Antimicrobial peptides (AMPs) are crucial effectors of the innate immune system. They provide the first line of defense against a variety of pathogens. AMPs display synergistic effects with conventional antibiotics, and thus present the potential for combined therapies. Insects are extremely resistant to bacterial infections. Insect AMPs are cationic and comprise less than 100 amino acids. These insect peptides exhibit an antimicrobial effect by disrupting the microbial membrane and do not easily allow microbes to develop drug resistance. Currently, membrane mechanisms underlying the antimicrobial effects of AMPs are proposed by different modes: the barrel-stave mode, toroidal-pore, carpet, and disordered toroidal-pore are the typical modes. Positive charge quantity, hydrophobic property and the secondary structure of the peptide are important for the antibacterial activity of AMPs. At present, several structural families of AMPs from insects are known (defensins, cecropins, drosocins, attacins, diptericins, ponericins, metchnikowins, and melittin), but new AMPs are frequently discovered. We reviewed the biological effects of the major insect AMPs. This review will provide further information that facilitates the study of insect AMPs and shed some light on novel microbicides.

Insect Antimicrobial Peptides, a Mini Review.

The black soldier fly Hermetia illucens is used for the bioconversion of organic waste into feed for livestock and aquaculture, and is economically among the most important farmed insects in the world The larvae can be fed on agricultural waste and even liquid manure, resulting in highly unpredictable pathogen levels and dietary conditions Here we show that H illucens larvae express a remarkably expanded spectrum of antimicrobial peptides (AMPs), many of which are induced by feeding on a diet containing high bacterial loads The addition of sulfonated lignin, cellulose, chitin, brewer's grains or sunflower oil revealed the diet-dependent expression profiles of AMPs in the larvae The highest number of AMPs and the highest levels of AMP expression were induced by feeding larvae on diets supplemented with protein or sunflower oil Strikingly, the diet-dependent expression of AMPs translated into diet-dependent profiles of inhibitory activities against a spectrum of bacteria, providing an intriguing example for the emerging field of nutritional immunology We postulate that the fine-tuned expression of the expanded AMP repertoire mediates the adaptation of the gut microbiota to the digestion of unusual diets, and this feature could facilitate the use of H illucens for the bioconversion of organic waste

Nutritional immunology: Diversification and diet-dependent expression of antimicrobial peptides in the black soldier fly Hermetia illucens.

In order to expand with validated scientific data the limited knowledge regarding the potential application of insects as innovative feed ingredients for poultry, the present study tested a partial substitution of soya bean meal and soya bean oil with defatted black soldier fly (Hermetia illucens) larvae meal (H) in the diet for growing broiler quails (Coturnix coturnix japonica) on growth performance, mortality, nutrients apparent digestibility, microbiological composition of excreta, feed choice, carcass and meat traits. With this purpose, a total of 450 10-day-old birds were allocated to 15 cages (30 birds/cage) and received three dietary treatments: a Control diet (C) and two diets (H1 and H2) corresponding to 10% and 15% H inclusion levels, respectively (H substituted 28.4% soya bean oil and 16.1% soya bean meal for H1, and 100% soya bean oil and 24.8% soya bean meal for H2, respectively). At 28 days of age, quails were slaughtered, carcasses were weighed, breast muscles were then excised from 50 quails/treatment, weighed, and ultimate pH (pHu) and L*, a*, b* colour values were measured. Breast muscles were then cooked to assess cooking loss and meat toughness. For the digestibility trial, a total of 15 28-day-old quails were assigned to the three feeding groups. The excreta samples were subjected to chemical and microbiological analysis. The same 15 quails were then simultaneously provided with C and H2 diets for a 10-day feed choice trial. Productive performance, mortality and carcass traits were in line with commercial standards and similar in all experimental groups. With the exception of ether extract digestibility, which was lower in H1 group compared with C and H2 (P=0.0001), apparent digestibility of dry matter, CP, starch and energy did not differ among treatments. Microbial composition of excreta was also comparable among the three groups. Feed choice trial showed that quails did not express a preference toward C or H2 diets. Breast meat weight and yield did not differ among C, H1 and H2 quails. Differently, the inclusion of H meal reduced meat pHu compared with C. In conclusion, this study demonstrated that H. illucens larvae meal can partially replace conventional soya bean meal and soya bean oil in the diet for growing broiler quails, thus confirming to be a promising insect protein source for the feed industry. Further research to assess the impact of H meal on intestinal morphology as well as on meat quality and sensory profile would be of utmost importance.

https://www.cambridge.org/core/services/aop-cambridge-core/content/view/4C4B9F297E2322C5814753EDA14EAE52/S1751731116001270a.pdf/div-class-title-black-soldier-fly-as-dietary-protein-source-for-broiler-quails-apparent-digestibility-excreta-microbial-load-feed-choice-performance-carcass-and-meat-traits-div.pdf

Black soldier fly as dietary protein source for broiler quails: apparent digestibility, excreta microbial load, feed choice, performance, carcass and meat traits.

An experiment was conducted to compare the suitability of chicken, pig, and cow manure as feed for larvae of the black soldier fly (Hermetia illucens (L.); Diptera: Stratiomyidae). Newly hatched larvae were inoculated on moistened manure (33% dry matter). Water and dried manure were added three times per week, until the first prepupae appeared. Survival was between 82 and 97%, indicating that the tested substrates were suitable. However, development time was much longer than on the control diet (144-215 vs 20 days). Efficiency of conversion of ingested nitrogen (N-ECI) was higher on pig manure than on chicken and cow manure, while the ECI for phosphorus was highest on cow manure. Substrate nitrogen content decreased in the chicken manure, but were stable in pig and cow manure. Phosphorus concentration, and N:P-ratio decreased in all treatments. Since a large proportion of the nitrogen from the manure (23–78%) was lost, the production system would require an air washer for instance, to make it ecologically sound. Shorter development times are required in order to improve economic viability. Drying of the manure could have decreased its nutritional value due to the destruction of microorganisms and heat-labile vitamins. A production system using fresh manure could result in a considerably shorter development time and increased conversion efficiency.

Nutrient utilisation by black soldier flies fed with chicken, pig, or cow manure

The oil fraction and partially defatted meal of black soldier fly larvae (Hermetia illucens) affect differently growth performance, feed efficiency, nutrient deposition, blood glucose and lipid digestibility of rainbow trout (Oncorhynchus mykiss)

In this study, we induced and purified a novel antimicrobial peptide exhibiting activity against Gram-positive bacteria from the immunized hemolymph of Hermetia illucens larvae . The immunized hemolymph was extracted, and the novel defensin-like peptide 4 (DLP4) was purified using solid-phase extraction and reverse-phase chromatography. The purified DLP4 demonstrated a molecular weight of 4267 Da, as determined using the matrix-assisted laser desorption/ionization–time-of-flight (MALDI–TOF) method. From analysis of DLP4 by N-terminal amino acid sequencing using Edman degradation, combined with MALDI–TOF and rapid amplification of cDNA ends–polymerase chain reaction (RACE–PCR), the amino acid sequence of the mature peptide was determined to be ATCDLLSPFKVGHAACAAHCIARGKRGGWCDKRAVCNCRK. In NCBI BLAST, the amino acid sequence of DPL4 was found to be 75% identical to the Phlebotomus duboscqi defensin. Analysis of the minimal inhibitory concentration (MIC) revealed that DLP4 have antibacterial effects against Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA). The expression of DLP4 transcripts in several tissues after bacterial challenge was measured by quantitative real-time PCR. Expression of the DLP4 gene hardly occurred throughout the body before immunization, but was mostly evident in the fat body after immunization.

Purification and characterization of a novel antibacterial peptide from black soldier fly (Hermetia illucens) larvae.

Maggots have become highly successful in the treatment of non-healing wounds and multidrug-resistant pathogen infections The main objective of this study was to extract antibacterial substances from larvae of the black soldier fly, Hermetia illucens To induce immune responses, we septically injured the larvae with a contaminated needle Lyophilized H illucens larvae were homogenized and extracted with acidic methanol We examined the antifungal and antibacterial effects of the low molecular weight antimicrobial factors within the larval extract on the growth of a broad range of microorganisms, including Gram-positive Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), and Gram-negative Pseudomonas aeruginosa Furthermore, we isolated the anti-MRSA substances from the larval extract using high performance liquid chromatography These investigations revealed that the larval extract possessed a broad-spectrum of antibacterial activity, demonstrating that secretions of H illucens larvae prove useful in the fight against MRSA and can potentially be a source of novel antibiotic-like compounds for infection control

Detection of antimicrobial substances from larvae of the black soldier fly, Hermetia illucens (Diptera: Stratiomyidae)

Cecropins are basic antibacterial peptides that have potent antimicrobial activities. We induced and purified a novel antimicrobial peptide exhibiting activity against Gram-negative bacteria from the immunized hemolymph of Hermetia illucens larvae. The immunized hemolymph was extracted, and the novel cecropin-like peptide 1 (CLP1) was purified using solid-phase extraction and reverse-phase chromatography. The purified CLP1 demonstrated a molecular weight of 4,840 Da, as determined by matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF). From analysis of CLP1 by N-terminal amino acid sequencing using Edman degradation, combined with MALDI-TOF and rapid amplification of cDNA ends-polymerase chain reaction (RACE-PCR), the amino acid sequence of the mature peptide was determined to be GWRKRVFKPVEKFGQRVRDAGVQGIAIAQQGANVLATARGGP PQQG. In NCBI BLAST, the amino acid sequence of CLP1 was found to be 60 % identical to the Drosophila melanogaster cecropin C. In silico analysis revealed that CLP1 was suggested to be part of the cecropin superfamily of AMPs characterized as cationic, linear, α-helical, and amphipathic polypeptides. Analysis of the minimal inhibitory concentration (MIC) and the minimal bactericidal concentration (MBC) showed that CLP1 exerted antibacterial effects against Gram-negative bacteria. The expression of CLP1 transcripts in several tissues after bacterial challenge was measured by quantitative real-time PCR. CLP1 expression was negligible throughout the body before immunization, and was mostly evident in the fat body after immunization.

A novel cecropin-like peptide from black soldier fly, Hermetia illucens: Isolation, structural and functional characterization

In this study, we report a novel member of the attacin family from Hermetia illucens. The cDNA clone encoding the attacin-like protein was isolated by screening a cDNA library prepared from immuniz...

Novel attacin from Hermetia illucens: cDNA cloning, characterization, and antibacterial properties.

An antibacterial peptide was isolated from a black soldier fly, Hermetia illucens. The molecular mass of this peptide was established as 4247.37 by matrix-assisted laser desorption/ionization-time of flight mass (MALD-TOF MS) spectrometry. The amino acid sequence of the mature peptide was determined by N-terminal sequencing using Edman degradation, combined with cDNA sequencing of the previously reported defensin-like peptide (DLP) 3. Analysis of the minimal inhibitory concentration (MIC) revealed that DLP3 had potent activity against Gram-positive and negative bacteria, but DLP4 had only anti-Gram-positive activity as previously reported. Recombinant DLP3 and DLP4 were overexpressed in Escherichia coli, and antibacterial activities were identical to DLPs purified from H. illucens hemolyph. In silico analysis revealed that only six amino acid sequences were different between DLP3 and DLP4, but antibacterial activity against Gram-negative bacteria differed. Therefore these amino acid variants may be key amino acids (Gly-10, Val-18, Met-23, Arg-25, Asp-32, Arg-40) related to killing Gram-negative bacteria.

Soon-Ik Park

Papers

Purification and characterization of a novel antibacterial peptide from black soldier fly (Hermetia illucens) larvae.

Detection of antimicrobial substances from larvae of the black soldier fly, Hermetia illucens (Diptera: Stratiomyidae)

A novel cecropin-like peptide from black soldier fly, Hermetia illucens: Isolation, structural and functional characterization

Novel attacin from Hermetia illucens: cDNA cloning, characterization, and antibacterial properties.

Defensin-like peptide3 from black solder fly: Identification, characterization, and key amino acids for anti-Gram-negative bacteria