THE CANADIAN SCIENCE FAIR JOURNAL ARTICLE

BSFL Frass: A Novel Biofertilizer for Improving Plant

Health While Minimizing Environmental Impact
Sarah Choi & Neelah Hassanzadeh
Age 17 | Langley, British Columbia
Canada-Wide Science Fair Excellence Award: Senior Silver Medal | Dalhousie University $2,500 Entrance Schol-
arship | UBC Science $2,000 Entrance Award | University of New Brunswick Canada-Wide Science Fair $2,500
Scholarship | University of Ottawa $2,000 Entrance Scholarship | Western University $2,000 Entrance Scholarship

The common synthetic fertilizer’s ability to increase nitrate levels is mitigated by the leaching of nitrate into aquatic ecosystems and wa-
terways; this contamination eventually causes eutrophication and pollutes groundwater (Dubey & Townsend, 2004). Synthetic fertilizer
application has also been a contributor to increased nitrous oxide emissions (Sanders, 2012). A novel solution is using black soldier fly
larvae (BSFL) frass as organic fertilizer.

INTRODUCTION this assimilation of nitrate to plant roots (Figure 1) makes soils

Black soldier flies are native to North America, and their frass more resilient to floods, droughts, and land degradation process-
—the solid excrement of insect larvae—contains the only es. Frass also consists of safer chemicals for human exposure,
plant-digestible form of chitin. This chitin naturally produc-
whereas synthetic fertilizers often are composed of harmful sub-
es antimicrobial peptides when under environmental stress
(Sistrunk, 2016), acting as a protective barrier. The chitin does stances such as cadmium, uranium, and arsenic, all of which are
this by making mineral nutrients inaccessible to pathogens carcinogens as well as triggers for developmental problems in
as well as by blocking the release of pathogenic mycotoxins children (Sharma & Singhvi, 2017). In addition to all these ben-
(Wan, Zhang, & Stacey, 2008); this capability has been prov- eficial components, if BSFL frass possesses defensive properties
en to be highly beneficial for insects, although it is not yet for plants, it could be a viable alternative to chemical fertilizers
fully understood in its effects for plants (Behie & Bidochka, for plant growth. This study aims to determine if BSFL frass
2013). Frass also contains nitrifying bacteria and nitrogen-fix- defends against plant pathogens, thereby increasing soil fertility.
ing bacteria, which partake in the nitrogen cycle and assist in
Although chitin has been shown to provide positive effects for
the plants’ uptake of nitrogen (Behie & Bidochka, 2013). It has
been shown that there are more greenhouse gases, like nitrous
oxide, in areas where more synthetic fertilizer is applied to the
soil (Hawkinson, 2005), whereas BSFL frass can store carbon
and nitrogen in the soil (Lovett et al., 2002). BSFL frass also
prevents atmospheric loss of nitrogen and groundwater con-
tamination through the nitrogen fixation by the bacteria (Lovett
et al., 2002). The atmospheric nitrogen cannot be directly as-
similated by plants, which is why there must be a nitrogen fix-
ation process by bacteria to make the nitrogen available for
plant uptake. In frass, bacteria like Bacillus and Pseudomonas
help fix atmospheric bacteria while other nitrifying bacteria
make nitrogen within the soil accessible to plants for photo-
synthesis (Zahn, 2017). Nitrifying bacteria convert nitrogen in
its ammoniacal form its nitrate form, which allows for more
efficient root uptake by the plant (Alattar, Alattar, and Popa,
2016). The nitrifying and nitrogen-fixing bacteria are critical
to horticultural production because fixed nitrogen is a limit-
ed nutrient in most environments (Zahn, 2017). Furthermore,

This work is licensed under:

https://creativecommons.org/licenses/by/4.0 Figure 1. Impact of nitrifying and nitrogen-fixing bacteria in frass
doi: 10.18192/csfj.v2i220194146 on the nitrogen cycle.

CSFJ | Volume 2 | Issue 2

© Choi & Hassanzadeh 2019 41
 THE CANADIAN SCIENCE FAIR JOURNAL ARTICLE

insects (Zahn, 2017), this study aims to solidify this role of Part IIa: Fertilizer Trial
chitin and bacteria in the protection of plants specifically, and
Treatment Group Composition
frass’s benefits as a fertilizer. The first objective of this study is
to investigate whether common fungi, Rhizoctonia solani and 1. Control 350.00 g soil + 300 mL RO
Fusarium oxysporum, will grow in frass. Secondly, this study water
examines the effects of frass as a fertilizer and as a protective 2. Soil + Humic acid 350.00 g soil + 300 mL RO
barrier against the common pathogen, Pythium myriotylum. water + 600 uL humic acid
This study also examines a combination of frass with other or- 3. Soil + Trichoderma 350.00 g soil + 300 mL RO
ganic fertilizers, such as Trichoderma, which is a non-patho- water + 0.665 g Trichoderma
genic fungus that stimulates plant growth and nutrient uptake, 4. Soil + 1.5% BSFL frass 344.75 g soil + 300 mL RO
and humic acid, which raises the nutrient-holding capacity of water + 5.250 g frass
soil (Gonsalves & Ferreira, 1993). The addition of other or-
5. Soil + Humic acid + 350.00 g soil + 300 mL RO
ganic fertilizers will allow for a comparison between frass and
Trichoderma water + 600 uL humic acid +
these more common fertilizers, in addition to an assessment of 0.665 g Trichoderma
their combined effects.
6. Soil + 1.5% BSFL frass + 344.75 g soil + 300 mL RO
HYPOTHESIS Humic Acid water + 600 uL humic acid +
It is our hypothesis that the treatments with frass will show 5.250 g frass
no bioaccumulation of pathogens in the petri dishes and will 7. Soil + 1.5% BSFL frass + 344.75 g soil + 300 mL RO
demonstrate better plant health and growth, whereas control Trichoderma water + 0.665 g Trichoderma
groups will have little to no positive effect. + 5.250 g frass
METHODS 8. Soil + 1.5% BSFL frass + 344.75 g soil + 300 mL RO
Part I: Transmission of Disease to BSFL Frass Humic acid + water + 600 uL of humic acid
Culturing the Plant Pathogens: A 7 mm plug of Fusarium ox- Trichoderma + 0.665g of Trichoderma +
ysporum was obtained from Kwantlen Polytechnic University. 5.250g of frass
The plug was transferred to a petri dish with potato dextrose 9. Soil + Water-soluble 350.00 g soil + 300 mL RO
agar (PDA) medium in a microbiological safety cabinet to cul- (chemical) fertilizer water + 0.600 g water-solu-
ture fungi; this process was repeated for Rhizoctonia solani. ble fertilizer
Five petri dishes were set up for each fungus and stored in an Part IIb: Disease Trial
incubator at 27°C for one week.
Treatment Group Composition
Retrieving Frass: Sixteen plastic cups were filled with 20
BSFL: eight for both F. oxysporum and R. solani, with four Control 350.00 g soil + 300 mL H2O
cups receiving the regular fruit and vegetable diet without fun- (mixed with V8 media)
gus (control), and four cups with the diet containing fungus. Soil + Pythium 350.00 g soil + 300 mL H2O
The amount of diet varied per day based off the Enterra diet (mixed with Pythium)
chart for BSFL larvae. The BSFL received food every two days. Soil + Pythium + 1.5% frass 344.75 g soil + 300 mL H2O
After a week, frass was collected from the cups and placed into (mixed with Pythium) +
16 petri dishes with PDA. The petri dishes were stored in an 5.250 g BSFL frass
incubator at 27°C for one week. Soil + Pythium + 1.5% frass 344.75 g soil + 200 mL H2O
Part I was repeated for a second trial to ensure precision in the + Humic acid + Trichoderma (mixed with Pythium) + 100
results. mL H2O (mixed with 600
Part II: Growth of Valentino Green Bush Bean Plants with µm of humic acid) + 5.25 g
Frass BSFL frass + 0.665 g Trich-
Part IIa: Arranging Plants for Fertilizer Trial (no Pythium) oderma
Valentino green bush bean plants were used for Part II and were Soil + Pythium + Water-solu- 350.00 g soil + 200 mL H2O
obtained as seeds. Nine different treatment groups of fertilizers ble (chemical) fertilizer (mixed with Pythium) + 100
(Table 1) were tested, with five replicates per treatment. All in- mL H2O (mixed with 0.600 g
dividuals were planted in pots (4 in tall) with sterile soil. Treat- of chemical fertilizer)
ments were mixed with 300 mL of water and given to the plant.
Each individual was given approximately 20 mL of water every Table 1. An outline of the composition of each treatment
three days. For 56 days, all plants were given 8 hours of light groups for Parts IIa and IIb.

CSFJ | Volume 2 | Issue 2

© Choi & Hassanzadeh 2019 42
 THE CANADIAN SCIENCE FAIR JOURNAL ARTICLE

and then 16 hours of darkness each day, which is a standard biomasses compared to both the control and the treatments with
light/dark interval for plant studies (Shin, Song, & Thompson, the chemical fertilizer (figures 2 and 3). In the fertilizer trial,
2011). They were held in an incubation room set to 25°C. the treatment of frass and humic acid showed the highest mean
Part IIb: Arranging Plants for Disease Trial (with Pythium) height of approximately 61.02 cm (table 2). In the disease trial, the
Five different treatment groups of fertilizers with Pythium myri- treatment of the combination of frass, humic acid, and Trichoderma
otylum (Table 1) were tested, with five replicates per treatment. showed the highest mean height of 52.34 cm. The treatment with
The cultured fungus was scraped using a lab spatula and mixed the greatest dry biomass was the treatment of frass, humic acid, and
evenly throughout the soil for the disease groups before the Trichoderma, with a mean dry biomass of 13.18 g. Treatments con-
plants were potted. All individuals were planted in pots (4 in tall) taining frass improved plant biomass with disease by five-fold in
with sterile soil and were given 300 mL of water. Each individual comparison to the positive control. A Tukey-Kramer test showed
was given approximately 20 mL of water every three days. Each that both biomass and height data for fertilizer and disease trials
disease group had one whole plate of P. myriotylum (90 mm di- demonstrated significant results (p < 0.05).
ameter). For 56 days, all plants were given 8 hours of light and Nitrate Concentration, pH, and Electrical Conductivity
then 16 hours of darkness. In the fertilizer trial, the nitrate concentration was highest in
All water used was processed through a reverse osmosis the treatment of frass and humic acid, with 224 ppm (table 2).
(RO) filtration system. For both the disease and fertilizer trials, In the disease trial, the treatment with frass, humic acid, and
heights of the plants were recorded every fourth day. Wet and dry Trichoderma had the highest mean nitrate concentration of 228
biomasses, number of leaves, and width of leaves were tested ppm, and the following treatment was only frass with a mean
after 56 days. The pH and electrical conductivity of the soil were nitrate concentration of 212 ppm. A Tukey-Kramer test showed
tested using a multiparam-
eter probe, and the nitrate
concentration was tested
using a nitrate meter.
RESULTS
Part I: Transmission of
Disease to BSFL Frass
Both trials demonstrated
that the F. oxysporum and
the R. solani fungi did not
grow in frass. In the first
trial, after about a week,
there was only frass in the
plate with no growth of
anything observed. In the
second trial, a grey fluffy
substance had grown. After
further analysis using PCR,
it was determined that the
substance was not linked to
Fusarium nor Rhizoctonia,
and it was also not a plant
pathogen.
Part II: Growth of Val-
entino Green Bush Bean
Plants with Frass
Biomass and Heights
After 56 days in both the
fertilizer and disease trials,
the treatments with frass
showed higher heights and
Figure 2. Plants’ growth in height from Part IIa.

CSFJ | Volume 2 | Issue 2

© Choi & Hassanzadeh 2019 43
 THE CANADIAN SCIENCE FAIR JOURNAL
Figure 3. Plants’ growth in height from Part IIb.
that both of those treatments were found to be in the same sta-
tistical group, indicating that the two treatments both increased
nitrate in similar amounts and are not significantly different.
Treatments with frass had higher mean nitrate amounts com-
pared to soil with no treatment. The optimum pH of 6.5 to 7 for
bacterial populations (Perry, 2003) was achieved for all treat-
ments with frass, while all other groups without frass, such as
the control group, had pH values less than 6.5. The optimum
EC is 0.6 to 0.7 mS/cm according to Behie & Bidochka (2013).
In comparison to the soil with disease, which had the electrical
conductivity (EC) of 0.362 mS/cm, the soil with disease and
treatments with frass had mean EC values ranging from 0.658
mS/cm to 0.662 mS/cm.
DISCUSSION
This study demonstrates that BSFL frass does not transmit dis-
ease and defends against it, thereby a consistent addition of
frass to soil can contribute to the prevention of fungal disease
from pathogens like Rhizoctonia, Fusarium, and Pythium. When
testing for the transmission of disease in Part I, there was no
growth of R. solani or F. oxysporum observed in either of the two
trials, suggesting that the disease from the fungi (in the diet) was
metabolized by the larvae and converted into safer chemicals for
the environment. Thus, it is our interpretation that BSFL frass can
CSFJ | Volume 2 | Issue 2
© Choi & Hassanzadeh 2019
ARTICLE
be safely used as organic fertilizer without it
being contaminated.
Part II of the study further demonstrated
the beneficial impacts of frass on plant health
against Pythium. Pythium blight causes high-
ly destructive turfgrass disease, which is one
of the most common reasons for crop failure.
While this condition was not observed in any
pots with treatments including frass, some pots
from the positive control showed the appear-
ance of the disease; this confirmed our hypoth-
esis regarding frass’s ability to prevent bioac-
cumulation of the pathogen. Figure 3 shows
that the heights were greater for treatment
groups with frass, suggesting that immune re-
ceptors within the chitin successfully blocked
pathogens from harming the plant (Zahn,
2017). The addition of different organic fertil-
izers with frass was effective as frass allows
carbon dioxide and nitrates to stay in the soil
(Lovett et al., 2002) (which allows the plant to
access nutrients) while the humic acid enables
the soil to have a greater capacity of holding
water (Gonsalves & Ferreira, 1993). This im-
proved capacity for accessing nutrients and
moisture may explain the increased biomass-
es and heights for treatments including frass.
These results comply with the theory that there
is a positive relationship between biomass and
the production of heavier and healthier fruits
(Ceresini, 1999). The greater amount of nitrogen in the treatments
containing both frass and humic acid compared to the treatment
with frass demonstrates the ability of humic acid to improve the
nitrogen uptake by plants. Treatments with frass also had more
optimal pH ranges, which is important for beneficial bacterial
populations in frass to develop and promote plant growth (Perry,
2003). The pH results correspond with the high amounts of nitrate
in the treatment groups with frass, because a greater amount of
nitrifying and nitrogen-fixing bacteria allows more nitrogen to be
absorbed by the plant. Since all treatments with frass had higher
nitrate amounts compared to soil with no treatment, it was shown
that frass was able to increase nitrogen content in soil.
CONCLUSION
Our study can conclude that frass can improve plant growth with-
out the transmission of plant-pathogenic disease from compost
waste diet. Not only does frass improve soil fertility and defend
against pathogens, but it does so without harming the environ-
ment. Furthermore, frass is available on the market for consumers
at a lower, more competitive price than competing conventional
synthetic fertilizers (Zahn, 2017). BSFL frass is inexpensive to
mass produce and consume because of how easily available it is
44
 THE CANADIAN SCIENCE FAIR JOURNAL ARTICLE

Wet biomass (g) pH EC (mS/cm) NO3- (ppm) Temperature (°C)

6.24 5.496 0.51 150 22.22
11.23 6.526 0.666 198 21.82
12.34 6.804 0.686 224 21.7
11.83 6.494 0.622 188 21.88
8.24 6.326 0.31 168 22.28
7.89 6.386 0.338 156 22.2
9.87 6.228 0.348 170 22
13.18 6.574 0.654 222 21.62
6.89 5.286 0.51 156 22.14
Part IIb: Disease Trial
6.12 5.868 0.418 142 22.06
3.84 5.648 0.362 98.48 22.28
12.19 6.842 0.658 212 22.28
13.05 6.756 0.662 146 22.86
8.21 5.954 0.542 228 22.94

Tukey-Kramer Test Categorization of Nitrate in Disease Trial Treatments

Treatment Level Mean (NO3-)
Soil + Pythium + F + HA + T A 228
Soil + Pythium + F A 212
Soil + Pythium + C B 146
Soil (control) B 142
Soil + Pythium B 120
Table 2. Collection of data from all trials and statistical analysis of nitrate concentration for Part IIb. F = frass, H = humic
acid, T = Trichoderma, C = chemical fertilizer.

throughout the year, allowing for mass quantities of larvae to be
bred in a controlled environment. Frass also contains safer chem-
icals for human exposure. In addition, the production of frass oc-
curs in a cycle where waste crops such as fruit can be used to feed
the BSFL that generate the frass, then that frass can then be used
in the production of new crops. Therefore, the production method
is more self-sustaining and economically advantageous than that
of synthetic fertilizers which use many non-renewable resources,
such as fossil fuels (Woods, Williams, Hughes, Black, & Murphy,
2010). Thus, BSFL frass is safer for the consumer, less expensive,
more effective, environmentally sustainable, and can be applied
less frequently for a long-lasting effect so that consumers do not
have to contribute much time to its usage. By using this organic
fertilizer that can defend against plant pathogens, the frass will
not only help with providing essential nutrients for the plant in a
sustainable way, but also act as a substitute for synthetic fertilizers
that pollute the environment.
ACKNOWLEDGEMENTS
We would like to thank Kwantlen Polytechnic University for pro-
viding support and facility for this project.
REFERENCES
Alattar, M. A., Alattar, F. N., & Popa, R. (2016). Effects of microaerobic fer-
mentation and black soldier fly larvae food scrap processing residues on
the growth of corn plants (Zea mays). Plant Science Today, 3(1), 57-62.
https://doi.org/10.14719/pst.2016.3.1.179
Behie, S. W., & Bidochka, M. J. (2013). Insects as a nitrogen source for plants.
Insects, 4(3), 423-424. https://doi.org/10.3390/insects4030413
Dubey, B., & Townsend T. (2004). Arsenic and lead leaching from the waste de-
rived fertilizer ironite. Environmental Science Technology, 38(20), 5400-

CSFJ | Volume 2 | Issue 2

© Choi & Hassanzadeh 2019 45
 THE CANADIAN SCIENCE FAIR JOURNAL
5404. https://doi.org/10.1021/es0493392
Gonsalves, A. K., and Ferreira, S. A. (1993). Fusarium oxysporum. Retrieved
from http://www.extento.hawaii.edu/kbase/crop/Type/f_oxys.htm
Hawkinson, C. (2005). Black soldier fly. Retrieved from https://aggie-horticul-
ture.tamu.edu/galveston/beneficials/beneficial-51_black_soldier_fly.htm
SARAH CHOI
I am a senior student at R.E. Mountain Secondary School inter-
ested in research and biological studies. This year was my first
time competing in the Canada-Wide Science Fair with my proj-
ect about how a North American species (Black Soldier Fly) of
insect frass can be used as a safe and effective biofertilizer for
many crops, providing benefits to both the plant and the envi-
ronment - all with a production process that is economically ad-
vantageous and self-sustainable through the recycling of food
compost wastes. I was able to conduct my trial as a research
intern in the Institute of Sustainable Horticulture at Kwantlen
Polytechnic University in Langley, BC. I have worked in this
lab since September of 2017, and my experiences there and
at the national science fair sparked my interest in biology and
research, which I hope to further pursue in the future. Along
with my partner, I was honoured to receive a silver medal at
the Canada-Wide Science fair, the NSERC Young Innovator
award, and the Association of Professional Biology Provincial
(BC) Award for the highest BC standing with a biology project.
CSFJ | Volume 2 | Issue 2
© Choi & Hassanzadeh 2019
ARTICLE
Lovett, G., Christenson, L. M., Groffman, P. M., Jones, C. G., Hart, J. E.,
& Mitchell, M. J. (2002). Insect defoliation and nitrogen cycling in
forests: Laboratory, plot, and watershed studies indicate that most of
the nitrogen released from forest foliage as a result of defoliation by
insects is redistributed within the ecosystem, whereas only a small
fraction of nitrogen is lost by leaching. BioScience, 52(4), 335-341.
NEELAH HASSANZADEH
My name is Neelah Hassanzadeh and I am a senior highschool
student. I started volunteering at the Institute of Horticulture
(KPU) in June 2018 and continued to work at ISH throughout
the year. Sarah and I decided to start our own project about Black
Soldier Fly Larvae frass as an organic fertilizer that is able to de-
fend against harmful pathogens in October 2018. We worked on
this project under the supervision of Dr. Sepideh Tahriri Adabi, a
research scientist at ISH. Now, I continue to work as a research
assistant at ISH and aspire to become a nurse.
46