Unraveling the significance of epithelial-associated bacteria in gastrointestinal diseases: Importance of choosing an optimal DNA extraction method

Understanding the significance of epithelial-associated bacteria in gastrointestinal diseases is essential for gaining insights into the complex host-microbiota interactions that influence disease development and progression. However, sequencing approaches face limitations due to the overwhelming presence of host DNA in the samples. PCR-based approaches like 16S rRNA gene amplicon sequencing enables taxonomic profiling and has been studied extensively in connection with a wide range of diseases and while still being a valuable method, microbiome research is moving towards metagenomic sequencing. This allows for the deciphering of both the bacterial community structure at a higher taxonomic resolution as well as insight into its functional potential. Here, we present a comparative study of three different DNA extraction methods for human colon biopsies: two commercially available kits (Qiagen Blood and tissue kit and Molzym ultra-deep microbiome prep) and one published optimized method (Saponin approach). The three methods were evaluated in terms of the ratio between host and bacterial DNA and their ability to retain the relative bacterial abundance at different taxonomic levels. Six colon biopsies (18 technical replicates) were sequenced with 16S rRNA gene amplicon and metagenomic sequencing resulting in distinct bacterial profiles dependent on the applied extraction method. The Saponin approach showed depletion of both host and bacterial DNA to an extent that the bacterial community structure was not retained when looking at 16S rRNA sequencing data. Furthermore, only the Molzym kit showed a satisfying sequencing depth suggesting that although Qiagens kit retained the community structure better than the Saponin approach, the vast amount of host DNA hampered the sequencing effort even after producing amplicons. When employing metagenomic sequencing to evaluate the bacterial:host DNA ratio, the Molzym kit showed up to a 10-fold enrichment of bacterial DNA compared to the Qiagen kit. Selecting an appropriate DNA extraction method is vital when studying epithelial-associated bacteria in gastrointestinal diseases and is essential for unravelling the intricate host-microbiota interactions underlying disease pathogenesis

Effect of Enzymatic Treatment of Different Starch Sources on the in Vitro Rate and Extent of Starch Digestion

Gelatinized wheat, potato and waxy maize starches were treated enzymatically in order to increase the degree of branching of the amylopectin fraction and thereby change the starch degradation profile towards a higher proportion of slowly digestible starch (SDS). The materials were characterized by single-pulse 1H HR-MAS NMR spectroscopy and in vitro digestion profile according to the Englyst procedure. Using various concentrations and incubation times with branching enzyme (EC 2.4.1.18) without or with additional treatment with the hydrolytic enzymes; β-amylase (EC 3.2.1.2), α-glucosidase (EC 3.2.1.20), or amyloglucosidase (EC 3.2.1.3) the proportion of α-(1–6) linkages was increased by up to a factor of 4.1, 5 and 5.8 in waxy maize, wheat and potato starches, respectively. The proportion of SDS was significantly increased when using hydrolytic enzymes after treatment with branching enzyme but it was only for waxy maize that the proportion of α-(1–6) bonds and the in vitro digestion profile was significantly correlated

Obesity Development and Signs of Metabolic Abnormalities in Young Göttingen Minipigs Consuming Energy Dense Diets Varying in Carbohydrate Quality

Consumption of fructose has been associated with a higher risk of developing obesity and metabolic syndrome (MetS). The aim of this study was to examine the long-term effects of fructose compared to starch from high-amylose maize starch (HiMaize) at ad libitum feeding in a juvenile Göttingen Minipig model with 20% of the diet provided as fructose as a high-risk diet (HR, n = 15) and 20% as HiMaize as a lower-risk control diet (LR, n = 15). The intake of metabolizable energy was on average similar (p = 0.11) among diets despite increased levels of the satiety hormone PYY measured in plasma (p = 0.0005) of the LR pigs. However, after over 20 weeks of ad libitum feeding, no difference between diets was observed in daily weight gain (p = 0.103), and a difference in BW was observed only at the end of the experiment. The ad libitum feeding promoted an obese phenotype over time in both groups with increased plasma levels of glucose (p = 0.005), fructosamine (p < 0.001), insulin (p = 0.03), and HOMA-IR (p = 0.02), whereas the clinical markers of dyslipidemia were unaffected. When compared to the LR diet, fructose did not accelerate the progression of MetS associated parameters and largely failed to change markers that indicate a stimulated de novo lipogenesis

Kasprzak, M.M., et al., Correction: Effect of Enzymatic Treatment of Different Starch Sources on the in Vitro Rate and Extent of Starch Digestion. Int. J. Mol. Sci. 2012, 13, 929-942.

The authors wish to change the description of preparation of samples at Experimental Section on their paper published in IJMS [1]. [...

Distinct Difference in Absorption Pattern in Pigs of Betaine Provided as a Supplement or Present Naturally in Cereal Dietary Fiber

The net absorption of betaine and choline was determined for 4 h after the first meal of the day in three experiments with porto-arterial catheterized pigs in which betaine was added as a supplement to a low-betaine diet (<i>n</i> = 4 pigs) and compared to the net absorption of betaine and choline from high-fiber breads differing in amount and source of dietary fiber (two experiments, <i>n</i> = 6 pigs each). Plasma betaine peaked after 30 min when betaine was fed as a supplement, whereas it peaked after 120–180 min when high-fiber breads were fed. Plasma betaine showed no diet × time interaction after feeding with high-fiber breads, indicating that the absorption kinetic did not differ between fiber sources. The net absorption of choline was not affected by the experimental diets. In conclusion, betaine in cereal sources has to be liberated from the matrix prior to absorption, causing delayed absorption