Gut symbiotic bacteria have a substantial impact on host physiology and ecology. However, the contribution of gut microbes to host fitness during long-term low-temperature stress is still unclear. This study examined the role of gut microbiota in host low-temperature stress resistance at molecular and biochemical levels in the oriental fruit fly Bactrocera dorsalis. The results showed that after the gut bacteria of flies were removed via antibiotic treatment, the median survival time was significantly decreased to approximately 68% of that in conventional flies following exposure to a temperature stress of 10˚C. Furthermore, we found that Klebsiella michiganensis BD177 is a key symbiotic bacterium, whose recolonization in antibiotic treated (ABX) flies significantly extended the median survival time to 160% of that in the ABX control, and restored their lifespan to the level of conventional flies. Notably, the relative levels of proline and arginine metabolites were significantly downregulated by 34and 10-fold, respectively, in ABX flies compared with those in the hemolymph of conventional flies after exposure to a temperature stress of 10˚C whereas recolonization of ABX flies by K. michiganensis BD177 significantly upregulated the levels of proline and arginine by 13-and 10-fold, respectively, compared with those found in the hemolymph of ABX flies. qPCR analysis also confirmed that K. michiganensis-recolonized flies significantly stimulated the expression of transcripts from the arginine and proline metabolism pathway compared with the ABX controls, and RNAi mediated silencing of two key genes Pro-C and ASS significantly reduced the survival time of conventional flies, postexposure low-temperature stress. We show that microinjection of L-arginine and L-proline into ABX flies significantly increased their survival time following exposure to temperature stress of 10˚C. Transmission electron microscopy (TEM) analysis further revealed that low-temperature stress caused severe destruction in cristae structures and thus resulted in abnormal circular shapes of mitochondria in ABX flies gut, while the recolonization of live K. michiganensis helped the ABX flies to maintain mitochondrial functionality to a normal status, which is important for the arginine and proline induction. Our results suggest that gut microbiota plays a vital role PLOS PATHOGENS PLOS Pathogens | https://doi.
Salmonella enterica serovar Enteritidis (SE) infection of chickens is a major contributing factor to non-typhoidal salmonellosis. The roles of the type-three secretion systems (T3SS-1 and T3SS-2) in the pathogenesis of SE infection of chickens are poorly understood. In this study, the functions of T3SS-1 and T3SS-2 during SE infection of primary chicken oviduct epithelial cells (COEC) and macrophages were characterized. The T3SS-1 and T3SS-2 mutants (sipB and ssaV), impaired in translocation and secretion, respectively, were significantly less invasive than their wild-type parent strain. The genes encoding effector proteins of T3SS-1 (SipA, SopB, and SopE2) and T3SS-2 (PipB) contributed equally to the entry of SE into COEC. The sipA mutant had reduced survival and the pipB mutant had enhanced replication in COEC. Mutations in the T3SS-2 genes ssaV and pipB reduced the survival of SE in chicken peripheral blood leukocyte-derived macrophages (PBLM), but not in the established chicken macrophage cell line HD11. A mutation in the ssaV gene also abolished SE-induced PBLM death between 1 h post-inoculation and 4 h post-inoculation. This study has shown that both T3SS-1 and T3SS-2 are required by SE to invade COEC; that SipA and PipB are necessary for the survival of SE in COEC and chicken PBLM, respectively; and that T3SS-2 triggers PBLM death during the early stages of SE infection, and this process does not depend on PipB.
In this work, hydroxyl-enriched glucose-based carbonaceous sphere (HEGCS) is prepared by hydrothermal method as an accelerator to enhance the hydrolytic efficiency of the treated cellulose to sugar in low-acidic aqueous system under microwave radiation. Due to the strong affinity of HEGCS to cellulose, during the hydrolysis, the treated cellulose can be flaked into small fragments by HEGCS, which is like a “microball milling”, and accordingly, the hydrolytic accessibility of cellulose is extremely improved. Highly efficient hydrolysis of cellulose to sugar is finally achieved in HEGCS containing 0.02 mol/L sulfuric acid. One hundred percent conversion of cellulose and 96.0 ± 4.0% total reducing sugar (TRS) yield was obtained within 15 min under microwave radiation. The average content of glucose in sugar products is over 70%. Our research not only obtains a catalytic accelerator to establish an effective method for hydrolyzing treated cellulose to sugar but also provides the idea for regulating cellulose accessibility during hydrolysis to achieve high efficiency in the hydrolysis of cellulose.
Nattokinase is a serine protease in the subtilisin family which is produced by Bacillus subtilis subsp. natto and exhibits vigorous fibrinolytic activity that has been suggested to be able to prevent and treat thromboembolic diseases. In this study, WTC016, a spore-forming and rod-shaped bacterium with fibrinolytic activity was successfully isolated from soil, which was identified as Bacillus subtilis subsp. natto based on morphological and physiological tests, and phylogenetic analysis of 16S rRNA and gyrA. According to the growth curve of WTC016, the nattokinase production reached the highest amount in the stationary phase. To optimize the liquid fermentation condition for nattokinase yield of WTC016, further optimal tests of four factors, including the temperature, pH, inoculum size, and loading volume, followed by orthogonal test of all these factors, was performed. The optimal fermentation conditions were determined as 30 °C, 7.0 pH, 2% inoculum size, and 60 mL of loading volume in 250 mL conical flask, which indicates the highest nattokinase production of 3284 ± 58 IU/mL while fermented for 26 h. This work laid the foundation for producing nattokinase using Bacillus subtilis subsp. natto WTC016.
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