1. Introduction
Soybean meal (SBM) is the main dietary protein source in poultry diets, however, the price of SBM has been soaring. For this reason, alternative dietary protein sources with comparable protein and amino acid content are needed to cope with the limited supply of soybean and increased demand for human consumption [
1,
2]. Moreover, managing organic waste by the human population is also a major issue [
2]. Organic waste can be decomposed by
Hermetia illucens larvae (HIL) and recycled to dietary protein and fat sources [
3,
4,
5]. However, the potential risk of heavy metal accumulation in the larval body from contaminated rearing substrates should be monitored [
6].
HIL has the potential to be an ingredient in poultry diets and HIL meal (HILM) has been investigated as an alternative protein source in poultry diets [
1,
7] as it contains similar or higher valuable protein and amino acid content than SBM [
7,
8]. However, with increased inclusion of HILM in broilers, chitin in the exoskeleton of larvae negatively impacted crude protein digestibility, and the growth performance decreased linearly, showing that low inclusion level of HILM was appropriate for diets [
9,
10]. Furthermore, the high level of inclusion of HILM showed negative effects on laying performances such as lay percentage, feed intake, egg mass, and feed conversion ratio (FCR) [
11,
12]. In contrast, chitin has been reported to have a positive effect by increasing cecal volatile fatty acid (VFA) concentration and decreasing blood cholesterol level [
13,
14]. Thus, proper ingestion levels of chitin in the HILM should be considered.
In previous studies, the focus was on the effect of HILM on the fatty acid (FA) profile of animal products such as meats and eggs [
7,
13,
14]. In comparison to the FA profile of SBM, HILM contains high levels of saturated fatty acid (SFA) and low levels of polyunsaturated fatty acid (PUFA) [
13]. It is reported that the FA profile differences between HILM and SBM are reflected in chicken meats and eggs [
7,
15,
16]. Furthermore, a positive effect of pigment in HILM on yolk color was observed, making it appear redder [
7,
15].
Apart from good nutrient composition and chitin effects, the drying process is also important for the chemical composition of nutrients [
17]. Particularly, the microwave drying method, which is commonly used for food processing, manages moisture content in the insect larvae and produces time and energy-efficient products [
17,
18]. Nevertheless, the effects of microwave-dried HILM on the diets of laying hens are not yet clear. In this study, SBM was replaced with microwave-dried HILM as a dietary protein source in the diets of laying hens to evaluate productive performance, cecal VFA profile, egg quality, FA profile, and heavy metal residue in the eggs.
4. Discussion
Appropriate energy and protein requirements are necessary for optimal egg production without increasing BW and plumpness during the laying period. In particular, an optimal level of crude protein is important to maintain growth and maximize productive performance in poultry [
30]. Hence, to be considered as a suitable protein ingredient in poultry, protein digestibility is an important factor [
14]. Although insects are a natural diet of poultry and HIL, being a high protein and amino acid source, is comparable to SBM [
3,
31], the exoskeletal chitin of larvae can negatively affect the protein digestibility [
11,
32].
Despite the nutritional concerns of HILM, our results showed that the dietary inclusion of HILM did not impair the BW changes during the experimental period. In laying quail diets, the addition of HILM up to 15% did not affect growth performance [
7,
31]. Laying pullets that fed on a 7.5% inclusion level of HILM during the experimental period (19 to 27 weeks of age) showed higher BW at 27 weeks [
12] and those that fed on a 15% inclusion level of HILM during the experimental period (28 to 43 weeks of age) had higher BW at 43 weeks [
33]. However, total replacement of SBM with HILM in the diets of Lohmann Brown Classic laying hens (24 to 45 weeks of age) decreased weight gain [
11]. In contrast, in laying hens (Hy-line Brown), the inclusion levels (7.3% and 14.6%) of HILM did not show the difference in BW at 40 weeks [
14]. In this study, laying hens that fed on HILM showed no negative effects of weight gain during the experimental period and the different BW results might be due to different inclusion levels of insect meal, experimental periods, species, or ages.
The productive performance of laying hens fed with HILM showed that there were no negative effects on FCR results; however, the lay percentage decreased in the 4HILM group. It was reported in a previous study that total replacement of SBM with HILM in a laying hen diet decreased laying performance by decreasing lay percentage, egg weight, and egg mass [
11]. In contrast, it was reported that soybean cake could be replaced with HILM in layer diets without negative effects on productive performance and health [
34]. The contrasting results can be attributed to different starting ages (24 and 64 weeks of age, respectively), experimental periods (21 and 10 weeks, respectively), or the dark coloration of insect meal diets [
11]. It is suggested that the lower feed intake was ascribed to the darker color of the HILM diet than that of the SBM diet [
11]. In another study, it is suggested that SBM can be substituted with microwave-dried HILM (<7%) in broiler diets [
21]. The suggested substitution level (7%) of the microwave-dried HILM was lower than the results (<10% in broiler diets), which were reported in previous studies [
10,
16]. In laying hens (19 to 27 weeks of age), it has been reported that a 7.5% inclusion level of HILM showed negative effects on FCR [
12]. Moreover, the FCR of older laying hens (28 to 43 weeks of age) that fed on HILM (0–15%) was also high [
33]. In this study, FCR was not affected by dietary treatments and this might be due to low inclusion levels of HILM and the short experimental period. Therefore, the low inclusion level of microwave-dried HILM seems to be appropriate for productive performance in younger laying hens.
The positive effects of chitin on gut health were confirmed by a linear increase in cecal BCFA level and a tendency to a higher total SCFA in the 4HILM group. In this study, total SCFA tended to increase with the inclusion level of HILM and increased by about 54% in the 4HILM group. Similar to our result, Cutrignelli et al. [
35] showed increased total VFA concentration (>36.8%) in HILM-fed broilers. In a previous study, the cecal content of butyrate in laying hens also increased with HILM [
35], in contrast, the concentration of butyrate was not affected in our result. Moreover, the concentration of the total BCFA in the 4HILM group drastically increased (approx. 89%) than in the CON group. Although the reason is unclear, it could be ascribed to different chemical compositions of the microwave-dried HILM compared to previous studies that used a different manufacturing process. The alteration of the chemical structure of the protein particle in the microwave-dried HILM might have contributed to lower protein digestibility [
36]. The microwave drying method makes the protein particle more compact by polymerization reaction [
36]. Furthermore, the characteristics of chitin combined with other nutrients in insect meals render it difficult to digest [
9,
14]. In this study, laying hens on 2HILM and 4HILM feeds ingested approximately 0.14 and 0.29 g/d of chitin, respectively, according to the method by Marono et al. [
9]. Increasing the ingestion level of chitin could affect digestibility, and therefore appropriate inclusion level of HILM should be considered for protein digestibility [
14]. Hence, proteolytic fermentation of the undigested protein in the cecum may have affected the concentrations of the BCFAs [
10]. The increase in the intestinal length of layers fed insect meal by a compensatory mechanism for increasing nutrient absorption surface and digestibility efficiency corroborates the availability of proteolytic fermentation in the cecum [
14,
37].
In this study, the egg quality parameters except for the yolk color were not different among the treatment groups. In a previous study, the eggshell thickness was increased by the inclusion levels (5% and 7.5%) of HILM [
12]. A similar result was reported in laying quails’ eggs where the physical parameters such as shell thickness improved [
7]. It was also suggested that the hindgut fermentation of chitin may have contributed to an increase in mineral absorption such as calcium [
12]. However, in our study, differences in physical traits of eggs were not observed and the results were due to low chitin ingestion levels in the 2HILM and 4HILM groups. The noticeable alteration was increased intensity of yolk color in the 4HILM group and a common result was reported in previous studies [
7,
12,
15]. The alteration of yolk color is related to carotenoids such as β-carotene and lutein in the HILM affecting the yolk color [
7,
15]. In a previous study, total carotenoid content was 2.15 mg/kg in the HIL [
15]. Furthermore, 5% inclusion level of HILM in laying pullet diets improved yolk color [
12]. These results indicate that eggs from laying hens that were fed HILM can contribute to consumer acceptability by enhancing the yolk color.
The proximate composition of eggs from laying hens fed on HILM did not differ among dietary treatments and suggests that it can be considered as a feed ingredient. Secci et al. [
15] also reported that total replacement (inclusion level of 17%) of SBM with HILM had no negative effect on the proximate composition of eggs in laying hens. In contrast, the protein content of eggs from laying quails fed 15% HILM in a diet was decreased [
7]. They suggested that the negative effect of chitin on nutrient digestibility resulting in lower protein availability could have affected the protein content in the eggs [
7]. Furthermore, in line with the finding of Secci et al. [
15], we also observed that cholesterol levels tended to decrease in eggs from laying hens fed on HILM. This reduction in cholesterol level can be ascribed to lower serum cholesterol in laying hens that fed on HILM, in addition to chitin contributing to decreased lipid absorption by binding to the lipids and fatty acids [
11,
14]. Furthermore, the bile acid-binding capacity of the chitin could inhibit bile reabsorption and enhance cholesterol excretion by hypocholesterolemia, resulting in lower cholesterol levels in the blood [
38,
39]. However, cholesterol levels in laying quails’ eggs and growing quail breast meats were not affected by the HILM [
7,
40]. This discrepancy might be due to different species.
The FA profiles of the larvae depend on their rearing system and the alteration of FA profiles in laying hen eggs reflected the FA profiles of the HILM. In this study, we observed increased total MUFA and decreased total PUFA. A similar result was also reported, where 10% and 15% inclusion levels of the HILM in laying quails’ diets increased total MUFA and decreased total PUFA in eggs, and increased total SFA content on the HILM fed groups [
7]. However, in our findings, total SFA content was not affected by dietary treatments and the discrepancy meant that inclusion levels in the 2HILM and 4HILM groups were too low to change the SFA content in the eggs. Moreover, the increase of the total MUFA content was associated with the SFA content and was contributed by elongation and desaturation of SFA to the MUFA [
7,
41]. There was also a reduction in important precursors (linoleic and linolenic acids) with inclusion levels of the HILM, which led to decreased
n-6 and
n-3 FAs such as DHA and eicosapentaenoic acid in the eggs [
7,
42].
Heavy metal concentrations in the HILM and animal products have to be monitored due to the residues of undesirable substances. The detrimental heavy metals can be accumulated in the HIL body from their contaminated feeding media [
5,
43,
44]. In our trial, undesirable substances such as F, As, Pb, Hg, and Cd in the HILM were under permissible limits [
26]. Furthermore, the hazardous heavy metals in the eggs did not exceed the permissible limits [
27,
28,
29]. Although the concentrations of Mg and Zn increased in the 4HILM group compared to the CON group, they were not applicable for restriction. These results indicate that in terms of safety, the HILM is suitable as a feed ingredient for animal diets.