Bactericidal and antibiofilm properties of Rumex japonicus Houtt. on multidrug-resistant Staphylococcus aureus isolated from milk.

Multidrug-resistant (MDR) Staphylococcus aureus and its biofilm formation have been challenging to control in milk and dairy industries. Biofilms formed by Staph. aureus may result in the failure of antibacterial agents and disinfectants to penetrate the biofilm in an attempt to control contamination. Novel natural antibacterial agents are required to combat MDR bacteria and biofilms. In this study, we evaluated the bactericidal, antibiofilm, and antimotility effects of Rumex japonicus Houtt. (RJH) extract on MDR Staph. aureus isolated from milk. The RJH extract exhibited good antibacterial activity against MDR strains with minimum inhibitory concentrations (MIC) ranging from 0.78 to 6.25 mg/mL and minimum bactericidal concentrations ranging from 3.125 to 12.5 mg/mL. The extract showed strong inhibition of biofilm formation (81.9%) at sub-MIC value and eradication of biofilm at higher concentrations. The motility of Staph. aureus was effectively blocked by the extract. Major compounds emodin, chrysophanol, and physcion were identified in RJH extract using HPLC-linear trap quadrupole (LTQ)/Orbitrap-mass spectrometry. The extract was nontoxic to human epithelial cell lines such as Caco-2 and HT-29 cell lines at concentrations ranging from 0.1 to 0.5 mg/mL, and from 0.1 to 0.75 mg/mL, respectively. These findings suggest that RJH extract could be an alternative to synthetic preservatives in milk and dairy products.

Microsatellites, single nucleotide polymorphisms and a sequence tagged site in starch-synthesizing genes in relation to starch physicochemical properties in nonwaxy rice (Oryza sativa L.).

Starch characteristics determine the quality of various products of rice, e.g., eating, cooking and processing qualities. Our previous study indicated that molecular markers inside or close to starch synthesizing genes can differentiate the starch properties of 56 waxy rices. Here we report microsatellite (or simple sequence repeat, SSR) polymorphism in the Waxy (Wx) gene, soluble starch synthase I gene (SS1) and starch branching enzyme 1 gene (SBE1), single nucleotide polymorphism (SNP) in Wx and starch branching enzyme 3 gene (SBE3), and a sequence tagged site (STS) in starch branching enzyme 1 gene (SBE1) among 499 nonwaxy rice samples and their relationships with starch physicochemical properties. The nonwaxy rice samples consist of landraces (n = 172) obtained from germplasm centers and cultivars and breeding lines (n = 327) obtained from various breeding programs. Ten (CT)( n ) microsatellite alleles, (CT)(8), (CT)(10), (CT)(11), (CT)(12, )(CT)(17), (CT)(18), (CT)(19), (CT)(20), (CT)(21), and (CT)(22), were found at the Wx locus, of which (CT)(11) was the most frequent, and (CT)(12), (CT)(21) and (CT)(22) were identified for the first time. Four (CT)( n ) microsatellite alleles were found at the SBE1 locus, (CT)(8), (CT)(9), and (CT)(10 )together with an insertion sequence of CTCTCGGGCGA, and (CT)(8) alone without the insertion, of which (CT)(9) and the insertion was a new allele identified in only one rice, IR1552. Multiple microsatellites clustered at the SS1 locus, and in addition to the three alleles previously detected (SSS-A = (AC)(2)...TCC(TC)(11)...(TC)(5)C(ACC)(11), SSS-B = (AC)(3)...TCT(TC)(6)...(TC)(4)C(ACC)(9), and SSS-C = (AC)(3)...TCT(TC)(6)...(TC)(4)C(ACC)(8)), one new allele (SSS-D = (AC)(2)...TCC(TC)(10)...(TC)(4)C(ACC)(9)) was found. Analysis of the starch physicochemical properties of the samples with different microsatellites, SNPs and STS groups indicated that these molecular markers can differentiate almost all the physicochemical properties examined, e.g., apparent amylose content (AAC), pasting viscosity characteristics, and gel textural properties. Wx SSR and Wx SNP alone explained more variations for all physicochemical properties than the other molecular markers. The total six markers could explain 92.2, 81 and 86% of total variation of AAC, gel hardness (HD), and gel cohesiveness (COH), respectively, and they could explain more than 40% of the total variation of hot paste viscosity (HPV), cool paste viscosity (CPV), breakdown viscosity (BD), setback viscosity (SB) and gel adhesiveness (ADH). However, only 29% of the total variation of peak viscosity (PV) and 37% of pasting temperature (PT) could be explained by all the molecular markers. Some of these markers can differentiate the starch physicochemical properties among the rice samples with the same Wx allele, indicating that the variation within Wx allele classes can be explained by other starch synthesizing genes. These SSRs, SNPs and STS are useful in marker-assisted breeding for the improvement of starch quality of rice.

Nucleotide diversity in starch synthase IIa and validation of single nucleotide polymorphisms in relation to starch gelatinization temperature and other physicochemical properties in rice (Oryza sativa L.).

The characteristics of starch, such as gelatinization temperature (GT), apparent amylose content (AAC), pasting temperature (PT) and other physicochemical properties, determine the quality of various products of rice, e.g., eating, cooking and processing qualities. The GT of rice flour is controlled by the alk locus, which has been co-mapped to the starch synthase IIa (SSIIa) locus. In this study, we sequenced a 2,051 bp DNA fragment spanning part of intron 6, exon 7, intron 7, exon 8 and part of 3' untranslated region of SSIIa for 30 rice varieties with diverse geographical distribution and variation in starch physicochemical properties. A total of 24 single nucleotide polymorphisms (SNPs) and one insertion/deletion (InDel) were identified, which could be classified into nine haplotypes. The mean pairwise nucleotide diversity pi was 0.00292, and Watterson's estimator theta was 0.00296 in this collection of rice germplasm. Tajima's D test for selection showed no significant deviation from the neutral expectation (D = - 0.04612, P > 0.10). However, significant associations were found between seven of the SNPs and peak GT (T (p)) at P < 0.05, of which two contiguous SNPs (GC/TT) showed a very strong association with T (p) (P < 0.0001). With some rare exception, this GC/TT polymorphism alone can differentiate rice varieties with high or intermediate GT (possessing the GC allele) from those with low GT (possessing the TT allele). In contrast, none of these SNPs or InDel was significantly associated with amylose content. A further 509 rice varieties with known physicochemical properties (e.g., AAC and PT) and known alleles of other starch synthesizing genes were genotyped for the SSIIa GC/TT alleles. Association analysis indicated that 82% of the total variation of AAC in these samples could be explained by a (CT)n simple sequence repeat (SSR) and a G/T SNP of Waxy gene (Wx), and 62.4% of the total variation of PT could be explained by the GC/TT polymorphism. An additional association analysis was performed between these molecular markers and the thermal and retrogradation properties for a subset of 245 samples from the 509 rice varieties. The SSIIa GC/TT polymorphism explained more than 60% of the total variation in thermal properties, whereas the SSR and SNP of Wx gene explained as much as the SSIIa GC/TT of the total variation in retrogradation properties. Our study provides further support for the utilization of the GC/TT polymorphism in SSIIa. As shown in our study of 509 rice varieties, the GC/TT SNP could differentiate rice with high or intermediate GT from those with low GT in about 90% of cases. Using four primers in a single PCR reaction, the GC/TT polymorphism can be surveyed on a large scale. Thus, this SNP polymorphism can be very useful in marker-assisted selection for the improvement of GT and other physicochemical properties of rice.

Analysis of the genetic behavior of some starch properties in indica rice ( Oryza sativa L.): thermal properties, gel texture, swelling volume.

Starch comprises about 90% of milled rice, so that the eating and cooking quality of rice is mainly affected by the starch properties. In the present paper, we analyzed the genetical behavior of gelatinization temperature tested by differential scanning calorimetry (DSC), gel texture, and the swelling volume (SV) of indica rice with an incomplete cross of 4x8 parents. A genetic model which can dissect the effects of triploid seed, the cytoplasm, and the maternal plant on the endosperm traits was used. The results indicated that peak temperature (T(p)), conclusion temperature (T(c)) and enthalpy (DeltaH) were controlled by three types of genetic effects: seed direct (endosperm) effects, cytoplasmic effects and maternal effects. No cytoplasmic effects for the onset temperature (T(o)), hardness and SV, and no maternal effects for cohesiveness were found. The additive variances ( V (A)+ V (Am)) were larger than the dominance variances ( V (D)+ V (Dm)) for all the traits except for T(c), which suggested that selection could be applied for the starch properties in early generations. The total narrow-sense heritability for each parameter was over 60%, indicating that selection advances were predictable in the early generations for these traits.

Microsatellites in starch-synthesizing genes in relation to starch physicochemical properties in waxy rice ( Oryza sativa L.).

Rice starch is composed of amylose and amylopectin. Amylose content, an important determinant of rice starch quality, is primarily controlled by the waxy gene, encoding granule-bound starch synthase (GBSS). The starch branching enzyme (SBE) and soluble starch synthase (SSS) play major roles in the synthesis of amylopectin. Microsatellite polymorphisms in the three genes, the wx gene encoding granule-bound starch synthase I, the SBE gene encoding starch branching enzyme I and the SSS gene encoding soluble starch synthase I, were studied for 56 accessions of waxy rice ( Oryza sativa L.). Four (CT)(n) microsatellite alleles, (CT)(16), (CT)(17), (CT)(18) and (CT)(19), at the wx locus were detected in this set of waxy rice, of which (CT)(17) was the most frequent. Three (CT)(n) microsatellite allele classes were found at the SBE locus, (CT)(8) or (CT)(10) together with an insertion sequence of CTCTCGGGCGA, and (CT)(8) alone without the insertion. There were multiple microsatellites clustered at the SSS locus. However, these alleles can also be grouped into three classes, i.e. the allele class SSS-A = (AC)(2) em leader TCC(TC)(11) em leader (TC)(5)C(ACC)(11), the allele class SSS-B = (AC)(3) em leader TCT(TC)(6) em leader (TC)(4)C(ACC)(9), and the allele class SSS-C = (AC)(3) em leader TCT(TC)(6) em leader (TC)(4)C(ACC)(8). The analyses of starch physicochemical properties among different microsatellite genotypes indicated that the waxy rice group with the (CT)(19) allele, the SBE-A allele and the SSS-B allele was quite different from other groups. Nine out of 15 accessions with a high gelatinization temperature (GT) belonged to the wx (CT)(19) group, all of them belonged to the SBE-A group and 13 of them belonged to the SSS-B group. These microsatellites might be useful in marker-assisted breeding for the improvement of rice grain quality.

Chemical stability and colorant properties of betaxanthin pigments from Celosia argentea.

The chemical stability and colorant properties of three betaxanthins recently identified from Celosia argentea varieties were evaluated. Lyophilized betaxanthin powders from yellow inflorescences of Celosia exhibited bright yellow color and high color purity with strong hygroscopicity. The aqueous solutions containing these betaxanthins were bright yellow in the pH range 2.2-7.0, and they were most stable at pH 5.5. The betaxanthins in a model system (buffer) were susceptible to heat, and found to be as unstable as red betacyanins (betanin and amaranthine) at high temperatures (>40 degrees C), but more stable at 40 degrees C with the exclusion of light and air. The three betaxanthins had slightly higher pigment retention than amaranthine/isoamaranthine in crude extracts at 22 degrees C, as verified by HPLC analysis. Lyophilized betaxanthins had much better storage stability (mean 95.0% pigment retention) than corresponding aqueous solutions (14.8%) at 22 degrees C after 20 weeks. Refrigeration (4 degrees C) significantly increased pigment retention of aqueous betaxanthins to 75.5%.

Betalains of Celosia argentea.

The betalains of yellow, orange and red inflorescences of common cockscomb (Celosia argentea var. cristata) were compared and proved to be qualitatively identical to those of feathered amaranth (Celosia argentea var. plumosa). In addition to the known compounds amaranthin and betalamic acid, the structures of three yellow pigments were elucidated to be immonium conjugates of betalamic acid with dopamine, 3-methoxytyramine and (S)-tryptophan by various spectroscopic techniques and comparison to synthesized reference compounds; the latter two are new to plants. Among the betacyanins occurring in yellow inflorescences in trace amounts, the presence of 2-descarboxy-betanidin, a dopamine-derived betacyanin, has been ascertained. The detection of high dopamine concentration may be of toxicological relevance in use of yellow inflorescences as a vegetable and in traditional Chinese medicine, common uses for the red inflorescences of common cockscomb.

Development of NMR and raman spectroscopic methods for the determination of the degree of substitution of maleate in modified starches.

In this paper we report the application of NMR spectroscopy and Raman spectroscopy to determine the degree of maleate substitution in maleinated starches. Five kinds of maleinated starches were investigated and calibration sets were constructed to derive linear regression equations that may be used to predict the degree of maleate substitution for starch samples with unknown amounts of chemical modification. The calibration sets reported have very high linearity (r > 0.99) for both the NMR and Raman methods. The NMR and Raman calibration sets allow fast and nondestructive measurement of the degree of maleate substitution for different starches with little need of sample preparation.

Identification and distribution of simple and acylated betacyanins in the Amaranthaceae.

Red-colored plants in the family Amaranthaceae are recognized as a rich source of diverse and unique betacyanins. The distribution of betacyanins in 37 species of 8 genera in the Amaranthaceae was investigated. A total of 16 kinds of betacyanins were isolated and characterized by HPLC, spectral analyses, and MS. They consisted of 6 simple (nonacylated) betacyanins and 10 acylated betacyanins, including 8 amaranthine-type pigments, 6 gomphrenin-type pigments, and 2 betanin-type pigments. Acylated betacyanins were identified as betanidin 5-O-beta-glucuronosylglucoside or betanidin 6-O-beta-glucoside acylated with ferulic, p-coumaric, or 3-hydroxy-3-methylglutaric acids. Total betacyanin content in the 37 species ranged from 0.08 to 1.36 mg/g of fresh weight. Simple betacyanins (such as amaranthine, which averaged 91.5% of total peak area) were widespread among all species of 8 genera. Acylated betacyanins were distributed among 11 species of 6 genera, with the highest proportion occurring in Iresine herbstii (79.6%) and Gomphrena globosa (68.4%). Some cultivated species contained many more acylated betacyanins than wild species, representing a potential new source of these pigments as natural colorants.

Determination of the degree of succinylation in diverse modified starches by raman spectroscopy.

A method using Raman spectroscopy was recently developed for the determination of the degree of substitution of succinate in waxy maize starch. In this paper it is demonstrated that the method can be generalized to a wide range of starches of different amylose contents and botanical origins. Raman calibration sets were used to form regression equations for five types of succinylated starches, that is, waxy, regular, and two high-amylose maize samples (47 and 66% amylose, respectively) and wheat. The derived calibration curves can be used to find the degree of substitution in samples with unknown levels of succinylation. The Raman calibration lines had linear correlation coefficients of 0.995 or better and enable the fast and nondestructive determination of the degree of substitution of succinate for different types of starches with minimal sample preparation. Also discussed is the potential utility of Raman spectroscopy to simultaneously determine the degree of substitution of succinate and amylose content, using previously determined calibration curves developed for the amylose content of maize starches.

Effect of beta-cyclodextrin on pasting properties of wheat starch.

Changes of viscosity characteristics of genetically diverse hexaploid wheat starches during pasting in water, 1% NaCl solution, or at pH 4 or pH 10 were studied using a Rapid Visco-Analyzer. Peak viscosity (PV), hot paste viscosity (HPV) and cool paste viscosity (CPV) of all the wheat starches was little affected in pH 4 and pH 10 treatments. In 1% NaCl, all starches showed substantial increases in all three parameters relative to pasting in water. The use of 1% beta-cyclodextrin (beta-CD) (cycloheptaamylose) solution increased PV of high-swelling starches, but generally slightly decreased that of low swelling starches in all treatment conditions. HPV was always reduced by addition of 1% beta-CD, but CPV was increased in most treatments for Anza and Yecora Rojo (low swelling), but decreased for Klasic (high swelling). Bacterial alpha-amylase was added to starch or flour. The effect of beta-CD was shown to be independent of alpha-amylase inhibition in wheat starch, but beta-CD strongly inhibited alpha-amylase in wheat flour.

Genetic variation in the physical properties of sweet potato starch.

Sweet potato starch, prepared from 44 genotypes adapted to Philippine conditions, showed wide variation and distinctly different pasting profiles in Rapid Visco-Analyzer (RVA) analysis at 11% and 7% starch concentration. At 11% starch concentration, the pasting profiles were type A, characterized by high to moderate peak with a major breakdown and low cold paste viscosity. At 7%, the pasting profile was generally type C, characterized by the absence of a distinct peak with none to very slight breakdown and high cold paste viscosity. However, differentiation among genotypes was better achieved from RVA pasting profiles at 11% starch concentration. Peak viscosity (PV) and hot paste viscosity (HPV) at 11% starch paste concentration had significant negative correlation with amylose content. PV, HPV, and setback ratio were significantly correlated to adhesiveness of the starch gel. Sweet potato starch generally had high swelling volume but low solubilities at 92.5 degrees C.

Physicochemical properties of maize starches expressing dull and sugary-2 mutants in different genetic backgrounds.

Isogenic lines carrying the du or su(2) genes in five different maize inbred lines were developed, and the effect of the starch-modifying genes on general, thermal, pasting, and gel textural properties of starch was studied. Swelling power of du and su(2) starches was significantly lower than that of normal starch. The peak viscosity of du starches was reduced, peaks were sharper, and the setback was lower than in normal starches. The hardness of du starch gels increased and the adhesiveness decreased severalfold during 7 days of storage, indicating unstable gel texture. The su(2)/su(2) starches had flatter viscosity peaks than normal starches, with extremely low viscosity and almost zero breakdown and setback. In the su(2) starch gels hardness and adhesiveness did not change significantly during 7 days of storage. For most measured properties, there was considerable variation among different inbred backgrounds carrying the same mutant genotypes. Genetic background and starch-modifying genes can both be manipulated to select specific desired starch properties.

Accurate estimation of sweetpotato amylase activity by flour viscosity analysis.

Sweetpotato flour (SPF), prepared from 44 genotypes adapted to Philippine conditions, showed wide variation in Rapid Visco-Analyzer (RVA) pasting characteristics due to its variation in composition and endogenous amylase activity. The RVA pasting parameters of peak viscosity determined in water (PV1) and that determined in 0.05 mM AgNO(3) (used as an amylase inhibitor) (PV2) were successfully used to estimate alpha-amylase activity. The correlation of the ratio (PV2-PV1)/PV1 to alpha-amylase activity was 0.96 (p < 0.01, N = 44). Swelling volume measurements were not found to be suitable for prediction of alpha-amylase activity.

Functional properties and enzymatic digestibility of cationic and cross-linked cationic ae, wx, and normal maize starch.

The functional properties and enzymatic digestibility of cationic and cross-linked cationic ae, wx, and normal maize starches were studied. Cationization reduced the endothermic transition temperatures (T(o), T(p), and T(c)), however, it increased peak viscosity, swelling power, solubility, clarity, and digestibility of all the starches compared to the corresponding native starch. After cationization, the enthalpy of waxy and normal starches was little changed but ae starch showed a decrease. For gel texture, cationization increased the hardness, adhesiveness, and springiness of all the starches, except for the hardness and adhesiveness of normal starch which showed a decrease, and the springiness of waxy starch did not show much change compared to the corresponding control starch. Cross-linking of cationic starch increased the endothermic transition temperatures, as well as peak viscosity. However, it reduced the swelling power and solubility, clarity, and enzymatic digestibility of all the cationic starches.

Time-dependent changes in dough color in hexaploid wheat landraces differing in polyphenol oxidase activity.

Time-dependent changes in the color of noodle sheets (using 2% NaCl or 1% alkaline salts in the formulation) made from 43 Iranian hexaploid wheat landrace accessions were measured. Pekar slick tests in water and in alkaline conditions were also carried out. A wide variation in color characteristics of the landraces was found, with L values of salted noodle sheets at 2 h ranging from 80.9 to 89.2 and b values of alkaline noodle sheets at 2 h ranging from 19.1 to 27.4, showing potential application in noodle wheat improvement programs. For initial rapid screening of samples it was observed that a single reading of the dough sheet after 2 h was adequate. The dough sheets should be kept at 5 degrees C during storage, to prevent microbiological activity in the dough, which would give erroneous results. The Pekar slick test results were not highly correlated to color measurements on the dough, so this test is not recommended for screening for noodle color potential in landraces.

Amaranthus and buckwheat protein concentrate effects on an emulsion-type meat product.

The utilization of Amaranthus (five genotypes) and buckwheat protein concentrates in an emulsion-type meat product comprising beef lean, pork fat, salt and water was studied. 15% of the beef protein was replaced with the protein concentrates and the resulting meat emulsions were evaluated by thermorheology and thermal analysis. The cooking loss and physical properties of the meat gel were determined. The use of Amaranthus and buckwheat protein concentrates considerably affected both the emulsion and the cooked meat gel properties. The most favorable outcome was obtained with the buckwheat protein, which had similar effects to soy proteins. The Amaranthus protein concentrates generally did not give favorable results, although that derived from genotype K112 showed some positive effects. Correlation analysis showed that most of the observed variation in meat product properties could be explained by the emulsifying activity of the protein additive used.

Population genetics of colonizing success of weedy rye in Northern California.

Genetic parameters of 11 weedy rye populations located in California's northern mountain area and the adjoining Oregon border were compared with those of the putative parents, wild species Secale montamim and cultivated rye S. cereale. All weedy populations exhibited high levels of genetic variation as determined by isozyme analysis. On average, 44% of the isozyme loci were polymorphic, total genetic diversity was 0.30; and number of alleles per locus was 1.65. High genetic identities, averaging 0.994 ± 0.005 between populations, indicated that little genetic differentiation has occurred among these weedy populations since the initial colonization. Lack of population differentiation could be attributed to a wind-pollinated, self-incompatible breeding system resulting in extensive gene flow among weedy populations, and between weedy populations and local cultivars of rye. Multilocus outcrossing rates of weedy populations ranged from 0.86 to 0.97. The estimated levels of gene flow using the private-alleles method were high among weedy populations, and between cv 'Merced' and weedy populations, with estimated Nm values of 14.50 and 8.21, respectively. The colonizing success of weedy rye is discussed and a strategy for its conservation recommended.

Effect of nitrogen nutrition on endosperm protein synthesis in wild and cultivated barley grown in spike culture.

In normal growth conditions, total protein percent (salt soluble plus hordein fractions) in the endosperm at maturity in barley cultivar Hordeum vulgare L. cv ;Ruth' was about 14%, whereas in an accession of wild barley, Hordeum spontaneum Koch line 297, it was about 28%. Spike culture experiments were conducted to ascertain whether there were basic differences between the two genotypes under conditions of widely different nitrogen supply. Spikes of each genotype were grown from 8 to 25 days after flowering in in vitro culture in a growth medium containing 0 to 4 grams per liter nitrogen supplied as NH(4)NO(3). Spikes were pulse-labeled at intervals from 12 to 24 days after flowering with 3.7 megabecquerel of [(3)H]leucine to determine relative rates of synthesis of hordein-1 and hordein-2 polypeptides. At low nitrogen levels ;Ruth' had a lower protein content than 297, but at increasing nitrogen levels its protein content increased rapidly and reached a maximum (35%) higher than 297 (30%). The relative contribution of the hordein fraction to total protein increased mainly with time, and hordein-1 to total hordein increased mainly with nitrogen level, in both genotypes. There appeared to be no fundamental limitations in the capacity of ;Ruth' to accumulate protein; 297 appears to have a greater basal level of nitrogen availability under normal conditions.