Carmine Crecchio

Short-term effects of municipal solid waste compost amendments on soil carbon and nitrogen content, some enzyme activities and genetic diversity

Abstract. Municipal solid waste (MSW) composts have been frequently used as N and C amendments to improve soil quality and to support plant growth, with the additional benefit of reducing waste disposal costs. However, attention has been paid to the risks of MSW use for the soil environment. The presence of heavy metals in MSW composts can affect some microbiological characteristics of soil such as the structure of the soil microbiota, which are responsible for the transformations making nutrients available to plants. The effects of MSW compost and mineral-N amendments in a 2-year field trial on some physical-chemical properties, some enzyme activities and the genetic diversity of cropped plots (sugar beet-wheat rotation) and uncropped plots were investigated. Variations of pH were not statistically related to MSW compost and mineral-N amendments, or to the presence of the crop. Amendment with MSW compost increased the organic C and total N contents, and dehydrogenase and nitrate reductase activities of soil. In cropped plots amended with MSW compost, dehydrogenase activity was positively correlated with β-glucosidase activity, and both enzyme activities with organic C content. No MSW compost dosage effect was detected. No effects were observed on denaturing gradient gel electrophoresis and amplified rDNA restriction analysis patterns, indicating that no significant change in the bacterial community occurred as a consequence of MSW amendment.

Microbial interactions in the rhizosphere: beneficial influences of plant growth-promoting rhizobacteria on nutrient acquisition process. A review

Plant growth-promoting rhizobacteria (PGPR) are soil bacteria that are able to colonize rhizosphere and to enhance plant growth by means of a wide variety of mechanisms like organic matter mineralization, biological control against soil-borne pathogens, biological nitrogen fixation, and root growth promotion. A very interesting feature of PGPR is their ability of enhancing nutrient bioavailability. Several bacterial species have been characterized as P-solubilizing microorganisms while other species have been shown to increase the solubility of micronutrients, like those that produce siderophores for Fe chelation. The enhanced amount of soluble macro- and micronutrients in the close proximity of the soil-root interface has indeed a positive effect on plant nutrition. Furthermore, several pieces of evidence highlight that the inoculation of plants with PGPR can have considerable effects on plant at both physiological and molecular levels (e.g., induction of rhizosphere acidification, up- and downregulation of genes involved in ion uptake, and translocation), suggesting the possibility that soil biota could stimulate plants being more efficient in retrieving nutrients from soil and coping with abiotic stresses. However, the molecular mechanisms underlying these phenomena, the signals involved as well as the potential applications in a sustainable agriculture approach, and the biotechnological aspects for possible rhizosphere engineering are still matters of discussion.

Duodenal and faecal microbiota of celiac children: molecular, phenotype and metabolome characterization

BackgroundEpidemiology of celiac disease (CD) is increasing. CD mainly presents in early childhood with small intestinal villous atrophy and signs of malabsorption. Compared to healthy individuals, CD patients seemed to be characterized by higher numbers of Gram-negative bacteria and lower numbers Gram-positive bacteria.ResultsThis study aimed at investigating the microbiota and metabolome of 19 celiac disease children under gluten-free diet (treated celiac disease, T-CD) and 15 non-celiac children (HC). PCR-denaturing gradient gel electrophoresis (DGGE) analyses by universal and group-specific primers were carried out in duodenal biopsies and faecal samples. Based on the number of PCR-DGGE bands, the diversity of Eubacteria was the higher in duodenal biopsies of T-CD than HC children. Bifidobacteria were only found in faecal samples. With a few exceptions, PCR-DGGE profiles of faecal samples for Lactobacillus and Bifidobacteria differed between T-CD and HC. As shown by culture-dependent methods, the levels of Lactobacillus, Enterococcus and Bifidobacteria were confirmed to be significantly higher (P = 0.028; P = 0.019; and P = 0.023, respectively) in fecal samples of HC than in T-CD children. On the contrary, cell counts (CFU/ml) of presumptive Bacteroides, Staphylococcus, Salmonella, Shighella and Klebsiella were significantly higher (P = 0.014) in T-CD compared to HC children. Enterococcus faecium and Lactobacillus plantarum were the species most diffusely identified. This latter species was also found in all duodenal biopsies of T-CD and HC children. Other bacterial species were identified only in T-CD or HC faecal samples. As shown by Randomly Amplified Polymorphic DNA-PCR analysis, the percentage of strains identified as lactobacilli significantly (P = 0.011) differed between T-CD (ca. 26.5%) and HC (ca. 34.6%) groups. The metabolome of T-CD and HC children was studied using faecal and urine samples which were analyzed by gas-chromatography mass spectrometry-solid-phase microextraction and 1H-Nuclear Magnetic Resonance. As shown by Canonical Discriminant Analysis of Principal Coordinates, the levels of volatile organic compounds and free amino acids in faecal and/or urine samples were markedly affected by CD.ConclusionAs shown by the parallel microbiology and metabolome approach, the gluten-free diet lasting at least two years did not completely restore the microbiota and, consequently, the metabolome of CD children. Some molecules (e.g., ethyl-acetate and octyl-acetate, some short chain fatty acids and free amino acids, and glutamine) seems to be metabolic signatures of CD.

Heat Shock Response in Lactobacillus plantarum

ABSTRACT Heat stress resistance and response were studied in strains of Lactobacillus plantarum. Stationary-phase cells of L. plantarum DPC2739 had decimal reduction times (D values) (D value was the time that it took to reduce the number of cells by 1 log cycle) in sterile milk of 32.9, 14.7, and 7.14 s at 60, 72, and 75°C, respectively. When mid-exponential-phase cells were used, the D values decreased. The temperature increases which caused a 10-fold reduction in the D value ranged from 9 to 20°C, depending on the strain. Part of the cell population treated at 72°C for 90 s recovered viability during incubation at 7°C in sterile milk for 20 days. When mid-exponential- or stationary-phase cells of L. plantarum DPC2739 were adapted to 42°C for 1 h, the heat resistance at 72°C for 90 s increased ca. 3 and 2 log cycles, respectively. Heat-adapted cells also showed increased growth at pH 5 and in the presence of 6% NaCl. Two-dimensional gel electrophoresis of proteins expressed by control and heat-adapted cells revealed changes in the levels of expression of 31 and 18 proteins in mid-exponential- and stationary-phase cells, respectively. Twelve proteins were commonly induced. Nine proteins induced in the heat-adapted mid-exponential- and/or stationary-phase cells of L. plantarum DPC2739 were subjected to N-terminal sequencing. These proteins were identified as DnaK, GroEL, trigger factor, ribosomal proteins L1, L11, L31, and S6, DNA-binding protein II HlbA, and CspC. All of these proteins have been found to play a role in the mechanisms of stress adaptation in other bacteria. Antibodies against GroES detected a protein which was induced moderately, while antibodies against DnaJ and GrpE reacted with proteins whose level of expression did not vary after heat adaptation. This study showed that the heat resistance of L. plantarum is a complex process involving proteins with various roles in cell physiology, including chaperone activity, ribosome stability, stringent response mediation, temperature sensing, and control of ribosomal function. The physiological mechanisms of response to pasteurization in L. plantarum are fundamental for survival in cheese during manufacture.

Biodegradation and insecticidal activity of the toxin from Bacillus thuringiensis subsp. kurstaki bound on complexes of montmorillonite- humic acids-Al hydroxypolymers

The equilibrium adsorption and binding of the active toxin from Bacillus thuringiensis subsp. kurstaki on complexes of montmorillonite‐ humic acids‐Al hydroxypolymers, as well as the biodegradation and the insecticidal activity of the bound toxin, were studied. Seventy percent of the total adsorption occurred within the first hour, and maximal adsorption occurred in ,8 h. Adsorption of the toxin on a constant amount of the complexes increased as the amount of the toxin added increased, and equilibrium adsorption isotherms of the L-type were obtained. There was essentially no desorption of the toxin after extensive washing of the toxin‐organomineral complexes with double distilled H2O and 1 M NaCl. The bound toxin was resistant to utilization by mixed microbial cultures from soil and to enzymatic degradation by Pronase E. Free and bound toxin were active against the larvae of Manduca sexta; the bound toxin retained the same activity after exposure to microbes or Pronase, whereas the toxicity of the free toxin decreased significantly. The results of these studies indicate that the release of transgenic plants and microorganisms expressing truncated genes that encode active insecticidal toxins from B. thuringiensis could result in the accumulation of these toxins in soil as a consequence of binding on surface-active soil particles. This persistence could pose a hazard to nontarget organisms, enhance the selection of toxin-resistant target species, and increase the control of target insect pests. q 2001 Elsevier Science Ltd. All rights reserved.

Different Fecal Microbiotas and Volatile Organic Compounds in Treated and Untreated Children with Celiac Disease

ABSTRACT This study aimed at investigating the fecal microbiotas of children with celiac disease (CD) before (U-CD) and after (T-CD) they were fed a gluten-free diet and of healthy children (HC). Brothers or sisters of T-CD were enrolled as HC. Each group consisted of seven children. PCR-denaturing gradient gel electrophoresis (DGGE) analysis with V3 universal primers revealed a unique profile for each fecal sample. PCR-DGGE analysis with group- or genus-specific 16S rRNA gene primers showed that the Lactobacillus community of U-CD changed significantly, while the diversity of the Lactobacillus community of T-CD was quite comparable to that of HC. Compared to HC, the ratio of cultivable lactic acid bacteria and Bifidobacterium to Bacteroides and enterobacteria was lower in T-CD and even lower in U-CD. The percentages of strains identified as lactobacilli differed as follows: HC (ca. 38%) > T-CD (ca. 17%) > U-CD (ca. 10%). Lactobacillus brevis, Lactobacillus rossiae, and Lactobacillus pentosus were identified only in fecal samples from T-CD and HC. Lactobacillus fermentum, Lactobacillus delbrueckii subsp. bulgaricus, and Lactobacillus gasseri were identified only in several fecal samples from HC. Compared to HC, the composition of Bifidobacterium species of T-CD varied, and it varied even more for U-CD. Forty-seven volatile organic compounds (VOCs) belonging to different chemical classes were identified using gas-chromatography mass spectrometry-solid-phase microextraction analysis. The median concentrations varied markedly for HC, T-CD, and U-CD. Overall, the r2 values for VOC data for brothers and sisters were equal to or lower than those for unrelated HC and T-CD. This study shows the effect of CD pathology on the fecal microbiotas of children.

Effects of conventional tillage on biochemical properties of soils

Abstract Modification of soil environment by different farming practices can significantly affect crop growth. Tillage causes soil disturbance, altering the vertical distribution of soil organic matter and plant nutrient supplies in the soil surface, and it may affect the enzyme activity and microbial biomass which are responsible for transformation and cycling of organic matter and plant nutrients. In this study, the influence of three conventional tillage systems (shallow plowing, deep plowing and scarification) at different depths on the distribution and activity of enzymes, microbial biomass and nucleic acids in a cropped soil was investigated. Analysis of variance for depth and tillage showed the influence of the different tillage practices on the activity of some enzymes and on the nucleic acids. Glucosidase, galactosidase, nitrate reductase and dehydrogenase activity were significantly affected by the three tillage modalities. Activity in the upper layer (0–20 cm) was higher in the plots tilled by shallow plowing and scarification than in those tilled by deep plowing. Positive relationships were observed between the soil enzymes themselves, with the exception of urease and pyrophosphatase activity. Moreover, significant correlations were found between DNA and β-galactosidase, and between RNA and β-glucosidase, β-galactosidase, alkaline phosphatase and phosphodiesterase. α-Glucosidase, β-galactosidase, alkaline phosphatase and phosphodiesterase were highly correlated with biomass C determined by the fumigation-extraction method.

Fractionation of sugar beet pulp into pectin, cellulose, and arabinose by arabinases combined with ultrafiltration

Incubation of beet pulp with two arabinases (alpha-L-arabinofuranosidase and endo-arabinase), used singularly or in combination at different units of activity per gram of beet pulp, caused the hydrolysis of arabinan, which produced a hydrolyzate consisting mainly of arabinose. Pectin and a residue enriched with cellulose were subsequently separated from the incubation mixture. The best enzymatic hydrolysis results were obtained when 100 U/g of beet pulp of each enzyme worked synergistically with yields of 100% arabinose and 91.7% pectin. These yields were higher than those obtained with traditional chemical hydrolysis. The pectin fraction showed a low content of neutral sugar content and the cellulose residue contained only a small amount of pentoses. Semicontinuous hydrolysis with enzyme recycling in an ultrafiltration unit was also carried out to separate arabinose, pectin, and cellulose from beet pulp in 7 cycles of hydrolysis followed by ultrafiltration. The yields of separation were similar to those obtained in batch experiments, with an enzyme consumption reduced by 3.5 times and some significant advantages over batch processes.

Identification of the mutation responsible for a case of plasmatic apolipoprotein CII deficiency (Apo CII-bari)

We studied a case of familial Apolipoprotein CII deficiency. By Southern hybridization, amplification and sequence analysis, the genetic defect was identified. It consists in a point mutation C- greater than G in the third exon of the gene causing a premature stop codon. Truncated at the aa. 36 of the mature form, the protein loses its functional domains, becomes inefficient and cannot be detected in the plasma, because of its high instability. The mutation destroys an RsaI site, present in the normal gene sequence. This point mutation is useful in the diagnosis of this Apolipoprotein CII deficiency.

Plant-microorganism-soil interactions influence the Fe availability in the rhizosphere of cucumber plants

Iron (Fe) is a very important element for plants, since it is involved in many biochemical processes and, often, for the low solubility of the natural Fe sources in soil, plants suffer from Fe - deficiency, especially when grown on calcareous soils. Among the numerous plant growth-promoting rhizobacteria (PGPR) that colonize the rhizosphere of agronomically important crops, Azospirillum brasilense has been shown to exert strong stimulating activities on plants, by inducing alterations of the root architecture and an improvement of mineral nutrition, which could result from an enhancement of ion uptake mechanisms as well as by increased bioavailability of nutrients. Some studies have also established that A. brasilense can act as biocontrol agent, by preventing the growth and/or virulence of phytopathogens, most likely through the production of microbial siderophores that sequester Fe from the soil. Despite microbial siderophores complexed with Fe could be an easily accessible Fe source for plants, the possible involvement of A. brasilense in improving Fe nutrition in plants suffering from the micronutrient deficiency has not been investigated yet. Within the present research, the characterization of the physiological and biochemical effects induced by Fe starvation and PGPR inoculation in cucumber plants (Cucumis sativus L. cv. Chinese Long) was carried out. The analyses of root exudates released by hydroponically grown plants highlighted that cucumber plants respond differently depending on the nutritional status. In addition, following the cultivation period on calcareous soil, also the root exudates found in the extracts suggested a peculiar behaviour of plants as a function of the treatment. Interestingly, the presence of the inoculum in soil allowed a faster recovery of cucumber plants from Fe-deficiency symptoms, i.e. increase in the chlorophyll content, in the biomass and in the Fe content of leaves. These observations might suggest a feasible application of A. brasilense in alleviating symptoms generated by Fe-limiting growth condition in cucumber plants.