Triticum aestivum L.)/faba bean (Vicia faba L.) intercropping shows significant overyielding and high nitrogen (N)-use efficiency, but the dynamics of plant interactions have rarely been estimated. The objective of the present study was to investigate the temporal dynamics of competitive N acquisition between intercropped wheat and faba bean with the logistic model. Wheat and faba bean were grown together or alone with limited N supply in pots. Data of shoot and root biomass and N content measured from 14 samplings were fitted to logistic models to determine instantaneous rates of growth and N uptake. The superiority of instantaneous biomass production and N uptake shifted from faba bean to wheat with their growth. Moreover, the shift of superiority on N uptake occurred 7–12 days earlier than that of biomass production. Interspecific competition stimulated intercropped wheat to have a much earlier and stronger superiority on instantaneous N uptake compared with isolated wheat. The modeling methodology characterized the temporal dynamics of biomass production and N uptake of intercropped wheat and faba bean in different planting systems, which helps to understand the underlying process of plant interaction for intercropping plants.Wheat ( Vicia faba L.) significantly facilitated phosphorus (P) uptake of neighbouring cereals by mobilization of sparsely soluble soil P Sorghum bicolor (L.) Moench)/pigeonpea (Cajanus cajan (L.) Millsp) and millet (Pennisetum glaucum (L.) R.Br.)/groundnut (Arachis hypogaea L.) intercropping systems showed temporal and spatial complementarity for light, water and nutrients In China, intercropping has been practiced for over 2000 years, and has made a great contribution to food security of northwest China, e.g. close to 43% of the total gain yield of Ningxia province was from intercropping in 1995 2 fixation of neighbuoring legumes Previous studies of intercropping focusing on belowground interactions have shown significant intercropping advantages of nutrient uptake Triticum aestivum L.)/faba bean intercropping system is widely used by famers, especially in south China, in which two plant species have the same sowing and harvesting time. The objective of this study was to assess the temporal dynamics of competitive N acquisition between intercropped wheat and faba bean by using a logistic model, and then understand the temporal dynamics of intra- and interspecific competition between different plant species.Due to high potential for overyielding, wheat  of 18.3 mg kg−1: nitrate N of 15.1 mg kg−1 and ammonium N of 3.2 mg kg−1, Olsen-P of 6.7 mg kg−1, NH4Ac-exchangeable K of 74.1 mg kg−1, and a pH in CaCl2 of 7.8 (the ratio of soil to CaCl2 solution is 1∶2.5). Basal nutrients were added in a solution to soil (mg kg−1 soil): N, 100 as Ca(NO3)2, P, 150 as KH2PO4, K, 188.7 as KH2PO4. The added nutrients were thoroughly mixed with the soil by shaking. There were no other micronutrients added and no any deficiency symptoms observed on plants at each sampling.A pot experiment was carried out in a glasshouse at China Agricultural University, Beijing, China. Air temperatures ranged from18–28°C. Evaporative cooling and shade cloth were used to prevent excessively high temperatures on sunny days. The soil used in the experiments was loam, and collected from fallow land of Shangzhuang Experimental Station, Beijing, China. Our collection was permitted by the Center for Resources, Environment and Food Security (CREFS), China Agricultural University. Air-dried soil was sieved (2 mm). Each pot (210×170 mm) was filled with 2 kg soil. The soil had an organic carbon concentration of 10.1 g kgTriticum aestivum L cv. Yunmai-42) and two seeds of faba bean (Vicia faba L. cv. 89–147) were grown separately or together in one pot. Plants were thinned to three per pot for wheat and one for faba bean based on plant density in field at 11 days after sowing (DAS) (Six germinated seeds of wheat (ng (DAS) . Another2SO4 and H2O2 using the micro-Kjeldahl procedure There were 14 destructive samplings taken every 3–5 days from 15 to 62 DAS when the wheat has been grain-filling and faba bean has been pod-filling. At each sampling, shoots were removed by cutting at the soil surface and roots were removed from the soil by washing them carefully. When washing the roots, deionized water and sieve with 1 mm aperture were used. All sampled material was oven-dried at 76°C for 72 h before weighing. Nitrogen concentration of plant material was determined by wet digestion with a mixture of concentrated HY; mg) of one herbaceous plant grown from seed for several weeks is typically a sigmoid function of time, tΔt, Y was computed using a discrete two-parameter form of the logistic equation as following r, the rate constant (d−1) for biomass production; Ymax, a maximum to which Y tends asymptotically) r and Ymax to maximize 2R with the SOLVER in Microsoft Excel 2010 The biomass −1, whichards see .c. 28 DAS, and for roots content at c. 31 DAS  for total plant biomass at 50 DAS which was delayed by 5 or 9 days compared with monocropped and intercropped wheat, respectively  much earlier than the corresponding brmax in total plant, shoot and root biomass, regardless of cropping designs  in the present study. It indicated that the available N was almost exhausted by uptake of intercropped plants which can be confirmed by low Nmin around 3–7 mg kg−1 in soil and near-zero instantaneous N uptake rate of plants in intercropping system after c. 41 DAS. Thus, N demanded by plants growth in intercropping was mainly from re-utilization of plant N after c. 41 DAS in the present study.The logistic model visualized competitive interaction in monocropping and intercropping. There was no competition for isolated plants, but intraspecific and interspecific competition arose with plant growth for monocropped and intercropped plants in the present study. The previous studies of intercropping usually focused on interspecific interactions, and ignored intraspecific interactions The changing trends of instantaneous growth rate and N uptake appeared like a dome for both faba bean and wheat which is consistent with previous studies Based on fundamental characteristics of N uptake that different plant species reached maximum instantaneous N uptake at different times, isolated faba bean and wheat showed slightly temporal niche separation, which allowed them to acquire available resources at separated times c. 41DAS in intercropping system. Intra-competition strongly reduced instantaneous rate of biomass production and N uptake of individual wheat plants in the monocropping system which was possibly due to suppression of root growth before c. 41 DAS. Nitrogen accumulation of wheat had a positive correlation with root length −1 at final harvest in the present study.The trajectories of biomass production and N uptake fully described the scale and nature of neighbour interaction, and showed that interaction strongly depended on sampling time. This trajectory analysis overcame the shortage of traditional experiments, with only one or two sampling times. Interactions between faba bean and wheat modified the dynamics of biomass production and N uptake compared with corresponding dynamics of isolated plants. As mentioned above, soil available N was close to be exhausted at Faba bean had superiority on biomass production and N uptake compared with wheat only at early stages in the intercropping system that resulted from fundamental characteristics of plant species which is consistent with the results of intercropped wheat and common vetch The information emerging from the present experiment shows that we can expect a temporal change of resource-capture dynamics if additional nutrients or other resources are considered at the same time. Moreover, it is necessary to extend this approach described above to explore competitive dynamics over a larger fraction of intercropped crops' long-term growth period in field. There is evidence that greater overyielding obtained through N fertilization or row ratio The logistic model visualized dynamics of biomass production and N uptake of faba bean and wheat with or without neighbours. When intercropped faba bean and wheat competed for N, there was a change-point of superiority on both biomass production and N uptake of faba bean and wheat with limited N supply. Faba bean had faster instantaneous rate of growth and N uptake only at early growth stages, and then was overtaken by wheat. Competition for soil N significantly modified these dynamics, not only temporally but also in intensity. This modeling methodology will be helpful to understand the underlying dynamic process of interaction in intercropping systems, and will be practicable to reveal the dynamic interaction of competitive resources utilization between intercropped plants in field.S1 TableP values using ANONA on biomass of plants.(DOCX)Click here for additional data file.S2 TableP values using ANONA on N uptake of plants.(DOCX)Click here for additional data file.


The effects of synthetic goethite (α-FeOOH) used as the catalyst in catalytic ozonation for the degradation of disinfection by-product (DBP) precursors are investigated. A biofiltration column applied following the catalytic ozonation process is used to evaluate the efficiency of removing DBP precursors via biotreatment. Ozone can rapidly react with aromatic compounds and oxidize organic compounds, resulting in a decrease in the fluorescence intensity of dissolved organic matter (DOM). In addition, catalytic ozonation can break down large organic molecules, which causes a blue shift in the emission-excitation matrix spectra. Water treated with catalytic ozonation is composed of low-molecular structures, including soluble microbial products (SMPs) and other aromatic proteins (APs). The DOM in SMPs and APs is removed by subsequent biofiltration. Catalytic ozonation has a higher removal efficiency for dissolved organic carbon and higher ultraviolet absorbance at 254 nm compared to those of ozonation without a catalyst. The use of catalytic ozonation and subsequent biofiltration leads to a lower DBP formation potential during chlorination compared to that obtained using ozonation and catalytic ozonation alone. Regarding DBP species during chlorination, the bromine incorporation factor (BIF) of trihalomethanes and haloacetic acids increases with increasing catalyst dosage in catalytic ozonation. Moreover, the highest BIF is obtained for catalytic ozonation and subsequent biofiltration. Disinfection by-products (DBPs) are regulated in many countries, due to their genotoxicity, mutagenicity and carcinogenicity . TrihaloOzone is a strong oxidant and disinfectant and, thus, is widely used in drinking water treatment. The advantage of using ozone in drinking water treatment is that it can effectively eliminate color and taste, as well as remove DBP precursors (such as humic substances) in water . Ozonati2, TiO2, Al2O3 and FeOOH) and metals or metal oxides on metal oxide supports . They proposed three main mechanisms for the HCOP reaction: (i) direct ozone reacts with surface functional groups on the surface of catalysts, initiating the production of hydroxyl radicals (·OH); (ii) the organic molecules adsorb onto the surface of catalysts and then react with ozone/·OH; and (iii) both ozone and organic molecules are simultaneously adsorbed onto the catalyst surface, and there is a subsequent interaction between the chemisorbed species [−) will influence the species of THM and HAA during chlorination and might cause the formation of bromate after the ozonation treatment [et al. [The heterogeneous catalytic ozonation process (HCOP) has recently received a lot of attention in water treatment due to its high effectiveness in the degradation and mineralization of refractory organic pollutants ,12,13. N species ,11,12,13 species ,14. Howe species ,15. Anotreatment . The Br-reatment , the pro [et al.  demonstr3) and trichloroacetic acid (TCAA) were analyzed in their study. Hassan et al. [et al. [−1). High reduction (>85%) of THMs and nine HAAs (HAA9) was obtained through adsorption with 6 g·L−1 of iron-coated pumice. They also reported that the bromide concentration (up to 550 µg·L−1) did not have an impact on the DOC uptake. With the exception of adsorption on FeOOH, Park et al. [A previous study used goethite (FeOOH), which is a byproduct released from the corrosion of iron pipes, as a reductant and an adsorbent in drinking water treatment to investigate its influence on DBP formation . They con et al.  investig [et al.  used FeOk et al.  demonstrIn this study, goethite (α-FeOOH) is used as the catalyst in a fluidized-bed reactor (FBR) for the purpose of catalytic ozonation followed by biofiltration. This system is constructed to investigate the transformation of DOM via emission-excitation matrix (EEM) spectra, to evaluate the removal efficiency of dissolved organic carbon (DOC), ultraviolet absorbance at 254 nm (UV254) and specific ultraviolet absorbance (SUVA) and C-DBP (THMs and HAAs) formation potential during chlorination. The bromine impact factor (BIF) of C-DBPs and bromate concentrations are also investigated in order to study the formation and speciation of DBPs under various operating conditions and treatment processes.A catalytic ozonation FBR system in a semi-continuous mode and a biofiltration column were used in this study. A schematic diagram of the catalytic ozonation FBR system is shown in The water treated with the FBR system was further treated in a biofiltration column in recirculation mode for 3 days to evaluate the removal efficiency of DBP precursors using the biotreatment process. The biofiltration column from a previous study was adopted . Untreatet al. [3)3·9H2O, 0.25 M) was added to a sodium hydroxide solution ; the pH was adjusted to >12, and the solution was allowed to stand for 48 h. The mixture was filtered through a 0.45-µm filter  to collect the precipitates. The precipitates were then rinsed with distilled water until the effluent remained at a neutral pH. Then, the precipitate was dried at 120 °C for 24 h before being used in the experiments. The catalysts were characterized using scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS) on a Hitachi S-3000N system. The results are shown in 2·g−1, as determined by N2 physisorption at 77 K.α-FeOOH catalyst was synthesized following a method developed by Sui et al. . A solutWater samples were collected from WLR, Pingtung, Taiwan, which is polluted with domestic and agricultural wastewaters. Water samples were collected in 25-L polyethylene tanks and stored in a refrigerator at 4 °C for less than 7 days. Samples were warmed back to the experimental operating temperature (20 °C) before being used and were passed through a 0.45-µm filter  to remove coarse suspended and colloidal solids.−1. During the course of the fluorescence analysis, the Raman scattering peak intensity for Milli-Q water  was recorded as a standard to verify the instrument stability. Detailed information regarding EEM analysis is available elsewhere [The fluorescence spectra of the water samples were measured and recorded on a fluorophotometer  to determine the characteristics of the organic matter. The excitation-emission matrix (EEM) of each sample was generated by scanning at excitation (ex) wavelengths of 200 to 450 nm in 5-nm steps and by detecting the emitted fluorescence (em) between 250 and 550 nm in 2-nm steps. The scanning speed was 5000 nm·minlsewhere .The DOC was analyzed in accordance with Standard Method 5310B . A total−1 after 48 h of incubation at room temperature according to the procedures delineated in Standard Method 2350B and Standard Method 5710C [A stock solution of sodium hypochlorite  was used for chlorination. It was standardized according to Standard Method 4500B . The watod 5710C . Chlorinod 5710C .63Ni electron capture detector (ECD), an auto-sampler and a Supelco EquityTM-5 column (30 m × 0.25 mm ID). The instrument operating conditions were set according to our previous work [THMs and nine HAAs HAA9) were analyzed using a gas chromatograph (Agilent HP 6890N) outfitted with a  were ana3), bromodichloromethane (CHBrCl2), dibromochloromethane (CHBr2Cl) and bromoform (CHBr3), were monitored in this study. The THMs were extracted with hexane in accordance with Standard Method 6232B [Four THM compounds, namely chloroform , mono bromoacetic acid (MBAA), dichloroacetic acid (DCAA), bromochloroacetic acid (BCAA), trichloroacetic acid (TCAA), bromodichloroacetic acid (BDCAA), dibromoacetic acid (DBAA), chlorodibromoacetic acid (CDBAA) and tribromoacetic acid (TBAA), were quantified according to the United States Environmental Protection Agency (USPEA) Method 552.2 .2CO3, Merck) and 15 mM NaOH. A pre-column  was connected to an analytical column . All samples were filtered using prerinsed 0.45-µm nylon syringe filters  prior to analysis and were analyzed in triplicate. An injection volume of 100 µL was used. A calibration mixture containing bromide and bromate was prepared from granular ACS -grade reagents.Bromide and bromate samples were analyzed following USEPA Method 300.1  using ioThe EEM spectra of the DOM fractions before and after ozonation and catalytic ozonation with α-FeOOH are shown in i,n) [etc., which are likely to affect the molecular size distribution of DOM. Her et al. [As can be seen in i,n)  of the ii,n) . Therefoi,n) ,7,29,30.i,n) ,7,31. Thi,n) ,21,32. Tr et al.  indicateet al. [et al. [The fluorescence index (FI), which is defined as the ratio of emission intensity at 450 and 500 nm for emission spectra measured at a excitation wavelength of 370 nm in EEM spectra ,34, of t [et al.  observedet al. [i,n) for each EEM region was calculated based on the FRI technique. The results are shown in i,n represents the composition of DOM and its distribution in the five excitation-emission regions. As shown in i,n of water treated with biofiltration following either ozonation or catalytic ozonation. It can be seen that the percentage of simple APs (Region I) and SMPs (Region II) decreased compared to that for ozonation or catalytic ozonation alone  technique developed by Chen et al. , which con alone a. This i−1 are 12%, 24% and 43%, respectively. The removal efficiencies of UV254 for these dosages are 69%, 77% and 83%, respectively. The removal efficiencies for DOC and UV254 both increased with increases in the catalyst dosage. Moreover, the removal efficiency of UV254 was higher than that of DOC during catalytic ozonation. This result reveals that only some DOC was oxidized to carbon dioxide. According to the EEM spectra, the DOM in the WLR raw water is abundant in FAs (Region III). It has been recognized that aromatic groups, which are susceptible to electrophilic attack, are the major structure in fulvic acids. Thus, ozone molecules are very effective in breaking the aromatic structure and, thus, rapidly decrease UV254 absorbance. Compared with ozone, which reacts with organic matter through a selective reaction mechanism, ·OH is a stronger and nonselective oxidant that reacts with organic contaminants quickly and effectively. It was found that catalytic ozonation has a better removal efficiency of UV254 than that obtained using ozonation alone. The removal percentage of UV254 increased with increases in the catalyst dosage in catalytic ozonation. Catalytic ozonation also has a higher DOC removal efficiency in comparison with that obtained using ozonation alone. This result is attributed to the increased formation of ·OH under catalytic ozonation [The concentrations of water quality parameters and DBPs in different experimental conditions are summarized in zonation ,35.The water treated with ozonation and catalytic ozonation was further treated using biofiltration in recirculation mode for three days b. The reFrom −1·m−1, which implies that the DOM in the raw water mainly consisted of hydrophilic acids [et al. [The THM and HAA9 formation potential were measured for both the raw water and the water treated with catalytic ozonation and subsequent biofiltration. The experimental results are shown in ic acids ,38,39. Z [et al.  found th−1, the removal efficiencies of THM and HAA9 precursors significantly increased from 37% to 56% and from 35% to 59%, respectively. These results indicate that the DBPFP can be decreased by increasing the catalyst dosage in the catalytic ozonation process. Many researchers have reported that an increase in the catalyst concentration benefits the degradation of organic substances [bstances ,41. Thisbstances ,41. Ther−1 after biofiltration. This result implies that catalytic ozonation not only decreased the formation potential of THMs and HAA9, but also promoted their removal efficiency in the subsequent biofiltration. This observation is in agreement with the findings reported by other researchers, who used a combination of catalytic ozonation and biotreatment [The removal efficiency of DBP precursors for catalytic ozonation followed by biofiltration was also evaluated b. The rereatment ,44.p < 0.0005) exists between DBPs in treated water and catalyst dosages. The HCOP followed by biofiltration effectively lowered the concentration of THMs and HAA9. In the case of the concentrations of individual THM species in different experimental conditions  (Equation (2) [tion (2) ), which tion (2) ) over chtion (2) ).(2)Br−−/DOC and Br−/Cl2. However, chlorine can be quickly consumed by NOM, resulting in less chlorine available for chlorine-bromide reactions when water samples contain high DOC concentrations and low bromide concentrations. This can lead to a decrease in the brominated DBP formation. In contrast, the formation of brominated THMs and HAA9 in treated water will increase if water samples are treated using catalytic ozonation to eliminate the DOC before chlorination. The subsequent use of biofiltration increased the BIF values as compared to those for ozonation alone and catalytic ozonation alone. The BIF value and catalyst dosage were also found to be directly proportional. The BIF values for THMs and HAA9 were in the range of 0.30–0.42 and 1.60–1.89, respectively. In our previous study [Many researchers have demonstrated that many parameters can directly influence brominated DBP formation in chlorination, including pH and the ratios of Br−1) was detected in the WLR raw water. This low concentration of bromide is insufficient for the chemical reaction to form bromate during ozonation in the treated water, even though the rate constants for Br− and HOBr/OBr− oxidation  with ·OH are high [ et al. [−1) in raw water resulted in no significant bromate formation during ozonation.A trace concentration of bromide  and DBP formation potential (THMs and HAA9) were investigated. Catalytic ozonation can break down large organic molecules, change the DOM composition and produce SMPs and APs in the treated water. Subsequent biofiltration can preferentially remove these low molecular weight compounds. The removal efficiencies of DOC and UV254 increased with increases in the catalyst dosage in catalytic ozonation. When a biofiltration unit was installed downstream of the catalytic ozonation, better water quality was obtained in terms of the DOC level and UV254 value compared to those obtained using catalytic ozonation alone. Lower THM and HAA9 formation potentials were obtained by catalytic ozonation and subsequent biofiltration. In the case of the DBP species during chlorination, the BIF values for THM and HAA9 increased for catalytic ozonation and subsequent biofiltration compared to those of the catalytic ozonation. No bromate was detected in the treated water for any treatment process, due to the low bromine concentration in the WLR water.


Aspergillus oryzae GB-107 (FSBMA), SBM fermented by Bacillus subtilis PP6 (FSBMB), UV sterilized SBM fermented by Bacillus subtilis PP6 (UVFSBMB), SBM containing Bacillus subtilis PP6 (PSBM), and soy protein concentrate (SPC). Six corn-based diets were used and each of soybean products was added. All diets contained 5.0 g/kg of chromic oxide as an indigestible indicator and an N-free diet was used to measure basal endogenous losses of CP and AAs. Ileal CP digestibility did not differ by different soybean products. However, SIDs of Ile, Phe and Val were improved in pigs fed the FSBMB, UVFSBMB and SPC diets and the pigs fed the FSBMA diet showed higher SIDs of Phe and Val compared with those fed the CSBM diet (P < 0.05). The FSBMB diet had higher SIDs in most AAs compared with the FSBMA diet (P < 0.05), and higher SIDs of Lys, Ala, Pro, Ser, and Tyr compared with PSBM diet (P < 0.05). However, there was no response of UV-sterilization on the FSBMB in the SIDs of AAs. These results suggest that SIDs of AAs could be improved by the supplementation of fermented soybean products in the diet for weaning pigs but fermentation with Bacillus subtilis is more efficient in improving ileal AA digestibility than that with Aspergillus oryzae. Furthermore, probiotics supplementation in the CSBM and UV-sterilization of the FSBMB had no effects on chemical composition and ileal AA digestibility.An experiment was conducted to evaluate apparent (AID) and standardized (SID) ileal digestibilities of crude protein (CP) and amino acids (AA) with 6 soybean products in weaning pigs. A total of 14 weaning barrows with an initial body weight of 6.54 ± 0.34 kg were fitted with T-cannula at the distal ileum and allotted to 7 diets containing various soybean products. The soybean products used in the experiment were conventional soybean meal (CSBM), SBM fermented by  Soybean meal (SBM) is widely used in swine diets as a protein source, but contains various anti-nutritional factors (ANFs) such as trypsin inhibitor, oligosaccharides, antigenic factors and lectin. Anti-nutritional factors limit nutrient availability in weaning pigs ,2. Thus,Aspergillus oryzae or Bacillus subtilis could degrade ANFs, produce small size peptide compared to conventional SBM (CSBM), and modify amino acid (AA) profiles by microbial AA synthesis and breakdown, resulting in improvement of the AA digestibility  × Duroc; average body weight (BW) of 6.54 ± 0.34 kg} were equipped with simple T-cannula at distal ileum followed by Stein et al. [2) in a room of steady temperature (27°C), controlled with a heating lamp. During a 14-d recuperation period, all pigs were fed the commercial starter diet containing 230 g/kg CP and allowed ad libitum access to feed and water. The initial BW for Groups 1 and 2 were 7.62 ± 0.52 and 10.44 ± 1.79 kg, respectively. After the collection phase of Group 1, the pigs were allotted to 7 dietary treatments again by CRD and the same adaptation and collection process were repeated. Each collection period was carried out over 7 d, consisting of 5 d of adaptation phase and 2 d of collection phase. The experimental diets were supplied twice a day at 0700 and 1900 with ad libitum access to water according to the rate of 2.0 times of the maintenance requirement for ME  based on initial BW of pigs. The ileal digesta were collected during 12 h from 0800 to 2000 for 2 d and the detailed collection procedure followed Jorgensen et al. [All pigs were housed in individual metabolic crates  and means were compared using least significant difference multiple range tests by the GLM procedure of SAS . Differences were declared significant at P < 0.05 and highly significant at P < 0.01. Single degree of freedom contrasts were conducted between different bacterial species for FSBM, between FSBMB and UVFSBMB, and between FSBMB and PSBM.Bacillus subtilis supplementation (PSBM) and ultraviolet (UV) sterilization (UVFSBMB) compared with CSBM and FSBMB, respectively.Fermented soybean products contained higher dry matter (DM), CP and AA contents compared to CSBM. Soy protein concentrate (SPC) had the greatest contents of DM, CP and AA among the soybean products Table . HoweverThe AID of CP was not different among dietary treatments. However, lower AIDs of Ile, Phe, Val, and Tyr, were observed in the CSBM diet compared with the FSBMB diet  had higher AIDs and SIDs of most AAs than FSBMA (fermented by Aspergillus oryzae). There is limited information about the effect of different fermenting microbial strains on the SIDs of AAs. Teng et al. [Bacillus subtilis (96%) than by Aspergillus oryzae (82%), and the amount of small peptide increased from 5% to 63% in the FSBM by Bacillus subtilis and from 5% to 35% in the FSBM by Aspergillus oryzae. The reduced trypsin inhibitor concentration in SBM can improve protein and AA digestion [Bacillus subtilis compared with those of Aspergillus niger was demonstrated when Parkia biglobosa was utilized as a fermented ingredient [Bacillus subtilis may have more potential to improve SBM availability than Aspergillus oryzae.Regarding the different bacterial species for SBM fermentation, FSBMB  showed slight improvements on the AIDs and SIDs of some AAs compared with the CSBM, but lower AIDs of Lys and Pro and SIDs of Lys, Ala, Pro, Ser, and Tyr compared with the FSBMB. The responses of supplementation of Bacillus subtilis on ileal AA digestibility of pigs are inconsistent [Bacillus subtilis supplementation was not provided. However, FSBM was supplemented to the diet after finishing the fermentation process. This difference in the pretreatment time might affect the AID and SID of AAs, resulting in little improvement of the ileal AA digestibilities in the PSBM diet.The supplementation of nsistent ,30. In tBacillus subtilis was killed by UV-sterilization, which means that the pigs fed UVFSBMB diet could not consume probiotics alive. Therefore, consumption of microbial residues after the fermentation process had no beneficial effect on the ileal AA digestibility because it is likely that the nutrients for Bacillus subtilis in the feed were already spent in fermentation.The UV-sterilization of FSBMB did not affect the ileal AA digestibilities. There is no evidence to demonstrate this effect. However, in this study, there is no difference in AA composition between the FSBMB and UVFSBMB. The only difference is that the supplemented Bacillus subtilis had more potential to improve ileal AA digestibility when compared with Aspergillus oryzae. However, supplementation of Bacillus subtilis had no positive effect on ileal AA digestibility in weaning pigs. Further study is needed to explain the possible mechanisms of different responses on ileal AA digestibility from different types of fermenting microbes for SBM.Supplementation of FSBM products in the diet of weaning pigs could improve AIDs and SIDs of AAs, and fermentation with


Balaenoptera musculus) that occurs in the Gulf of California, we genotyped a 172 bp fragment of exon 2 of the MHC Class II DQB locus for 80 members of this population. Twenty-two putatively functional DQB allotypes were identified, all of which were homologous with DQB sequences from other cetacean species. Up to 5 putative alleles per individual were identified, suggesting that gene duplication has occurred at this locus. Rates of non-synonymous to synonymous substitutions (ω) and maximum likelihood analyses of models of nucleotide variation provided potential evidence of ongoing positive selection at this exon. Phylogenetic analyses of DQB alleles from B. musculus and 16 other species of cetaceans revealed trans-specific conservation of MHC variants, suggesting that selection has acted on this locus over prolonged periods of time. Collectively our findings reveal that immunogenic variation in blue whales is comparable to that in terrestrial mammals, thereby providing no evidence that marine taxa are subject to reduced pathogen-induced selective pressures.The genes of the Major Histocompatibility Complex (MHC) play an important role in the vertebrate immune response and are among the most polymorphic genes known in vertebrates. In some marine mammals, MHC genes have been shown to be characterized by low levels of polymorphism compared to terrestrial taxa; this reduction in variation is often explained as a result of lower pathogen pressures in marine habitats. To determine if this same reduction in variation applies to the migratory population of blue whales ( The major histocompatibility complex (MHC) is a multigene family that plays a fundamental role in the vertebrate immune response ,2. SpeciBalaenoptera physalus; sei whales, B. borealis; beluga, Delphinapterus leucas), which are characterized by lower levels of MHC variation than most terrestrial mammals surveyed to date [Lipotes vexillifer [Megaptera novaeangliae [Delphinapterus leucas eight DQB alleles in 233 individuals were reported [Balaenoptera physalus three alleles were found in 36 individuals [DQB locus has been reported in some marine species  by finding more than two alleles per individual [Presumably as a result of pathogen-mediated balancing selection, MHC genes are typically among the most polymorphic loci in vertebrate genomes ,2. Appar to date ,7–9. The to date ,9. Due txillifer  and Megaeangliae  have revividuals ; the freividuals . Thus, tividuals ,11 is lidividual ,11.DQB locus in a population of blue whales  from the Gulf of California, Mexico. This locus was chosen as among mysticetes the DQB locus is reportedly more variable than other commonly studied MHC genes such as the DRB gene, which seems to be more variable in odontocetes [DQB gene encodes the peptide binding region (PBR) of the DQ β chain [To explore determinants of variation at cetacean MHC loci in greater detail, we characterized patterns of variability and selection at the MHC Class II ntocetes ,7). Furt β chain ,10,11,13DQB locus for a subset of the individuals overwintering in the Gulf of California. Our analyses generate new insights into patterns of immunogenetic variability in these animals as well as the impacts of selection on cetacean MHC genes over multiple time scales.Although cosmopolitan in distribution, this species is divided into four geographically and genetically distinct subgroups: Northwest Atlantic, Indic Ocean, South Ocean, and East Pacific . After eBiopsy samples were collected under D. Gendron annual research permits issued by the Secretaría de Medio Ambiente Recursos Naturales y Pesca  from 1997 to 2001, and by the Dirección General de Vida Silvestre, Secretaría de Medio Ambiente y Recursos Naturales  from 2002 to 2012, which now represents the only approved Government institution to issue research permits on endangered species in Mexico. By issuing these annual research permits they approved the methods and the number of blue whale skin-blubber biopsies collection for each year of the permit. A more detailed description of biopsy sampling it could be found in Costa-Urrutia et al. 2013 [Balaenoptera musculus) over-wintering in the Gulf of California, Mexico, were genotyped at the MHC Class II DQB locus. Skin samples (5–10 mm) were obtained from Diane Gendron Laniel and colleagues at the Laboratorio de Ecología de Cetáceos y Quelonios, Centro Interdisciplinario de Ciencias Marinas (CICIMAR), Instituto Politécnico Nacional, Mexico. Samples were collected from 1999 to 2012 using a crossbow to shoot a small dart (distance ~ 10 m) at the whale, the tip of dart was designed to secure a skin-blubber biopsy from an individual before falling free so that the dart could be retrieved from the water, this procedure required no direct contact with the study animals [2 portion of each biopsy was isolated and stored in 70% EtOH at 4°C until sequencing analysis. The remainder of each biopsy was placed in 20% dimethylsulfoxide (DMSO), and then frozen in liquid nitrogen to create a tissue archive for the study population [Skin biopsies (35–40 mm) from 80 blue whales  following the protocol of the manufacturer.We examined a 172 bp fragment of exon 2 of the MHC Class II DQB locus was performed using primers DQB-F and DQB-R from Murray et al.[Amplification of the ay et al.. Per-samDQB exon 2 was limited to a single variant per sequencing reaction, all PCR products of the appropriate size (225 bp) were cloned using the TOPO® TA Cloning® kit (Invitrogen Inc.) following the instructions of the manufacturer. Sequencing of eight clones per PCR reaction is expected to have a high probability (p = 0.992) of detecting both alleles in heterozygous individuals [To insure that sequencing of the ividuals  and thusAll sequences were aligned and edited using the program Geneious .5.6.6 . To deteDQB locus, we performed BLAST searches [At the nucleotide level, we defined sequence variants as distinct alleles following the criteria of Kennedy et al. : variantsearches  using thDQB alleles identified for B. musculus followed Kennedy et al.[Bamu (corresponding to the two first letters of the genus and the species names) and the suffix GC, which represents the Gulf of California locality where the tissues samples were collected [DQB sequences generated during this study were accessioned into GenBank as numbers: KJ179618 to KJ179639.Nomenclature for the dy et al.; each alollected . DQB seqS) and non-synonymous (dN) base pair substitutions  for all codons as well as for the subset of codons corresponding to the PBR of this exon. Ratios greater than one (ω > 1) indicate an excess of non-synonymous substitutions relative to neutral expectations and such values are typically interpreted as evidence of positive selection [DQB locus [B. musculus. We calculated values of ω for the putative PBR following the above procedure. Significance tests of differences between ω values were conducted using the Z-test contained in MEGA 5.2 [Evidence of selection at the molecular level was assessed using three distinct procedures. First, we calculated rates of synonymous  tree reconstruction was conducted by using the matrix of distances compound as implemented in PAUP* 4.0b10 [DQB alleles from B. musculus and other cetaceans, we constructed a median-joining haplotype network using Network 4.6 [To assess evidence for trans-species conservation of  GenBank , DQB all* 4.0b10 , genetic* 4.0b10 . These a* 4.0b10 , the runwork 4.6 , which iwork 4.6 .DQB locus was identified from the 80 blue whales examined , were sequenced. More than two DQB alleles were found in 36 (45%) of the surveyed animals , suggesting that members of the population possessed multiple copies of this DQB locus.A total of 757 cloned PCR products  being the most similar[A BLAST search indicated that the allotypes detected in our population displayed 90% to 99% concordance with exon 2 of the t similar. VariatiDQB. At the amino acid level, our sequenced fragment contained 57 amino acids of DQB exon 2 as indicated by the P value (P> 3.05).We obtained a positive value of the Tajima’s D (D = 1.078), however statistics calculations did not show a significant departure from neutrality at p < 0.01) in the entire 172 bp DQB sequence. Otherwise comparisons of ω for the PBR and non-PBR portions of exon 2 revealed that 10 (71.4%) of the 14 amino acid changes detected occurred in the PBR. Within the PBR, ω was significantly greater than one , in contrast, ω did not differ significantly from neutral expectations  for the non-PBR portion of the exon  of this exon. The DQB locus in our study population appears to be subject to selection at the nucleotide level.Class II MHC loci are typically thought to be subject to strong positive selection due to their role in detecting and responding to pathogens–4. Our fDQB alleles from our study animals and other cetaceans revealed evidence of trans-specific preservation of alleles, particularly between blue and humpback whales. Similar trans-specific conservation of MHC alleles has been reported for a variety of mammalian species including non-human primates [DQB locus in blue whales from the Gulf of California have been subject to selection over multiple scales.Comparisons of primates , bank voprimates , and tucprimates  as well primates ,13,14,42L. vexillifer: 33 functional alleles in 18 individuals [T. truncates: 27 functional alleles in 48 individuals [B. musculus from the Gulf of California are characterized by considerable MHC variation, the population of fin whales  from this locality displays low levels of MHC variation [In contrast to our findings, studies of a number of species of cetaceans have reported limited variability at MHC loci ,7–9,11, ividuals ), and Atividuals . FurtherDQB locus per individual in our study population. None of the alleles detected contained insertions, deletions, or stop codons, suggesting that some of these copies might represent functional sequences. We consider that our findings suggest that one or more gene duplication events have resulted in multiple functional copies of DQB locus, this assumption supports the hypothesis that cetacean DQB sequences are paralogous genes separated by gene duplication events, which is thought to be one of the major forces for MHC evolution [DQB locus has been reported for several other cetacean species  as well indicus , Cervus  elaphus , Bubalus bubalis , Mastomytalensis ), thereftalensis ,41,49,50One factor that may contribute to interspecific differences in immunogenetic variability among marine mammals is demography. For example, the population of fin whales in the Gulf of California is resident in that area and is isolated from a population of conspecifics . The GulDQB locus in blue whales from the Gulf of California revealed levels of immunogenetic variability comparable to those of many terrestrial mammals. Further, our data revealed evidence of selection on this exon at both the nucleotide and trans-specific levels, thereby contradicting the hypothesis that marine taxa are subject to less intense selective pressures than terrestrial mammals. Instead, we suggest that differences in current and historical demography may better explain the differences in immunogenetic variability evident among marine mammals. Future studies of MHC variation in blue whales and other marine taxa will benefit from exploring differences in pathogen-induced selective pressures as a function of both habitat and variation in current and historical demography.Our analyses of variation at exon 2 of the


The field work was carried out near Corrientes, Argentina. Lime treatments were 0, 625, and 1250 kg ha−1 dolomite, and each liming dose was applied to a 1.7 ha field. Ninety-three soil samples per treatment were first collected in aerobic conditions and then two more times after flooding, at bunch formation and flowering. Soil NH4+-N increased along time, whereas P was highest at bunch formation and K steadily decreased along the rice growth period. Dolomite addition increased macronutrient availability at the first and second samplings, but its effects at the third sampling depended on the element. The three soil nutrients analyzed displayed strong patterns of spatial dependence for the three lime treatments and at the three periods studied. The areas with relative high or low macronutrient concentrations within each field were not stable throughout the rice growth period. Seasonality in the spatial distribution of macronutrients may be of agronomic value for site specific management.Proper and effective management of soil nutrients requires assessment of their variability at the field scale. We compare the effects of lime amendment rate on the spatial variability of three macronutrient forms (NH Macronutrients  play an important role in crop production. Inadequate fertilizer management limits crop yield, results in nutrient mining, and causes loss of soil productivity. A satisfactory nutrient status saves the soil, a limited natural resource, and also prevents environmental pollution . Proper The spatial variability of soil properties is the outcome of the interaction of several soil forming factors and processes and in agricultural and forest fields involves also effects of management practices –5. ThereIn paddy soils, spatial variability has been shown to occur over distances of meters or tens of meters. Because these soils are characterized by a relatively flat topography and seasonal flooding, in the past they have been thought to be apparently homogeneous. However, large spatial variability in soil properties has been reported for paddy fields of different countries , 14, 16,4+), rather than nitrate (NO3−), has been found to be the main source of N for crop growth. This is because NO3− (if present) can be depleted rapidly after flooding due to denitrification, while NH4+ tends to accumulate with time of submergence due to lack of O2 for nitrification. Therefore, ammonium is likely to be the main form of nitrogen in flooded soils [3−, are very mobile and may show a marked seasonal change in concentrations during the growing season, owing to the unstable physicochemical properties, for example, redox potential, or to variations in microbial activity.Successful rice production requires an adequate soil fertility status and especially satisfactory levels of N, P, and K. In flooded soils major chemical and electrochemical changes occur that have a profound influence on soil nutrient availability. In this way, under submerged condition as in paddy rice cultivation ammonium . The experimental site and the sampling procedure have been described before [The study was conducted on a paddy field located near Corrientes, Argentina, cropped to rice , 625 kg hachannels . The expSoil samples were collected at three different stages during the rice growth period. The first sampling was performed in aerobic conditions, just before sowing, the second sampling at bunch formation, that is, 28 days after flooding, and the third sampling at flowering stage, that is, 56 days after flooding. Ninety-three samples per liming treatment were taken at each of the three sampling dates. The basic sample grid was 11.9 × 20 m . TherefoThe soil was collected to a depth of 15 cm. Soil samples were air-dried and sieved (2 mm mesh). Ammonium-N was extracted with 2 M KCl and determined using methods described in . Availab−1 dolomite <1250 kg ha−1 dolomite, at the three sampling dates. Redox potential decreased along the rice growth period with increasing duration of the anaerobic conditions; the effect of dolomite addition was to decrease mean soil redox potential on the three sampling dates.Mean values of pH and redox potential (Eh), reported in , are sumThe 27 data sets studied (3 nutrients × 3 lime treatments × 3 sampling dates) were first analyzed for normality by means of the Kolmogorov-Smirnov test. Then, descriptive statistics including mean, variance, coefficient of variation, maximum, minimum, skewness, and kurtosis were determined. Pearson correlation coefficients were also calculated to determine the relationship between soil properties.γ(h), which are given by the following equation:Z(xi) and Z(xi + h) are the actual values of the variable, Z, at places xi and (xi + h), and N(h) is the total number of data pairs separated by a distance h. Thus, the experimental or sample variogram is obtained by changing h.Geostatistical analysis is based on the assumption that measurements separated by small distances are more likely to be similar to each other than those farther apart, meaning that spatial autocorrelation exists. This hypothesis can be verified through examination of semivariograms for the attributes under investigation. Moreover, the statistical tool used to measure the autocorrelation between samples is called the semivariogram. An experimental semivariogram can be obtained from semivariance values calculated as a function of the distance, r2), mean error (ME), and no dimensional mean square error (NMSE).The experimental semivariogram must be fitted by a model that mathematically describes the spatial variation. Several standard models  are currently available. In this work, calculation of sample variogram and fitting of models were made using accredited criteria and procedures , 33. Theh, the sill variance (C0 + C1), and the range of spatial dependence (a). The sill variance eventually reaches an upper bound; this parameter consists of a spatially correlated or structural variance (C1) and may include a nugget variance (C0), which represents the variation that has not been resolved at the scale of sampling. The degree of spatial dependence can be expressed by the ratio of nugget variance (C0) to the threshold (C0 + C1) variance. The nugget to sill ratio was used to qualitatively classify the spatial dependence into strong (<25%), moderate (25 to 75%), and weak (>75%) following accredited criteria [Semivariogram models were described using the following basic parameters: the distance criteria .Based on the best fitting semivariogram model, the kriging approximation was used for interpolation and mapping purposes. Kriging is an optimal method of prediction in the sense that it is unbiased and the predictions have minimum variance , 28.4+-N just before flooding ranged from 20.07 to 28.87 mg kg−1 and those at rice flowering (56 days after flooding) ranged from 37.74 to 43.92 mg kg−1. The latter were significantly higher than the former (P < 0.05) and this was for the three liming treatments. The average increase in NH4+-N concentrations between the first and the third sampling period was 88%. Ammonium-N accumulation in anaerobic conditions is an expected result, due to the lack of conversion of organic N into mineral N in paddy soils [−1, were not much different from those of the first sampling date, even if the rate of NH4+-N accumulation was not negligible for the control treatment during this period. Moreover, comparison between the first and the second sampling dates showed small but significant (P < 0.05) ammonium-N losses and increases at the 625 kg ha−1 and 1250 kg ha−1 treatments, respectively. The net increase in NH4+-N after flooding is the result of the net balance from accumulation (owing to the lack of O2 for nitrification) and losses .Descriptive summary statistics of the 27 data sets studied are presented in dy soils , 22, as 4+-N concentration, both at sowing and at bunch formation, as the differences between the control treatment and the two treatments amended with dolomite were significant (P < 0.05). Nevertheless, concentrations of NH4+-N within each sampling period were considerably stable, as shown by the smaller range of values among liming treatments than among sampling dates.There were positive effects of lime amendment on NH4+-N data sets studied ranged from 9.7 to 20.3% and showed a trend to decrease for the successive sampling periods. This seasonal reduction in the statistical variability of NH4+-N concentrations throughout the rice growth period could be the result of more uniformity in microbiological activity across the field, which in turn may be due to a higher homogeneity of the redox conditions  increased Olsen P availability in aerobic conditions before flooding. Also, at the second sampling the mean concentration of Olsen P was significantly lower (P < 0.05) in the control treatment than in the two dolomite amended treatments. However, at the third sampling date, the highest mean Olsen P values were recorded at the 625 kg ha−1 dolomite treatment. In general, and in agreement with the trends observed before for ammonium-N, Olsen P concentrations showed smaller differences among liming treatments than among sampling dates  with rate of dolomite application. Hence, in the two first sampling dates, mean Mehlich-1 K values for the three dolomite treatments ranked as control < 625 kg ha−1 < 1250 kg ha−1. At the third sampling date, however, the mean extractable K concentrations measured at the control treatment were significantly lower than those at the two dolomite amended treatments, but no significant differences were found between the two treatments with dolomite amendment. The control treatment had the lowest mean extractable K level before flooding and the highest after 58 days of flooding; in contrast the 1250 kg ha−1 treatment had the highest mean extractable K level before flooding and the lowest after 58 days of flooding. Hence, the rate of diminution in soil available K throughout the crop growth period was greater for the dolomite amended treatments than for the control treatment. Therefore, seasonal changes in available K under paddy rice cultivation were not similar for all the treatments, suggesting specific K fixation or depletion depending on soil reaction.Mean extractable K concentrations in the soil measured at sowing and at bunch formation also increased significantly /(C0 + C1). The geostatistical parameters for the best fitted semivariogram models describing the spatial dependence of the soil macronutrients studied are listed in h), until a more or less constant value was reached, at a given separation distance, that is, the sill or total semivariance (C + C0); this separation distance (a) is called the range of spatial dependence. Note that the nugget variance, C0, is the semivariance at h = 0. Experimental semivariograms of the NH4+-N concentrations measured at the three lime treatments and during the three periods studied could be described quite well by either spherical or exponential models with a nugget component (C0) plus a spatial component (C1) with a range of spatial dependence varying between 40.9 and 84.6 m. For Olsen P and Mehlich-1 K, all experimental semivariograms were described by spherical models with a range between 51.4 and 76.8 m.Selected examples of semivariograms are presented in r2), mean errors (ME), and no dimensional mean square errors (NMSE). Goodness of fit was assessed by proximity to ideal values of these parameters for a perfect fit, that is; r2 = 1, ME = 0, and NMSE = 1 [r2 ≥ 0.639, mean errors were ME ≤ 0.059, and no dimensional mean square errors were 0.813 < NMSE < 1.189. Therefore, the values of these parameters suggest that all models adequately fitted the spatial dependence of the studied macronutrient data sets.Theoretical models were fitted to experimental semivariograms based both on subjective visual inspection and on the following parameters obtained by cross-validation: determination coefficients of the regression between calculated and kriging estimated values  also have been found to fit mostly spherical models and otherwise exponential models [Note also that in most of the cases studied the experimental semivariograms were best described by a spherical model. The exception was for two NHl models , 17.C0/(C + C0), ranged between 0 and 43.7% for all the data sets studied. This ratio was zero at four data sets, was smaller than 25% at 11 data sets, and ranged from 25 to 50% at 12 data sets, indicating in most of the cases a strong to moderate degree of spatial dependence, according to the authorized criteria [The nugget variance represents either experimental variability induced by potential laboratory errors or field random variability that can not be detected at the scale of sampling. The nugget to sill ratio, criteria  previousC + C0) reflects the total variance for very large distances; the sill is given by the sum of the nugget semivariance (C), which may be zero, plus the portion of the semivariance that is spatially structured (C0). As expected, for each data set, the value of the sill modeled was of the same order of magnitude as the value of statistical variance. The sill value ranged from 12.35 to 32.63 (mg kg−1)2 for NH4+-N, from 1.82 to 6.67 (mg kg−1)2 for Olsen P, and from 6.17 to 89.81 (mg kg−1)2 for Mehlich-1 K, indicating that Olsen P had the weakest pattern of spatial variability. Mean sill values were lowest for NH4+-N at the second sampling date; however in this sampling they were highest for available P and K. Main factors responsible for the differences in semivariance for very large distances, that is, for the sill value, of the data sets studied here could be: (a) seasonal variations, driven by the different biogeochemical processes during aerobic and anaerobic conditions, (b) nutrient uptake by the rice crop, and (c) management effects such as liming.The sill value  and during each of the three sampling dates.Examples of kriging maps for ammonium-N, Olsen P, and Mehlich-1 K are shown in Figures The patterns of spatial distribution of the three nutrients presented in the maps of Figures Our results also showed that seasonality changed the patterns of distribution of ionic species with different degrees of mobility such as ammonium, phosphate, or potassium. Subsequently, seasonal variability in soil macronutrient concentration  was not only present but potentially of agronomic importance.Our macronutrient maps were based on an intensive sampling on a 50 × 11.9 m grid. Farmers cannot afford this level of investment in sampling and often rely on data from satellites or aircraft or from tractor-borne equipment. However, and in spite of significant technological advances, nowadays a lack of decision criteria still remains for an efficient site specific management of fertilizer inputs on several crops, and this is the case for lowland rice. To overcome this deficit nutrient maps based on intensive soil sampling are required. Therefore, kriging maps are useful for optimizing soil sampling and for delineating management units. They have been widely used in rainfed agriculture as a straightforward approach for precise management of phosphorus and potassium, since these nutrients show comparatively stable spatial distribution patterns in aerobic conditions , 3, 6, 7In our lowland paddy soil, marked seasonal changes in mean concentrations and in the spatial distribution of ions with wide differences in mobility such as ammonium-N, phosphate, and potassium were detected. This information may be potentially very useful both for site specific management and for environmental purposes. In general, recommendations for fertilizer application to rice crops are based on correlation between soil test values of the target nutrient measured on samples collected before flooding. The above results suggest that the efficiency of site specific management practices in rice fields would increase if the observed patterns of seasonal changes in macronutrient concentration are taken into account. This is in agreement also with recent work .On the other hand, analysis of soil spatial variability also provides valuable knowledge for environmental management. For example, if only mean values of macronutrients were taken into account, the low concentrations of ammonium-N and available P and K across the rice growth period were unlikely to have any adverse effects on the environment; however the presence of spots with nutrient concentrations higher than the average could be critical for ponding water and ground water quality.Mean values of ammonium-N, Olsen P, and Mehlich-1 K sampled during three different dates along the rice growth period, and under three dolomite treatments, in an acid paddy soil, varied both seasonally and as a function of the liming rate. Increasing rates of dolomite amendment increased ammonium-N, Olsen P, and Mehlich-1 K, both before sowing (aerobic conditions) and 28 days after flooding (anaerobic conditions). Meanwhile, after 56 days of flooding effects of dolomite amendment on the concentration of the studied nutrients were dissimilar.Ammonium-N accumulated in anaerobic conditions and was highest at the third sampling date, 56 days after flooding. Olsen P availability increased with increasing pH after flooding, but this effect was reversed for pH values higher than 6.5 measured at the flowering stage of the rice crop. Mehlich-1 K steadily decreased along the growth period of rice.4+-N, P, and K on rice fields with three different rates of dolomite amendment and at three different crop stages was best described mainly by spherical models and otherwise with exponential models. The nugget to sill ratio ranged from 0 to 43.7% for all the data sets studied, showing strong to moderate pattern of spatial dependence. The small to moderate nugget effects also indicate that the sampling grid used was proper to reflect the spatial dependence of the studied macronutrients.The spatial variability of NH4+-N, Olsen P, and Mehlich-1 K within each liming treatment and during each of the three sampling dates. Also the position of patches with high and low concentrations of the studied macronutrients changed between successive sampling dates, providing evidence of the lack of temporal stability in the patterns of spatial distribution. Seasonality in the spatial distribution of macronutrients should be considered as an important factor for an efficient site specific management.Kriging maps clearly showed the presence of small scale variability for NH


In secondary progressive Multiple Sclerosis (SPMS), global neurodegeneration as a driver of disability gains importance in comparison to focal inflammatory processes. However, clinical MRI does not visualize changes of tissue composition outside MS lesions. This quantitative MRI (qMRI) study investigated cortical and deep gray matter (GM) proton density (PD) values and T1 relaxation times to explore their potential to assess neuronal damage and its relationship to clinical disability in SPMS.11 SPMS patients underwent quantitative T1 and PD mapping. Parameter values across the cerebral cortex and deep GM structures were compared with 11 healthy controls, and correlation with disability was investigated for regions exhibiting significant group differences.PD was increased in the whole GM, cerebral cortex, thalamus, putamen and pallidum. PD correlated with disability in the whole GM, cerebral cortex, putamen and pallidum. T1 relaxation time was prolonged and correlated with disability in the whole GM and cerebral cortex.Our study suggests that the qMRI parameters GM PD  and cortical T1  are promising qMRI candidates for the assessment of disease status, and are related to disability in SPMS. MS is an immune-mediated chronic disease of the CNS. The diagnosis and treatment of MS is based on clinical observations and supported by MRI . As WM lThe transition to secondary progression is important clinically, because therapy options are limited for this phase. Importantly, the transition to Secondary Progressive Multiple Sclerosis (SPMS) can be delayed by disease modifying therapy . In SPMSQuantitative MRI (qMRI) techniques allow for the direct measurement of tissue parameters in vivo, providing a window into the microstructural tissue composition and its changes in MS. qMRI detects pathological processes in tissue that appears normal on conventional MR images, and the measured tissue parameters have been shown to correlate with disability in RRMS and mixed collectives:Prolongation of the T1 relaxation time in normal appearing brain tissues has been shown for various collectives of MS patients, including primary-progressive MS patients ,10, SPMSIn RRMS patients, the proton density (PD) in the frontal white matter inside and adjacent to the corpus callosum was found to correlate strongly with EDSS .In summary, a correlation between T1 values measured in various regions and the degree of disability has been shown in mixed collectives including RRMS and SPMS patients, but it is unclear whether these results are representative for SPMS patients, or whether the correlation with disability was driven by the combination of patients at early and late stages of disease. For the parameter PD, which indicates replacement of neural tissue by water in the cortex, no data specific to SPMS patients are available.There is an as yet unmet need for better imaging surrogates of disease state in SPMS, where information from conventional MRI does not sufficiently reflect the ongoing pathology. In this study, we explore qMRI measurements taken from cortical and subcortical GM regions of interest (ROIs) in SPMS patients. We assess parameter changes relative to a collective of healthy subjects, and explore the relationship between qMRI tissue parameters and clinical disability in SPMS. The hope is to provide clinicians and researchers with a means to measure neurodegeneration directly, and to provide a suitable and clinically relevant imaging surrogate of neuronal damage in SPMS.The study presented here aims at an assessment of microstructural changes in cerebral tissue apart from MS lesions. T1 and PD values in the cerebral cortex and deep GM of SPMS patients were systematically investigated with advanced qMRI and segmentation methods, comparing the results with values measured on a matched collective of healthy controls and evaluating the correlation with the EDSS.11 patients  with SPMS and 11 age-matched healthy control subjects  participated in the study. Healthy controls were free from neurological or psychiatric disease, as assessed by systematic history evaluation and clinical examination. Further exclusion criteria were: brain surgery, drug abuse, diabetes mellitus and severe arterial hypertension. Data from an overlapping collective have been reported earlier, evaluating the predictive value of qMRI regarding gadolinium enhancing lesions . PatientMeasurements were performed on a 3 Tesla whole body MR scanner .3, TR/TE/α1/α2 = 16.4 ms/6.7 ms/4°/24°, BW = 222 Hz/Pixel. The total scan duration for both data sets was 9:48 min.The variable flip angle method was applied for T1 and PD mapping . This meNon-uniformities of the transmitted radiofrequency field (B1) were measured as described before : two mulFor the evaluation of residual signal losses induced by T2* relaxation effects during the finite TE of 6.7 ms, two multi-slice gradient echo data sets with different TE were recorded with the following parameters: isotropic spatial resolution: 2 mm, field-of-view as above, TR/TE1/TE2/α = 1336 ms/4.3 ms/11 ms/50°, BW = 292 Hz/Pixel. The scan duration for both data sets was 5 min.Data processing was performed with custom-built functions written for Matlab  and shell scripts using tools from the FMRIB Software Library .i/tan(αi) was plotted versus Si/sin(αi) where S1/2 are the local signal amplitudes in the PD and T1 weighted data sets, respectively, and α1/2 are the respective excitation angles, corrected for B1 inhomogeneities. Additionally, data were corrected for the effect of insufficient spoiling of transverse magnetization [B1 was obtained as described above and in the literature . Quantittization . PD mapstization : in summtization . PD valuSynthetic T1-weighted anatomical MP-RAGE data sets were calculated from the quantitative T1 and PD maps, following the procedure described in . The calAfter skull-stripping the data sets using the FMRIB Software Library tool “BET” for every participant, the software “FAST”  was appl3 kernel. The respective deep GM structures of both hemispheres were combined to yield one ROI per structure. Additionally, a global GM map was created from the deep GM and cortex maps. After removal of lesions as described below, the respective maps were applied to the T1 and PD maps and mean values for T1 and PD across each ROI were calculated. The study focused on GM changes, as GM pathology is not well respresented by clinical MRI. Still, in order to also investigate normal appearing WM (NAWM) separately, WM masks were obtained with ‘FAST’ from the FSL toolbox and eroded with a 6x6x6 mm3 kernel in order to reduce partial volume effects. After manual removal of any remaining MS lesions (see below), the masks were applied to the T1 and PD maps and the mean T1 and PD values in NAWM were extracted.Additionally, for every single participant, a segmentation of the deep GM structures, which are anatomically best suited for an accurate segmentation , was generated with “FIRST” . To avoiIn detail, the manual removal of lesions still remaining in the GM and WM ROIs was performed as follows: the respective GM and WM ROIs were carefully inspected together with the MP-RAGE datasets by a researcher experienced in the management of patients with MS. If a remaining MS lesion was identified within those ROIs, these were manually edited with ‘FSLVIEW’ and all voxels of the ROI overlapping with the identified MS lesion were removed. To assess T1 and PD values in the whole brain, voxels containing CSF only needed to be excluded. For this purpose, a threshold was applied to the skull-stripped quantitative maps removing voxels with T1 values above 1600ms ,27 from Statistical analyses were carried out with SPSS for Windows (20.0.0). Non-parametric testing was used (Mann-Whitney-U) for group comparisons of the respective mean parameter values. As evaluation of the relationship between quantitative and clinical parameters is only meaningful for ROIs in which T1 or PD are significantly increased in the MS group as compared to the control group, correlations of T1 and PD with EDSS, disease duration, gender and age were assessed only for the respective regions, using Spearman’s rank correlation coefficient. For all statistical procedures, tests where p was <0.05 were considered significant.Average age, EDSS and disease duration of the 11 patients examined (mean ± standard deviation) were: Age 46.8±11.03 years, EDSS 5.8±1.88, range 3–8.5, disease duration 15.3±6.75 years. The healthy control group did not differ in age . There was no significant correlation between the tissue parameters and age, gender, or disease duration for any ROI.For the whole NAGM, Figs This study investigates cortical and deep GM damage in SPMS patients using qMRI measurements of absolute PD and T1 values as potential imaging surrogates of disease state and disability. When MS patients enter the secondary chronic progressive phase, neurodegeneration, rather than localized inflammatory disease activity, appears to drive progression of clinical disability ,7. DiffuSo far, radiological assessment in SPMS patients relies on conventional MRI, usually by comparing the lesion load with previous scans and visually estimating the global brain atrophy. However, it is the tissue outside of these lesions where the pathogenic processes mainly occur in SPMS–indeed, disability is driven by damage to tissue outside overt lesions that remains inaccessible to routine MRI assessment . ClearlyOur data show that PD in the cerebral cortex, thalamus, putamen and pallidum is increased in our cohort of SPMS patients. Previous histopathological studies have identified neurodegeneration and subsequent loss of neurons as defining pathological processes in SPMS. The quantitative parameter PD assesses the tissue water content. For SPMS patients, PD might be sensitive to replacement of neural tissue by interstitial water, thus being sensitive to neuronal loss, but also to loss of supporting glial structures or myelin, or cell swelling. Gross edema is unlikely to occur in SPMS, but could form an alternative explanation for increased PD in other conditions. Attractively, our observations suggest that PD may be a suitable parameter to assess neurodegenerative changes in SPMS.T1 measurements in the cortex inform about the actual composition of tissue, including, but also reaching beyond water content. T1 reflects a complex interplay of factors such as myelin content, iron deposits, axonal density and cellularity. T1 changes in MS indicate loss of myelin, neuronal/axonal loss and gliosis ,36. CortWhile both T1 and PD changes were observed in the cerebral cortex, the question arises why PD only was increased in three out of four deep GM regions studied . A commoA nother factor for the differences observed for T1 and PD increases might be that, even though T1 values are affected by the water content, other processes driving T1 changes in SPMS, such as gliosis, myelin loss and reduced axonal density, play a more prominent role in the cerebral cortex than in the deep gray. Moreover, an increased deposition of iron in the deep GM  could shIn contrast to the situation with T1, increased water content in tissue, as reflected by increased PD, appears to be widespread in both the cortex and deep GM ROIs. While it is not possible to isolate a single cause of the T1 changes from our data, they suggest that the pathological processes in cortical and deep grey are not identical. An alternative explanation would be a higher sensitivity of PD in the detection of neurodegeneration in SPMS.To investigate the clinical relevance of our findings, we performed correlation analysis between the qMRI measurements in the GM regions identified above and EDSS, a score for the assessment of disability in MS . EDSS waIn the study presented here, a significant correlation with the EDSS could also be shown for cortical and whole NAGM T1. These results are congruent to a previous study showing cortical T1 to be correlated with disability in a mixed RRMS and SPMS group . InteresNaturally, patients with higher EDSS scores tend to be older than patients with lower EDSS scores. However, as GM T1 and PD values decrease with age ,40, thisInterestingly, T1 and PD values were increased in NAWM, but a relationship with the degree of disability could only be observed for the respective values in NAGM. In NAWM, no such relationship was found, which indicates that GM might be a more relevant target for the assessment of global neurodegenerative processes with qMRI techniques in SPMS.We also found T1 and PD increases across the whole brain. These, however, would strongly depend on the macroscopic lesion load. No correlation of T1 and PD values with the EDSS could be observed for the whole brain. These results suggest that the analysis of clearly defined anatomical regions with similar microstructural properties outside of MS lesions is of advantage for the assessment of neurodegeneration in relation to the clinical status in SPMS.While the present study focuses on GM pathology, previous work performed a more complex WM analysis subdividing the WM outside of MS lesions into NAWM and diffusely abnormal white matter, which seem to reflect independent pathological entities in MS .Our study has some methodological innovations that help avoid partial volume effects and missegmentation, especially in the thin cortical ribbon. Segmentation and ROI definition was performed on synthetic MP-RAGE data sets directly calculated from the T1 and PD maps. Therefore, the quantitative maps share the coordinate space with the anatomical data set, and no registration was required. This reduces the risk of misregistration, partial volume effects, and also of misclassification due to applying segmentation steps in different reference frames. Furthermore, artifacts and misclassified voxels were carefully removed, which is of special importance for the cortical volume which is close to CSF cf. . The remIn contrast to magnetization transfer ratio , the tisConventional MRI is widely available and represents standard in clinical routine. Clinical studies might benefit from including qMRI protocols to assess properties of normal appearing tissues, opening a separate window for assessment of the disease. One benefit of the qMRI technique is the ability to provide synthetic anatomies from the data, such as performed in the study presented here. This approach reduces the total MR examination time , becauseOur study is not without limitations. The number of patients examined is small and the cohort examined might therefore not be representative for SPMS patients in general. Nevertheless, correlations of PD and T1 relaxation time with disability measurements were present in our study group. This fact is encouraging, as the use of MRI surrogates which become significant only in a very large collective would have been more limited. Furthermore, increased partial volume effects in the SPMS group caused by atrophy can never be fully excluded when analyzing the thin cortical ribbon, which might have contributed to the cortical T1 and PD increase. However, this argument would not apply for the deep GM ROIs, where partial volume effects were avoided by performing morphological erosion.Identification of lesions for exclusion from the ROIs was based on routine MR images, but did not involve special sequences to facilitate the detection of cortical lesions, such as double inversion recovery. However, even DIR detects only 37% of the GM lesions identified by histopathology . AccordiWhile the degree of disability was quantified with the EDSS in our work, future studies might benefit from more detailed neuropsychological data and other clinical scores such as the Multiple Sclerosis Functional Composite Measure (MSFC) when assessing the relationship of behavioural measures to GM T1 and PD changes in SPMS. Moreover, imaging at higher field strengths might even allow a discrimination of different cortical layers, and their discrete analysis in SPMS.The study presented here focuses on T1 relaxometry and PD mapping. Future studies may also benefit from incorporating the parameter T2, which has been shown to decrease in the deep GM of MS patients , but to In conclusion, this study assessed the parameters PD, a potential indicator of replacement of tissue by water, and T1, an indicator of compositional changes including but also beyond increased water content such as gliosis, demyelination and axonal loss. Our results suggest that, cortical and deep GM PD values and cortical T1 values provide biologically meaningful insights into disease status in SPMS. Quantitative T1 and PD mapping helps elucidate pathological processes in SPMS beyond the lesion load, and could potentially allow for the measurement of ongoing pathology in SPMS.In the future, we hope that clinical studies in SPMS can use qMRI tools to assess ongoing tissue damage, and perhaps even efficacy of treatment as new treatment strategies for SPMS become available. Eventually, qMRI may even help to guide the therapy for individual patients.S1 FigEfforts were taken to exclude lesions (some are marked with red arrows) from the ROIs.(PDF)Click here for additional data file.S1 File(SAV)Click here for additional data file.


AbstractLithophaga hastasia, Adula soleniformis, Mactrellona subalata, and Strigilla ervilia. The number of species increases from the upper (44) and lower intertidal (53) to the shallow subtidal (76), but only 11 (17%) have a wide distribution in the bay . The bivalve assemblages are composed of four main life forms: 27 epifaunal species, 26 infaunal, 16 semi-infaunal, and 20 endolithic. A taxonomic distinctness analysis identified the sampling sites and environments that contribute the most to the taxonomic diversity (species to suborder categories) of the bay. The present work increased significantly (31%) to 132 species previous inventories of bivalves of Bahía de Mazatlán. These species represent 34% of the bivalve diversity of the southern Golfo de California and approximately 15% of the Eastern Tropical Pacific region.We describe the composition and distribution of bivalve molluscs from the sandy and rocky intertidal and the shallow subtidal environments of Bahía de Mazatlán, México. The bivalve fauna of the bay is represented by 89 living species in 28 families, including 37 new records and four range extensions:  Studies on molluscs from Bahía de Mazatlán, located in the Mexican Pacific, have focused mainly on the conspicuous species of gastropods and bivalves from the rocky intertidal  and rockAccording to Many studies on marine biodiversity have used species accumulation curves  to evaluate the sampling effort; this technique indicates when a sufficiently large percentage of species has been observed with a definite number of samples with respect to a theoretical expected total number of species of a given community . The evaPageBreakbiodiversity at a regional scale is occasionally difficult, but the taxonomic distinctness parameter facilitates this measurement (+ and Λ+) are an easy-to-use tool to measure biodiversity in the time and space scales  (06°25'W) . The bay06°25'W) .PageBreakor no rainfall, occurs from November to June , lower intertidal (LI), and shallow subtidal (SS) (3–10 m depth) adjacent to each beach. The intertidal zones were defined according to the natural zonation of benthic invertebrates (tebrates .Olas Altas (OA), a partially-protected beach with a well-developed rocky area of approx. 150 m long and smaller areas of medium-to-coarse sand; 2) Los Pinos (LP), a protected rocky beach approx. 100 m long, with some areas of medium-to-coarse sand; 3) Casa del Marino (CM), a semi-protected to exposed rocky beach approx. 250 m long mixed with small sandy areas of fine-to-medium size grains; 4) Venados Island (VI), the east side of the island has an extensive protected sandy beach approx. 850 m long and 30–60 m wide with medium-to-coarse sand; towards the northern part of the beach there is a rocky beach approx. 200 m long with many tidal pools and boulders; 5) Malecon (MA), an exposed and very dynamic sandy beach approx. 400 m long of medium-to-coarse PageBreaksand and pebbles. The beach runs along the urban sector of the city of Mazatlán and it is frequently visited by locals and tourists; and 6) Zona Dorada (ZD), a protected beach of medium-to-fine sand approx. 300 m long located in the tourist hub of the city just in front of hotels and extensively visited by tourists. The adjacent shallow subtidal environments of all these beaches include mixed substrates composed by coarse and fine sand, rocky reef areas, and many shell fragments. In Venados Island and Los Pinos small patches of live coral are also frequent.Some of the general characteristics of these beaches are: 1) x = 3) quadrats (0.5 m2) were placed equidistant along each transect and all bivalves found in each quadrat identified in situ or collected for taxonomic identification in the laboratory. In the shallow subtidal, sampling was performed during SCUBA diving. The total sampling effort was 126 quadrats (63 m2) in the rocky intertidal and 52 quadrats (26 m2) in the sandy intertidal. A total of 90 quadrats (45 m2) were sampled in the shallow subtidal environments of all beaches. Additionally, in order to increase the inventory of bivalves, the specimens found in the areas immediately adjacent to the quadrats were also identified in situ or collected during direct searches in the intertidal and shallow subtidal environments. Dredges (24) were carried out in the shallow subtidal zone (8–15 m depth) of the six beaches, using a naturalist´s dredge   were applied during four expeditions in December 2008, and March, June, and August 2009. The transects (15 m long) were set parallel to the coastline, two in each environment  of each beach. Two to four ( 1.3 cm)  during 1Laboratory methods. A detailed examination of each sample was conducted to search for bivalves. Only living specimens were considered. Endolithic specimens  were obtained by breaking rocks and shells, coral fragments, polychaete tubes, and rodoliths. Epifaunal specimens  were obtained by scraping the surface of rocks. Semi-infaunal specimens  and infaunal specimens (< 4 mm)  were obtained by screening the sandy sediment . Six taxonomic levels  were considered based on the classification schemes of ERMANOVA .The species accumulation curves show a trend towards an symptote in all environments . The obsPageBreakMytilidae (14 species), Veneridae (10), and Arcidae (8). Ten families (35%) included only one species. The number of species increased from the upper (44) and lower intertidal (53) to the shallow subtidal (76). In addition, the numbers of unique species were 7 (upper), 4 PageBreak(lower), and 18 . The species richness was similar in the adjacent shallow subtidal zone of all beaches (28–36 species) except for Venados Island (55 species), which had the highest number of species restricted to this island (7)  .Crassinella coxa, Crassinella nuculiformis, Liralucina approximata, Ctena mexicana, Kellia suborbicularis, Neolepton subtrigonum, Nutricola cf. humilis, Pitar cf. omissa, Sphenia fragilis, Chioneryx squamosa, and Transennella cf. puella. Only three species were collected in the sandy intertidal environment: Strigilla cicercula, Strigilla dichotoma, and Donax punctatostriatus.Several species of small size (5–10 mm) were recorded in rocky and sandy substrates: PageBreakPageBreakPageBreakPageBreakParapholas calva was found only in sedimentary rocks; all other species were found in two or more types of hard substrates.More species were recorded with the quadrat-transect technique in the shallow subtidal (64 species) than with either dredges (42 species) or direct searches (38 species). However, the species composition was different since some were collected only with quadrat-transects (10 species), dredges (7) or direct searches (1). The life-forms recorded included epifaunal (27), infaunal (26 species), endolithic (20), and semi-infaunal (16). Endolithic species were found in various hard substrates such as sedimentary rocks, corals, polychaete tubes, bivalve shells, and rodoliths. The rock-drilling bivalve Lithophaga hastasia, Adula soleniformis, Mactrellona subalata, and Strigilla ervilia.Compared to previous studies in the region, this study includes 37 new records for Bahía de Mazatlán , and geoAcar rostae, Carditamera affinis, Gregariella coarctata, Hiatella arctica, Lithophaga aristata, Lithophaga plumula, Leiosolenus spatiosus, Ostrea conchaphila, Saccostrea palmula, Striostrea prismatica, and Sphenia fragilis. However, a large number of species (27) were unique to one environment and sampling site. Only nine species were recorded in the three environments and in five or six sampling sites: Acar rostae, Arcopsis solida, Brachidontes adamsianus, Carditamera affinis, Isognomon janus, Ostrea conchaphila, Plicatulostrea anomioides, Saccostrea palmula, and Striostrea prismatica (Twelve (13.5%) of the 89 species recorded were widely distributed in the bay , and 11 were recorded at three of the environments : ismatica .There are several distribution patterns revealed by the life forms recorded in the different environments of Bahía de Mazatlán. Epifaunal bivalves were more frequent in the upper and lower rocky intertidal (12–17 species), followed by the endolithic species (1–8), whereas the number of infaunal species was very similar in all six sites (4–5), except for Venados Island. Similarly, epifaunal species dominated the shallow subtidal and intertidal zones (10–22), followed by the endolithic (7–14), infaunal (3–8) and semi-infaunal species (2–10) .+ for Los Pinos and Olas Altas fell within the probability funnel , indicating a greater contribution to the mean taxonomic diversity of Bahía Mazatlan. However, values of Λ+ fell within the probability funnel for the four sites, suggesting that these are significantly representative of the bay’s bivalve assemblage , except for Olas Altas upper intertidal zone and Los Pinos and Venados Island lower PageBreakPageBreakPageBreakPageBreakPageBreakPageBreakintertidal zone  the inventory of bivalve species of Bahía de Mazatlán to an updated total number of 132 species, including 37 new records . Accordiroximata ; and theroximata .PageBreakthis size range are common. Therefore, it was important to address these factors for a more complete inventory of bivalves.Our surveys yielded a substantial increase in the number of infaunal (29%) and endolithic (23%) species of bivalves; most of them (67%) not recorded previously in the bay. This is particularly important since frequently the species richness of molluscs has been underestimated in ecological investigations due to two main factors that, alone or combined, contribute to incomplete inventories . The firLithophaga hastasia ; Strigilla ervilia ; Mactrellona subalata ; and Adula soleniformis   .A total of 83 additional species were collected during field work; these are not reported here because they were not living specimens however they were identified from complete and well preserved shells. Interestingly, most of these species (64) have not been recorded previously in the bay, thus raising the total inventory (living specimens plus empty shells) to 196 species. Many ecological investigations include the species recorded from empty mollusc shells assuming that they are components of the regional community i.e., . Howeverconfer which means “compare with”, that is, similar to and probably the same as, the parent taxon) are worth mentioning. These specimens corresponded to juvenile stages , which restricts their taxonomic determination because the keys and photographs in the literature consistently refer to adult specimens  and 57%  higher than the number of species we actually collected. This difference relates to the large number of rare species recorded and it is a good estimator of the potential number of species expected in these environments at Bahía de Mazatlán. Even so, the species accumulation curves confirmed that our sampling effort was sufficient to calculate the theoretical total number of bivalve species in the bay.Although different sampling techniques were used in the bay’s different environments, the sampling effort was estimated only for the quadrat-transect technique. Thus, whether all the bivalve species that inhabit the bay were collected in this study was not satisfactorily demonstrated. Some bivalves may be present either only in some seasons or impermanently, so these will not be recorded irrespective of the sampling intensity, which in turn is reflected in the sampling effort outcome , 3.PageBreakrichness in Golfo de California has been documented by The high marine biodiversity of Golfo de California has been related to its irregular coastal geomorphology , the local dynamics of the surface currents and the seabed heterogeneity . AccordiPageBreakare related to substrate heterogeneity, wave exposure and particle size of the sediments in the intertidal and shallow subtidal environments  and its variation (Λ+). Their inclusion within the probability funnel indicates that they involve a good representativeness of the bay’s taxonomic diversity.Our analysis combined data from three different sampling techniques, which was a major advantage, as the average taxonomic distinctness analysis is not affected by the various techniques and sampling effort used . With th+ and Λ+ when rocky shores and shallow subtidal adjacent zones were taken into consideration. This is because, although the three environments are clearly different from each other, all the sites contribute towards the taxonomic diversity of the bay. For example, Venados Island and Casa del Marino had the highest number of taxa in the shallow subtidal zone; Los Pinos and Venados Island, in the lower intertidal zone; and Olas Altas, in the upper intertidal zone. Theoretically, populations with a high genetic diversity have a high evolutionary potential or ability to adapt to changing environmental conditions .The present work demonstrates that the bivalve fauna in Bahía de Mazatlán is well represented by various life forms  in all the sites studied. Venados Island is an area protected by two government agencies; this is significant because it displayed high species richness and a large number of unique species. Since the bay is now a popular destination for tourists, efforts to preserve its ecosystems and species are essential, including those bivalves of economic importance such as the rock oyster PageBreaktermine their relative distribution at different spatial scales. A quantitative analysis investigating the relationship between bivalve assemblage structure and local and seasonal environmental parameters is also required. Such an analysis, would contribute to a comprehensive framework on the ecology of these bivalves, which is essential for further studies on the conservation of the bay.The information on bivalve assemblages in Bahía de Mazatlán should be supplemented with analysis including an assessment of α, γ, and β diversity in order to de


To the Editor: Human granulocytic anaplasmosis is a tick-borne disease caused by Anaplasma phagocytophilum, an obligate intracellular gram-negative bacterium that infects granulocytes. The usual clinical signs and symptoms include nonspecific fever, chills, headache, and myalgia. Infection is usually mild or asymptomatic, but severe systemic complications can occur, leading to a need for intensive care and estimated fatality rates of 0.5%–1.0%  and other nonspecific symptoms such as malaise, myalgia, and severe headache 3 days after becoming aware of several tick bites received while on a hiking vacation in Scotland. The man had removed the 3 tick nymphs from his legs immediately after their discovery and stored them in a plastic container; they were later sent to the Consultant Laboratory for Tick-borne Encephalitis in Berlin, Germany, for analysis. When the patient returned to Germany, 5 days after the onset of symptoms and 8 days after tick removal, a blood sample was collected (sample 1) and the bite sites were swabbed with a sterile cotton bud. By that time, the fever was gone, but malaise and other symptoms persisted. The patient began taking doxycycline, and within 2 days all symptoms subsided and the patient recovered completely.A second blood sample was collected 28 days after tick removal (sample 2). Complete blood counts and chemistry panels were performed for both samples. All values were within the reference range except that for lactate dehydrogenase (248 U/L), which was moderately increased over the baseline value of <245 U/L in sample 1. Values did not differ substantially between the 2 samples.A. phagocytophilum, Babesia spp., Borrelia spp., and Rickettsia spp. by using commercially available rapidSTRIPE assays for Anaplasma, Babesia, Borrelia, Rickettsia . DNA extracted from blood and swab samples was negative for all tested pathogens.DNA from whole-blood samples and swabs was extracted  and tested for I. ricinus, DNA/RNA was extracted  and tested for the same pathogens. All 3 ticks were negative for Babesia spp., Borrelia spp., and Rickettsia spp., but 2 were positive for A. phagocytophilum.After the tick specimens were taxonomically identified as A. phagocytophilum–specific IgM titer, from 20 at 5 days after symptom onset to 80 at 20 days later; the A. phagocytophilum–specific IgG titer rose from a high titer of 800 to >3,200 over this period.Indirect immunofluorescence assays  performed on the paired serum samples revealed an increased A. phagocytophilum–specific IgM and IgG in paired serum samples confirmed the diagnosis of human granulocytic anaplasmosis in accordance with Centers for Disease Control and Prevention criteria (A. phagocytophilum DNA in 2 of the 3 ticks removed from the patient’s skin. PCR amplification failed to detect A. phagocytophilum DNA in the patient’s blood, consistent with previous studies documenting frequent lack of A. phagocytophilum DNA detection in whole blood and a substantial drop in PCR positivity after the acute phase of illness (The presence and 4-fold increase of Human granulocytic anaplasmosis is not usually reported in Scotland like it is in the rest of Europe. The case originated from an area with long-established disease occurrence in ruminants, but the literature reports only 1 case of human infection in southwestern Scotland (


Proteus syndrome (PS) is an extremely rare and complex disease characterized by malformations and overgrowth of different tissues. Prognosis of affected patients may be complicated by premature death, mostly due to pulmonary embolism and respiratory failure. To date, immunological data in Proteus syndrome are scarse.We report on the novel immunologic findings of a 15 years old girl affected with PS. Detailed T and B cell evaluation revealed maturational alterations for both subsets and functional hyperactivation for the latter. Such findings have not been reported previously in PS and may be the spy of more complex immune abnormalities in this syndrome. The Proteus syndrome (PS) is an extremely rare and complex disease characterized by malformations and overgrowth of various tissues, mainly connective tissue, bone, skin, and central nervous system, although any tissue may be involved –6. CliniThe syndrome has an incidence of less than 1 per 1,000.000 live births and is estimated that 120 individuals with PS are currently alive worldwide –5. NewboPublished immunological data for PS are scarse: in fact, only one patient has been reported to date with mild hypogammaglobulinemia and lymphopenia leading to reduction of total T and B cell numbers .left panel) with greater growth of the calcaneus and cuboid, especially of the left foot . The patient also presented severe kyphoscoliosis . Diagnosis of PS was made at the age of 2 years, with progressive overgrowth over time. The index patient presented multiple lesions affecting the central nervous and circulatory system, lung, skeleton, limbs and abdominal organs. The phenotypic examination showed dolichocephaly, hyperostosis of the skull in the right fronto-parietal area, facial dysmorphic features with facial asymmetry and ptosis of right eye, depressed nasal bridge, wide and anteverted nares, oligodontia and multiple caries, long neck. The patient presented thin skin, adipose dysregulation with poorly represented subcutaneous adipose and a disproportionate, asymmetric overgrowth of the limbs. The index patient presented important overgrowth of the feet  and recent thymic emigrants (RTE) (CD45RA+CCR7+CD31+) CD4 T cell percentages, an increase in the Terminally differentiated CD45RA+CCR7− CD4 T cells and a reduction of the CD8 T cells (Table hiCD21dim/loCD27−) subset and in the IgM memory subset (IgD+CD27+), while all other subsets appeared normally distributed in terms of percentages  Ethical Committee). Written informed consent was obtained from the patient's parent for the publication of this report and any accompanying images.