Dataset Information

Revealing the respiratory system of the coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae: Scolytinae) using micro-computed tomography.

ABSTRACT: The coffee berry borer (Hypothenemus hampei) is the most economically important insect pest of coffee globally. Micro-computed tomography (micro-CT) was used to reconstruct the respiratory system of this species for the first time; this is the smallest insect (ca. 2?mm long) for which this has been done to date. Anatomical details of the spiracles and tracheal tubes are described, images presented, and new terms introduced. The total volume and the relationship between tracheal lumen diameter, length and volume are also presented. The total length of the tracheal tubes are seventy times the length of the entire animal. Videos and a 3D model for use with mobile devices are included as supplementary information; these could be useful for future research and for teaching insect anatomy to students and the public in general.

SUBMITTER: Alba-Tercedor J

PROVIDER: S-EPMC6882887 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Revealing the respiratory system of the coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae: Scolytinae) using micro-computed tomography.

Alba-Tercedor Javier J Alba-Alejandre Ignacio I Vega Fernando E FE

Scientific reports 20191128 1

The coffee berry borer (Hypothenemus hampei) is the most economically important insect pest of coffee globally. Micro-computed tomography (micro-CT) was used to reconstruct the respiratory system of this species for the first time; this is the smallest insect (ca. 2 mm long) for which this has been done to date. Anatomical details of the spiracles and tracheal tubes are described, images presented, and new terms introduced. The total volume and the relationship between tracheal lumen diameter, l ...[more]

PMID: 31780747

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Project description:BackgroundThe coffee berry borer, Hypothenemus hampei, reproduces and feeds exclusively on the mature endosperm of the coffee seed, which has a cell wall composed mainly of a heterogeneous mixture of hemicellulose polysaccharides, including arabinoxylans. Xylanases are digestive enzymes responsible for the degradation of xylan based polymers, hydrolyzing them into smaller molecules that are easier to assimilate by insects. We report the cloning, expression and enzymatic characterization of a xylanase gene that was identified in the digestive tract of the coffee berry borer.MethodsThe complete DNA sequence encoding a H. hampei xylanase (HhXyl) was obtained using a genome walking technique in a cDNA library derived from the borer digestive tract. The XIP-I gene was amplified from wheat (Triticum aestivum variety Soisson). A Pichia pastoris expression system was used to express the recombinant form of these enzymes. The xylanase activity and XIP-I inhibitory activity was quantified by the 3,5-dinitrosalicylic (DNS). The biological effects of XIP-I on borer individuals were evaluated by providing an artificial diet enriched with the recombinant XIP-I protein to the insects.ResultsThe borer xylanase sequence contains a 951 bp open reading frame that is predicted to encode a 317-amino acid protein, with an estimated molecular weight of 34.92 kDa and a pI of 4.84. Bioinformatic analysis revealed that HhXyl exhibits high sequence homology with endo-β-D-xylanases of Streptomyces bingchenggensis from glycosyl hydrolase 10 (GH10). The recombinant xylanase showed maximal activity at pH 5.5 and 37°C. XIP-I expressed as a recombinant protein inhibited HhXyl activity in vitro and caused individual H. hampei mortality in bioassays when included as a supplement in artificial diets.ConclusionA xylanase from the digestive tract of the coffee berry borer was identified and functionally characterized. A xylanase inhibitor protein, XIP-I, from wheat was shown to be a potent inhibitor of this xylanase, suggesting that its deployment has potential as a strategy to control coffee berry borer colonization of coffee plants.

Project description:BackgroundThe study of coffee polysaccharides-degrading enzymes from the coffee berry borer Hypothenemus hampei, has become an important alternative in the identification for enzymatic inhibitors that can be used as an alternative control of this dangerous insect. We report the cloning, expression and biochemical characterization of a mannanase gene that was identified in the midgut of the coffee berry borer and is responsible for the degradation of the most abundant polysaccharide in the coffee bean.MethodsThe amino acid sequence of HhMan was analyzed by multiple sequence alignment comparisons with BLAST (Basic Local Alignment Search Tool) and CLUSTALW. A Pichia pastoris expression system was used to express the recombinant form of the enzyme. The mannanase activity was quantified by the 3,5-dinitrosalicylic (DNS) and the hydrolitic properties were detected by TLC.ResultsAn endo-1,4-β-mannanase from the digestive tract of the insect Hypothenemus hampei was cloned and expressed as a recombinant protein in the Pichia pastoris system. This enzyme is 56% identical to the sequence of an endo-β-mannanase from Bacillus circulans that belongs to the glycosyl hydrolase 5 (GH5) family. The purified recombinant protein (rHhMan) exhibited a single band (35.5 kDa) by SDS-PAGE, and its activity was confirmed by zymography. rHhMan displays optimal activity levels at pH 5.5 and 30°C and can hydrolyze galactomannans of varying mannose:galactose ratios, suggesting that the enzymatic activity is independent of the presence of side chains such as galactose residues. The enzyme cannot hydrolyze manno-oligosaccharides such as mannobiose and mannotriose; however, it can degrade mannotetraose, likely through a transglycosylation reaction. The K(m) and k(cat) values of this enzyme on guar gum were 2.074 mg ml(-1) and 50.87 s(-1), respectively, which is similar to other mannanases.ConclusionThis work is the first study of an endo-1,4-β-mannanase from an insect using this expression system. Due to this enzyme's importance in the digestive processes of the coffee berry borer, this study may enable the design of inhibitors against endo-1,4-β-mannanase to decrease the economic losses stemming from this insect.

Dataset Information

Revealing the respiratory system of the coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae: Scolytinae) using micro-computed tomography.

Publications

Revealing the respiratory system of the coffee berry borer (Hypothenemus hampei; Coleoptera: Curculionidae: Scolytinae) using micro-computed tomography.

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