The ectomycorrhizal status of European larch (Larix decidua Mill.) seedlings from bare-root forest nurseries

In this paper, we report molecular investigation of an ectomycorrhizal fungi (EMF) community of European larch (Larix decidua Mill.) seedlings grown in a bare-root nursery. In total, we surveyed 32 nurseries that were each active in supplying planting stocks to restock forests and for afforestation of post-agricultural land. Sequence-based approach was used to identify EMF taxa, quantify EMF richness, and document differences in the relative abundance of individual taxa. We identified seven fungal species that might contribute to the mycorrhizal community structure of 1 to 3-year-old L. decidua seedlings. The species richness in the examined larches varied between one and four fungal taxa, depending on both the nursery stock samples (NSS) and age class of the seedlings. The average was 1.4 for 1-year-old seedlings and 2.3 for 2- and 3-year-old plants. The dominance of Ascomycota over Basidiomycota and the prevalence of two species, Wilcoxina mikolae and Suillus grevillei, as EMF partners were characteristic features of nursery-grown L. decidua seedlings. S. grevillei was the only one basidiomycetes colonized roots of tested seedlings. The rest of the mycorrhizal pool from forest nurseries was typically dominated by pioneer fungal ascomycetes. W. mikolae was the most common mycorrhizal ascomycete present at a high frequency on NSS from both age classes and with very high abundance (average 90%) on 1-year-old seedlings. Some other ascomycetes (Pezizales 1, Pezizales 2 and Pezizales 3) appeared on tested larches at a low frequency, but sometimes in high abundance. Tuber spp. appeared at a low frequency and low abundance. The relative abundance of S. grevillei was positively correlated with the age of seedlings, while W. mikolae was negatively correlated with age. Tuber sp. 1 and 2, Pezizales 2, and W. mikolae were positively associated with the basic soil pH values. However, forward selection of the environmental variables showed that only the age of the larch seedlings contributed significantly (F = 11.45, P = 0.02) to the variance in the ECF community.     

Paralytic shellfish poisoning toxin analysis of the genus <Emphasis Type="Italic">Alexandrium</Emphasis> (Dinophyceae) occurring in Korean coastal waters

ABSTRACT:   Paralytic shellfish poisoning (PSP) toxins produced by Alexandrium isolates from Korea were analyzed by high-pressure liquid chromography. Species designation of the regional isolates was determined by morphological criteria and ribotyping inferred from sequences of the 28S rDNA D1-D2 region. Toxin analysis performed at the exponential growth phase, revealed that the two strains of A. fraterculus were non-toxic, while the strains of A. tamarense and A. catenella were toxic. Toxic isolates DPC7 and DPC8 of A. catenella produced GTX1, 2, 3, 4, 5, dcGTX2, 3, C1, 2, neoSTX and STX with trace or non-detectable levels of C3 and C4, while isolates UL7, KDW981, SJW97043, SJW97046, KJC97111 and KJC97112 of A. tamarense produced GTX1, 2, 3, 4, dcGTX3, C1, 2, neoSTX with trace or non-detectable levels of C3, 4, dcSTX and STX, and no GTX5 and dcGTX2. The major toxins produced by A. catenella were C1 +2, and those of A. tamarense were C1 +2 and GTX4 in most of the isolates. A. tamarense strains other than SJW97046 produced a relatively high proportion of carbamate toxins, reflecting the high toxicity scores of shellfish intoxication in sampled coastal areas. Two representative toxic isolates, A. tamarense SJW97043 and A. catenella DPC7, were cultured for 30 days in batch mode and subjected to toxin analysis at 5-day intervals. Comparison of toxin productivity in terms of total toxin content, toxin components, and their variations with culture age revealed marked differences between the two strains.     

Identification of Arcanobacterium pluranimalium by matrix-assisted laser desorption ionization-time of flight mass spectrometry and,as novel target,by sequencing pluranimaliumlysin encoding gene pla

In the present study 13 Arcanobacterium pluranimalium strains isolated from various animal origin could successfully be identified phenotypically by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and genotypically by sequencing 16S rDNA and the pluranimaliumlysin encoding gene pla. The detection of mass spectra by MALDI-TOF MS and the novel genotypic approach using gene pla might help to identify A. pluranimalium in future and might elucidate the role this species plays in infections of animals.     

Mycobacterium salmoniphilum infection in farmed Atlantic salmon, Salmo salar L

Multiple greyish‐white visceral nodules containing abundant rapidly growing and acid‐fast bacteria, subsequently identified as Mycobacterium salmoniphilum, were detected in moribund and newly dead market‐sized fish during a period of increased mortality in an Atlantic salmon, Salmo salar, farm in western Norway. Isolates cultured from diseased fish were phenotypically consistent with Mycobacterium sp. previously isolated from Atlantic salmon [MT 1890 (= NCIMB13533), MT1892, MT1900 and MT1901] in the Shetland Isles, Scotland. Partial sequences of 16S rDNA, ribosomal RNA internal transcribed spacer (ITS1), 65‐kDa heat‐shock protein (Hsp65) and β subunit of RNA polymerase (rpoB) revealed 97‐99% similarity with M. salmoniphilum type strain ATCC 13758^T. The source of infection was not confirmed. Koch’s postulates were fulfilled following experimental challenge of Atlantic salmon with field isolate NVI6598 ( FJ616988 ). Mortality was recorded in experimentally infected fish; however, the infection remained subclinical in the majority of affected fish over the 131‐day challenge period.     

Members of soil bacterial communities sensitive to tillage and crop rotation

Pyrosequencing was used to study the effect of rotation and tillage on total bacterial communities. We designed primers to the bacterial 16s rDNA and amplified DNA from soil samples from a long-term tillage/rotation trial in Kansas for two seasons. The 2 × 2 factorial trial had two rotation treatments (wheat-wheat and wheat-soybean) and two tillage treatments (conventional and no-till). A total of 20,180 16s rDNA sequences were generated and 2337 operational taxonomic units (OTUs) were assembled using a 97% similarity cut-off. The phylum Proteobacteria represented 38% of 299 identified taxa. The second most abundant phylum was Acidobacteria, making up 20% of the sequences, the majority of which were Acidobacteria Group 1. The phyla Actinobacteria and Gemmatimonadetes comprised 12% and 3.5% of the sequences. Other groups detected included TM7, Nitrospira, Verrucomicrobia, and Bacteroidetes. Some clusters of Acidobacteria Group 1 were more frequent in continuous wheat versus wheat-soybean rotation, some Acidobacteria Group 2 were more frequent in no-till, and some Acidobacteria Group 4 were more frequent in wheat-soybean rotation. These results were validated by quantitative real-time PCR. Pyrosequencing provided taxonomic information about the overall bacterial community, and detected community shifts resulting from different cropping practices.     

上海临港滨海河道夏季周丛生物群落演替特征

于2021年6月26日-7月10日对上海浦东地区芦潮引河3段河流进行连续挂板采样,研究不同污染程度河流中周丛生物的分布特征及时间变化。共采集12次,获得36个样品,对18S rDNA V4可变区进行高通量测序。结果发现,主要类群为轮虫、绿藻、纤毛虫。不同污染程度河流周丛生物群落组成呈现显著性差异(p<0.01),污染最严重的Y点样品优势属为Pentatrocha、Lagenidium、Dinovorax、Symbiodinium等,其特有属有Calycofera、Crucigeniella等;污染较为严重的H点样品的优势属为Ptygura、Oedogonium、Philodina、Physocypria等,特有属有Eccrinidus、Hyphochytrium等;无污染的G点样品优势属为Cyclotella、Prunus、Vorticella、Chlamydomonas等,特有属有Actinochloris、Adelina、Albugo等。生物群落与环境因子的趋势对应显示,TDS、盐度、pH和污染程度为影响周丛生物群落变化的显著性因素。不同生物对于环境因子的响应程度不同,优势属对于环境因子的选择性趋势更加明显。主要周丛生物类群（纤毛虫、轮虫、绿藻）在不同污染程度河流中群落丰度变化不同,调查周期内,在受到污染的G点和Y点周丛生物群落出现了绿藻代替纤毛虫的群落演替趋势。本研究为受污染水体周丛生物生态功能评估及环境评估提供基础数据,也为周丛生物（膜）作为一种环保绿色、造价低廉的污染物去除方法提供理论基础。     

Molecular phylogenetical studies of the thermophilic spore-forming desulfotomaculum isolated from oil-field

A novel thermophilic and heterotrophic sulfate-reducing bacteria, strain CW-03, was isolated from crude oil well whose depth was 3.2 kilometer. The bacterium was strictly anaerobic; it does not endure acid and its maximum surviving temperature was 70℃. Many short chain organic compounds can be utilized as electron donors, which were acetate, formate, lactate, propionate, pyruvate, butyrate, succinate, malate, fumarate, valerate, caproate, heptanoate, octanoate, decanoate, tridecanoate, pentadecanoate, palmitate, heptadecanoate or ethanol, while sulfate and sulfite were used as electron acceptors. The following substrates were not utilized： benzoate undecanoate, dodecanoate, tetradecane, propanol, butanol, H2＋CO2 （80/20%; v/v） and acetate （1mM） ＋ H2. When lactate was used as electron donors, sulfite and thiosulfate, but not sulfer and nitrate, can be used as electron acceptors. Strain CW-03 was motile, curved rod, Gram-positive, pole flagellum and spore-forming. On the basis of 16S rRNA sequence alignment （accession numbers： AY703032）, CW-03 should be included in the genus Desulfotomaculum with BLAST analysis on line. However, some of its physiology and multiple sequence alignments were different from other members of this genus. Therefore, CW-03 should be recognized as a new species, for which we propose the name Desulfotomaculum chinamiddle （Bacteria, Firmicutes, Clostridia, Clostridiales, Peptococcaceae）.     

草莓褐色轮斑病病原鉴定及室内药效研究*

草莓褐色轮斑病近年越来越严重,尤其在草莓育苗阶段。从草莓发病叶片、匍匐茎上分离得到病原菌,对病原菌进行形态特征观察、生物学特性研究、ITS序列分析以及室内药效试验。结果表明：该病原菌为Sphaeronaemella fragariae。该菌菌丝生长最适温度范围是25～28 ℃;适宜pH为6;在供试的几种碳、氮源中,最适的碳源是蔗糖,最适的氮源是酵母浸出液。在供试的9种药剂中,以咪鲜胺1 000倍液对病菌的抑制作用最好。     

Molecular Monitoring of SRB Community Structure and Dynamics in Batch Experiments to Examine the Applicability of in situ Precipitation of Heavy Metals for Groundwater Remediation (15 pp)

Background, Aims and Scope   Sulfate-reducing bacteria (SRB) are known for their capacity to reduce and precipitate heavy metals (HM) as metal sulfides, offering the opportunity to create an in situ reactive zone for the treatment of heavy metal-contaminated groundwater, a process called in situ metal precipitation (ISMP). The applicability of the ISMP technology first has to be investigated at a laboratory scale before going into an on site application. The evaluation and optimization of the ISMP process is facilitated when physical/chemical analysis techniques are combined with molecular tools that specifically monitor the abundance, diversity and dynamics of the indigenous sulfate reducing microbial community. In this study, batch experiments were conducted in order to investigate the feasibility of ISMP as a groundwater remediation strategy for an industrial site contaminated with elevated levels of Zn, Cd, Co and Ni. Methods   The potential of different types of carbon source/ electron donor (lactate, acetate, methanol, ethanol, Hydrogen Release Compound?, molasses) to stimulate the sulfate reduction and metal precipitation activity of the naturally present (or indigenous) SRB community was explored. In addition, the effect of amending vitamin B12 and yeast extract was evaluated. The ISMP process was monitored by combining analytical analyzes of process parameters (SO42&#61485;-concentration, heavy metal concentrations, pH, Eh) with molecular tools such as SRB subgroup and genus specific PCR, denaturing gradient gel electrophoresis (DGGE), and phylogenetic analysis of clone sequences, based on either the 16S rRNA or the dsr (dissimilatory sulfite reductase) gene. Results and Discussion   The efficiency of different carbon-sources to stimulate the ISMP process followed the order HRC&#61650; 〉 molasses 〉 methanol 〉 lactate 〉 ethanol 〉 acetate. Within 10 weeks, the highest sulfate and metal removal efficiencies ranged from 85% to 99%. Addition of yeast extract boosted the ISMP process, whereas vitamin B12 negligibly affected SRB activity. Analysis of the sulfate reducing population by SRB subgroup and genus specific PCR demonstrated that members of the genus Desulfosporosinus dominated in all batch tests, while 16S rDNA DGGE profiles additionally revealed the presence in the microbial communities of non-sulfate reducing bacteria within the family Clostridium and the &#61541;-proteobacteria. The dsrB-based DGGE profiles allowed us to assess the diversity and dynamics of the sulfate reducing community and added to a better understanding of the effects of different batch conditions on the ISMP process. Remarkably, all dsrB sequences affiliated with the dsrB gene sequence cluster found in Desulfotomaculum, which received their xenologous dsrB gene from the &#61540;-proteobacteria. Conclusions   The batch experiments, which aimed at stimulating the activities of the indigenous SRB communities, demonstrated that these communities were present and that their activities could be used to obtain efficient in situ precipitation of the contaminating heavy metals. This opens the possibility to test this concept in the future as an on site demonstration as part of the groundwater strategy for the heavy metal contaminated site. Although batch setups are suitable for preliminary feasibility studies for ISMP, they do not reflect the in situ situation where sulfate and heavy metal and metalloid polluted groundwater are supplied continuously. A sulfate reducing strain JG32A was isolated from whose 16S rRNA gene affiliated with the genus Desulfosporosinus, while its dsrB gene sequence clustered with Desulfotomaculum dsrB gene sequences, which received their xenologous dsr genes from &#61540;-proteobacteria. Therefore we hypothesize that the batch experiments enrich members of the Desulfosporosinus genus that possess a non-orthologous dsrB gene. Recommendation and Perspective   The next step towards an on site pilot test for ISMP will be the setup of a series of column experiments, with process conditions that are selected based on the above mentioned results. This will allow to define optimal ISMP process conditions and to test its long-term efficacy and sustainability before going into an on site bioremediation application. By applying the described molecular tools together with physical-chemical analyzes, it can be investigated whether the same SRB community is enriched and which type of C-source is most effective in promoting and sustaining its growth and sulfate-reduction activity.