isolated at the hepatic hilum, then the liver was dissected from IVC by dissection of ligation and short hepatic veins. Total hepatectomy was performed after side clamping the IVC. The liver graft was implanted into the hepatic cavity by hepatic vein anastomosis. The vascular reconstruction was performed in end-to-end fashion in the portal vein with 7–0 polyglyconate (PDS). The hepatic artery anastomosis was accomplished with 8–0 or 9–0 polypropylene (Prolene) with the surgical microscope . Biliary reconstruction was performed with a Roux-en-Y limb or interposed jejunal conduit previously existing in patients treated with Kasai portoenterostomy.V ≤ 1 (V3 as the dominant hepatic vein). The distance between two orifices should smaller than the length of retrohepatic inferior vena cava. As shown in The modified dual hepatic vein anastomosis technique is a better choice for hepatic vein reconstruction in LLS grafts with two wide orifices (>20 mm) and RV2 is anastomosed end-to-end to the left of the common stump of MHV and LHV.An oval incision orifice is created close to the central axis of the IVC in recipients. The diameter of incision orifice should not be smaller than the size of V3. Whereas, V3 is anastomosed end-to-side to the longitudinal incision orifice in the anterior wall of IVC with 5–0 polyglyconate (PDS).Eversion suture was not suggested to expand outflow size. Care was taken to adjust the cut surface of liver graft slightly oblique upward to avoid liver regeneration-related hepatic vein occlusion.Doppler ultrasound (US) was used to estimate the outflow status at the completion of hepatic vein reconstruction and the completion of transplantation. We then performed Doppler US tests in all recipients daily in the 1st week after LT, every 2 days in the 2nd week, monthly during the first 6 months, and every 3 months thereafter. Oral administration of FK506 is initially at doses of 0.1–0.15 mg/kg every day. Trough levels of FK 506 were maintained in the range of 8~12 ng/ml for the 1st month and gradually decreased to 5 ng/ml after 1 year. Cyclosporine was initial at a dose of 4 mg/kg/day, and the target C0 and C2 levels were 150–200 and 800–1,200 ng/ml, respectively. CYP3A5 genotypes in both recipients and donors were performed to guide the usage of FK506 and cyclosporine . The antAll data analysis in this research was performed using R software. All results were expressed as mean ± standard error of the mean.n = 341, 78.57%); (II) two close orifices ; (III) two wide-spaced orifices . According to the distance between two separated openings, type III can be classified as: (a) orifices distances <20 mm  and (b) orifice distances >20 mm . RV was defined as the ratio of diameter of V2 and V3 which can be used to determine the dominant hepatic vein in LLS. For type IIIb LLS grafts with RV ≤ 1, modified dual hepatic vein anastomosis is preferable to ensure satisfactory hepatic venous drainage.The LHV anatomy of 434 LLS graft donors was classified on the basis of the patterns of the graft hepatic veins. The mean age of the donors was 31.42 years (range = 18–58 years), and 210 (48.39%) were male. The patterns of the LLS graft hepatic veins were classified into types according to the number, size, and location of orifices at the cut surface : (I) a sA consecutive series of recipients using modified dual hepatic vein anastomosis included four boys and fouor girls with a median age of 22.3 months  and a median weight of 10.2 kg . Four of the children suffered from biliary astresia (BA), and the other four children's primary diseases were acute hepatic failure, hyperhomocysteinemia, tyrosinemia, and ornithine transcarboxylase deficiency (OTCD), respectively. The profile of donors are summarized in V was 0.85 (0.77–0.97). All of the grafts showed good venous outflow, and the Doppler US results at the completion of transplantation are summarized in For the eight LDLTs using modified dual hepatic anastomosis, the grafts were all left lateral segments and the median GRWR was 2.5% (1.8–5.0%). The median time for hepatic vein reconstruction was 22.88 min (17–31 min). The median hepatic phase was 47.38 min (44–54 min). The median distance between two orifices was 25.30 mm (22.82–29.57 mm). The median diameter of V2 was 6.55 mm (5.23–7.33 mm), and the median diameter of V3 was 7.72 mm (6.63–8.69 mm). The median RFor the eight patients using modified dual hepatic anastomosis, all of them have satisfactory venous flow after LDLT, and no HV-related complications were observed in the subsequent follow-up. Graft function recovered within 2 weeks. The median period for drainage tube indwelling was 15 days (7–37), and the details of abdominal drainage are summarized in Adequate venous outflow has been recognized to be a vital element for the success of liver transplantation . HepaticIn this research, LLS graft donors were classified into three types according to the hepatic vein orifice variations at the cut surface , 19, 20.V ≤ 1). Therefore, we introduced modified dual hepatic vein anastomosis technique for type IIIb variant LLS graft with RV ≤ 1.Venoplasty has been applied to form a single opening to facilitate the vein anastomosis in type IIIa (orifices distance <20 mm) , 24. ForDual hepatic vein anastomosis was adopted for grafts with two wide orifices which cannot form a common anastomotic stump. Two independent anastomosis can provide a stable dual-axis structure to prevent twisting or kinging. Reconstruction of V2 and V3, respectively, also prevent the whole LLS venous obstruction from interposition conduit congestion. However, traditional dual hepatic vein anastomosis were made between the graft HVs and the recipient MHV and LHV or RHV and the common stump of MHV/LHV . This anV ≤ 1.In the LDLTs performed from September 2018 to December 2019, we identified 27 LLS grafts with two wide separate hepatic orifices at the cut surface. A common of eight children used modified dual hepatic vein anastomosis to reconstruct venous outflow. During the maximal follow-up period of 50 months, no HV complications occurred and all the eight children recovered smoothly. Although longer follow-up and larger sample size are required, the encouraging results in this research confirm the efficacy and safety of the modified dual hepatic vein anastomosis for type IIIb grafts with RV ≤ 1 to reconstruct venous outflow. For type IIIb LLS grafts with RV > 1, conduit is also available considering that small V3 conduit occlusion has a limited effect on graft survival.In conclusion, our modified dual hepatic vein anastomosis technique provides a concise method for type IIIb grafts with RThe raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.Written informed consent was obtained from the individual(s), and minor(s)' legal guardian/next of kin, for the publication of any potentially identifiable images or data included in this article.QX and PW designed and performed the surgery. YH collected the data and wrote the paper. MF and TZ participated in arterial anastomosis. BQ and JZhu analyzed the data. YL and JZha participated in donor operations. All authors contributed to the article and approved the submitted version.This study was supported by Shanghai Sailing Program (18YF1412700 [PW]), the Project of Medical Key Specialty of Shanghai Municipality (shslczdzk05801 [QX]), and the National Key Research and Development Program of China (2017YFC0908100 [QX]).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.


Free gingival graft (FGG) is a highly predictable method to increase the width of keratinized gingiva. Various materials have been reported to accelerate the wound healing process. Considering the positive effect of EPO on dermal wound healing this study aimed to investigate the effects of EPO on the rate of healing and degree of inflammation in free gingival grafts.Seventeen patients with bilateral lack of keratinized gingiva in mandible were selected for this clinical trial. The surgical intervention was performed after phase I periodontal therapy.Recipient site was prepared apical to the mucogingival line, and FGG was harvested from the palate. Before graft placement, the test side and control side were treated with 1 ml of EPO 4000 IU/ml and distilled water, respectively, for 2 min. On days 7, 14, 21, 28, 60, and 90 after surgery, the grafted sites were examined by blinded observers to compare the healing and inflammation of the areas.2 and 71.35 ± 15.62 mm2 in the EPO and control groups, respectively. The difference was not significant, though.All the 17 patients completed the surgeries and follow-up examinations. Direct examination revealed significantly better healing in EPO group only on the 28th day. Assessment of the photographs showed a significant value in favor of the test group at some other time points as well. The EPO group demonstrated less inflammation, which was statistically significant in many time points. The graft area was 80.88 ± 30.21 mmTopical application of erythropoietin can accelerate the healing of gingival grafts and reduce the inflammation during healing period. The final graft outcome, nevertheless, does not seem to be influenced by EPO.Trial registration This was a split-mouth randomized controlled clinical trial (IRCT201201278830N1). The first registration date: 2016-10-22 Free gingival graft (FGG) is a relatively common procedure among periodontal surgeries. FGG is usually performed to gain wider keratinized gingiva , increasAccording to the histological study by Oliver et al. , the heaTraditionally, various materials with mechanical protection, hemostatic and bacteriostatic properties are used to protect the graft in the early stages of graft healing . EugenolCurro et al.  also revErythropoietin (EPO) is a glycoprotein (molecular weight: 30.4 kDa) essential for hematopoiesis and a member of Class I Cytokines . Apart fThe basal cells of the oral mucosa also express receptors for erythropoietin . Thus, iIn the present study, the effects of topical application of EPO on clinical healing of FGG recipient site have been investigated.This was a split-mouth randomized controlled clinical trial (IRCT201201278830N1). The study protocol was in accordance with the Helsinki Declaration of 1975, revised in 2013. The study was approved by the ethical committee of Tehran University of Medical Sciences (119225).All study cases were selected from the patients who were referred to the periodontics department, Tehran University of Medical Sciences. The inclusion criteria were: (1) age of 18 years old or above; (2) periodontal health; and (3) need for gingival graft in order to augment keratinized gingiva in at least two mandibular areas. If any of the following criteria were present, the patient would be excluded: (1) compromised systemic health, such as diabetes mellitus and immune suppression; (2) consumption of anti-inflammatory or antibiotics during last 3 months; (3) smoking, and (4) O’Leary plaque index >15% after phase I periodontal therapy. Phase I periodontal treatment, including oral hygiene instructions, scaling, root planing and prophylaxis, if necessary, was performed for the patients meeting the inclusion criteria. Subjects compliant with oral hygiene measures who reached the O’Leary plaque index of less than 15% in the re-evaluation session were selected for the study. Informed consent was obtained from the patients who agreed to participate in the study.To determine the sample size, using tab analysis of PASS 11, α was set as 0.05 and β was set as 0.2. The clinically relevant difference between the test and control groups was considered, and according to the standard deviation from the study by Yaghobee et al., the sample size was calculated as 17 in each group.Balanced block randomization was used to allocate each patient into one of the following categories:Right side surgery first, treated with distilled water.Right side surgery first, treated with EPO.Left side surgery first, treated with distilled water.Left side surgery first, treated with EPO..The patient, the surgeon, the first and last examiner, and the data analyzer were blinded to the intervention.The classic technique was used for free gingival graft . SurgicaAnalgesics  and antibiotics  were prescribed for all patients. Routine post-surgical instructions were given to patients in a pamphlet. The surgical dressing was removed 7 days after surgery, and the sutures were removed at 14th day. Professional tooth cleaning and prophylaxis were provided for patients every 2 weeks for 12 weeks.To evaluate the healing and the inflammation of the surgical site, direct (by one observer) and indirect (by 3 observers through photographs) examinations were performed. For direct observation patients were visited 7, 14, 21, 28, 60, and 90 days after surgery. Each day, one examiner (N. R.), blinded to the test and control sides, compared both surgical sites in each patient in terms of healing status and inflammation severity by direct observation. A standard clinical photograph was also taken from each site for indirect observation by 3 other observers. The photographs were always taken by one camera (Canon SX210 IS) in a specific dental chair at about 10 AM each day, with the chair light off, and at the distance of 30 cm from the surgical site. To compare the test and control sides, two photos were placed side by side in one slide using PowerPoint . The magnification values of the photographs were similar. Observers of the photographs were blind to the allocated intervention.Degree of healing was assessed based on the graft color match, contour, and texture. The inflammation status was evaluated according to the size of the red inflammatory halo around the graft. Direct and indirect examiners decided which side had better conditions, if different at all. At the last visit, the graft dimensions (width and length) were also measured in millimeters through a periodontal probe .All statistical analysis was performed using SPSS software (IBM SPSS Statistics 20.0). The mean graft dimensions in two groups were compared using Student t-test. Sign test was used to statistically compare the healing and inflammation results in the test and control groups. A p-value of smaller than 0.05 was considered significant difference between the groups.Seventeen patients 5 male, 12 Female) with the mean age of 45 ± 6 years completed the study. Figure 2 Female No adverse effect was seen related to the surgery. The clinical photographs of one patient are shown in Fig. The results of the evaluation of healing by direct examination and observing the photographs are presented in Table Table 2 in the EPO side, and 71.35 ± 15.62 mm2 in the control side .The mean graft width in the last evaluation was 6.47 ± 1.37 mm and 6.0 ± 1.27 mm in the study and control groups, respectively . The mean graft size was 80.88 ± 30.21 mmThis study aimed to evaluate the effects of the topical application of erythropoietin on the outcome of free gingival graft performed to increase the keratinized tissue width. The results of this study showed that EPO could accelerate the rate of healing and reduce the degree of inflammation in free gingival grafts. The areas treated with EPO gel generally healed faster and experienced less inflammation compared to the control sites. Similar results have been reported in terms of improved wound healing and reduced inflammation using topical or systemic EPO in previous articles. In an animal study by Hamed et al. , topicalIn a clinical study, Yaghobee et al.  evaluateBader et al.  also repThe positive effects of EPO on wound healing is likely to be mediated by improved angiogenesis , 27, 32,Considering the results of this study on days 60 and 90, no significant difference in terms of tissue color match, contour, and texture between the test and control groups was found. This means that erythropoietin does not affect the final outcome of graft healing, but simply causes the graft to go through the healing process slightly faster. This, however, equates less pain and discomfort for the patient and sooner achievement of the desired result.Based on direct and/or indirect examinations, the inflammation halo was significantly less evident around the test side compared to the control side at all timepoints, except for the 7th day, during which no significant difference was found between the groups. Although some degrees of inflammation are essential for the wound repair, excessive inflammatory response will delay healing . AccordiOther growth factors, individually or in comparison to erythropoietin or in addition to it have been used in some studies. Curro et al.  pioneereSorg et al.  studied Overall, the results of multiple studies including the present study, indicates that EPO has a considerable effect in wound healing. However, the reported effect size has been different which may have been influenced by the drug administration route and the circumstances of the area to be treated. For example, the presence of saliva and tongue movements in the oral cavity causes the drug to be washed away in a short time, and as a result the drug effects are limited compared to dermal applications. The dose and frequency of application in oral cavity, therefore, has to be specified to the region to gain optimum results of healing in gingival/mucosal wounds.Due to the limited human trials regarding the effects of topical EPO, further research is still needed on different types of wounds , with various administration routes, doses and frequencies. Also future research may focus on the synergistic effects of EPO with other growth factors needs to be investigated as well.Erythropoietin may accelerate healing and reduce inflammation in free gingival graft recipient sites, which could be related to improved angiogenesis and epithelialization mediated by this factor. However, it does not affect the final graft outcome regarding the size, contour, texture, and color match in short term. More investigation is needed to find the optimum dose and frequency of EPO application.


Also, the redox balance during dormancy appeared to be maintained mainly by catalase and, to a lesser extent, by glutathione (GSH). Overall, the present study concludes that differences in ROS and the bud redox balance could, at least partially, explain the differences in dormancy duration and bloom date among apple cultivars.Understanding the biochemical mechanisms underlying bud dormancy and bloom time regulation in deciduous woody perennials is critical for devising effective strategies to protect these species from spring frost damage. This study investigated the accumulation profiles of carbohydrates, ROS and antioxidants during dormancy in ‘Cripps Pink’ and ‘Honeycrisp’, two apple cultivars representing the early and late bloom cultivars, respectively. Our data showed that starch levels generally declined during dormancy, whereas soluble sugars increased. However, the present study did not record significant alternations in the carbohydrate accumulation profiles between the two cultivars that could account for the differences in their bloom dates. On the other hand, H Temperate woody perennials annually cycle between growth cessation and growth resumption phases in synchrony with seasonal changes in temperature and photoperiod. Bud dormancy is critical for the tree to survive under potential freezing and subfreezing temperatures in winter. Bud dormancy can be triggered either by low temperatures and short days, or by low temperatures only, such as in apples and pears . The dorDormancy release and bud burst involve numerous physiological and biochemical changes that include, but are not limited to, changes in hormone hemostasis, lipid mobilization, carbohydrate interconversion and transport, and redox processes ,8,9,10. 2O2) and superoxide (O2•−), which are key components in the redox signaling pathways when present below the lethal levels [2O2 and O2•− in the buds [2O2 content of flower buds with the onset of endodormancy release, whereas insufficient chilling exposure fails to change H2O2 content [2O2 leading to dormancy release in grapes [2•− is generated by plasma membrane-based NADPH oxidase that transfers electrons from cytoplasmic NADPH to oxygen. NADPH oxidase has been reported to promote dormancy release in Arabidopsis seeds and potato tubers [2O2. These genes include superoxide dismutase, catalase, glutathione-s-transferase, glutathione reductase, ascorbate peroxidase as well as redox-related genes [The exposure of plants to stressful conditions promotes the production of ROS, such as hydrogen peroxide  catabolism and gibberellin (GA) biosynthesis [2O2 production, decreases endogenous ABA levels in grape [2O2, O2•− and hydroxyl (•OH) radicals can play central roles in signal transduction pathways by changing the oxidation and reduction of proteins cysteinyl thiols, and subsequently altering protein functions and characteristics [ROS’s role in regulating dormancy and bud burst is also tightly linked to the relative abundance of carbohydrates and phytohormones . For insynthesis . Similarynthesis . Along tin grape ,46 and sin grape  by promoin grape . Indeed,eristics . These sMalus × domestica). To this end, we used two apple cultivars with contrasting bloom dates, ‘Cripps Pink’ and ‘Honeycrisp’, and collected flower buds from these cultivars at different times of endodormancy and ecodormancy to quantify the levels of sugars and ROS related compounds and enzymes. These two cultivars were previously used by Sapkota et al., 2021, to monitor the levels of major plant hormones  throughout the dormancy cycle. Given the anticipated interactions among these plant hormones, ROS and carbohydrate metabolism, the present investigation can deepen our understanding of the regulation of dormancy and flowering in apple and lay the foundation for effective frost mitigation strategies.The aim of this study was to investigate the accumulation patterns of sugars, ROS and antioxidants in relation to bud dormancy progression and flowering time in apple  was used to record the field temperatures at 10-min intervals A. CH werg) for 10 min. The supernatant was collected and this extraction step was repeated twice. Soluble sugars  were determined using commercial enzyme assays . To determinate the starch levels, the pellet was dried overnight to remove ethanol completely. The dry pellet was suspended in 500 µL of deionized water and the solution was incubated at 80 °C for 20 min. The solution was then cooled and mixed with 400 µL of 200 mM acetate buffer (pH 5.1). A 100 µL of 2 U of amyl glucosidase and 40 U of α-amylase was added to the solution, followed by incubation at 50 °C for 24 h and centrifugation at 14,000 rpm  for 10 min. The glucose content in the supernatant was measured and starch content was estimated as glucose times 0.9.Carbohydrate analysis was performed according to Edwards et al. (2011) . Briefly2O2) and superoxide (O2•−) were prepared according to Lei et al. (2006). In brief, 100 mg ground sample was mixed with 1.0 mL of 50 mM K-PO4 (pH 7) and incubated on ice for 10 min. The solution was then centrifuged at 12,000 rpm  at 4 °C for 10 min. The supernatant was used for H2O2 and O2•− content evaluation.The samples for hydrogen peroxide  was mixed with 200 µL of 0.1% TiCl4 in 20% H2SO4 (v/v) and incubated for 5 min. The solution was centrifuged at 10,000 rpm  for 5 min at room temperature and the absorbance was recorded at 410 nm using a microplate reader . The H2O2 content in the tissue was calculated using the extinction coefficient (e = 0.28 µ mol−1 cm−1).For H2•− analysis, an aliquot of supernatant (100 µL) was mixed with 25 µL of 10 mM NH2OH-HCl and incubated at 25 °C for 1 h. After incubation, 50 µL of 7 mM sulfanilamide and 50 µL of 7 mM α-naphthylamine were added to the solution and incubated at 25 °C for 20 min. The absorbance was recorded at 530 nm spectrophotometrically. The O2•− content was calculated based on a standard curve prepared by serial dilutions of NaNO2 (2–250 ppm).For Og) at 4 °C for 10 min. The content of reduced glutathione (GSH) and glutathione disulfide (GSSG) was determined using a GSH: GSSG kit following the manufacturer’s protocol . The ratio of GSH: GSSG was calculated by simple division of the concentrations of GSH and GSSG.For the extraction of glutathione, 500 mg of ground bud tissue was mixed with 0.6 mL 5% sulfosalicylic acid, and centrifuged at 12,000 rpm , superoxide dismutase(SOD), glutathione peroxidase (GPX), and glutathione reductase (GR) enzymes were determined spectrophotometrically using the colorimetric kits  and following the manufacturer’s instructions.100 mg ground bud tissue was extracted with 1 mL of cold potassium phosphate buffer . After centrifugation at 4 °C, the supernatant was directly used for protein content measurement and enzyme activity assays. Total soluble proteins were quantified according to the Bradford assay  and usint-test was used to compare the means between the cultivars at each sampling time point. All values were expressed as mean ± SEM. Probabilities of p < 0.05 were considered statistically significant. Person’s correlation table was developed using metaboanalyst version 5.0  at p < 0.05. The data of plant hormones  to build the correlation matrix was retrieved from our already published article [ROS, sugar quantification and enzymatic activities data were analyzed using R statistical programming language (version 3.6.3). Student’s  article .The phenological events in this study were recorded according to CH and GDH calculated for ‘Honeycrisp’ and ‘Cripps Pink’ during the 2019 and 2020 dormancy-regrowth cycle as detailed in our previous investigation . In brie−1, approximately 2.3 mg g−1 higher than ‘Honeycrisp’. The starch content in ‘Cripps Pink’ showed two remarkable rises at 600 and 1000 CH, whereas, ‘Honeycrisp’ showed only one rise at the time of endodormancy release (1000 CH). The starch content of the two cultivars both reached 0.5 mg g−1 at their respective budburst time of 3000 and 4000 GDH. The total soluble sugars (TSS) in ‘Cripps Pink’ showed a remarkable peak in the middle of endodormancy, while the TSS in ‘Honeycrisp’ remained relatively stable throughout both endodormancy and ecodormancy . In contrast, the TSS in ‘Honeycrisp’ only showed some small changes during the dormancy, reaching a lowest point at 600 CH, followed by a slight rise at 800 CH, which was approximately half of ‘Cripps Pink’ at the respective time point.The starch content in the floral buds declined in both cultivars over the entire dormancy period, with some noticeable fluctuations A. At thedormancy B. The TSTo gain insights into the relative contributions of sugar forms to the TSS dynamics, we examined the concentrations of sucrose, glucose and fructose separately. In ‘Honeycrisp’, similar dynamic patterns were found in the three sugar forms, with glucose and fructose both peaking at 800 CH, corresponding with the peak of TSS C–E. In ‘2O2) content was high and started to decrease with the progression of endodormancy in both cultivars (2O2 level reached its peak and was 3 times higher in ‘Cripps Pink’ than ‘Honeycrisp’. The H2O2 level started to decrease with the progression of ecodormancy and reached its lowest point (~3 nmol g−1) at budburst in both cultivars. However, the decrease in H2O2 during ecodormancy started early (1000 CH) in ‘Cripps Pink’ compared to 2000 GDH for ‘Honeycrisp’.At endodormancy induction, hydrogen peroxide (Hultivars A. Howeve2•−) in both cultivars showed a general increase from the beginning of endodormancy to budburst , followed by a dramatic increase at 1000 CH and a dramatic decrease at 1000 GDH. The levels of O2•− increased again with the progression of ecodormancy reaching its peak of 40 µg g−1 at budburst.The level of superoxide  activities were highly similar between the two cultivars, especially during the ecodormancy period B. The in−1 mg−1 for ‘Honeycrisp’ and ‘Cripps Pink’, and both increased rapidly to 20.0 µmol min−1 mg−1 at 400 CH, plateauing until 600 CH, at which both declined sharply to 2.0 µmol min−1 mg−1 at 800 CH. After remaining at the low level until 1000 GDH, the GPx activities of ‘Cripps Pink’ (12 µmol min−1 mg−1) and ‘Honeycrisp’ (22 µmol min−1 mg−1) both reached a peak at 2000 GDH, followed by a rapid decrease to 2.0 µmol min−1 mg−1, which remained until budburst. It is noticeable that the GPx activity in ‘Cripps Pink’ was higher than in ‘Honeycrisp’ at 1000 CH, corresponding to the time of endodormancy release.Dramatic fluctuations of the glutathione peroxidase (GPx) activities were observed in both cultivars throughout the dormancy period C. The GP−1 mg−1 at 200 CH to 0.03 nmol min−1 mg−1 at 600 CH, while ‘Honeycrisp’ increasing from 0.05 to 0.07 nmol min−1 mg−1 during the same period. After reaching 0.3 nmol min−1 mg−1 at 800 CH, the GR activity in ‘Cripps Pink’ peaked (1.3 nmol min−1 mg−1) at 1000 CH, 3 times higher than ‘Honeycrisp’, then declined to 0.5 nmol min−1 mg−1 at 1000 GDH, remaining at this level until budburst. In contrast, the GR activity in ‘Honeycrisp’ increased steadily since 800 CH until 3000 GDH, and declined slightly at budburst (4000 GDH).The glutathione reductase (GR) activities showed different trends in the two cultivars during endodormancy and ecodormancy D. At ear2O2 and ABA. ABA positively correlated with starch, but negatively correlated with the other molecules in both cultivars , starch , NADPH oxidase , superoxide , and GSSG  in ‘Honeycrisp’ and ‘Cripps Pink’ respectively but no significant correlation was observed between Hultivars .Understanding the physiological mechanisms underlying apple bud dormancy is critical for developing strategies that can be used to modulate bloom time and thus avoid spring frost damage. In the present study, we used two apple cultivars, ‘Cripps Pink’ and ‘Honeycrisp’ with contrasting bloom dates to further our understanding of the biochemical pathways associated with bud dormancy and bloom time regulation in apple, aiming at providing a framework for devising agronomic strategies for modulating bud burst and bloom time.Carbohydrates and their accumulation profiles in floral buds are tightly linked to the status of dormancy. Particularly, soluble sugars have been reported to act as signaling molecules  or physi2O2 has been suggested as a key signaling molecule associated with dormancy release. The characteristics of H2O2 that make it a potential molecule marker for chilling accumulation and dormancy status include membrane permeability, relatively long-life span, and oxidization of the critical thiol groups in some redox-sensitive proteins [2O2 is implicated in the control of plasmodesmata conductivity [2O2 level in ‘Cripps Pink’ peaked markedly when chilling requirement was fulfilled, confirming its role as an inducer of endodormancy release. However, such peak of H2O2 was not as prominent in ‘Honeycrisp’, which maintained at relatively high level until 2 weeks before bloom. Such divergent H2O2 profile during ecodormancy may, at least partially, explains the differences in bloom dates between the two cultivars. Indeed, in grapevine floral buds, maintaining a high level of H2O2, by inhibiting H2O2 degradation, leads to delayed budbreak [2O2 from the buds through antioxidative mechanisms is required for growth resumption [2O2 differentially regulate the release of endodormancy and ecodormancy warrants further investigation.Accumulating evidence has suggested that the bud redox status resulting from the balance between generation and scavenging of ROS plays a critical role in bud dormancy transition . As an iproteins , which cuctivity , a membructivity . In thisbudbreak . Togethesumption . NeverthSuperoxide is generated ubiquitously in cells during aerobic metabolism. High levels of superoxide have been shown to enhance energy production and amino acid metabolism, which are both required for growth resumption ,67. Supe2O2 scavenging systems require glutathione (GSH) that exists in the cells of dormant apple buds [2O2 detoxification through glutathione cycle requires NADPH that is generated via the pentose phosphate pathway (PPP). In this metabolic system, GSH participates as a reducing agent catalyzed by glutathione peroxidase (GPx), which results in reduction of H2O2 and formation of GSSG. The GSSG can then be reduced to GSH by glutathione reductase (GR) at the expense of NADPH [2O2 can also be detoxicated by CAT. Interestingly, CAT activity was found to be inhibited by the treatment of HC, leading to the accumulation of H2O2. In this study, the profiles of H2O2 levels in both cultivars bear high resemblance to the CAT activity, and to a lesser extent to the GSH activity, especially during the ecodormancy, suggesting that H2O2 may induce an immediate detoxication response, mediated primarily by CAT. In addition, this response appeared to be transient, rather than retained, as the CAT activity in ‘Honeycrisp’ decreased about 2 weeks (between 1000 and 2000 CH) earlier than the H2O2 levels during ecodormancy. In contrast to CAT, the profile of GPX activities was in opposite to H2O2 levels during both endodormancy and ecodormancy in both cultivars, indicating that GPX may operate in an antagonistic way with CAT in regulating H2O2.Antioxidants, both in non-enzymatic and enzymatic forms play a key role in cellular redox homeostasis. One of the important non-enzymatic antioxidants in plants is glutathione. Metabolic activities of Hple buds . H2O2 deof NADPH ,70. In a2 and H2O2. In apple buds, the SOD activity has been reported to rise toward the onset of budbreak [Superoxide dismutase (SOD) quenches ROS by catalyzing the dismutation of superoxide radicals to Obudbreak ,71. In tBetula pubescens) showed that the starch content in the buds declines in concomitance with ABA during ecodormancy [Dormancy is a complex process that involves several interactions among key metabolic components including, but not limited to ROS, carbohydrate, and hormones. In our previous study, we showed that plant hormones abscisic acid (ABA) and cytokinin (CK) are critical regulators of bud dormancy and bloom in apple . Comparidormancy . The clodormancy . In this2O2 during the early stage of endodormancy corresponds to the increase of ABA in both cultivars [2O2 reduces ABA content by enhancing ABA-8′-hydroxylase, an ABA catabolic enzyme [2O2 accumulation, protecting cells against oxidative damage [ROS and ABA are both essential mediators of stress responses, and are also critical regulators of bud dormancy. In this study, the gradual decline of Hultivars . Additioultivars  and enhaultivars . In addic enzyme . Similare damage . Howevere damage , which ie damage .It is well established that CK is involved in the initial steps triggering growth resumption in floral buds . Our pre2O2 accumulation patterns closely correlated with the dormancy stage and bloom time in both cultivars, peaking early in the early-blooming cultivar while remaining at high levels for a longer time in the late-blooming cultivar. The findings of the present study match almost precisely similar results in grape and Japanese pear where the peak of H2O2 during dormancy marks the achievement of chilling requirements or the transition to another phase; reinforcing a notion that H2O2 either as a signaling molecule and/or a metabolite could serve as a molecular marker for the intensity of dormancy in deciduous woody perennials. In the present study, we also found that redox balance during dormancy was achieved mainly by catalase and, to a lesser extent, by GSH. The differential levels of superoxide in both apple cultivars, especially during ecodormancy, add another line of evidence that bloom time regulation in apple could, at least partially, be modulated by disrupting the bud redox balance. This study also revealed that the carbohydrate profiles between the two apple cultivars- though differing sporadically- may not sufficiently account for the differences in their bloom dates. However, it should also be noted that other forms of carbohydrates, such as sorbitol, were not studied in the present study, which warrants further investigation. Overall, the present study emphasizes that ROS and the redox balance in apple buds may serve as mediators of dormancy and bloom phenology among apple cultivars.The present study showed that differences between ‘Honeycrisp’ and ‘Cripps Pink’ apple cultivars in bloom time were associated with changes in the ROS and, to a lesser extent, carbohydrate accumulation profiles during the dormancy period. The H


The ever-increasing competitiveness in the academic publishing market incentivizes journal editors to pursue higher impact factors. This translates into journals becoming more selective, and, ultimately, into higher publication standards. However, the fixation on higher impact factors leads some journals to artificially boost impact factors through the coordinated effort of a “citation cartel” of journals. “Citation cartel” behavior has become increasingly common in recent years, with several instances being reported. Here, we propose an algorithm—named CIDRE—to detect anomalous groups of journals that exchange citations at excessively high rates when compared against a null model that accounts for scientific communities and journal size. CIDRE detects more than half of the journals suspended from Journal Citation Reports due to anomalous citation behavior in the year of suspension or in advance. Furthermore, CIDRE detects many new anomalous groups, where the impact factors of the member journals are lifted substantially higher by the citations from other member journals. We describe a number of such examples in detail and discuss the implications of our findings with regard to the current academic climate. Measuring academic impact is a controversial and challenging task2. Yet, the evaluation of research has increasingly been operationalized in terms of the citations received by research papers and citation-based bibliometric indicators, such as the h-index and the journal impact factor (JIF), which are widely used to evaluate individual researchers, academic institutions, and the research output of entire nations6.The volume of published research is growing at exponential rates7 and other bibliometric indicators. Such indicators are widely recognized as proxies for a journal’s quality and prestige8, and have a considerable impact on the journal’s readership numbers and subscription base. This fact, in turn, incentivizes editors to devise strategies aimed at increasing citation numbers. Such strategies may ultimately result in publications of a higher quality. However, there have been multiple reports of malicious practices merely aimed at boosting citation numbers.Editors and academic publishers are under increasing pressure to ensure that their journals achieve and sustain high values of JIF9 or by writing editorial reviews10. Such self-citations are relatively easy to spot because they involve only one journal. Concerns have grown for less detectable forms of manipulation which involve the coordinated effort of a number of journals, a practice known as citation cartels. Such a practice—also referred to as citation stacking—consists of groups of journals exchanging citations at excessively high rates12. For example, one instance of a citation cartel attracted attention in 2011. In this example, two papers published in different journals provided a number of citations to a single journal, increasing its JIF by 25%13. Since then, new instances of citation stacking have been reported every year16.Editors of some journals have generated citations for their journals by coercing the authors of submitted papers17. Such citation cartels are easy to launch and hard to detect. To tackle this issue, Journal Citation Reports (JCR), which is owned by Clarivate Analytics and was owned by Thomson Reuters until 2016, has excluded from its annual journal ranking some pairs of journals when at least one of the two journals cited the other excessively and the journal pair satisfied some additional criteria18. As of 2019, JCR has suspended from its annual journal ranking 46 pairs of journals—featuring 55 journals in total—due to excessive pairwise citations18. Alternatively, a previous study regarded citation cartels as groups of densely interconnected nodes  in journal citation networks12. However, the approach based on network communities may suffer from false positives because communities are the norm rather than the exception in journal citation networks: journals tend to cite other journals in the same research field, which forms densely connected communities21.Journal editors may set up citation cartels by informally agreeing with other journal editors and colleagues to coerce citationsi) proximity (in terms of research areas) and (ii) size . Then, CIDRE finds groups of journals with excessive within-group citations relative to the null model. To the best of our knowledge, no empirically validated tool has been proposed for identifying groups of journals whose citation practices can be regarded—with statistical confidence—as anomalous.Detecting anomalous citation groups is inherently challenging because the abundance and prominence of citation communities may overshadow the anomalous citation patterns. To address this challenge, we propose the CItation Donors and REcipients (CIDRE) algorithm. The key idea of CIDRE is to discount the amount of citations between communities using a null model of networks with communities. The null model accounts for the citation rates that can be expected under healthy citation practices due to journals’  released on January 30th, 2020 to construct citation networks of journals15.In citation networks, a citation cartel is manifested as a group of journals that excessively cite papers published in other journals within the group. Although not all such groups are necessarily citation cartels, we aim to identify journal groups with excessive within-group citations. Specifically, we assume that an anomalous citation group is composed of donor journals and recipient journals. A donor journal provides excessive citations to papers published in recipient journals in the previous 2 years i.e., the time window for the JIF. In cases where two journals exchange citations at excessively high rates, they simultaneously behave as both donors and recipients. Although donor journals have no apparent direct benefit in providing citations to recipient journals, we consider them as a member of the anomalous citation group because some previously identified instances contain journals giving excessive citations to particular journals, which often share the publishers or editors25 as the null model. The dcSBM generates randomized networks that preserve the number of citations between groups of journals , and the outgoing and incoming citations of each journal on average. We determine the blocks by fitting the dcSBM using a non-parametric Bayesian method25. Community detection methods for networks including the dcSBM have been shown to provide a reasonable partitioning of journal citation networks into research fields21. Therefore, the networks generated by the dcSBM are considered to be random networks that roughly preserve the patterns of citations within and across research fields.We identify excessive citations between journals using a null model for citation networks. Specifically, we use the degree-corrected stochastic block model (dcSBM)U, that contains journal i. Journal i’s donor score, denoted by i provides to the other journals in U. Journal i’s recipient score, denoted by i receives from other journals in U. CIDRE considers a journal as a donor journal and a recipient journal if CIDRE removes from the given network all the edges that are statistically compatible with the null model and then computes a donor score and a recipient score for all journals based on the residual edges in the network . In the following, we refer to the weights of such edges as excessive citations. Consider a journal group, denoted by U to be the set of all nodes in the network. Then, CIDRE removes from U the journals that are neither a donor nor a recipient and recomputes the donor and recipient scores for the journals remaining in U. CIDRE iterates the removal of nodes and the recomputation of scores until no journal is further removed. We partition U into disjoint groups U and the residual edges. We regard each weakly connected component To find anomalous citation groups, CIDRE initializes CIDRE detected 184 citation groups between years 2010 and 2019  come from a single paper, (B) go to a single paper, (C) come from a single author, or (D) go to a single author, or (E) two journals in the group share at least one editorial board member  were made to the papers published in the JIF time window. Removing the citations from the two review papers decreases the JIFs for the 4 recipient journals, Anthropological Quarterly, Cultural Anthropology, Focaal, and Journal of the Royal Anthropological Institute, by more than 26%.An instance of category A is group 1, which CIDRE detected in the network in 2018 and is composed of 17 journals on anthropology Fig. a. Two reActa Crystallographica Section C  and Entrepreneurship and Sustainability Issues (ESI), which account for at least 25% of the editorial board members in each of the two journals. The editor-in-chief of ESI, who also serves JSSI as an editor, provided and received the largest number of citations  among the authors of papers published in this journal group.An instance of category E is group 5, which CIDRE detected in the network in 2016 and is composed of seven journals on business Fig. e. There Laser Physics, provided 1984 citations to the recipient journal, Laser Physics Letters. We did not find any concentration of citations; neither a single paper nor a single author provided or received more than 8% of citations within the group. In 2011, the number of citations from the donor journal to the recipient journal increased more than double, from 987 citations in 2010 to 1984 citations in 2011. CIDRE identified the increase in the citations to be excessive and detected this group.An instance of category F is group 6, which CIDRE detected in the network in 2011 and is composed of two journals on laser science Fig. f. The doInternational Journal of Automotive and Mechanical Engineering, provided 81 citations to a recipient journal, Journal of Mechanical Engineering and Sciences, of which 53 (65%) citations were provided to the papers published in the JIF time window. Removing the 53 citations decreases the JIF of the recipient journal by 46%. The donor and recipient journals have 32 and 30 editorial board members, respectively, of which three editors overlap. Two of the overlapping editors serve as the advisor and associate editor of both journals.In addition to groups 1–6, two citation groups caught our attention, which we refer to as groups 7 and 8. Group 7 is present in the network in 2017. This group belongs to category E and consists of two journals on engineering Fig. g. CIDRE Keats Shelley Journal, provided 119 citations to the seven recipient journals, of which 110 (92%) citations were provided to the papers published in the JIF time window. Removing the 119 citations decreases the JIF of the recipient journals by at least 57%. A single paper titled “Annual Bibliography for 2015” provided all the within-group citations from the donor journal to the recipient journals. This paper consists of 42 pages, of which 40 pages are the reference list. In each of year 2012, 2013, and 2014, the donor journal published a paper with a similar title  that cited many papers in the recipient journals. In these years, CIDRE detected the groups that consisted mostly of the donor journal and the recipient journals in group 8.Group 8 is present in the network in 2016. This group belongs to category A and consists of eight journals on literature Fig. h. A donoWhile JCR has not reported any anomalous pairwise citations in the 2019 citation data, CIDRE detected seven citation groups in 2019. Six out of these seven groups belong to category A, B, C, D, or E Fig. . We refeInternational Journal of Surgery  from 34 papers published in the donor journal. The 29 out of the 34 papers are included in a special issue of which the author was the guest editor. The special issue consists of 74 papers.Group 10 is composed of two journals and belongs to category D Fig. b. The doSocial Anthropology, cited 95 papers published in the recipient journals, all of which were published in the time window for the JIF. If one removes the citations from that review paper, the JIF of each of the five recipient journals decreases by more than 18%. In the review paper, the author acknowledged the editors of the two recipient journals, Social Analysis and Focaal, owned by a publisher, Berghahn Journals, for granting access.Group 11 is composed of seven journals on anthropology and belongs to category A Fig. c. A singCrystallography Reviews, cited 124 papers published in the recipient journal, IUCrData, all of which were published in the time window for the JIF. If one removes these 124 citations, the JIF of the recipient journal decreases by 57%.Group 12 consists of two journals on crystallography and belongs to category A Fig. d. A singRegulation and Governance, provided 95 citations to the recipient journal, Annals of American Academy of Political and Social Science. Removing the citations from the donor decreases the JIF of the recipient by 26%. The 89 out of 95 (93%) citations from the donor to recipient journals pointed to the papers included in a special issue of the recipient journal, i.e., “Regulatory Intermediaries in the Age of Governance.” The special issue consists of 16 papers, each of which received less than two citations on average from journals outside group 11 in 2019. The special issue was edited by 3 guest editors who are on the editorial board of the donor journal. The three editors wrote a paper in the special issue. The paper received 26 citations in 2019, of which 13 citations (50%) came from the donor journal. The paper was highlighted as the most cited paper in the last 3 years in the recipient journal in 2019.Group 13 is composed of two journals on political science and belongs to category D Fig. e. The doJournal of Mathematical Sciences and Cryptography, published 150 papers, of which 52 papers cited 36 papers published in the recipient journal, Journal of Information and Optimization Science, in the time window for the JIF. We did not find a single author or a single paper that was exclusively cited or was cited within the group. The 52 papers published in the donor journal were written by 126 authors, of which 107 authors (84%) had never cited the recipient journal before. Both donor and recipient journals have the same chief editor.Group 14 consists of two mathematical journals and belongs to category E Fig. f. The doIn this paper, we put forward an algorithm—named CIDRE—to identify groups of journals that cite each other at excessively high rates. CIDRE detects a majority of journal groups suspended from JCR. Notably, in several cases, it does so years in advance. In addition, it detects a number of anomalous groups, whose members increased their JIFs by 17–130% via within-group citations. The inspection of such groups reveals a variety of mechanisms leading to such inflation. Specifically, more than half of the anomalous groups are due to one paper or one author that singlehandedly provides or receives many citations within the group.30. CIDRE could also be applied in different contexts, e.g., to detect the manipulation of ratings in e-commerce platforms and social media31.The algorithm’s practical value lies in that it is deterministic and scalable to large networks, which makes it possible to apply it in an online fashion to incoming streams of new citation data. Furthermore, it can be applied to different types of networks. For instance, CIDRE could be applied to bipartite author–journal networks, where a directed edge indicates a publication by an author in the journal, in order to detect potential predatory practices, such as the publication of papers with little peer review33, reciprocity between peers34, and editorial preferences36, rather than to outright malicious citation practices. In this respect, CIDRE should not be considered as a tool for automated decision-making or a substitute for expert judgment, but rather a support tool to extract interpretable information from the complexity of journal citation networks.One should be careful when drawing conclusions from the application of CIDRE. The comparison against the ground-truth data provided by JCR, and the manual inspection of the groups detected by CIDRE support that the groups flagged by CIDRE warrant consideration as potential citation cartel candidates. That being said, we ought to acknowledge that some of such candidates may arise due to unintended biases such as geographical proximity37. Then, we manually inspected each group detected by CIDRE to pinpoint individual papers, authors, editors, and specific journals associated with excessive citations. However, manual inspection is a costly task and hard to scale up when dealing with large numbers of groups. This problem will manifest itself when one analyzes citation groups composed of authors because an author network can be much larger than a journal network. Therefore, in practice, it may be useful to prioritize groups that survive with higher thresholds. With CIDRE, one can easily determine the ranking of groups according to this criterion because gradually increasing CIDRE has a parameter—the threshold 38. In this respect, we believe that our results should encourage a more critical and nuanced approach to the use and interpretation of citation-based bibliometric indicators.Regardless of the conclusions that one may draw on specific anomalies, our findings reveal the widespread presence of journals whose JIFs are substantially hoisted by the citations received from a small group of other journals. It would be hard not to relate this with the ever-increasing emphasis on citations and bibliometric indicators, and the pressure it puts on journal editors to boost growth in such numbers. We believe our findings to be a rather direct consequence of this environment, where actors are incentivized to act on the very same metrics according to which they are ranked, in a feedback loop that closely echoes Goodhart’s Law: “when a measure becomes a target, it ceases to be a good measure”We assume that an anomalous citation group is composed of journals that act as donors, recipients, or both. A donor journal gives excessive citations to the journals in the same group. A recipient journal receives excessive citations from the journals in the same group.U, composed of the donor and recipient journals. We quantify the extent to which a journal i acts as donor or recipient within group U using the donor score i, respectively, and N is the number of journals. Function h is an indicator function, where we set i to journal j are excessive relative to the null model; otherwise i, i.e., i cites papers in other journals in U more often than expected under a null model; similar for the recipient score.Algorithm CIDRE finds groups of journals, i to journal j are deemed to be excessive if and only if they satisfy the following two conditions. First, more than half of citations made to papers published in any previous years from i to j were made to papers published in the last 2 years . Second, the number of citations, i to node j, we compute the p-value as the probability w that is larger than or equal to the actual weight of edge  in the given network, i.e., The citations from journal 39 to suppress the false positives due to the multiple comparison problem. In other words, one regards m edges with the smallest p-values as significant . Using exact string matching for the manually inspected names, we matched the editors in different journals.There are 641 journals in the 184 groups detected by CIDRE. We manually identified the web pages listing the editorial board members for 525 among the 641 journals. Extracting human names, particularly non-Latin names, from web pages is challenging. In addition, spelling variation makes it difficult to match editors in different journals. Therefore, we did not aim to calculate the precise number of editorial board members shared by different journals but to calculate its lower bound. Specifically, we extracted person names with the Spacy package


The contributions of processed foods to the overweight and obesity problem in Latin America are well known. Engagement with the private and public sectors on possible solutions requires deeper insights into where and how these products are sold and the related implications for diet quality. This article characterizes the diversity of wheat and maize processed foods (WMPFs) available to consumers in Mexico City. Data were gathered across nine product categories at different points of sale  in high and low socioeconomic (SE) areas. We assessed WMPFs based on Nutri-Score profile, price, and health and nutrition claims. Roughly 17.4% of the WMPFs were considered healthy, of which 62.2% were pastas and breads. Availability of healthy WMPFs was scarce in most stores, particularly in convenience stores Compared to supermarkets in the low SE area, those in the high SE area exhibited greater variety in access to healthy WMPFs across all product categories. In the low SE area, healthy WMPFs were priced 16–69% lower than unhealthy WMPFs across product categories. The extensive variety of unhealthy WMPFs, the limited stock of healthy WMPFs in most retail outlets, and the confusing health and nutrition claims on packaging make it difficult for urban consumers to find and choose healthy WMPFs. Food processors and food retailers target specific products to specific subgroups of consumers in response to heterogeneity in economic, demographic, and behavioral factors. In providing food products tailored to meet consumer demands ,2,3, segWhile conceptualized and interpreted in different ways , a commoThe personal domain “relates to individual level dimensions such as food affordability, accessibility, convenience, and desirability”. The external domain considers “the world of opportunities and constraints that are out there within a given context”, which is shaped by price, vendors, food availability and product properties. Various studies have focused on the internal and external domains and how these are shaped by food processors and retailers. Some found that in-store healthy foods were less likely to be found in low-income neighborhoods in cities in the United States , AustralEvidence suggests that maintaining a healthy diet is relatively expensive given the cost of items that comprise a lower quality diet which tends to include more processed, energy-dense foods . Food veResearchers have called for policies and interventions  to incentivize the consumption of foods of higher nutritional quality ,26. OtheThis study examines, across different types of retail outlets, access by urban consumers to processed wheat and maize products—major occupiers of food retail shelf space that include crackers and chips, tortillas, breakfast cereals, breads, pastas, and frozen foods. As shown in This study seeks insights into the following two questions: (1) What are the characteristics of WMPFs in terms of nutritional quality, health and nutrition claims, and price? (2) How do these vary by levels of socioeconomic attainment and points of sale? The findings here will allow for insights on the nutritional challenge and possible entry points for future engagement with public and private sectors on potential options for promoting increased access to healthy WMPFs for different types of urban consumers.This cross-sectional study presents data on access to WMPFs collected in Mexico City and its surrounding area between March and June 2019. Principal component analysis was used to characterize “basic geographical areas” (BGAs)—administrative units in Mexico City for which data were available. The analysis included 14 variables on socioeconomic (SE) characteristics: housing materials and access to public utilities , material goods , and population characteristics . BGA-level data and information on frequency and location of stores were obtained from the National Statistic Directory of Economic Units and the National Population and Household Census 2010. Those BGAs which did not contain at least two supermarkets and two independently owned grocery stores were excluded. BGAs were categorized into low and high SE BGAs using their first and last deciles of the first principal component of SE characteristics. These criteria resulted in eight high SE areas and eight low SE areas in Mexico City and its surroundings; i.e., the high SE was in the Polanco area, the low SE was in the Chimalhuacán area. Additionally, field reconnaissance of each selected area was conducted to assess diversity in the type of food retailer and ensure the minimum number of food retail outlets, as well as their distribution, for the final selection of the two areas for the study. Four types of food retail outlets were considered for each area: chain supermarkets, independently owned grocery stores, convenience stores, and corner stores. Trained personnel visited each store in a selected SE area and included adjacent BGAs if a specific type of food retailer was not accessible to the data collection team. Twenty-seven food retail outlets were confirmed within each SE area. We were able to increase the final sample to 2 convenience stores , 2 neighborhood corner stores, and 20 independently owned grocery stores in both areas and 3 supermarkets in the high SE area and 2 supermarkets in the low SE area. Most low SE areas did not contain a chain supermarket, thus requiring adjustments in the field to ensure adequate supermarket coverage. We distinguished nine categories of WMPFs for inclusion in the study: ready-to-eat foods, breakfast cereals, salty snacks, cookies, flour, breads, bars and pastries, pastas, and maize tortillas and tortilla products. Within each category, we considered only packaged and labeled products, regardless of origin, which listed wheat or maize among the first three ingredients. We excluded nonpackaged products from traditional stores such as bakeries and tortillerias. Data were collected by taking photographs of information available on each product in the selected product categories at the store. A standardized methodology  was applEach product was classified according to its Nutri-Score profile . The scoPrices obtained from each food category were converted from Mexican pesos to US dollars (USD) by using the exchange rate during the collection period (USD 1 to MXN 19.15) . Prices The study protocol and procedures were approved by the Ethics Committee of the International Maize and Wheat Improvement Center (CIMMYT). Consent to conduct in-store data collection was obtained by email or in person from store representatives. We described the diversity of access to WMPFs by SE area and the type of store with the percentage of distinct WMPFs that were found in both SE areas, in low SE areas exclusively, or in high SE areas exclusively. The same analysis was used to describe the availability of WMPFs by Nutri-Score and food product categories by SE area including all products in the sample. We applied a specified a multiple linear regression model for analyzing price differences between SE areas. The log of price of WMPFs per 100 g was specified as the dependent variable. Explanatory variables included three categorical variables and all their two and three way interactions, these variables were: SE area (low/high), Nutri-Score profile  and food group. We obtained geometric means for each combination of SE area, Nutri-Score profile, and food group category, as well as geometric mean ratios between SE areas, by exponentiating the corresponding linear combination of model coefficients. We derived 95% confidence intervals by back transforming the confidence limits of the linear combinations. Additionally, we fitted a separate model for each type of retail store with the log of price per package as a dependent variable and the same specification of predictors as previously described and obtained the aforementioned differences by SE area and their confidence intervals. We reported the use of health and nutrition claims as percentages for each combination of food group category, SE area, and Nutri-Score profile, with their corresponding logit-transformed 95% confidence intervals , and caltortillas and tortilla products, flour, and breads were the WMPF categories with the least variety of products . Independently owned grocery stores provided the most homogeneous set of WMPFs in both low and high SE areas (from 86.1% to 100% similarity within WMPF categories). Across WMPF categories, cereal bars and pastries (38.7%), salty snacks (36.7%), and breakfast cereals (29.1%) were more frequently found in both SE areas. We recorded a total of 7567 WMPFs available in both SE areas, including unique and repeated products. Of these, 2462 were unique WMPFs. Among unique WMPFs, only 28.8% of them were classified as healthy . SupermaLooking across the sample, only 11.7% and 5.6% of available WMPFs were included in the A and B Nutri-Score profiles, respectively. We found considerably greater availability for healthy WMPFs  in the high SE area . Indeed,The geometric mean (GM) prices per 100 g and per package of healthy and unhealthy WMPFs food profiles were significantly higher in the high SE area compared to the low SE area . For exaHealth and nutrition claims were found on 23.2% of all WMPFs. They were more frequently found on healthy products in the high SE area compared with the low SE area for cookies  and maize tortillas and tortilla products  . A consiIn this study, we collected data on the diversity of WMPFs available to consumers in Mexico City paying attention to variation in access by socioeconomic areas, retail outlets, nutrient profile, and health and nutrition claims employed. The study was motivated by the need for data on what WMPFs were accessible to consumers, as an input for future discussions with the private sector on their role in marketing available healthy and nutritious WMPFs for urban consumers. Our findings highlighted various issues in the retail landscape for WMPFs that have important implications on the dietary options available to consumers, especially those in lower-income neighborhoods. Cereals, including wheat and maize, can contribute essential amino acids, minerals, and vitamins and beneficial phytochemicals and dietary fiber components to the human diet. Although wheat and maize have traditionally been processed for human consumption, current WMPFs normally exceeded the daily recommended intake of sodium, saturated fat, trans fats, and added sugars established by WHO to prevent overweight, obesity, and chronic diseases ,39, and The high SE area had much more diversity in terms of WMPFs available as compared to the low SE area. Previous studies had found that in-store healthy foods were less likely to be found in low-income neighborhoods in cities, and our findings were consistent with these ,17,18,19Mexico’s 8% sales tax on unhealthy foods resulted in a 5.8% decline in the purchase of taxed foods among households of medium SE status and a 10.2% decline among households of lower SE status ; thus, iWe found specialization in retail stores, with convenience stores and neighborhood corner stores specializing in WMPFs with Nutri-Score profiles D and E  WMPFs. Supermarkets had the greater stock of products with profiles A and B . In the low SE area, neighborhood corner stores and supermarkets could provide, in theory, access to healthier WMPFs; however, supermarkets are scarce in low SE areas. Similarly, in high SE areas, neighborhood corner stores and supermarkets could provide greater access to healthier WMPFs, but neighborhood corner stores, as well as independently owned grocery stores, tended to be scarce. Yet, increasing in-store healthy products in small food retail outlets is feasible and can be achieved with support and understanding of healthy product criteria  or incenInterventions to improve healthy food offerings in small stores should consider the diverse environments, stocking practices, and supply mechanisms with innovative distribution practices of small stores, particularly nontraditional food retailers . For exaThe consumption of healthy food is mostly consistently related to income, contrary to the intake of unhealthy ones . HoweverAlthough prices varied greatly across WMPF categories (as expected), higher prices for some foods also may predict lower consumption of such groups, as has been previously demonstrated in several countries . While hThe use of health and nutrition claims can inform consumers through short and clear messages about the energy and nutrient content of a product and its potential health benefits . Howevertortilla, bread, and pastas; others comprised other nonbasic food commodities such as snacks and ready-to-eat foods, which have become part of the modern diet in Mexico. The WMPFs from the first categories tended to be lower in price and of better nutritional value in both SE areas. Maintaining healthy properties and ingredients of WMPFs, including dietary fiber, prebiotics [Some WMPF categories included in our study comprised traditional food ingredients and foods such as flour, ebiotics , resistaebiotics . Moreoveebiotics . Healthy WMPFs have an important role to play in healthy diets , given tThe findings raise important questions about the private sector’s formulation and marketing of WMPFs and the implications for promoting healthier diets. In many developing regions, income growth will continue to push demand towards more processed foods and more foods purchased at supermarkets. The Food Systems Summit in 2021 stated that “the food choices people are faced with and the choices they make are profoundly determined by the food system of which they are part” , and addThis study represents a much-needed initial step to initiate dialogue between researchers, the private sector, and government agencies on how to improve access to healthier WMPFs. It highlights issues of differential access by consumers from higher and lower socioeconomic strata to healthier processed foods. Strengths in our study include the careful selection of the high and low socioeconomic areas  and the variety of food retail outlets visited. To the best of our knowledge, no other study has included all available food retail outlets that comply with our inclusion criteria in a given area. Future work in this area should look to engage the consumers, private sector, government agencies, and civil society on options for increasing access to and purchases of healthy WMPFs.