1. Introduction
Tilletia contraversa Kühn (commonly abbreviated TCK) belongs to Fungi, Basidiomycotina, Teliomycetes, Ustilaginales, Tilletiaceae, Tilletia. TCK is a plant quarantine fungus that is greatly harmful and causes 20% to 75% reduction in wheat production. In the epidemic outbreak years, it can eventually result in no harvest at all [1]. TCK grows up with the wheat together and it invades wheat inflorescence at booting stage. Finally, TCK forms mycocecidium which replaces the whole kernel. The shape of mycocecidium is similar to a ball and the surface of mycocecidium is highly tough. Mycocecidium is full of black teleutospore which owns a stronger survival ability. Teleutospore is primary infection resources and it will germinate at least three weeks if the temperature (3˚C - 8˚C), sunlight and humidity are suitable. The viability of TCK fungus galls scattered in the field lasts up to 10 years, and the germination rate of TCK teliospores gone through animal’s digestive tract can be as high as 64% [2]. If the TCK-infected straws, wheat bran, and other wastes remain active, once the infectious source has formed, or the fungus has accumulated in the fields to a relatively high density in an area under years of nonstop crop rotation, TCK diseases will become more and more severe. Moreover, in the annual harvest season, the thick smoke-generating straw burning not only greatly harms the environment, but also wastes the straw resources. The inspection and quarantine management department in China requires that the storage and treatment of waste materials generated during the grain process should be centralized, to prevent the potential spreading of pests. Therefore, treating TCK-infected straws, wheat bran, and other wastes, in a biosecure way to transform them into usable products, becomes the best approach to both block the epidemics and control the hazard. This study has applied microbial fermentation-based composting techniques to inactivate TCK in wheat wastes via fermentation and composting maturation with mixed crop straws and manures.
2. Materials and Methods
2.1. The Tested Fungal Strain and Culture Medium
The TCK fungus galls collected in 2009 were provided by the Xinjiang Entry-Exit Inspection and Quarantine Bureau of China. The water agar (WA) medium contained 30 g of agar and 1 L of distilled water.
2.2. The Instruments
Autoclave, superclean bench, low temperature incubator, microscope, temperature sensor, pH meter, elemental analyzer, freeze-drying machine, and compost bin (dimension: 280 cm × 120 cm × 100 cm; thickness: 10 cm) were used in this study.
2.3. The Establishment of Composting System
The cow dungs were collected freshly from a cattle farm in Dalian city of China and made homogeneous by thorough mix. The C/N ratio of the compost was adjusted to 19:1 with straws, and its water content was adjusted to 63.7% by drying. The composting materials were piled into the bottomless compost bin to start the static composting. The compost bin was loosely covered with lid to not only provide a shelter but also allow the gas exchange between the pile and the environment. A pad of 20 cm thick paddy straw was placed on the bottom of compost bin to absorb the exudate. Under sterile condition, the intact TCK gall was placed into 1.5 mL sterilized tube, which was then buried 40 - 50 cm deep inside the compost. Multiple thermocouples were also placed close to the sample tube, and the compost temperature was measured daily. After 10 days’ composting fermentation, compost was required to turn over to make sure the mixture of various ingredients in composting system.
2.4. The Detection of Physicochemical Parameters
The TCK gall samples were randomly collected from multiple sites in the compost after 0, 2, 4, 6, 10, 14, 18, 26, 32, and 40 days of composting, respectively, to detect the pH value, moisture, and C/N ratio. Briefly, 2 g of compost sample were weighed, put into a 50 mL test tube, added with 18 mL of distilled water, and placed on a Vortex mixer to shake for 5 min and stay static for another 30 min. The pH value of the supernatant was then measured using a pH meter. The moisture was determined by freeze-drying method. The freeze-dried compost sample was ground evenly and used to detect the C/N ratio.
2.5. The Germination Rate of Composting-Treated TCK
The germination rate of treated TCK were detected after 2, 4, 6, 10, 14, 18, 26, 32, and 40 days of composting, respectively, and compared with that in control group. Briefly, the teliospores removed from the fungal gall samples were suspended in appropriate amount of sterile water and spread evenly onto WA plate with approximately 50 teliospores per field. The plate was then placed at 4˚C and cultured under continuous light condition. The observation was started 3 weeks later. The germination rate was calculated 6, 8, and 12 weeks, respectively, after the cultivation. The average germination rate was calculated from triplicated tests. The length of promycelium greater than or equal to the diameter of TCK teliospore was recognized as germinating.
3. Results
3.1. The Changes in Compost Temperature
The temperature is an important factor affecting the microbial activities and the composting processes. High temperature contributes to the rapid inactivation of the pathogens, and an appropriate temperature range is conducive to humus formation. As shown in Figure 1, 2 - 8 days after the composting, the temperature was maintained between 50˚C - 60˚C, without influence from the ambient temperature. In the 5th day of composting, the temperature reached the maximum approximately 55˚C. In the 10th day, the temperature dropped to approximately 30˚C, and the compost was mixed again. The temperature of mixed compost decreased initially, then began to rise, and finally dropped to room temperature after 14 days of maintaining between 40˚C - 50˚C.
3.2. The Changes of pH in the Compost
Figure 2 shows that in the beginning, the pH of the composting materials increases rapidly. After reaching 8.9 at the 2nd day, the pH began to decline slowly until the 10th day. The pH values were between 8.5 and 8.9. After the 10th day’s mixing, until the 40th day, the compost pH maintained in the range of 8.5 to 9.1. Microorganisms survive within certain pH range; a stable, suitable pH facilitates the microbial fermentation and accelerates the composting process.