Histological changes induced by Polyglycolic-Acid (PGA) scaffolds seeded with autologous adipose or muscle-derived stem cells when implanted on rabbit bladder

Purpose: To evaluate the morphological and histological changes induced by PGA scaffold seeded with autologous adipose or muscle derived stem cells implanted on rabbit bladder wall. Material and Methods: Adipose derived stem cells (ADSCs) were obtained from the inguinal fat of eight rabbits and muscle derived stem cells (MDSCs) from the anterior tibial muscle of other eight rabbits. After culture and isolation, the cells were stained with Vybrant Red CM DiI and then implanted at third passage. Two PGA scaffolds were implanted on the bladder submucosa of each animal. On the right bladder side was implanted unseeded PGA scaffold while on the left side was implanted ADSCs or skeletal MDSCs seeded PGA scaffold. ADSCs were implanted in eight animals and MDSC in other eight animals. The animals were sacrificed at four and eight weeks. Histological evaluation was performed with Hematoxylin and Eosin, Masson's Trichrome and smooth muscle α-actin. Results: We observed a mild inflammatory response in all the three groups. Seeded scaffolds induced higher lymphocytes and lower polimorphonuclear migration than controls. Fibrosis was more pronounced in the control groups. Smooth muscle α-actin was positive only in ADSC and MDSC seeded scaffolds. At four and eight weeks ADCSs and skeletal MDSCs labeled cells were found at the implant sites. Conclusions: The implantation of PGA scaffolds seeded with ADSC and MDSC induced less fibrosis than control and smooth muscle regeneration.     

Focus Issue on Roots: Duplicate and Conquer: Multiple Homologs of PHOSPHORUS-STARVATION TOLERANCE1 Enhance Phosphorus Acquisition and Sorghum Performance on Low-Phosphorus Soils

Low soil phosphorus (P) availability is a major constraint for crop production in tropical regions. The rice (Oryza sativa) protein kinase, PHOSPHORUS-STARVATION TOLERANCE1 (OsPSTOL1), was previously shown to enhance P acquisition and grain yield in rice under P deficiency. We investigated the role of homologs of OsPSTOL1 in sorghum (Sorghum bicolor) performance under low P. Association mapping was undertaken in two sorghum association panels phenotyped for P uptake, root system morphology and architecture in hydroponics and grain yield and biomass accumulation under low-P conditions, in Brazil and/or in Mali. Root length and root surface area were positively correlated with grain yield under low P in the soil, emphasizing the importance of P acquisition efficiency in sorghum adaptation to low-P availability. SbPSTOL1 alleles reducing root diameter were associated with enhanced P uptake under low P in hydroponics, whereas Sb03g006765 and Sb03g0031680 alleles increasing root surface area also increased grain yield in a low-P soil. SbPSTOL1 genes colocalized with quantitative trait loci for traits underlying root morphology and dry weight accumulation under low P via linkage mapping. Consistent allelic effects for enhanced sorghum performance under low P between association panels, including enhanced grain yield under low P in the soil in Brazil, point toward a relatively stable role for Sb03g006765 across genetic backgrounds and environmental conditions. This study indicates that multiple SbPSTOL1 genes have a more general role in the root system, not only enhancing root morphology traits but also changing root system architecture, which leads to grain yield gain under low-P availability in the soil.Increasing food production is one of the major global challenges in dealing with continuously growing population and food consumption (Godfray et al., 2010). One of the major obstacles to improve crop production in tropical regions is phosphorus (P) deficiency caused by P fixation in the soil clays. P is one of the most important plant nutrients, contributing approximately 0.2% of a plant’s dry weight, and is a component of key organic molecules such as nucleic acids, phospholipids, and ATP (Schachtman et al., 1998). On tropical soils, even when the total amount of soil P is high, its bioavailability is low due to P fixation by aluminum and iron oxides in clay minerals (Marschner, 1995) and immobilization into organic forms (Schachtman et al., 1998). Approximately half of the world’s agricultural lands occurs on low-P soils (Lynch, 2011); hence, crop adaptation to P insufficiency should be a major breeding target to enable sustainable agricultural production worldwide. In addition, because phosphate rock fertilizer is a nonrenewable resource that is being depleted by agricultural demands, increasing fertilizer prices negatively impact agriculture, particularly for small-holder farmers in developing countries in the tropics and subtropics (Cordell et al., 2009; Sattari et al., 2012). In sorghum (Sorghum bicolor), breeding strategies for low-P adaptation have been developed based on multienvironment trials in West Africa, indicating the importance of undertaking selection in low-P soil conditions (Leiser et al., 2012a, 2012b). Therefore, developing crops with greater ability to grow and maintain satisfactory yields on soils with reduced P availability is expected to substantially improve food security worldwide.Tolerance to P deficiency in plants can be achieved by mechanisms underlying both P acquisition and P internal utilization efficiency (Parentoni and Souza Junior, 2008). One of the major mechanisms that plants have evolved to overcome low-P availability is to maximize the ability of the roots to acquire and absorb P from the soil. Plants can mobilize P through the exudation of organic acids, acid phosphatases, and ribonucleases, resulting in enhanced P availability and uptake (Hinsinger, 2001; Ryan et al., 2001; Dakora and Phillips, 2002; Hammond and White, 2008; Ma et al., 2009; Pang et al., 2009). Another strategy to cope with low-P availability is to increase the soil volume accessed by root systems by forming mycorrhizal symbioses (Li et al., 2012; Smith and Smith, 2012; Rai et al., 2013). Due to low-P mobility on tropical soils, changes in root architecture and morphology enhance P uptake by facilitating soil exploration (Williamson et al., 2001; Ho et al., 2005; Walk et al., 2006; Svistoonoff et al., 2007; Lynch, 2011; Ingram et al., 2012; Niu et al., 2013). Root structural changes leading to higher P uptake include increased root hair growth (Yan et al., 2004; Haling et al., 2013; Lan et al., 2013) and length and enhancing lateral root over primary root growth (Williamson et al., 2001; Wang et al., 2013). In addition, increased root surface area is achieved by a combination of reduced root diameter and enhanced elongation of relatively thinner roots (Fitter et al., 2002). There is both intraspecific and interspecific genetic variation for P deficiency tolerance in crop species (Lynch and Brown, 2001, 2012; Mudge et al., 2002; Paszkowski et al., 2002; Rausch and Bucher, 2002; Huang et al., 2011; Zhang et al., 2011; Leiser et al., 2014a) that can be explored to develop P-efficient cultivars.In rice (Oryza sativa), Phosphorus uptake1 (Pup1), a major quantitative trait locus (QTL) for P deficiency tolerance donated by an aus-type Indian variety, Kasalath, was mapped to the long arm of chromosome 12 (Ni et al., 1998; Wissuwa et al., 1998, 2002; Heuer et al., 2009). Near-isogenic lines bearing the Kasalath allele at Pup1 showed 3-fold higher P uptake and grain yield in low-P trials compared with the recurrent parent, cv Nipponbare, which is intolerant to P starvation (Wissuwa and Ae, 2001). Following high-resolution mapping of Pup1, comparative sequence analyses of homologous bacterial artificial chromosomes showed that a Kasalath genomic fragment contained several genes not present in cv Nipponbare, highlighting an approximately 90-kb deletion in the cv Nipponbare reference genome that encompassed the Pup1 locus (Heuer et al., 2009). Within this insertion/deletion, OsPupK46-2, a gene encoding a Ser/Thr kinase of the Receptor-like Protein Kinase LRK10L-2 subfamily, was found to enhance grain yield and P uptake in rice lines overexpressing this gene, indicating that this protein kinase underlies the Pup1 locus (Gamuyao et al., 2012). OsPupK46-2, which is now designated PHOSPHORUS-STARVATION TOLERANCE1 (OsPSTOL1), was found to be up-regulated in the root tissues of tolerant near-isogenic lines under P-deficient conditions and was shown to increase P uptake by a physiological mechanism based on the enhancement of early root growth and development. Furthermore, lines overexpressing OsPupK46-2 showed an approximately 30% grain yield increase in comparison with the null lines, suggesting that PSTOL1 has potential for molecular breeding applications to improve crop performance under low-P conditions. Consistent with the proposed physiological mechanism underlying OsPSTOL1, the superior performance of the transgenic lines was related to enhanced root dry weight, root length, and root surface area (Gamuyao et al., 2012).Sorghum is the world’s fifth most important cereal crop and is a staple food for more than half a billion people. It is widely adapted to harsh environmental conditions, and more specifically, to arid and semiarid regions of the world (Doumbia et al., 1993, 1998). It has been estimated that rice diverged from its most recent common ancestor with sorghum and maize (Zea mays) approximately 50 million years ago (Kellogg, 1998; Paterson et al., 2000, 2004; Paterson, 2008). About 60% of the genes in the sorghum genome are located in syntenic regions to rice (Paterson et al., 2009), emphasizing the potential for using comparative genomics for cross-species identification of genes underlying abiotic stress tolerance in the grass family. Here, we applied association analysis to specifically study the role of sorghum homologs of rice OsPSTOL1 in tolerance to P starvation in sorghum. Single-nucleotide polymorphisms (SNPs) within PSTOL1 homologs in sorghum, collectively designated SbPSTOL1, were significantly associated with grain yield under low-P conditions and also root morphology and root system architecture (RSA) traits phenotyped from hydroponically grown plants. Under low P, SbPSTOL1 genes increased biomass accumulation and P content in the African landrace panel and grain yield in the sorghum association panel phenotyped in a low-P Brazilian soil. This suggests a stable effect across environments and sorghum genotypes that potentially can be used for molecular breeding applications. QTL mapping with a large sorghum recombinant inbred line population was used to validate the association results, indicating that SbPSTOL1 homologs colocalize with QTLs related to root morphology and performance under low P. Our results indicate that SbPSTOL1 homologs have the ability to enhance P uptake and sorghum performance in low-P soils by a mechanism related not only to early root growth enhancement, as was the case for rice OsPSTOL1, but also by modulating RSA.     

Nitric oxide increases tolerance responses to moderate water deficit in leaves of Phaseolus vulgaris and Vigna unguiculata bean species

Drought stress is one of the most intensively studied and widespread constraints, and nitric oxide (NO) is a key signaling molecule involved in the mediation of abiotic stresses in plants. We demonstrated that a sprayed solution of NO from donor sodium nitroprusside increased drought stress tolerance responses in both sensitive (Phaseolus vulgaris) and tolerant (Vigna unguiculata) beans. In intact plants subjected to halting irrigation, NO increased the leaf relative water content and stomatal conductance in both species. After cutting leaf discs and washing them, NO induced increased electrolyte leakage, which was more evident in the tolerant species. These leaf discs were then subjected to different water deficits, simulating moderate and severe drought stress conditions through polyethylene glycol solutions. NO supplied at moderate drought stress revealed a reduced membrane injury index in sensitive species. In hydrated discs and at this level of water deficit, NO increased the electron transport rate in both species, and a reduction of these rates was observed at severe stress levels. Taken together, it can be shown that NO has an effective role in ameliorating drought stress effects, activating tolerance responses at moderate water deficit levels and in both bean species which present differential drought tolerance.     

Seroprevalence and Spatial Epidemiology of Lymphatic Filariasis in American Samoa after Successful Mass Drug Administration

Background

As part of the Global Programme to Eliminate Lymphatic Filariasis (LF), American Samoa conducted mass drug administration (MDA) from 2000–2006, and passed transmission assessment surveys in 2011–2012. We examined the seroprevalence and spatial epidemiology of LF post-MDA to inform strategies for ongoing surveillance and to reduce resurgence risk.

Methods

ELISA for LF antigen (Og4C3) and antibodies (Wb123, Bm14) were performed on a geo-referenced serum bank of 807 adults collected in 2010. Risk factors assessed for association with sero-positivity included age, sex, years lived in American Samoa, and occupation. Geographic clustering of serological indicators was investigated to identify spatial dependence and household-level clustering.

Results

Og4C3 antigen of >128 units (positive) were found in 0.75% (95% CI 0.3–1.6%) of participants, and >32 units (equivocal plus positive) in 3.2% (95% CI 0.6–4.7%). Seroprevalence of Wb123 and Bm14 antibodies were 8.1% (95% CI 6.3–10.2%) and 17.9% (95% CI 15.3–20.7%) respectively. Antigen-positive individuals were identified in all ages, and antibody prevalence higher in older ages. Prevalence was higher in males, and inversely associated with years lived in American Samoa. Spatial distribution of individuals varied significantly with positive and equivocal levels of Og4C3 antigen, but not with antibodies. Using Og4C3 cutoff points of >128 units and >32 units, average cluster sizes were 1,242 m and 1,498 m, and geographical proximity of households explained 85% and 62% of the spatial variation respectively.

Conclusions

High-risk populations for LF in American Samoa include adult males and recent migrants. We identified locations and estimated the size of possible residual foci of antigen-positive adults, demonstrating the value of spatial analysis in post-MDA surveillance. Strategies to monitor cluster residents and high-risk groups are needed to reduce resurgence risk. Further research is required to quantify factors contributing to LF transmission at the last stages of elimination to ensure that programme achievements are sustained.     

Early Standard Electroencephalogram Abnormalities Predict Mortality in Septic Intensive Care Unit Patients

IntroductionSepsis is associated with increased mortality, delirium and long-term cognitive impairment in intensive care unit (ICU) patients. Electroencephalogram (EEG) abnormalities occurring at the acute stage of sepsis may correlate with severity of brain dysfunction. Predictive value of early standard EEG abnormalities for mortality in ICU septic patients remains to be assessed.MethodsIn this prospective, single center, observational study, standard EEG was performed, analyzed and classified according to both Synek and Young EEG scales, in consecutive patients acutely admitted in ICU for sepsis. Delirium, coma and the level of sedation were assessed at the time of EEG recording; and duration of sedation, occurrence of in-ICU delirium or death were assessed during follow-up. Adjusted analyses were carried out using multiple logistic regression.ResultsOne hundred ten patients were included, mean age 63.8 (±18.1) years, median SAPS-II score 38 (29–55). At the time of EEG recording, 46 patients (42%) were sedated and 22 (20%) suffered from delirium. Overall, 54 patients (49%) developed delirium, of which 32 (29%) in the days after EEG recording. 23 (21%) patients died in the ICU. Absence of EEG reactivity was observed in 27 patients (25%), periodic discharges (PDs) in 21 (19%) and electrographic seizures (ESZ) in 17 (15%). ICU mortality was independently associated with a delta-predominant background (OR: 3.36; 95% CI [1.08 to 10.4]), absence of EEG reactivity (OR: 4.44; 95% CI [1.37–14.3], PDs (OR: 3.24; 95% CI [1.03 to 10.2]), Synek grade ≥ 3 (OR: 5.35; 95% CI [1.66–17.2]) and Young grade > 1 (OR: 3.44; 95% CI [1.09–10.8]) after adjustment to Simplified Acute Physiology Score (SAPS-II) at admission and level of sedation. Delirium at the time of EEG was associated with ESZ in non-sedated patients (32% vs 10%, p = 0.037); with Synek grade ≥ 3 (36% vs 7%, p< 0.05) and Young grade > 1 (36% vs 17%, p< 0.001). Occurrence of delirium in the days after EEG was associated with a delta-predominant background (48% vs 15%, p = 0.001); absence of reactivity (39% vs 10%, p = 0.003), Synek grade ≥ 3 (42% vs 17%, p = 0.001) and Young grade >1 (58% vs 17%, p = 0.0001).ConclusionsIn this prospective cohort of 110 septic ICU patients, early standard EEG was significantly disturbed. Absence of EEG reactivity, a delta-predominant background, PDs, Synek grade ≥ 3 and Young grade > 1 at day 1 to 3 following admission were independent predictors of ICU mortality and were associated with occurence of delirium. ESZ and PDs, found in about 20% of our patients. Their prevalence could have been higher, with a still higher predictive value, if they had been diagnosed more thoroughly using continuous EEG.     

Incomplete transfer of accessory loci influencing SbMATE expression underlies genetic background effects for aluminum tolerance in sorghum

Impaired root development caused by aluminum (Al) toxicity is a major cause of grain yield reduction in crops cultivated on acid soils, which are widespread worldwide. In sorghum, the major Al‐tolerance locus, Alt_SB, is due to the function of SbMATE, which is an Al‐activated root citrate transporter. Here we performed a molecular and physiological characterization of various Alt_SB donors and near‐isogenic lines harboring various Alt_SB alleles. We observed a partial transfer of Al tolerance from the parents to the near‐isogenic lines that was consistent across donor alleles, emphasizing the occurrence of strong genetic background effects related to Alt_SB. This reduction in tolerance was variable, with a 20% reduction being observed when highly Al‐tolerant lines were the Alt_SB donors, and a reduction as great as 70% when other Alt_SB alleles were introgressed. This reduction in Al tolerance was closely correlated with a reduction in SbMATE expression in near‐isogenic lines, suggesting incomplete transfer of loci acting in trans on SbMATE. Nevertheless, Alt_SB alleles from the highly Al‐tolerant sources SC283 and SC566 were found to retain high SbMATE expression, presumably via elements present within or near the Alt_SB locus, resulting in significant transfer of the Al‐tolerance phenotype to the derived near‐isogenic lines. Allelic effects could not be explained by coding region polymorphisms, although occasional mutations may affect Al tolerance. Finally, we report on the extensive occurrence of alternative splicing for SbMATE, which may be an important component regulating SbMATE expression in sorghum by means of the nonsense‐mediated RNA decay pathway.     

Comparative proteomics between natural Microcystis isolates with a focus on microcystin synthesis

ABSTRACT: BACKGROUND: Microcystis aeruginosa is a species of cyanobacteria commonly found in a number of countries and frequently related to animal poisoning episodes due to its capacity to produce the cyanotoxin known as microcystin. Despite vast literature on microcystin structures and their deleterious effects, little is known about its synthesis by cyanobacteria. Therefore, this study used proteomic tools to compare two M. aeruginosa strains, contrasting them for microcystin production. RESULTS: 2-DE gels were performed and 30 differential protein spots were chosen. Among them, 11 protein spots were unique in the toxin producing strain and 8 in the non-toxin producing strain, and 14 protein spots were shown on both 2-DE gels but expressed differently in intensity. Around 57% of the tandem mass spectrometry identified proteins were related to energy metabolism, with these proteins being up-regulated in the toxin producing strain. CONCLUSIONS: These data suggest that the presence of higher quantities of metabolic enzymes could be related to microcystin metabolism in comparison to the non-toxin producing strain. Moreover, it was suggested that the production of microcystin could also be related to other proteins than those directly involved in its production, such as the enzymes involved in the Calvin cycle and glycolysis.