Anaerobic Ammonium Oxidation in Acidic Red Soils

Anaerobic ammonium oxidation (anammox) has been proven to be an important nitrogen removal process in terrestrial ecosystems, particularly paddy soils. However, the contribution of anammox in acidic red soils to nitrogen loss has not been well-documented to date. Here, we investigated the activity, abundance, and distribution of anammox bacteria in red soils collected from nine provinces of Southern China. High-throughput sequencing analysis showed that Candidatus Brocadia dominates the anammox bacterial community (93.03% of sequence reads). Quantification of the hydrazine synthase gene (hzsB) and anammox 16S rRNA gene indicated that the abundance of anammox bacteria ranged from 6.20 × 106 to 1.81 × 109 and 4.81 × 106 to 4.54 × 108 copies per gram of dry weight, respectively. Contributions to nitrogen removal by anammox were measured by a 15N isotope-pairing assay. Anammox rates in red soil ranged from 0.01 to 0.59 nmol N g−1 h−1, contributing 16.67–53.27% to N2 production in the studied area, and the total amount of removed nitrogen by anammox was estimated at 2.33 Tg N per year in the natural red soils of southern China. Pearson correlation analyses revealed that the distribution of anammox bacteria significantly correlated with the concentration of nitrate and pH, whereas the abundance and activity of anammox bacteria were significantly influenced by the nitrate and total nitrogen concentrations. Our findings demonstrate that Candidatus Brocadia dominates anammox bacterial communities in acidic red soils and plays an important role in nitrogen loss of the red soil in Southern China

Effects of KN-42 on Growth Performance, Diarrhea and Faecal Bacterial Flora of Weaned Piglets

This research focused on the effects of different doses of Bacillus subtilis KN-42 on the growth performance, diarrhea incidence, faecal bacterial flora, and the relative number of Lactobacillus and Escherichia coli in faeces of weaned piglets to determine whether the strain can serve as a candidate antimicrobial growth promoter. A total of 360 piglets (initial body weight 7.14±0.63 kg) weaned at 26±2 days of age were randomly allotted to 5 treatment groups (4 pens per treatment with 18 pigs per pen) for a 28-day trial. Dietary treatments were basal diet without any antimicrobial (negative control; NC), basal diet supplemented with 120 mg/kg feed of neomycin sulfate (positive control; PC) and basal diet supplemented with 2×109 (L), 4×109 (M) and 20×109 (H) CFU/kg feed of B. subtilis KN-42. During the overall period, average daily gain and feed efficiency of piglets were higher in groups PC, M, and H than those in group NC (p<0.05), and all probiotics and antibiotics groups had a lower diarrhea index than group NC (p<0.05). The 16S rDNA gene-based methods were used to analyze faecal bacterial flora on day 28 of experiment. The result of denaturing gradient gel electrophoresis analysis showed that supplementation of B. subtilis KN-42 to the diet changed the bacterial communities, with a higher bacterial diversity and band number in group M than in the other four groups. Real-time polymerase chain reaction analysis showed that the relative number of Lactobacillus were higher in groups PC and H than in group NC (p<0.05), and the supplemented B. subtilis KN-42 to the diet also reduced the relative number of E. coli (p<0.05). These results suggest that dietary addition of B. subtilis KN-42 can improve the growth performance and gastrointestinal health of piglets

Optimization of Saccharomyces boulardii production in solid-state fermentation with response surface methodology

Saccharomyces boulardii preparations are promising probiotics and clinical agents for animals and humans. This work focused on optimizing the nutritional conditions for the production of S. boulardii in solid-state fermentation by using classical and statistical methods. In single-factor experiments, the S. boulardii production was significantly increased by the addition of glucoamylase and the optimal carbon and nitrogen sources were found to be soluble starch and NH4Cl, respectively. The effects of the glucoamylase, soluble starch and NH4Cl on S. boulardii production were evaluated by a three-level three-factor Box–Behnken design and response surface methodology (RSM). The maximal yeast count (4.50 ×109CFU/g) was obtained under the optimized conditions (198 U/g glucoamylase, 2.37% soluble starch and 0.9% NH4Cl), which was in a good agreement with the predicted value of the model. This study has provided useful information on how to improve the accumulation of yeast cells by RSM

Bacterial diversity index calculated from the DGGE banding patterns (Fig. 1A).

<p>N (negative control, basal diet); P (positive control, diet supplemented with neomycin); L, M, H (diets supplemented with probiotics 0.5×10⁹, 1.0×10⁹ and 2.5×10⁹ CFU/kg feed, respectively);</p><p>*1/D, the reciprocal of Simpson diversity index.</p><p>Bacterial diversity index calculated from the DGGE banding patterns (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0116635#pone.0116635.g001" target="_blank">Fig. 1A</a>).</p

Dietary <i>Enterococcus faecalis</i> LAB31 Improves Growth Performance, Reduces Diarrhea, and Increases Fecal <i>Lactobacillus</i> Number of Weaned Piglets

<div><p>Lactic acid bacteria (LAB) have been shown to enhance performance of weaned piglets. However, few studies have reported the addition of LAB <i>Enterococcus faecalis</i> as alternatives to growth promoting antibiotics for weaned piglets. This study evaluated the effects of dietary <i>E. faecalis</i> LAB31 on the growth performance, diarrhea incidence, blood parameters, fecal bacterial and <i>Lactobacillus</i> communities in weaned piglets. A total of 360 piglets weaned at 26 ± 2 days of age were randomly allotted to 5 groups (20 pens, with 4 pens for each group) for a trial of 28 days: group N (negative control, without antibiotics or probiotics); group P (Neomycin sulfate, 100 mg/kg feed); groups L, M and H (supplemented with <i>E. faecalis</i> LAB31 0.5×10⁹, 1.0×10⁹, and 2.5×10⁹ CFU/kg feed, respectively). Average daily gain and feed conversion efficiency were found to be higher in group H than in group N, and showed significant differences between group H and group P (<i>P₀</i> < 0.05). Furthermore, groups H and P had a lower diarrhea index than the other three groups (<i>P₀</i> < 0.05). Denaturing gradient gel electrophoresis (DGGE) showed that the application of probiotics to the diet changed the bacterial community, with a higher bacterial diversity in group M than in the other four groups. Real-time PCR revealed that the relative number of <i>Lactobacillus</i> increased by addition of probiotics, and was higher in group H than in group N (<i>P₀</i> < 0.05). However, group-specific PCR-DGGE showed no obvious difference among the five groups in <i>Lactobacillus</i> composition and diversity. Therefore, the dietary addition of <i>E. faecalis</i> LAB31 can improve growth performance, reduce diarrhea, and increase the relative number of <i>Lactobacillus</i> in feces of weaned piglets.</p></div

<i>Lactobacillus</i> diversity index calculated from the DGGE banding patterns (Fig. 2A).

<p>N (negative control, basal diet); P (positive control, diet supplemented with neomycin); L, M, H (diets supplemented with probiotics 0.5×10⁹, 1.0×10⁹ and 2.5×10⁹ CFU/kg feed, respectively);</p><p>*1/D, the reciprocal of Simpson diversity index.</p><p><i>Lactobacillus</i> diversity index calculated from the DGGE banding patterns (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0116635#pone.0116635.g002" target="_blank">Fig. 2A</a>).</p

Bacterial community of weaned piglets fed with neomycin or <i>E. faecalis</i>.

<p>(A) DGGE profiles of the V6~V8 regions of the 16S rDNA gene fragments from the samples. The denaturant gradient range is from 42% to 58%. The major difference bands are numbered. Lane S (Standard ladder, which are PCR products generated from different bacterial 16S rDNA genes with primers 968F-GC and 1401R); N (negative control, basal diet); P (positive control, diet supplemented with neomycin); L, M, H (diets supplemented with probiotics 0.5×10⁹, 1.0×10⁹ and 2.5×10⁹ CFU/kg feed, respectively); (B) UPGMA cluster analysis of Dice similarity indices from DGGE profiles.</p

Identification of band fragments in DGGE gels (Fig. 1A).

<p>* Bands are numbered according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0116635#pone.0116635.g001" target="_blank">Fig. 1A</a>.</p><p>^◆Identity represents the sequence identity (%) compared with that in the GenBank database.</p><p>Identification of band fragments in DGGE gels (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0116635#pone.0116635.g001" target="_blank">Fig. 1A</a>).</p