Mycelial elongation and sporulation of two fungi on amended media in light or dark

Botrytis allii andCollectotrichum dematium are onion pathogens which can infect in the field and cause decay in storage. Some phenolics can hinder development of these fungi, but the effect of cytokinins is not clear. Cytokinins (kinetin or 6-benzyladenine) or phenolics (caffeic or chlorogenic acids) were added to agar at concentrations of 0 to 10^–3 M. Cultures were continuously irradiated with fluorescent light or maintained in the dark for 6 days. On unamended media, final mycelial elongation was 45 or 17.8 mm and sporulation was 28 or 10.6 × 10⁴ spores/ml forBotrytis andColletotrichum, respectively. ForBotrytis, mycelial elongation was slightly (5%) but significantly increased and sporulation increased by 21% by incubation on phenolics as compared to cytokinins. Mycelial extension ofColletotrichum was not affected by amendment. Sporulation ofColletotrichum on kinetin was 16 to 28% greater than on the other amendments. As amendments concentration increased elongation of mycelia of both fungi decreased. Sporulation ofBotrytis increased by 60% as amendment concentration increased from 0 to 10^–5 M and then decreased 25% at 10^–3 M. As amendment concentration increased from 0 to 10^–3 M, sporulation ofColletotrichum increased by 45%. Incubation in light increased mycelial extension 3 to 17% forBotrytis andColletotrichum respectively, and sporulation was increased approximately 78% for both fungi. These compounds do not appear to inhibit development of theseBotrytis orColletotrichum species in culture.     

Regulatory mutations affecting the synthesis of pectate lyase in Xanthomonas campestris

Four classes of Xanthomonas campestris mutants were identified with respect to pectate lyase. Pectate lyase production in the wild-type and classes I and IIb mutants was partially dependent on the growth-phase whereas in classes IIa and III it was totally dependent. Enzyme activity in some of the mutants was constitutive and resistant to catabolite repression.     

N-phenyl ureidobenzenesulfonates,a novel class of promising human dihydroorotate dehydrogenase inhibitors

N-phenyl ureidobenzenesulfonates (PUB-SOs) is a new class of promising anticancer agents inducing replication stresses and cell cycle arrest in S-phase. However, the pharmacological target of PUB-SOs was still unidentified. Consequently, the objective of the present study was to identify and confirm the pharmacological target of the prototypical PUB-SO named 2-ethylphenyl 4-(3-ethylureido)benzenesulfonate (SFOM-0046) leading to the cell cycle arrest in S-phase. The antiproliferative and the cytotoxic activities of SFOM-0046 were characterized using the NCI-60 screening program and its fingerprint was analyzed by COMPARE algorithm. Then, human dihydroorotate dehydrogenase (hDHODH) colorimetric assay, uridine rescuing cell proliferation and molecular docking in the brequinar-binding site were performed. As a result, SFOM-0046 exhibited a mean antiproliferative activity of 3.5 μM in the NCI-60 screening program and evidenced that leukemia and colon cancer cell panels were more sensitive to SFOM-0046. COMPARE algorithm showed that the SFOM-0046 cytotoxic profile is equivalent to the ones of brequinar and dichloroallyl lawsone, two inhibitors of hDHODH. SFOM-0046 inhibited the hDHODH in the low nanomolar range (IC₅₀ = 72 nM) and uridine rescued the cell proliferation of HT-29, HT-1080, M21 and MCF-7 cancer cell lines in the presence of SFOM-0046. Finally, molecular docking showed a binding pose of SFOM-0046 interacting with Met43 and Phe62 present in the brequinar-binding site. In conclusion, PUB-SOs and notably SFOM-0046 are new small molecules hDHODH inhibitors triggering replication stresses and S-phase arrest.     

Chemophenetics of Azorean Leontodon taxa (Cichorieae,Asteraceae)

The genera Leontodon s.str. and Hedypnois are so far the only known sources of hydroxyhypocretenolides, a rare subclass of guaianolide type sesquiterpene lactones. In this study the three endemic species from the Azorean Archipelago, L. filii, L. hochstetteri, and L. rigens, were analyzed together with L. hispidus and L. × grassiorum, a hybrid originating from L. hispidus and L. hochstetteri. Flowering heads were analyzed by UHPLC-DAD-MS with regards to their phenolics' profiles, establishing qualitatively identical profiles for all taxa. The following phenolics were detected in flowering heads of all investigated taxa: caffeoyltartaric acid, cichoric acid, chlorogenic acid, 3,5-di-O-caffeoylquinic acid, luteolin, luteolin 7-O-β-d-glucopyranoside, luteolin 4′-O-β-d-glucopyranoside, and luteolin 7-O-β-d-glucuronide.In UHPLC-DAD-MS analyses of the rhizomes, no flavonoids were detected. In rhizomes, caffeoyltartaric acid was only detected in L. hispidus. However, in addition to caffeoylquinic acid derivatives already found in the flowering heads, 1,5-di-O-caffeoylquinic acid, 3,4-di-O-caffeoylquinic acid, and 4,5-di-O-caffeoylquinic acid were detected in rhizomes of all investigated taxa.The chemophenetically most interesting group of hydroxyhypocretenolides was detected in rhizomes of all investigated taxa. 11,13β-Dihydro-14-dihydroxyhypocretenolide was detected in L. filii and L. hochstetteri, while 11,13β-dihydro-14-hydroxyhypocretenolide-β-d-glucopyranoside was present in all Azorean taxa. 1,10-Epoxy-14-hydroxyhypocretenolide-β-d-glucopyranoside and 1,10-epoxy-14-hydroxyhypocretenolide-β-d-glucopyranoside-6′-O-p-hydroxyphenylacetic acid ester were restricted to the Azorean taxa and the hybrid L. × grassiorum, while the dimeric sesquiterpenoid 14-hydroxyhypocretenolide-β-d-glucopyranoside-4′,14″-hydroxyhypocretenoate ester was restricted to L. hispidus and L. × grassiorum.     

Transaminase encoded by NCgl2515 gene of Corynebacterium glutamicum ATCC13032 is involved in γ-aminobutyric acid decomposition

Corynebacterium glutamicum that expresses an exogenous l-glutamate decarboxylase (GAD) gene can synthesize γ-aminobutyric acid (GABA). GABA is decomposed to succinic semialdehyde (SSA) by GABA transaminase (GABA-T) and to succinate thereafter by SSA dehydrogenase (SSADH). However, deletion of the gabT gene encoding GABA-T could not prevent GABA from decomposing at neutral pH. In this study, an additional transaminase gene, NCgl2515, was deleted in a gabT-deleted GAD strain, and GABA fermentation in this gabT NCgl2515-deleted GAD strain was investigated. GABA concentration remained at 22.5–24.0 g/L when pH was maintained at 7.5–8.0, demonstrating that GABA decomposition was reduced. Activity assay indicated that unlike GabT, which exhibits high GABA-T activity (1.34 ± 0.06 U/mg) and utilizes only α-ketoglutarate as amino acceptor, the purified NCgl2515 protein exhibits very low GABA-T activity (approximately 0.03 U/mg) only when coupled with the SSADH, GabD, but can utilize both α-ketoglutarate and pyruvate as amino acceptor. The optimum pH for coupled NCgl2515–GabD was 8.0, similar to that of GabT (7.8). Therefore, NCgl2515 has weak GABA-T activity and is involved in GABA decomposition in C. glutamicum. Deletion of gabT and NCgl2515 could effectively reduce GABA decomposition at neutral pH.     

The oxalyl‐CoA synthetase‐regulated oxalate and its distinct effects on resistance to bacterial blight and aluminium toxicity in rice

• Oxalic acid is widely distributed in biological systems and known to play functional roles in plants. The gene AAE3 was recently identified to encode an oxalyl‐CoA synthetase (OCS) in Arabidopsis that catalyses the conversion of oxalate and CoA into oxalyl‐CoA. It will be particularly important to characterise the homologous gene in rice since rice is not only a monocotyledonous model plant, but also a staple food crop.
• Various enzymatic and biological methods have been used to characterise the homologous gene.
• We first defined that AAE3 in the rice genome (OsAAE3) also encodes an OCS enzyme. Its K_m for oxalate is 1.73 ± 0.12 mm , and V_m is 6824.9 ± 410.29 U·min^?1·mg protein^?1. Chemical modification and site‐directed mutagenesis analyses identified thiols as the active site residues for rice OCS catalysis, suggesting that the enzyme might be regulated by redox state. Subcellular localisation assay showed that the enzyme is located in the cytosol and predominantly distributed in leaf epidermal cells. As expected, oxalate levels increased when OCS was suppressed in RNAi transgenic plants. More interestingly, OCS‐suppressed plants were more susceptible to bacterial blight but more resistant to Al toxicity.
• The results demonstrate that the OsAAE3‐encoded protein also acts as an OCS in rice, and may play different roles in coping with stresses. These molecular, enzymatic and functional data provide first‐hand information to further clarify the function and mechanism of OCS in rice plants.

     

New pyranonaphthazarin and 2-naphthoic acid derivatives from the mangrove endophytic fungus Leptosphaerulina sp. SKS032

Two new compounds, named leptospyranonaphthazarin A (1) and leptosnaphthoic acid A (2), together with four known compounds (3–6) were isolated from an endophytic fungus Leptosphaerulina sp. SKS032. Their structures were assigned using spectroscopic methods, computational methods, and single-crystal X-ray diffraction analysis. In the antibiotic assay, compounds 1, 2, and 6 exhibited antibacterial activities against Staphylococcus aureus with minimal inhibitory concentration (MIC) values of 25.0, 50.0, and 50.0 μg/mL, respectively.     

一株耐受低pH、高浓度硒菌株的筛选鉴定及两阶段pH调控高效除硒的研究

通过酸性含硒平板和摇瓶筛选出一株对低pH、高浓度硒有很好耐受性的菌株Y1,通过菌落形态特征分析和26S rDNA测序,鉴定该菌株为Pichia kudriavzevii,多抗性实验结果显示该菌是一株多重耐受性毕赤酵母。通过摇瓶实验研究了温度、接种量、摇床转速、pH对菌株除硒性能的影响,结果显示当温度为25℃,接种量为12%(v/v),摇床转速为250 r/min,pH为3.0时,菌株对硒的去除率最高为58.3%。基于不同pH发酵过程中菌体生物量及富硒量的不同表现:pH 3.0时生物量最高,pH 5.0时富硒量最高,提出两阶段pH调控策略:发酵0 h~14 h将pH控制在3.0,14 h~28 h将pH控制在5.0,最终除硒率可达78.6%,分别比pH恒定在3.0及5.0条件下提高了15.4%和21.7%。