Elevation of Francisella philomiragia subsp. noatunensisMikalsen et al. (2007) to Francisella noatunensis comb. nov. [syn. Francisella piscicida Ottem et al. (2008) syn. nov.] and characterization of Francisella noatunensis subsp. orientalis subsp. nov., two important fish pathogens

Aims: This study was conducted to clarify the taxonomic status of Francisella sp. strain Ehime‐1, a fish pathogen, in relation to the fish pathogens F. piscicida and F. philomiragia subsp. noatunensis and to F. philomiragia subsp. philomiragia. Methods and Results: Francisella sp. Ehime‐1 was compared to F. piscicida, F. philomiragia subsp. noatunensis and several F. philomiragia subsp. philomiragia isolates through sequencing of the 16S rRNA‐gene and several house‐keeping genes and determination of biochemical and phenotypic properties. Results show that F. piscicida is indistinguishable from F. philomiragia subsp. noatunensis by sequence and phenotypic traits. Francisella sp. Ehime‐1 and F. philomiragia subsp. noatunensis are clearly separated from F. philomiragia. Francisella sp. Ehime‐1 is biochemically, phenotypically and genetically different from F. philomiragia subsp. noatunensis (=F. piscicida), but DNA–DNA hybridization does not clearly support establishment as a separate species (level of relatedness 64% and 73·4%, mean 68·7%). Conclusions: We propose to elevate F. philomiragia subsp. noatunensis to species rank as F. noatunensis comb. nov., while F. piscicida is considered a heterotypic synonym of F. noatunensis comb. nov. Evidence suggests that Francisella sp. Ehime‐1 represents a novel subspecies of F. noatunensis, for which the name F. noatunensis subsp. orientalis subsp. nov. is proposed (=DSM21254^T, = LMG24544^T). Significance and Impact of the Study: This study contributes to the taxonomy and characteristics of fish‐pathogenic Francisella spp.     

Bacterial (Bacillus thuringiensis subsp. thuringiensis and B. thuringiensis subsp. kurstaki) Entomocidal Preparations Decrease Chlorophyll Accumulation in Larch Needles

Chlorophyll a plus b content and absorption spectra of the homogenates from the cotyledonary leaves of 30-day-old seedlings of two larch species, Larix gmelinii (Rupr.) Rupr. and L. sibirica Ldb. were studied. The seedlings were grown on Perlite containing aqueous solutions of entomocidal biopreparations isolated from Bacillus thuringiensis subsp. thuringiensis (bitoxybacillin) and B. thuringiensis subsp. kurstaki (lepidocide) at various final concentrations (2, 6, and 12 g/l). Changes in the form of chlorophyll absorption spectra induced by biopreparations were established. A marked inhibition of pigment accumulation in the needles dependent on the biopreparation concentration was noted. At a low concentration (2 g/l), the biopreparations virtually did not affect the chlorophyll content; an increase in their concentrations resulted in a decrease in chlorophyll content in leaves by 20% (at 6 g/l) and 40% (at 12 g/l). It is concluded that bitoxybacillin and lepidocide inhibited the chlorophyll accumulation in larch needles to a similar extent.     

Pectobacterium carotovorum subsp. brasiliense Causing Soft Rot on Paprika in Korea

During summer 2011 in South Korea, severe fruit rot of paprika was observed, causing severe economic losses in paprika production. Symptoms of fruit and pedicel decay were consistent with symptoms caused by Pectobacterium carotovorum subsp. brasiliense (Pcb) as recently described in Brazil, the United States, Israel and South Africa. Physiological analysis and pathogenicity test of strains isolated from paprika fruit revealed that the pathogen was the bacterium Pcb. Sequencing and phylogenetic analysis of the 16S rDNA and partial 16S–23S rDNA intergenic spacer region confirmed that the isolates were Pcb. This is the first report of Pcb in Korea, which has a significant economic impact on Korean paprika production.     

Characterization of Pectobacterium carotovorum subsp. carotovorum Causing Watermelon Soft Rot Disease in Iran

The aim of this study was characterized Pectobacterium carotovorum subsp. carotovorum (Pcc) the causal pathogen of watermelon soft rot disease in Iran. Of fifty bacterial isolates with white grey and convex colonies on nutrient agar obtained from symptomatic watermelon, ten isolates were selected for further tests. Pathogenicity tests results showed that all test isolates developed typical water‐soak symptoms after 2 days and signs of soft rot began 4 days after inoculation on watermelon fruits. Based on the phenotypic properties, the isolates were identified as Pectobacterium carotovorum subsp. carotovorum. The 16S rDNA sequences of isolates were 99% similar to the corresponding 16S rDNA sequence of the reference Pcc isolate. BOX and ERIC‐PCR analysis indicated that genetic diversity was present among the isolated Pcc isolates did not relate to the geographic location isolated from. To the best of our knowledge, this is the first study of biochemical and genotypic characterization of Pcc isolates the causal agents of soft rot disease on watermelon, in Iran.     

Micromorphological and chemical characterisation of Stachys recta L. subsp. serpentini (Fiori) Arrigoni in comparison to Stachys recta L. subsp. recta (Lamiaceae)

Stachys recta L. is a very polymorphous species in which numerous subspecies were recognised. S. recta L. subsp. serpentini (Fiori) Arrigoni is a typical endemism growing on serpentine soils in northern Apennines and particularly in Tuscany (Italy). In order to contribute to a better knowledge of this plant and to its differentiation with respect to S. recta L. subsp. recta, the micromorphological characters (non-glandular and glandular trichomes) and the essential oil composition of the two subspecies were investigated. Micromorphological characters were studied using scanning and transmission electron microscopy, while light microscopy was used for histochemical observations. Essential oil analysis was carried out by gas chromatography and mass spectrometry.In the two examined taxa, the morphology and distribution of glandular and non-glandular trichomes, and the different essential oil composition, may be considered distinctive characters at subspecies level. This is consistent with the taxonomic classification considering S. recta subsp. serpentini a subordinate taxon of S. recta.     

Exopolysaccharide production by Lactobacillus delbruckii subsp. bulgaricus and Streptococcus thermophilus strains under different growth conditions

Summary The optimal temperature, pH and incubation time for production of exopolysaccharide (EPS) by Lactobacillus delbruckii subsp. bulgaricus and Streptococcus thermophilus strains in MRS and M17 media, respectively, were determined. In all strains, the temperature and incubation time for EPS production were 45 °C and 18 h, respectively. At 45 °C, L. delbruckiisubsp. bulgaricus B3 and G12 and S. thermophilus W22 strains produced 263, 238 and 127 mg/l, respectively. At 18 h, B3, G12 and W22 strains produced 220, 152 and 120 mg/l, respectively. While the pH for highest EPS production by L. delbruckii subsp. bulgaricus strains was 6.2 (in B3 strain: 211 mg/l, in G12 strain: 175 mg/l), for highest EPS production byS. thermophilus strain it was 6.8 (114 mg/l).     

Cloning of the ribokinase gene of Staphylococcus hyicus subsp. hyicus in Staphylococcus carnosus

Summary A gene library with DNA of Staphylococcus hyicus subsp. hyicus was established in S. carnosus by using the plasmid vector pCT20. Two clones of S. carnosus were isolated which were able to ferment d-ribose. The two hybrid plasmids (pRib 1) and (pRib 2) were isolated and characterized. They contained inserted DNA fragments of S. hyicus subsp. hyicus with sizes of 10.2 and 8.2 kb, respectively. d-Ribose uptake and enzyme activities were studied. All strains tested [S. hyicus subsp. hyicus, S. carnosus (wild type) and the two S. carnosus clones] possessed an inducible uptake system for d-ribose. S. hyicus subsp. hyicus possessed in addition enzyme activities of d-ribokinase and d-ribose-5-P isomerase. None of these enzyme activities could be detected in S. carnosus (wildtype). Only in the S. carnosus clones containing (pRib 1) or (pRib 2) could a d-ribokinase activity be demonstrated, indicating that the gene for d-ribokinase of S. hyicus subsp. hyicus was cloned in S. carnosus.Abbreviations bp base pairs - C-TLC cellulose-thin layer chromoatography - kb kilo base pairs - pR_ib 1 and 2 ribokinase activity conferring hybridplasmids - MBq megabequerel - wt wild type     

Secondary Metabolites,Glandular Trichomes and Biological Activity of Sideritis montana L. subsp. montana from Central Italy

Sideritis montana subsp. montana is a small annual herb occurring in countries bordering the Mediterranean and Balkan regions. The secondary metabolism of this plant has not been fully explored so far. The aim of the present study was to understand the complex mixture of secondary metabolites and the type of secretory structures. The polar constituents were isolated by column chromatography from the ethanolic extract, and their structure was elucidated by NMR and MS. The essential oil was isolated by hydrodistillation and analysed by GC/MS. The plant indumentum was studied by light and scanning electron microscopy. To complete the work, the essential oil antioxidant activity and cytotoxicity on tumor cells were evaluated by DPPH, ABTS, FRAP, and MTT methods. Four different classes of secondary metabolites were isolated, namely flavonoids, caffeoylquinic derivatives, glycosidic hydroquinones and iridoids. The essential oil was mainly characterized by sesquiterpenene hydrocarbons. Peltate and long‐capitate hairs were the main sites where terpenes and polar constituents are produced. The secondary metabolites found in S. montana subsp. montana are of chemotaxonomic interest, some of them being typical of the genus Sideritis. The trichomes types observed partially differ from those described in other members of the genus Sideritis. The essential oil showed noteworthy inhibition on tumor cells.     

Interpopulation Variability in the Essential Oil Composition of Cistus creticus subsp. eriocephalus from Sardinia

Cistus genus is widespread in the Mediterranean regions with several species and is traditionally known as a natural remedy, but few previous phytochemical researches have been reported on Cistus species growing in Sardinia. The aim of this work was to study the interpopulation variability of the volatiles of C. creticus subsp. eriocephalus to find out the natural chemotypes of this taxon for chemotaxonomic purposes. Plant material was collected from seven wild populations in Sardinia. The oils from aerial part were obtained by hydrodistillation and analyzed by GC/MS enabling to identify 185 compounds. The seven samples showed appreciable variations in their chemical composition. MA was found almost exclusively linear hydrocarbons (85%) and MN presented fatty acids (36.97%) as major fraction. CP and MN had high quantity of non‐terpenic carbonylic compounds (63% and 69%, respectively) and in CP was found also sclareol, a well‐known antimicrobial compound. PCA analysis showed the existence of a high interpopulation variability within the essential oils of C. creticus subsp. eriocephalus growing in Sardinia. For example, MN and CP are very close to each other as CG is close to PM. BN is isolated from the others Cistus populations due to lacking essential oil. These data suggest that the basis of variation in the volatile composition of seven C. creticus subsp. eriocephalus populations depends on hybridization and that the sample without essential oil is the only one no‐hybridized.     

A note onArmillaria mellea subsp.nipponica subsp. nov. in Japan

Basidiomes of homothallicArmillaria mellea complex were discovered in the forests of Hokkaido. Their monosporous isolates showed partial compatibility with North AmericanA. mellea s. str. haploid testers. The morphological characteristics of basidiomes differed from other temperateA. mellea s. str. in several aspects. Here, JapaneseA. mellea s. str. is reclassified asA. mellea subsp.nipponica and its basidiome morphologies and habitats are described.     

Selection of Escherichia coli-inhibiting strains of Lactobacillus paracasei subsp. paracasei

The aim of this study was to select Escherichia coli-inhibiting strains among lactic acid bacteria. On the basis of phenotypical and technological characteristics, 20 strains of lactic acid bacteria were screened from a total of 225 isolates, obtained from nine samples of artisanal Caprino d'Aspromonte cheese, made from raw goats' milk. The antagonistic activity of these 20 strains was detected in plates against three different strains of E. coli. Two strains of Lactobacillus paracasei subsp. paracasei showed a marked anti-E. coli activity against all three strains tested; the other lactic acid bacteria did not exhibit inhibiting activity. The E. coli inhibition can be ascribed to production of bacteriocin-like compounds. The use of L. paracasei subsp. paracasei strains to increase the safety of the cheeses made from raw milk is recommended because these cultures strongly inhibit E. coli, without foreseeable adverse sensory changes. Received 27 December 2001/ Accepted in revised form 28 June 2002     

In vitro antifungal and antibacterial activities of extracts of Galium tricornutum subsp. longipedunculatum

The aim of this study was to evaluate the antimicrobial activity of crude ethanolic extracts and fractions of the ariel parts and the fruits of Galium tricornutum subsp. longipedunculatum, traditionally used in northern areas of Pakistan for treating microbial infections of skin. Extracts and their fractions were tested against six bacteria and six fungal strains using the hole diffusion method and macrodilution method. All extracts and fractions possessed significant antimicrobial effect. Four fungal strains, Candida albicans, Trichophyton longifusus, Fusarium.solani and Candida glabrata, showed interesting susceptibility profiles when evaluated using the extracts and fractions with MICs ranging from 0.18 to 200 mg/mL. In case of bacterial strains, Staphylococcus aureus, Pseudomonas aeruginosa and Salmonella typhi were significantly susceptible to the extracts and fractions with MICs ranging from 0.12 to 200 mg/mL. Comparative results were carried out using imepenem, miconazole and amphotericin B as standard antibiotics.     

Bicarbonate is a stimulus in the inter-species induced sporulation of strict anaerobic Syntrophomonas erecta subsp. sporosyntropha

Previously Syntrophomonas species had been described as the bacteria those did not form spores, however, in our previous studies, a newly isolated S. erecta subsp. sporosyntropha JCM13344^T was found to form spores in the co-culture with methanogens, while not in mono-culture or in co-culture with sulfate reducer. In this study, we examined the sporulation stimulus conferred by methanogens in the co-culture. By reducing bicarbonate in mono-culture and sulfate-reducing co-culture, we could induce S. erecta subsp. sporosyntropha JCM13344^T to form spores, so that bicarbonate at lower concentration was determined as another stimulus for sporulation. Based on the substrate degradation by strain JCM13344^T in different concentration of bicarbonate vs at different pHs, it was suggested that bicarbonate could stimulate the sporulation by mediating a nutrient deprivation but not pH drop. To further confirm the sporulation potential of this group of bacteria, spo0A fragments were amplified from strain JCM13344^T as well as all the recognized Syntrophomonas species, confirming that they were members of the spore-forming group. Since sporulation is a kind of response of spore-forming bacteria to environmental stresses, the observation in this work implies that stresses can be created even between the mutual beneficial partners, in this case, inducing sporulation.     

Syntrophomonas wolfei subsp. methylbutyratica subsp. nov., and assignment of Syntrophomonas wolfei subsp. saponavida to Syntrophomonas saponavida sp. nov. comb. nov

A spore-forming bacterium strain 4J5(T) was isolated from rice field mud. When co-cultured with Methanobacterium formicicum DSM 1535(T), strain 4J5(T) could syntrophically degrade saturated fatty acids with 4-8 carbon atoms, including 2-methylbutyrate. Phylogenetic analysis based on 16S rRNA gene similarity showed that strain 4J5(T) was most closely related to Syntrophomonas wolfei subsp. wolfei DSM 2245(T) (98.9% sequence similarity); however, it differed from the latter in the substrates utilized and its genetic characteristics. Therefore, a new subspecies Syntrophomonas wolfei subsp. methylbutyratica is proposed. The type strain is 4J5(T) (=CGMCC 1.5051(T)=JCM 14075(T)). Furthermore, based on 16S rRNA sequence divergence and substrate utilization, we propose the assignment of Syntrophomonas wolfei subsp. saponavida DSM 4212(T) to Syntrophomonas saponavida sp. nov. comb. nov.     

Comparative analysis of C-band karyotypes inCrepis praemorsa subsp.praemorsa and subsp.dinarica

Crepis incarnata subsp.dinarica (G. Beck)Hayek is nowadays considered as a subspecies ofC. praemorsa (L.)Tausch. Comparative analyses of Feulgen karyotypes demonstrate great similarities, but remarkable differences in the presence and the distribution of the constitutive heterochromatin in the two taxa are detected by using the Giemsa differential staining technique (C-banding). This favours their specific distinctness.     

两种不同生境中国沙棘种子内生细菌的多样性

植物内生菌广泛分布于植物的各种组织及器官中,对植物的生长表现出各种作用,而植物种子中的内生菌对植物的作用也越来越受到人们的关注。以榆中县和秦安县两种不同生境的中国沙棘种子为材料,分析中国沙棘种子内生细菌多样性,以期探究生境对种子内生菌多样性的影响,并为进一步研究种子内生菌与沙棘的相互作用提供依据。研究利用纯培养方法和高通量测序方法分别进行中国沙棘种子内生细菌多样性分析。对纯培养分离得到的内生细菌,利用16S rRNA基因序列分析法结合形态学特征进行内生细菌的鉴定;对高通量测序得到的数据进行基于OTUs(Operational Taxonomic units,可操作分类单元)的物种注释分析。通过纯培养方法从榆中县中国沙棘种子中分离得到4株内生细菌,分属于芽孢杆菌属(Bacillus)、葡萄球菌属(Staphylococcus)和假单胞菌属(Pseudomonas);秦安县中国沙棘种子中分离得到5株内生细菌,分属于芽孢杆菌属(Bacillus)、葡萄球菌属(Staphylococcus)和不动杆菌属(Acinetobacter)。采用高通量测序方法检测到榆中县中国沙棘种子内生细菌分属于7个门、68个属,秦安县中国沙棘种子内生细菌分属于5个门、30个属。在门分类水平,榆中县中国沙棘种子内生细菌的主要优势门类是蓝藻门(Cyanobacteria)和变形菌门(Proteobacteria),相对丰度分别为95.62%和2.03%;秦安县中国沙棘种子的主要优势门类是变形菌门(Proteobacteria)和蓝藻门(Cyanobacteria),相对丰度分别为91.68%和8.06%。在属分类水平,榆中县中国沙棘种子内生细菌的优势菌属为蓝藻细菌属(Cyanobacteria),相对丰度为95.09%;秦安县中国沙棘种子内生细菌的优势菌属为寡养单胞菌属(Stenotrophomonas),相对丰度为85.60%。榆中县和秦安县两种不同生境中国沙棘种子内生细菌有着丰富的多样性,且内生细菌的多样性和丰富度存在明显差异,表现为榆中县中国沙棘种子内生细菌的多样性和丰富度均高于秦安县中国沙棘种子内生细菌。     

The allopolyploid origin ofSedum rupestre subsp.rupestre (Crassulaceae)

InSedum rupestre L. a polyploid series (x = 16) occurs in which aneuploid chromosome numbers and odd levels of ploidy prevail. The most common and widely distributed cytotype,S. rupestre subsp.rupestre, is 2n = 112. Plants resemblingS. rupestre subsp.rupestre can be obtained by hybridizing the tetraploid cytotypes ofS. forsterianum Sm. (2n = 48) andS. rupestre subsp.erectum 't Hart (2n = 64). Comparison of these artificial hybrids with their parents and a large number of plants ofS. rupestre subsp.rupestre (2n = 112) from nature showed thatS. rupestre subsp.rupestre and the artificial hybrids are morphologically indistinguishable, and intermediate betweenS. forsterianum andS. rupestre subsp.erectum. MorphologicallyS. rupestre subsp.rupestre is closer to subsp.erectum than toS. forsterianum. Chloroplast DNA restriction patterns ofS. rupestre subsp.rupestre, however, resembleS. forsterianum more closely. The combined results of the hybridization experiments, the analysis of the cpDNA restriction patterns, and the morphological variation indicate the allopolyploid origin ofS. rupestre subsp.rupestre. Natural hybrids inSedum (Crassulaceae) 4.     

Bacillus subtilis subsp. spizizenii MB29 controls alfalfa root rot caused by Fusarium semitectum

Medicago sativa L. is the most important forage legume in China. Reducing production losses caused by disease is an essential aspect of maximising alfalfa production. In the current study a Fusarium semitectum isolate collected from alfalfa roots exhibiting symptoms of root rot was proven to infect alfalfa by fulfilling Koch's postulates. A bacterial strain, MB29, also collected from alfalfa roots, was evaluated as a potential biocontrol agent against F. semitectum and a range of other alfalfa pathogens using in vitro tests. It was found that MB29 reduced the mycelia growth of all the pathogens assessed, and in the case of F. semitectum by as much as 84.47%. Furthermore, in vivo test showed that MB29 reduced the severity of rot symptoms in alfalfa seedlings resulting from F. semitectum infection. Strain MB29 was subsequently classified as Bacillus subtilis subsp. spizizenii using the Biolog MicroLog microbial identification system and sequence analysis of its 16S rDNA gene. Taken together these results indicate that B. subtilis subsp. spizizenii MB29 has great potential for the control of root rot diseases in alfalfa.     

Characterisation of Pseudomonas chlororaphis subsp. aurantiaca strain Pa40 with the ability to control wheat sharp eyespot disease

This study details the isolation and characterisation of Pseudomonas chlororaphis subsp. aurantiaca strain Pa40, and is the first to examine P. chlororaphis for use in suppression of wheat sharp eyespot on wheat. Pa40 was isolated during an investigation aimed to identify biocontrol agents for Rhizoctonia cerealis. Over 500 bacterial strains were isolated from the rhizosphere of infected wheat and screened for in vitro antibiosis towards R. cerealis and ability to provide biocontrol in planta. Twenty‐six isolates showed highly antagonistic activity towards R. cerealis, in which Pseudomonas spp. and Bacillus spp. were predominant members of the antagonistic community. Strain Pa40 exhibited clear and consistent suppression of wheat sharp eyespot disease in a greenhouse study and suppression was comparable to that of chemical treatment with validamycin A. Pa40 was identified as P. chlororaphis subsp. aurantiaca by the Biolog identification system combined with 16S rDNA, atpD, carA and recA sequence analysis and biochemical and physiological characteristics. To determine broad‐spectrum applicability and the specific mechanisms involved in Pa40's pathogen suppression this strain was tested for antibiosis towards various phytopathogens and assayed for many biocontrol activities and plant‐beneficial traits. Strain Pa40 inhibited the growth of 10 of 13 phytopathogenic fungal strains and six of eight phytopathogenic bacteria tested. This original work characterises HCN, protease and siderophore production in P. chlororaphis. Each of these characteristics likely contributed to Pa40's biocontrol capabilities as well as stimulation of the hypersensitive response in tobacco and the presence of genes involved in the biosynthesis of phenazine, 2‐hydroxylated phenazine and pyrrolnitrin.