Filamentous fungi in wrapped forages determined with different sampling and culturing methods

The use of wrapped forage bales with high dry‐matter (DM) content implies risk of fungal growth inside the wrapping, and impaired feed quality. Since fungi may be unevenly distributed in bales, the method of sampling can influence the outcome of the analysis. Three common sampling methods for detection of fungi in wrapped forage were compared: direct plating of visible mouldy patches on bale surfaces (Method I), direct plating of pieces of foliage from drilled core samples (Method II) and quantitative analysis dilution series of drilled core samples (Method III). All samples were cultured on two media at two temperatures. Samples were collected from 124 farms in Sweden and Norway. Using Methods I, II and III, fungi were detected on 52%, 77% and 56% of the farms respectively. Fifty‐two fungal species were identified using a combination of culturing and molecular methods. The most predominant species were Arthrinium spp., found on 55% of the farms, followed by Penicillium roqueforti at 48% of the farms. Incubation at 25°C resulted in higher numbers of all genera, except Aspergillus, compared to incubation at 37°C. The different methods of sampling and culturing did not identify the same fungal species within the same bale. Analysing one bale per farm resulted in detection of less than half the number of species compared to analysing three bales. Of the sampling and culturing methods compared, direct plating of drilled core samples cultivated at malt extract agar at 25°C performed best in terms of qualitative analysis of fungi species.     

植被恢复对黄土高原矿区重构土壤理化性质、酶活性以及真菌群落的影响

[目的]研究黄土高原矿区重构土壤植被恢复对土壤理化性质、酶活性以及真菌群落的影响,准确评价植被恢复对该地区的土壤生态恢复效果。[方法]在内蒙古自治区准格尔旗选择3种典型植被恢复类型(灌木、乔木、草地)和未复垦地采集0—20 cm土壤样品,采用扩增子测序技术分析了土壤真菌多样性和群落组成,同时结合土壤理化性质探讨了不同植被恢复类型影响真菌群落多样性的重要因素。[结果]①与未复垦地相比,3种典型植被恢复类型不仅对土壤理化性质和酶活性均有明显提升,还提高了土壤真菌可操作分类单元(OUTs)和多样性指数,且差异显著,以灌木的效果最好。②真菌主要菌门相对丰度在各样地变化趋势为子囊菌门(Ascomycota)担子菌门(Basidiomycota)球囊菌门(Glomeromycota),相对丰度之和已超过50%,土壤真菌群落结构在4个样地之间保持了一定的稳定性。③土壤含水量为影响真菌群落结构的最主要因素。[结论]植被恢复影响土壤理化性质、酶活性及真菌群落多样性,黄土高原矿区生态恢复是一个漫长的过程,在生态恢复过程中,宜种植灌木来改善重构土壤的生态环境。     

黔中地区马尾松杜鹃群落杜鹃根围真菌多样性

选取黔中地区的贵阳市乌当区百宜乡(WD)、花溪区孟关(MG)和贵州省黔南龙里县龙架山森林公园(LL)3个地区的马尾松-杜鹃群落为研究对象,采集杜鹃根围土壤进行高通量测序,分析物种多样性,并通过数量生态学原理分析植物群落物种α多样性及土壤养分对真菌群落的影响.结果表明:从杜鹃的根围土壤中获得有效序列505 444条,隶属于9门23纲84目169科296属真菌(3 435 OTU),其中LL有真菌9门19纲53目90科127属(495 OTU)、 WD有8门21纲78目150科246属(1 076 OTU)、 MG有6门17纲55目99科129属(522 OTU).菌群α多样性分析显示, Chao1,ACE,Shannon指数均表现为WD大于MG和LL.菌群群落结构受生境植物多样性、土壤养分的综合作用,土壤有机质、 pH值以及速效磷、速效氮、全氮含量较大程度解释了真菌对土壤养分变化的响应.     

Microbial composition and diversity of an upland red soil under long-term fertilization treatments as revealed by culture-dependent and culture-independent approaches

Background, aim, and scope  Fertilization is an important agricultural practice for increasing crop yields. In order to maintain the soil sustainability, it is important to monitor the effects of fertilizer applications on the shifts of soil microorganisms, which control the cycling of many nutrients in the soil. Here, culture-dependent and culture-independent approaches were used to analyze the soil bacterial and fungal quantities and community structure under seven fertilization treatments, including Control, Manure, Return (harvested peanut straw was returned to the plot), and chemical fertilizers of NPK, NP, NK, and PK. The objective of this study was to examine the effects on soil microbial composition and diversity of long-term organic and chemical fertilizer regimes in a Chinese upland red soil. Materials and methods  Soil samples were collected from a long-term experiment station at Yingtan (28°15′N, 116°55′E), Jiangxi Province of China. The soil samples (0–20 cm) from four individual plots per treatment were collected. The total numbers of culturable bacteria and fungi were determined as colony forming units (CFUs) and selected colonies were identified on agar plates by dilution plate methods. Moreover, soil DNAs were extracted and bacterial 16S rRNA genes and fungal 18S rRNA genes were polymerase chain reaction amplified, and then analyzed by denaturing gradient gel electrophoresis (DGGE), cloning, and sequencing. Results  The organic fertilizers, especially manure, induced the least culturable bacterial CFUs, but the highest bacterial diversity ascertained by DGGE banding patterns. Chemical fertilizers, on the other hand, had less effect on the bacterial composition and diversity, with the NK treatment having the lowest CFUs. For the fungal community, the manure treatment had the largest CFUs but much fewer DGGE bands, also with the NK treatment having the lowest CFUs. The conventional identification of representative bacterial and fungal genera showed that long-term fertilization treatments resulted in differences in soil microbial composition and diversity. In particular, 42.4% of the identified bacterial isolates were classified into members of Arthrobacter. For fungi, Aspergillus, Penicillium, and Mucor were the most prevalent three genera, which accounted for 46.6% of the total identified fungi. The long-term fertilization treatments resulted in different bacterial and fungal compositions ascertained by the culture-dependent and also the culture-independent approaches. Discussion  It was evident that more representative fungal genera appeared in organic treatments than other treatments, indicating that culturable fungi were more sensitive to organic than to chemical fertilizers. A very notable finding was that fungal CFUs appeared maximal in organic manure treatments. This was quite different from the bacterial CFUs in the manure, indicating that bacteria and fungi responded differently to the fertilization. Similar to bacteria, the minimum fungal CFUs were also observed in the NK treatment. This result provided evidence that phosphorus could be a key factor for microorganisms in the soil. Thus, despite the fact that culture-dependent techniques are not ideal for studies of the composition of natural microbial communities when used alone, they provide one of the more useful means of understanding the growth habit, development, and potential function of microorganisms from soil habitats. A combination of culture-dependent and culture-independent approaches is likely to reveal more complete information regarding the composition of soil microbial communities. Conclusions  Long-term fertilization had great effects on the soil bacterial and fungal communities. Organic fertilizer applications induced the least culturable bacterial CFUs but the highest bacterial diversity, while chemical fertilizer applications had less impact on soil bacterial community. The largest fungal CFUs were obtained, but much lower diversity was detected in the manure treatment. The lowest bacterial and also fungal CFUs were observed in the NK treatment. The long-term fertilization treatments resulted in different bacterial and fungal compositions ascertained by the culture-dependent and also the culture-independent approaches. Phosphorus fertilizer could be considered as a key factor to control the microbial CFUs and diversity in this Chinese upland red soil. Recommendations and perspectives  Soil fungi seem to be a more sensitive indicator of soil fertility than soil bacteria. Since the major limitation of molecular methods in soil microbial studies is the lack of discrimination between the living and dead, or active and dormant microorganisms, both culture-dependent and culture-independent methods should be used to appropriately characterize soil microbial diversity.     

Can placement of seed away from relic stubble limit Rhizoctonia root rot in direct-seeded wheat?

Rhizoctonia root rot of wheat can be a problem in no-till systems, especially during the transition from conventional tillage. There are no effective chemical controls or resistant varieties, leaving only cultural methods to manage this disease. In a no-till system, residue and inoculum of soilborne pathogens are not moved by cultivation, therefore the inoculum may be concentrated in the seeding row of the previous year. Using GPS tracking systems with sub-meter accuracy, the seeding row could be placed away from the row of the previous year. We tested the hypothesis that seeding away from the relic row may reduce Rhizoctonia root rot. In two field experiments, plants were sampled at three distances from the seed row, as well as from fumigated plots. Intact soil cores were also removed from the field, planted with seeds at various distances from the previous row, and grown in the greenhouse under controlled conditions. Pasteurized cores served as controls. Disease levels were higher in the field in the second year, but there was no consistent effect of seed row placement on disease or plant parameters. However, soil fumigation and pasteurization had significant effects, indicating that soilborne pathogens were active. Inoculum of Rhizoctonia is not produced in the crowns and lower stems of the plant, but the pathogen survives in living and dead roots of the previous year crop, volunteers, and grassy weeds. Thus, high inoculum densities may be present in between the relic rows, as well as within the rows. If this is the situation with Rhizoctonia, precision placement of seed rows would not be efficacious.     

真菌诱导子对虎杖愈伤组织中白藜芦醇合成的影响

用源于黑曲霉(Aspergillum niger)、啤酒酵母(Saccharomyces cerevisiae)、青霉(Penicillium sp.)及灰色葡萄孢(Botrytis cinerea)的真菌诱导子对虎杖愈伤组织进行诱导处理,研究其对愈伤组织中白藜芦醇合成的影响,采用HPLC测定白藜芦醇含量。结果表明,4种真菌诱导子中,以黑曲霉诱导子的诱导效果最好,在愈伤组织继代培养后第14天加入100mg/L的黑曲霉诱导子,引入后第6天愈伤组织中白藜芦醇含量比对照高2.25倍。黑曲霉诱导子对虎杖愈伤组织中白藜芦醇的积累有很好的促进作用。     

Australian Uromyces viciae-fabae: Host and nonhost interaction among cultivated grain legumes

Uromyces viciae-fabae, rust of faba bean, parasitizes other legume crops such as lentils (Lens culinaris) and field peas (Pisum sativum) in some environments. In this study we examined the host range of two Australian isolates of U. viciae-fabae collected and purified from a faba bean crop and classified as U. viciae-fabae ex V. faba. Field pea (P. sativum), chickpea (Cicer arientinum), lupin (Lupinus spp.), lentil (L. culinaris), and mung bean (Vigna radiata) genotypes were tested with these isolates, as well as resistant and susceptible genotypes of the faba bean host. Race specificity for these two pathogen isolates was observed on Vicia faba, with two faba bean genotypes showing partial resistance. Both U. viciae-fabae isolates also colonized field pea seedlings and successfully produced uredinia under glasshouse conditions, despite this fungus not being known as a pathogen of Australian field pea crops. No sporulation of either isolate of U. viciae-fabae ex V. faba was observed on any of the remaining legume species tested. However, obvious differences in fungal growth were observed, ranging from small infection sites with very rare haustorium formation in mung bean to more extensive growth and the development of potential uredinial structures in chickpea. These observations are discussed in relation to the phylogenetic relationship of these host and nonhost species.     

Identification of resistance to Aspergillus flavus infection in cotton germplasm

Natural resistance in cottonseed to Aspergillus flavus infection has not been explored to date. A green fluorescent protein (GFP) expressing A. flavus strain was used to assess the resistance of seed from 35 cotton varieties from Gossypium arboreum, G. barbadense, and G. hirsutum. Mature cotyledons devoid of seed coat were wounded, inoculated, and assessed for innate resistance to A. flavus infection. Of the initial 35 varieties tested, we observed a range of resistance to infection in representatives of all three species. A subset of 15 representatives was further analyzed. Within this group, G. arboreum cultivar A2 186 and G. hirsutum cultivar SA 1582 were most resistant to fungal infection. The most susceptible cultivar in this group was G. hirsutum SA 1595. The remaining 12 representatives tested in the secondary screen (3 G. arboreum, 3 G. barbadense, and 6 G. hirsutum lines) exhibited intermediate resistance. We did not observe any relationship between species and resistance. Within each species, there was a range of responses to fungal infection. Future studies using this methodology to screen additional diploid and tetraploid cotton lines may enable us to identify naturally resistant germplasm that can be used to develop cotton with enhanced resistance to fungal infection.