Effect of nitrate and ammonium concentration on nitrate reductase activity in five species of mycorrhizal fungi

The nitrate reducing capacity of pure cultures of Cenococcum geophilum (Sow.) Ferd. & Winge, Paxillus involutes (Batsch: Fr.) Fr. (strains 1 and 2), Piloderma croceum Erikss. & Hjortst., Suillus variegatus (Fr.) O. Kuntze (strains 1 and 2) and an ectendomycorrhizal (E-strain) fungus was measured using an in vivo nitrate reductase (EC 1.6.6.3) assay. Differences between species and strains were established. The nitrate concentration of the culture medium influenced the nitrate reductase activities of the E-strain fungus and one strain of S. variegatus. The nitrate reductase activity of certain species and strains was a function of nitrate concentration. Addition of ammonium to the growth medium did not have any significant effect on the in vivo or in vitro nitrate reductase activity. The in vivo nitrate reductase activity in the mycelia of C. geophilum and the E-strain fungus decreased during 28 day growth in modified Melin-Norkrans medium. For mycelia of Paxillus involutus, Piloderma croceum and S. variegatus grown on agar the in vitro assays showed higher nitrate reductase activity than the in vivo assays.     

Bacterial contaminants in liquid packaging boards: assessment of potential for food spoilage

Liquid packaging boards and blanks were examined for microbial contaminants. A total of 218 strains were identified and representatives of the most frequent species were characterized for their potential for food spoilage. Contaminants found were aerobic spore-forming bacteria, mostly Bacillus megaterium, B. licheniformis, B. cereus group, B. pumilus, Paenibacillus macerans, P. polymyxa, P. pabuli and B. flexus. Production of amylolytic, proteolytic, lipolytic and phospholipolytic enzymes was common. Approximately 50% of the B. cereus group strains were positive in the diarrhoeal enterotoxin immunoassay test or in the enterotoxin reversed passive latex agglutination test. Strains capable of growth at 6°C were found among B. cereus group, P. pabuli, P. validus, B. megaterium and P. polymyxa. All B. licheniformis strains grew at 55°C. The spores of B. licheniformis were most resistant to hydrogen peroxide. The B. cereus group strains were recognizable by fatty acid components not present in any of the other paperboard strains, 11-methyldodecanoic acid (13:0 iso) and trans-9-hexadecenoic acid (16:1 ω 7 trans), each contributing 7% or more to the total cellular fatty acids.     

Mycorrhiza formation is not needed for early growth induction and growth-related changes in polyamines in Scots pine seedlings in vitro

Ectomycorrhizal (ECM) fungi have been shown to improve growth of the host plant before the formation of physical ECM structures, i.e. during the so-called pre-mycorrhizal phase. In the present study, changes in growth and the concentrations of individual polyamines (PAs) were followed during the mycorrhiza formation in Scots pine (Pinus sylvestris) seedlings in the presence of two ECM fungi, Pisolithus tinctorius and Paxillus involutus. The two fungus stains were chosen because they differed in infection characteristics as well as in PA and auxin production. The results were compared to our earlier study with two Suillus variegatus strains forming ECMs with Scots pine seedlings in vitro. Paxillus was not able to form ECMs whereas Pisolithus formed ECM association with Scots pine seedlings within two weeks. However, Paxillus enhanced the growth of the seedlings more than Pisolithus. Paxillus also increased putrescine (Put) concentrations of the seedlings in the pre-mycorrhizal phase much more than Pisolithus. A similar trend was observed in the free spermidine (Spd) in stems, whereas in the needles Paxillus decreased free Spd concentration. Pisolithus caused a threefold greater increase in root free Spd than Paxillus. Effects of Paxillus on the growth and PA fluctuation, excluding root free Spd, of the host plant resembled that observed in our previous in vitro study on S. variegatus–Scots pine interaction. Therefore, changes in specific PA concentrations in the pre-mycorrhizal phase seem to be related to growth induction by the ECM fungus rather than to mycorrhiza formation. Moreover, we suggest that growth induction in host plants is not necessarily followed by ECM formation.     

Suillus variegatus causes significant changes in the content of individual polyamines and flavonoids in Scots pine seedlings during mycorrhiza formation in vitro

Changes in the concentrations of individual flavonoids and polyamines (PAs) in Scots pine (Pinus sylvestris L.) cotyledonary seedlings were studied during the establishment of an ectomycorrhizal (ECM) symbiosis with two Suillus variegatus strains in vitro. Both flavonoids and PAs were analysed after 3, 7, and 14 d in dual culture, and changes in concentrations were compared with growth of the seedlings. Both S. variegatus strains caused similar responses in Scots pine seedlings. Free putrescine accumulated immediately but only transiently after inoculation. This was followed by continuous accumulation of PA conjugates in needles and stems, and free spermidine and spermine in roots, which was accompanied by mycorrhiza formation and improved growth. The fungi induced lateral root formation and main root and primary needle elongation. Inoculation caused no qualitative changes in flavonoid composition, while quantitative changes in flavonols, catechins, and condensed tannins were observed in shoots during mycorrhiza formation. These results indicate that in this in vitro system conjugated PAs and specific flavonoids, generally related to the plant's defence reactions, did not play a major role in the regulation of the establishment of the ectomycorrhizal (ECM) symbiosis in Scots pine roots. The results also clearly show that positive growth responses in shoots and roots due to S. variegatus were supported by different and highly specific changes in the synthesis of both primary and secondary metabolites in these parts of the seedling.     

Coronary flow reserve and heart failure in experimental coxsackievirus myocarditis. A transthoracic Doppler echocardiography study

The objective of this study was to apply transthoracic Doppler echocardiography (TTDE) in mice to study coronary flow reserve (CFR), an index of coronary microvascular function, in mild and severe forms of experimental viral myocarditis. Regarding methodology, BALB/c mice were infected with cardiotropic coxsackieviruses causing either a mild (Nancy strain) or a severe (Woodruff strain) myocarditis. Left ventricular dimensions, fractional shortening, and CFR (ratio of left coronary artery flow velocity during maximal adenosine-induced vasodilatation to rest) were measured by TTDE before infection and again 1 or 2 wk after infection. As a result, the resting flow velocity did not change after infection. In contrast, CFR reduced significantly 1 wk after infection with either virus variant [from 2.5 (SD 0.3) to 1.4 (SD 0.1) in severe and from 2.4 (SD 0.4) to 2.1 (SD 0.3) in mild myocarditis], being significantly lower in the severe than mild myocarditis. CFR remained low in severe myocarditis 2 wk after infection. Fractional shortening decreased to the same levels 1 wk after infection with either virus variant [from 0.54 (SD 0.02) to 0.43 (SD 0.03) in severe and from 0.51 (SD 0.03) to 0.44 (SD 0.02) in mild myocarditis, P < 0.05]. However, 2 wk after infection, mice with severe myocarditis had enlarged left ventricles and lower fractional shortening [0.31 (SD 0.03)] than mice with mild myocarditis [0.47 (SD 0.02), P < 0.01]. In conclusion, CFR measured with TTDE is reduced in coxsackievirus myocarditis in mice. Low CFR is associated with progressive heart failure, indicating that dysfunction of coronary microcirculation is a determinant of poor outcome in viral myocarditis.     

Potassium nutrition and free polyamines of Betula pendula Roth and Betula pubescens Ehrh.

Potassium and free polyamine concentrations in the leaves of silver birch (Betula pendula Roth) and downy birch (Betula pubescens Ehrh) were followed during three successive growing seasons 1996, 1997 and 1998 in order to define K deficiency levels.The highest foliar K concentrations were found in June. In August, the K concentrations were lower and remained quite stable in Betula pendula but varied in Betula pubescens. In addition to a common diamine, putrescine, and the polyamines, spermidine and spermine, also a less common diamine and polyamine, 1,3-diaminopropane and norspermidine were found in the birch leaves. The accumulation of both diamines, putrescine and 1,3-diaminopropane, was used to define the critical levels of K nutrition in birch leaves. Foliar K concentrations below 7–8 mg g ^–1 DW were found to correlate with the accumulation of diamines at most sampling dates.     

Ectomycorrhization of Tricholoma matsutake and two major conifers in Finland—an assessment of in vitro mycorrhiza formation

This study aimed to test the ability of Tricholoma matsutake isolates to form mycorrhizas with aseptic seedlings of Pinus sylvestris L. and Picea abies (L.) Karst. Germinated seedlings of Scots pine and Norway spruce were separately inoculated with either isolates originating from Finland or Japan. Eight months after inoculation, the Finnish isolate had formed a sheath and Hartig net on both host species. Ectomycorrhizal Scots pine seedlings inoculated with the Finnish isolate showed the same shoot height and dry mass as the controls. Ectomycorrhizal Norway spruce seedlings inoculated with the Finnish isolate had similar shoot height but slightly less dry mass than the control seedlings. For both tree species, inoculation with the Finnish isolate resulted in reduced total nitrogen content per seedling, but carbon content was unaffected. Inoculation with the Japanese isolate resulted in an initial Hartig net-like structure in pine but not in spruce. No typical Hartig net was observed on either tree species. Furthermore, seedlings of both species inoculated with the Japanese isolate showed significantly reduced growth, dry mass, nitrogen, and carbon content per seedling and shoot height (in spruce) compared to the controls. This study documents and describes the in vitro ectomycorrhization between T. matsutake and Scots pine or Norway spruce and the variable mycorrhizal structures that matsutake isolates can form.     

The Evolution of Vp1 Gene in Enterovirus C Species Sub-Group That Contains Types CVA-21, CVA-24, EV-C95, EV-C96 and EV-C99

Genus Enterovirus (Family Picornaviridae,) consists of twelve species divided into genetically diverse types by their capsid protein VP1 coding sequences. Each enterovirus type can further be divided into intra-typic sub-clusters (genotypes). The aim of this study was to elucidate what leads to the emergence of novel enterovirus clades (types and genotypes). An evolutionary analysis was conducted for a sub-group of Enterovirus C species that contains types Coxsackievirus A21 (CVA-21), CVA-24, Enterovirus C95 (EV-C95), EV-C96 and EV-C99. VP1 gene datasets were collected and analysed to infer the phylogeny, rate of evolution, nucleotide and amino acid substitution patterns and signs of selection. In VP1 coding gene, high intra-typic sequence diversities and robust grouping into distinct genotypes within each type were detected. Within each type the majority of nucleotide substitutions were synonymous and the non-synonymous substitutions tended to cluster in distinct highly polymorphic sites. Signs of positive selection were detected in some of these highly polymorphic sites, while strong negative selection was indicated in most of the codons. Despite robust clustering to intra-typic genotypes, only few genotype-specific ‘signature’ amino acids were detected. In contrast, when different enterovirus types were compared, there was a clear tendency towards fixation of type-specific ‘signature’ amino acids. The results suggest that permanent fixation of type-specific amino acids is a hallmark associated with evolution of different enterovirus types, whereas neutral evolution and/or (frequency-dependent) positive selection in few highly polymorphic amino acid sites are the dominant forms of evolution when strains within an enterovirus type are compared.