Polyphasic characterization of Brazilian Rhizobium tropici strains effective in fixing N2 with common bean (Phaseolus vulgaris L.)

Common bean (Phaseolus vulgaris) is native to the Americas, and Rhizobium etli is the dominant microsymbiont in both the Mesoamerican and the Andean centers of genetic diversification. Wild common beans are not found in Brazil, although the legume has been cropped in the country throughout time and all but one of the rhizobial species that nodulate it (Rhizobium gallicum) have been broadly detected in Brazilian soils. However, the majority of the effective rhizobial strains isolated so far from field-grown plants belong to R. tropici. This study describes the analysis of symbiotic and non-symbiotic genes of 15 effective R. tropici strains, isolated from four geographically distant regions in Brazil. With RFLP-PCR of the 16S and 23S rRNA genes and sequence analysis of 16S rRNA, two clusters were observed, one related to R. tropici type A and another to type B strains. Diversity in ribosomal genes was high, indicating that type A strains might represent a new species. High intraspecies diversity was also observed in the rep-PCR analysis with BOX, ERIC and REP primers. However, in the RFLP-PCR analysis of nifH and nodC genes, all R. tropici showed unique combinations of profiles, which might reflect an evolutionary strategy to maximize N₂ fixation.     

Campylobacter jejuni, Campylobacter coli, and cytolethal distending toxin (CDT) genes in common teals (Anas crecca)

To evaluate the presence of Campylobacter spp. and related cdt genes, cloacal swabs were collected from 70 common teals (Anas crecca) and analyzed by culture methods and polymerase chain reaction. In addition, C. jejuni were examined also for the presence of wlaN gene. This is believed to be the first report of Campylobacter spp. in common teal and our results confirm the very common occurrence of C. jejuni (n=40) and C. coli (n=13) in waterfowls. Furthermore, the cdt genes were frequently present in both C. jejuni and C. coli isolated. Moreover, seven C. jejuni isolates carried also the wlaN gene which is presumably involved in the expression of ganglioside mimics in Guillain-Barré syndrome.     

Modelling diapause termination and phenology of the Japanese beetle,Popillia japonica

We deve?loped a mechanistic, stage-structured model simulating the phenology of Popillia japonica. The model simulates the influence of soil temperature on the larval diapause termination and on the development rate function of post-overwintering larvae and pupae. Model parameters are estimated based on literature evidence for pupae development and on a parameterisation process that allows estimating parameters for larval diapause termination and for the development rate function (and the related uncertainty) of post-overwintering larvae. Data used for model parameterisation and validation refer to time-series adult trap catches collected during the P. japonica monitoring programme performed by the Phytosanitary Service of Lombardy Region within the infested area in Lombardy (Italy) from 2015 to 2019. A total of 12 randomly selected locations are used to estimate biologically realistic model parameters (parameterisation dataset). We applied a Jackknife nonparametric resampling procedure on the parameterisation dataset to quantify uncertainty associated with parameters’ estimates. Parameterised model is then validated on time-series adult trap catches data referring to a different set of 12 randomly selected locations (validation dataset) surveyed in Lombardy. The model successfully predicted the beginning of adult emergence and the overall curve of adult emergence in the validation dataset. The model presented can support the definition of the best timing for the implementation of monitoring and control activities for the local and the area-wide management of P. japonica.

     

Diversity and symbiotic effectiveness of rhizobia isolated from field-grown soybean nodules in Paraguay

Soybean was introduced in Paraguay in the 1920s and commercial crops have been grown since the 1970s. Root nodulation occurs at the majority of the producing sites, although inoculation has been practiced in only 15-20% of the cropping areas. The diversity and symbiotic effectiveness of soybean rhizobia was studied using 78 isolates obtained from root nodules of field-grown plants at 16 sites located in the two main producing states. The rhizobial isolates were characterized in relation to several parameters in vitro (colony morphology, tolerance to high temperature and salinity, intrinsic resistance to antibiotics, synthesis of indole acetic acid, profiles of proteins and lipopolysaccharides) and in vivo (nodulation, plant growth and total N accumulated in shoots). Fifty-eight isolates had slow growth rates and alkaline reaction in medium containing mannitol as the carbon source, whereas 20 had fast growth rates and an acid reaction. Most isolates did not tolerate acidity (pH 4.5) or high temperature (40°C). Very few isolates shared similar protein and lipopolysaccharide profiles; therefore a high level of diversity was detected, with most of the isolates representing unique strains. Some of the isolates with an outstanding symbiotic performance were identified, and will now be tested under field conditions in a search for efficient and competitive strains for use in commercial inoculants in Paraguay.     

Effects of different management practices on arbuscular mycorrhizal fungal diversity in maize fields by a molecular approach

As obligate mutualistic symbionts, arbuscular mycorrhizal fungi (AMF) colonize the roots of many agricultural crops, and it is often claimed that agricultural practices are detrimental to AMF. As a result, agroecosystems impoverished in AMF may not get the fully expected range of benefits from these fungi. Using molecular markers on DNA extracted directly from soil and roots, we studied the effects of different management practices (tillage and N fertilization) on the AMF communities colonizing an experimental maize field in Central Italy. Our molecular analysis based on three different nuclear rRNA regions (18S, 28S and ITS) allowed us to assess AMF biodiversity. Glomeraceae members were the main colonizer, and they co-occurred with Gigasporaceae and Paraglomus regardless of the management practices applied. Diversisporaceae and Entrophosporaceae members were instead detected in the N-fertilized soils and in the untreated soil, respectively. The results obtained indicated that the general AMF assemblages structure and composition in the maize field plots appear to be primarily influenced by N fertilization and, to a lesser extent, by tillage. This study also validates the usefulness of multiple molecular markers to consolidate and refine the assessment of the environmental AMF diversity.     

Antioxidant activity and phenolic composition of wild, edible, and medicinal fennel from different Mediterranean countries

Fennel (Foeniculum vulgare Mill.) is a typical aromatic plant of the Mediterranean area, long used as a medicinal and spice herb. Fennel is also well-known for its essential oil, which has been extensively studied for many years owing to its commercial importance. In this work, the antioxidant activity and the total phenolic and flavonoid contents, as well as the quantitative determination of individual flavonoids and phenolic acids of wild, edible, and medicinal fennel from different Mediterranean countries, have been determined. The antioxidant activity was measured as the free radical (DPPH), hydroxyl radical, and superoxide anion scavenging activities. Wild fennel was found to exhibit a radical scavenging activity, as well as a total phenolic and total flavonoid content, higher than those of both medicinal and edible fennels.     

Characterization of Neofusicoccum parvum causing canker and dieback on Brachychiton species

Brachychiton species are planted in Italy as ornamental trees. A survey in a nursery in Sicily (Italy) revealed the presence of young B. acerifolius and B. populneus showing severe trunk cankers, massive gummosis from the bark cracks, wood discolouration, and twig dieback. Morphological characterization was based on conidia measurement, and molecular characterization was carried out performing phylogenetic analysis (Maximum Parsimony) based on multi-locus approach of partial ITS, EF-1α, and tub2. Identification of the causal agent resulted in Neofusicoccum parvum (Botryosphaeriaceae). Pathogenicity tests were conducted in order to fulfill the Koch’s postulates. Five and three-years-old potted Brachychiton plants were inoculated (indoor and outdoor) using mycelial plugs. All the inoculated plants showed severe symptoms similar of those observed in the nursery during the survey. Some of the inoculated plants were completely dead. Re-isolations fully confirmed the causal agent. For our knowledge, this is the first report worldwide of N. parvum attacking Brachychiton spp.

     

Microbiological parameters as indicators of soil quality under various soil management and crop rotation systems in southern Brazil

The objective of this work was to identify soil parameters potentially useful to monitor soil quality under different soil management and crop rotation systems. Microbiological and chemical parameters were evaluated in a field experiment in the State of Paraná, southern Brazil, in response to soil management [no-tillage (NT) and conventional tillage (CT)] and crop rotation [including grain (soybean, S; maize, M; wheat, W) and legume (lupin, L.) and non-legume (oat, O) covers] systems. Three crop rotation systems were evaluated: (1) (O/M/O/S/W/S/L/M/O/S), (2) (O/S/L/M/O/S/W/S/L/M), and (3) (O/S/W/S/L/M/O/M/W/M), and soil parameters were monitored after the fifth year. Before ploughing, CO₂-emission rates were similar in NT and CT soils, but plough increased it by an average of 57%. Carbon dioxide emission was 13% higher with lupin residues than with wheat straw; decomposition rates were rapid with both soil management systems. Amounts of microbial biomass carbon and nitrogen (MB-C and MB-N, respectively) were 80 and 104% higher in NT than in CT, respectively; however, in general these parameters were not affected by crop rotation. Efficiency of the microbial community was significantly higher in NT: metabolic quotient (qCO₂) was 55% lower than in CT. Soluble C and N levels were 37 and 24% greater in NT than in CT, respectively, with no effects of crop rotation. Furthermore, ratios of soluble C and N contents to MB-C and MB-N were consistently lower in NT, indicating higher immobilization of C and N per unit of MB. The decrease in qCO₂ and the increase in MB-C under NT allowed enhancements in soil C stocks, such that in the 0–40 cm profile, a gain of 2500 kg of C ha⁻¹ was observed in relation to CT. Carbon stocks also varied with crop rotation, with net changes at 0–40 cm of 726, 1167 and −394 kg C ha⁻¹ year, in rotations 1, 2 and 3, respectively. Similar results were obtained for the N stocks, with 410 kg N ha⁻¹ gained in NT, while crop rotations 1, 2 and 3 accumulated 71, 137 and 37 kg of N ha⁻¹ year⁻¹, respectively. On average, microbial biomass corresponded to 2.4 and 1.7% of the total soil C, and 5.2 and 3.2% of the N in NT and CT systems, respectively. Soil management was the main factor affecting soil C and N levels, but enhancement also resulted from the ratios of legumes and non-legumes in the rotations. The results emphasize the importance of microorganisms as reservoirs of C and N in tropical soils. Furthermore, the parameters associated with microbiological activity were more responsive to soil management and crop rotation effects than were total stocks of C and N, demonstrating their usefulness as indicators of soil quality in the tropics.     

Benefits of inoculation of the common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis>) crop with efficient and competitive<Emphasis Type="Italic"> Rhizobium tropici</Emphasis> strains

Cropping in low fertility soils, especially those poor in N, contributes greatly to the low common bean (Phaseolus vulgaris L.) yield, and therefore the benefits of biological nitrogen fixation must be intensively explored to increase yields at a low cost. Six field experiments were performed in oxisols of Paraná State, southern Brazil, with a high population of indigenous common bean rhizobia, estimated at a minimum of 10³ cells g^–1 soil. Despite the high population, inoculation allowed an increase in rhizobial population and in nodule occupancy, and further increases were obtained with reinoculation in the following seasons. Thus, considering the treatments inoculated with the most effective strains (H 12, H 20, PRF 81 and CIAT 899), nodule occupancy increased from an average of 28% in the first experiment to 56% after four inoculation procedures. The establishment of the selected strains increased nodulation, N₂ fixation rates (evaluated by total N and N-ureide) and on average for the six experiments the strains H 12 and H 20 showed increases of 437 and 465 kg ha^–1, respectively,in relation to the indigenous rhizobial population. A synergistic effect between low levels of N fertilizer and inoculation with superior strains was also observed, resulting in yield increases in two other experiments. The soil rhizobial population decreased 1 year after the last cropping, but remained high in the plots that had been inoculated. DGGE analysis of soil extracts showed that the massive inoculation apparently did not affect the composition of the bacterial community.