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Lipid metabolism, fatty acid composition and meat quality in broilers supplemented with increasing levels of defrosted black soldier fly larvae

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Abstract

A feeding experiment was conducted to investigate the impact of feeding defrosted whole black soldier fly larvae (BSFL) to broilers in increasing levels in the ration on blood metabolites, carcass characteristics (CC) and on changes in fatty acid (FA) composition in plasma, muscle and abdominal fat. Day-old chicks (Ross-308; n=252) were assigned to one of four groups each with 6 replicate pens (10-11 birds/pen). The birds were fed either a demand-oriented age-specific control (CON) diet and had no access to BSFL, or fed CON plus BSFL at 10% (L10), 20% (L20) or 30% (L30) of CON feed intake. At weeks (wk) 4 and 6, birds (2 per pen) were slaughtered to collect blood, breast muscle, and abdominal fat samples and to determine CC. Plasma triglyceride concentrations increased in a dose dependent manner with increasing levels of whole BSFL compared with CON (P<0.05). The L30 and L20 had higher plasma non-esterified FA concentrations than CON (P<0.05). There were no differences in slaughter weight and CC between groups (P>0.05). Broilers fed 30% BSFL had the highest saturated FA proportion in plasma, muscle and abdominal fat and the lowest monounsaturated FA proportion in abdominal fat tissue (P<0.05). The levels of total polyunsaturated FA in plasma and abdominal fat were lower in L30 than in CON (P<0.05). In plasma, muscle and abdominal fat, the proportion of conjugated linoleic acid (isomer C18:2cis-9, trans-11) was highest in L30 followed by L20 and L10 compared with CON (P<0.05). Overall, whole BSFL could be included in broiler diets up to 20% to promote sustainability in broiler farming without adverse effects on slaughter weight, meat quality and FA compositions, whereas, the highest inclusion level (i.e. 30%) of whole BSFL in the daily ration was associated with altered FA composition in plasma, fat and meat.
Journal of Insects as Food and Feed, 2022; ##(##): 1-16 Wageningen Academic
Publishers
ISSN 2352-4588 online, DOI 10.3920/JIFF2022.0125 1
1. Introduction
Heavy reliance on soybean and fishmeal as protein feed for
commercial livestock is no longer considered sustainable
(Dörper et al., 2020). To support a more sustainable and
environmentally friendly production of feed for livestock,
including poultry, insects meals such as from black soldier
fly larvae (BSFL; Hermetia illucens), have been suggested
as alternative protein feed in exchange for soybean meal
(Cutrignelli et al., 2018; De Souza Vilela et al., 2021a;
Maurer et al., 2016; Schiavone et al., 2017). Whole insect
larvae are a protein-rich source of natural feed consumed
by wild birds and free-range poultry (Rumpold et al., 2017).
Whole BSFL contain 33 to 59% crude protein (CP), and
11 to 34% lipids in dry matter (DM), which make them
an interesting ingredient for poultry feed (Bava et al.,
2019; Maurer et al., 2016; Shumo et al., 2019). In recent
years, increasing attention has been paid to using BSFL in
the form of meal or oil in broiler diets. Previous studies
demonstrated a successful incorporation of BSFL meal
in broiler diets without negative effects on key carcass
traits and meat quality including carcass composition,
meat pH and colour, cooking loss, or lipid oxidation in
broiler meat (De Souza Vilela et al., 2021a; Popova et al.,
Lipid metabolism, fatty acid composition and meat quality in broilers supplemented
with increasing levels of defrosted black soldier fly larvae
M.M. Seyedalmoosavi1, D. Dannenberger2, R. Pfuhl2, S. Görs1, M. Mielenz1, S. Maak2, P. Wolf3, G. Daş1* and
C.C. Metges1
1Research Institute for Farm Animal Biology (FBN), Institute of Nutritional Physiology, Wilhelm Stahl Allee 2, Dummerstorf,
18196, Germany; 2Research Institute for Farm Animal Biology (FBN), Institute of Muscle Biology and Growth, Wilhelm
Stahl Allee 2, Dummerstorf, 18196, Germany; 3University of Rostock, Nutrition Physiology and Animal Nutrition, Faculty
of Agricultural and Environmental Sciences, Justus-von-Liebig-Weg 6, Rostock, 18059, Germany; gdas@fbn-dummerstorf.de
Received: 27 August 2022 / Accepted: 15 October 2022
© 2022 Wageningen Academic Publishers
RESEARCH ARTICLE
Abstract
A feeding experiment was conducted to investigate the impact of feeding defrosted whole black soldier fly larvae
(BSFL) to broilers in increasing levels in the ration on blood metabolites, carcass characteristics (CC) and on changes
in fatty acid (FA) composition in plasma, muscle and abdominal fat. Day-old chicks (Ross-308; n=252) were assigned
to one of four groups each with 6 replicate pens (10-11 birds/pen). The birds were fed either a demand-oriented
age-specific control (CON) diet and had no access to BSFL, or fed CON plus BSFL at 10% (L10), 20% (L20) or
30% (L30) of CON feed intake. At weeks (wk) 4 and 6, birds (2 per pen) were slaughtered to collect blood, breast
muscle, and abdominal fat samples and to determine CC. Plasma triglyceride concentrations increased in a dose
dependent manner with increasing levels of whole BSFL compared with CON (P<0.05). The L30 and L20 had higher
plasma non-esterified FA concentrations than CON (P<0.05). There were no differences in slaughter weight and
CC between groups (P>0.05). Broilers fed 30% BSFL had the highest saturated FA proportion in plasma, muscle
and abdominal fat and the lowest monounsaturated FA proportion in abdominal fat tissue (P<0.05). The levels of
total polyunsaturated FA in plasma and abdominal fat were lower in L30 than in CON (P<0.05). In plasma, muscle
and abdominal fat, the proportion of conjugated linoleic acid (isomer C18:2cis-9, trans-11) was highest in L30
followed by L20 and L10 compared with CON (P<0.05). Overall, whole BSFL could be included in broiler diets up
to 20% to promote sustainability in broiler farming without adverse effects on slaughter weight, meat quality and
FA compositions, whereas, the highest inclusion level (i.e. 30%) of whole BSFL in the daily ration was associated
with altered FA composition in plasma, fat and meat.
Keywords: carcass, chicken, fatty acid profile, Hermetia illucens, defrosted whole larvae
OPEN ACCESS
Journal of Insects as Food and Feed, 2022 online ARTICLE IN PRESS
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M.M. Seyedalmoosavi et al.
2 Journal of Insects as Food and Feed ##(##)
2020). Moreover, using full-fat BSFL meal in the diet of
growing layer chickens improved growth performance,
nutrient digestibility, plasma antioxidant ability and gut
health (Chu et al., 2020). In addition, inclusion of BSFL
fat in broiler diets did not affect blood metabolites, and
carcass traits and chemical composition of broiler meat
were satisfactory (Dabbou et al., 2021; Schiavone et al.,
2017). However, the industrial processing of BSFL to
produce protein meal, implies additional costs which might
constrain expansion of using BSFL in poultry rations. This
may be particularly important for low-input poultry farming
systems particularly in developing countries. There are
few reports on the successful inclusion of whole BSFL in
poultry diets supporting growth performance (Ipema et
al., 2020a,b; Star et al., 2020; Tahamtani et al., 2021) and
meat quality including FA profile and protein content in
meat (Moula et al., 2018).
BSFL fat is rich in saturated fatty acids (SFA) and contains
considerable amounts of medium chain fatty acids (MCFA),
particularly lauric acid (C12:0) (Li et al., 2022). Lauric acid,
which can constitute up to 50% of BSFL fat (Franco et al.,
2021; Li et al., 2022), can also be converted to monolaurin
(or glycerol monolaurate), which has growth-promoting
potential and could explain the antibacterial activity
of BSFL fat (Almeida et al., 2020; Franco et al., 2021).
Moreover, MCFA can have positive effects on energy
availability without increasing lipid deposition (Li et al.,
2016). However, BSFL fatty acid (FA) composition is highly
variable as it heavily depends on the FA composition of the
feeding substrates for larvae (Ewald et al., 2020). The FA
composition of broiler meat is also dependent on the feed
source (Cullere et al., 2019; De Souza Vilela et al., 2021a;
Dörper et al., 2020), therefore feeding BSFL to broilers has
the potential to modulate FA composition of the particular
meat products (e.g. breast meat), and may enrich MCFA in
edible tissues (Franco et al., 2021). It has been reported that
BSFL fat can be used to incorporate considerable amounts
of monounsaturated fatty acids (MUFA) contents in chicken
breast and leg meats of broilers (Cullere et al., 2019).
Moreover, it has been suggested that lower proportions
of polyunsaturated fatty acids (PUFA) in BSFL fat may
increase antioxidant capacity in BSFL oil-fed chickens
(Kim et al., 2020).
Despite the reported benefits of BSFL in broiler diets,
there seem to be trade-offs in the meat FA composition,
characterised by increased SFA at the expense of PUFA
(Cullere et al., 2019; Schiavone et al., 2017). There is only
one study available in which whole BSFL were used in
poultry diets (8% of commercial feed) but no difference
in the FA composition of meat between BSFL-fed and
control birds were observed (Moula et al., 2018). There
is a lack of knowledge on the amount of whole BSFL that
can be included in broiler diets without adverse effects
on metabolism and FA compositions in different tissues
of chickens. Therefore, the objective of this study was to
investigate the effects of feeding whole BSFL to broilers in
increasing dietary levels for 42 d on carcass characteristics,
blood metabolites, and particularly on changes of FA
compositions in plasma, muscle and abdominal fat tissues.
2. Material and methods
Chickens and management
The feeding experiment was registered under A.Z.
202022_70_A28_anz. A total of 252 mixed-sex newly
hatched chicks (Ross 308) was obtained from a commercial
hatchery and housed at the experimental poultry facility
at the Research Institute for Farm Animal Biology (FBN),
Dummerstorf, Germany. The chicks were weighted at
arrival (42±0.38 g/bird) and randomly allocated to one of
24 pens (n=10-11 chicks / pen) in four adjacent rooms of
the facility, which resulted in 63 birds in 6 pens for each
group. Pens of each room (n=6) were separated from
each other with solid panels. Each pen was equipped
with one feeder, a line of drinking nipples, and a deep
layer of wood shavings as litter material. Throughout the
experiment, birds in different rooms were raised under
the same environmental conditions. Climate conditions
in the rooms were automatically controlled based on
recommendations of the Aviagen Ross broiler handbook
(Aviagen, 2018) with some modification by a ventilation
and heating system, ensuring uniform ambient temperature,
light and ventilation conditions across the pens within and
between 4 experimental rooms. Ambient temperature at
the start of the experiment was 33°C and was gradually
decreased to 21°C at week (wk) 6, whereas humidity was
gradually increased from 37 to 70% until wk 6.
Experimental design, diets and black soldier fly larvae
provision
A completely randomised design with 4 treatments was
used in this study. All birds received the same basal diet
in mesh feed form. The basal diet was designed to meet or
exceed age-specific nutrient recommendation of broilers
(Aviagen, 2019) in three phases, i.e. starter (d 1-14), grower
(d 15-28) and finisher (d 29-42) diets (Table 1). Equal
numbers of pens (n=6 per group) and birds (n=63 per
group) were randomly allocated to each of the 4 dietary
treatments with each treatment occurring in each room at
least once. Broilers in the control group (CON) received the
age-specific basal diet, and had no access to BSFL. Birds
in the remaining 18 pens received defrosted whole BSFL
in addition to the basal diet at increasing levels, i.e. 10%,
20% or 30% of the feed intake (FI) of CON birds (hereafter
referred to as groups L10 (n=63), L20 (n=63), and L30
(n=63), respectively). Except for the first day (d1), the daily
amount of BSFL to be fed to the broilers in L10 to L30
groups was calculated based on FI of the CON birds on
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Defrosted whole black soldier fly larvae in broiler diets
Journal of Insects as Food and Feed ##(##) 3
the previous day. On d1, FI of broiler birds from previous
experiments was used as a reference. Pen based daily FI
was measured in the mornings before feeding larvae, and
the average FI per bird was then calculated. The thawed
whole larvae to be given to the birds of a pen were weighed
and placed on a feeding plate which was then placed on
the ground of the recipient pen at the same time each day
(07:30 h). Cumulative total fresh matter intake (FMI; the
sum of feed and larvae intake as is) per an average bird over
42 d of each pen was calculated. Based on the amounts
of cumulative feed and BSFL intakes, and the nutrient
and energy contents of the diets and BSFL, pen based
cumulative fat and energy intakes trough both feed and
larvae consumption were calculated for an average bird over
the complete fattening period (42 d). Live BSFL (a mix of
5
th
-6
th
instars) were purchased from Hermetia Deutschland
GmbH & Co. KG, Baruth/Mark, Germany. The larvae used
in this experiment originated from the same rearing batch,
and had been fed a company-specific feeding substrate
in accordance with feed regulations. As soon as the live
larvae were received, they were snap frozen using liquid
nitrogen and stored at -20°C until fed to broilers. Twelve
hours before broilers were offered, the frozen larvae were
thawed in a refrigerator (4°C). Broilers received defrosted
BSFL at approximately room temperature.
Chemical analysis of feed and black soldier fly larvae
During the experiment feed and larvae sub-samples were
collected at regular intervals and stored at -20°C for
analysis. At the end of the experiment, all of the sub-samples
were pooled by feed type (e.g. starter, grower, and finisher)
and representative samples were analysed for their nutrient
contents. Analyses of nutrient composition in larvae and
feed samples were performed for DM content, crude ash, CP,
crude fat, starch, crude fibre, total sugar, neutral detergent
fibre (NDF), acid detergent fibre (ADF), acid detergent
lignin (ADL) and macro minerals by the accredited feed
laboratory of Landwirtschaftliche Untersuchungs-und
Forschungsanstalt der LMS Agrarberatung GmbH (LUFA,
Rostock, Germany) using standard methods (Naumann
and Bassler, 1997). Metabolizable energy contents of feed
and BSFL were then calculated (Naumann and Bassler,
1997). Chitin content of BSFL was calculated as ADF –
ADL as described (Hahn et al., 2018). Table 1 presents the
ingredients and chemical compositions of the age-specific
basal diets, and summarises the nutrient composition of
BSFL. The basal diet and drinking water were provided
to the birds ad libitum throughout the experimental
period. The FA composition of the feed and BSFL samples
were determined as explained in next sections. The
FA composition of the age-specific diets and BSFL are
summarised in Supplementary Table S1. BSFL fat contained
71% of SFA followed by 18% MUFA and 11% of PUFA
(Supplementary Table S1).
Slaughtering and sample collection
In the end of wk 4 and wk 6, two birds per pen (n=48 birds/
time point) were randomly chosen from their pens, weighed
and slaughtered after electrical stunning at the institute’s
EU approved abattoir. The stunning and slaughtering of
the birds were conducted according to the German animal
welfare regulations. From each slaughtered bird, blood was
collected in K3-EDTA-coated vacutainers (Sarstedt AG &
Co., Nümbrecht, Germany) to obtain plasma. Plasma was
harvested after centrifugation at 2,500×g for 20 min at 4°C,
aliquoted in 2 ml reaction tubes, and frozen at -20°C until
the determination of cholesterol, glucose, non-esterified
FA (NEFA), triglycerides and FA compositions.
Carcass characteristics
Immediately after slaughter, the carcasses were labelled
and weighed to determine hot carcass weight. Wings, legs,
breast, and abdominal fat were dissected and weighed. In
addition, breast muscle and abdominal fat were sampled
from individual birds and stored at -20°C for FA analysis.
The pH values of the breast muscle samples were measured
at 30 min, 45 min and 24 h post mortem (p.m.) with a
portable pH meter (pH-Star, Matthäus, Eckelsheim,
Germany) in the thickest part of the muscle. The colour of
the breast muscle samples was measured at 24 h p.m. using
a chromameter (Minolta CR 200, Ahrensburg, Germany)
with triplicate measurements from a freshly cut surface
using the parameters L* (brightness), a* (red-green) and b*
(yellow-blue). Chroma ((a
*2
+b
*2
)
1/2
) and hue (arctan (b
*
/a
*
))
indexes were calculated (Kongsup et al., 2022). Water
holding capacity was measured following descriptions of
Grosse et al. (1975).
The breast muscle DM was determined according to AOAC
methods (AOAC, 2000). The nitrogen content in breast
muscle samples was measured using an elemental analyser
(FlashEA 1112 NC Analyzer by Thermo Fisher Scientific,
Bremen, Germany). For this, 1 to 1.5 mg of sample was
weighed in tin capsules (IVA-Analysentechnik GmbH
und Co. KG, Meerbusch, Germany (Size: 3.3×5 mm)) and
sealed airtight. The samples were combusted at 900°C to
carbon dioxide and nitrogen oxides; the latter were reduced
to elemental nitrogen at 680°C. Combustion gases were
separated via gas chromatography with helium as the carrier
gas and quantified by a thermal conductivity detector. The
nitrogen content was then multiplied with the factor of 6.25
to calculate the crude protein content.
Blood metabolites
Plasma NEFA, cholesterol, glucose and triglycerides
concentrations of the birds were analysed with an automatic
enzymatic analyser (ABX Pentra 400, Horiba Medical,
Montpellier, France), using commercial kits [Glucose:
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M.M. Seyedalmoosavi et al.
4 Journal of Insects as Food and Feed ##(##)
Kit No. A11A01667, cholesterol: Kit No. A11A01634;
triglyceride: Kit No. A11A01640 (Horiba ABX); NEFA:
Kit No. 434-91795 (Wako Chemicals GmbH, Neuss,
Germany)].
Lipid extraction and transesterification of feed, black
soldier fly larvae, and chicken plasma, breast muscle and
abdominal fat samples
Feed and larvae
Larvae and feed samples were freeze dried and the samples
were finely ground using liquid nitrogen in a mortar with
a pestle. For extraction and direct FA methylation of feed
and larvae samples, a modified method from Sukhija and
Palmquist (1988) was used. The samples were treated
with 2 ml toluene (containing 19:0 methyl ester as internal
standard) and 4 ml of 5% methanolic HCl. The mixture
was shaken in a water bath at 60°C for 2 h. After cooling,
fatty acid methyl ester (FAMEs) were extracted with 3.5 ml
toluene in the presence of 8.75 ml 6% K
2
CO
3
solution and
vortexed. After centrifugation (ScanSpeed 40, LaboGene,
Allerød, Denmark) at 1,200×g for 5 min (at 4°C), the toluene
phase was separated, and 1 g Na
2
SO
4
and activated charcoal
were added and the mixture was stored overnight until the
organic phase became colourless. After filtration, an aliquot
of the toluene extract was taken and stored at -18°C until
GC analysis (Kalbe et al., 2019).
Table 1. Ingredients and analysed chemical composition of the age-specific broiler diets and defrosted whole black soldier fly
larvae (BSFL) offered to the broilers during the experimental period.1
Age-specific basal diets
Starter (d 1-14) Grower (d 15-28) Finisher (d 29-42) BSFL (d 1-42)
Ingredients, %
Soybean meal 48% 36.0 34.0 26.5 -
Wheat 31.0 28.0 35.0 -
Maize 21.5 28.0 28.0 -
Barley 5.0 4.0 5.0 -
Linseed oil 3.0 3.0 3.0 -
Vit-min. premix22.5 2.5 2.5 -
Oyster shells 1.0 0.5 0.0 -
Chemical analysis, g/kg DM
Dry matter 893 891 892 312
Crude ash 61.6 48.3 44.8 80.4
Crude protein 228 218 203 435
Crude fat 47.0 47.1 42.6 278
Crude fibre 24.6 25.8 34.8 72.7
Starch3464 508 517 14.3
Total sugar (calculated as sucrose) 43.7 46.0 42.6 2.2
NDF 118 11 4 110 121
ADF 41.4 43.8 38.1 77.8
ADL n.d. n.d. n.d. 4.42
Chitin4n.d. n.d. n.d. 73.4
ME, MJ/kg DM 13.4 14.0 13.8 16.5
Minerals, g/kg DM
Calcium 12.1 7.6 6.1 18.1
Phosphorus 6.6 5.2 5.2 9.3
Magnesium 2.1 1.9 2.0 3.9
1 ADF = acid detergent fibre; ADL = acid detergent lignin; n.d. = not determined; NDF = neutral detergent fibre.
2 Amount of vitamin and minerals provided through premix per kg of feed were as following; Vit. A 10,000 IU, Vit. D3 2,000 IU, Vit. E 20 mg, Vit. K3 3 mg, Vit.
B1 1 mg, Vit. B2 6 mg, Vit. B6 3 mg, Vit. B12 30 µg, Niacin 30 mg, Pantothenic acid 10.8 mg, Folic acid 0.4 mg, Biotin 24 µg, Cholin 300 mg, Fe 55 mg, Cu 18
mg, Zn 80 mg, Mn 93 mg, I 0.66 mg, Se 0.34 mg, Co 0.05 mg, phytase 250 FTU.
3 For BSFL it is glycogen.
4 Calculated based on Hahn et al. (2018) (i.e. Chitin = ADF – ADL).
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Defrosted whole black soldier fly larvae in broiler diets
Journal of Insects as Food and Feed ##(##) 5
Muscle, abdominal fat and plasma
The frozen breast muscle and adipose tissue samples were
cut into small pieces and homogenised using a mill (Model
A 11 Basic, IKA GmbH; Staufen, Germany). For lipid
extraction, approx. 1 g of muscle (0.5 g adipose tissue) was
weighed in a tube. Each Precellys-tube contained 20 pieces
of 2.8 mm bulk beads and 2 pieces of 5 mm bulk beads
(Zirconium oxide Precellys beads, Bertin Instruments,
Montigny-le-Bretonneux, France). After addition of 3
ml methanol and nonadecanoic acid (19:0) as an internal
standard, the extracts (in duplicate) were homogenised
3 times at 25 s intervals at 4°C and 6,500 rpm using a
homogeniser (Precellys Evolution, Bertin Instruments).
The homogenates were vortexed and transferred to Pyrex
tubes (Pyrex, Hayes, UK) containing 8 ml of chloroform.
Thereafter, the Precellys-Tubes were washed two times with
1 ml methanol and added to the Pyrex tubes (Töniβen et
al., 2022). Plasma (1.5 ml) was added dropwise to a tube
containing 8 ml of chloroform/methanol (2:1, v/v) with
60 μl C19:0 as an internal standard (60 mg/ml) at room
temperature. The plasma sample preparation was described
in detail elsewhere (Dannenberger et al., 2017). All solvents
used for feed, tissue and plasma lipid extraction contained
0.005% (w/v) of t-butylhydroxytoluene to prevent oxidation
of PUFA.
After filtration, the lipid extracts of tissues and plasma
samples were stored at 5°C for 18 h in the dark and
subsequently washed with 0.02% CaCl
2
solution. The
organic phase was separated and dried with a mixture
of Na
2
SO
4
and K
2
CO
3
(10:1, w/w), and the solvent
was subsequently removed using a vacuum centrifuge
(ScanSpeed 40; LaboGene) at 2,000 rpm/min, 30°C, 30
min. The lipid extracts were redissolved in 300 μl of toluene,
and a 25 mg aliquot was used for methyl ester preparation
(Kalbe et al., 2019). Briefly, for transmethylation, 2 ml of
0.5 M sodium methoxide in methanol were added to the
lipid extracts, which were shaken in a 60°C water bath
for 10 min. Subsequently, 1 ml of 14% boron trifluoride
in methanol was added to the mixture, which was then
shaken for an additional 10 min at 60°C. The FAMEs were
extracted twice with 2 ml of n-hexane and stored at -18°C
until use for high-resolution gas chromatography analysis.
Fatty acid analysis
The fatty acid analysis of all sample lipid extracts was
performed using a capillary gas chromatograph (GC) with
a CP-Sil 88 CB column (100 m × 0.25 mm, Agilent, Santa
Clara, CA, USA) that was installed in a PerkinElmer GC
CLARUS 680 with a flame ionisation detector and split
injection (PerkinElmer Instruments, Waltham, MA, USA)
as described earlier (Dannenberger et al., 2012). Briefly,
hydrogen was used as the carrier gas at a flow rate of 1
ml/min while the split ratio was 1:20, with the injector and
detector were set at 260 and 280°C, respectively. The GC
oven temperature program was 150°C for 5 min; heating
rate of 2°C/min until 200°C and kept for 10 min; heating
rate of 1°C/min until 225°C and kept for 20 min. For the
calibration the reference standard mixture ‘Sigma FAME’
(Sigma-Aldrich, Deisenhofen, Germany), the methyl ester
of C18:1cis-11, C22:5n-3, and C18:2cis-9,trans-11 (Matreya,
State College, PA, USA), C22:4n-6 (Sigma-Aldrich), and
C18:4n-3 (Larodan, Limhamn, Sweden) were used. The
five-point calibration of single fatty acids ranged between
16 and 415 µg/ml and was assessed after GC analysis of
five samples. Fatty acid proportions are presented as % of
total fatty acids or as concentration in mg/100 g tissue.
In addition to composition of FA groups (SFA, MUFA,
PUFA), five nutritional indices (i.e. nutritional value index,
NVI; peroxidability index, PI; atherogenicity index, AI;
thrombogenicity index, TI; and hypocholesterolemic /
hypercholesterolemic ratio, HH) were calculated (Chen
and Liu, 2020; Dabbou et al., 2017; Dal Bosco et al., 2022)
to assess nutritional quality of fatty acid composition in
breast meat only.
Statistical analysis and presentation of the results
The pen was considered as the experimental unit for the
cumulative feed and energy intake variables (n=6), and
for the single-point measurements, birds sampled at the
slaughterhouse (n=96) were considered as the experimental
unit. Data related to the parameters measured on individual
animals, i.e. blood metabolites, meat quality, carcass
characteristics, and FA composition of plasma, breast
meat and abdominal fat were analysed by the general
linear model (PROC GLM) of SAS (SAS Institute, Cary,
NC, USA). The statistical model included fixed effects of
treatment (1-4) and slaughter week (4 and 6), interaction
term, and the blocking effects of room (1-4) and pens (1-6).
The Tukey test was used to separate means and identify
significant differences among treatments. The significance
level was preset at P<0.05, and a tendency was declared at
0.05<P≤0.10. Data for the cumulative intake variables (i.e.
cumulative feed, larvae, feed+larvae, fat and ME intake)
up to d 42 were analysed with the same statistical model
omitting pen effect as pen replaced individual animals
as the experimental unit for these variables. Similarly,
week was omitted from this model as cumulative intakes
corresponded to week 6 only.
A hierarchical two-way clustering analysis was conducted
using JMP statistical software v.10 (SAS Institute) to
investigate patterns in the FAs compositions in three
different tissues in response to the dietary treatments.
Data used for this analysis comprised 43 individual FAs
across plasma, fat and muscle tissues of 96 birds in two
age groups (weeks 4 and 6). Furthermore, alterations in
the composition of FA groups (i.e. SFA, MUFA, PUFA) in
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M.M. Seyedalmoosavi et al.
6 Journal of Insects as Food and Feed ##(##)
response to dietary treatments were visualised using relative
differences of each treatment to the CON.
As no significant interaction effects between the two
main experimental factors (treatment and time) were
encountered, the results were presented in relevant tables
and figures as least square means (LSMEANS) and their
standard errors (SE) as overall differences between relevant
treatment groups, and additional information was provided
in supplementary Tables.
3. Results
Cumulative feed, black soldier fly larvae, fat and energy
intakes
Birds in all BSFL groups fully consumed pre-determined
proportions of offered larvae (data are not shown).
Cumulative intakes over the whole fattening period (42
d) including feed, larvae, feed plus larvae (total), and fat
and ME intakes trough feed and larvae consumption are
presented in Table 2. The L30 group had a lower cumulative
FI compared to CON (P<0.05). Total intake did not differ
among the other groups, while there was a tendency that
L20 birds had a higher intake than CON (P=0.052). Birds
in the L20 and L30 groups had a higher total fat intake than
in the L10 and CON groups (P<0.05). Despite the high fat
intake, L30 had lower total ME intake than CON (P<0.05).
Slaughter weight and carcass characteristics
There was no slaughter weight difference between treatment
groups (Table 3; P>0.05). Incorporation of different levels
of BSFL in the rations did not affect carcass characteristics
of the birds (P>0.05). Dressing percentage, percentages
of breast and wings significantly differed between wk
4 to wk 6 (P<0.001), but no treatment or treatment by
week interaction effect (not shown in Table 3) could be
quantified (P>0.1). Similarly, no treatment or treatment
by week interaction effect could be observed for pH and
color traits of breast meat (P>0.1). Dry matter contents of
breast muscle were higher in L30 than in L10 and CON
birds (P<0.05). In line with this, the CP content of breast
meat was higher in L30 than in L10 and CON (P<0.05).
These differences disappeared when CP contents were
adjusted for DM of the meat tissue (P=0.133).
Plasma metabolites concentrations
The plasma NEFA concentration was higher in L30 and
L20 compared to the CON birds (Table 3; P<0.05). In
addition, birds in L10 tended to have a higher NEFA level
than CON birds (P<0.10). As compared to CON, provision
of BSFL increased plasma triglyceride concentration in all
BSFL groups with a significant increase from L10 to L30
(Table 3; P<0.05). Plasma cholesterol concentration was
not affected by the dietary treatments (Table 3; P>0.05).
No significant treatment × week effect was found for the
plasma metabolites concentrations (P>0.05). Plasma glucose
Table 2. Cumulative intakes (g/bird) over 42 days through feed and defrosted whole black soldier fly larvae (BSFL) consumption
in broilers offered either only regular feed (CON) or increasing levels of BSFL (10-30%) in addition to the regular feed.1
Cumulative intakes, g/bird Dietary treatments2SE P-value3, ≤
CON L10 L20 L30 T
Feed 3,953a† 3,580ab 3,569ab† 3,034b100.0 0.001
BSFL n.a. 382 764 1,145 - -
Feed+BSFL 3,9533,961 4,3324,180 99.6 0.052
Fat (Feed) 157.5a† 142.3ab 141.5ab† 120.4b3.96 0.001
Fat (BSFL) n.a. 33.1 66.2 99.3 - -
Fat (feed+BSFL) 157.5c175.4b207.7a219.7a3.96 0.001
ME (feed), MJ 48.8a† 44.2ab 44.0ab† 37.4b1.23 0.001
ME (BSFL), MJ n.a. 2.0 3.9 5.9 - -
ME (feed+BSFL), MJ 48.8a46.1ab 48.0ab† 43.3b† 1.23 0.027
1 ME = metabolizable energy; n.a. = not applicable; SE = standard error.
2 Dietary treatments: ad libitum feeding without access to BSFL (CON), or with BSFL amounting to 10% (L10), 20% (L20) or 30% (L30) of the feed intake of CON
birds. Total number of observations used for statistical analyses, n=24 (4 treatments each with 6 replicate pens; Number of birds, n=63 per treatment. Data are
presented as LSEMANS and their SE. For the sake of a succinct presentation, only the most conservative (i.e. the largest) SE is presented.
3
T: treatment effect. a-b: Values in a row denoted with different letters differ significantly (Tukey, P<0.05). Symbol † in a row indicates a tendency of two treatments
to differ (Tukey, 0.05<P≤0.10).
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Defrosted whole black soldier fly larvae in broiler diets
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concentration tended be higher in L30 than in L10 (Table
3; P=0.062).
Plasma fatty acids compositions
The composition of fatty acids (g/100 g of total fatty
acids) in plasma of broilers, is presented in Table S2.
The proportion of capric acid (C10:0) in plasma FA was
higher in L30 than in CON birds (Supplementary Table
S2; P<0.05). In addition, the L20 tended to have a higher
proportion of C10:0 than the CON group (P<0.05). The
plasma C12:0 proportion was higher in L30 than those in
L10 and CON birds (P<0.05). Similarly, C12:0 was higher
in L20 than in CON (P<0.05). The L30 and L20 birds had
higher plasma myristic acid (C14:0) proportion than L10
(P<0.05), followed by CON (P<0.05). The total saturated
FA level (∑SFA) was higher in L30 than in L10 and CON
birds (P<0.05). Accordingly, both L20 and L10 groups had
higher ∑SFA levels in plasma as compared to CON (P<0.05).
No significant difference was found among the 4 treatment
groups for the FA proportion of MUFA in plasma (P>0.05).
Plasma levels of linoleic acid (C18:2n-6), α-linolenic acid
(C18:3n-3), total n-3 PUFA (∑n-3), and total n-6 PUFA (∑n-
6) were lower in L30 than in CON (P<0.05). The proportion
of the conjugated linoleic acid (CLA) C18:2cis-9, trans-11
Table 3. Effects of increasing levels of defrosted whole black soldier fly larvae (BSFL) in broiler diets on carcass characteristics
and breast meat quality traits, as well as on blood metabolites of broilers1.
Dietary treatments2SE P-values3, ≤
CON L10 L20 L30 T W
Slaughter weight, g 1,821 1,732 1,971 1,817 114.5 0.602 0.001
Hot carcass weight, g 1,169 1,096 1,254 1,143 82.1 0.628 0.001
Breast weight, g 339 326 365 337 30.1 0.859 0.001
Breast DM, % 23.2b23.2b23.9ab 24.2a0.22 0.006 0.077
% of carcass weight
Dressing percentage 63.0 62.3 62.2 62.5 0.77 0.848 0.001
Breast 27.8 28.2 27.6 29.0 0.78 0.571 0.001
Legs 33.4 33.4 33.4 32.9 0.49 0.806 0.258
Wings 10.7 10.8 10.5 10.5 0.20 0.736 0.001
Abdominal fat 2.1 2.0 2.0 2.4 0.48 0.885 0.318
Breast meat quality parameters
Water holding capacity, % 40.7 39.5 39.7 38.7 1.01 0.534 0.001
pH at 30 min 6.4 6.4 6.3 6.4 0.03 0.384 0.001
pH at 45 min 6.2 6.2 6.2 6.2 0.03 0.791 0.002
pH at 24 h 5.5 5.5 5.5 5.5 0.02 0.484 0.512
L* (lightness) 55.3 54.8 54.2 53.8 0.63 0.409 0.605
a* (redness) 3.3 3.3 3.1 3.2 0.25 0.923 0.659
b* (yellowness) 4.7 4.7 4.9 4.8 0.32 0.982 0.001
C* (Chroma) 5.9 5.9 5.9 5.9 0.30 0.999 0.001
H* (Hue) 53.7 53.1 56.2 56.3 2.68 0.801 0.001
Crude protein, % FM 20.8b20.6b21.4ab 21.7a0.23 0.002 0.315
Crude protein, % DM 89.4 88.8 89.4 90.0 0.36 0.133 0.160
Blood metabolites
NEFA, mM 132.9c† 192.8bc† 271.6ab 308.8a19.5 0.001 0.667
Triglyceride, mM 1.04c1.40b1.59ab 1.81a0.101 0.001 0.630
Cholesterol, mM 3.04 3.17 3.26 3.33 0.109 0.272 0.002
Glucose, mM 13.86 13.7614.03 14.450.197 0.062 0.972
1 NEFA = non-esterified fatty acid; FM = fresh matter; DM = dry matter; SE = standard error.
2 Dietary treatments: ad libitum feeding without access to BSFL (CON), or with BSFL amounting to 10% (L10), 20% (L20) or 30% (L30) of the feed intake of CON
birds. Total number of observations used for statistical analyses, n=96 (i.e. 2 birds sampled from each of 6 pens allocated to each of 4 treatments at weeks 4 and 6).
3 T: treatment effect; W = time effect (weeks 4 and 6). Symbol † in a row indicates a tendency of two treatments to differ (Tukey, 0.05<P≤0.10). P-values for T×W
= treatment by time interaction were all P>0.1, and are not shown. Data are presented as LSEMANS and their SE, and the values refer to average of weeks 4 and
6 as there was no significant T×W interaction. For the sake of a succinct presentation, only the most conservative (i.e. the largest) SE is presented. Values in a
row denoted with different letters differ significantly (Tukey, P<0.05).
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FA increased in the plasma lipids of larvae fed birds in
response to increasing levels of BSFL in contrast to CON
(P<0.05). The eicosapentaenoic acid (EPA) was lower
in plasma samples of L30 an L20 than in L10 and CON
(P<0.05). However, proportions of docosapentaenoic acid
(DPA) and docosahexaenoic acid (DHA) were not affected
by the dietary groups (P>0.05). Similarly, the n-6/n-3 ratio
did not differ among the groups (P>0.05; Table S2). No
treatment × week interaction effect was found for individual
or groups of FA in plasma (P>0.05).
Breast muscle fatty acids compositions
The proportions of fatty acids with relatively high
concentrations in the breast muscle lipids of broilers are
presented in Supplementary Table S3. The proportion of
C10:0 in breast muscle total FA was higher in L30 than in
the other groups (Supplementary Table S3; P<0.05). With
an increasing BSFL level in the diet, the proportions of
C12:0 and C14:0 increased linearly (P<0.05), so that higher
proportions of C12:0 and C14:0 in the breast muscle total
FA in L30 birds than in L20 followed by L10 and CON
were observed (P<0.05). Provision of BSFL at 30% of FI
(L30) increased ∑SFA in breast muscle as compared to
CON (P<0.05), whereas ∑MUFA levels were not affected
by the provision of BSFL (P>0.05). Furthermore, there was
no difference between treatment groups for total PUFA
and ∑n-6 levels (P>0.05), while ∑n-3 FA were reduced
with increasing BSFL levels (P<0.05). Although C18:3n-6
and C18:3n-3 proportions were reduced in response to
increasing levels of BSFL, the C18:2cis-9, trans-11 fractions
in breast muscle total FA were significantly increased in
treatment group L10 to L30 as opposed to CON (P<0.05).
The L30 had a higher n-6/n-3 ratio compared to CON
(P<0.05). Supplementation with BSFL at any of the three
levels did not influence NVI of breast muscle (P>0.05;
Supplementary Table S3), whereas the L30 led to a lower
PI than those in CON (P<0.05). Increasing BSFL levels in
the diet resulted in increasing AI and TI indices, with L30
having the highest impact (P<0.05), while the opposite was
the case for the HH ratio (P<0.05). No treatment × week
interaction was found for breast muscle FA proportions,
and the nutritional indices (P>0.05).
Fatty acids compositions in abdominal fat tissue
The proportions of fatty acids with relatively high
concentrations in the abdominal fat tissue of broilers are
presented in Supplementary Table S4. The FA analysis
of abdominal tissue showed that C10:0, C12:0 and C14:0
fractions increased in the BSFL-fed treatment groups in a
dose dependent manner, which resulted in higher ∑SFA levels
in BSFL-fed groups than those in CON (Table S4, P<0.05).
Broilers fed BSFL had lower C18:1cis-9 and C18:1cis-11
fractions in abdominal fat (P<0.05), which caused a higher
percentage of ∑MUFA in L30 and L20 than in CON groups
(P<0.05). The ∑PUFA, ∑n-3, and ∑n-6 levels were reduced
in response to feeding increasing levels of BSFL (P<0.05).
Among PUFAs, C18:2n-6, γ linolenic acid (C18:3n-6) and
C18:3n-3 proportions decreased with increasing BSFL level
(P<0.05). However, the C18:2 cis-9, trans-11 FA fraction
was significantly increased in the abdominal fat in broilers
which received BSFL compared to those in the CON group
(P<0.05). The L30 treatment had a lower proportion of DPA
than in CON and L20 (P<0.05), however, EPA and DHA
were not affected by the dietary treatments (P>0.05). The
n-6/n-3 ratio did not differ among the treatment groups
(P>0.05). No treatment × week interaction effect was found
for abdominal fat tissue FAs (P>0.05).
Overall changes in fatty acid compositions across
different tissues in response to feeding black soldier fly
larvae
Changes in relative abundance of FA groups in BSFL-fed
groups as compared to CON in abdominal fat (A), plasma
lipids (B) and breast muscle (C) at wk 4 (Supplementary
Figure S1) and wk 6 (Figure 1) were linearly influenced by
increasing BSFL levels in the ration. These results collectively
indicate that as BSFL level increased in the ration, total SFA
proportions increased at the expense of ∑MUFA, ∑PUFA,
∑n-3 and ∑n-6 in all three tissues. The alterations were more
pronounced in abdominal fat tissue followed by breast muscle
and plasma tissues (Figure 1).
A two-way hierarchical analysis was conducted to investigate
the FAs compositions of different groups across different
tissues (Figure 2). As shown with intensifying red and blue
colours in the heatmap of the dendrogram, clustering of
different variables showed dependency on the treatment
groups across three tissues. Our results showed a distinct
clustering of the animals from L20 and L30 groups, which
had higher SFA levels, whereas CON stood either alone or
in most cases clustered together with L10. On the left side
of the heatmap, there is a clear clustering for SFA, indicating
that L10 and CON had the lowest SFA contents, and by
scrolling down the heatmap, higher SFA contents in L20 and
L30 can be observed. At the bottom of the heatmap and on
the right side, unsaturated fatty acids appear, and in some
places they lead to a smaller clustering. Furthermore, at the
mid-top of the heatmap, a clear grouping is shown in red,
which indicates the presence of more unsaturated fatty acids
in the CON and L10 groups, and at the bottom, this grouping
can be detected in blue, which indicates lesser proportions
of unsaturated fatty acids in L30 and L20 groups.
4. Discussion
We investigated the effect of feeding defrosted whole
BSFL to broilers at increasing dietary levels on carcass
characteristics, blood metabolites, and FA composition
in plasma and breast muscle lipids and abdominal fat. We
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Defrosted whole black soldier fly larvae in broiler diets
Journal of Insects as Food and Feed ##(##) 9
-10
0
10
20
-10
0
10
20
L10 vs CON
L20 vs CON
L30 vs CON
-10
0
10
20
∑ SFA ∑ MUFA ∑ PUFA ∑ n-3 n-6
Abdominal fat
Plasma
Breast muscle
Fatty acid abundance in relation to control, % of total FA
Figure 1. Relative changes in fatty acid (FA) groups abundance in plasma lipid, breast muscle and abdominal fat of broilers fed
increasing dietary levels of black soldier fly larvae compared with broilers in the CON group without access to larvae. n=48 (i.e.
2 birds sampled from each of 6 pens allocated to each of the 4 treatment groups at week 6). SFA = saturated fatty acids; MUFA =
monounsaturated fatty acids; PUFA = polyunsaturated fatty acids; n-3, n-6 = n-3 and n-6 fatty acids.
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hypothesised that increasing levels of defrosted whole
BSFL can be used in broiler diets with no adverse effects on
meat quality, metabolism, and FA compositions of plasma,
muscle and depot fat tissues. Our results indicated that up
to 30% inclusion of defrosted whole BSFL in broiler diets did
not negatively affect carcass characteristics, water holding
capacity, meat pH and meat colour of broilers. However,
inclusion of 30% defrosted whole BSFL in broiler diets
increased plasma NEFA and TG concentrations. In addition,
the FA compositions in plasma lipids, breast muscle and
abdominal fat of broilers fed 30% BSFL showed increased
SFA levels at the expense of MUFA and PUFA. Increasing
dietary levels of BSFL caused a dose dependent increase of
the CLA fraction (C18:2 cis9, trans11) in total FA of plasma,
muscle and fat tissues. However, lower proportions of ∑n-3
FA in BSFL than in the age-specific diets was reflected by an
increased n-6/n-3 FA ratio in the L30 group. In the following,
we discuss possible mechanisms affecting changes in carcass
traits, plasma metabolites, and FA composition in plasma,
muscle and fat tissues in BSFL-fed broilers.
Slaughter weight and carcass characteristics
We could not find indications that the inclusion of
defrosted whole BSFL up to 30% in broiler diets affected
meat quality and carcass traits. In line with our results, it
has been reported that using up to 20% full-fat BSFL in
the diet did not influence key meat characteristics and
broiler performance (De Souza Vilela et al., 2021a,b).
Similarly, Cullere et al. (2019) reported that breast and
leg physical meat quality and nutritional composition
remained substantially unaffected in broilers that received
diets in which 50% or 100% of the soybean oil was replaced
with BSFL fat. However, Murawska et al. (2021) found
Figure 2. Two-way hierarchical cluster analysis of fatty acid proportions in plasma lipid, breast muscle and abdominal fat of
broilers fed increasing dietary levels of black soldier fly larvae. The heat map is based on concentrations of fatty acids (FAs)
(x-axis) in plasma, breast muscle and abdominal tissues across 4 experimental groups (y-axis). The clusters are presented on the
respective opposite axes, overlapping clusters due to similarity in diets and the resulting fatty acid proportions are then shown
with similar colour regions on the colour-map. The deeper red represents the higher concentration and the deeper blue represents
the lower concentration. The first left-column of the heatmap with 4 different colours indicates groups to which individual birds,
each shown on a single line, belong to.
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Journal of Insects as Food and Feed ##(##) 11
that replacement of soybean meal with high levels (75 or
100%) of full-fat BSFL meal in broiler diets compromised
growth performance and carcass traits (lower juiciness
and taste intensity). We observed that despite high levels
of fat in BSFL, abdominal fat (% of carcass weight) did not
significantly differ among the groups. Despite the alterations
of the fatty acid composition in abdominal fat tissue, the
abdominal fat percentage in the carcass was similar among
groups. However, despite the higher fat intake, L30 had a
lower metabolizable energy intake. Lower ME intake in
L30 birds may explain the lack of a significant difference in
abdominal fat content in the carcass between the groups.
The consumed fat had a high proportion of MCFA which
can be utilised as a rapid energy source, improves energy
availability and reduces the deposition of adipose tissue
(Li et al., 2016). Therefore, the lack of differences in the
amount of abdominal fat depots observed in this study
could be attributed to the lower ME intake in L30 birds
and the high MCFA content of BSFL. Lower ME intake
observed in the L30 group could be due to the overall drop
in feed intake over all treatment groups which was most
pronounced at the highest inclusion level (i.e. 30%). Lower
feed intake in L30 could be attributed to the high dietary fat
content which might affect the response of hypothalamic
appetite-related peptides (Obrosova et al., 2007; Wang et
al., 2017b), and might consequently cause the reduced feed
intake. Moreover, the physical constrains in the digestive
tract of broilers might negatively affect feed intake of birds
(Brickett et al., 2007; Dozier III et al., 2006).
Blood metabolites concentrations
Plasma triglycerides concentrations increased linearly with
increasing levels of BSFL intake, whereas a higher plasma
NEFA concentration in L30 compared to L10 and the CON
groups was observed. High plasma NEFA concentrations
in the L30 group can be explained by either a high lipolysis
rate due to a low dietary energy intake or a high intake of
dietary fat (Wang et al., 2017a). Birds of the L30 group
showed a lower ME intake which might be associated to
higher plasma NEFA levels. Although we did not measure
serum insulin concentrations, previous studies in chicken
(Crespo and Esteve-Garcia, 2003) and humans (Lee et al.,
2006) showed that increased intake of n-6 FA and SFA is
positively associated with insulin resistance (Sears and
Perry, 2015). Plasma cholesterol concentration remained
unaffected by the inclusion of BSFL in the ration. This is in
line with results of Kim et al. (2020) that diets containing
5% of coconut oil or BSFL oil did not change the serum
cholesterol levels of broilers compared to those of birds
fed diets contained 5% of corn oil. It has been shown that
increasing intakes of saturated fat (such as lard) can be
associated with higher plasma triglyceride and cholesterol
levels in chicken (Peña-Saldarriaga et al., 2020; Velasco
et al., 2010). This might indicate that the higher intake
of dietary fat/SFA in the L30 group (219.7/92.5 g over 42
d) compared to CON was not high enough to trigger an
increase in plasma cholesterol. This could be attributed to
the chitin content of BSFL, which might attract negatively-
charged bile acids and free FA and thus make them less
available for cholesterol synthesis in the liver (Secci et al.,
2018).
Fatty acid compositions in different tissues
It is well known that the FA composition of muscle and
adipose tissues in broilers can be modified by dietary factors
(Bostami et al., 2017; Kim et al., 2020; Semwogerere et al.,
2019). In this experiment, FA analysis of BSFL revealed a
high SFA (71%) and low PUFA content (i.e. approximately
12%). Plasma, breast muscle and abdominal fat FA profiles
were similar to that of the FA profile of BSFL. Studies on
the effect of feeding BSFL on FA composition in chicken
meat are scarce. Schiavone et al. (2019) reported that using
defatted BSFL-meal in broiler diets caused remarkable
differences in the FA composition, with an increased MUFA
content at the expense of PUFA. Also, De Souza Vilela et al.
(2021a) reported that feeding up to 20% dried full-fat BSFL
to broilers reduced total PUFA levels, while an increase in
EPA was observed, coupled with an increase in total SFA
and, in particular the C12:0 fraction. In the present study we
observed an increased SFA level in plasma, breast muscle
and adipose tissue and a reduced PUFA level in plasma and
adipose tissue of the broilers fed BSFL, which is probably
due to the carry-over effect from BSFL fat. In addition to
carry-over effects of BSFL, we assume that biochemical
mechanisms may be involved in the alteration of FAs profile.
In chickens, C18:3 n-3 and C18:2 n-6 are essential FA and
precursors for long chain PUFA synthesis (Cherian, 2015)
with EPA (20:5 n-3), docosapentaenoic acid (DPA; 22:5
n-3), and docosahexaenoic acid (DHA; 22:6 n-3) derived
from C18:3 n-3 and arachidonic acid (20:4 n-6) derived
from C18:2 n-6, respectively (Murff and Edwards, 2014).
In our study, the lower plasma C18:3 n-3 and C18:2 n-6
proportions in L30 birds reflects their lower dietary intake,
and might also be associated with the lower ∑PUFA level
in abdominal fat. However, we did not find this effect in
breast muscle tissue which could be due to the low fat
content of the breast muscle. The conversion of C18:3 n-3
and C18:2 n-6 into long-chain PUFAs is carried out by Δ-6
desaturase, Δ-5 desaturase and elongases (Gonzalez-Soto
and Mutch, 2021). It has been shown that rats fed a diet
supplemented with SFA (20% w:w partially hydrogenated
coconut oil) had a reduced Δ-5 desaturase activity compared
with a non-purified diet (5% w:w fat) (Dang et al., 1989). In
addition, Valenzuela et al. (2017) reported that a very high
fat diet (i.e. 60% fat) decreased the activity of Δ-5 and Δ-6
desaturase and PUFA accretion in liver and other tissues of
mice. Whether this mechanism is also effective in poultry
at a much lower fat and SFA intake needs to be clarified.
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We found a higher n-6/n-3 ratio in breast muscle of L30
birds, which is likely due to the many fold higher n-6/n-3
ratio in BSFL than in age-specific diets (Table S1). However,
we could not detect this difference in plasma and abdominal
fat tissues. An increased n-6/n-3 ratio larger than 5 in
breast muscle is undesirable for human consumption (De
Souza Vilela et al., 2021a). However, the n-6/n-3 ratio of
the broiler breast meat in our study was far below this
threshold so that the consumption of broiler meat produced
with up to 30% BSFL in the ration seems unlikely to be
harmful to health in this respect. Except for the NVI, the
indices assessing nutritional quality of FA composition in
the breast muscle indicated significant and mostly linear
changes in lipid quality as response to increasing levels of
BSLF in the diet. As compared to control diet, L30 reduced
PI, an index assessing the lipid peroxidation susceptibility
in breast muscle tissue (Henriques et al., 2015), which
may be considered as a positive effect, but increased AI
and TI. The AI represents the relationship between the
sum of the main SFA which favours the adhesion of lipids
to cells of the immunological and circulatory system and
that of the main classes of unsaturated FA inhibiting
the accumulation of plaque and reduces the levels of
esterified fatty acid, cholesterol, and phospholipids. The
TI characterises the thrombogenic potential of FA, showing
the tendency to form clots in the blood vessels. Therefore,
consumption of foods with a lower AI and TI are associated
with reduced levels of circulating total cholesterol and
associated cardiovascular disorders (Chen and Liu, 2020).
The HH ratio is an indication of the relationship between
hypocholesterolemic FA (cis-C18:1 and PUFA) and
hypercholesterolemic FA (Chen and Liu, 2020). Despite
the favourable PI, the higher AI and TI and lower HH in
breast muscle of broilers fed with a high amount of larvae
in their diets (i.e. 30%) might limit nutritional quality of the
lipids in the breast meat for human consumption.
As confirmed in this study, fat of BSFL contains a high
content of SFA with a considerable amount of MCFA,
particularly C12:0 (De Souza Vilela et al., 2021a). The
antimicrobial effects of MCFA such as C12:0 are well studied
(Kabara et al., 1972). Likewise, it has been suggested that
C12:0 induces immuno-regulatory functions and alleviates
inflammation during infections in broilers (Wu et al., 2021).
Whether an antimicrobial effect of C12:0 is relevant in
our study cannot be determined from the available data.
However, it should be taken into account that a high
intake of SFA is positively related to plasma cholesterol
and cardiovascular disease in humans (EFSA Panel on
Dietetic Products and Allergies, 2010). Considering that
the SFA especially in breast muscle is higher in the L30
treatment group, high inclusion levels of BSFL (i.e. 30%) in
broiler diets seem to result in unfavourable FA composition
in chicken meat for human consumption.
Our results showed that C18:2 cis-9, trans-11 FA, a CLA
isomer, was higher in BSFL (0.50% of total FAs) than in
the soybean-grain based feed, which was associated to
an increasing proportion of CLA in total FA of plasma,
breast muscle and abdominal fat in response to increasing
levels of dietary BSFL intake. The occurrence of CLA in
BSFL was previously reported by Hoc et al. (2020). Meat of
monogastric farmed animals is a poor source of CLA (0.1
to 0.2% of total FA) (Chin et al., 1992), and enrichment of
poultry meat with CLA has been of interest for humans
consumers (Den Hartigh, 2019; Lehnen et al., 2015).
Although the proportion of CLA in BSFL-fed broilers’ meat
was not dramatically increased in our study, it indicates the
potential of feeding BSFL to modulate the CLA content in
broiler meats. Dietary CLA can be readily incorporated in
broiler muscle (Du and Ahn, 2002); therefore, we assume
that the increased CLA level in BSFL-fed broilers is due to
a carry-over effect from BSFL. The results of the current
study suggest that inclusion of whole BSFL in broiler diets
up to 20% of the voluntary FI did not compromise the meat
quality traits including FA composition. However, higher
inclusion rates (i.e. 30%) are not recommended as it might
be associated with negative effects on FA composition in
plasma, breast muscle and adipose tissues which reduces
the nutritional quality of meat for human consumption.
5. Conclusions
The current study provides concrete information on the
consequences of different levels of feeding un-processed
whole BSFL to broilers. Dietary inclusion of defrosted BSFL
did not influence slaughter weight, meat quality and carcass
characteristics, suggesting that BSFL can be included in
broiler diets without compromising consumer acceptance.
With an inclusion level of larvae (30%) we however observed
pronounced alterations in the FA compositions of plasma,
breast and abdominal fat resulting in an increased SFA
content at the expense of MUFA and PUFA. Increasing
dietary levels of BSFL resulted in a dose dependent increase
in the proportion of the CLA isomer C18:2 cis-9, trans-11
in total FA of plasma, muscle and fat tissues. Although the
n-6/n-3 ratio observed in breast muscle of L30 birds was
higher than in CON birds, it is still far below the threshold
ratio of <5 considered favourable for human health. The
overall nutritional quality of fats in the breast meat for
human consumption may decrease linearly in response
to increasing dietary levels of BSFL, particularly at the
highest inclusion level (i.e. 30%). In conclusion, up to 20% of
defrosted whole BSFL could be used in broiler diets without
considerably negative effects on the birds’ metabolism,
slaughter weight and carcass traits, and fatty acid profile
in plasma, muscle and fat tissues.
Please cite this article as 'in press' Journal of Insects as Food and Feed
https://www.wageningenacademic.com/doi/pdf/10.3920/JIFF2022.0125 - Friday, December 02, 2022 4:39:44 AM - Leibniz-Institut für die Biologie landwirtschaftlicher Nutztiere IP Address:195.37.182.216
Defrosted whole black soldier fly larvae in broiler diets
Journal of Insects as Food and Feed ##(##) 13
Supplementary material
Supplementary material can be found online at https://doi.
org/10.3920/JIFF2022.0125
Table S1. Fatty acids compositions of the age-specific
broiler diets and black soldier fly larvae (BSFL) offered
to broilers.
Table S2. Effects of increasing levels of defrosted whole
black soldier fly larvae in broiler diets on plasma fatty acid
compositions of broilers.
Table S3. Effects of increasing levels of defrosted whole
black soldier fly larvae in broiler diets on breast muscle
fatty acid compositions of broilers.
Table S4. Effects of increasing levels of defrosted whole
black soldier fly larvae in broiler diets on abdominal tissue
fatty acid compositions of broilers.
Figure S1. Relative changes in fatty acid abundance in
plasma lipid, breast muscle and abdominal fat of broilers fed
increasing dietary levels of black soldier fly larvae compared
with broilers in the CON group without access to larvae.
Raw data availability: Raw data used in this study are
available at https://doi.org/10.5281/zenodo.7347351
Authors’ contributions
CCM and GD conceived and designed the study. PW and
CCM acquired funding. MMS and GD contributed to
rearing and sampling of the chickens. MMS, GD, SG, and
DD contributed to the analyses of the sample and collection
of data. MMS and GD performed statistical analysis of the
data and drafted the manuscript. CCM, GD, MMS, MM,
SG, DD, SM and PW reviewed the manuscript. GD had the
final responsibility for the content of the manuscript. All
authors read the article and approved the submitted version.
Acknowledgements
This research project is part of the Leibniz Science Campus
Phosphorus Research Rostock and is co-funded by the
funding line strategic networks of the Leibniz Association.
The authors are deeply grateful to F. Feldt and co-workers in
the FBN slaughterhouse, as well as B. Mielenz, K. Kàrpàti,
U. Lüdtke, E. Wünsche, K. Wilke, R. Gaeth and S. Dwars
for their support of various laboratory analyses and with
sample collection. The authors wish to thank Birgit Jentz
and Maria Dahm in the Institute of Muscle Biology and
Growth who assisted with sample preparation and GC
analysis of fatty acids. The open access publication of this
article was funded by the Open Access Fund of the Research
Institute for Farm Animal Biology (FBN).
Conflict of interest
The authors declare no conflict of interest.
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Please cite this article as 'in press' Journal of Insects as Food and Feed
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