rDNA Sequencing and Preliminary Analysis of Microorganisms from Diseased Edible Fungus Using Non-culture Technology

The metagenomic DNA of disease tissue samples from four kinds of major edible fungus was extracted by CTAB method combined with DNA gel recovery kit. The genomie DNA was amplified by polymerase chain reaction using the universal primers of 16S rDNA and 18S rDNA, and then mone, elonal sequenced after ligated and transformed, rDNA sequences of 20 positive clones were selected randomly from each pair of primers for sequence alignment. The results showed that there were two bacterial diseases and two fungul diseases in the samples, respectively.

Mycosphaerella areola—The Teleomorph of Ramularia areola of Cotton in Brazil, and Its Epidemiological Significance

While Ramularia leaf blight of cotton caused by Ramularia areola is of top most importance for Brazil, information is lacking regarding the survival mechanism of this pathogen during the cotton-free period. The teleomorph of R. areola is expected to belong to the genus Mycosphaerella. In the present study attempts were made to verify occurrence of this teleomorph in the State of Mato Grosso, Brazil. Decaying cotton leaves were collected two months after harvest of 2014 from 44 commercial and experimental fields where aerial fungicidal applications were made or not during the crop cycle to control the Ramularia leaf blight. Examination of the decaying cotton leaves revealed presence of abundant sclerotia, spermagonia and ascoma of Mycosphaerella sp. intermingled with each other during the cotton-free period in most of the leaf samples. Mono-ascospore isolations were obtained from the ascoma and considering their cultural, morphological, pathological and DNA sequence analysis they were identified as Mycosphaerella areola. M. areola and R. areola isolates produced similar symptoms under glasshouse inoculations. Reisolation of the pathogen from the symptoms produced by M. areola isolates yielded R. areola. Some cotton leaves showing such symptoms were kept on the soil surface on plastic trays for two months under natural field condition. After this period the decaying leaves showed abundant perithecia identical to their original M. areola. ITS rDNA sequence analyses revealed identical sequences from M. areola and R. areola isolates. Occurrence and the viability of the perfect stage M. areola during the cotton-free period on the left-over stubble from one season to another were interpreted as the survival mechanism of the pathogen and were considered responsible for the Ramularia blight epidemics in the State of Mato Grosso. Disease management practices for the State of Mato Grosso are discussed. This is the first report about the occurrence of the M. areola in Brazil.

Bioleaching of spent Ni-Cd batteries and phylogenetic analysis of an acidophilic strain in acidified sludge

Biohydrometallurgy is a novel method to recycle discarded batteries,in which sewage sludge is used as microorganisms and culture due to the presence of indigenous Thiobacilli.A two-step continuous flow leaching system consisting of an acidifying reactor and a leaching reactor was introduced to achieve the bioleaching of spent nickel-cadmium(Ni-Cd)batteries.The acid supernatant produced in the acidifying reactor by the microorganisms with ferrous ions as the substrate was conducted into the leaching reactor to dissolve electrode materials.The efficiency of a batch treatment of batteries was examined.The results showed that the complete dissolution of two AA-sized Ni-Cd batteries with 0.6 L/d acid supernatant took about 30,20,and 35 days for Ni,Cd,and Co,respectively.But the dissolution ability of the three metals was different.Cd and Co can be leached mostly for pH below 4.0 while the complete dissolution of Ni can be achieved for pH below 2.5.Meanwhile,a strain(named Thiooxidans.WL)accounting for the reduction of pH in the acidified sludge was isolated and sequenced.It was identified to be 100%similar to Acidithiobacillus ferrooxidans strain Tf-49 based on 16S rDNA sequence analysis.The relevant phylogenetic tree constructed indicates that the strain should be classified into genus Acidithiobacillus ferrooxidans.

Screening and Denitrification Characteristics of a Slight Halophilic Heterotrophic Nitrobacteria

A slight halophilic heterotrophic nitrobacteria named gs1 was separated from the matured activated sludge. According to the morphological observation,physiological biochemical tests and sequence analysis of the 16S rDNA,strain gs1 was identified to be as Pseudomonas sp. Sodium acetate and ammonium chloride were used as carbon and nitrogen sources,respectively,to investi-gate the characteristics of the bacterium. When cultured for 24 h under aerobic conditions,with the removal rates of the NH4+-N and COD being 82.2% and 74.73%,respectively,strain gs1 will have a nitrification function of producing NO2--N. When cultured for 24 h under aerobic conditions in nitrite medium,the removal rate of the NO2--N became 100%,and when cultured for 24 h under aerobic conditions in nitrate medium,the removal rate of the NO3--N became 97%. The result shows that this strain functions for either nitrification or denitrification,i.e.,it can complete the full process of biological deoxidation.

The diversity of soil culturable fungi in the three alpine shrub grasslands of Eastern Qilian Mountains

To understand the diversity of culturable fungi in soil at alpine sites, Rhododendron fruticosa shrubland, Salix cupularis fruticosa shrubland, and Dasiphoru fruticosa shrubland of the Eastern Qilian Mountains were selected to investigate. Three methods, including tradi- tional culturing, rDNA internal transcribed spacer （ITS） sequence analysis, and economical efficiency analysis, were carried out to estimate the diversity of soil culturable fungi of these three alpine shmblands. A total of 35 strains of culturable fungi were cultured by dilution plate technique and were analyzed by rDNA ITS sequence. The diversity indices such as species abundance （S）, Shannon-Wiener index （H）, Simpson dominance index （D）, and Pielou evenness index （3） of Rhododendron fruticosa shrubland, Salix cupularis fruticosa shrubland, and Dasiphoru fruticosa shrubland were ranged between 16 and 17, 2.66-2.71, 0.92, 0.95~）.97 respectively. The results showed that the diversity of soil fungi were abundant in these three types of alpine shrub grasslands, while further study should be done to explore their potential value.