一株淡紫拟青霉GDIZM-2的分离鉴定及其对柑橘木虱的毒力评价

昆虫病原真菌是一类能够侵染昆虫并造成其死亡的病原微生物,在自然界中广泛存在,是农业害虫绿色防控的无公害优质资源。本研究从田间采集到的柑橘木虱Diaphorina citri成虫僵虫上分离出一株对柑橘木虱具有强致病性的病原真菌菌株。利用形态学和rDNA-ITS序列分析相结合的方法鉴定其为淡紫拟青霉Purpureocillium lilacinum,命名为GDIZM-2,并在室内测定了该菌株对柑橘木虱若虫和成虫的毒力以及在温室条件下对柑橘木虱成虫的致病力。室内毒力试验结果表明,菌株GDIZM-2可侵染柑橘木虱若虫和成虫,且随着孢子浓度和处理时间的增加,柑橘木虱若虫和成虫的校正死亡率逐渐上升,LC50和LT50值逐渐递减。当浓度为1×108孢子/mL处理第7天时,各龄期柑橘木虱的死亡率均高于75%,具有较高的致病力。温室致病力试验结果表明,淡紫拟青霉GDIZM-2在1×108孢子/mL处理第14天时,柑橘木虱成虫的存活率为1.67%,显著低于金龟子绿僵菌Metarhizium anisopliae GDIZMMa-3和无菌水。本研究结果表明,淡紫拟青霉GDIZM-2对柑橘木虱有良好的生物防治效果,具有开发成为一种新型微生物真菌制剂的应用潜力。     

Pichia koratensis sp. nov., a new ascomycetous yeast related to Pichia acaciae isolated from insect frass in Thailand

Two yeast strains (ST-235 and ST-237) isolated from insect frass collected in northeastern Thailand, were identified as Pichia acaciae and a hitherto undescribed species based on the sequence analysis of the D1/D2 domain of 26S rDNA, ITS regions, and DNA-DNA reassociation experiments. The latter strain, ST-237 (=BCC 11769=NBRC 103638=JCM 12576), was described as Pichia koratensis sp. nov. Pichia koratensis is closely related to Pichia acaciae in the D1/D2 domain sequence of 26S rDNA and phenotypical characteristics but differs by 5 nucleotides (0.9%) from the type strain of P. acaciae and is clearly distinguished by the assimilation and fermentation of sucrose.     

Identification and biodegradation potential of a novel strain of Dietzia cinnamea isolated from a petroleum-contaminated tropical soil

A bacterial strain, named P4, isolated previously from microcosms containing oil-contaminated soil collected from an environmentally protected area of a tropical Atlantic forest (Biological Reserve of Poço das Antas) located in Brazil was identified as Dietzia cinnamea by morphological, biochemical and genotypic tests. Arabian Light and Marlin oils were both degraded when strain P4 was tested for oil degradation ability in microplates. Total Petroleum Hydrocarbons (TPH) analysis, determined by gas chromatography, showed that strain P4 degraded a wide range of n-alkanes, and also pristane and phytane. Furthermore, this strain was also able to grow in mineral liquid media amended with carbazole, quinoline, naphthalene, toluene, gasoline and diesel as the sole carbon sources. The species D. cinnamea has been previously described with only one representative strain isolated from a perianal swab of a patient with a bone marrow transplant. With the results presented here this species is implicated not only as a human pathogen but also as a potential strain for further studies concerning its role for bioremediation of oil contaminated soil.     

Influence of histidine-198 of the D1 subunit on the properties of the primary electron donor, P680, of photosystem II in Thermosynechococcus elongatus

The influence of the histidine axial ligand to the PD1 chlorophyll of photosystem II on the redox potential and spectroscopic properties of the primary electron donor, P680, was investigated in mutant oxygen-evolving photosystem II (PSII) complexes purified from the thermophilic cyanobacterium Thermosynechococcus elongatus. To achieve this aim, a mutagenesis system was developed in which the psbA1 and psbA2 genes encoding D1 were deleted from a His-tagged CP43 strain (to generate strain WT*) and mutations D1-H198A and D1-H198Q were introduced into the remaining psbA3 gene. The O2-evolving activity of His-tagged PSII isolated from WT* was found to be significantly higher than that measured from His-tagged PSII isolated from WT in which psbA1 is expected to be the dominantly expressed form. PSII purified from both the D1-H198A and D1-H198Q mutants exhibited oxygen-evolving activity as high as that from WT*. Surprisingly, a variety of kinetic and spectroscopic measurements revealed that the D1-H198A and D1-H198Q mutations had little effect on the redox and spectroscopic properties of P680, in contrast to the earlier results from the analysis of the equivalent mutants constructed in Synechocystis sp. PCC 6803 [B.A. Diner, E. Schlodder, P.J. Nixon, W.J. Coleman, F. Rappaport, J. Lavergne, W.F. Vermaas, D.A. Chisholm, Site-directed mutations at D1-His198 and D2-His197 of photosystem II in Synechocystis PCC 6803: sites of primary charge separation and cation and triplet stabilization, Biochemistry 40 (2001) 9265-9281]. We conclude that the nature of the axial ligand to PD1 is not an important determinant of the redox and spectroscopic properties of P680 in T. elongatus.     

Reassortant rotaviruses as potential live rotavirus vaccine candidates.

A series of reassortants was isolated from coinfection of cell cultures with a wild-type animal rotavirus and a "noncultivatable" human rotavirus. Wild-type bovine rotavirus (UK strain) was reassorted with human rotavirus strains D, DS-1, and P; wild-type rhesus rotavirus was reassorted with human rotavirus strains D and DS-1. The D, DS-1, and P strains represent human rotavirus serotypes 1, 2, and 3, respectively. Monospecific antiserum (to bovine rotavirus, NCDV strain) or a set of monoclonal antibodies to the major outer capsid neutralization glycoprotein, VP7 (of the rhesus rotavirus), was used to select for reassortants with human rotavirus neutralization specificity. This selection technique yielded many reassortants which received only the gene segment coding for the major neutralization protein from the human rotavirus parent, whereas the remaining genes were derived from the animal rotavirus parent. Single human rotavirus gene substitution reassortants of this sort represent potential live vaccine strains.     

一株花生根际铁载体产生菌的分离鉴定及耐药性分析

目的：从花生根际筛选铁载体产生能力较强的菌株,对其进行鉴定及耐药性分析。方法：用梯度稀释法从花生根际中分离出细菌,在刃天青（CAS）检测平板上依显色圈的大小从中筛选出一株铁载体产生能力较强的菌,并对其进行生理生化鉴定和16S rDNA序列分析,用抗生素梯度平板检测其对9种常见抗生素的抗性。结果：筛选出一株产铁载体的菌株D15,13项生理生化指标除甲基红试验呈阳性外,其他均与恶臭假单胞菌相同;其16S rDNA与恶臭假单胞菌的同源性为100%;该菌对氨苄青霉素、氯霉素、利福平、红霉素、新霉素、链霉素、四环素都有不同程度的的抗性,对卡那霉素和庆大霉素表现较强的敏感性。结论：获得铁载体产生菌D15,经鉴定为恶臭假单胞菌,该菌耐药性符合用转座子诱变法研究铁载体合成的相关基因的条件。     

Antibacterial metabolites from the Actinomycete <Emphasis Type="Italic">Streptomyces</Emphasis> sp. P294

The Actinomycete strain P294 was isolated from soil and identified as Streptomyces sp. based upon the results of 16S rRNA sequence analysis. Three compounds obtained from the solid fermentation products of this strain have been determined by 1D, 2D NMR and HRMS experiments. These compounds include two new compounds angumycinones C (1) and D (2), and the known compound X-14881 E (3). All compounds were assayed for antibacterial and nematicidal activity. The results showed the three compounds had different degrees of inhibitory activity against several target bacteria but no significant toxicity against the nematode Caenorhabditis elegans.     

一株分离于酸性矿坑水的异养菌的特性与分子鉴定

采用平板划线分离法,从江西某铜矿酸性矿坑水中分离出一株极端嗜酸、又能耐受中度碱性条件的异养微生物,命名为HJM菌株。该菌株能在pH1.5~10.0的范围内生长。形态学以及18SrDNA和26S rDNAD1/D2区序列分析表明,HJM菌株属于P.guilliermondii这个种。金属抗性试验表明,该菌株对重金属铜离子的抗性可高达45mmol/L,因此,它的分离为研究极端环境中微生物的抗铜机制提供了材料。     

Taxonomy and secondary metabolites of <Emphasis Type="Italic">Pseudobotrytis</Emphasis> sp. FKA-25

Fungal strain FKA-25, isolated from forest soil collected on Yakushima Island, Kagoshima Prefecture, Japan, was assigned to genus Pseudobotrytis based on its morphological characteristics. Conidiophores were erect, slightly swollen at the end of the tip, and gave rise to umbellate conidiogenous cells that were in an expanded denticulate portion at the end and formed ellipsoidal to clavate conidia in sympodial succession. Identification as species P. terrestris was made on the basis of the character of 1-septate conidium. Although no secondary metabolites have been reported from the genus Pseudobotrytis, four secondary metabolite compounds (designated A to D) were isolated from the culture broth of strain FKA-25. Compounds B to D have been reported previously as FK-17-p2a, lunatinin, and 3,4-dihydro-3,4,8-trihydroxy-1(2H)-naphthalenone, respectively. Compound A was designated sespendole and possessed a novel indole-sesquiterpene skeleton.     

Complete genome sequence of Pasteurella multocida HN06, a toxigenic strain of serogroup D

Pasteurella multocida is an important etiological agent that can cause many economically important diseases in a wide range of mammals and birds. Here, we report the complete genome sequence of P. multocida HN06, a toxigenic serogroup D strain isolated from a diseased pig in China.     

斑马鱼肠道中一株红酵母(Rhodotorula)的分离与鉴定

目的:分离及鉴定来自于斑马鱼肠道中的酵母菌.方法:采用马铃薯葡萄糖培养基( PDA),从斑马鱼肠道中分离到1株酵母菌,编号为ZF -2,进行形态学观察、生理特征、PCR扩增26S rDNA D1/D2区以及基因测序分析,并构建系统发育进化树.结果:ZF -2分离株为粉红色菌落、菌体呈椭圆形,芽殖;可以同化利用葡萄糖、乳糖、蔗糖等糖类;PCR扩增获得26S rDNA D1/D2区序列长度为642bp( HQ323251),序列比对分析表明与斯鲁菲亚红酵母(Rhodotorula slooffiae)相似性在99％-100％之间,构建的系统发育进化树显示菌株ZF-2与R.slooffiae( EU583485)关系最近,且位于同一分支.结论:首次从斑马鱼肠道中分离到斯鲁菲亚红酵母.     

Penicillium chrysogenum var. halophenolicum, a new halotolerant strain with potential in the remediation of aromatic compounds in high salt environments

A halotolerant phenylacetate-degrading fungus Penicillium CLONA2, previously isolated from a salt mine at Algarve (Portugal), was identified as a variant of P. chrysogenum using the ITS-5,8S rDNA and the D1/D2 domain of 28S rDNA sequences. The metabolic features and genetic characteristics suggest that this strain belongs to a subgroup of P. chrysogenum, named var. halophenolicum. The presence of the penicillin biosynthetic cluster was proven by Southern hybridizations using probes internal to the pcbAB and penDE genes and sequencing of the pcbAB-pcbC intergenic region. However the pcbAB-pcbC divergent promoter region contained 20 point modifications with respect to that of the wild type P. chrysogenum NRRL1951. The CLONA2 strain produced non-aromatic natural penicillins rather than benzylpenicillin in a medium containing potassium phenylacetate (the precursor of benzylpenicillin) and was able to grow well on phenylacetatic acid using it as sole carbon source. Due to the ability of P. chrysogenum CLONA2 to degrade aromatic compounds, this strain may be an interesting organism for aromatic compounds remediation in high salinity environments.     

Interconversion of Type C and D Strains of Clostridium botulinum by Specific Bacteriophages

These studies show that Clostridium botulinum types C and D cultures can be cured of their prophages and converted to either type C or D depending on the specific phage used. Strains of types C and D were cured of their prophages and simultaneously ceased to produce their dominant toxins designated as C(1) and D, respectively. Cured nontoxigenic cultures derived from type C strain 162 were sensitive to the phages from the toxigenic type C strain 162 and type D strain South African. When cured nontoxigenic cultures derived from strain 162 were infected with the tox(+) phages from the 162 strain of type C and the South African strain of type D, they then produced toxin neutralized by types C and D antisera, respectively. Cured nontoxigenic cultures isolated from the type D South African strain were only sensitive to the parent phage, and, when reinfected with the tox(+) phage, they produced toxin neutralized by type D antiserum. Type C strain 153 and type D strain 1873, when cured of their respective prophages, also ceased to produce toxins C(1) and D, but, unlike strain 162 and the South African strain, they continued to produce a toxin designated as C(2). When the cured cultures from strains 153 and 1873 were infected with the tox(+) phage from type D strain 1873, the cultures simultaneously produced toxin that was neutralized by type D antiserum. When these cured cultures were infected with the tox(+) phage from type C strain 153, the cultures produced toxin that was neutralized by type C antiserum. These studies with the four strains of C. botulinum confirm that the toxigenicity of types C and D strains requires the continued participation of tox(+) phages. Evidence is presented that types C and D cultures may arise from a common nontoxigenic strain.     

Anaerobic degradation of aniline and dihydroxybenzenes by newly isolated sulfate-reducing bacteria and description of Desulfobacterium anilini

A new, rod-shaped, Gram-negative, non-sporing sulfate reducer (strain Ani1) was enriched and isolated from marine sediment with aniline as sole electron donor and carbon source. The strain degraded aniline completely to CO2 and NH3 with stoichiometric reduction of sulfate to sulfide. Strain Ani1 also degraded aminobenzoates and further aromatic and aliphatic compounds. The strain grew in sulfide-reduced mineral medium supplemented only with vitamin B12 and thiamine. Cells contained cytochromes, carbon monoxide dehydrogenase, and sulfite reductase P582, but no desulfoviridin. Strain Ani1 is described as a new species of the genus Desulfobacterium D. anilini. Marine enrichments with the three dihydroxybenzene isomers led to three different strains of sulfate-reducing bacteria; each of them could grow only with the isomer used for enrichment. Two strains isolated with catechol (strain Cat2) or resorcinol (strain Re10) were studied in detail. Both strains oxidized their substrates completely to CO2, and contained cytochromes, carbon monoxide dehydrogenase, and sulfite reductase P 582. Desulfoviridin was not present. Whereas the rod-shaped catechol oxidizer (strain Cat2) was able to grow on 18 aromatic compounds and several aliphatic substrates, the coccoid resorcinol-degrading bacterium (strain Re10) utilized only resorcinol, 2,4-dihydroxybenzoate and 1,3-cyclohexanedion. These strains could not be affiliated with existing species of sulfate-reducing bacteria. A further coccoid sulfate-reducing bacterium (strain Hy5) was isolated with hydroquinone and identified as a subspecies of Desulfococcus multivorans. Most-probable-number enumerations with catechol, phenol, and resorcinol showed relatively large numbers (10(4)-10(6) per ml) of aryl compound-degrading sulfate reducers in marine sediment samples.     

Low antiplasmodial activity of alkaloids and amides from the stem bark of Zanthoxylum rubescens (Rutaceae)

The stem bark of Zanthoxylum rubescens (syn. Fagara rubescens) is used for treating fevers associated with malaria in the Ivory Coast. Three alkaloids: N-nornitidine, 7,9-dimethoxy-2,3-methylenedioxybenzophenanthridine, and bis[6-5,6-dihydrochelerythrinyl)] ether; and two amides: zanthomamide and lemairamide, were isolated from the stem bark of this plant. These compounds were screened in vitro against the chloroquine-sensitive 3D7 strain and the chloroquine-resistant FCM29 strain of P. falciparum. N-nornitidine was found to be inactive. 7,9-dimethoxy-2,3-methylenedioxybenzophenanthridine, lemairamide and zanthomamide showed weak activity with average IC50 values ranging from 45.6 microM to 149.9 microM. Bis[6-15,6-dihydrochelerythrinyl)] ether was the most active of the tested compounds with mean IC50s of 14.9 +/- 1.4 microM in FCM29 strain and 15.3 +/- 3.4 microM in 3D7 strain (approximately 58 to approximately 1130 times less active than chloroquine respectively). The anti-Plasmodium activities of the tested alkaloids of Z. rubescens were low; and do not encourage the use of this plant as antimalarial.     

Seven complementation groups of respiratory syncytial virus temperature-sensitive mutants.

Fifteen temperature-sensitive mutants of the RSN-2 strain of respiratory syncytial virus have been classified into six complementation groups, two of which appeared to be homologous with two of the three complementation groups of the A2 strain described by Wright et al. (P. F. Wright, M. A. Gharpure, D. S. Hodes, and R. M. Chanock, Arch. Gesamte Virusforsch, 41:238--247). Thus seven complementation groups of respiratory syncytial virus, designated A, B, C, D, E, F, and G, have been defined. The frequency and type of mutant isolated varied according to strain; group C was unique to the A2 strain, and groups D, E, F, and G were unique to the RSN-2 strain. The highest complementation indexes were obtained by preincubation for 7 h at permissive temperature, followed by incubation at restrictive temperature for 40 to 50 h in the case of A2 strain mutants or 80 to 90 h for RSN-2 strain mutants. Genetic recombination was not detected.     

Isolation of an effective strain of Frankia from nodules of Discaria trinervis (Rhamnaceae)

Frankia strain BCU110501 was isolated from root nodules of the native Patagonian actinorhizal plant Discaria trinervis. The strain was grown on BAP medium with sodium propionate or glucose as carbon sources. Colonies grown in nitrogen-free medium showed branched hyphae bearing polymorphic sporangia and vesicles, which were capable of nitrogen fixation. Old cultures produced a red pigment. The infectivity and effectivity of a Frankia strain isolated from Discaria on its own host, D. trinervis and also in D. chacaye, is reported for the first time. Frankia BCU110501 has physiological properties that are intermediate between categories proposed by Lechevalier et al. (1983) to classify Frankia.     

Structure of a highly phosphorylated lipopolysaccharide core in the Delta algC mutants derived from Pseudomonas aeruginosa wild-type strains PAO1 (serogroup O5) and PAC1R (serogroup O3)

Lipopolysaccharides (LPS) were isolated from rough-type mutant strains of Pseudomonas aeruginosa (Delta algC) derived from wild-type strains PAO1 (serogroup O5) and PAC1R (serogroup O3). Structural studies of the LPS core region with a special focus on the phosphorylation pattern were performed by 2D NMR spectroscopy, including a 1H,(31)P HMQC-TOCSY experiment, MALDI-TOF MS, and Fourier-transform ion cyclotron resonance ESIMS using the capillary skimmer dissociation technique. Both LPS were found to contain two residues each of 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) and L-glycero-D-manno-heptose (Hep), one residue of N-(L-alanyl)-D-galactosamine and one O-carbamoyl group (Cm) on the distal Hep residue. The following structures of a tetrasaccharide trisphosphate from strain PAC1R Delta algC and that carrying an additional ethanolamine phosphate group (PEtN) from strain PAO1 Delta algC were elucidated: [carbohydrate structre: see text] where R=P in PAC1R Delta algC and PPEtN in PAO1 Delta algC. To our knowledge, in this work the presence of ethanolamine diphosphate is unambiguously confirmed and its position established for the first time in the LPS core of a rough-type strain of P. aeruginosa. In addition, the structure of the complete LPS core of wild-type strain P. aeruginosa PAO1 was reinvestigated and the position of the phosphorylation sites was revised.     

不同来源鼠李糖乳杆菌的随机扩增多态DNA分析

[目的]建立鼠李糖乳杆菌(Lactobacillus rhamnosus,Lr)菌株之间的分子鉴别方法并分析不同分离株之间的遗传多样性.[方法]从56份采集自中国新疆和田和广西巴马瑶族自治县的长寿老人粪便样本中分离得到的乳酸菌中,经生理生化分析和API 50CHL试验条鉴定,获得10株Lr.对10株Lr分离株和1株Lr标准株ATCC7469进行了随机扩增多态DNA分析,从50条随机引物中筛选到5条在菌株水平上具有鉴别力的引物P14、OPG28、OPG25、P7和P4并建立和优化了Lr菌株RAPD指纹图谱扩增方法.根据RAPD结果计算菌株间的遗传相似系数并进行聚类分析.[结果]获得了清晰稳定的DNA指纹图谱,扩增产物大小在100～2000bp之间,菌株间呈现显著的DNA多态性,不同来源的Lr分离株的遗传相似系数在0.581～0.935之间,在相似系数0.80水平上可以将11株Lr菌株分为5个类群,其中分离自新疆和田的Lr菌株归在类群B和类群C,而分离自广西巴马瑶族自治县的Lr菌株归在类群D和类群E.[结论]应用RAPD方法对Lr菌株进行分子鉴别是可行的,不同来源的Lr之间存在着较大的种内遗传多态性和不同的亲缘关系.     

Coexistence of antibiotic-producing and antibiotic-sensitive bacteria in biofilms is mediated by resistant bacteria

Antibiotic-sensitive bacteria have been found to coexist with antibiotic-producing bacteria in biofilms, but little is known about how the former develop in such an environment. Here we isolated pyocyanin-sensitive bacteria belonging to the genus Brevibacillus from a biofilm derived from soil extract and based on the preestablished biofilm of a pyocyanin producer, Pseudomonas aeruginosa strain P1. In addition, pyocyanin-resistant strains belonging to the genus Raoultella were isolated from the same biofilm. Microbial relationships within biofilms were examined by using three strains, strain P1, Brevibacillus strain S1, and Raoultella strain R1, each of which individually formed a biofilm within 2 days in a flow cell. Strain S1 did not fully develop on the preestablished biofilm of strain P1 during 4 days of cultivation, whereas a mutant of strain P1 which was deficient in pyocyanin production allowed strain S1 to cocolonize within a biofilm. On the other hand, strain R1 developed on the biofilm of strain P1 regardless of pyocyanin production. When mixed 1:1 inocula of strains S1 and R1 were introduced into the strain P1 biofilm, all three species were found in the 4-day biofilm. In the mixed biofilm, strain S1 was surrounded by the layer of strain R1 and seemed to be separated from strain P1 and the outflow solution. However, strain S1 did not survive in a three-species mixed culture under planktonic conditions. These results indicate that the survival of sensitive bacteria in biofilm with a pyocyanin producer is achieved by covering them with a layer of resistant bacteria. We also evaluated the influence of antibiotic production on the producer.