Search Results (10)

The identification and quantification of chitinolytic activity in microorganisms is critical for advancing biological control strategies against arthropod pests and fungal pathogens. However, current laboratory methods designed for fast detection of chitinolytic microorganisms are often time-consuming, produce low-quality results and lack sensitivity. Here, we report the development of a novel fluorogenic culture medium incorporating a chemically modified chitinase substrate, N-fluoresceyl poly-D-glucosamine, which allows for a highly sensitive chitinase assay, enabling both qualitative and quantitative fluorescent detection of chitinase activity in situ. This substrate is synthesized through covalent conjugation of poly-D-glucosamine with fluorescein isothiocyanate under alkaline conditions, resulting in an insoluble polymer that becomes fluorescent upon enzymatic hydrolysis by chitinases. When supplemented with culture media, the modified fluorogenic substrate serves as the sole carbon source, selectively supporting the growth of chitinolytic microorganisms. Enzymatic activity is visualized under longwave UV light and can be quantitatively measured via spectrophotometric (493 nm) or fluorometric (530 nm) methods. Validation using characterized entomopathogenic chitinolytic strains of the fungi Aspergillus flavus, Beauveria bassiana, and Metarhizium anisopliae demonstrated a detection sensitivity that was at least three orders of magnitude greater than that of conventional methods. In contrast, the non-chitinolytic fungi Penicillium notatum and Fusarium venenatum presented no detectable fluorescent signals. This fluorogenic medium provides a rapid, cost-effective, and highly sensitive tool for screening chitinolytic microorganisms with potential applications in agriculture, veterinary parasitology, and environmental microbiology. Full article

This study examines the effects of reduced nitrogen (N) application on rice straw N depolymerization in coastal saline paddy soil to establish a scientific basis for optimizing N application strategies during straw incorporation in coastal paddy systems. A 360-day field straw bag burial experiment was conducted using four N application levels: N0 (control, without N fertilizer), N1 (225 kg N/ha), N2 (300 kg N/ha), and N3 (375 kg N/ha). The results indicated that applying 300 kg N/ha significantly (p < 0.05) increased dissolved organic N (DON) content, apr and chiA gene copies, and the activities of alkaline protease, chitinase, leucine aminopeptidase, and N-acetylglucosaminidase. In addition, the application of 300 kg N/ha enhanced the synergistic effects of alkaline protein- and chitin-degrading microbial communities. Pseudomonas, Brevundimonas, Sorangium, Cohnella, and Thermosporothrix were identified as keystone taxa predominant in straw N depolymerization. Straw N depolymerization occurred by two primary pathways: direct regulation of enzyme activity by straw properties of total carbon and electrical conductivity, and indirect influence on N hydrolase activity and DON production through modified microbial community structures. The findings suggest that an application rate of 300 kg N/ha is optimal for promoting straw N depolymerization in coastal saline paddy fields. Full article

Plant Growth Promoting Rhizobacteria, PGPR, can protect plants against soil-borne diseases and abiotic stress conditions. The primary objective of this study was to evaluate the effects of different PGPRs (TF1, TF2, TF3, and TF4) on the rhizosphere microbial community of silage maize in a saline–alkaline field via Illumina MiSeq high-throughput sequencing technology. Results demonstrated that different PGPRs significantly increased the harvest density (by 21.31–45.16%), plant height (by 9.12–19.98%), stem diameter (by 30.07–45.78%), and biomass (by 33.20–65.36%) of silage maize, TF3 treatment significantly increased the fresh weight (by 32.50%), while the other treatments could increase the fresh weight but not significantly. Four microbial agents significantly reduced the contents of soil available phosphorus (AP), electrical conductivity (EC), and neutral phosphatase activity (NPA), while significantly increasing the contents of available potassium (AK), ammonium nitrogen (NH₄⁺-N), nitrate nitrogen (NO₃⁻-N), chitinase activity (ChtA), and urease activity (UA). Specifically, TF2 and TF3 treatments significantly decreased the soil pH value, while not for TF1 and TF4. Microbiome analysis showed that four microbial agents significantly increased the relative abundances of beneficial microorganisms, such as Arthrobacter, Blastococcus, MNDI, Chaetomidium, Alternaria, Sarocladium, Acremonium, and Clonostachys, and significantly decreased the relative abundances of Gibberella and Fusarium. Mental analysis showed that the soil bacterial community structure did not significantly correlate with soil biochemical properties, while the soil fungal community structure significantly and positively correlated with pH. Maize yield significantly and positively correlated with NH₄⁺-N, OM, AP, EC, UA, ChtA, and NPA. Full article

Bacterial endophytes (120) were isolated from six halophytes (Distichlis spicata, Cynodon dactylon, Eragrostis obtusiflora, Suaeda torreyana, Kochia scoparia, and Baccharis salicifolia). These halophiles were molecularly identified and characterized with or without NaCl conditions. Characterization was based on tests such as indole acetic acid (IAA), exopolysaccharides (EPS), and siderophores (SID) production; solubilization of phosphate (P), potassium (K), zinc (Zn), and manganese (Mn); mineralization of phytate; enzymatic activity (acid and alkaline phosphatase, phytases, xylanases, and chitinases) and the mineralization/solubilization mechanisms involved (organic acids and sugars). Moreover, compatibility among bacteria was assessed. Eleven halophiles were characterized as highly tolerant to NaCl (2.5 M). The bacteria isolated were all different from each other. Two belonged to Bacillus velezensis and one to B. pumilus while the rest of bacteria were identified up to the genus level as belonging to Bacillus, Halobacillus, Halomonas, Pseudomonas, Nesterenkonia, and three strains of Oceanobacillus. The biochemical responses of nutrient solubilization and enzymatic activity were different between bacteria and were influenced by the presence of NaCl. Organic acids were involved in P mineralization and nutrient solubilization. Tartaric acid was common in the solubilization of P, Zn, and K. Maleic and vanillic acid were only detected in Zn and K solubilization, respectively. Furthermore, sugars appeared to be involved in the solubilization of nutrients; fructose was detected in the solubilization tests. Therefore, these biochemical bacterial characteristics should be corroborated in vivo and tested as a consortium to mitigate saline stress in glycophytes under a global climate change scheme that threatens to exacerbate soil salinity. Full article

The purpose of this research is to investigate the interactions among wheat varieties and microorganisms within rhizosphere and how conventional agriculture affects these dynamics during the growing season. Indeed, little is known about how commercial bread varieties modulate root exudates and how agrochemicals affect the microbiological processes. Therefore, this study investigated the changes in soil microbiological features such as enzyme activities (β-glucosidase, xylosidase, glucoronidase, chitinase, leucine-aminopeptidase, acid and alkaline phosphomonoesterases, inositol phosphatase, phosphodiesterase, pyrophosphatase–phosphodiesterase, arylsulphatase) and microbial biomass as a function of treatment (fungicides and plant growth regulator—PGR) and wheat varieties (Skyfall, SY Moisson, Aquilante, Bandera, Tintoretto, Antille, and Bologna) at the sowing, heading, and harvesting stage. A total of 168 samples (2 treatments × 7 varieties × 3 field replicates × 4 sub-samples taken in each plot) were collected in each period and analyzed. We found that soil microbial biomass was a sensible indicator in the fungicide/PGR application, with reduced values in treated plots at the heading. At this stage, the soil enzymatic activities were in general more expressed, confirming that the microbial processes are more proactive due to the growth of plants. Overall, the soil enzymatic activities responded differently according to the wheat varieties, highlighting specific capabilities to interact with microbes. Full article

The substrate in the aquatic environment plays a crucial role in nutrient deposition and recovery for the growth of aquatic organisms. In order to optimize the culture medium of Procambarus Clarkii, culture media from different sources were selected in this study to explore their effects on the growth and immune performance of red swamp crayfish. The results showed that the weight gain rate (WGR), body length growth rate (BLGR) and specific growth rate (SGR) in group I2 were the highest, followed by group I1 and group I3. The WGR and SGR of crayfish in the I1 and I2 groups were significantly higher than those in the I3 group (p < 0.05). The activities of acid phosphatase (ACP), alkaline phosphatase (AKP) and superoxide dismutase (SOD) were the highest in group I2, followed by group I3, and the lowest in group I1. The expression trends in growth-related genes, nuclear hormone receptor (E75), molt-inhibiting hormone (MIH) and chitinase genes were similar, and the expression levels in the I2 group were higher than those in the I1 and I3 groups. It was noted that the expression levels of E75 and MIH genes in the I2 group were significantly higher than those in the I3 group (p < 0.05). α diversity analysis of 16S rRNA data showed that there was no statistically significant difference in the abundance of intestinal flora among the three culture substrate groups. The β diversity in the Xitangni group, crayfish Tangni group and Shuitangni group was significantly different. These changes in microbiota suggest that using different substrates to culture crayfish leads to differences in gut microbiota diversity. To sum up, the growth in crayfish and immune performance influenced by the culture substrate condition and aquatic breeding sediment substrates, rather than crab pool and paddy field pond sediment substrates, showed a better effect. Full article

This study examined the biological activity and genome of Bacillus cereus CDHWZ7 isolated from the root of Lycium ruthenicum in the Dachaidan saline area, Haixi Prefecture, Qinghai Province, China. The results revealed that B. cereus CDHWZ7 exhibited strong inhibition activity against the pathogenic fungi Fusarium graminearum, F. acuminatum, and F. oxysporum. CDHWZ7 also demonstrated cellulose-degrading activity, nitrogen-fixing activity, and the ability to secrete indole-3-acetic acid (IAA) at 55.00 mg∙L⁻¹. The strain CDHWZ7 can grow at a salt concentration of 3–11%, a pH range of 5–11, and a temperature of 4 °C–18 °C, and shows good salt tolerance, acid and alkaline tolerance, and low-temperature fitness. The genome of strain CDHWZ7 was sequenced using Illumina HiSeq + PacBio, revealing a circular structure of 5,648,783 bp in length, containing two intact plasmids with an average GC content of 35.2%, and a total number of 5672 encoded genes. It contained 106 tRNA genes, 42 rRNA genes, and 134 sRNA genes. A total of 137 genes were annotated as carbohydrases, with a total base length of 3,968,396,297 bp. The numbers of coding sequences assigned to the Kyoto Encyclopedia of Genes and Genomes, Clusters of Orthologous Groups of Proteins, and Gene Ontology Databases were 4038, 4133, and 2160, respectively. Further analysis of the genome identified genes encoding chitinase activity, cellulases, secondary metabolites, phytohormone production, volatile compounds, nitrogen and phosphate metabolism, and resistance responses to biotic stresses (glycine betaine transporter protein, catalase, superoxide dismutase, low-affinity potassium transporter protein, cold-shock protein, heat-shock protein), as well as genes related to proliferation, stress response, and resistance to pathogenic fungi. Therefore, this study determined that strain CDHWZ7 has several excellent biological traits, such as antagonism to pathogenic fungi, nitrogen-fixation ability, cellulose-degradation ability, and IAA-production ability. The genome sequence of strain CDHWZ7 and several biodefense functional genes were also analyzed, revealing the potential use of strain CDHWZ7 in the development of biological agents. Full article

Four chitinases were cloned and characterized from three strains isolated from a mudflat: Aeromonas sp. SK10, Aeromonas sp. SK15, and Chitinibacter sp. SK16. In SK10, three genes, Chi18A, Pro2K, and Chi19B, were found as a cluster. Chi18A and Chi19B were chitinases, and Pro2K was a metalloprotease. With combinatorial amplification of the genes and analysis of the hydrolysis patterns of substrates, Chi18A and Chi19B were found to be an endochitinase and exochitinase, respectively. Chi18A and Chi19B belonged to the glycosyl hydrolase family 18 (GH18) and GH19, with 869 and 659 amino acids, respectively. Chi18C from SK15 belonged to GH18 with 864 amino acids, and Chi18D from SK16 belonged to GH18 with 664 amino acids. These four chitinases had signal peptides and high molecular masses with one or two chitin-binding domains and, interestingly, preferred alkaline conditions. In the activity staining, their sizes were determined to be 96, 74, 95, and 73 kDa, respectively, corresponding to their expected sizes. Purified Chi18C and Chi18D after pET expression produced N,N′-diacetylchitobiose as the main product in hydrolyzing chitooligosaccharides and colloidal chitin. These results suggest that Chi18A, Chi18C, and Chi18D are endochitinases, that Chi19B is an exochitinase, and that these chitinases can be effectively used for hydrolyzing natural chitinous sources. Full article

Red maple leaf extracts (RME) were tested for their plant defense inducer (PDI) properties. Two extracts were obtained and compared by different approaches: RME1 using ethanol–water (30–70%, v/v, 0.5% HCl 1N) and RME2 using pure water. Both extracts titrated at 1.9 g L⁻¹ in polyphenols and infiltrated into tobacco leaves efficiently induced hypersensitive reaction-like lesions with topical accumulation of auto-fluorescent compounds noted under UV and scopoletin titration assays. The antimicrobial marker PR1, β⁻¹,3-glucanase PR2, chitinase PR3, and osmotin PR5 target genes were all upregulated in tobacco leaves following RME1 treatment. The alkaline hydrolysis of RME1 and RME2 combined with HPLC titration of gallic acid revealed that gallate functions were present in both extracts at levels comprised between 185 and 318 mg L⁻¹. HPLC-HR-MS analyses and glucose assay identified four gallate derivatives consisting of a glucose core linked to 5, 6, 7, and 8 gallate groups. These four galloyl glucoses possessed around 46% of total gallate functions. Their higher concentration in RME suggested that they may contribute significantly to PDI activity. These findings define the friendly galloyl glucose as a PDI and highlight a relevant methodology for combining plant assays and chemistry process to their potential quantification in crude natural extracts. Full article

The aim of this study was to determine the effect of a Douglas fir plantation along a stand chronosequence in the North Apennine (Italy) on soil carbon and nitrogen stocks, as well as on soil chemical and biochemical properties involved in the nutrients biogeochemical cycle. In 2014, three sites of Douglas fir stands, aged 80, 100, and 120 years, were selected in Vallombrosa forest to study the dynamics of soil nutrients in the ecosystem. Along the Douglas fir chronosequence, general evidence of surface element accumulation was found, including a conspicuous increase of alkaline element with respect to Al, which was attributed to the increase of soil pH along the Douglas fir stand age classes. A general increase of specific enzyme activity (per unit of organic carbon) and functional diversity were observed in the epipedon of the Douglas fir stand over 100 years of age. Moreover, the (chitinase + leucine aminopeptidase) to acid phosphatase ratio progressively increased from 0.15 to 0.31 in the epipedon of the chrononsequence, while the β-glucosidase to (chitinase + leucine aminopeptidase) ratio decreased from 1.45 to 0.83, suggesting nitrogen limitation with respect to carbon. In fact, the soil carbon stock progressively increased along the chronosequence, in the epipedon from 17 to 53 Mg C ha⁻¹ and in the endopedon from 17 to 37 Mg C ha⁻¹. Conversely, the soil nitrogen stock increased from 1.2 to 2.4 Mg N ha⁻¹, but not over the 100-year-old stand class. In conclusion, soil organic matter accumulation became sufficient to define the umbric horizon in the Northern Apennines when the Douglas fir plantation reached the age of 100 years. Over this age class of plants, a limitation of soil nitrogen may occur, affecting enzyme activities regulating the biogeochemical cycle of nutrients. Full article