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Background 9α-hydroxyandrost-4-ene-3,17-dione (9-OHAD) refers to a significant intermediate for synthesis of glucocorticoid drugs. Microbial transformation of phytosterols to manufacture 9-OHAD has obvious advantages in terms of lead time, cost control, environmental friendliness, etc. However, the product degradation and the formation of by-products restrict their application. In this study, a combined strategy of blocking Δ1‑dehydrogenation and regulating metabolic flow was investigated to construct a stable high-yield 9-OHAD producer. Results Five 3-Ketosteroid-Δ1-dehydrogenase (KstD) were identified in ATCC 35855 and each of them was functionally confirmed by heterologous expression, transcriptional response and knockout analysis. KstD2 showed the highest catalytic activity on 3-ketosteroids, followed by KstD3, KstD1, KstD4 and KstD5. In particular, KstD2 has a much higher catalytic activity for C9 hydroxylated steroids than for C9 non-hydroxylated steroids, whereas KstD3 showed the opposite characteristics. The deletion of kstDs indicated that KstD2 and KstD3 are the main assailants of 9-OHAD degradation. Compared with ATCC 35855, MFΔkstD, the five KstDs deficient train, realized the stable accumulation of 9-OHAD, and its yield increased by 42.57%. The knockout of Opccr or the overexpression of Hsd4A alone could not reduce the metabolic flow of C22 pathway, while the overexpression of Hsd4A based on the knockout of Opccr in MFΔkstD could remarkably reduce the contents of 9-OHHP by-products. The inactivation of FadE28-29 leads to a large amount of accumulation of incomplete side chain degradation products. Therefore, hsd4A and fadE28-29 were co-expressed in MFΔkstDΔopccr successfully eliminated two by-products. Compared with MFΔkstD, the purity of 9-OHAD was improved from 80.24% to 90.14%. Ultimately, 9‑OHAD production reached 12.21 g/L (83.74% molar yield) and the productivity of 9-OHAD was 0.0927 g/L/h. Conclusions Five putative KstDs have been identified and functionally confirmed in ATCC 35855. KstD2 and KstD3 were the main murderers of 9-OHAD degradation. Hsd4A and Opccr are key enzymes regulating the metabolic flux of C19 and C22-pathways. The overexpression of fadE28-29 can reduce the accumulation of incomplete degradation products of side chains. According to the findings above, MF-FA5020 transformant was successfully constructed to rapidly and stably accumulate 9-OHAD from phytosterols. These results contribute to understanding the diversity and complexity of steroid catabolism regulation in actinobacteria and providing a theoretical basis for further optimizing industrial microbial catalyst.

SUMMARY On 2019 March 21, an explosion accidentally occurred at a chemical plant in Xiangshui, Yancheng City, Jiangsu Province, China. Using broad-band digital seismic data from East China, South Korea and Japan, we investigate properties of the Xiangshui explosion as well as two nearby chemical explosions and four nearby natural earthquakes in Jiangsu Province, East China. From Lg and Rayleigh waves recorded by regional networks, both body wave magnitude mb (Lg) and surface wave magnitude Ms (Rayleigh) are calculated for these events. The magnitudes of the Xiangshui explosion are mb (Lg) = 3.39 ± 0.24 and Ms = 1.95 ± 0.27, respectively. Both the empirical magnitude–yield relation for buried explosion and empirical yield–crater dimension relation for open-pit explosion are adopted for investigating the explosive yield. The result from the yield–crater dimension relation is approximately 492 ton, which is consistent with the ground truth and considerably larger than that from the buried source model. This also reveals that, for Xiangshui explosion, the explosion to seismic energy conversion rate is approximately one-third compared to a similar sized fully confined explosion. By comparing the body wave and surface wave magnitudes from explosions and nearby earthquakes, we find that the mb:Ms discriminant calculated at regional distances cannot properly distinguish explosions from natural earthquakes. However, the P/S spectral ratios Pg/Lg, Pn/Lg and Pn/Sn from the same data set can be good discriminants for identifying explosions from earthquakes.

Simple Summary Airflow is very important for the long-distance migration of O. decorus asiaticus, and wind shear, in particular, is the main factor related to forced landing. Analyzing the weather records, we found that the northwest wind prevailed when the population invaded. Specifically, from July to August, a large number of emerging adults appeared in the source areas of Mongolia, and the large-scale northwest wind was the decisive condition for the successful long-distance migration of O. decorus asiaticus. The species has a strong migratory ability, flying along the airflow for several nights. If the northwest air current meets the southwest warm current going north, a large number of O. decorus asiaticus will drop due to wind shear, and then a major outbreak will occur. Analysis of the source of the insects shows that the O. decorus asiaticus break outs in China may have originated from Mongolia. They were brought into China by the southerly airflow at night, and they likely made a forced landing in Beijing due to wind shear, sinking airflow, rainfall and other reasons. In summary, through analysis of the insect’s prevalence and the meteorological conditions in Mongolia, we can provide a basis for predicting the occurrence of O. decorus asiaticus in China. Abstract Oedaleus decorus asiaticus (Bey-Bienko) is a destructive pest in grasslands and adjacent farmland in northern China, Mongolia, and other countries in Asia. It has been supposed that this insect pest can migrate a long distance and then induce huge damages, however, the migration mechanism is still unrevealed. The current study uses insect light trap data from Yanqing (Beijing), together with regional meteorological data to determine how air flow contributes to the long-distance migration of O. decorus asiaticus. Our results indicate that sinking airflow is the main factor leading to the insects’ forced landing, and the prevailing northwest wind was associated with O. decorus asiaticus taking off in the northwest and moving southward with the airflow from July to September. Meanwhile, the insects have a strong migratory ability, flying along the airflow for several nights. Thus, when the airflow from the northwest met the northward-moving warm current from the southwest, a large number of insects were dropped due to sinking airflow, resulting in a large outbreak. Our simulations suggest that the source of the grasshoppers involved in these outbreaks during early 2000s in northern China probably is in Mongolia, and all evidence indicates that there are two important immigrant routes for O. decorus asiaticus migration from Mongolia to Beijing. These findings improves our understanding of the factors guiding O. decorus asiaticus migration, providing valuable information to reduce outbreaks in China that have origins from outside the country.

BackgroundSteroid drugs are particularly important for disease prevention and clinical treatment. However, traditional chemical methods are rarely implemented during the whole synthetic process to generate steroid intermediates due to the intricate steroid structure. Novel steroid drug precursors and their ideal bacterial strains for industrial production have yet to be developed. Among these, 9-OH-4-HP is a potential steroid drug precursor for the synthesis of corticosteroids. In this study, a combined strategy of blocking Δ1-dehydrogenation and the C19 pathway as well as improving the intracellular environment was investigated to construct an effective 9-OH-4-HP-producing strain.ResultsA Δ1-dehydrogenation-deficient strain of wild-type Mycobacterium neoaurum DSM 44074 produces 9-OH-4-HP with a molar yield of 4.8%. hsd4A, encoding a β-hydroxyacyl-CoA dehydrogenase, and fadA5 encoding an acyl-COA thiolase, were separately knocked out to block the C19 pathway in the Δ1-dehydrogenation-deficient strain. The two engineered strains could accumulate 0.59 g L-1 and 0.47 g L-1 9-OH-4-HP from 1 g L-1 phytosterols. Furthermore, hsd4A and fadA5 were knocked out simultaneously in the Δ1-dehydrogenation-deficient strain. The 9-OH-4-HP production from the Hsd4A and FadA5 double-deficient strain was 11.9% higher than that of the Hsd4A -deficient strain and 40.4% higher than that of the strain with FadA5 deficiency, and its selectivity reached 94.9%. Subsequently, the catalase katE from Mycobacterium and an NADH oxidase, nox, from Bacillus subtilis were overexpressed to improve the intracellular environment. Ultimately, 9-OH-4-HP production reached 3.58 g L-1 from 5 g L-1 phytosterols, and the selectivity of 9-OH-4-HP improved to 97%.Conclusionhsd4A and fadA5 are key enzymes in the C19 pathway for phytosterol side chain degradation. Deletion of hsd4A and fadA5 could almost entirely block the C19 pathway. Improving the intracellular environment of Mycobacterium during phytosterol bioconversion could accelerate the conversion process and enhance the productivity of target sterol derivatives.

This study explored how a thermophilic microbial agent altered nitrogen transformation, nitrogen functional genes, and bacterial communities during bean dregs composting with (T) and without (CK) a thermophilic microbial agent for 15 days. The results showed that the maximum temperature in T reached 73 °C and remained above 70 °C for 8 days, while that in CK was only 65 °C. The pH in T had essentially stabilized on day 7, while that in CK was still increasing. On day 15, the seed germination index (GI) of T (95%) reached maturity (defined by GI ≥ 85%), while the GI of CK was only 36%. The concentrations of total nitrogen, water-soluble nitrogen, ammonia nitrogen, and nitrate nitrogen in T (2.5%, 18.9 g/kg, 8.75 g/kg, and 1.69 g/kg) were all lower than those in CK (3.6%, 28.9 g/kg, 12.75 g/kg, and 6.82 g/kg). During composting, Bacillus played a major role in nitrogen reduction, nitrogen mineralization, denitrification, and the conversion between nitrite and nitrate. Weissella played a major role in nitrogen assimilation. Komagataeibacter and Bacillus played a major role in nitrogen fixation in CK and T, respectively. Nitrification was not observed during composting. The nosZ gene, which converts nitrous oxide to nitrogen, was found only in T. Network analysis suggested that the average number of neighbours in T was 3.30% higher than that in CK and the characteristic path length in T was 14.15% higher than that in CK. Therefore, the thermophilic microbial agents could cause nitrogen loss but promote the maturity of bean dregs, which have great potential application.

Ensiling has been widely applied to cope with agricultural solid waste to achieve organic waste valorization and relieve environmental pressure and feedstuff shortage. In this study, co-ensiling of cabbage leaf byproduct and rice straw was performed with inoculation of Lactiplantibacillusplantarum (LP) to investigate the effects of inoculation on ensiling performance and microflora profiles. Compared to the control, LP inoculation preserved more dry matter (DM) content (283.4 versus 270.9 g·kg−1 fresh matter (FM) on day 30), increased lactic acid (LA) content (52.1 versus 35.8 g·kg−1 dry matter on day 15), decreased pH (3.55 versus 3.79 on day 15), and caused accumulation of acetic acid (AA), butyric acid (BA), and ammonia. The investigation showed that LP inoculation modified microflora composition, especially resisting potential pathogens and enriching more lactic acid bacteria (LAB) (p &lt, 0.05). Moreover, Lactiplantibacillus and Lacticaseibacillus were identified as the keystone taxa that influenced physicochemical properties and interactions in microflora. They were also the main functional species that directly restrained undesirable microorganisms (p &lt, 0.05), rather than indirectly working via metabolite inhibition and substrate competition (p &gt, 0.05). The results of this present study improve the understanding of the underlying effect of LP inoculation on improving silage quality and facilitate the bio-transformation of cabbage byproduct and rice straw as animal feed.

Hydrogeochemical characteristics, controlling factors, and recharge sources of the seismic observation wells in Shandong Province were investigated by analyzing cation and anion concentrations, hydrogen and oxygen isotopes in well water. A total of 17 water samples in seismic observation wells were collected on April 25-29, 2018. The results show that temperatures of seismic observation wells were in the range of 14.8 to 52.1°C, and the values of δD and δ 18 O ranged from -72.4‰ to -37.9‰ and from -9.4‰ to -4.3‰, respectively. Using C.A. ЩукаЛев’s classification method, the water samples of 17 seismic observation wells were classified into 7 types: Cl·SO 4 -Na·Ca, SO 4 -Na, Cl-Na, HCO 3 -Na·Ca, HCO 3 -Mg·Na·Ca, HCO 3 -Na and HCO 3 -Mg·Ca·Na. The results indicate that the hydrogeochemical characteristics of 17 seismic observation wells, with a certain spatial distribution pattern, are affected by several factors, such as the tectonic, topography, stratigraphy, hydrology and meteorology. The analyses of ratio coefficients, Schooller diagram, hydrogen and oxygen isotopes compositions, Giggenbach and Gibbs diagram suggest that the atmospheric precipitation is the main recharge source of 17 observation wells. The recharge sources of deep lateral runoff and sedimentation water, moreover, play a significant role in some seismic observation wells. Combined with the amount of precipitation, the distance from recharge areas, the closure degree of observation wells and the stage of water-rock reactions, the development directions of faulting and topography control the directions of groundwater recharge, runoff and discharge, which make hydrogeochemical characteristics represent complex spatial distribution rules.

In order to explore the synergistic control effect of crude extracts of Artemisia sieversiana and Metarhizium anisopliae on Oedaleus asiaticus, we used different doses of M. anisopliae and crude extracts of A. sieversiana singly and in combination, to determine their toxicities to fourth instar O. asiaticus. The results showed that the combination of 10% crude extract of A. sieversiana with 107 and 108 spores/g M. anisopliae concentrations and the combination of 20% crude extract of A. sieversiana with 107 and 108 spores/g M. anisopliae concentrations had significant effects on the mortality, body weight gain, body length gain, growth rate, and overall performance of O. asiaticus than those of the crude extract of A. sieversiana and M. anisopliae alone. Among them, the 20% A. sieversiana crude extract mixed with 108 spores/g M. anisopliae and 10% A. sieversiana crude extract combined with 107 spores/g M. anisopliae, had the best control efficacy. In order to clarify the biochemical mechanism underlying the immune responses of O. asiaticus to the pesticide treatments, we monitored the activities of four enzymes: superoxidase dismutase (SOD), peroxidase (POD), catalase (CAT), and polyphenol oxidase (PPO). The results showed that the activities of three enzymes (SOD, CAT, and PPO) were significantly increased from the treatment with the combination of M. anisopliae mixed with crude extract of A. sieversiana. Interestingly, compared to the crude extract, the combination treatment did not significantly induce the expression of POD enzyme activity, which may be a biochemical factor for increasing the control effect of the combination treatment. Our results showed that the combination treatment had synergistic and antagonistic effects on host mortality, growth, development, and enzyme activities in O. asiaticus.

Composting has been a rational method to dispose of horse or wildlife animal manures, especially in the developed cities with horse clubs and wildlife parks. However, few studies have focused on the mechanism and improvement methods for composting the horse or wildlife animal manures. In this study, we investigated the effect of exogenous compound enzymes on thermophilic composting, which could potentially support the management of horse and wildlife animal manures. With the presence of exogenous enzymes, the duration of high temperature ( 60 °C) was significantly prolonged (p 0.05), and the germination index was significantly improved (p 0.05). More-efficient improvement of composting maturity was associated with the addition of that exogenous enzyme that might influence microflora succession and the interaction among microorganic communities, especially fungal, during the composting process. Furthermore, redundancy and canonical correspondence analyses indicated that the C/N ratio, temperature, and germination index were significant variations to influence bacterial communities (p 0.05). The dominant Flavobacterium, Thermopolyspora, Thermomonospora, and Chaetomium and Saccobolus could play an essential role in carbohydrate and phytotoxin degradation, while Thermobispora and norank_f_Limnochordaceae could lead to temperature rising.

Migratory insect pests pose a substantial challenge to global food security. These issues are particularly acute when pest incursions occur considerably beyond the expected range, through natural migration or human-aided transport, as the lack of species-specific control strategies and a potential absence of species-specific natural enemies in the newly-invaded area, may lead to rapid establishment of a new pest. One such threat is posed by the Oriental migratory locust (Locusta migratoria manilensis) in China, which historically has been restricted to eastern China from the Bohai Gulf southwards, but now threatens to expand its range into the agriculturally important region of northeast China. We analysed data from a recent outbreak of migratory locusts in Heilongjiang (HLJ) province (extreme northeast China), >700 km north of its current known range, and identified the source region, timing of arrival and probable migratory routes of this incursion. We further show that warming temperatures in this region will likely allow subsequent invasions to establish permanent populations in northeast China, and thus authorities in this important crop-producing region of East Asia should be vigilant to the threat posed by this species.

The spotted alfalfa aphid (Therioaphis trifolii (Monell)) is a known destructive pest that can significantly reduce alfalfa yields. Two differentially up-regulated alfalfa trypsin inhibitors &lsquo, Msti-94&rsquo, and &lsquo, Msti-16&rsquo, in transcriptome were verified in terms of their mRNA levels using RT-qPCR. The prokaryotic expression vector was constructed and its biological functions, including phenotypic and physiological responses, were verified through feeding spotted alfalfa aphids with active recombinant protein mixed with an artificial diet. Gene clone and gene prokaryotic expression confirmed that Msti-94 had a size of 651 bp, encoded 216 amino acids with a predicted protein weight of 23.5 kDa, and a pI value of 6.91. Similarly, the size of Msti-16 was 612 bp, encoded 203 amino acids, and had a predicted protein weight of 22.2 kDa with a pI value of 9.06. We concluded that both Msti-94 and Msti-16 acted as a stomach poison with survival rates reduced to 21.7% and 18.3%, respectively, as compared to the control, where the survival rate was significantly (p &lt, 0.05) higher (60.0%). Aphid reproduction rates were significantly reduced, after 72 h of feeding, in both the Msti-94 and Msti-16 treatments compared to the controls. A concentration of 800 &mu, g/mL (0.8 mg/mL) of recombinant protein and 5000 &mu, g/mL (5 mg/mL) of recombinant expressing bacteria that inhibits the total protease, which ultimately disrupted the activity of trypsin, chymotrypsin, and aminopeptidase.

The fermentation of leaf vegetable waste to produce animal feed reduces the environmental impact of vegetable production and transforms leaf vegetable waste into a commodity. We investigated the effect of exogenous probiotics and lignocellulose enzymes on the quality and microbial community of fermented feed (FF) produced from cabbage waste. The addition of exogenous probiotics resulted in increased crude protein (CP) content (p &lt, 0.05), better odor (moderate organic acid and ethanol, with low ammonia-N, p &lt, 0.05), and a lower relative abundance (RA) of pathogens (below 0.4%, p &lt, 0.05) in FF, compared to without. With the addition of exogenous probiotics, only Pediococcus and Saccharomyces were enriched and symbiotic in FF, these were the keystone taxa to reduce the abundance of aerobic, form-biofilms, and pathogenic microorganisms, resulting in an efficient anaerobic fermentation system characterized by facultative anaerobic and Gram-positive bacterial communities, and undefined saprotroph fungal communities. Thus, inoculation of vegetable waste fermentation with exogenous probiotics is a promising strategy to enhance the biotransformation of vegetable waste into animal feed.

This study was supported by the National Natural Science Foundation of China, 31672485, the Earmarked Fund for China Agriculture Research System, CARS-34-07, and the Innovation Project of Chinese Academy of Agricultural Sciences. Livestock grazing can affect insects by altering habitat quality; however, the effects of grazing years and intensities on insect abundance and trophic level during manipulative sheep grazing are not well understood. Therefore, we investigated these effects in a large manipulative experiment from 2014 to 2016 in the eastern Eurasian steppe, China. Insect abundance decreased as sheep grazing intensities increased, with a significant cumulative effect occurring during grazing years. The largest families, Acrididae and Cicadellidae, were susceptible to sheep grazing, but Formicidae was tolerant. Trophic primary and secondary consumer insects were negatively impacted by increased grazing intensities, while secondary consumers were limited by the decreased primary consumers. Poor vegetation conditions caused by heavy sheep grazing were detrimental to the existence of Acrididae, Cicadellidae, primary and secondary consumer insects, but were beneficial to Formicidae. This study revealed variations in insect abundance and trophic level in response to continuous sheep grazing in steppe grasslands. Overall, our results indicate that continuous years of heavy- and over- sheep grazing should be eliminated. Moreover, our findings highlight the importance of more flexible sheep grazing management and will be useful for developing guidelines to optimize livestock production while maintaining species diversity and ecosystem health. Publisher PDF

The diagenetic processes of the Tabei sandstones in the Tarim Basin include compaction, cementation (quartz overgrowths, calcite, clay minerals and a minor amount of pyrite), and dissolution of the feldspar and calcite cement. Porosity was reduced by compaction from an assumed original 40% to about 22.1%. Cementation reduced porosity to 26.6%. The Tabei sandstones lost a little more porosity by compaction than by cementation. Quartz cementation, especially syntaxial quartz overgrowth, is a major cause of porosity-loss in many reservoirs in moderately to deeply buried sandstone. Calcite cementation played a key role in the porosity evolution of sandstones. At the early stage of burial, the early calcite cement occupied most of the pore spaces resulting in significant porosity. On the other hand, some primary porosity has been preserved due to incomplete filling or the presence of scattered patches of calcite cement. In addition to calcite, several clay minerals, including illite and chlorite occurred as pore-filling and pore-lining cements. The pore-lining chlorite may have helped in retaining the porosity by preventing the precipitation of syntaxial quartz overgrowths. Illite, which largely occurred as hair-like rims around the grains and bridges on the pore throats, caused a substantial deterioration of penetrability of the reservoir. Calcite cement dissolution was extensive and contributed significantly to the development of secondary porosity.