MINISTRY OF EDUCATION AND TRAINING CAN THO UNIVERSITY SUMMARY OF DOCTORAL DISSERTATION Major in Soil Science Identification code: 9620103 NGUYEN THI PHUONG COMPOSTING MICROBIAL ORGANI
Trang 1MINISTRY OF EDUCATION AND TRAINING
CAN THO UNIVERSITY
SUMMARY OF DOCTORAL DISSERTATION
Major in Soil Science Identification code: 9620103
NGUYEN THI PHUONG
COMPOSTING MICROBIAL ORGANIC FERTILIZER OF SLUDGES FROM
WASTEWATER TREATMENT PLANTS OF BEER AND SEAFOOD PROCESSING
FACTORIES
Can Tho, 2019
Trang 2This study was achieved at Can Tho University
Scientific Supervisor: Assoc Prof Dr Nguyen My Hoa
This dissertation was defended at the University
The dissertation is available at:
Learning Resource Center, Can Tho University
National Library of Viet Nam
Trang 3PUBLICATIONS RELATED WITH THE
DISSERTATION
[1] Nguyen Thi Phuong, Nguyen My Hoa, Do Thi Xuan,
Lam Ngoc Tuyet, Vo Thi Thu Tran, 2016 “Characteristics of sludges from wastewater treatment plants of beer and seafood processing factories” Journal of Science, Can Tho University, 45A/2016, p.74-81
(In Vietnamase)
[2] Nguyen Thi Phuong, Nguyen My Hoa, Do Thi Xuan,
Lam Ngoc Tuyet, 2017.“Composting of sludge from wastewater treatment plants of seafood processing factories” Science and
Technology Journal of Agriculture & Rural Development, 5, p.54-61 (In Vietnamase)
[3] Nguyen Thi Phuong, Nguyen My Hoa, Do Thi Xuan,
Lam Ngoc Tuyet, 2017 “Composting of sludge from wastewater treatment plants of beer processing factories” Journal of Vietnam soil
science, 50, p.47-52 (In Vietnamase)
[4] Nguyen Thi Phuong, Nguyen My Hoa, Do Thi Xuan,
2018 " Effect of micro - organic composts from beer and seafood sludge on growth and yield of Okra (Abelmoschus esculentus)" Journal
of Agricultural Science and Technology of Vietnam, 2(87), p.7-10 (In Vietnamase)
[5] Nguyen Thi Phuong, Nguyen My Hoa, Do Thi Xuan,
2018 “Production and assessment efficiency of microbial-organic fertilizers from beer and seafood factories’ sludge on vegetable yield”
Journal of Science, Can Tho University, special issue Agriculture 54 p.81-90 (In Vietnamase)
[6] Nguyen Thi Phuong, Nguyen My Hoa, Do Thi Xuan,
2018 “Effect of microbial organic fertilizers from seafood sludge on growth and yield of winter melon (Benincasa hispida)” Journal of
Vietnam soil science, 54, p.19-24 (In Vietnamase)
Trang 4CHAPTER 1 INTRODUCTION 1.1 Rationale of the study
In many wastewater treatment facilities, the bottleneck of the sludge handling system is the dewatering operation Beer (BS) and seafood sludges (SS) were effluents dewatered in waste water treatment plants of beer and seafood factoires The annual sludge production from beer breweries and seafood factories is about 6
million tons and 313,170 tons, respectively (Fillaudeau et al (2006);
Ministry of Industry and Trade, 2009, 2016; Vo Phu Duc, 2013 )
According to Senthilraja et al (2013), Feng et al (2008), and Vo Phu
Duc (2013) beer and seafood sludges are nontoxic and contained much of the organic matter and valuable nutrients For these reasons, recycling of sludges for agricultural purposes seems to be an appealing solution that enables using of valuable components and resolve pollutants ( Ministry of Natural Resources and Environment,
2013; Oanh and Dieu,2016; Saviozzi et al., 1994; Stocks et al., 2002;
Thomas and Rahman, 2006)
Both BS and SS possessed poor compressibility and settleability
(Jin et al., 2003; Li et al., 2012) Therefore, it is necessary to mix BS
and SS with organic materials such as sugarcane sludge, bagasse and straw that are high in cellulose and porosity to provide energy for microorganisms during composting process, and increase compost quality However, the most suitable ratio mixture of sludge and organic materials to produce high quality microbial-organic fertilized to meet the standard of Vietnam and to increase crop yield is still questionableAt present, there are not many research papers on beer and seafood sludges for production of microbial-organic compost (Vo Phu Duc, 2013; Oanh and Dieu, 2016) In addition, isolating fungal strains from rice straw, bagasse and coconut fiber was also carried out to find out new fungi that can decompose organic materials, resisted to pathogenic fungi and promote organic composting process
Because of these issues, study on production of microbial-organic fertilizers from beer and seafood factories’ sludge is necessary as an effective treating of sludge
1.2 Objectives of study
Determination of optimal composting formula for the production
of microorganic compost from beer and seafood sludges;
Trang 5Assessment effetiveness of microbial-organic fertilizers from beer and seafood factories’ sludge on vegetable yield;
Isolation and selection of fungal strains capable of decomposing organic materials for further use as a microbe in production microbial organic fertilizer
1.3 Research activity
Determination of physical, chemical, nutritional and biological characteristics of BS and SS
Efficiency of dried beer and seafood sludge on growth and yield
of vegetable grown under the greenhouse condition;
Determination of decomposability and suitable composting ratio of beer/seafood sludge and organic materials at bag scale;
Production microbial-organic fertilizers from beer and seafood sludge at scale of 0,5 meter cubic;
Assessing efficiency of microbial-organic fertilizers from beer and seafood factories’ sludge on vegetable yield, consisting of mustard, okra, cucumber, and winter melon in the field conditions;
Isolation and selection of organic material decomposable-fungal strains
1.4 Scope of the study
The study focused on sludges from wastewater treatment of beer production and seafood processing plants in the Mekong delta to
produce microbial-organic fertilizer using Trichoderma spp from
Trico-DHCT product as the beneficial microorganism The crops used for testing the effect of microbial-organic fertilizer were vegetables for short growth duration The isolation of the fungal strain focused on organic material decomposability of the fugus for the aim of the wide use in production of microbial-organic fertilizer
1.5 Scientific significance and applicability of study
The results showed that BS and SS are two sources rich in nitrogen, phosphorus, micro nutrients and do not contain heavy metal contaminants
Sugarcane cake was suggested to mix with beer/seafood sludges for the production of bio-organic fertilizer The optimal mixing ratio of sludge and sugarcane sludge with 20:80 is recommended for the production of microbial-organic fertilizers with high quality
The research has proved the efficiency of microbial organic fertilizer produced from both sludges on vegetable yields such as
Trang 6mustard, okra, cucumber, and winter It is recommended to use 5 tons /ha of microbial organic fertilizer from the sludges with recommended dose of inorganic fertilizer to increase vegetable yield
Four strains of fungi which possessed organic material decomposability and fungal pathogens antagonistic character were
selected Two strains were identified as Neurospora crassa and Neurospora intermedia, which decomposed well mixture of sludge and
sugarcane cake with ratio of 20:80
Table 2.1 Analytical methods of input materials
Parameter Unit Analytical methods
Moisture % Dry at 105oC to constant weight Bulk density g/cm3 Samples are taken in a 100 cm
3
box and then dried at 1050C for 24 hours
pHH2O - Extract with water with a soil ratio of
1: 5 Measure by pH meter
EC mS/cm Extract with water with a soil ratio of
1: 5 Measure by EC meter Organic cabon %C The samples were dried at 105
Total potassium %KO Anaerobic digestion with salicylic acid +
Trang 7concentrated H2SO4 + H2O2 and measured on atomic absorption
N available %N Extract with H2SO4 0.5N
Dw Colony forming unit method
2.2 Efficiency of sun-dried beer and seafood sludges on growth and yeild of vegetable under the greenhouse condition
2.2.1 Germination of mustard (Brassica juncea) on substrates of
sun-dried sludges
Each sludge were put in a plastic tray with a thickness of 4 cm Then each tray would directy exposure the Sun from 9 am up to 3 pm, and rotate the sample every 30 minutes The sludge wastes were treated by directly exposing under the sunshine at 4 hours, 3 hours and
2 hours to get three levels of moistures i.e 10%, 30% and 50%, respectively to treat human pathogens
The experiment was performed in trays with a completely randomized block with eight treatments (Table 2.2) and three replications for each The volume of soil, sun-dried sludges, organic fertilizer of sugarcane cake (SF) used for each tray is 0.5 kg/tray 100 green mustard were sowing into the trays and sprinkle with 60% humidity to ensure germination in all trays After 14 days, germinated percentage, height, fresh weight, dry weight of mustard were recorded
Trang 8Table 2.2: Substrates of sun-dried sludges on mustard germination
2.2.2 Evaluation efficiency of sun-dried sludges on growth and yield of green mustard (Brassica juncea)
As a result of experiment at section 2.2.1, two samples of BS-30
and SS-50 were used as compost on growth and yield of green mustard The experiment was completely randomized in a pot with six treatments and three replications (Table 2.3) Each pot contains 7kg dry land The BS-30 and SS-50 mixed with sugarcane cake (SC), organic fertilizer of sugarcane cake used for each pot is 5 tons/ha and distributed a week before sowing Green mustard seed is sown 10 seeds into pots and watered to maintain moisture Each pot would choose well three plants when the plant highthened about
5 cm Fertilizers are used according to the recommended rate of Tran Thi Ba (1999) with 55N-32P2O5-46K2O (kg/ha).
Table 2.3: Experiment on growth and yield of green mustard
Order Treatment Mixture Ratio
and Salmonella The analyzing method was described in section 2.1
Treatment Types of fertilizer
1 Control (Soil)
2 Beer sludge at 10% humidity (BS-10)
3 Beer sludge at 30% humidity (BS-30)
4 Beer sludge at 50% humidity (BS-50)
5 Seafood sludge at 10% humidity (SS-10)
6 Seafood sludge at 30% humidity (SS-30)
7 Seafood sludge at 50% humidity (SS-50)
8 Organic fertilizer of sugarcane cake (SF)
Trang 92.3 Determination of decomposition ability, suitable composting ratio of sludges in the bag
2.3.1 Evaluation decomposable capacity of organic materials
Beer sludge (BS) was collected from Tien Giang beer factory and seafood sludge (SS) was collected from Hau Giang seafood company Both sugarcan cake (SC) and bagasse were sampled from Vi Thanh sugarcane factory Experimental equipment and chemicals used department of soil science, Can Tho University, including of
Trichoderma-DHCT fungus.The experiment was arranged in a
completely randomized block of 13 treatments with 3 replications for each treatment (Table 2.4)
For incubation, 2 g a mixture of sludge and organic materials at moisture of 65%, and placed in 150-mL plastic jars One vial containing NaOH solution (3N) were placed into 150-mL plastic jars After this, the vial containing the NaOH solution was renewed weekly Moisture was checked by weighing at each sampling date The amount
of Trichoderma fungus was injected into each treatment with 100g /m3
(Dw) at initial compost After this, all of the jars were placed in the dark at a constant temperature of 300C for 45 days Carbon mineralisation was measured at 7, 14, 21, 30 and 45 day after incubation The CO2 was titrated with H2SO4 (0.01 N) Mass loss was determined by weighing the sample after 45 days of incubation
Table 2.4: Treatments, mixture ratio and C/N ratio
Order Treatments Ratio (%Dry weight) C/N
Trang 102.3.2 Composting beer and seafood sludges in bag condition
The samples of BS, SS, rice straw, sugarcane cake, bagassem and
cow dung were collected and disposed as similarly as sections 2.1.1 and 2.3.1 Incubation equipments consisted of plastic bags of 50kg,
weighing scale of 30kg, canvas cover plastic bag, thermometer, and
Trichoderma-DHCT fungus The experiment was arranged in a
completely randomized block of 14 treatments with 3 replications for each treatment (Table 2.5) The treatments were mixed by dry weight The amount of Trichoderma fungus was injected into each treatment with 100g /m3 (Dw) at initial and after 15-day incubation with 50 for
each injection The basis on the results of section 2.3.1 and initial C/N
ratio, combination sludge with sugarcane cake and straw or bagasse with ratio of 10:60:30; 10:70:20, 20:60:20, and 20:80 was studied
Table 2.5 treatment and mixture ratio
Order Treatments Ratio (%Dw) C/N
1 BS: SC:bagasse 10:60:30 30
2 BS: SC:bagasse 10:70:20 23
3 SS: SC:bagasse 10:60:30 30
4 SS: SC:bagasse 10:70:20 23
5 Cow dung: SC:bagasse 10:60:30 21
6 Cow dung: SC:bagasse 10:70:20 20
7 BS: SC: rice straw 20:60:20 24
8 BS: SC: rice straw 10:70:20 21
9 SS: SC: rice straw 20:60:20 23
10 SS: SC: rice straw 10:70:20 20
11 Cow dung: SC: rice straw 20:60:20 23
12 Cow dung: SC: rice straw 10:70:20 21
Trang 11Trichoderma and pathogens such as E coli, Coliforms, Salmonella after 49-days incubation The analyzing method was described in
section 2.1 Weight loss also observed at final composting
2.4 Production microbial-organic fertilizers from beer and seafood factories’ sludge in 0.5 meter cubic
Based on the results of section 2.3, the optimal mixtue ratio of
sludge and sugarcane cake at 20:80 was chosen to product organic fertilizers in 0.5 meter cubic
microbial-The samples of BS, SS, straw, sugarcane cake (SC), bagasse and
cow dung were collected and disposed as similarly as sections 2.1.1 and 2.3.1 Composts were a mixture of BS or SS and SC by dry weight
and 3 replication for each The composting experiments were conducted at the experimental Farm of the Agriculture and Applied Biology, Can Tho university Six square piles were prepared on an experimental scale (1m high and 1m basal diameter) in a shed, to prevent excessive drainage and nutrient leaching The piles were manually turned at 7, 14, and 30 days after establishment to promote aeration and ensure that all material was exposed to high temperatures The amount of Trichoderma fungus was injected into each treatment with 200 g/0.5m3 (Dw) at initial (with 100 g), 28 and after 15-day incubation with 50g for each injection
At days 0, 21, 28, 49, and 63 from initiation, one composite sample (~1 kg dry matter) of three subsamples was collected from each pile These samples were used for analyses of temperature, humidity, pH, and EC Total C, N, phosphorus (P), potassium (K), available N, P, K, calcium (Ca), magnesium (Mg), micro nutrients (Zn,Cu, Mn), heavy
metal contaminants (total Cd, Pb, As, Hg) and amount of Trichoderma
and pathogens were analysied after composting days of 0, 49 and 63
The analyzing method was described in section 2.1 Weight loss also
observed at final composting
2.5 Assessment efficiency of microbial-organic fertilizers from beer and seafood sludges on vegetable yield in the field conditions
Assessment efficiency of microbial-organic fertilizers from beer and seafood sludges on vegetable yield of mustard, cucumber, okra, and winter melon in the field conditions was carried out All experimental designs were arranged randomly block with 6 treatments and three replications for each, listed as follows: T1: Farmer rate
Trang 12fertilizer (FR) (control); T 2: Recommended rate chemical fertilizer (RR): T3: RR+ 5 tons/ha micro-organic fertilizer from BS (bioF-BS); T4: 70% RR + 5 tons/ha bioF-BS; T5: RR+ 5 tons/ha micro-organic fertilizer from SS (bioF-SS); T 6: 70% RR + 5 tons/ha bioF-SS The research on mustard, cucumber and winter mellon was carried out at Long Tuyen commune, Binh Thuy district, Can Tho city Okra research was carried out at My Hoa commune, Binh Minh district, Vinh Long province Mustard plants spacing were 40 cm between the plants within each row and 30 cm between the rows The area of one treatment plot was 15.75 m2 (10.5 long × 1.5 m wide) For cucumber, plants spacing were 40 cm between the plants within each row and 1.5
m between the rows The area of one treatment plot was 10.5 m × 2 5
m Winter melon plants spacing were 40 cm between the plants within each row and 60 cm between the rows The area of one treatment plot was 9,6 x 2,6 m For okra, each block was 30 m long and equally divided into two treatment plots, with each treatment 15 m long and a width of 1.5 m
Yield (tons/ha) of vegetables was determined at harvest
2.6 Isolation and selection of fungal strains capable of decomposing organic materials
2.6.1 Isolation fungal strains decomposed cellulose and chitin
The samples rice straw (R) was collected in Dong Thap and Vinh Long provinces Coconut fiber (D) was sampled in Vinh Long province Bagasse (M) sample was collected in Vinh Long province and Can Tho city The stubble was cut further into small pieces and aseptically incubated on sterile Hagen agar amended with sodium carboxy methyl cellulose (10g/l) and Streptomycin 30 ppm for isolation of cellulolytic fungi In addition, crude chitin was washed and neutralized with acid for pH = 7
To examine if all colonies appearing on cellulose amended media were cellulolytic, fresh pure cultures were treated with 0.1% congo red stain followed by procedure described by Alström (2000) Presence of chitinolytic activity of fungi was evaluated according to Arora et al (2005) The degree of enzymatic activity was assessed by measuring the radius of the clear zone appearing around each colony
Trang 132.6.2 Assessment effect of isolated fungi on rice straw and bagasse decomposition
Rice straw was cut into approximately 5 cm pieces and bagasse was verificated 5mm The microcosms used for the experiment consisted of aucto clavable polyethylene bags (50 cm x 31 cm) containing straw and bagasse (20 gram dry weight) and distilled water enough to give 60% moisture The bag was closed with a cotton plug and autoclaved twice for one hour each
A total of 8 isolated fungi, including D91B7, M-LT3, M-LT4,
R-ĐT1, R-NVT1, M-2HA1, M-NK1, M-TA3, Trichoderma-ĐHCT
(positive control), and non fungus (negative control) The suspensions were diluted before aseptically inoculated to microcosms (106 CFU/1g gram dry weight) using as syringe and sealing the needle hole with tape
After eight weeks, the rice straw and bagasse were harvested and dried over night in an oven at 60oC The biomass from each microcosm was calculated and transformed to % weight loss compared with the weight at the beginning of the experiment
2.6.3 Resistance of isolated fungi to the R solani fungus
Rice straw and basgass decomposed by four advanced strains were used as substrates to assess the antagonistic capacity of R solani pathogen over a 4 week period as well as the resistance of these fungal strains against R solani on PDA agar The experiment was arranged in
a randomized form with 6 treatments and 3 replications for each treatment After 4 weeks of incubation, the diameter of R solani was measured and compared with the control diameter of R solani
2.6.4 Identify fungal strains isolated by molecular biological
Four fungal cellulosic and chitin decomposed strains in section
2.6.2 were selected to verify their degree of decomposition on mixture
of sludge and sugarcane cake All equipment and researched method
were similar to Section 2.3.1
Trang 142.7 Measurements and Statistical Analysis
Data was analyzed using the SPSS package version 16.0 Data were analyzed by analysis of variance (ANOVA) and Duncan’s multiple range tests National standards is used to evaluate the quality
of sludge and microbial organic compost
CHAPTER 3 RESULTS AND DISCUSSION
3.1 Determination physical, chemical, nutrient and biological characteristics of beer and seafood sludges and other materials
3.1.1 Physical characteristics of materials
As sludges do not contain cellulose compounds but they contain many protein compounds, sugars, starches and organic acids (Feng et al., 2008; Mook et al., 2012; Olajire, 2012) Whether BS and SS were not only good porosity but also high moisture, so it is necessary to mixure these sludges with other high cellulose materials such as sugarcane sludge, bagasse, or straw (Table 3.1)
Table 3.1 Bulk density and moisture of materials before compost
(g/cm3)
Moisture
EC (mS/cm)
Note: Mean ±SD, BS: beer sludge, SS: seafood sludge, “-“: not data
The pH value of BS and SS varied from 5.71 to 7.6, which was suitable for reuse in composting organic fertilizers The EC value was also high in all materials, including both sludges (table 3.1) because several chemicals and salts were mixed milled malt, gelatinized adjunct and water to obtain a high yield of extract product (Olajire, 2012; Park et al 2010) For seafood sludge, high levels of EC could be attributed to the origin of aquatic species as saltwater aquaculture