High Acid Phosphatase Activity Research Articles

The effect of Astragalus polysaccharides on the repair of adverse effects in largemouth bass (Micropterus salmoides) under enrofloxacin stress

Antibiotics have important role in controlling the outbreak of bacterial diseases in fish farming, but long-term use or persistent existence of antibiotics in water can cause adverse effects on aquatic animals. In order to evaluate repairing effect of Astragalus polysaccharides (APS) on damaged intestine of fish under enrofloxacin (ENR) stress, ENR-pretreated (1.0 mg/L, 7 days) largemouth bass were randomly divided into two groups, and subsequently were fed basal diet and diet supplemented with 1.0 g/kg APS for 14 days. Intestine tissue and content were sampled on day 0 (D0), days 7 (D7), days 21 (D21_APS and D21) to detect the intestinal structure, intestinal microbiota structure, oxidative stress, non-specific immune and inflammatory response.Intestinal tissue section showed that the height, width of intestinal villi, and muscular thickness in D21_APS were significantly greater than those on D0, D7, and D21 (P < 0.05). APS-supplemented diet was significantly increased expression of tight junction protein related genes occludin, claudin-1, ZO-1 (P < 0.05). APS addition significantly improved the activities of catalase (CAT) and peroxidase (POD) and decreased superoxide dismutase (SOD) activity (P < 0.05). The activity of CAT and SOD in D21_APS and D0 were similar. The highest activities of acid phosphatase (ACP), lysozyme (LZM) and alkaline phosphatase (AKP) were found in D21_APS group. The expression of proinflammatory factors IL-1β and TNF-α in D21_APS and D21 were significantly down-regulated compared to that of D7, while anti-inflammatory factor IL-10 was up-regulated (P < 0.05). Compared with D7 group, quinolone resistance gene (qnrB) in D21 and D21_APS groups were significantly down-regulated, and in contrast to D21 group, qnrB and qnrS genes in D21_APS group were significantly down-regulated (P < 0.05).High-throughput 16SrRNA gene sequencing revealed proportion of Cyanobacteria in intestinal microbiota increased from 8.23% to 52.66%, while Firmicutes decreased from 28.54% to 2.03% after ENR treatment. In D21 group, Proteobacteria was notable decrease compared to D0, while Firmicutes increased. In D21_APS, the dominant bacteria were Proteobacteria, Fusobacteriota and Firmicutes, and Acinetobacter, Cetobacterium and Aeromonas were the predominant genera. Cluster analysis indicated that bacterial community structure in D21_APS has a similar level to D0 at phylum level. Feeding APS increased abundance of genes related to styrene, isoleucine and lysine degradation; propanoate, butanoate phenylalanine, and tryptophan metabolism; lipopolysaccharide biosynthesis; bacterial secretion system; cell motility and secretion at KEGG pathway Level 3 compared to the basal diet. Taken together, APS can repair the damage of intestinal structure and intestinal microbiota caused by ENR.

Phosphorus absorption kinetics and exudation strategies of roots developed by three lupin species to tackle P deficiency.

Different lupin species exhibited varied biomass, P allocation, and physiological responses to P-deprivation. White and yellow lupins had higher carboxylate exudation rates, while blue lupin showed the highest phosphatase activity. White lupin (Lupinus albus) can produce specialized root structures, called cluster roots, which are adapted to low-phosphorus (P) soil. Blue lupin (L. angustifolius) and yellow lupin (L. luteus), which are two close relatives of white lupin, do not produce cluster roots. This study characterized plant responses to nutrient limitation by analyzing biomass accumulation and P distribution, absorption kinetics and root exudation in white, blue, and yellow lupins. Plants were grown in hydroponic culture with (64µM NaH2PO4) or without P for 31days. Under P limitation, more biomass was allocated to roots to improve P absorption. Furthermore, the relative growth rate of blue lupin showed the strongest inhibition. Under + P conditions, the plant total-P contents of blue lupin and yellow lupin were higher than that of white lupin. To elucidate the responses of lupins via the perspective of absorption kinetics and secretion analysis, blue and yellow lupins were confirmed to have stronger affinity and absorption capacity for orthophosphate after P-deprivation cultivation, whereas white lupin and yellow lupin had greater ability to secrete organic acids. The exudation of blue lupin had higher acid phosphatase activity. This study elucidated that blue lupin was more sensitive to P-scarcity stress and yellow had the greater tolerance of P-deficient condition than either of the other two lupin species. The three lupin species have evolved different adaptation strategies to cope with P deficiency.

Multi-omics analysis reveals the roles of purple acid phosphatases in organic phosphorus utilization by the tropical legume Stylosanthes guianensis.

Stylo (Stylosanthes guianensis) is a tropical legume known for its exceptional tolerance to low phosphate (Pi), a trait believed to be linked to its high acid phosphatase (APase) activity. Previous studies have observed genotypic variations in APase activity in stylo; however, the gene encoding the crucial APase responsible for this variation remains unidentified. In this study, transcriptomic and proteomic analyses were employed to identify eight Pi starvation-inducible (PSI) APases belonging to the purple APase (PAP) family in the roots of stylo and seven in the leaves. Among these PSI-PAPs, SgPAP7 exhibited a significantly positive correlation in its expression levels with the activities of both internal APase and root-associated APase across 20 stylo genotypes under low-Pi conditions. Furthermore, the recombinant SgPAP7 displayed high catalytic activity toward adenosine 5'-diphosphate (ADP) and phosphoenolpyruvate (PEP) in vitro. Overexpression (OE) of SgPAP7 in Arabidopsis facilitated exogenous organic phosphorus utilization. Moreover, SgPAP7 OE lines showed lower shoot ADP and PEP levels than the wild type, implying that SgPAP7 is involved in the catabolism and recycling of endogenous ADP and PEP, which could be beneficial for plant growth in low-Pi soils. In conclusion, SgPAP7 is a key gene with a major role in stylo adaptation to low-Pi conditions by facilitating the utilization of both exogenous and endogenous organic phosphorus sources. It may also function as a PEP phosphatase involved in a glycolytic bypass pathway that minimizes the need for adenylates and Pi. Thus, SgPAP7 could be a promising target for improving tolerance of crops to low-Pi availability.

Exploring the Phosphate Solubilizing Potential and Acid-Alkaline Phosphatase Activity of Two Multitrait Heavy Metal Tolerant Bacterial Strains Isolated from North Gujarat, India

A study was conducted to isolate and characterize phosphate solubilizing bacteria (PSB) from soils of various locations in North Gujarat. A total of 40 PSB isolates were obtained, exhibiting a phosphate solubilizing index (PSI) ranging from 2.11 to 4.33. Interestingly, among 40 PSB isolates, a subset of 16 displayed an additional trait of potassium solubilization, as evidenced by their potassium solubilizing index (KSI) values falling within the range of 2.10 to 3.10. These 16 PSB isolates were screened for tolerance to heavy metal stressors, namely CdCl2, HgCl2, Pb(NO3)2, and ZnSO4, spanning a concentration gradient from 0.1 mM to 10 mM. Remarkably, among the tested isolates, two, namely, D4 and G2, exhibited exceptional resilience against the heavy metal stressors, as indicated by their minimum inhibitory concentration (MIC). Specifically, isolate D4 showed MIC values of 5 mM, 0.2 mM, 18 mM, and 12 mM, respectively, against CdCl2, HgCl2, Pb(NO3)2, and ZnSO4. For isolate G2, the corresponding MIC values were 6 mM, 0.2 mM, 21 mM, and 11 mM. The bacterial isolates were molecularly identified as strains Klebsiella pneumoniae PHmD4 (abbreviated as D4) and Enterobacter chuandaensis PHmG2 (abbreviated as G2), based on sequence homology obtained from 16S rDNA gene sequencing and subsequent BLAST analysis. In Pikovskaya’s broth, K. pneumoniae PHmD4 and E. chuandaensis PHmG2 had phosphate solubilization values of 4.30 μ moles mL⁻¹ and 3.51 μmoles mL⁻¹, respectively, with terminal pH values of 3.06 and 4.07. Enzymatic activity assays conducted after 96 hours of incubation in Pikovskaya’s broth revealed that the highest acid phosphatase activity was 21.62 U mL⁻¹ for K. pneumoniae PHmD4 and 21.09 U mL⁻¹ for E. chuandaensis PHmG2. The highest alkaline phosphatase activity was 5.85 U mL⁻¹ and 6.38 U mL⁻¹, respectively. In the tomato seed germination study, both non-bacterized and bacterized tomato seeds achieved a 100% germination rate. However, increasing metal concentrations diminished the germination percentage of heavy metal-treated seeds. Interestingly, bacterization with K. pneumoniae PHmD4 and E. chuandaensis PHmG2 ameliorated germination under cadmium stress but had no mitigative effect under mercury and zinc stress conditions.

Crop domestication disrupts intercropping benefits: A case study from barley-faba bean mixture under contrasting P inputs.

How crop domestication mediates root functional traits and trait plasticity in response to neighboring plants is unclear, but it is important for selecting potential species to be grown together to facilitate P uptake. We grew two barley accessions representing a two-stage domestication process as a sole crop or mixed with faba bean under low and high P inputs. We analyzed six root functional traits associated with P acquisition and plant P uptake in five cropping treatments in two pot experiments. The spatial and temporal patterns of root acid phosphatase activity were characterized in situ with zymography at 7, 14, 21, and 28 days after sowing in a rhizobox. Under low P supply, wild barley had higher total root length (TRL), specific root length (SRL), and root branching intensity (RootBr) as well as higher activity of acid phosphatase (APase) in the rhizosphere, but lower root exudation of carboxylates and mycorrhizal colonization (MC), relative to domesticated barley. In response to neighboring faba bean, wild barley exhibited larger plasticity in all root morphological traits (TRL, SRL, and RootBr), while domesticated barley showed greater plasticity in root exudates of carboxylates and colonization by mycorrhiza. Wild barley with greater root morphology-related trait plasticity was a better match with faba bean than domesticated barley, indicated by higher P uptake benefits in wild barley/faba bean than domesticated barley/faba bean mixtures under low P supply. Our findings indicated that the domestication of barley disrupts the intercropping benefits with faba bean through the shifts of root morphological traits and their plasticity in barley. Such findings provide valuable information for barley genotype breeding and the selection of species combinations to enhance P uptake.

Distribution and Characterization of Staphylococci Isolated From Healthy Canine Skin

Despite increasing interest to study skin microbiota with progressive methods, there are almost no data on staphylococcal species distribution on skin of healthy dogs available. Therefore we decide to characterize staphylococci isolated from 8 different body sites (inner pinna, chin, nasal skin, back, axilla, abdomen, interdigital area and perianal region) of healthy canine skin. A total of 91 staphylococci were isolated from 30 dogs living in East Slovakia. Swabs of each dog were cultivated and colonies analysed using MALDI-TOF spectrometry. The vast majority of isolated staphylococci belonged to S pseudintermedius species (48%) followed by S hominis (15%) and S aureus (10%). S haemolyticus, S warneri, S epidermidis, S capitis, S xylosus, S pasteuri, S intermedius and S succinus were also isolated (<10%). The most frequent resistance in staphylococcal isolates was observed for chloramphenicol (73%) and penicillin (67%) followed by erythromycin (42%), tetracycline (26%), and oxacillin (20%). Multi-drug resistance was found in 50% of isolates. All strains were gentamicin and vancomycin sensitive and were strong or moderate biofilm producers with high acid and alkaline phosphatase activities. Over half of strains were haemolytic (57%) and produced gelatinase (54%), DNAse (84%) and lipase (64%). It seems, multiresistant biofilm forming staphylococci could be commonly detected also in healthy dogs and could probably serve as reservoir for other dogs or owners because of constant exchange of their microbiota.

Site-Effects Dominate the Plant Availability of Nutrients under Salix Species during the First Cutting Cycle

Fast-growing willows (Salix spp.) provide alternative sources of renewable energy generation, but need an adequate nutrient availability in the soil for high biomass production. In general, species mixtures can be more nutrient-efficient than pure cultures, but this is scarcely known for Salix spp. Therefore, this study evaluates the nutrient availability and P mobilization under two willow species, Salixdasyclados var. ‘Loden’ and S. schwerinii × viminalis var. ‘Tora’, grown as pure and mixed cultures at non-fertilized former arable sites in Germany (Stagnic Cambisol) and Sweden (Vertic Cambisol). The plant availability of potassium (K), magnesium (Mg) and phosphorus (P) and soil phosphatase activities in the topsoil were measured in spring of the year of planting (initial) and under 4 years-old stocks (one year after the first 3-year cutting cycle). The initial plant availability of the nutrients significantly differed between the sites and the two sampling dates at both sites. The plant availability of K and Mg was optimal to high at both sites and sampling dates, but rather low for P (after 4 years ≤5 mg P 100 g−1 soil). The plant-available P and K content in soil significantly decreased within the 4 years of willow growth at both sites. The acid and alkaline phosphatase activity in the soil of the German site (Rostock) was significantly lower after 4 years of willow growth, but differed not significantly between the two sampling dates at the Swedish site (Uppsala). Higher activity of acid phosphatase compared to alkaline phosphatase was recorded in the soils at both test sites based on the site-specific soil pH (&lt;7). The slight decrease of plant availability of P after 4 years of Salix growth in pure culture differed not significantly between the different species. Mixed growth did not decrease the plant availability of P within this period, although no significant difference in the biomass production of pure and mixed growth was observed. This was valid at both sites, and therefore, seems independent of the site-specific differences in soil and climate conditions. The general validity of the assumptions should be tested also for other species mixtures and soil conditions in the future before site-adapted growth designs can be recommended in biomass production of Salix.

Calcite in combination with olive pulp biochar reduces Ni mobility in soil and its distribution in chili plant

The presence of Ni above the permissible limit in agriculture soils poses negative effects on soil health, crop quality, and crop productivity. Surprisingly, the usage of various organic and inorganic amendments can reduce Ni mobility in the soil and its distribution in the crops. A pot experiment was conducted to elucidate the effects of olive pulp biochar (BR), calcite (CAL), and wheat straw (WS), as sole amendments and their mixtures of 50:50 ratio, added to Ni polluted soil on Ni mobility in the soil, Ni immobilization index (Ni − IMi), soil enzymatic activities, Ni distribution in parts of chili plant, Ni translocation factor and bioaccumulation factor in fruit, plant growth parameters and oxidative stress encountered by the plants. Outcomes of this pot experiment revealed that amendments raised soil pH, improved soil enzymatic activities, values of Ni − IMi, while significantly reduced bioavailable Ni fraction in the post-harvest soil. However, the highest activities of acid phosphatase, urease, catalase, and dehydrogenase by 50, 70, 239, and 111%, respectively, improvement in Ni − IMi up to 60% while 60% reduction in the bioavailable Ni fraction was observed in BR + CAL treatment, compared to control was noted. Among all amendments, the top most reduction in Ni concentrations in shoots, roots, fruit, Translocation Factor (TF), and Bioaccumulation Factor (BAF) values of fruit by 72%, 36%, 86%, 72%, and 86%, in BR + CAL treatment, compared to control. Moreover, the plants growing on BR + CAL amended Ni contaminated soil showed the topmost improvement in plant phonological parameters while encountered the least oxidative stress. Such findings refer to the prospective usage of BR + CAL at 50:50 ratio than BR, CAL, WS alone, and BR + WS as well as WS + CAL for reducing Ni mobility in the soil, improving Ni − IMi, soil enzymatic activities, plant phonological and oxidative stress while reducing Ni distribution in plant parts. Novelty statement In this experiment, it was hypothesized that amending Ni polluted soil with olive pulp biochar (BR), CAL, and WS as alone soil amendments and their combinations at 50:50 ratios can reduce Ni bioavailability in soil, Ni distribution in chili plant and oxidative stress encountered by the plants. Moreover, these amendments may improve, soil enzymatic activities, Ni immobilization index, plant phenological traits. Therefore, it was aimed to undertake useful scientific planning and research, to restore and rehabilitate the dwellings, biological resources and to minimize the sufferings of the peoples in nutrient-poor Ni contaminated soils, by improving soil health and chili productivity.

Growing Medium Type Affects Organic Fertilizer Mineralization and CNPS Microbial Enzyme Activities

Managing plant fertilization is a major concern of greenhouse growers to achieve sustainable production with growing media (GM). Organic fertilization is popular but is more difficult to control, since organic compounds need first to be mineralized by microbes. After 7, 14, 28, and 56 days of incubation, we investigated the response of microbial activities and nutrient releases from three frequently used organic fertilizers (horn and two plant-based fertilizers) in three frequently employed GM types (peat, coir, and bark). We measured pH, electrical conductivity, nutrient contents (NH4+-N, NO3−-N, PO43−-P, SO42−-S), and enzyme activities (β-1.4-glucosidase, urease, acid phosphatase, arylsulfatase). After fertilization, microbes in coir expressed all the C, N, P, and S functions studied, making related nutrients available. In peat and bark, some C, N, P, and S-related pathways were locked. Peat presented high NH4+-N and PO43−-P releases linked to high acid phosphatase and β-glucosidase activities, while bark showed high nitrification rates but weak enzyme activities. Fertilizer types modulated these responses with lower activities and nutrient releases with horn. Our results contributed to better understanding mineralization processes in GM, showing different microbial responses to fertilization. This study pointed out the necessity to look deeper into microbial functions in GM optimizing biological and physicochemical properties.

Processes at the soil–root interface determine the different responses of nutrient limitation and metal toxicity in forbs and grasses to nitrogen enrichment

Abstract Nutrient limitation and metal toxicity have been implicated in changes of grassland communities by nitrogen (N) deposition. Below‐ground processes, especially those at the soil–root interface, play important roles in determining variation in nutrient concentrations in plants. However, few studies have specifically focused on the roles of these processes in mineral‐element acquisition in grassland plants in response to N enrichment. Here we investigated the contributions of below‐ground processes at the soil–root interface to the differential acquisition of phosphorus (P), calcium (Ca) and manganese (Mn) by forbs and grasses of a temperate steppe in response to N addition by combining field and glasshouse experiments. Nitrogen addition increased the concentrations of both leaf P ([P]) and Mn ([Mn]) and decreased leaf [Ca] of forbs while it had little effects on leaf concentrations of these elements in grasses. Nitrogen addition led to a higher activity of acid phosphatase in the rhizosphere of forbs, and greater release of protons and carboxylates from forb roots than grass roots, contributing to the differential [P], [Ca] and [Mn] in the leaves of forbs and grasses. Applying oxalate to soil to simulate the release of carboxylates by N enrichment enhanced [P] and [Mn], and decreased [Ca] in the soil solution. However, addition of hydrogen‐ion increased [P], [Mn] and [Ca] in the soil solution. Lime addition mitigated the N‐addition‐induced soil acidification while it did not abolish the stimulatory effect of short‐term N addition on leaf [P] and [Mn] of forbs. Therefore, we conclude that differences in the eco‐physiological processes at the soil–root interface account for changes in leaf [P], [Ca] and [Mn] under short‐term N addition, and that soil acidification aggravates the responses of these elements, especially [Ca] and [Mn], to long‐term N enrichment. Synthesis. Our results highlight the contribution of below‐ground processes, especially those at the soil–root interface, to variation in plant element concentrations between dominant forbs and grasses in the temperate steppe. These findings greatly enhance our mechanistic understanding of the effects of N deposition on grassland communities.

Response of Soil Microbes and Soil Enzymatic Activity to 20 Years of Fertilization

Fertilization is a worldwide agricultural practice used in agronomy to increase crop yields. Fertilizer application influences overall soil characteristics, including soil microbial community composition and metabolic processes mediated by microbial enzymatic activity. Changes in the structure of microbial communities and their metabolic activity after long-term fertilization were studied in this research. We hypothesized that the different types of fertilization regimes affect nutrient levels in the soil which subsequently influence the metabolic processes and microbial diversity and community structure. Manure (MF; 330 kg N/ha), sewage sludge at two application doses (SF; 330 kg N/ha and SF3x; 990 kg N/ha) and chemical (NPK; N-P-K nutrients in concentrations of 330-90-300 kg/ha) fertilizers have been applied regularly to an experimental field since 1996. The microbial diversity increased in all soils amended with both organic (MF, SF, SF3x) and chemical (NPK) fertilizers. The shifts in microbial communities were observed, which were mainly caused by less abundant genera that were mostly associated with one or more fertilization treatment(s). Fertilization also influenced soil chemistry and the activity of β-xylosidase, β-N-acetylglucosaminidase (NAG), acid phosphatase and FDA-hydrolases. Specifically, all fertilization treatments were associated with a higher activity of β xylosidase and lower NAG activity. Only the NPK treatment was associated with a higher activity of acid phosphatase.

VILLOUS LYMPHOCYTES IN BLOOD AND BONE MARROW IN SOME FORMS OF B-CELL LYMPHOID MALIGNANCIES

The cytological and immunocytochemical features of the lymphocytes with villous morphology in peripheral blood and bone marrow in some B-lymphoproliferative disorders were studied. The diagnosis of hairy cell leukemia, a hairy cell leukemia variant, splenic marginal zone lymphoma and splenic diffuse red pulp small B-cell lymphoma was ascertained in accordance with the new revision of the WHO classification (2016). The neoplastic cells of hairy cell leukemia were determined by the presence of high tartrate resistant acid phosphatase (TRAP) activity. Cell surface expression of CD19, CD20 and CD21 antigens was detected. Also, the expression of CD25, CD103 and CD200, and in some cases cyclin D1, was found out. CD5, CD10 and CD23 were not detected. The immunophenotype of cells in splenic marginal zone lymphoma with villous processes also corresponded to the mature B cells. The expression of CD19, CD20 and CD21 was observed in all cases, CD11c – in 50% of patients, CD25 or CD5 – in 10% of patients. In 80% of patients, the pathologic cells did not show TRAP activity. In the bone marrow and peripheral blood cells of patients with diffuse red pulp lymphoma, TRAP activity was not detected. An immunophenotype in the hairy cell leukemia variant was different from those of classic HCL (CD19+CD20+CD22+CD103+CD11c+CD5–CD10–CD23–). Characterized immunophenotypical markers, which have differential diagnostic values in several forms of lymphoid tumors of B cell origin, will be important for the choice of treatment methods and prognosis

Overexpression of MdPHR1 Enhanced Tolerance to Phosphorus Deficiency by Increasing MdPAP10 Transcription in Apple (Malus × Domestica)

Phosphorus is an essential nutrient during plant growth and development. It is involved in the formation of important compounds in plants, such as phospholipids, ATP, and nucleic acids. However, phosphorus is distributed unevenly in most soils, and exists primarily in the form of organophosphorus, which plants cannot use. In this study, the AtPHR1 homolog MdPHR1 was identified in apple. Our results showed that MdPHR1 contained a MYB domain and a coiled-coil domain, which were conserved in AtPHR1. The MdPHR1 transgenic lines had higher acid phosphatase activity and phosphorus content than the wild-type under phosphorus limited conditions. Overexpression of MdPHR1 enhanced tolerance to phosphorus deficiency in apple calli and Arabidopsis seedlings, and MdPHR1 could bind to the P1BS cis-element of MdPAP10 and activated its transcription. Collectively, our experimental evidence suggests that MdPHR1 may be the central regulator of the system controlling transcriptional responses to Pi starvation in apple.

Effects of graded levels of starch on the non-specific immune responses, antioxidant capacities and intestinal health in Chinese mitten crab, Eriocheir inensis

A 9-week feeding trial was conducted to investigate the effects of graded levels of dietary starch (12%, 17%, 22%, 27% and 32%) on growth, non-specific immune responses, antioxidant capacities, immunity genes expression levels and pathogen resistance in Chinese mitten crab, Eriocheir inensis (initial body weight: 10.5 ± 0.5 g). Results showed that the highest weight gain rate of crabs was obtained in group containing 22% dietary starch. The highest activity of acid phosphatase, phenoloxidase and lysozyme in blood was found in crabs fed with 22–27% dietary starch. Additionally, 17%–27% dietary starch significantly increased the activities of superoxide dismutase and glutathione peroxidase, reduced malondinaldehyde content and then increased the total antioxidant capacities in hepatopancreas of crabs. The highest activity of alanine aminotransferase and aspartate aminotransferase was found in crabs fed with 32% dietary starch, indicating that excess starch had a negative effect on the liver function of crabs. With the dietary starch level increased, the transcription factors gene expression of the pro-inflammatory factors were significantly up-regulated, and the highest ILF2, IL-16, Relish and ADAM10 was found in crabs fed with dietary 32% starch, which may potentially promote the inflammatory response in intestines. Moreover, with the dietary starch increased, the activity of phenoloxidase and lysozyme, as well as the gene expression of crustin, were all increased in crabs after challenge against Citrobacter freundii, which demonstrated that additional dietary starch could provide immune-protection and help crabs improve their resistance against pathogens. In conclusion, these results suggest that adequate dietary starch can increase growth, enhance innate immune responses and promote disease resistance, reduce oxidative stress and inflammatory response in E. inensis. Taken together, 22–27% dietary starch (25.9–30.8% dietary carbohydrate) was suggested as a digestible energy source in crabs feed.

E-POSTERS – MITOCHONDRIAL / METABOLIC DISEASES –CASE REPORTS: EP.01Becker muscular dystrophy with acid α-glucosidase pseudodeficiency

Diagnosis of Pompe disease is sometimes challenging because it exhibits clinical similarities to muscular dystrophy. We describe a case of Becker muscular dystrophy (BMD) with a remarkable reduction in activity of the acid α-glucosidase (GAA) enzyme, caused by a combination of pathogenic mutation and polymorphism variants resulting in pseudodeficiency in GAA. The three-year-old boy demonstrated asymptomatic creatinine kinase elevation. Neither exon deletion nor duplication was detected on multiplex ligation-dependent probe amplification (MLPA) of dystrophin. GAA enzyme activity in both dried blood spots and lymphocytes was low, at 11.7% and 7.7% of normal, respectively. However, genetic analysis of GAA detected only heterozygosity for a nonsense mutation (c.118C>T, p.Arg40*). Muscle pathology showed no glycogen deposits and no high acid phosphatase activity. Hematoxylin-eosin staining detected scattered regenerating fibers; the fibers were faint and patchy on immunochemistry staining of dystrophin. The amount of dystrophin protein was reduced to 11.8% of normal, on Western blotting analysis. Direct sequencing analysis of dystrophin revealed hemizygosity for a nonsense mutation (c.72G>A, p.Trp24*). The boy was diagnosed with BMD, despite remarkable reduction in GAA activity; further, he demonstrated heterozygosity for [p.Gly576Ser; p.Glu689Lys] polymorphism variants that indicated pseudodeficiency on another allele in GAA. Pseudodeficiency alleles are detected in approximately 4% of the Asian population; these demonstrate low activity of acid α-glucosidase (GAA), similar to levels found in Pompe disease. Clinicians should be careful in their interpretations of pseudodeficiency alleles that complicate diagnosis in cases of elevated creatinine kinase.

Activity of rhizobacteria antagonistic to Fusarium oxysporum f. sp. passiflorae in soils cultivated with monocotyledonous plants1

ABSTRACT In spite of the importance of passion fruit cultivation in Brazil, this activity has been forced to be itinerant, due to the occurrence of diseases in producing regions, in particular fusariosis. The use of rhizobacteria is a potential control alternative, but other improvements are necessary to guarantee the activity of these bacteria in the field. This study aimed at evaluating the influence of three monocotyledonous groups on the activity of rhizobacteria antagonistic to Fusarium oxysporum f. sp. passiflorae (Fop). Rhizobacteria were isolated from passion fruit plants and tested for the production of diffusible and volatile compounds, as well as chitinases that inhibit Fop. From a total of 167 isolates, two produced volatile compounds, 21 diffusible compounds and 32 chitinases. Three compatible combinations of isolates that had at least two action mechanisms against Fop and could grow in a wide pH range were applied in pots with sorghum, millet or maize, in addition to pots left fallow (control), in a greenhouse. The microbial biomass carbon, soil basal respiration, metabolic quotient and activity of the acid phosphatase enzyme were evaluated. The interaction between the cultivated species and rhizobacterial combinations was significant only for the metabolic quotient, whose values tended to be higher under millet cultivation in combination 3 (R77/R95/R104/R120), the most metabolically diversified one. The highest activity of acid phosphatase was also obtained with millet. The results for enzymatic activity and metabolic quotient indicate that millet provides a greater activity of the antagonists in the soil.

Influence of a tropical grass (Brachiaria brizantha cv. Mulato) as cover crop on soil biochemical properties in a degraded agricultural soil

The inclusion of tropical grass forage as a cover crop (CC) could be a useful tool to improve microbiological activity and, consequently, soil quality. The aim of this study was to evaluate the effect of Brachiaria brizantha cv. Mulato and maize (Zea mays) as CC on soil microbial communities and their contributions to a degraded common bean (Phaseolus vulgaris L.). monoculture system. Soil sampling was carried out in 2016 after six years of cumulative effect across different treatments: B. brizantha-B. brizantha-common bean (B2), B. brizantha-common bean (B1), maize-common bean (M) and common bean monoculture (control). B2 and B1 showed higher fluorescein diacetate hydrolysis (108.1% and 78.6%, respectively) and higher acid phosphatase activity (304.5% and 181.6%, respectively) compared with the control treatment. The metabolic efficiency was higher in treatments containing B. brizantha as CC, with a significantly lower metabolic quotient (respiration rate per unit microbial biomass carbon) in B2 (1.65) compared with the control (5.46). The B2 treatment also showed higher values of soil organic carbon, which was correlated with soil microbial activities. In contrast, qPCR analysis of microbial structure did not show significant differences in response to the evaluated treatments. Thus, fungal and bacterial abundance probably had less influence on the differentiation of treatments compared to microbial activity and soil chemical properties. In context of this research, the use of B. brizantha as CC increased soil fertility and generated a greater microbial metabolic efficiency. Our research demonstrates that B. brizantha cv. Mulato as CC is a suitable agricultural tool to restore soil biochemical properties.

Osteoclastic differentiation and resorption is modulated by bioactive metal ions Co2+, Cu2+ and Cr3+ incorporated into calcium phosphate bone cements.

Biologically active metal ions in low doses have the potential to accelerate bone defect healing. For successful remodelling the interaction of bone graft materials with both bone-forming osteoblasts and bone resorbing osteoclasts is crucial. In the present study brushite forming calcium phosphate cements (CPC) were doped with Co2+, Cu2+ and Cr3+ and the influence of these materials on osteoclast differentiation and activity was examined. Human osteoclasts were differentiated from human peripheral blood mononuclear cells (PBMC) both on the surface and in indirect contact to the materials on dentin discs. Release of calcium, phosphate and bioactive metal ions was determined using ICP-MS both in the presence and absence of the cells. While Co2+ and Cu2+ showed a burst release, Cr3+ was released steadily at very low concentrations (below 1 μM) and both calcium and phosphate release of the cements was considerably changed in the Cr3+ modified samples. Direct cultivation of PBMC/osteoclasts on Co2+ cements showed lower attached cell number compared to the reference but high activity of osteoclast specific enzymes tartrate resistant acid phosphatase (TRAP), carbonic anhydrase II (CAII) and cathepsin K (CTSK) and significantly increased gene expression of vitronectin receptor. Indirect cultivation with diluted Co2+ cement extracts revealed highest resorbed area compared to all other modifications and the reference. Cu2+ cements had cytotoxic effect on PBMC/osteoclasts during direct cultivation, while indirect cultivation with diluted extracts from Cu2+ cements did not provoke cytotoxic effects but a strictly inhibited resorption. Cr3+ doped cements did not show cytotoxic effects at all. Gene expression and enzyme activity of CTSK was significantly increased in direct culture. Indirect cultivation with Cr3+ doped cements revealed significantly higher resorbed area compared to the reference. In conclusion Cr3+ doped calcium phosphate cements are an innovative cement modification because of their high cytocompatibility and support of active resorption by osteoclasts.

Effects of prey, pitcher age, and microbes on acid phosphatase activity in fluid from pitchers of Sarracenia purpurea (Sarraceniaceae).

Carnivory in pitcher plants generally involves digestion of prey, by the plant itself, by symbionts, or both. While symbionts appear to be important in the digestion of prey in Sarracenia purpurea, the importance of pitcher-derived enzymes is less well documented. Our goal was to reduce microbial numbers in pitcher fluid in order to measure the acid phosphatase activity attributable to the pitchers themselves. Preliminary experiments indicated that various antibiotics were minimally effective at reducing microbial populations and that antibiotic-resistant microbes were easily cultured from pitcher fluid. Consequently, we measured the abundance of culturable microbes in every sample taken for the measurement of acid phosphatase activity. Pitchers fed with one sterilized ant had higher levels of acid phosphatase activity than unfed pitchers. Older pitchers were more responsive to feeding than young pitchers. Pitchers with high levels of microbes (on Day 5) had higher acid phosphatase activity than pitchers with low levels of microbes. However, fed pitchers were not more likely to have higher microbe levels and microbe levels were not related to pitcher age. When fluid samples from inside the pitcher were compared to appropriate controls incubated outside the pitcher, acid phosphatase activity was higher inside the pitcher. Results from the feeding experiments are consistent with a primary role of microbes in the digestion of prey in pitchers of S. purpurea. However, the relationship between pitcher age and enzyme activity is not a function of microbes in the pitcher fluid and may depend on enzymes produced by the plant. Our methods would not detect microbes embedded on the inner surface of the pitcher; and if they survived the alcohol rinse and antibiotics, we cannot rule out microbes as the source of the relationship between pitcher age and acid phosphatase activity.

Effects of dietary supplementation ofHaematococcus pluvialispowder on gonadal development, coloration and antioxidant capacity of adult male Chinese mitten crab (Eriocheir sinensis)

The green algae Haematococcus pluvialis is an important source of natural astaxanthin as feed additive. This study was conducted to investigate the effects of dietary supplementation of H. pluvialis powder on gonadal development, coloration and antioxidant capacity of adult male Chinese mitten crab Eriocheir sinensis, and four experimental diets were formulated to contain 0, 0.2%, 0.4% and 0.6% of H. pluvialis powder. There were four treatments (defined as D1 similar to D4) in this study and each treatment had three replicates. Dietary H. pluvialis contents had no significant effects on survival, body weight gain rate and gonadal development of male E.sinensis. For colour parameters, the total carotenoids content in carapace and hepatopancreas as well as hepatopancreatic lightness (L*) and carapace redness (a*) increased significantly with increasing dietary H. pluvialis (p<0.05). For the antioxidant indices in the serum, D4 had the lowest activities of superoxide dismutase (SOD) and peroxidase (POD), but the highest glutathione peroxidase (GSH-Px) and lactic dehydrogenase (LDH), while the malondialdehyde (MDA) in serum and hepatopancreas decreased significantly with the rising content of dietary H. pluvialis (p<0.05); D1 had the highest levels of SOD, POD and GSH-Px in hepatopancreas. For the non-specific immune indices, the highest activities of acid phosphatase (ACP) and -glutamyl transpeptidase (-GT) were found on the serum of D3 and D4 (p<0.05). D1 had the highest levels of ACP and alkaline phosphatase (ALP) in hepatopancreas, while D2 and D3 had the lowest levels of ALP and ACP respectively. These results suggested the optimal dietary natural astaxanthin level was around 40mg/kg diets.