Cryptococcus neoformans using a random insertional mutagenesis strategy, screening for mutants with defective growth at 37°C. Among several thermosensitive mutants, we identified one bearing a disruption in a gene predicted to encode the Ape4 aspartyl aminopeptidase protein. Ape4 metalloproteases in other fungi, including Saccharomyces cerevisiae, are activated by nitrogen starvation, and they are required for autophagy and the cytoplasm-to-vacuole targeting (Cvt) pathway. However, none have been previously associated with altered growth at elevated temperatures. We demonstrated that the C. neoformans ape4 mutant does not grow at 37°C, and it also has defects in the expression of important virulence factors such as phospholipase production and capsule formation. C. neoformans Ape4 activity was required for this facultative intracellular pathogen to survive within macrophages, as well as for virulence in an animal model of cryptococcal infection. Similar to S. cerevisiae Ape4, the C. neoformans GFP-Ape4 fusion protein co-localized with intracytoplasmic vesicles during nitrogen depletion. APE4 expression was also induced by the combination of nutrient and thermal stress. Together these results suggest that autophagy is an important cellular process for this microbial pathogen to survive within the environment of the infected host.In order to survive and cause disease, microbial pathogens must be able to proliferate at the temperature of their infected host. We identified novel microbial features associated with thermotolerance in the opportunistic fungal pathogen  C. neoformans is a fungal pathogen with worldwide distribution; it is found in the environment on rotting wood and is often associated with bird excreta. This yeast can cause fatal respiratory and neurological infections, especially in immunocompromised populations. Recent surveys estimate more than 500,000 deaths from C. neoformans every year, especially in patients with AIDS and other diseases that compromise the immune system [e system . The imme system , 3.C. neoformans to survive and multiply in the host. Among these virulence factors, the most well studied are the production of polysaccharide capsule, melanin, phospholipase and growth at 37°C [Agrobacterium tumefaciens gene delivery system, in order to identify mutants unable to growth at 37°C. Among several thermo-sensitive mutants, we explored one bearing an insertion in the aspartyl aminopeptidase (APE4) gene. In S. cerevisiae the protein aspartyl aminopeptidase (Ape4) plays a role during autophagy [The options for antifungal therapies for cryptococcosis are limited. The most commonly used drugs for treatment are polyenes (amphotericin B-based drugs), antimetabolites (flucytosine) and azoles . However at 37°C . The abi at 37°C –15. To iutophagy .In eukaryotes, autophagy is defined as a group of processes that occurs inside the vacuoles leading to degradation of cytoplasmic components such as parts of the cytosol, macromolecular complexes and organelles. This process is important to maintain the balance between catabolism and anabolism, allowing the cell to recycle organelles and other cellular components, ensuring availability of basic nutrients that can be used for growth, cell development and favorable energy balance during nutrient shortage, such as nitrogen starvation . PrimariS. cerevisiae. The Cvt pathway occurs in the setting of starvation, sequestering hydrolases that are transferred to the vacuole which is the site of their activity [S. cerevisiae, six proteins were found to be involved in this process: the hydrolases Ape1 (aspartyl aminopeptidase 1), Ape4 (aspartyl aminopeptidase 4) and Ams1 (Alpha-Mannosidase); the phosphorylated cargo receptor Atg19 (AuTophaGy related); the Atg11 adapter protein; and Atg8 which is involved in phagophore expansion [Autophagy can be divided into microautophagy or macroautophagy, and both can further divide in selective or nonselective processes , 23. Selactivity . In S. cxpansion , 23, 25.C. neoformans virulence has been established [APE4 gene and drive the co-localization of C. neoformans GFP-Ape4 fusion protein and intracytoplasmic vesicles. We demonstrate how Ape4 is important for C. neoformans survival within macrophages and virulence in murine animal model. Moreover, we present how other C. neoformans autophagy-related genes are modulated during nitrogen starvation and thermal stress.The requirement of autophagy in ablished , 27, howape4 mutant and ape4+APE4 reconstituted strains were generated in the C. neoformans var. grubii background (KN99α). The standard medium for growth was YPD . To evaluate the influence of nitrogen source, synthetic dextrose (SD) medium was used, either with or without amino acids and ammonium sulfate (SIGMA cat. Y1250 and cat. Y1251 respectively).The In order to evaluate non-preferred nitrogen sources, the SD medium without amino acids and ammonium sulfate was supplemented with 10mM/mL of L-proline or uric acid. To verify the effect of non-preferred carbon source, the SD medium with amino acid and ammonium sulfate was supplemented with galactose (2%). For selection of mutant strains, the growth media were supplemented with Geneticin (Invitrogen–Cat. #11811–023) at 200 μg/ml final concentration.APE4 (CNAG_01169) gene deletion was performed by substituting part of the coding region with an antibiotic resistance marker. The mutant allele construction was designed using PCR overlap extension as previously described [ape4+APE4, were confirmed by lack of growth on YPD supplemented with 200 μg/ml of Geneticin G418 and for the restoration of the ability to grow at 37°C. Reconstituted strains were confirmed by diagnostic PCR and Southern blot. All primers used in the gene deletion procedure are described in supplementary escribed  and intrescribed . The traAPE4 coding region was amplified by PCR using oligonucleotides described in the GFP-APE4 construct was transformed into C. neoformans (KN99α) by biolistic transformation. The mutants were selected on YPD plate supplemented with 100 μg/mL of nourseothricin (Jena Bioscience).The C. neoformans  genomic DNA extractions were performed as described previously [APE4) at its original locus was performed by Southern blot, using the techniques described by Sambrook et al. (1989). The detection of the band patterns of the wild individuals (KN99α), mutant (ape4) and reconstituted (ape4+APE4) was performed by chemiluminescence using a dig-based labeling kit,  and detection system  according to the manufacturer's instructions. We used as a probe the PCR fragment amplified with primers MAV143 and MAV144  grown in YPD for 12 hours were centrifuged and the pelleted cells were washed with saline (NaCl 0.9%). The cell number was determined by counting in a Neubauer chamber, the cell concentration was adjusted to 2x106 cells/ml, followed by 10-fold serial dilution (four dilutions), from where 5μL aliquots were spotted on Petri dish plate containing appropriate media follow by incubation at the required temperature (30°C or 37°C).Cultures of Cryptococcus strain  was incubated at 30° C for 16h to 18h in YPD medium, pelleted, and the inoculum washed three times with PBS. Cells were inoculated to a final OD600nm of 0.5 into either YPD or SD-N-AA liquid media. Cultures were grown under agitation for 24 hours at 30°C and 37°C. This experiment was performed in duplicate. One hundred cells per experiment were measured using a 40x objective. Employing the Anatomic MIPro Standard software, the cell size (μm) was determined. As the Cryptococcus cell is not a perfect circle, we used the software ellipse area tool, where the cell edges were defined and the radius marked, to estimate the cell size (μm). The data generated by the software was treated statistically using ANOVA (GraphPad Prism 5 program). Melanin production was evaluated at 30°C on Niger seed agar, as described [et al. [2-Independent Medium  at 150rpm [The thermotolerance was evaluated in rich medium (YPD) at temperatures of 30°C and 37°C. The cell size assay was performed as follows: each escribed . The abi [et al. . Inductit 150rpm . Cell saThe urease activity was assayed at 30°C in urea agar base as described before .The analMulti-stress sensitivity was evaluated with YPD medium supplemented with 0.75M and 1.5M of NaCl or KCl. The cell wall sensitivity was evaluated on YPD plus 0.5% Congo Red plates. All plates were incubated at 30°C.Sensitivity to antifungal drugs was determined for wild type, mutant and reconstituted strains by standard minimal inhibitory concentration (MIC) assays according to Clinical and Laboratory Standards Institute (CLSI M27-A2) protocols with slight modifications. Briefly, fresh cultures were diluted in saline (0.9% NaCl) and standardized to the turbidity level 1 on the McFarland scale, then inoculated on agar plates containing RPMI-1640 medium (Sigma) buffered with MOPS (Sigma) pH 7. Plates were air dried and E-test strips with fluconazole  or amphotericin B  were carefully laid on the top of the agar.https://techlib.biomerieux.com/wcm/techlib/techlib/documents/docLink/Package_Insert/35904001-35905000/Package_Insert_-_9305056_-_D_-_en_-_Etest_-_AFST_WW.pdf). The results were submitted to statistical analysis. .The biological assays were performed in triplicate at 30°C for 72 hours. This protocol was performed as described by the E-test manufacturer .To localize the Gfp-Ape4 fusion protein, the transformants were incubated in liquid YPD for 18 hours at 30°C with 150rpm. The cells were collect by centrifugation and washed twice with pre-heated (30°C) saline (0.9% NaCl). Ten milliliter of suitable liquid medium (SD without nitrogen source or YPD) supplemented with 10μg/mL of FM4-64 dye (Invitrogen) as described before , was inoThe cultures were incubated in YPD at 30°C and 37°C and SD (without nitrogen source) at 30°C and 37°C for 2 hours. The samples from each treatment were analyzed using a fluorescent microscopy . Fifty cells per sample distributed in eight different visual fields were analyzed. Captured images were processed with the program ZEN, 2012 and subjected to statistical analysis .ape4 and ape4+APE4 grown in YPD broth overnight were washed twice in 0.9% NaCl at room temperature. After washing, the cells were suspended in rich medium (YPD) or Synthetic Dextrose (SD) supplemented with ammonium sulfate and amino acids (Y1250 Sigma), or 10 mM of either uric acid or L-proline. The effect of a non-preferred carbon source was evaluated in SD medium supplemented with galactose 2%. Cultures were standardized to OD600 0.3 and incubated at 30°C with constant shaking (150 rpm—TECNAL mod. TE 420). The optical density (OD600) was determined at 0, 6, 8, 24, 48 and 72 hours . The biological assay was performed in triplicate. The data registered in a Microsoft® Excel worksheet were subjected to statistical analysis .Cultures of KN99α, C. neoformans (KN99α) according to previously described protocols [Total RNA was extracted from rotocols . InductiGPDH1, Varma and Know-Chung, 1999) and 1X Power SYBR Green master mix (Life Technologies). The quantification of transcripts was done on a comparative manner, using the ΔΔCt method [GPDH1 as previously described by [The Real-Time PCR reactions were performed using cDNA (diluted 1:10), with 800 μM of target primers (MAV 205 and MAV 206) and 300 μM primers (AA 301 and AA 302) for the internal control t method  and stanribed by .J774A.1 macrophages (ATCC TIB-67) were co-cultivated with yeast cells as described by  with mod2), the medium from each well was collected into a fresh tube, macrophages were quickly lysed with 0.5% SDS in PBS, the medium plus lysate were diluted 200-fold in PBS and 100 μl of the dilution were platted on YPD agar for CFU counting. This experiment was performed four times. Statistical validation: for each macrophage co-cultivation experiment, CFU counts were compared among strains by one-way ANOVA followed by Tukey’s post-test for comparison of pairs. Differences were considered valid at a 95% confidence interval (p<0.05).Each yeast strain was added to three wells. After 24 hours of incubation . Prior to infection, animals were briefly anesthetized by isoflurane inhalation. The mice were then infected via nasal inhalation with the wild-type strain KN99α, mutant (ape4) or reconstituted (ape4+APE4) strains. The inoculum (105 cells in a 25μL per strain) was prepared from cultures which were incubated overnight in YPD at 30°C. For each strain ten mice were infected.The murine inhalation model of cryptococcal infection , 43 was ape4-infected mice were sacrificed at 40 days post-infection; lungs from two of the surviving mice were harvested, homogenized, serially diluted, and plated on YPD to access the number of viable cells in the lungs. These experiments followed the Duke University institutional guidelines for animal experimentation.Mice were examined twice daily and sacrificed according to clinical measures predicting mortality . Survival data were analyzed by the Kruskal–Wallis test. The Cryptococcus neoformans genome database formerly located at the Broad Institute.The CNAG_01169 sequence was retrieved from the http://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi) and Uniprot (http://www.uniprot.org/blast).The search for protein domains where performed using the tools available at NCBI (S. cerevisiae were obtained from the Saccharomyces Genome Database (SGD), and for the other microorganisms we used the following database available at: http://www.ncbi.nlm.nih.gov/; http://www.broadinstitute.org/; http://www.candidagenome.org/; and http://www.aspergillusgenome.org/.The sequences for the autophagy related genes from in silico analysis.The supplementary C. neoformans gene sequenced retrieved from the data base described above were employed as query in a reciprocal BLASTx to confirm that the best hit would be what we present at the supplementary All C. neoformans using Agrobacterium tumefaciens, as previously reported [APE4 gene, encoding an aspartyl aminopeptidase (CNAG_01169).We created a collection of insertional mutants in reported . In one Saccharomyces Genome Database) revealed that CNAG_01169 has an average amino acid identity of 44.6% compared to the S. cerevisiae aspartyl aminopeptidase 4 (YHR113w/APE4). When we used the CNAG_01169 nucleotide sequence in a BLASTx search against the S. cerevisiae at the SGD, the first hit found was YHR113 . The t-DNA bearing the drug resistance cassette (Neomycin) was inserted after the nucleotide sequences that encode the amino acid glutamine (Q209) in C. neoformans Ape4.BLAST search analysis at NCBI and SGD . The disruption of the APE4 locus was confirmed by a Southern blot assay . To confirm that the ot assay  for two C. neoformans putative Ape4 protein (523 amino acids) is encoded by the CNAG_01169 gene located on chromosome 5. Protein domain analysis at Uniprot suggests that this protein has domains characteristics of the M18 family of metalloproteases, which have protease activity against aspartate and glutamate amino acids located at the amino-terminus of various proteins [The proteins . ConsistC. neoformans Ape4 amino acid sequence was compared to its putative orthologues in Coprinopsis cinerea, Ustilago maydis, Neurospora crassa, Saccharomyces cerevisiae (S288c), Zea mays, Oryza sativa, Arabidopsis thaliana, Chlamydomonas reinhardtii, Homo sapiens, Mus musculus, Xenopus laevis, Danio rerio, Caenorhabiditis. elegans, Toxoplasma gondii and Plasmodium falciparum. The alignment showed that the Ape4 protein from C. neoformans has a high value of similarity to C. cinerea (57%), whereas, the lowest percentage of similarity (31.3%) was observed against the aspartyl aminopeptidase from P. falciparum , mutant (ape4) and reconstituted (ape4+APE4) strains were spotted onto YPD agar medium and incubated at 30°C and 37°C for 72 hours. ape4 mutation. Another C. neoformans mutant, ras1, has its growth impaired at 37°C, and this mutant also has increased cell size consistent with defects in cytoskeleton polarization and cell division [ape4 mutation on cell size. We compared the morphology of the three strains after incubation at 30°C and 37°C in two separate media (YPD and SD-N-AA). We observed that at 30°C in YPD medium the ape4 mutant strain was statistical indistinguishable from wild-type and reconstituted strains where the average cell size was about 4 μm. In the same medium at 37°C, the average cell diameter was increased about 25% for all strains (about 5 μm) with no statistical difference observed among the strains (ape4 mutant showed a significant increase (50%) in cell diameter when nutrition deprivation was combined with high temperature incubation (6 μm). Therefore, we observed that the lack of a functional APE4 gene led to a cell size increase at high temperature, which is more pronounced with concomitant nutritional deprivation. , mutant (ape4) and reconstituted (ape4+APE4) strains. However, with longer incubations, the ape4 mutant displayed a marked decrease in surface capsule compared to control strains. At 37°C, the difference in capsule volume was observed earlier, after 24 hours of incubation. The polysaccharide capsule of al model –47. Prevtriction . Therefoape4 mutant (Pz = 0.39 ± 0.01) was significantly reduced compared to wild type (p<0.05), similar to previously reported ura4 mutant with decreased phospholipase activity [The production of phospholipase, a secreted virulence factor, was evaluated on egg yolk agar. The precipitation zones (Pz) as a measure of phospholipase activity were identical for wild type (Pz = 0.27 ± 0.004) and reconstituted (Pz = 0.26 ± 0.01) strains. However, the phospholipase activity for the activity  Fig 3)..ape4 mutAPE4 gene did not have any impact on production of urease, melanin or mating. Therefore, we conclude that Ape4 is important for capsule and phospholipase production in C. neoformans.Deletion of ape4 and ape4+APE4 strains were incubated on YPD medium containing Congo Red (0.5%w/v) at 30°C for 72 hours. The ape4 mutant failed to growth in the presence of this dye  when compared to the wild and reconstituted strains. Similarly, the growth of the ape4 mutant was not impaired in the presence of 0.75M NaCl  to investigate the synergistic impact that the ape4 mutation would have in association to fluconazole and amphotericin B on yeast cell growth. The wild type (KN99α), mutant (ape4) and reconstituted (ape4+APE4) strains were equally sensitive to amphotericin B  compared to wild type and the reconstituted strains (12 μg/ml)   . These rC. neoformans in macrophages, we co-incubated the three strains with J774A.1 cells and quantified fungal viability by quantitative culture . The ape4 mutant displayed a significant decrease in survival within these cells  showed that autophagy mechanisms were defective in the atg7 mutant, including its survival within the macrophage [To assess the role of autophagy for survival of se cells , similarignaling . Also, Ocrophage .C. neoformans putative autophagy protein Ape4 on virulence trait expression and in vitro survival in macrophages, we tested whether the ape4 mutant would be virulent in an animal model of cryptococcal infection. Using a murine inhalational model of cryptococcosis [ape4), and the reconstituted (ape4+APE4) strains. As shown in ape4) strain-infected mice survived 40 days post-infection. We harvested and homogenized lungs from two of these surviving mice, followed by serial dilution and quantitative cultures. Both animals appeared clinically healthy, however they had viable yeast fungal cells in their lungs (1.15×103±1.2×103 CFU/mg). This finding of minimally symptomatic infection was also observed by other authors studying other C. neoformans mutants with altered virulence [C. neoformans strain with a mutation in the putative autophagy regulator Atg7 also had its virulence attenuated in a murine model.Given our data indicating the role of coccosis , we compirulence , 12, 51.S. cerevisiae, nitrogen starvation modulates the expression of genes involved in autophagy [C. neoformans putative autophagy gene APE4 transcription pattern under nutritional and temperature stress. The expression pattern of APE4 does not change at different temperatures when C. neoformans is incubated in either rich medium (YPD) or synthetic dextrose medium supplemented with an abundant nitrogen source . The analysis of C. neoformans growth patterns on rich medium (YPD) revealed that at 30°C there is no growth difference among wild type (KN99α), mutant (ape4) and reconstituted (ape4+APE4) strains  was observed for growth in uric acid  significantly decreased the growth of the ape4 compared to wild type and reconstituted strains in the same medium condition , had a different effect on the growth of ric acid . Even inondition . In the dextrose  this mutS. cerevisiae the expression of the genes encoding vacuolar hydrolases is influenced by nutritional status since, during nitrogen starvation, there is a significant increase in the amount of vacuolar proteins [In proteins .S. cerevisiae Ape4 localizes in the vesicles; however, its vacuolar transport is accelerated by nutrient starvation, when cytosolic Ape4 redistributes to the vacuole to attend the yeast cells need for more active vacuolar degradation [C. neoformans Ape4 would behave like S. cerevisiae Ape4, we expressed a GFP-Ape4 fusion protein in KN99α (wild type). Fluorescent microscopic analysis showed that GFP-Ape4 was dispersed in the cytoplasm in 100% of cells analyzed at the optimal temperature and nutrition conditions  genes were reported [http://www.yeastgenome.org/) and used BlastX to identify likely homologs using the C. neoformans genome data base at Broad Institute (http://www.broadinstitute.org/). Out of these 34 autophagy-related (ATG) genes reported for S. cerevisiae, we found 21 homologs in C. neoformans  the temperature change from 30°C to 37°C resulted in a significant increase in transcription (p<0.05) (two-fold and above) for 13 genes out of 21 evaluated,  without nitrogen at 30°C leads to at least two-fold transcriptional induction for 15 genes out of 21, whereas among the fifteen, eight genes had more than five-fold induction and three had over ten-fold induction    . When thnd AMS1) . These rC. neoformans [S. cerevisiae APE4, which belongs to a M18 metalloprotease family [S. cerevisiae the Ape4 protein is part of the selective autophagy Cvt pathway, which is unique to this yeast [Previously, we reported the use of an insertional mutagenesis library to uncover traits involved in high temperature growth in oformans . We furte family , 44. In is yeast . Also Apis yeast , 44. Culis yeast .C. neoformans Ape4 in high temperature growth by independently generated ape4 mutants. These finding were consistent with Hu et al‘s (2008) description of autophagy-related genes impairing C. neoformans growth at 37°C. Exploring further the effects of the ape4 mutation in this yeast we found that C. neoformans GFP-Ape4 fusion protein is also located within a vesicle, which is formed upon nitrogen starvation. Yuga et al (2011) reported that S. cerevisiae Ape4 transport from the cytoplasm to the vacuole is accelerated by nutrient starvation to meet the requirements for more active vacuolar degradation [C. neoformans, but due to its nature as a possible protease we believe that C. neoformans Ape4 is somewhat involved with nutrient recycling during starvation in this yeast. It seems likely that its function takes place within vesicles during nitrogen deprivation, and this effect seems more dramatic when the cells are at higher temperature (37°C), a situation where more protein is recruited to the vesicle. Also, the combination of elevated temperature and nitrogen deprivation leads to a 9-fold increase in APE4 transcript abundance, reiterating its role in response to nutritional and temperature stress.We confirmed the role of radation . Our datC. neoformans cells encounter a hostile environment upon host infection, confronting challenges including nutrient depletion and high temperature [C. neoformans genes, ATG3 and ATG9, are up-regulated 2.6- (2 hours) and 3.2-fold (24 hours), during murine macrophage infection [C. neoformans, first implicating autophagy in cell adaptation in an unfavorable environment. Our results confirmed that the transcription of these two genes is indeed induced 8- and 22-fold, respectively, when C. neoformans is incubated under nitrogen shortage and high temperature. Recently, Oliveira et al. (2016) showed that the C. neoformans ATG7a putative autophagy regulator is important for virulence in this yeast [The perature . The autperature . In 2005is yeast . Our traAspergillus fumigatus, Richie et al. (2007) showed that lack of a functional autophagy process led to impaired radial growth and conidiophore development at 37°C [C. albicans the atg9 mutant was deficient for autophagosome formation during nitrogen starvation, and it also showed defects in yeast-hyphae differentiation. However, this mutant was able to survive within the macrophage and was fully virulent in a mouse model of hematogeneously disseminated candidiasis, suggesting that this autophagy-related protein is dispensable for pathogenesis [C. albicans revealed that autophagy is important for ER stress response and tolerance to antifungal drugs [C. neoformans ape4 mutant is more sensitive to fluconazole, implicating autophagy in antifungal tolerance. Basse’s group studying autophagy in Ustilago maydis uncovered that atg11 and atg8 are important for mitochondrial recycling [U. maydis atg8 rendered the mutant strain less virulent, with reduced teliospore production. A similar but less pronounced phenotype effect was observed for atg1. However the combination of both atg mutations exacerbated these phenotypes [Studies in other fungi species have demonstrated a complex interplay between autophagy related proteins and microbial pathogenesis/antifungal tolerance. In  at 37°C . In C. aogenesis . A recenal drugs . Similarecycling , 64. Alsenotypes .in silico analysis for C. neoformans genes predicted to play a role in autophagy, using the S. cerevisiae genes in a genome BLAST search Click here for additional data file.S2 Table(DOC)Click here for additional data file.S1 Fig(PPT)Click here for additional data file.S2 Fig(PPT)Click here for additional data file.S3 Fig(PPT)Click here for additional data file.


Corrections to be noted in Volume 91 of the JOURNAL OF RESEARCH of the National Bureau of Standards


PLOS Neglected Tropical Diseases marked its 10th anniversary. As highlighted during a celebratory anniversary event held at the World Health Organization in Geneva, PLOS Neglected Tropical Diseases was founded to coincide with the shaping and creation of the modern framework of neglected tropical diseases (NTDs) [In 2017, s (NTDs) . It was PLOS Neglected Tropical Diseases articles are tracked on social media and highlighted in the world’s most widely circulating newspapers. Many of our papers have influenced the global policy makers from both disease-endemic nations and donor countries as well as WHO and other major international health agencies. Overall, our journal has raised awareness about the NTDs and played an important role in promoting access to NTD medicines and treatments as well as access to innovation by supporting research and development on these diseases.Some of our “big-picture” metrics and achievements were highlighted previously . They inPLOS Neglected Tropical Diseases, and together with my friend and colleague Professor Serap Aksoy, to guide the journal through its first decade. There is no question that PLOS Neglected Tropical Diseases has been one of my greatest passions, and it’s been a fulfilling experience serving as Co-Editor-in-Chief (EIC).It has been a great honor for me to help lead efforts that launched the founding of PLOS Neglected Tropical Diseases through its next journey. Beginning in January 2018, I will transition away from day-to-day journal management in order to serve as the founding EIC. In that role, I hope to continue contributing editorials and viewpoints for PLOS Neglected Tropical Diseases, providing overall guidance for future directions, and representing our journal at future summits and meetings when asked to do so. I also hope to continue submitting research articles on our NTD vaccine development program.Like with any important journal, I also think it’s wise to refresh and look to new leaders who can shepherd PLOS Neglected Tropical Diseases will continue to remain at the vanguard of new approaches for peer review and open access. I’m excited to see how our journal progresses as it takes on new and innovative approaches and projects, and I welcome the opportunity to help it move into its second decade!Professor Judd Walson will now join Professor Aksoy as Co-EIC. Many in the NTD community already know Professor Walson through his clinical trial and implementation research in infectious disease or in his current role as Deputy EIC. I want to thank Professor Walson for agreeing to take on the role of Co-EIC as I transition to founding EIC. There continue to be many opportunities to innovate and improve biomedical publishing, particularly as it relates to diseases of the poor. I expect and hope that


The changes of the morphology during heating and the degree of crystallinity of dry and hydrated starch granules are investigated using second harmonic generation (SHG) based Stokes polarimetry. A spatial distribution of various polarization parameters, such as the degree of polarization (DOP), the degree of linear polarization (DOLP), and the degree of circular polarization (DOCP) are extracted and compared with the two dimensional second harmonic (SH) Stokes images of starch granules. The SH signal from hydrated and dry starch on heating differed significantly in DOLP and DOCP values, indicating that hydrated starch has a greater degree of ultrastructural amylopectin disorder. The detail of denaturation and the phase transition of hydrated starch demonstrate the significant influence of thermal processing. In order to optimize the processing operations and obtain the desired quality of starch-based foods, a thorough understanding of the starch-water interaction through the gelatinization process is required16Second harmonic generation (SHG) microscopy is an effective analytical tool for detailed investigation of microscopic structure of non-centrosymmetric molecules11121411161711In pixel by pixel Stoke vector based SH image analysis, it is found that at room temperature the double helical amylopectin is self-organized upon hydration within starch granulesThe polarization-resolved SH images were obtained from potato starch in both dry (laboratory condition) and hydrated (suspended in water) condition using a Stokes-polarimeter. The starch granules were extracted from homegrown potatoes  and 90° (I90) polarization intensities, S2 is the difference between the +45° (I45) and −45° (I135) polarization intensities, and S3 is the difference between the left . A series of MATLAB  programs were developed to fully reconstruct the data.The measured Stokes vectors of SH light can be expressed as STherefore, using Stokes matrix based formalism, crucial physical parameters of the SH light can be inferred from the measured Stokes parameters1112The physical significance of these parameters are indicates the polarization property of the light. If light is perfectly polarized, then DOP = 1 and 0 for an unpolarized light. For partially polarized light, DOP is in between 0 and 1, depending on the degree of polarization. The DOLP indicates the alignment of molecules parallel to the linear polarization states; DOLP is in between 0 and 1. The DOCP is a measure of how effectively the molecules flip the circularly scattered lights within the focal volume and the value ranges from 0 to 1.0), Stoke vector (S3) and DOCP of dry and hydrated starch granules at room temperature. SHG images of dried potato starch granules were compared to the granules embedded in distilled water and a better behavior was observed for the hydrated polarization marker0) granules . The SH image of hydrated starch is more uniform than that of the dry starch, which is due to the increase in crystalline order of the starch granules during hydration230) are due to the scattering and crystalline order of the amylopectin during hydration. Recently, Richard et al. also found out the role of scattering by measuring the SH signal only of dry and hydrated potato starch253 and DOCP). In another study, the image histogram of helical angle of SHG active molecule (amylopectin) remains unchanged under hydration3 and the DOCP which correlate to the helicity of the amylopectin molecules are unchanged during hydration compared to dry starch.We investigated the polarization states and reconstructed polarization parameters of SH light from both dry and hydrated starch in laboratory conditions. The SH signal depends on the geometrical characteristics and the relative phase difference of SH active molecules within the molecular arrangements1412The detail characterization of SH light from potato starch at different temperatures was performed using Stokes vector based polarimetry. Stokes vectors and polarization parameters are revealed in a pixel by pixel analysis which are visualized in the respective color bar as shown in 0), Stokes vectors  and DOP, DOLP, DOCP of hydrated starch granules at two different temperatures.The effect of temperature on the swelling power of starch acts differently2727282930310) of dry starch is observed at 84 °C, which is due to the presence of atmospheric water molecules inside the laboratory. Again, it is reported that in the case of dry starch granules, the hydroxide bonds are retained due to the loss of water and hydrogen bonds oriented closer to the helical axis of crystal domain15The SH signal and the respective Stokes vectors of dry starch granules are measured with increasing temperature. As shown in The amylopectin lamellae are arranged in the A, B-type crystalline form in potato starch and3436726The origin of chirality in starch has been determined using second-harmonic-generation circular dichroism (SHG-CD) microscopyWe investigated the starch gelatinization process through a Stokes vector based four-channel polarimeter integrated within a SH microscopy. The technique was shown to yield the polarization states of the SH signal, as quantified by the Stokes parameters, for a fixed input polarization state. Various polarization parameters of SH signals were calculated and analyzed, further elucidating structural properties of individual starch granules. The degradation of intermolecular integrally occurs in hydrated starch whereas disruption of surface in dry only. Thus, hydrated starch upon heating changes the polarization properties of the SH signal significantly, which suggests the deformation of crystallinity in starch granules. In the case of dry starch, there is no significant change of polarization properties observed at higher temperatures. Therefore, the fundamental understanding of the optical nonlinearity of starch granules will provide a valid basis for future studies in food science and insights into the energy transformation dynamics.How to cite this article: Mazumder, N. et al. Investigating starch gelatinization through Stokes vector resolved second harmonic generation microscopy. Sci. Rep.7, 45816; doi: 10.1038/srep45816 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


Flat enamel surfaces, mounted in acrylic resin, were bonded to the zirconia discs (diameter: 4.7 mm). All specimens were subjected to shear bond strength testing using a universal testing machine with a crosshead speed of 1 mm/min. All data were statistically analyzed using one-way analysis of variance and multiple-comparison least significant difference tests (α = 0.05). Hot-etching treatment produced higher bond strengths than the other treatment with both resin cements. The shear bond strength of all groups significantly decreased after the thermal cycling test; except for the hot-etching group that was cemented with Panavia F2.0 (p < 0.05). Surface treatment of zirconia with hot-etching solution enhanced the surface roughness and bond strength between the zirconia and the resin cement.This study was designed to evaluate the effect of hot-etching surface treatment on the shear bond strength between zirconia ceramics and two commercial resin cements. Ceramic cylinders  were randomly divided into 12 groups ( High-strength materials such as zirconia can offer improved fracture resistance and longer-term viability ,2. Howev2O3 particles and different air pressures can be used to roughen the surface of zirconia [There has been extensive research concerning the adhesion between zirconia and resin cements . Particlzirconia ,8. Tribozirconia ,10. Howezirconia ,12. Recezirconia . Our prezirconia  revealedMetals and zirconia have some similarities. For this reason, hot-etching technology, which was initially used with nickel chromium-alloy Maryland bridge wings , was expin vitro study was to actualize hot-etching technology with an effective and simple application using a reaction kettle, and to investigate changes in the surface morphology and bond strength between zirconia and resin cements after the etching procedure. The null hypotheses of this study were that the use of the surface hot-etching treatment would not influence the surface roughness of zirconia and would not affect the adhesion between zirconia and the resin cements.The aim of this 2O3; Tosho, Tokyo, Japan) by the Biomaterials Department of Tsinghua University : 12.5 GPa). The specimens were sequentially surface polished with SiC abrasive papers from grit #180, 240, 480, 600, 1000, 1200 to 2000  under flowing water, rinsed with tap water for 1 min, ultrasonically cleaned in a deionized water bath for 30 min and gently air-dried.A total of 120 specimens of zirconia were compacted and sintered into dense disc specimens  from yttria-stabilized zirconia powder  according to the different surface treatments performed: Group 1: blank control;2O3 particles applied for 10 s at 0.25 MPa of air pressure; andGroup 2: airborne-particle-abrasion with 50 μm AlGroup 3: hot-etching treatment for 1 h. Specimens were placed in a reaction kettle  that was filled with hot-etching solution  . The ketSpecimens were divided into three groups  and evaluated under a SEM  at 25,000× magnification to assess the changes in surface topography.Each zirconia disc was imaged using Atomic Force Microscopy (AFM) . Five random 5 μm × 5 μm spots on the surface of each disc were imaged at a probe speed of 1 Hz  at 256 × 256 resolution. SPI4000 software  was used for analyzing the image and the surface roughness. The average surface roughness (Ra) values and standard deviation for each specimen were calculated.m (m (111) and tetragonal t (111) phases were used to determine the extent of phase transformations after the different surface treatments. The relative amount of the monoclinic phase (Xm) was calculated according to the formulas proposed by Garvie and Nicholson [XRD analysis  using Cu Kα radiation was conducted to examine the influence of the different surface treatments on the phase composition and transformation of the zirconia. Zirconia surfaces were scanned with Cu Kα X-rays over an angular range of 2θ = 15°–75° with a step size of 0.026° and a 5 s step interval. The maximum peak intensities of the monoclinic icholson .The Ethics Committee of the Hospital of Stomatology, Peking University (PKUSSIRB-201520021) granted permission to use isolated teeth.A total of 120 human middle incisors that were free of caries, fluorosis, cracks, or previous endodontic treatments were selected from incisors recently extracted for heavily reduced periodontal support at the Hospital of Stomatology, Peking University. The teeth were stored in 0.5% chloramine solution at 4 °C and external debris was removed using a hand scaler. The root part of each tooth was sectioned 1 mm below the cement-enamel junction with a diamond burr  to obtain the crown. Each crown was embedded in self-curing resin in a Teflon mold to form a cylindrical specimen . The labial surface of the crown was exposed at the top of the cylinder. After solidification of the self-curing resin, the labial surface of the crown was polished with SiC abrasive paper  under flowing water to achieve a uniform 5 mm × 5 mm enamel surface. The isolated tooth specimens were discarded when the area of the exposed enamel was less than 5 mm × 5 mm or if a chalky color surface was not observed after etching by 35% phosphoric acid in the step described below.The zirconia specimens from the three different treatment groups were further subdivided according to the two different resin cements  and whether or not the thermocycling fatigue testing was performed. Each subgroup contained 10 specimens.The treated surfaces of the zirconia were cemented to the enamel surfaces according to the manufacturer’s instructions. In the Superbond C and B subgroup, the enamel surfaces were etched with a layer of red Activator Superbond C & B for 30 s, rinsed and then dried. The Superbond C and B monomer, catalyst and polymer were mixed at a ratio of 4:1:1 and the mixture was then applied to the surface of each zirconia surface. In the Panavia F2.0 subgroup, the enamel surfaces were etched with a layer of phosphoric acid  for 30 s, rinsed for 15 s and air-dried. ED Primers A and B were combined and painted on the enamel surface for 30 s and air-dried. The combined A and B paste was applied to the surface of the zirconia.The zirconia specimens were cemented on the surface of the enamels and then compressed at a constant load of 10 N for 20 s; excess cement was then wiped off. The specimens of the Panavia F2.0 group were light-cured for 40 s, then all of the specimens were stored in distilled water for 24 h.Half of each group was subjected to a thermocycling test  in deionized water for 5000 cycles between 5 and 55 °C. The dwell time at each temperature was 30 s and the transfer time was 2 s (ISO 10477-1996) .Each specimen was loaded onto a universal test machine  equipped with a 1000 kg load cell. The shear bond strength test was performed according to the guidelines specified in the ISO/TS 11405-2003 . The sheAfter debonding, the fractured interfaces of the specimens were examined with a stereomicroscope  at 10× magnification to determine the debonding modes as follows: ZR, failure between the zirconia surface and resin cement; ER, failure between the enamel and resin cement; MM, mixed mode; and CF, cohesive fracture inside the resin cement.One-way analysis of variance (ANOVA) in combined with least significant difference (LSD) tests performed with the IBM SPSS Statistics  were used to analyze differences in surface roughness and the shear bond strength values among groups (α = 0.05), after the normal distribution and homogeneity of variance were checked using the Shapiro-Wilk test and Levene tests, respectively.p < 0.01). No significant difference was detected between the airborne-particle-abrasion and hot-etching groups (p > 0.05). The shear bond strength of each group was listed in p < 0.01). The bond strengths were significantly lower after the thermal cycling test in all groups (p < 0.05) except for the hot-etching group that was cemented with Panavia F2.0 (p = 0.08).Representative SEM and AFM images of each group were shown in The distribution of the different failure modes of each group was shown in M was calculated from these patterns. The amount of monoclinic phase was negligible in the control and hot-etching groups, and the airborne-particle-abrasion group had a higher relative amount of the monoclinic zirconia phase.The use of hot-etching treatment on zirconia has been recently studied . The actIn the present study, a simple kind of reaction kettle was used to carry out the hot-etching process. The reaction kettle was of a type widely used in the chemical field for acid etching. The inner tank of the kettle was made of polytetrafluoroethylene (PTFE), which is an extremely stable material, and it was covered with a shell made of stainless steel. The test temperature (100 °C) was suitable for conditioning the substrate in a water bath application. The boiling point of methanol is 64.7 °C, so a pressure-tight reaction kettle was needed to heat the solution to the reaction temperature. The etching rate inside the kettle was accelerated by using a rotor that was activated by the magnetic agitator.p < 0.01); these results are in agreement with Casucci et al. [p < 0.01).Strong resin bonding relies on micromechanical interlocking and adhesive chemical bonding to the ceramic surface, requiring surface roughening for mechanical bonding and surface activation for chemical adhesion . The prei et al. . The etcp < 0.05).XRD analysis revealed that the airborne-particle-abrasion group contained a higher relative amount of the monoclinic zirconia phase, in agreement with other studies ,26. The p < 0.05). The cellular structure etched by the acid solution was of a form favorable for micromechanical interlocking with the resin cement as well as to the enamel after a three-step etch-and-rinse process. This result contradicts those of El-Korashy [The present study demonstrated that the hot-etching treatment significantly enhanced the shear bond strength of resin cement to zirconia when compared with no treatment and airborne-particle-abrasion treatment for both immediate testing and testing after thermal cycling (-Korashy  who concp < 0.05). This decrease might relate to the different coefficients of thermal expansion of the bonded substrates. Since Panavia F2.0 contains the phosphate ester monomer MDP (methacryloyloxydecyl dihydrogen phosphate), some reaction may have occurred between the adhesive monomer and the zirconium oxide layer coated on the zirconia surface [After thermal cycling, the shear bond strength significantly decreased for all of the groups except for the hot-etching group bonded with Panavia F2.0 ( surface . Combinep > 0.05).Superbond resin cement containing 4-META, and Panavia F2.0 containing 10-MDP, are commonly used in the clinic. In previous studies, these two cements were recommended for bonding to zirconia having different surface treatments ,30,31. TThe failure mode observed was mostly failure between the ceramic surface and the resin cement. The shear bond strength could reach its ultimate value under this failure condition.p < 0.05). This treatment is an alternative treatment to airborne-particle-abrasion and avoids micro-crack formation and phase transitions that are detrimental to the longevity of the ceramic restoration. However, the hot-etching treatment still requires further study before being used as a step in the processing of zirconia restorations: control of the hot-etching area, alteration of cytotoxicity and biaxial flexural strength after hot-etching are all issues that need attention. The limitation of this study was that only one hot-etching reaction time was used, the optimal reaction time and other technology parameters need to be explored in future work: only one zirconia quality, combined with two resin composite cements, were tested; the universal property of this hot-etching method needs to be researched.The present study indicated that the use of the hot etching technology as a pre-treatment of zirconia significantly enhanced the adhesion between zirconia and resin cement ; andHot-etching treatment produced significantly higher shear bond strength between zirconia and resin cement compared to no treatment and airborne-particle-abrasion treatment (Hot-etching treatment could provide a rougher zirconia surface while avoiding the detrimental tetragonal-to-monoclinic surface phase transition.Within the limitations of this laboratory study, the following conclusions are drawn:


This study analyzed the maintenance of lateral incisors in the dental rehabilitation of individuals with cleft lip and palate.The study was conducted on a tertiary craniofacial center and comprised retrospective analysis of panoramic and periapical radiographs of Caucasoid individuals with non-syndromic complete unilateral cleft lip and palate, analyzing all radiographs available on the records of each individual, from the first to the last up to 12 years of age. Overall, 2,826 records were reviewed to achieve a sample of 1,000 individuals. Among these, 487 individuals presented the permanent lateral incisors on both cleft and non-cleft sides, which were included in this study.The results were evaluated in percentages and by descriptive statistics. The association between maintenance of the lateral incisor and timing of alveolar bone graft were analyzed by the t test. Among the 487 individuals, 265 had not completed treatment, 62 presented insufficient information, and 44 concluded the treatment elsewhere. Among the remaining 116 individuals, the lateral incisor was extracted from 88 (75.86%) of them on the cleft side (CS) and from 23 (19.83%) people on the non-cleft side (NCS). The age at accomplishment of alveolar bone graft was significantly associated with maintenance of the lateral incisor on the cleft side (p<0.01). Most extractions were indicated because of the inadequate positioning on the CS and for midline correction on the NCS. Rehabilitation was primarily completed by orthodontic movement .In conclusion, the lateral incisor on the cleft side was not maintained in most individuals. Positive relationship was observed between extraction of the lateral incisor and age at accomplishment of the alveolar bone graft, suggesting the need to anticipate the initial radiographic evaluation to enhance its maintenance and reduce the procedures required for rehabilitation. Individuals with cleft lip and palate present complex skeletal deformities and are subjected to a treatment load that requires several procedures, which begin in childhood and continue up to adulthood, aiming to restore the normal morphology and function.,,,,,The treatment of alveolar defects usually requires alveolar bone graft,Despite the high success rates reported in the literature for the secondary alveolar bone graft, there are controversies concerning the age of accomplishment, suggesting the need to establish a specific treatment protocolThis study was approved by the Institutional Review Board of HRAC/USP (protocol no. 241/2011).The inclusion criteria were: 1) Caucasoid individuals with non-syndromic complete unilateral cleft lip and palate, 2) presence of panoramic and periapical radiographs in the individual's files, from the first radiographs obtained up to the last up to 12 years of age, in addition to thorough dental history in the records to analyze the presence or absence of the permanent lateral incisors on the cleft and non-cleft sides, 3) individuals originally presenting permanent lateral incisors on both cleft and non-cleft sides.A single examiner reviewed 2,826 records of individuals with non-syndromic complete unilateral cleft lip and palate, regularly registered in the institution. Among these individuals, we selected those whose records contained panoramic and periapical radiographs from the first to the last up to 12 years of age, which led to a sample of 1,000 individuals. Among these, an additional selection was performed to include only individuals whose radiographs available and dental history allowed reliable analysis of the presence or absence of permanent lateral incisors on both cleft and non-cleft sides. This led to a sample of 487 individuals who presented the permanent lateral incisors on the cleft and non-cleft sides, who were included in this study .post-hoc power of the study was calculated using the following parameters: sample size 487 individuals/ percentage of extraction of lateral incisors in this sample 76%/putative percentage of extraction of lateral incisors in the overall population 1%/alpha error of 0.05, which revealed a post-hoc power of 100%.The authors did not find any study analyzing the extraction of lateral incisors for orthodontic/dental rehabilitation. For this reason, sample size calculation could not be performed. Thus, after study completion, the This study comprised retrospective analysis of such records, searching for information about maintenance of the permanent lateral incisor on the cleft and noncleft sides for completion of dental rehabilitation. In cases with indication of lateral incisor extraction, information about the specialist who indicated the extraction and the reason for such decision was achieved, in addition to the treatment indicated for space closure in the cleft region. Secondarily, the study also evaluated the association between maintenance of the lateral incisor on the cleft side and age at accomplishment of alveolar bone graft, using the t test, at a significance level of p<0.05. The other results were evaluated in percentages and by descriptive statistics.Among the 487 individuals previously selected who presented the permanent lateral incisor on both cleft and non-cleft sides, 265 individuals had not completed the dental rehabilitation until the onset of this study, 62 individuals exhibited insufficient information on the records, and 44 completed the treatment elsewhere. Thus, the other analyses were conducted on a sample of 116 individuals with complete unilateral cleft lip and palate, with presence of one permanent lateral incisor on both cleft and non-cleft sides, and dental rehabilitation concluded before the onset of this study.The lateral incisor on the cleft side was maintained in 28 individuals (24.14%), while the lateral incisor on the non-cleft side was maintained in 93 individuals (80.17%).The mean age at accomplishment of alveolar bone graft was 14.30 years (SD=4.63), ranging from 8 to 30 years. Statistically significant association was found between age at bone graft and maintenance of the lateral incisor only for the cleft side .In individuals submitted to extractions, the specialist indicating them was informed only on the records of 26 individuals, with predominance of orthodontists (21), followed by maxillofacial surgeons (4) and prosthodontist (1). The reason for extraction indication was described in 16 cases , includiThe specialist who indicated the extraction of the lateral incisor on the non-cleft side was described in the records of 13 individuals, all of which were indicated by the orthodontist. The reason for extraction indication was described in nine cases, including midline correction (4), inadequate positioning (3) and achievement of space at the posterior region (2). In cases of extraction of the lateral incisor, the corresponding space was rehabilitated by orthodontic movement with mesial movement of the canine (13) and fixed prosthesis (1), without information available for the other nine individuals.,,The rehabilitation of individuals with complete unilateral cleft lip and palate is complex and involves several stages, including alveolar bone graft, which is fundamental to join the alveolar segments,,,,Bone graft should be performed whenever possible to facilitate the eruption of teeth close to the cleft or orthodontic movement, either of the canine or lateral incisorThe eruption and adequate positioning of the permanent lateral incisor in the cleft area maintains the bone graft by mechanical stimulation, but also achieves stable functional occlusion, allowing normal maxillary growth and more harmonious facial and dental estheticsIn this study, the mean age at accomplishment of alveolar bone graft was 14.30 years (SD=4.63), ranging from 8 to 30 years. This may have been influenced by the continental dimensions of the country and predominantly low socioeconomic status of the population assisted at the institution; and this may have contributed to the low percentage of maintenance of the lateral incisor on the cleft side, as demonstrated by the statistically significant association between maintenance of the lateral incisor and age at accomplishment of alveolar bone graft . However,Our results revealed indication of 17 prostheses and 10 implants, and the space corresponding to the maxillary lateral incisor on the cleft side was closed by orthodontic mesial movement in 53 individuals . When neUnfortunately, several records were incompletely filled by the different professionals treating the individuals in the institution. This is an inherent limitation of retrospective studies, especially those conducted in large institutions involving different specialists, such as in this case.Based on these results, the authors concluded that 1) most of the lateral incisors (76%) adjacent to the cleft were extracted; 2) the main reason for extracting the lateral incisor was its inadequate position; 3) the earlier the bone grafting procedure is accomplished, the greater are the chances of maintaining the lateral incisor; and 4) the rehabilitation of the cleft area in most of the cases was achieved by means of orthodontic space closure.Thus, analyzing the maintenance of lateral incisor for dental rehabilitation of individuals with complete unilateral cleft lip and palate, we observed that evaluation for alveolar bone graft is mostly performed considering the canine, i.e. when the lateral incisor is already erupted, thus missing the ideal timing of bone graft for this tooth, despite present and in good conditions. Considering the reports of success of secondary alveolar graft performed before eruption of the lateral incisor and the low rate of use of this tooth when bone graft is performed after eruption of the lateral incisor, as demonstrated in this study, the possibility of a slight anticipation in the timing of alveolar bone graft might be considered to increase the possibility of utilization of the lateral incisor. These findings suggest the need to customize the timing of orthodontic evaluation and alveolar bone graft in individuals with lateral incisor in the cleft area, considering the possibility of its eruption through the bone graft to increase the maintenance of the lateral incisor, when present, and reduce the burden of care for dental rehabilitation of individuals with cleft lip and palate.The lateral incisor on the cleft side was not maintained in most individuals. There was positive relationship between extraction of the lateral incisor and age at accomplishment of the alveolar bone graft, suggesting the need to anticipate the initial radiographic evaluation to enhance its maintenance and reduce the steps required for rehabilitation.


The American College of Medical Physics (ACMP) and the Journal of Applied Clinical Medical Physics (JACMP) are very pleased to announce that with this issue the JACMP is now an official journal of the International Organization for Medical Physics (IOMP). As an all‐electronic journal and with the current non‐subscription policy the journal is available and accessible to medical physicists world‐wide. From the beginning of publication over two years ago, it was clear that the journal was being accessed in many different parts of the world. About 20% of the manuscripts submitted are from outside the United States, and to date papers from over half‐a‐dozen countries have been published. It has been the editorial policy of the journal to encourage this participation, and this agreement to become an official journal of the IOMP will further strengthen our ties with the international community of medical physicists.This concept was initiated by the Board of Editors a year ago and was presented to the Board of Chancellors of the ACMP at the annual meeting in June of 2001. With their concurrence the Editor‐in‐Chief proceeded with discussions with Gino Fallone, Chairman of the IOMP Publications Committee. This resulted in the invitation by Oskar Chomicki, the President of the IOMP, for the JACMP to become an official journal of the IOMP. The ACMP and the JACMP wish to express their thanks to the IOMP and its officials for their constructive discussions that made this possible.Under the terms of the agreement the Editor‐in‐Chief of the JACMP will be an ex officio member of the IOMP Publications Committee and a representative of the IOMP will be invited to attend the Editorial Board meetings of the JACMP.The ACMP and the JACMP look forward to many years of fruitful cooperation with the IOMP.Peter R. Almond, Ph.D.Editor‐in‐Chief


Dengue in Tokyo and Key West. Leishmaniasis in Madrid and Texas. Chikungunya in Ravenna and Puerto Rico. SARS in Toronto. Ebola in Dallas. And measles everywhere. Population growth, globalization, climate change and urbanization together are reshaping the context upon which we must consider the global spread of emerging and historically tropical infectious diseases and their interface with non-communicable diseases of transitioning societies.We live on a crowded and interconnected planet, with a projected global human population of at least 9 billion by 2050 . Public International travel and immigration increase each year, with more than 1 billion humans crossing international borders in 2013 alone [E. coli infection associated with Middle Eastern fenugreek sprouts, [Aedes albopictus  via imported used-tires from Asia [Our current age of globalization also involves the increasingly rapid and direct transportation of food and other products that can introduce or facilitate disease transmission, whether it be cyclosporiasis associated with Latin America raspberries,  enterohesprouts,  monkeyposprouts,  or introrom Asia .Evolving global climate change and severe weather events can also be expected to have significant impacts on the distribution, spread and burden of many tropical and global infections; unfortunately, we just do not yet have the ability to accurately predict what these impacts will be. Two obvious areas that warrant special surveillance are vector-borne and water-borne diseases. For instance, regional burdens of malaria, dengue and chikungunya may all increase with the effect of increasing temperature on their mosquito vectors, and cholera and leptospirosis can increase in the setting of flooding associated with severe weather events . The effesa.un.org/unpd/wup/FinalReport/WUP2014.pdf Accessed 12 June 2015). This dense concentration of people lacking appropriate infrastructure support facilitates the transmission of contagious diseases while providing a ready source of susceptible individuals, whether it be influenza, typhoid fever or Ebola.The urbanization of the human population is impacting the risk of transmission of many infectious agents. Currently, more than 80 % of humans live in developing countries, and more than half of these people now live in urban settings, many in informal settlement or slum conditions  was created with the hope of providing a forum where important discovery, observation and insights may find a global audience. As an open access journal, TDTMV articles will be freely available, and through an article-processing charge waiver program for researchers from low and low-middle income countries (as defined by the World Bank in September 2014) [TDTMV will endeavor to provide a much-needed forum for targeted updates in clinical tropical and travel medicine to serve as an easily-accessible source for the busy practitioner and trainee.The journal of er 2014) , we hopeTropical Diseases, Travel Medicine and Vaccines, not only to help the journal become a top journal in its field, but also improve the lives of many through shared knowledge. Our many thanks in advance.Finally, the launch and future success of this journal is dependent upon the esteemed Editorial Board, the peer reviewers and you. Thus, we welcome you to submit your research, observations, experience and perspectives to


The right gastroepiploic artery is commonly used in coronary artery bypass grafting. Appropriate strategies are required when performing upper abdominal surgeries after the right gastroepiploic artery has been used in coronary artery bypass grafting because compressing or injuring the graft may cause myocardial ischemia and fatal arrhythmias. To our knowledge, this is the first reported case of surgery for achalasia performed after coronary artery bypass grafting using the right gastroepiploic artery. We have discussed the surgical procedure and particular intraoperative considerations.A 62-year-old man who had undergone coronary artery bypass grafting using the right gastroepiploic artery presented with achalasia. Because medication and balloon dilation had been ineffective and he was having difficulty ingesting food, we performed a Heller–Dor procedure via laparotomy. The right gastroepiploic artery was not damaged during this surgery, and there were no perioperative cardiovascular complications. Adequate control of symptoms was achieved.When performing upper abdominal surgeries after coronary artery bypass grafting with the right gastroepiploic artery, it is necessary to investigate the patient carefully preoperatively and adapt the intraoperative procedure to minimize risk of injury to the graft and consequent cardiovascular complications. The right gastroepiploic artery (RGEA) is sometimes used in coronary artery bypass grafting (CABG) as an alternative source of an arterial graft. It is recognized as a reliable conduit with superior long-term patency and is easy to anastomose to the right coronary artery . AccordiAchalasia is a rare disease, its incidence being approximately 1/100,000 annually and its prevalence 10/100,000 . AlthougThe patient was a 62-year-old man. Twelve years previously, he had undergone CABG for angina pectoris and a bypass between the right coronary artery and the RGEA had been constructed. Six years previously, he had been referred because of difficulty swallowing. He was diagnosed as having achalasia and treated with nicardipine hydrochloride and balloon dilatation in the Gastrointestinal Medicine Department of our hospital. He was referred to our surgical department again because he remained symptomatic, and we were consulted about surgical treatment for achalasia. Gastrointestinal endoscopy showed dilatation of the whole esophagus and food residues within it Fig. . A bariuWe discussed the surgical procedure with cardiac surgeons preoperatively and ordered them to back up, considering the possibility of re-anastomosis at the time of the damage of the graft and the onset of cardiovascular complications. Additionally, we prepared papaverine hydrochloride against the graft spasm.We also discussed the operative procedure in detail. It was expected that there is adhesion of intraperitoneal tissue because of the previous surgery. We could not deny presence of adhesion of the RGEA bypass graft and abdominal wall through the preoperative examination. Therefore, we decided to perform laparotomy in view of a risk of the graft damage at the time of the port insertion. Additionally, because it took time to cope with damage of the graft in laparoscopic surgery, we selected laparotomy after discussion with cardiac surgeons.An oblique incision was made below the left costal arch in order to avoid the adhesion because of the previous surgery. The extent of adhesion was mild. Searching intraperitoneal at the time of laparotomy, we identified the RGEA graft wrapped by soft tissues due to palpation. The RGEA graft ran along the left edge of the round ligament of the liver and reached to the heart via the ventral and lateral side of the liver. We performed surgical procedure so as not to damage this graft while checking the graft beats by palpation appropriately. Especially when we operated the cardiac region, we performed carefully because we needed to retract the liver to secure the visual field. The second assistant dedicated to observation around the graft. After dissecting around the hiatus esophagus, the abdominal esophagus was pulled up by using a vessel tape and as much dissection as possible was performed around the cranial esophagus. The dilated esophagus was then pulled caudally and straightened under observation by gastrointestinal endoscopy (GIS). The vagal nerves, including the hepatic branch, were preserved. Myotomy was performed for 6 cm on the esophageal side and 2 cm on the gastric side (Heller myotomy) and dilation of the narrow segment observed via GIS. Having recognized that the fornix was mobile, a Dor fundoplication was performed. Additionally, two shoulder stitches were placed in the crus of the diaphragm. Finally, straightening and dilation of the esophagus was confirmed by GIS. The RGEA was not damaged during the procedure, and no fluctuation in circulatory dynamics or electrocardiographic changes occurred. On postoperative day 3, he started drinking water and on day 5 started feeding. His postoperative course was good, and he was discharged on day 12. Two months later, a barium esophagogram showed that the failure of the esophagogastric junction to relax had improved and barium was flowing smoothly into the stomach after deglutition. Postoperative esophageal manometry showed the stillness pressure of the esophagogastric junction had decreased, confirming the therapeutic effect of the operative procedure. At present, his symptoms are adequately controlled.When performing upper abdominal surgery after CABG using RGEA, it is necessary to assess whether the graft is contributing to the coronary artery blood flow. If the graft is obstructed or coronary angiography shows a “to and fro” phenomenon despite the graft being patent, the graft is not contributing to the coronary artery blood flow and its inherent blood flow is being maintained. In such cases, standard lymphadenectomy with resection of the RGEA can be performed, for example, during gastrectomy for gastric cancer.Conversely, if the graft is contributing to the coronary artery blood flow, injury or compression, including indirect injury or compression, may cause myocardial ischemia, coronary failure, and fatal arrhythmias. It has been reported that stretching of the abdominal wall can depress the ST segment in an electrocardiogram . AdditioTo avoid graft injuries, it is necessary to assess the route of the graft by enhanced CT or abdominal angiography preoperatively: it can be difficult to identify the graft intraoperatively because of adhesions after CABG. In such cases, indocyanine green fluorescent angiography may help with identification of the graft.It is also important to recognize how the graft for CABG has been procured. Two major techniques are employed, namely pedicle grafts that include surrounding soft tissues and skeletonized grafts that lack them. Pedicle grafts can be difficult to distinguish from the falciform ligament of the liver and mesentery, which may result in injury to a pedicle graft.During both achalasia surgery and other abdominal surgeries, it is necessary to be careful not to injury the graft with a wound retractor used during laparotomy. There are two standard routes for RGEA grafts, namely an ante-gastric and a retro-gastric route; the latter is preferred by most cardiac surgeons . TherefoThe other possible countermeasure is to first perform an angioplasty on the coronary artery. In one report of a patient having advanced gastric cancer after CABG using the RGEA, rerouting revascularization of the RGEA graft was performed via thoracoscopic surgery prior to gastric surgery . In any To our knowledge, this is the first reported case of surgery for achalasia performed after CABG using the RGEA. Surgical procedures for achalasia comprise Heller–Dor surgery (laparotomy or laparoscopic) and per-oral endoscopic myotomy . In our For these reasons, we believed that Heller–Dor via laparotomy was the procedure of choice for this patient.We here present a case of surgery for achalasia after CABG with RGEA. When performing upper abdominal surgeries in such cases, it is necessary to investigate the patient carefully preoperatively and plan the intraoperative procedure so as to minimize risk of injury to the graft and consequent cardiovascular complications.


Liver grafts from donation after circulatory death (DCD) donors are increasingly accepted as an extension of the organ pool for transplantation. There is little data on the outcome of liver transplantation with DCD grafts from a pediatric donor. The objective of this study was to assess the outcome of liver transplantation with pediatric DCD grafts and to compare this with the outcome after transplantation of livers from pediatric donation after brain death (DBD) donors.All transplantations performed with a liver from a pediatric donor (≤16 years) in the Netherlands between 2002 and 2015 were included. Patient survival, graft survival, and complication rates were compared between DCD and DBD liver transplantation.In total, 74 liver transplantations with pediatric grafts were performed; twenty (27%) DCD and 54 (73%) DBD. The median donor warm ischemia time (DWIT) was 24 min (range 15–43 min). Patient survival rate at 10 years was 78% for recipients of DCD grafts and 89% for DBD grafts (p = 0.32). Graft survival rate at 10 years was 65% in recipients of DCD versus 76% in DBD grafts (p = 0.20). If donor livers in this study would have been rejected for transplantation when the DWIT ≥30 min (n = 4), the 10-year graft survival rate would have been 81% after DCD transplantation. The rate of non-anastomotic biliary strictures was 5% in DCD and 4% in DBD grafts (p = 1.00). Other complication rates were also similar between both groups.Transplantation of livers from pediatric DCD donors results in good long-term outcome especially when the DWIT is kept ≤30 min. Patient and graft survival rates are not significantly different between recipients of a pediatric DCD or DBD liver. Moreover, the incidence of non-anastomotic biliary strictures after transplantation of pediatric DCD livers is remarkably low. There is a growing discrepancy between the extensive number of patients waiting for liver transplantation and the availability of organs . TherefoTransplantation of DCD donor livers was introduced in the Netherlands in 2001 . The firThe aim of this study was to analyze the outcome after transplantation of pediatric (age ≤16 years) DCD liver grafts since the introduction of a national protocol for the procurement of DCD livers in the Netherlands. For this purpose, the outcome after transplantation of pediatric DCD livers was compared with that of pediatric DBD livers in the same time period.A retrospective cohort study was performed including all liver transplantations with grafts recovered from pediatric DCD donors aged 16 years or younger in the Netherlands between January 2002 and December 2015. The results of liver transplantation with pediatric DCD grafts were compared with those of pediatric DBD grafts performed in the same time period. The examined parameters included patient survival rate, graft survival rate, and rate of complications including PNF. Follow-up was until August 2016. High urgency and split liver transplantations were excluded since they were only performed with DBD grafts.The DCD liver grafts were all procured from controlled donors (Maastricht category III). Liver procurement was cancelled when a) the period between withdrawal of life support and circulatory arrest exceeded one hour, b) the period of hypoperfusion  in the donor exceeded 15 minutes, or c) the asystole time (the period between circulatory arrest and start of cold aortic perfusion) exceeded 30 minutes ,18,19.The technique of DCD and DBD organ procurement was performed according to national protocol and was described in detail elsewhere ,18,19. IIn DBD, brain death was determined according to a standard procedure . An intaIn both donor types, after initiating perfusion, the abdominal and thoracic cavity was filled with ice-cold 0.9% sodium chloride solution and slushed ice for topical cooling. Once procured, the livers were packed and stored on melting ice.The Eurotransplant organization allocated the liver grafts to adult and pediatric recipients, according to their position on the waiting list. Centers were allowed to refuse a liver graft, resulting in an allocation to the next recipient on the waiting list. DCD grafts were not transplanted in Germany due to legislation stipulating that organs could only be recovered from DBD donors and DCD transplantation was prohibited. Standard piggy-back orthotopic liver transplantation was performed if possible. Immunosuppressive regimen evolved over the study period and mainly consisted of induction with basiliximab and maintenance immunosuppression with a calcineurin inhibitor (tacrolimus or cyclosporine) and a rapid taper of steroids, either with or without mycophenolate mofetil.Detailed information regarding the Dutch donors and recipients was obtained from the Dutch Organ Transplant Registry, to which data was prospectively submitted by all organ procurement and transplant centers. The Dutch Transplantation Foundation maintained the registry. The information regarding foreign recipients was obtained from transplant centers within the Eurotransplant region.Donor characteristics that were collected included age, sex, and cause of death. Graft and preservation information included donor warm ischemia time (DWIT), asystole time, cold ischemia time (CIT), and anastomosis time. DWIT was defined as period between withdrawal of life support and in situ aortic cold perfusion; asystole time was defined as time between circulatory arrest and in situ aortic cold perfusion; CIT was defined as the time between in situ aortic cold perfusion and removal of the liver from the ice-cold preservation fluid for implantation into the recipient. Anastomosis time was defined as the time between removing the liver graft from the cold preservation fluid to revascularization of the liver.Recipient characteristics that were collected included age, sex, indication for liver transplantation, time spent waiting for a liver transplantation, earlier transplantation, cause of graft loss, cause of recipient death, and complications. MELD (model for end-stage liver disease) score was calculated as laboratory based MELD score with additional points for standard exceptions according to Eurotransplant criteria.Recipient survival was defined as the time from transplantation to recipient death. Graft survival was defined as the time from transplantation to retransplantation or recipient death. Complications included PNF, infection, hepatic artery thrombosis, portal vein thrombosis, and rejection. PNF was defined as liver failure requiring retransplantation or leading to death within seven days after transplantation without any identifiable cause and other causes of failure such as surgical problems, hepatic artery thrombosis, portal vein thrombosis and acute rejection .http://www.federa.org) and conducted in accordance with the Declaration of Helsinki. This study was approved by the Dutch Transplantation Foundation which is responsible for maintaining the Dutch Organ Transplant Registry. This type of research is compliant with Dutch legislation and this study was retrospectively approved by the Medical Ethical Review Board of the University Medical Center Groningen. The reference number of the approval statement is M16.204932. None of the transplant donors were from a vulnerable population and all donors or next of kin provided informed consent that was freely given.Collection, storage and use of patient data were performed in agreement with the ‘Code of Conduct for health research’, put forward by the federation of Dutch medical scientific societies . Data was presented as median with interquartile range in parenthesis or as number with percentages. Continuous data was compared with Mann-Whitney U test and proportions with Fisher’s exact or chi square test, when appropriate. Graft and recipient survival analyses were determined with the Kaplan-Meier method and significance of survival differences was determined with the log rank test. Rates of complications were compared between groups with univariate logistic regression analysis. Tests were all 2-sided and p-values less than 0.05 were considered statistically significant.Between 2002 and 2015, a total number of 74 liver transplantations with pediatric grafts were performed with 20 (27%) DCD and 54 (73%) DBD grafts. The median follow-up of functioning grafts was 85 months (43–125 months), 36 months (24–113 months) for the DCD group and 93 months (61–126 months) for the DBD group. The minimum follow-up was 8 months.Donor, preservation, graft and recipient characteristics are summarized in Patient survival rate was similar for recipients of DCD versus DBD liver grafts  . After tGraft survival rate was 65% at 1 year in the DCD group, compared to 82% at 1 year in the DBD group (p = 0.20) . At 10 yMost cases of graft failure (84%) occurred within three months after transplantation. Graft loss occurred in seven of twenty (35%) recipients of DCD grafts and twelve of 54 (22%) recipients of DBD grafts. The etiology of graft loss of the DCD grafts is summarized in In the DCD group, the DWIT exceeded 30 minutes in four patients. Graft failure occurred in 100% of patients who received a DCD liver with DWIT ≥30 minutes, whereas it occurred in 19% of patients who received a DCD livers with DWIT <30 minutes (p = 0.007). At present the DWIT is kept below 30 minutes as it was recently reported that an extended DWIT of more than 30 minutes is associated with a significantly increased risk of graft failure ,23. If dThe rate of complications within the first year after transplantation was not different between DCD recipients and DBD recipients . PNF occThis multicenter study with the largest series of pediatric DCD liver transplantation reports good long-term outcome with 78% patient survival and 65% graft survival at 10 years after transplantation. Patient survival, graft survival, and complication rates were similar between recipients of a pediatric DCD or DBD liver. Moreover, the observed rate of biliary complications and NAS after transplantation of DCD liver grafts was relatively low and no differences were noted between pediatric DCD and DBD livers.The patient survival rate of pediatric DCD livers in the current study was in line with that of adult DCD liver grafts (78% versus 80–92% at 1 year respectively) ,12,24–26In comparison with the current study, the graft and patient survival rates were higher in the previously reported ten cases of transplantation of pediatric DCD grafts (100% in the UCLA group [n = 7] and in the Birmingham group [n = 3]) –17. The In the present study the incidence of NAS in pediatric DCD grafts was relatively low and similar to the incidence in pediatric DBD grafts (5% versus 4% respectively). Interestingly, the incidence of NAS in pediatric DCD livers was considerably lower than that reported in adult DCD livers ,28–30. AAlthough the low number of cases warrants careful interpretation of the results of the current study, this study triples the amount of reported transplantations with pediatric DCD liver grafts. Furthermore, the results of pediatric DCD grafts were compared with pediatric DBD grafts to obtain the best estimate of the effect of warm ischemia on these relatively small size pediatric livers. However, as result of small group size a multivariable analysis was not appropriate and survival analyses could not be corrected for differences in baseline characteristics. One of the differences in baseline characteristics was the recipient age which was higher in the DCD than in the DBD grafts. In the current study pediatric DCD livers were generally not transplanted in younger recipients with a long life expectancy. The DCD grafts were probably considered as suboptimal organs because long-term graft survival of DCD livers was considered to be inferior to DBD livers. However, in DCD liver transplantation with adult livers, survival rate of grafts functioning after 1 year is equivalent to that of functioning DBD livers, which is illustrated by graft survival curves of DCD grafts that run parallel to that of DBD grafts at 1 year after transplantation –5,24. AlIn conclusion, this paper describes the largest series of liver transplantation with pediatric DCD grafts and triples the number of reported cases. The results of this multicenter study demonstrate good long-term patient and graft survival rates after transplantation of pediatric DCD livers, especially when DWIT is limited to 30 minutes. Also, the results of this study indicate that risk of NAS is relatively low in pediatric DCD liver grafts. These are important findings in the current era of organ shortage and high mortality rate on the waiting list.


SRCs of the oat flours increased when the solvents turned from neutral (water and NaCl) to acidic (5% lactic acid) or alkaline , and rose as the metal ion valencies of the metal salts  increased. The β-glucan contents were significantly positively correlated with the SCSRC (0.83**), CCSRC (0.82**), SCHSRC (0.80**) and FCSRC (0.78*). SRC measurements of β-glucan in oat flours revealed that the CCSRC values were related with β-glucan (0.64*) but not related with protein and starch. CaCl2 could therefore potentially be exploited as a reagent for β-glucan assay.This study measured the solvent retention capacities (SRCs) of flours from eight oat varieties and one wheat variety against different solvents to explore the swelling volume of oat flour with different solvents, and thus provide a theoretical basis for quick β-glucan analysis. The SRC profile consists of water SRC (WSRC), 50% sucrose SRC (SSRC), 5% lactic acid SRC (LASRC), 5% Na The refractive index of the isolated oligosaccharides is measured with water as the mobile phase and then quantitively determined by HPLC. However, the HPLC method requires expensive analytical instruments and manual operation [2CO3 and 5% lactic acid, commonly adopted to measure flour solvent retention capacity (SRC), resulting in four evaluation indexes for wheat flour qualities. For wheat flours, glutelin is lactic acid SRC (LASRC)-related, damaged starch is CO3 SRC (SCASRC)-related, pentosan is sucrose SRC (SSRC)-related, and all flour compositions are water SRC (WSRC)-related [Currently, β-glucan is mainly determined by enzyme-catalyzing, flow injection analysis (FIA) using fluorescence spectroscopy and High Performance Liquid Chromatography (HPLC) ,7,8. Forperation . Therefoperation . There a-related .+ and Ca2+ and presents an increasing water retention capacity in a solution of which the pH is regulated from a neutral one to an acidic one.Oat β-glucan possesses all physiological properties of common dietary fibers, such as water retention, chelating and absorption to cholesterol and bile acid molecules and improving environments for microbe communities in intestinal tract ,14,15,16In this study, we determined the retention capacities of oat flours in different solvents, elicited differences in the retention capacities among several solvents, investigated the relationships between solvent retention capacities and β-glucan contents, and screened proper solvents to predict β-glucan contents in oat flours. All these would contribute to the development of a quick method for β-glucan analysis.As shown in Of the oat flours, the Calibrations of unit Chopin Dubois (UCDc) of damaged starch is averaged at 9.78, with a varying range within 4.35–12.80. Samples containing larger amounts of damaged starch (expressed by UCDc) are from Bayou 1 (12.80) and Bayou 12 (12.10) and that containing a lower amount is Baiyan 19 (4.35). Remarkably, UCDc of Zhengmai 129 (25.10) was significantly higher than those of oat flours (9.78). Particle size of oat flour (60–80 mesh sieve) is larger than that of wheat flour (100–120 mesh sieve), thus, damaged starch content of oat flour is lower than that of wheat flour. Damaged starch is defined as the starch of which surrounding cell membrane and granules are damaged by extruding, cutting, shearing, rubbing, ripping and other action forces in the process of milling . Native The WSRC, SSRC, LASRC, SCASRC values of oat flours were measured using conventional SRC measurements are summarized in 3 SRC (FCSRC), CaCl2 SRC (CCSRC), NaCl SRC (SCSRC), sodium cholate SRC (SCHSRC) values of oat flours are 157.09%, 99.65%, 88.21% and 64.13%, respectively. The flour with the higher SCSRC values are from Baiyan 9 (95.95%) and Baiyan 2 (95.60%), while the flour with the lowest SCSRC value is from Zhengmai 129 (70.70%). The flour with the highest CCSRC value is from Baiyan 2 (110.85%), and the flour with the lowest CCSRC value is from Zhengmai 129 (85.95%). The flour with the highest FCSRC value is from Bayou 9 (181.10%), and the flour with the lowest FCSRC value is from Zhengmai 129 (109.80%). The flour with the highest SCHSRC value is from Zhengmai 129 (77.45%), and the flour with the lowest SCHSRC value is from Bayou 1 (53.55%).Modified SRC measurements were also used to identify the effects of metal ions, SRCs against different solvents are summarized in Modified SRC measurements were used to identify the effect of different solvent with the same pH, SRCs against different solvents are summarized in 2CO3 (pH 10) (11.38), CaCl2 (8.25), 5% Na2CO3 (8.13), FeCl3 (8.10), SDS (pH 10) (7.84), NaCl (6.77), 50% sucrose (3.89). The coefficient of variation (CV) is a statistic that measures the degree of variability in the data, and can determine whether there are significant differences among samples. Significant differences among samples can help expected the experimental errors.As shown in 2CO3), the SRCs of the oat flours slightly increased. Zhang [With their solvent pH turned acid (lactic acid) or alkaline , β-glucan (with a content of 88.20%) and starch (with a content of 90.33%) were extracted from the oat flours and then added into the flours of Baiyan 2, Baiyan 15 and Bayou 9 . The preWith the addition of β-glucan, flours from the three oat varieties present the values of WSRC, SSRC, LASRC, SCASRC, CCSRC, SCHSRC that tended to increase slightly . SimilarWith more protein added, the flours tend to show varying but generally increasing SRCs . The valWith more starch added, all the SRCs of the three oat flours tend to decrease . HoweverWith both β-glucan and protein added at 2%, the former exerts significantly stronger influence on the SRCs than the latter , indicatBased on the Pearson’s correlation coefficients in 2 has the potential to be used as a β-glucan assay reagent for the solvent retention capacities method.In one word, the CCSRC values are only related with the β-glucan content and have no relation with the protein and starch contents. As a result, CaClEight oat varieties, including Baiyan 2, 13, 14, 15 and 19 were obtained by Baicheng Academy of Agricultural Sciences in Jilin Province, and Bayou 1, 9 and 12 were obtained by Zhangjiakou Academy of Agricultural Sciences in Hebei Province. One wheat variety, Zhengmai 129 were provided by Henan Provincial Academy of Agricultural Sciences. After the foreign matters removed, wheat and oat grains were milled into whole flours with lab mill  and stored at 4 °C for future use. All other chemicals were reagent grade.β-Glucan , protein2CO3 and 5% lactic acid; NaOH, Na2CO3 and SDS with the pH of 10; and 1 M NaCl, CaCl2, FeCl3 and sodium cholate.The following reagent solutions were prepared: 50% sucrose, 5% Naβ-Glucan was measured using a β-glucan assay kit ; moisture content was measured following approved method 44-15A ; total pWSRC, SSRC, LASRC and SCASRC values of eight oat flours and one wheat flour were done by conventional SRC measurements. SRC measurements were conducted using CHOPIN SRC machine . The measurement procedure was conducted as follows: 5.000 ± 0.005 g flour samples were weighed and placed into 50 mL centrifuge tubes. Tubes and samples in combination were weighed using a balance and the 25 g required solutions were drawn into syringes. The tubes and syringes were placed in corresponding positions as indicated on touch screen. After the instrument was turned on, the solutions in the syringes were injected into the tubes and the shaking and static system was in operation. Afterwards, they were nonstop centrifuged at 1000 g for 15 min and allowed to gradually come to a standstill. Next, the tubes were drained upside down for 10 min, taken out and weighed. Finally, the SRCs were automatically calculated. The SRCs were calculated by the following formula :SRC(%)=In which wet pellet is the sample weight after centrifugation and draining. All SRC analyses were performed in triplicate and the coefficient of variation of the SRC value was less than 2.0%.Modified SRC measurements keep the conventional SRC measurement procedure, but changed the solvents. SCSRC, CCSRC, FCSRC, SCHSRC, SHSRC, SCABSRC and SDSSRC values of eight oat flours and one wheat flour were measured in triplicate.Flours of Baiyan 2, 15 and Bayou 9 were sampled, and the extracted β-glucan, protein and starch were added in them separately to increase the contents of β-glucan (β-glucan content was increased by 1% and 2%), protein (protein content was increased by 2% and 4%) and starch (starch content was increased by 5% and 10%) in flours; and WSRC, SSRC, LASRC, SCASRC, CCSRC and SCHSRC values were measured in triplicate.p < 0.01 and p < 0.05.The values were provided as means ± standard deviation (SD). The statistical significance of the differences among the parameters was assessed using analysis of variance (ANOVA) by Statistical Analysis System software 9.2 , and group means were considered to be significantly different at 2CO3, CaCl2, FeCl3, pH 10 NaOH and pH 10 Na2CO3), and rose as the metal ion valencies of the metal salts  decreased.The SRCs of the oat flours increased when the solvents turned from neutral (Water and NaCl) to acidic (5% lactic acid) or alkaline ; the SCSRC, SCASRC, SDSSRC, WSRC, FCSRC, SHSRC, SCABSRC values were significantly positively or extremely significantly positively correlated with the protein contents ; only the FCSRC values (−0.69*) were significantly negatively correlated with the starch contents; and the SRCs against all the solvents were not correlated with the damaged starch contents.In the same individual oat samples with different β-glucan, protein and starch contents, the CCSRC values were significantly positively correlated with β-glucan (0.64*), but not related with protein (0.39) and starch (−0.39), so that it could be employed as being a reagent for β-glucan assay. This result provided a theoretical basis for fast β-glucan assay and laid down a better foundation for β-glucan used in food industry.