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Comparative Analysis of the Composition Of Comparative Analysis of the Composition of Intestinal Bacterial Communities in Dastarcus helophoroides Fed Different Diets Author(s): Wei-Wei Wang, Cai He, Jun Cui, Hai-Dong Wang, and Meng-Lou Li Source: Journal of Insect Science, 14(111):1-13. 2014. Published By: Entomological Society of America DOI: http://dx.doi.org/10.1673/031.014.111 URL: http://www.bioone.org/doi/full/10.1673/031.014.111 BioOne (www.bioone.org) is a nonprofit, online aggregation of core research in the biological, ecological, and environmental sciences. BioOne provides a sustainable online platform for over 170 journals and books published by nonprofit societies, associations, museums, institutions, and presses. Your use of this PDF, the BioOne Web site, and all posted and associated content indicates your acceptance of BioOne’s Terms of Use, available at www.bioone.org/page/terms_of_use. Usage of BioOne content is strictly limited to personal, educational, and non-commercial use. Commercial inquiries or rights and permissions requests should be directed to the individual publisher as copyright holder. BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit publishers, academic institutions, research libraries, and research funders in the common goal of maximizing access to critical research. Journal of Insect Science: Vol. 14 | Article 111 Wang et al. Comparative analysis of the composition of intestinal bacterial communities in Dastarcus helophoroides fed different diets Wei-Wei Wang,1a Cai He,2b Jun Cui,1c Hai-Dong Wang,1d and Meng-Lou Li1e* 1Laboratory of Forestry Pests Biological Control, College of Forestry, Northwest A&F University, Yangling, Shaanxi, 712100, P. R. China 2Wuwei Academy of Forestry Sciences, Wuwei, Gansu, 733000, P. R. China Abstract The diversity of the intestinal bacterial communities in Dastarcus helophoroides (Fairmaire) (Coleoptera: Bothrideridae) larvae and adults was assayed by PCR-DGGE to determine whether different artificial diets could influence these bacterial communities. Two diets were used for feeding the larvae and four for the adults. Escherichia, Desemzia, Staphylococcus, Asticcacaulis, Cellvibrio, Aurantimonas, and Planomicrobium were isolated from the gut of the adults, with Escherichia and Staphylococcus being the main bacterial communities, and the quantities of in- testinal bacterial were different in the adults fed different diets. Specifically, the amount of intestinal bacteria from the adults fed different diets had the following ranking according to the major component of the diet: ant powder > darkling beetle pupa powder > cricket powder > silk- worm pupa powder. Escherichia, Bacillus, Staphylococcus, Kurthia, Planococcaceae, Ralstonia, Leptothrix, Acinetobacter, and Pseudomonas were isolated from the gut of the larvae. The quanti- ty of intestinal bacteria from the larvae fed the darkling beetle pupae was greater than that from the larvae fed other artificial diets. This study, for the first time, investigated the effect of artifi- cial diets on the bacterial community and the intestinal microbial diversity of D. helophoroides. Keywords: parasitoid beetle, nested PCR, DGGE, larva, adult Correspondence: a [email protected], b [email protected], c [email protected], d [email protected], e [email protected], *Corresponding author Editor: Allen Cohen was editor of this paper. Received: 1 October 2012 Accepted: 20 February 2013 Published: 12 August 2014 Copyright: This is an open access paper. We use the Creative Commons Attribution 3.0 license that permits unrestricted use, provided that the paper is properly attributed. ISSN: 1536-2442 | Vol. 14, Number 111 Cite this paper as: Wang WW, He C, Cui J, Wang HD, Li ML. 2014. Comparative analysis of the composition of intestinal bacterial communities in Dastarcus helophoroides fed different diets. Journal of Insect Science 14(111). Available online: http://www.insectscience.org/14.111 Journal of Insect Science | http://www.insectscience.org 1 Journal of Insect Science: Vol. 14 | Article 111 Wang et al. Introduction Kane and Breznak (1991) and Kaufman et al. (2000) found that using different food sources Vast numbers of microorganisms inhabit the could promote the development of alternative insect gut, and many occupy the alimentary bacterial communities in cockroaches and tract and play an important role in the physio- crickets. Abundant microorganisms inhabit in logical function and behavior of their host the gut of Costelytra zealandica (White) of (Dillon and Dillon 2004). It is well known that Coleoptera, an important group in insects intestinal tract flora plays an important role in (Zhang and Jackson 2008), and are considered maintaining the health of the host by main- to play important roles in coleopteran nutri- taining a normal ecological equilibrium, tion and metabolism (Jeon et al. 2011). combining absorption, digestion, and protec- tion against the invasion of pathogenic Dastarcus helophoroides (Fairmaire) (Coleop- microorganisms (Rio et al. 2004, Dillon et al. tera: Bothrideridae) is one of the most 2005, Jeon et al. 2011). At present, the re- effective natural enemies against the large- search on the insect intestinal microflora and bodied longhorn beetle (Yang 2004). The lar- dominant bacterial communities is focused va of D. helophoroides is an ecto-parasitoid of mainly on termites belonging to Isoptera and the larvae and pupae of various trunk borers, on Bombyx mori (L.), which belongs to Lepi- such as Anoplophora glabripennis doptera (Eutick et al. 1978, Brune and (Motschulsky), Monochamus alternatus Friedrich 2000, Xiang et al. 2007). (Hope), Mallambyx radei Blessig, Apriona swainsoni (Hope), Batocera horsfieldi (Hope), Denaturing gradient gel electrophoresis and A. germari (Wang et al. 1996, Yang (DGGE) has been used widely to identify and 2004). Although the adult lifespan of D. helo- monitor the microbes common to different phoroides exceeds three years, the larval stage individuals and environments (Reeson et al. is only approximately one week, and the pupal 2003, Hayashi et al. 2007). Using polymerase stage is 20 to 30 days. The previous reports on chain reaction and denaturing gradient gel D. helophoroides were focused mainly on its electrophoresis (PCR-DGGE), Zhang and biological characteristics (Lei et al. 2003), ar- Jackson (2008) studied the digestive tract of tificial diets, the release of adults for scarabs and found that the dominant bacterial biological control (Wang and Ogura 1999), communities were mainly Clostridiales, fol- and the adult host-identification mechanism of lowed by Proteobacteria and Bacteroides. using informational chemicals (Wei et al. Twenty-one bacteria, belonging to thirteen 2008). In this study, we examined the micro- microflora, were isolated from the larval gut bial communities of larval and adult guts of D. of Apriona germari (Hope) (Zhang et al. helophoroides by using DGGE to separate 2004). Different food resources provide nutri- amplified 16S rDNA gene fragments by PCR. ents to the different bacterial communities in the intestines of insects, and the diet of a Because the narrow distribution and low den- number of aquatic invertebrates is known to sity of D. helophoroides in nature limit its influence their intestinal microflora (Harris function in biological control, it is necessary 1993). The effect of diet on the intestinal bac- to develop artificial diets for D. helophoroid- teria in crickets (Kaufman et al. 2000), es. However, if the composition of the cockroaches (Kane and Breznak 1991), and artificial diet were inappropriate, it would af- wasps (Reeson et al. 2003) has been reported. fect the intestinal bacterial community and Journal of Insect Science | http://www.insectscience.org 2 Journal of Insect Science: Vol. 14 | Article 111 Wang et al. thus the digestive function and healthy growth 2005, Shang et al. 2009), but whether these of the insect. To date, artificial diets for D. diets have an effect on the intestinal bacteria helophoroides larvae and adults have been remains unknown. Hence, this study investi- investigated (Wang et al. 1999, Lei et al. gated the larval and adult intestinal microflora of D. helophoroides fed different arti- Table 1. Composition of the four diets used for feeding D. helophoroides adults. ficial diets under the same conditions. Ingredient Diet I II III IV Materials and Methods Silkworm pupa powdera 10 g a Darkling beetle pupa powder 10 g Diets and insects Cricket powdera 10 g All of the insects used in this study Ant powdera 10 g were housed in the laboratory under Wood powderb 2.5 g optimal growing conditions. The adult Dry yeastsc 10 g 10 g 10 g 10 g artificial diets (Diet I, II, III, and IV) Lyophilized chicken yolkd 14 g 14 g 14 g 14 g Sucrosec 3.3 g 3.3 g 3.3 g 3.3 g were prepared as shown in Table 1, Agarc 3.3 g 3.3 g 3.3 g 3.3 g and all insects used as the major com- Benzoic acidc 0.02 g 0.02 g 0.02 g 0.02 g ponent of four diets were collected Distilled water 56.88 mL 56.88 mL 56.88 mL 56.88 mL from the wild, dried, and ground into Sunflower seed oild 8 mL powder in the lab. The larval Diet V a Silkworm pupae, crickets, darkling beetle pupae, and ants were collected from consisted of darkling beetle pupae. the wild and then dried and ground into powder in the lab. The artificial Diet VI was prepared as b Poplar xylem was dried and ground into powder in the lab. c Dry yeasts, sucrose, agar, and benzoic acid were purchased from TIANGEN shown in Table 2, and the contents
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