3.1. Amino Acid Composition of Larval Saliva
One of a few examples for the application of wasp materials to food is the development of a supplementary beverage for physical exercise inspired from the amino acid composition of
V. mandarinia larval saliva [
24]. The beverage products do not contain the actual larval saliva of
V. mandarinia, but an amino acid mixture composed of the same amino acid composition of
V. mandarinia larval saliva. The products were developed on the basis of a series of investigations on exercise endurance enhancement effects by supplementation of the amino acid mixture of which the composition is the same as the larval saliva of
V. mandarinia [
25,
26,
27]. This idea has come from trophallaxis, a unique way of survival observed in social wasps. Trophallaxis is common among wasps; carnivorous larvae which feed on meat pellets prepared by adult workers, while larvae produce and pass an oral exudate to adult workers in response [
27,
28]. For social wasps, larval saliva is considered a source of energy and motivation for adult workers that fly a distance of about 100 km daily only to prepare meat pellets for larvae [
29]. Takashi and colleagues reported the composition of amino acids in the larval saliva from five species of hornets including
Vespa mandarinia, Vespa crabo, Vespa tropica, Vespa analis and
Vespa xanthoptera, collected in central Japan, which has a temperate climate [
27]. Based on this research, other related researches were performed subsequently, which were focused on observations of the metabolism of animals administered an amino acid mixture of an identical composition to the larval saliva of
V. mandarinia [
25,
26,
29,
30,
31,
32]. The vespa amino acid mixture (VAAM) was coined, based on the free amino acid composition of the larval saliva of
V. mandarina, which may be related to aggressiveness and the longest distance flown for hunting [
25,
27,
29]. Studies on the metabolism using of exercising animals revealed that amino acids in larval saliva may act as energy sources and enhance exercise performance through controlling the metabolism [
25,
26,
29,
30,
31].
Vespa mandarinia, the most dominant hornet species in central Japan, has been used as a reference, as it boasts the longest flight distance and the most extensive hunting domains [
25,
27,
29].
Reports on
V. v. nigrithorax invasions to temperate regions sparked an increasing interest in them as their distribution is expanding [
1,
16,
33,
34]. However, studies on trophallaxis and the larval saliva of
V. v. nigrithorax has not been performed despite the significance of these topics. According to previous research by Takashi et al. [
27], each wasp species has a peculiar larval saliva amino acid composition and larval saliva is the principal component of trophallaxis in hornets. There may be a number of factors that influenced the expansion of the
V. v. nigrithorax territory beyond the tropical region. In addition,
V. v. nigrithorax are found in numbers of 1500–2500 individuals per nest, and their nests are 2–3 times larger than those of other
Vespa spp [
2,
35,
36]. The amino acid contents in the larval saliva of
V. v. nigrithorax might be correlated with the energy required for exercise and metabolism and could have influenced the growth of their population, which might inspire us to coin a supplementary diet formula. Thus, the amino acid composition of the larval saliva of
V. v. nigrithorax was investigated.
As
V. mandarinia has been the standard for the physiological studies involved in vespa amino acid mixture (VAAM) supplementation, our results on the amino acid composition of the larval saliva of
V. v. nigrithorax were also presented alongside the amino acid composition of the larval saliva of
V. mandarinia reported by Takashi et al. [
27] (
Table 2). A notable feature of the amino acid composition of the larval saliva of
V. v. nigrithorax was the high proportion of the physiological amino acids compared to the total amount of amino acids measured from the hydrolyzed product of larval saliva. The total amount of physiological amino acids in the larval saliva of
V. v. nigrithorax was 38.29 μmol/mL and the total amount of amino acids deduced from the hydrolyzed product was 67.24 μmol/mL; these results suggest that the larval saliva of
V. v. nigrithorax has a higher ratio of free amino acids than that of
V. mandarinia. Physiological amino acids are valuable nutrients for adult wasp workers; adult wasps are able to consume only liquid foods and hardly digest polymer-type nutrients, owing to their short and simplified gut structure and narrow waists [
37]. Worker wasps in particular, spend most of their time flying for hunting purposes; therefore, free amino acids in larval saliva could help them prolong their flight time, as has been reported in previous studies on animal exercise physiology and VAAM supplementation [
25,
29]. The amino acids that composed the physiological amino acid mixture of the larval saliva of
V. v. nigrithorax were similar to those of saliva from other wasps, as revealed by Takashi et al. [
27]. A notable feature of the physiological amino acid composition of the larval saliva of
V. v. nigrithorax was the high proportion of proline. The content of proline is believed to be correlated with the daily flight distance and hunting domain size of
Vespa spp. [
27], as proline is metabolized by insects to produce energy for hunting [
38]. Branched chain amino acids (BCAAs) such as Ile, Val, and Leu were detected in substantial amounts (3.28, 5.45, and 3.75 mol%, respectively). In animals, BCAAs are directly metabolized in muscles as energy sources and the intake of BCAAs helps prevent muscular loss caused by endurance exercise [
39]. The other essential amino acids (threonine, phenylalanine, lysine, and histidine) were detected in fair amounts (5.48, 5.34, 9.88, and 4.28 mol%, respectively), except for tryptophan and methionine.
The amino acid content of the hydrolyzed product of the larval saliva of
V. v. nigrithorax revealed that a few amino acids were present as peptide constituents (
Table 3). Acidic amino acids aspartate and glutamate were detected in significantly different amounts in the physiological and hydrolyzed amino acid mixtures. Aspartate and glutamate were detected in amounts of 0.06 and 1.71 μmol/mL, respectively, in the physiological amino acid mixture, while they were detected in amounts of 3.21 and 10.71 μmol/mL, respectively, in the hydrolyzed amino acid mixture. Other amino acids such as glycine, alanine, isoleucine, and lysine were detected in the hydrolyzed amino acid mixture in quantities that were about 2–3 times higher than those in the physiological amino acid mixture. The larval saliva of
V. v. nigrithorax did not contain higher amino acid and protein amounts than that of
V. mandarinia. Takashi et al. [
27] suggested that high amino acid and protein contents in larval saliva may be correlated with the body weight and the ecological dominance of wasps in nature. However, the larval saliva of the invasive alien species
V. v. nigrithorax has not been subjected to amino acid analysis up to this point.
3.2. Nutritional Composition of the Larvae
Table 4 represents the nutrient composition of the lyophilized
V. v. nigrithorax larvae compared to other representative edible insect larvae,
Protaetia brevitarsis seulensis and
Tenebrio molitor. The nutrient compositions of
V. mandarinia, P. b. seulensis and
T. molitor larvae were quoted from previous reports [
35,
36,
37]. Among carbohydrates in the larvae, the sugar content was 6.08 g/100 g. And the content of saturated fat was 4.31 g/100 g, while the content of trans fat was detected under the limit of quantification. Total cholesterol content was 31.97 mg/100 g. Sodium content was 60.91 mg/100 g.
The content of protein in
V. v. nigrithorax larvae was similar to that of
T. molitor larvae, and less than in
P. b. seulensis larvae. The content of fat in
V. v. nigrithorax larvae was slightly less than in
P. b. seulensis larvae and far less than in
T. molitor larvae. Interestingly, carbohydrate content in
V. v. nigrithorax larvae was nearly three times of the content in other larvae. Overall nutrient contents in
V. v. nigrithorax larvae imply the larvae may provide a nutritionally balanced diet, as they contain sufficient and balanced amounts of essential nutrients compared to other representative edible insect larvae,
P. b. seulensis and
T. molitor. Compared to dry whole milk, the protein content in
V. v. nigrithorax larvae (48.64%) was an overwhelmingly great amount, as protein takes 26.3% of dry milk weight [
42]. The protein content in
V. v. nigrithorax larvae was even more than commercial dried whey protein concentrate products (protein contents 35–39%) [
43].
In 2010, the South Korean government enacted the “Act of Fosterage and Support of the Insect Industry” to establish a legal basis for supporting the growth of the insect industry [
44]. Along with the approved insect food ingredients (crickets,
T. molitor larvae,
P. b. seulensis larvae, and silkworm), wasps are often ingested in Korea even if they are not approved as food ingredients yet [
13,
45]. However, the nutritional analysis may provide firm grounds for approval of wasp larvae as a food ingredient along with the entomophagic records of wasp larvae in Japan [
46], Papua New Guinea [
7] and Laos [
9].
3.3. Amino Acid Composition in Larvae
Composition of free amino acids in the larvae and amino acids in the hydrolyzed products of the larvae (
Table 5.) demonstrated that the larvae of
V. v. nigrithorax might be a balanced source for essential amino acids. Total content of free amino acids was nearly a quarter of constitutional amino acids of the larvae, which reflected that the larvae might be good resources for amino acid supplement. Twenty-seven amino acids were found in the free amino acid mixture, and sixteen amino acids were detected in hydrolyzed products of the larvae. In particular, essential amino acid contents, except Met and Trp (Arg, His, Ile, Leu, Lys, Phe, Thr and Val), were detected in substantial amounts in hydrolyzed products of the larvae (7.74–24.86 mg/g).
The amount of each hydrolyzed amino acid reflects that
V. v. nigrithorax larvae could be a good supplementation for essential amino acids. Compared to the guideline of the World Health Organization (WHO) for daily adult essential amino acid requirement [
47], the essential amino acids that might be provided by
V. v. nigrithorax larvae are balanced and sufficient.
Table 6 represents daily adult amino acid requirement suggested by the WHO, the amount of each essential amino acid required for an adult of 70 kg weight, and the amount of each essential amino acid equivalent to amino acid contained in 100 g of the larvae. On the basis of the guideline from the WHO, the essential amino acid requirements of a 70 kg adult was deduced, and could be mostly fulfilled by supplementation of amino acids provided from
V. v. nigrithorax larvae. The amounts of Thr, Cys, Phe, Tyr, Lys, and His contained in 100 g of the larvae were more than the daily amino acid requirements for a 70 kg adult (104–143% of the requirement). Val and Ile provided by 100 g of the larvae were estimated to account for 99% and 97% of the requirements, respectively. The Leu content in 100 g of the larvae was 80% of the requirement. Except Trp and Met which were not detected in the larvae, the majority of the essential amino acids in the larvae are sufficient to fulfil the daily requirements of amino acids in adults.
3.5. General Discussion
The habitat of
V. v. nigrithorax is quickly expanding northward, particularly in Korea [
48], since its first reported appearance in 2003 [
16].
V. v. nigrithorax has serious negative public health, economic, and ecological impacts [
1]. In recent years, invasion of alien species has gradually increased in Korea. However, in most of the cases, the controls over the invasive species were not successful, and various damages are rapidly increasing through the nationwide spread [
49,
50,
51,
52]. However, in some cases, the control of alien species is made when the valuable utilization of the invasive species as a potential resource are discovered [
53].
V. v. nigrithorax was designated an ecological disturbance species in 2019, causing comprehensive impact in Korea, but the attempts to discover the utilization as a useful bioresource from this species are rare [
54].
Owing to environmental pressures, food and feed insecurity, and a growing demand for protein, the consumption of insects to ease the problem of global food shortages has been advocated since 1975 [
55]. Along with the approved insect food ingredients (crickets, mealworm,
Protaetia brevitarsis larvae, and silkworm), wasps are also often ingested in Korea even if they are not yet approved as food ingredients by KFDA. The history of utilizing the social wasps as a food and pharmaceutical resources suggests that
V. v. nigrithorax could be a valuable bioresource.