Development of functional consortia for the pretreatment of compostable lignocellulosic waste: A simple and effective solution to a large-scale problem.

Microorganisms drive the degradation of organic matter thanks to their enzymatic versatility. However, the structure of lignocellulose poses a great challenge for the microbiota inhabiting a compost pile. Our purpose was to increase the biodegradability of vegetable waste in the early stages of the composting process by applying a microbial consortium with lignocelllulolytic capacity. For this, a previous screening was performed among the culturable microbiota from different composting processes to find inoculants with ligninocellulolytic activity. Selected strains were applied as a pure culture and as a microbial consortium. The starting material was composed of tomato plant and pruning remains mixed in a ratio (50:50 v/v), whose humidity was adjusted to around 65%. To determine the ability of both treatments to activate the biodegradation of the mixtures, moisture, organic matter, ash, C/N ratio, 4-day cumulative respirometric index (AT4) and degradation rates of cellulose, hemicellulose and lignin were evaluated. Subsequently, a real composting process was developed in which the performance of the microbial consortium was compared with the composting process without inoculum (control). According to our tests, three microbial strains (Bacillus safensis, Bacillus licheniformis and Fusarium oxysporum) were selected. The results showed that the application of the bacteria strains at low doses (104 CFU g-1 on the complete residual material of the pile) resulted in higher rates of lignocelullose degradation after 10 days of treatment compared to that observed after application of the fungus in pure culture or untreated controls. The implementation of the strategy described in this work resulted in obtaining compost with better agronomic quality than the uninoculated controls. Therefore, the application of this consortium could be considered as an interesting tool for bioactivation of lignocellulosic waste prior to the composting process.

Sustainable approach to the control of airborne phytopathogenic fungi by application of compost extracts.

The search for new sustainable alternatives for plant disease control has gained interest in the last decades. Compost extracts are nowadays considered a remarkable alternative to agrochemicals due to their biopesticidal properties. However, these properties could be affected by the different variables of extraction protocols and by starting compost. This work focused on the physicochemical and biological characterization of compost extracts obtained from a wide range of composted materials and different extraction protocols (CEP). CEP-1 and CEP-4 involved incubation at 20 °C for 48 h and 14 days, respectively; CEP-2 incubation for 24 h at 40 °C; while CEP-3 were incubated for 12 h at 70 °C. Electrical conductivity, pH, total organic carbon (TOC) and phenolic content were determined as well as the actinobacterial count and enzyme profiles related to plant pathogen suppression. Additionally, the influence of the different materials and protocols on the in vitro growth inhibition of Alternaria alternata and Botrytis cinerea was determined. The starting materials and extraction protocols significantly influenced the physicochemical and biological characteristics of extracts. Treatments based on long incubation times at 20 °C, as well as those based on short incubation times at 40 °C, resulted in extracts with increased suppressive properties. However, extracts derived from CEP-3 protocol were characterized by high phenolic and TOC content, low functional biodiversity, and a more discreet antagonistic capacity. Therefore, the development and optimization of suitable extraction protocols could lead to compost extracts with increased phytoprotective capacities, thus becoming an effective and sustainable alternative to chemical inputs.

Mitigation of phytotoxic effect of compost by application of optimized aqueous extraction protocols.

The abuse of chemical fertilizers in recent decades has led the promotion of less harmful alternatives, such as compost or aqueous extracts obtained from it. Therefore, it is essential to develop liquid biofertilizers, which in addition of being stable and useful for fertigation and foliar application in intensive agriculture had a remarkable phytostimulant extracts. For this purpose, a collection of aqueous extracts was obtained by applying four different Compost Extraction Protocols (CEP1, CEP2, CEP3, CEP4) in terms of incubation time, temperature and agitation of compost samples from agri-food waste, olive mill waste, sewage sludge and vegetable waste. Subsequently, a physicochemical characterization of the obtained set was performed in which pH, electrical conductivity and Total Organic Carbon (TOC) were measured. In addition, a biological characterization was also carried out by calculating the Germination Index (GI) and determining the Biological Oxygen Demand (BOD5). Furthermore, functional diversity was studied using the Biolog EcoPlates technique. The results obtained confirmed the great heterogeneity of the selected raw materials. However, it was observed that the less aggressive treatments in terms of temperature and incubation time, such as CEP1 (48 h, room temperature (RT)) or CEP4 (14 days, RT), provided aqueous compost extracts with better phytostimulant characteristics than the starting composts. It was even possible to find a compost extraction protocol that maximize the beneficial effects of compost. This was the case of CEP1, which improved the GI and reduced the phytotoxicity in most of the raw materials analyzed. Therefore, the use of this type of liquid organic amendment could mitigate the phytotoxic effect of several composts being a good alternative to the use of chemical fertilizers.

Biorecovery of olive mill wastewater sludge from evaporation ponds.

Olive mill wastewater (OMW) resulting from the olive oil extraction process is usually disposed of in evaporation ponds where it concentrates generating a sludge that pollutes the ponds nearby area. In this study, four bio-treatments were applied for the in-situ bioremediation and valorization of OMW sludge: Landfarming, phytoremediation, composting and vermicomposting. In all cases, the OMW sludge was added with organic residues (mushroom compost, rabbit manure, and chicken manure). The bio-treatments were carried out in duplicate, inoculated and non-inoculated, to determine the effect of a specialized fungal consortium (Aspergillus ochraceus H2 and Scedosporium apiospermum H16) on the efficacy of the bio-treatments. The evaluation of chemical parameters, toxicity, and functional microbial biodiversity revealed that the four techniques depleted the toxicity and favored the stimulation of functional microbiota. Landfarming and phytoremediation allowed the decontamination and improvement of soils. Composting and vermicomposting also offered high-quality products of agronomic interest. Inoculation improved the bioremediation effectiveness. Biological treatments are effective for the safe recovery of contaminated OMW sludge into high-quality services and products.

Design and validation of cyanobacteria-rhizobacteria consortia for tomato seedlings growth promotion.

The use of rhizobacteria provide great benefits in terms of nitrogen supply, suppression of plant diseases, or production of vitamins and phytohormones that stimulate the plant growth. At the same time, cyanobacteria can photosynthesize, fix nitrogen, synthesize substances that stimulate rhizogenesis, plant aerial growth, or even suppose an extra supply of carbon usable by heterotrophic bacteria, as well as act as biological control agents, give them an enormous value as plant growth promoters. The present study focused on the in vitro establishment of consortia using heterotrophic bacteria and cyanobacteria and the determination of their effectiveness in the development of tomato seedlings. Microbial collection was composed of 3 cyanobacteria (SAB-M612 and SAB-B866 belonging to Nostocaceae Family) and GS (unidentified cyanobacterium) and two phosphate and potassium solubilizing heterotrophic bacteria (Pseudomonas putida-BIO175 and Pantoea cypripedii-BIO175). The results revealed the influence of the culture medium, incubation time and the microbial components of each consortium in determining their success as biofertilizers. In this work, the most compatible consortia were obtained by combining the SAB-B866 and GS cyanobacteria with either of the two heterotrophic bacteria. Cyanobacteria GS promoted the growth of both rhizobacteria in vitro (increasing logarithmic units when they grew together). While Cyanobacteria SAB-B866 together with both rhizobacteria stimulated the growth of tomato seedlings in planta, leading to greater aerial development of the treated seedlings. Parameters such as fresh weight and stem diameter stood out in the plants treated with the consortia (SAB-B866 and both bacteria) compared to the untreated plants, where the values doubled. However, the increase was more discrete for the parameters stem length and number of leaves. These results suggest that the artificial formulation of microbial consortia can have positive synergistic effects on plant growth, which is of enormous agro-biotechnological interest.

Olive mill wastewater-evaporation ponds long term stored: Integrated assessment of in situ bioremediation strategies based on composting and vermicomposting.

During the last two decades, the method most widely used to manage olive mill wastewater (OMW) derived from olive oil production has been its disposal in evaporation ponds. Long-term storage of OMW leads to the accumulation of toxic sediments (OMWS) rich in recalcitrant compounds with phytotoxic and antimicrobial properties, which limit their use for agronomic purpose. The aim of this study was to compare the effect of two in situ bioremediation strategies (composting and a combination of composting followed by vermicomposting) to remove the potential toxicity of the sediments derived from long-term stored OMW. The results obtained showed that the composting method assisted with the earthworms enhanced the depletion of phenolic compounds and OMWS ecotoxicity more than composting, especially during the maturation stage. Moreover, vermicomposting was more effective in the reduction of the OMWS salinity. However, a pre-composting process to the OMWS is necessary prior to vermicomposting to provide the suitable conditions for earthworms survival and activity. Furthermore, the final compost showed a phytostimulating effect. Therefore, these in situ bioremediation strategies can be considered potential tools for decontamination and recovery of long-term stored OMWS in evaporation ponds, which currently poses an unsolved environmental problem.

Diet and cooling interactions on physiological responses of grazing dairy cows, milk production and composition.

The objective of this trial was to evaluate the effects of diet and cooling in the holding pen before milking on rectal temperature, respiration rate and milk production and composition. Fifty-eight lactating Holstein cows were used in a factorial split-plot design, at Rafaela Experimental Station from 12 January to 3 March 2003. The treatments were combinations of two diets: control (CD) and balanced (BD) with two levels of cooling before milking: none (NSF) and a sprinkler and fans (SF). Forage:concentrate ratios for CD and BD were 81:19 and 68:32, respectively. Cows were milked twice daily. Milk production was recorded daily, and milk composition (fat, protein, lactose and urea) was analysed twice a week. The physiological data were recorded once a week, before the cattle entered the holding pen and after milking, in the afternoon. Average maximum weekly temperature humidity index was 75.4 and ranged from 61.4 to 83. There were highly significant effects of cooling on physiological responses. Milk production was affected by diet and cooling, with no interaction; the highest and lowest production of milk was 22.42 and 20.07 l/cow per day, for BD+SF and CD+NSF, respectively. Protein was affected by diet, and was higher for BD (3.17 vs. 3.08%). There were interaction effects on milk fat at the 8% level, the highest concentration being 3.65% for BD+NFS. It was concluded that under grazing conditions, cooling by sprinkler and fans before milking improves the comfort of dairy cows, and that the effects on milk production and composition are enhanced when diets are specially formulated for heat-stress periods.

Monensin for lactating dairy cows grazing mixed-alfalfa pasture and supplemented with partial mixed ration.

The effect of monensin on milk production was evaluated in 58 lactating Holstein cows (48 multiparous; 10 primiparous) grazing a mixed-alfalfa pasture and supplemented with a partial mixed ration in a completely randomized design with repeated measurements. Cows were paired by calving date, lactation number, previous lactation milk production, body weight, and body condition score and were assigned to one of 2 treatments: control or monensin. Cows on the monensin treatment received 2 monensin controlled-release capsules (335 mg/d for 90 d), one 30 d before the expecting calving date and the other 60 d after calving. Short-term (0 to 150 d in milk) and long-term (305-d adjusted lactation) effects of monensin were evaluated. Pasture (measured by difference between pre- and postgrazing pasture mass), supplements, and total dry matter intake did not differ between treatments and averaged 8.7, 14.1, and 22.9 kg/d, respectively. In the short-term, monensin increased milk production (27.7 vs. 26.6 kg/d) and milk protein yield (0.890 vs. 0.860 kg/d); milk fat yield was not affected (0.959 kg/d). Monensin decreased milk fat content (3.51 vs. 3.60%) with no changes in milk protein content (3.25%). In the long term, milk production and milk protein yield were also increased by monensin: 214 and 7 kg, respectively. Monensin reduced the loss of body condition score and increased percentage of pregnancy at first service (44.8 vs. 20.7%). Monensin improves production and reproduction performance of dairy cows grazing a mixed-alfalfa pasture and supplemented with a partial mixed ration.

Effects of feeding rations with genetically modified whole cottonseed to lactating Holstein cows.

Two experiments were conducted to evaluate dry matter intake (DMI), milk yield, and milk composition from feeding rations that contained different sources of genetically modified whole cottonseed to Argentinean Holstein dairy cows. Twenty-four lactating multiparous Argentinean Holstein dairy cows were used in 2 experiments with a replicated 4 x 4 Latin square design, with cows averaging 565 kg body weight and 53 d in milk at the beginning of the experiments. Treatments in Experiment 1 were: Bollgard cotton containing the cry1Ac gene, Bollgard II cotton containing cry1Ac and cry2Ab genes, Roundup Ready cotton containing the cp4 epsps gene, and a control nongenetically modified but genetically similar cottonseed. In Experiment 2, two commercial sources, a parental control line, and the transgenic cotton containing both cry1Ac and cp4 epsps genes were used as treatments. All cows received the same total mixed ration but with different whole cottonseed sources. Cottonseed was included to provide 2.50 kg per cow daily (dry matter [DM] basis) or about 10% of the total diet DM. The ingredient composition of the total mixed ration was 32% alfalfa hay, 28% corn silage, 22% corn grain, 17% soybean meal, and 2% minerals and vitamins. In addition, genomic DNA was extracted from a subset of milk samples and analyzed by polymerase chain reaction followed by Southern blot hybridization for small fragments of the cry1Ac transgene and an endogenous cotton gene, acp1. No sample was positive for transgenic or plant DNA fragments at the limits of detection for the assays following detailed data evaluation criteria. The DMI, milk yield, milk composition, body weight, and body condition score did not differ among treatments. Cottonseed from genetically modified varieties used in these studies yielded similar performance in lactating dairy cows when compared to non-transgenic control and reference cottonseed.

Hydrogenated fish fat for grazing dairy cows in summer.

Thirty-two multiparous Holando argentino cows in mid lactation were randomly assigned to two treatments: control or HFF (hydrogenated fish fat) at Rafaela, 31 degrees 11' South, during summer 1997/1998, to evaluate the effect of using HFF as a supplement under grazing conditions. Animals in both treatments grazed an alfalfa pasture, and were confined from 1000 hours to 1700 hours daily in a shaded pen where water was provided ad libitum. During each milking, animals in the control group received 3.73 kg dry matter per cow each day (DM cow(-1) day(-1)) concentrate (15% crude protein; 8.69 MJ energy for location/kg DM). Cows in the HFF group received 3.25 kg DM cow(-1) day(-1) concentrate, plus 0.200 kg DM cow(-1) day(-1) HFF. Both diets presented similar energy, protein and neutral detergent fibre contents. The trial was performed during a strong "El Niño" event, which resulted in a total rainfall of 396.3 mm (80% higher than normal). The mean temperature was 23.7 (SD 3.2) degrees C and the mean temperature humidity index was 72.9 (SD 4.96). Production data were analysed using a completely randomised design with analysis of covariance. Supplementation with HFF produced an increase in daily milk production (26.4 (SD 2.46) l/cow compared to 23.9 (SD 2.68) l/cow for the controls; P<0.05). Milk fat production was higher for HFF (P<0.05): 941 (SD 96) g cow(-1) day(-1) as compared to controls, which yielded 846 (SD 95) g cow(-1) day(-1). Milk protein yields also differed significantly (P<0.05), the respective values for HFF and controls being 795 (SD 72) g cow(-1) day(-1) and 715 (SD 83) g cow(-1) day(-1). It was concluded that hydrogenated fish fat could be a good ingredient to sustain high yields and elevated maintenance requirements in a grazing system during hot conditions.

Evaluation of different shades to improve dairy cattle well-being in Argentina.

Two tree shades (TS1 and TS2) and an artificial shade structure (AS) were evaluated using black globe temperatures (BGTs) to assess their effectiveness in reducing heat load. The artificial structure consisted of a black woven polypropylene cloth providing 80% shade, mounted on 2.5-m-high eucalyptus posts. The work was carried out at Rafaela Experimental Station, Argentina, during the summer (January and February) 1994. BGTs and floor temperatures were measured in concrete floor holding pens with and without artificial shade. The results showed no difference between TS1, TS2 and AS, their average BGTs being 30.2 (SD 0.58), 29.0 (SD 0.70) and 30.2 (SD 0.74) degrees C, respectively. BGTs under all three shades were significantly lower (P < 0.01) than the average BGT recorded outside: 35.5 (SD 1.12) degrees C. Average BGTs in holding pens were 32.4 (SD 1.38) and 39.9 (SD 1.91) degrees C for shaded and non-shaded areas (P < 0.01). The corresponding average floor temperatures were 27.8 (SD 0.68) degrees C and 47.7 (SD 2.13) degrees C (P < 0.01). To assess the effects of shade on animal well-being, afternoon rectal temperatures (RT) and respiratory rate (respirations per minute, RR) of lactating cows were recorded twice a week. Rectal temperatures were significantly higher for non-shaded cows (P < 0.01), mean RT being 40.1 (SD 0.65) degrees C vs 39.3 (SD 0.42) degrees C for the shaded animals. Corresponding RRs were 78.9 (SD 18.0) and 60.7 (SD 10.6) (P < 0.05). It was concluded that: (1) tree and artificial shades produced similar effects, (2) shading the holding pen with an 80% shading cloth was effective in reducing heat load and floor temperatures, and (3) access to shade in our pasture-based system improved animal well-being.

Hydroxyindole-O-methyl-transferase activity in the pineal gland of the rabbit.

Rabbit pineal HIOMT activity shows a diurnal rhythm. The highest values are observed at night and the lowest during the daytime. Animals kept in permanent darkness show higher values than those in constant light. Bilateral superior cervical ganglionectomy results in a tenfold decrease in pineal HIOMT activity.