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<front>
<journal-meta>
<journal-id journal-id-type="publisher-id">Front. Plant Sci.</journal-id>
<journal-title>Frontiers in Plant Science</journal-title>
<abbrev-journal-title abbrev-type="pubmed">Front. Plant Sci.</abbrev-journal-title>
<issn pub-type="epub">1664-462X</issn>
<publisher>
<publisher-name>Frontiers Media S.A.</publisher-name>
</publisher>
</journal-meta>
<article-meta>
<article-id pub-id-type="doi">10.3389/fpls.2023.1220339</article-id>
<article-categories>
<subj-group subj-group-type="heading">
<subject>Plant Science</subject>
<subj-group>
<subject>Original Research</subject>
</subj-group>
</subj-group>
</article-categories>
<title-group>
<article-title>Larvicidal proficiency of volatile compounds present in <italic>Commiphora wightii</italic> gum extract against <italic>Aedes aegypti</italic> (Linnaeus, 1762)</article-title>
</title-group>
<contrib-group>
<contrib contrib-type="author">
<name>
<surname>Patel</surname>
<given-names>Krupal</given-names>
</name>
<xref ref-type="aff" rid="aff1">
<sup>1</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/1325482"/>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Akbari</surname>
<given-names>Divya</given-names>
</name>
<xref ref-type="aff" rid="aff2">
<sup>2</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Pandya</surname>
<given-names>Rohan V.</given-names>
</name>
<xref ref-type="aff" rid="aff3">
<sup>3</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Trivedi</surname>
<given-names>Jigneshkumar</given-names>
</name>
<xref ref-type="aff" rid="aff4">
<sup>4</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/1325464"/>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Mevada</surname>
<given-names>Vishal</given-names>
</name>
<xref ref-type="aff" rid="aff5">
<sup>5</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/940807"/>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Wanale</surname>
<given-names>Shivraj Gangadhar</given-names>
</name>
<xref ref-type="aff" rid="aff6">
<sup>6</sup>
</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Patel</surname>
<given-names>Rajesh</given-names>
</name>
<xref ref-type="aff" rid="aff7">
<sup>7</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/1772537"/>
</contrib>
<contrib contrib-type="author" corresp="yes">
<name>
<surname>Yadav</surname>
<given-names>Virendra Kumar</given-names>
</name>
<xref ref-type="aff" rid="aff4">
<sup>4</sup>
</xref>
<xref ref-type="author-notes" rid="fn001">
<sup>*</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/193265"/>
</contrib>
<contrib contrib-type="author" corresp="yes">
<name>
<surname>Tank</surname>
<given-names>Jigna G.</given-names>
</name>
<xref ref-type="aff" rid="aff2">
<sup>2</sup>
</xref>
<xref ref-type="author-notes" rid="fn001">
<sup>*</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/769464"/>
</contrib>
<contrib contrib-type="author" corresp="yes">
<name>
<surname>Sahoo</surname>
<given-names>Dipak Kumar</given-names>
</name>
<xref ref-type="aff" rid="aff8">
<sup>8</sup>
</xref>
<xref ref-type="author-notes" rid="fn001">
<sup>*</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/225187"/>
</contrib>
<contrib contrib-type="author" corresp="yes">
<name>
<surname>Patel</surname>
<given-names>Ashish</given-names>
</name>
<xref ref-type="aff" rid="aff4">
<sup>4</sup>
</xref>
<xref ref-type="author-notes" rid="fn001">
<sup>*</sup>
</xref>
<uri xlink:href="https://loop.frontiersin.org/people/2189536"/>
</contrib>
</contrib-group>
<aff id="aff1">
<sup>1</sup>
<institution>Marine Biodiversity and Ecology Laboratory, Department of Zoology, Faculty of Science, The Maharaja Sayajirao University of Baroda</institution>, <addr-line>Vadodara, Gujarat</addr-line>, <country>India</country>
</aff>
<aff id="aff2">
<sup>2</sup>
<institution>University Grants Commission-Career Advancement Scheme (UGC-CAS) Department of Biosciences, Saurashtra University</institution>, <addr-line>Rajkot, Gujarat</addr-line>, <country>India</country>
</aff>
<aff id="aff3">
<sup>3</sup>
<institution>Department of Microbiology, Atmiya University</institution>, <addr-line>Rajkot, Gujarat</addr-line>, <country>India</country>
</aff>
<aff id="aff4">
<sup>4</sup>
<institution>Department of Life Sciences, Hemchandracharya North Gujarat University</institution>, <addr-line>Patan, Gujarat</addr-line>, <country>India</country>
</aff>
<aff id="aff5">
<sup>5</sup>
<institution>DNA Division, Directorate of Forensic Science</institution>, <addr-line>Gandhinagar</addr-line>, <country>India</country>
</aff>
<aff id="aff6">
<sup>6</sup>
<institution>School of Chemical Sciences, Swami Ramanand Teerth Marathwada University</institution>, <addr-line>Nanded, Maharashtra</addr-line>, <country>India</country>
</aff>
<aff id="aff7">
<sup>7</sup>
<institution>Department of Biosciences, Veer Narmad South Gujarat University</institution>, <addr-line>Surat</addr-line>, <country>India</country>
</aff>
<aff id="aff8">
<sup>8</sup>
<institution>Department of Veterinary Clinical Sciences, College of Veterinary Medicine, Iowa State University</institution>, <addr-line>Ames, IA</addr-line>, <country>United States</country>
</aff>
<author-notes>
<fn fn-type="edited-by">
<p>Edited by: Petr Mad&#x11b;ra, Mendel University in Brno, Czechia</p>
</fn>
<fn fn-type="edited-by">
<p>Reviewed by: Lucie Vanickova, Mendel University in Brno, Czechia; Seema Ramniwas, Chandigarh University, India</p>
</fn>
<fn fn-type="corresp" id="fn001">
<p>*Correspondence: Ashish Patel, <email xlink:href="mailto:uni.ashish@gmail.com">uni.ashish@gmail.com</email>; Dipak Kumar Sahoo, <email xlink:href="mailto:dsahoo@iastate.edu">dsahoo@iastate.edu</email>; Jigna G. Tank, <email xlink:href="mailto:jignagtank@gmail.com">jignagtank@gmail.com</email>; Virendra Kumar Yadav, <email xlink:href="mailto:yadava94@gmail.com">yadava94@gmail.com</email>
</p>
</fn>
</author-notes>
<pub-date pub-type="epub">
<day>30</day>
<month>08</month>
<year>2023</year>
</pub-date>
<pub-date pub-type="collection">
<year>2023</year>
</pub-date>
<volume>14</volume>
<elocation-id>1220339</elocation-id>
<history>
<date date-type="received">
<day>10</day>
<month>05</month>
<year>2023</year>
</date>
<date date-type="accepted">
<day>07</day>
<month>08</month>
<year>2023</year>
</date>
</history>
<permissions>
<copyright-statement>Copyright &#xa9; 2023 Patel, Akbari, Pandya, Trivedi, Mevada, Wanale, Patel, Yadav, Tank, Sahoo and Patel</copyright-statement>
<copyright-year>2023</copyright-year>
<copyright-holder>Patel, Akbari, Pandya, Trivedi, Mevada, Wanale, Patel, Yadav, Tank, Sahoo and Patel</copyright-holder>
<license xlink:href="http://creativecommons.org/licenses/by/4.0/">
<p>This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.</p>
</license>
</permissions>
<abstract>
<p>
<italic>Aedes</italic> mosquitoes are the major cause of several vector-borne diseases in tropical and subtropical regions. Synthetic pesticides against these mosquitoes have certain limitations; hence, natural, eco-friendly, and safe larvicides obtained from plant resources are used to overcome these. In the present study, the larvicidal efficiency of <italic>Commiphora wightii</italic> against the fourth instar stage of the dengue fever mosquito <italic>Aedes aegypti</italic> (Linnaeus, 1762) was studied. The gum resin of <italic>C. wightii</italic> was collected using the borehole tapping method, and hexane extracts in different concentrations were prepared. The fourth-instar larvae were exposed to the extracts, and percent mortality, as well as LC<sub>20</sub>, LC<sub>50</sub>, and LC<sub>90</sub>, was calculated. Volatile compounds of the hexane gum extract were analyzed by Headspace GC/MS, and the sequence of the acetylcholine, Gamma-aminobutyric acid (GABA) receptor, and octopamine receptor subunit of <italic>A. aegypti</italic> was obtained. It was found that the hexane gum extract was toxic and lethal for larvae at different concentrations. Minimum mortality was observed at 164 &#xb5;g mL<sup>&#x2212;1</sup> (10%/h), while maximum mortality was at 276 &#xb5;g mL<sup>&#x2212;1</sup> (50%/h). The lethal concentrations LC<sub>20</sub>, LC<sub>50</sub>, and LC<sub>90</sub> were 197.38 &#xb5;g mL<sup>&#x2212;1</sup>, 294.13 &#xb5;g mL<sup>&#x2212;1</sup>, and 540.15 &#xb5;g mL<sup>&#x2212;1</sup>, respectively. The GC/MS analysis confirmed the presence of diterpenes, monoterpenes, monoterpene alcohol, and sesquiterpenes in the gum samples, which are lethal for larvae due to their inhibitory activity on the acetylcholinesterase enzyme, GABA receptor, and octopamine receptor subunit. The use of commonly occurring plant gum for the control of mosquitoes was explored, and it was found that the gum of <italic>C. wightii</italic> had larvicidal activities and could be potentially insecticidal.</p>
</abstract>
<kwd-group>
<kwd>
<italic>Aedes aegypti</italic>
</kwd>
<kwd>crude extracts</kwd>
<kwd>larvicidal activity</kwd>
<kwd>volatile compounds</kwd>
<kwd>plant gum extract</kwd>
<kwd>viral vector</kwd>
</kwd-group>
<counts>
<fig-count count="7"/>
<table-count count="2"/>
<equation-count count="2"/>
<ref-count count="45"/>
<page-count count="10"/>
<word-count count="3951"/>
</counts>
<custom-meta-wrap>
<custom-meta>
<meta-name>section-in-acceptance</meta-name>
<meta-value>Plant Metabolism and Chemodiversity</meta-value>
</custom-meta>
</custom-meta-wrap>
</article-meta>
</front>
<body>
<sec id="s1" sec-type="intro">
<title>Introduction</title>
<p>Mosquitoes are a well-known insect vector, transmitting various diseases that lead to millions of deaths every year (<xref ref-type="bibr" rid="B1">Acock et&#xa0;al., 2022</xref>; <xref ref-type="bibr" rid="B22">Kaavya et&#xa0;al., 2022</xref>). Globally, there is a documented presence of approximately 3,540 species (41 genera) of mosquitoes (<xref ref-type="bibr" rid="B16">Harbach and Besansky, 2014</xref>). Among these, <italic>Aedes</italic> mosquitoes are the main cause of viral vector-borne diseases such as chikungunya, Zika virus infection, yellow fever, hemorrhagic fever, and dengue fever in the tropical and subtropical regions (<xref ref-type="bibr" rid="B44">Yang et&#xa0;al., 2009</xref>; <xref ref-type="bibr" rid="B22">Kaavya et&#xa0;al., 2022</xref>). Out of these, dengue is a dominant and rapidly spreading disease caused by <italic>Aedes aegypti</italic> mosquitoes (<xref ref-type="bibr" rid="B14">Guzman and Harris, 2015</xref>). The transmission of dengue fever is facilitated by female <italic>A. aegypti</italic> mosquitoes, which deposit their eggs in stagnant water to raise the immature stages of their biphasic life cycle (<xref ref-type="bibr" rid="B6">Benelli and Mehlhorn, 2016</xref>).</p>
<p>Presently, the absence of efficacious dengue vaccines necessitates reliance on mosquito control as the sole preventive measure against the disease. This can be achieved through the wide use of different insecticides available. Synthetic larvicides are widely used in <italic>Aedes</italic> mosquitoes&#x2019; breeding places to control their vector-containing larva growth. However, deficiency in the control plans has resulted in the development of resistance by larvae against these synthetic larvicides (<xref ref-type="bibr" rid="B8">Braga et&#xa0;al., 2004</xref>; <xref ref-type="bibr" rid="B27">Medronho, 2008</xref>; <xref ref-type="bibr" rid="B10">Dusfour et&#xa0;al., 2011</xref>). In a study using WHO protocol to determine insecticide resistance and its mechanisms in the primary dengue vector in dengue-endemic districts of West Bengal, India, it was found that most <italic>Aedes</italic> populations were resistant to multiple synthetic insecticides (<xref ref-type="bibr" rid="B7">Bharati and Saha, 2018</xref>).</p>
<p>The development of insecticidal resistance has become a major issue in controlling vectors and, hence, vector-borne diseases. Such resistance can be induced due to changes in mosquitoes&#x2019; enzymatic system, leading to rapid detoxification and sequestration of exposed insecticide. Moreover, mutations in the target site can also prevent insecticide&#x2013;target interaction (<xref ref-type="bibr" rid="B17">Hemingway et&#xa0;al., 2004</xref>). Synthetic larvicides also have an adverse effect on the environment.</p>
<p>In order to overcome these issues, many researchers have explored natural, eco-friendly, and safe larvicides obtained from plant resources. Various studies have documented biologically active plant compounds having efficient larvicidal and insecticidal effects on mosquitoes. It has been observed that these biologically active plant compounds are evolving and that they have no negative effects on the environment. They present the best alternative to synthetic pesticides, as they are environment-friendly, least toxic to human health, and convenient to use. Approximately 2,000 plant species are known for producing compounds that can be used for pest management, out of which 344 species are reported to be used against mosquitoes. Although these many species are known for mosquito control, phytochemical extracts of only a few plants have been known to act against mosquitoes in the form of growth regulators, repellents, and ovipositional barriers.</p>
<p>Guggulu is an oleo-gum resin that exudes from the bark of <italic>Commiphora wightii</italic> (Arnott) Bhandari or &#x201c;Indian bdellium.&#x201d; It has been used in Ayurveda since time immemorial to treat a variety of disorders, such as inflammation, gout, rheumatism, obesity, and lipid metabolism disorders (<xref ref-type="bibr" rid="B33">Sarup et&#xa0;al., 2015</xref>). <italic>Commiphora wightii</italic> resin incense was burned to repel mosquitoes traditionally by the tribes of Rajasthan, India (<xref ref-type="bibr" rid="B23">Kantheti and Padma, 2017</xref>).</p>
<p>The larvicidal activity of <italic>C. wightii</italic> gum resin and the effect of the identified volatile compounds on the development of <italic>A. aegypti</italic> larvae were analyzed. Moreover, the phytochemicals were screened against a homology model of the nervous system protein target acetylcholinesterase, the GABA receptor subunit, and the octopamine receptor of <italic>A. aegypti</italic> larvae.</p>
</sec>
<sec id="s2" sec-type="materials|methods">
<title>Materials and methods</title>
<sec id="s2_1">
<title>Collection of gum and preparation of hexane gum extract</title>
<p>The borehole tapping method was used to obtain gum resin from the bark of <italic>C. wightii</italic>. Pure hexane (Sigma Aldrich; purity: &#x2265;99%; CAS Number: 592-41-6) was used to prepare the extract from the gum sample using the Soxhlet extraction procedure for 3 h (<xref ref-type="bibr" rid="B40">Vasishth and Guleria, 2017</xref>). Later, 200-mL samples were extracted and stored in a vacuum-tight glass reagent bottle at 4&#xb0;C until further analysis.</p>
</sec>
<sec id="s2_2">
<title>Culturing of <italic>Aedes aegypti</italic> larva</title>
<p>Dengue fever mosquitoes (<italic>A. aegypti</italic>) were cultured in the small mosquito insectary under laboratory conditions at the botanical garden of the Department of Biosciences, Saurashtra University, Rajkot, Gujarat, India. The colony was maintained in breeding cages 40 cm &#xd7; 40 cm &#xd7; 40 cm with standard conditions like temperature (28&#xb0;C &#xb1; 1&#xb0;C), humidity (80% &#xb1; 5% RH), and photoperiod (14L:10D) (<xref ref-type="bibr" rid="B24">Kumar et&#xa0;al., 2010</xref>; <xref ref-type="bibr" rid="B38">Udayanga et&#xa0;al., 2019</xref>). The fourth instar stage of <italic>A. aegypti</italic> larvae was used for bioassays.</p>
</sec>
<sec id="s2_3">
<title>Larvicidal activity</title>
<p>The larvicidal activity was carried out at a laboratory scale. Different concentrations of the plant gum extracts in hexane were prepared to range from 164 &#xb5;g mL<sup>&#x2212;1</sup> to 284 &#xb5;g mL<sup>&#x2212;1</sup>. For each hexane gum extract, 50 larvae of <italic>A. aegypti</italic> (fourth instar stage) were exposed for 60 min. The controls were exposed to the hexane solvent. The duration required for the mortality of half the number of larvae and all of the larvae in each concentration of hexane gum extract was recorded. Each bioassay was replicated in triplicates. The percent mortality of larvae was calculated using the following formula:</p>
<disp-formula>
<mml:math display="block" id="M1">
<mml:mrow>
<mml:mtext>Mortality</mml:mtext>
<mml:mo>&#xa0;</mml:mo>
<mml:mrow>
<mml:mo stretchy="false">(</mml:mo>
<mml:mo>%</mml:mo>
<mml:mo stretchy="false">)</mml:mo>
</mml:mrow>
<mml:mo>=</mml:mo>
<mml:mfrac>
<mml:mrow>
<mml:mtext>No.&#xa0;of&#xa0;dead&#xa0;larvae</mml:mtext>
</mml:mrow>
<mml:mrow>
<mml:mtext>No.&#xa0;of&#xa0;total&#xa0;larvae</mml:mtext>
</mml:mrow>
</mml:mfrac>
<mml:mo>&#xa0;</mml:mo>
<mml:mo>&#xd7;</mml:mo>
<mml:mo>&#xa0;</mml:mo>
<mml:mn>100</mml:mn>
</mml:mrow>
</mml:math>
</disp-formula>
</sec>
<sec id="s2_4">
<title>Statistical analysis</title>
<p>The percent mortality data were subjected to probit analysis for the calculation of LC<sub>20</sub>, LC<sub>50</sub>, and LC<sub>90</sub>. The regression analysis at the 95% confidence level and chi-square values were acculated by considering a <italic>p</italic>-value&lt;0.05 to be statistically significant. The analyses were performed using Past software (4.03 version).</p>
</sec>
<sec id="s2_5">
<title>Headspace GC/MS analysis of gum samples</title>
<p>The volatile compounds present in each hexane gum extract were analyzed by Headspace GC/MS. Their total ion chromatograms were obtained using a GERSTEL DHS System (Germany) connected to an Agilent 7890A GC and 5975C MS equipped with an Inert MSD with Triple Axis Detector (Germany). Each gum sample (1 g) was placed in a 20-mL standard headspace vial, which was placed in a tray of a GERSTEL Multipurpose Sampler. Volatile compounds were injected automatically into a CP-WAX 52 GC column (60 m &#xd7; 25 &#x3bc;m film thickness &#xd7; 0.25 mm) inner diameter (INNOWax, Germany). In the analytical conditions, the flow rate of helium gas was 1.2 mL/min, inlet pressure was 25 kPa, linear velocity was 1 mL/min at 210&#xb0;C, injector temperature was 250&#xb0;C, and injection mode was splitless. In the MS scan conditions, the source temperature was 200&#xb0;C, the interface temperature was 250&#xb0;C, the electron energy was 70 eV, and the mass scan was in the range of 40&#x2013;350 amu. The temperature program was from 40&#xb0;C to 240&#xb0;C at a rate of 5&#xb0;C/min. The relative percentage of each identified compound was calculated from the GC peak area. The compounds in each sample were identified through a comparison of spectral mass patterns and linear retention indices based on the n-alkane standards from C8&#x2013;C32 with data reported in the Wiley and NIST databases. The retention index was calculated as per the retention index system proposed by <xref ref-type="bibr" rid="B39">Van Den Dool and Kratz (1963)</xref> using the following formula for temperature-programmed retention index:</p>
<disp-formula>
<mml:math display="block" id="M2">
<mml:mrow>
<mml:mtext>Ix</mml:mtext>
<mml:mo>=</mml:mo>
<mml:mn>100</mml:mn>
<mml:mrow>
<mml:mo>[</mml:mo>
<mml:mrow>
<mml:mfrac>
<mml:mrow>
<mml:mrow>
<mml:mo stretchy="false">(</mml:mo>
<mml:mrow>
<mml:mi>t</mml:mi>
<mml:mi>x</mml:mi>
<mml:mo>&#xa0;</mml:mo>
<mml:mo>&#x2212;</mml:mo>
<mml:mi>t</mml:mi>
<mml:mi>n</mml:mi>
</mml:mrow>
<mml:mo stretchy="false">)</mml:mo>
</mml:mrow>
</mml:mrow>
<mml:mrow>
<mml:mrow>
<mml:mo stretchy="false">(</mml:mo>
<mml:mrow>
<mml:mi>t</mml:mi>
<mml:mrow>
<mml:mo stretchy="false">(</mml:mo>
<mml:mrow>
<mml:mi>n</mml:mi>
<mml:mo>+</mml:mo>
<mml:mn>1</mml:mn>
</mml:mrow>
<mml:mo stretchy="false">)</mml:mo>
</mml:mrow>
<mml:mo>&#x2212;</mml:mo>
<mml:mi>t</mml:mi>
<mml:mi>n</mml:mi>
</mml:mrow>
<mml:mo stretchy="false">)</mml:mo>
</mml:mrow>
</mml:mrow>
</mml:mfrac>
<mml:mtext>&#xa0;</mml:mtext>
<mml:mo>+</mml:mo>
<mml:mi>n</mml:mi>
</mml:mrow>
<mml:mo>]</mml:mo>
</mml:mrow>
</mml:mrow>
</mml:math>
</disp-formula>
<p>where <italic>I<sub>x</sub>
</italic> is the temperature-programmed retention index, and <italic>t<sub>n</sub>
</italic>, <italic>t<sub>n</sub>
</italic>
<sub>+1</sub>, and <italic>t<sub>x</sub>
</italic> are the retention time (in min) of the two <italic>n</italic>-alkanes containing <italic>n</italic> and <italic>n</italic> + 1 carbons and the compound of interest, respectively.</p>
</sec>
<sec id="s2_6">
<title>Homology modeling</title>
<p>The acetylcholinesterase and GABA receptor subunit protein and octopamine receptor sequences of <italic>A. aegypti</italic> were retrieved in FASTA format from the National Center for Biotechnology Information (<ext-link ext-link-type="uri" xlink:href="https://www.ncbi.nlm.nih.gov/protein">https://www.ncbi.nlm.nih.gov/protein</ext-link>) with accession number AAB35001.1, AAA68961.1, and XP021692997.1, respectively. The protein&#x2019;s three-dimensional structure was computationally generated with the homology modeling technique as described by the developer (<xref ref-type="bibr" rid="B43">Waterhouse et&#xa0;al., 2018</xref>).</p>
</sec>
<sec id="s2_7">
<title>Molecular docking study</title>
<p>All molecular structures of phytochemicals reported in various fractions during LC-MS analysis were acquired from the PubChem database (<ext-link ext-link-type="uri" xlink:href="https://pubchem.ncbi.nlm.nih.gov">https://pubchem.ncbi.nlm.nih.gov</ext-link>). The ligand library included natural substrates and established inhibitors. The protein structures and ligands were transformed to pdbqt format using Openbabel software, and the energy was minimized by implementing the Autodock Vina Tool&#x2019;s Python script. Autodock Vina 1.2.0 was used for docking following the developers&#x2019; default parameters (<xref ref-type="bibr" rid="B11">Eberhardt et&#xa0;al., 2021</xref>). The software iGEMDOCK version 2.1 was employed to investigate postdocking ligand interactions (<xref ref-type="bibr" rid="B19">Hsu et&#xa0;al., 2011</xref>).</p>
<p>To model the target sequence, ProMod3 3.2.1 was used, and the SWISS-MODEL template library (SMTL version 2023-04-05, PDB release 2023-03-31) was searched for evolution-related structures that matched the target sequence.</p>
<p>The 6huj, 6xyy, and 2z73 PDB structures were used as a template for acetylcholinesterase, GABA receptor subunit, and octopamine receptor, respectively. The obtained model had 94.74%, 93.29%, and 79.88% amino acid residue in the Ramachandran plot favored region.</p>
</sec>
</sec>
<sec id="s3" sec-type="results">
<title>Results</title>
<sec id="s3_1">
<title>Larvicidal properties against <italic>A. aegypti</italic>
</title>
<p>The larvicidal activity was examined in the range of 164&#x2013;284 &#xb5;g mL<sup>&#x2212;1</sup> (<xref ref-type="fig" rid="f1">
<bold>Figure&#xa0;1</bold>
</xref>). Minimum mortality was observed at the concentration of 164 &#xb5;g mL<sup>&#x2212;1</sup> (10%/h), while maximum mortality was observed at the concentration of 276 &#xb5;g mL<sup>&#x2212;1</sup> (50%/h). The lethal concentrations LC<sub>20</sub>, LC<sub>50</sub>, and LC<sub>90</sub> were 197.38 &#xb5;g mL<sup>&#x2212;1</sup>, 294.13 &#xb5;g mL<sup>&#x2212;1</sup>, and 540.15 &#xb5;g mL<sup>&#x2212;1</sup>, respectively (&#x3c7;2 = 0.905, <italic>p</italic>&lt; 0.001) (<xref ref-type="table" rid="T1">
<bold>Table&#xa0;1</bold>
</xref>). There was no significant mortality observed in the control group treated with only hexane solvent (<xref ref-type="fig" rid="f1">
<bold>Figure&#xa0;1</bold>
</xref>).</p>
<fig id="f1" position="float">
<label>Figure&#xa0;1</label>
<caption>
<p>Mortality at different concentrations of <italic>Commiphora wightii</italic> gum hexane extracts against the fourth instar stage of <italic>Aedes aegypti</italic> larvae.</p>
</caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fpls-14-1220339-g001.tif"/>
</fig>
<table-wrap id="T1" position="float">
<label>Table&#xa0;1</label>
<caption>
<p>Mosquito larvicidal effectiveness of hexane gum extract against <italic>Aedes aegypti</italic> (95% confidence interval) (***<italic>p&lt;</italic> 0.001).</p>
</caption>
<table frame="hsides">
<thead>
<tr>
<th valign="top" align="left">Sr. No.</th>
<th valign="top" align="center">Hexane gum extract</th>
<th valign="top" align="center">LC<sub>20</sub> in &#xb5;g mL<sup>&#x2212;1</sup>
</th>
<th valign="top" align="center">LC<sub>50</sub> in &#xb5;g mL<sup>&#x2212;1</sup>
</th>
<th valign="top" align="center">LC<sub>90</sub> in &#xb5;g mL<sup>&#x2212;1</sup>
</th>
<th valign="top" align="center">Chi-square<break/>&#x3c7;<sup>2</sup>
</th>
</tr>
</thead>
<tbody>
<tr>
<td valign="middle" align="left"/>
<td valign="middle" align="center">
<italic>Commiphora wightii</italic>
</td>
<td valign="middle" align="center">197.38***</td>
<td valign="middle" align="center">294.13***</td>
<td valign="middle" align="center">540.15***</td>
<td valign="middle" align="center">0.905</td>
</tr>
</tbody>
</table>
</table-wrap>
</sec>
<sec id="s3_2">
<title>Identification of volatile compounds from plant gum extracts prepared in hexane</title>
<p>Headspace GC/MS analysis was conducted for the hexane gum extract of <italic>C. wightii</italic>, which revealed the following types of volatile compounds: monoterpenes [<italic>&#x3b1;</italic>-Pinene (11.725%), 2-Norpinene 3,6,6-trimethyl- (10.607%), <italic>&#x3b1;</italic>-Ocimene (2.309%), <italic>&#x3b1;</italic>-Myrcene (5.289%), Limonene (5.324%), <italic>&#x3b1;</italic>-Terpineol (1.056%), <italic>&#x3b2;</italic>-Acoradiene (2.262%), and Sesquisabinene (1.140%)], sesquiterpenes [Caryophyllene (6.128%), <italic>&#x3b2;</italic>-Elemene (2.381%), <italic>&#x3b1;</italic>-Curcumene (3.961%), Sesquithujene (1.693%), and <italic>&#x3b1;</italic>-Bisabolene (1.033%)], and diterpenes [Cembrene (1.757%) and Neocembrene (9.950%)] (<xref ref-type="supplementary-material" rid="SM1">
<bold>Supplementary Figure S1</bold>
</xref>; <xref ref-type="table" rid="T2">
<bold>Table&#xa0;2</bold>
</xref>). The refraction index of all identified compounds was calculated as per the retention index system proposed by <xref ref-type="bibr" rid="B39">Van Den Dool and Kratz (1963)</xref>. The calculated retention indices of each identified compound were compared with the retention indices available in the NIST Chemistry WebBook, SRD 69 (<xref ref-type="table" rid="T2">
<bold>Table&#xa0;2</bold>
</xref>). Previous studies have reported the presence of phytoconstituent groups such as monoterpenoids, sesquiterpenoids, diterpenoids, and triterpenoids; steroids; flavonoids; guggultetrols; lignans; sugars; and amino acids in the resin of <italic>C. wightii</italic>. Specific phytochemicals, <italic>viz</italic>., monoterpene (Limonene, <italic>&#x3b1;</italic>-Terpineol, <italic>&#x3b1;</italic>-Pinene, and <italic>&#x3b1;</italic>-Myrcene) and diterpene (Cembrene), have been reported in the gum resin of <italic>C. wightii</italic> (<xref ref-type="bibr" rid="B33">Sarup et&#xa0;al., 2015</xref>; <xref ref-type="bibr" rid="B9">Chandran et&#xa0;al., 2020</xref>).</p>
<table-wrap id="T2" position="float">
<label>Table&#xa0;2</label>
<caption>
<p>Volatile compounds from the hexane extract of <italic>Commiphora wightii</italic> gum.</p>
</caption>
<table frame="hsides">
<thead>
<tr>
<th valign="middle" align="center">Type of volatile compound</th>
<th valign="middle" align="center">Volatile compound</th>
<th valign="middle" align="center">RT</th>
<th valign="middle" align="center">Peak area %</th>
<th valign="top" align="center">RI (calculated)</th>
<th valign="top" align="center">RI<break/>(NIST webbook database)</th>
</tr>
</thead>
<tbody>
<tr>
<td valign="middle" rowspan="2" align="center">
<bold>Diterpene</bold>
</td>
<td valign="middle" align="center">Cembrene</td>
<td valign="middle" align="center">21.129</td>
<td valign="middle" align="center">1.757</td>
<td valign="top" align="center">1,941</td>
<td valign="top" align="center">1,930</td>
</tr>
<tr>
<td valign="middle" align="center">Neocembrene</td>
<td valign="middle" align="center">21.518</td>
<td valign="middle" align="center">9.950</td>
<td valign="top" align="center">1,954</td>
<td valign="top" align="center">1,960</td>
</tr>
<tr>
<td valign="middle" rowspan="7" align="center">
<bold>Monoterpene</bold>
</td>
<td valign="middle" align="center">2-Norpinene, 3,6,6-trimethyl-</td>
<td valign="middle" align="center">3.517</td>
<td valign="middle" align="center">10.607</td>
<td valign="top" align="center">916</td>
<td valign="top" align="center">Not available in database</td>
</tr>
<tr>
<td valign="middle" align="center">
<italic>&#x3b1;</italic>-Pinene</td>
<td valign="middle" align="center">3.406</td>
<td valign="middle" align="center">11.725</td>
<td valign="top" align="center">907</td>
<td valign="top" align="center">909</td>
</tr>
<tr>
<td valign="middle" align="center">Limonene</td>
<td valign="middle" align="center">5.103</td>
<td valign="middle" align="center">5.324</td>
<td valign="top" align="center">1,032</td>
<td valign="top" align="center">1,039</td>
</tr>
<tr>
<td valign="middle" align="center">Sesquisabinene</td>
<td valign="middle" align="center">13.190</td>
<td valign="middle" align="center">1.140</td>
<td valign="top" align="center">1,463</td>
<td valign="top" align="center">1,461</td>
</tr>
<tr>
<td valign="middle" align="center">
<italic>&#x3b1;</italic>-Ocimene</td>
<td valign="middle" align="center">4.097</td>
<td valign="middle" align="center">2.309</td>
<td valign="top" align="center">809</td>
<td valign="top" align="center">805</td>
</tr>
<tr>
<td valign="middle" align="center">
<italic>&#x3b1;</italic>-Myrcene</td>
<td valign="middle" align="center">4.313</td>
<td valign="middle" align="center">5.289</td>
<td valign="top" align="center">981</td>
<td valign="top" align="center">986</td>
</tr>
<tr>
<td valign="middle" align="center">
<italic>&#x3b2;</italic>-Acoradiene</td>
<td valign="middle" align="center">11.821</td>
<td valign="middle" align="center">2.262</td>
<td valign="top" align="center">1,454</td>
<td valign="top" align="center">1,462</td>
</tr>
<tr>
<td valign="middle" align="center">
<bold>Monoterpene alcohol</bold>
</td>
<td valign="middle" align="center">
<italic>&#x3b1;</italic>-Terpineol</td>
<td valign="middle" align="center">7.860</td>
<td valign="middle" align="center">1.056</td>
<td valign="top" align="center">1,184</td>
<td valign="top" align="center">1,172</td>
</tr>
<tr>
<td valign="middle" rowspan="5" align="center">
<bold>Sesquiterpene</bold>
</td>
<td valign="middle" align="center">
<italic>&#x3b2;</italic>-Elemene</td>
<td valign="middle" align="center">10.630</td>
<td valign="middle" align="center">2.381</td>
<td valign="top" align="center">1,362</td>
<td valign="top" align="center">1,366</td>
</tr>
<tr>
<td valign="middle" align="center">Caryophyllene</td>
<td valign="middle" align="center">11.179</td>
<td valign="middle" align="center">6.128</td>
<td valign="top" align="center">1,456</td>
<td valign="top" align="center">1,467</td>
</tr>
<tr>
<td valign="middle" align="center">
<italic>&#x3b1;</italic>-Curcumene</td>
<td valign="middle" align="center">12.228</td>
<td valign="middle" align="center">3.961</td>
<td valign="top" align="center">1,460</td>
<td valign="top" align="center">1,469</td>
</tr>
<tr>
<td valign="middle" align="center">Sesquithujene</td>
<td valign="middle" align="center">12.511</td>
<td valign="middle" align="center">1.693</td>
<td valign="top" align="center">1,410</td>
<td valign="top" align="center">1,413</td>
</tr>
<tr>
<td valign="middle" align="center">
<italic>&#x3b1;</italic>-Bisabolene</td>
<td valign="middle" align="center">12.801</td>
<td valign="middle" align="center">1.033</td>
<td valign="top" align="center">1,446</td>
<td valign="top" align="center">1,443</td>
</tr>
</tbody>
</table>
</table-wrap>
</sec>
<sec id="s3_3">
<title>Molecular docking study</title>
<sec id="s3_3_1">
<title>Acetylcholinesterase molecular docking</title>
<p>Acetylcholinesterase was docked against 15 unique phytochemicals, including the substrate acetylcholine and the inhibitor carbamate insecticide propoxur. The docking binding energy for phytochemicals ranged between -5.4 and -8.2. The binding energy of the substrate acetylcholine and the inhibitor carbamate insecticide propoxur was -4.5 and -7.8, respectively. The least binding energy was obtained for <italic>&#x3b1;</italic>-Bisabolene (-8.2), Caryophyllene (-8.1), and <italic>&#x3b2;</italic>-Acoradiene (-7.9), followed by other compounds (<xref ref-type="fig" rid="f2">
<bold>Figure&#xa0;2</bold>
</xref>). Acetylcholinesterase and docked phytochemical interaction analysis displayed diverse interaction profiles into five clusters. Acetylcholine and the inhibitor propoxur have been classified into different clusters. The three highest ranking <italic>&#x3b1;</italic>-Bisabolene, Caryophyllene, and <italic>&#x3b2;</italic>-Acoradiene had similar interaction profiles and shared a common bond pattern with substrate acetylcholine and inhibitor propoxur, indicating the potential of inhibition activity at the molecular level along with the obtained best docking score (<xref ref-type="fig" rid="f3">
<bold>Figure&#xa0;3</bold>
</xref>).</p>
<fig id="f2" position="float">
<label>Figure&#xa0;2</label>
<caption>
<p>Molecular docking binding energy of Commiphora wightii phytochemicals against the acetylcholinesterase receptor.</p>
</caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fpls-14-1220339-g002.tif"/>
</fig>
<fig id="f3" position="float">
<label>Figure&#xa0;3</label>
<caption>
<p>Interaction profile of <italic>Commiphora wightii</italic> phytochemicals best pose against the acetylcholinesterase receptor. Numbers in the bracket indicate the docking rank, black color indicates zero value, and green color indicates negative energy.</p>
</caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fpls-14-1220339-g003.tif"/>
</fig>
</sec>
<sec id="s3_3_2">
<title>GABA receptor molecular docking</title>
<p>In addition to 15 phytochemicals for GABA receptors, natural substrate Gamma-Aminobutyric Acid and organochlorine insecticide endosulfan were also identified. The docking binding energy for phytochemicals ranged between -2.5 and -7.4. The minimum docking binding energy was obtained for Cembrene (-7.4), Bisabolene (-7.2), and Neocembrene (-7.2), followed by other compounds (<xref ref-type="fig" rid="f4">
<bold>Figure&#xa0;4</bold>
</xref>). The interaction analysis for GABA receptor molecular docking against phytochemicals revealed four distinct clusters (<xref ref-type="fig" rid="f5">
<bold>Figure&#xa0;5</bold>
</xref>). Substrate Gamma-Aminobutyric Acid was grouped in a unique cluster with a single member, and the inhibitor endosulfan was grouped in a different cluster with three other phytochemicals. Cembrene, the top-ranking compound, was also classified in a unique cluster with only one member, but it shared a few binding interactions with the inhibitor endosulfan. Conversely, Neocembrene, which is ranked second, pertains to the endosulfan cluster and exhibits a comparable binding interaction. On the contrary, the third compound with the highest rank, Bisabolene, exhibited a binding interaction that was comparable to that of Gamma-Aminobutyric Acid and was also found to be in close proximity (<xref ref-type="fig" rid="f5">
<bold>Figure&#xa0;5</bold>
</xref>).</p>
<fig id="f4" position="float">
<label>Figure&#xa0;4</label>
<caption>
<p>Molecular docking binding energy of <italic>Commiphora wightii</italic> phytochemicals against the GABA receptor.</p>
</caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fpls-14-1220339-g004.tif"/>
</fig>
<fig id="f5" position="float">
<label>Figure&#xa0;5</label>
<caption>
<p>Interaction profile of <italic>Commiphora wightii</italic> phytochemicals best pose against the GABA receptor. Numbers in the bracket indicate the docking rank, black color indicates zero value, and green color indicates negative energy.</p>
</caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fpls-14-1220339-g005.tif"/>
</fig>
</sec>
</sec>
<sec id="s3_4">
<title>Octopamine receptor docking</title>
<p>For the octopamine receptor protein docking experiment, the binding energy ranged from &#x2013;4.5 to -6.8 (<xref ref-type="fig" rid="f6">
<bold>Figure&#xa0;6</bold>
</xref>). The overall binding interaction for octopamine was relatively lower compared to that of acetylcholinesterase and GABA receptor, which may be due to the lower quality of the model. The minimum docking binding energy was obtained for 2-Norpinene, 3,6,6-trimethyl (-6.8), <italic>&#x3b1;</italic>-Bisabolene (-6.4), and Cembrene (-6.4), followed by other compounds (<xref ref-type="fig" rid="f6">
<bold>Figure&#xa0;6</bold>
</xref>). The interaction analysis for Octopamine receptor molecular docking against phytochemicals revealed four clusters, generated with a single member of with first-ranking 2-Norpinene, 3,6,6-trimethyl, <italic>&#x3b1;</italic>-Curcumene, <italic>&#x3b2;</italic>-Elemene, and inhibitor Formamidine hydrochloride. Second- and third-ranking compounds <italic>&#x3b1;</italic>-Bisabolene and Cembrene are also assorted in a separate group (<xref ref-type="fig" rid="f7">
<bold>Figure&#xa0;7</bold>
</xref>). In contrast, substrate octopamine was grouped together with the other three phytochemicals with a lower rank. The substrate octopamine had a lower binding energy of -5.4 when compared to the -2.5 of acaricide insecticide Formamidine hydrochloride.</p>
<fig id="f6" position="float">
<label>Figure&#xa0;6</label>
<caption>
<p>Molecular docking binding energy of <italic>Commiphora wightii</italic> phytochemicals against the octopamine receptor.</p>
</caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fpls-14-1220339-g006.tif"/>
</fig>
<fig id="f7" position="float">
<label>Figure&#xa0;7</label>
<caption>
<p>Interaction profile of <italic>Commiphora wightii</italic> phytochemicals best pose against the octopamine receptor. Numbers in the bracket indicate the docking rank, black color indicates zero value, and green color indicates negative energy.</p>
</caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fpls-14-1220339-g007.tif"/>
</fig>
<p>Grouping of the three top-ranking compounds against generated leads the group of a total of six compounds, <italic>&#x3b1;</italic>-Bisabolene, Caryophyllene, Neocembrene, <italic>&#x3b2;</italic>-Acoradiene, 2-Norpinene, and 3,6,6-trimethyl that had better binding energy in comparison to their natural substrate and commercial inhibitor.</p>
</sec>
</sec>
<sec id="s4" sec-type="discussion">
<title>Discussion</title>
<p>Since ancient times, extracts prepared from different parts of plants like flowers, bark, roots, leaves, and seeds have been used as insecticides (<xref ref-type="bibr" rid="B36">Sir et&#xa0;al., 2020</xref>). Various studies have reported the efficacy of plant extracts against mosquitoes. Studies have proven that these natural extracts have certain compounds that help in controlling the pest population (<xref ref-type="bibr" rid="B21">Jamio&#x142;kowska and Kopacki, 2020</xref>). These extracts have certain volatile compounds that can act on insects as insect repellents, insecticides, insect antifeedants, insect growth inhibitors, acetylcholinesterase enzyme inhibitors, oviposition inhibitors, ovicides, or insect growth-reducing agents (<xref ref-type="bibr" rid="B18">Hikal et&#xa0;al., 2017</xref>).</p>
<p>Numerous studies have reported the larvicidal activity of <italic>Commiphora</italic> species. The larvicidal activity of the following <italic>Commiphora</italic> species against different species of mosquitoes has been reported that includes oil-resins of <italic>Commiphora molmol</italic> (1,500 ppm acetone and aqueous extracts) against mosquito larvae of <italic>Culex pipiens</italic> (<xref ref-type="bibr" rid="B5">Baz et&#xa0;al., 2021</xref>); arabinofuranosidetridecanol from <italic>C. merkeri</italic> against <italic>A. aegypti</italic> (LC<sub>50</sub> = 40.66&#x2009; &#xb5;g mL<sup>&#x2212;1</sup>), <italic>Anopheles gambiae</italic> (LC<sub>50</sub> = 22.86&#x2009; &#xb5;g mL<sup>&#x2212;1</sup>), and <italic>C. quinquefasciatus</italic> (LC<sub>50</sub> = 15.88&#x2009; &#xb5;g mL<sup>&#x2212;1</sup>) (<xref ref-type="bibr" rid="B32">Samwel et&#xa0;al., 2021</xref>); essential oil of <italic>C. erythraea</italic> against <italic>C. restuans</italic>, <italic>C. pipiens</italic>, and <italic>A. aegypti</italic> (<xref ref-type="bibr" rid="B29">Muturi et&#xa0;al., 2020</xref>). As per the researchers, LC<sub>50</sub>&lt;50 &#xb5;g mL<sup>&#x2212;1</sup> is very active; LC<sub>50</sub> 50 &#xb5;g mL<sup>&#x2212;1</sup>&#x2013;100 &#xb5;g mL<sup>&#x2212;1</sup> is active, and LC<sub>50</sub> &gt;100 &#xb5;g mL<sup>&#x2212;1</sup> is a weak or inactive larvicidal compound (<xref ref-type="bibr" rid="B28">Milugo et&#xa0;al., 2021</xref>).</p>
<p>It has been suggested that monoterpenoids including Terpineol are very effective in inhibiting insect reproduction (<xref ref-type="bibr" rid="B31">Regnault-Roger et&#xa0;al., 2004</xref>), and the occurrence of monoterpenes and sesquiterpenes leads to fumigant toxicity (<xref ref-type="bibr" rid="B15">Hamraoui and Regnault-Roger, 1997</xref>; <xref ref-type="bibr" rid="B31">Regnault-Roger et&#xa0;al., 2004</xref>). Gum extracts are known as potential sources of larvicides due to their ability to alter the activity of the insect&#x2019;s acetylcholinesterase enzyme (<xref ref-type="bibr" rid="B2">Ashokkumar et&#xa0;al., 2021</xref>), which is required for neuro-neuronal and neuromuscular junctions in insects. Among these, Limonene and <italic>&#x3b1;</italic>-Pinene are the most effective compounds that alter the activity of the acetylcholinesterase enzyme (<xref ref-type="bibr" rid="B26">Marei et&#xa0;al., 2012</xref>; <xref ref-type="bibr" rid="B41">Vieira et&#xa0;al., 2018</xref>).</p>
<p>Monoterpenes have natural pesticidal properties that are effective, safe, and biodegradable, which make them the best alternative to synthetic pesticides. Various pesticidal activities of monoterpenes, including bactericidal (<xref ref-type="bibr" rid="B3">Badawy et&#xa0;al., 2019</xref>), fungicidal, herbicidal (<xref ref-type="bibr" rid="B25">Liu et&#xa0;al., 2022</xref>), and insecticidal properties (<xref ref-type="bibr" rid="B12">Gad et&#xa0;al., 2022</xref>), are known. The larvicidal activity of monoterpenes against certain mosquito species (<xref ref-type="bibr" rid="B12">Gad et&#xa0;al., 2022</xref>), including toxic effects on <italic>A. aegypti</italic> (<xref ref-type="bibr" rid="B13">Giang An et&#xa0;al., 2020</xref>) and anti-oviposition effects on <italic>A. aegypti</italic> (<xref ref-type="bibr" rid="B42">Waliwitiya et&#xa0;al., 2009</xref>), has also been reported. It has been observed that monoterpenes and terpenoids are efficient acetylcholinesterase enzyme inhibitors in different insects and larvae (<xref ref-type="bibr" rid="B42">Waliwitiya et&#xa0;al., 2009</xref>), whereas diterpenes like Cembrene have antifeed activity against herbivores (<xref ref-type="bibr" rid="B37">Sun et&#xa0;al., 2019</xref>).</p>
<p>Previous research conducted on 24 species of the genus <italic>Commiphora</italic>, including <italic>C. wightii</italic>, identified 230 phytoconstituents, including phytoconstituents <italic>&#x3b1;</italic>-Bisabolene, Caryophyllene, Cembrene, Limonene, <italic>&#x3b1;</italic>-Pinene, <italic>&#x3b2;</italic>-Elemene, <italic>&#x3b1;</italic>-Myrcene, and <italic>&#x3b1;</italic>-Ocimene (<xref ref-type="bibr" rid="B34">Selvamani et&#xa0;al., 2008</xref>), which have also been obtained in the present study. However, Sesquisabinene and 2-Norpinene, 3,6,6-Trimethyl have not been reported from any <italic>Commiphora</italic> species so far.</p>
<p>The research conducted on five <italic>Piper</italic> species (<italic>Piper nigrum</italic>, <italic>Piper mutabile</italic>, <italic>Piper longum</italic>, <italic>Piper montium</italic>, and <italic>Piper caninum</italic>) showed excellent larvicidal activities with LC<sub>50</sub> and LC<sub>90</sub> values&lt;10 &#x3bc;g mL<sup>-1</sup> and were correlated with <italic>&#x3b2;</italic>-Caryophyllene, <italic>&#x3b2;</italic>-Bisabolene, <italic>&#x3b2;</italic>-Pinene, and <italic>&#x3b1;</italic>-Pinene concentrations (<xref ref-type="bibr" rid="B20">Huong et&#xa0;al., 2019</xref>), which support our molecular docking findings for <italic>&#x3b2;</italic>-Caryophyllene and <italic>&#x3b2;</italic>-Bisabolene.</p>
<p>
<italic>&#x3b1;</italic>-Bisabolene was best docked against the acetylcholinesterase with binding energy &#x2212;8.2. It has been observed that niloticin (&#x2212;8.4 kcal/mol) had higher binding affinities and energy values than temephos (&#x2212;4.75 kcal/mol), which is a commercial larvicide (<xref ref-type="bibr" rid="B30">Reegan et&#xa0;al., 2016</xref>). Similarly, the inhibitory effect of natural alkaloids on the acetylcholinesterase present in <italic>A. aegypti</italic> had the best fit into the AChE1 binding pocket with a minimum binding energy of &#x2212;8.13 (<xref ref-type="bibr" rid="B4">Balachandran et&#xa0;al., 2021</xref>).</p>
<p>The gamma-aminobutyric acid receptor, also known as the GABA receptor, has four distinct but overlapping and coupled targets of pesticide action. These targets are associated with very little or no cross-resistance (<xref ref-type="bibr" rid="B35">Setlur et&#xa0;al., 2023</xref>). In the present research work, besides acetylcholinesterase, <italic>&#x3b1;</italic>-Bisabolene also ranked second and third for the octopamine receptor and GABA receptor, indicating its possibility to act as a multitarget ligand inhibiting the nervous system. Thus, <italic>&#x3b1;</italic>-Bisabolene emerged as one of the potent multitarget inhibitors for application as a bio-insecticide to control the resistant <italic>A. aegypti.</italic>
</p>
<p>Bisabolene, a plant-based Sesquiterpene, is used as a precursor for the synthesis of several industrially relevant chemicals (<xref ref-type="bibr" rid="B45">Zhao et&#xa0;al., 2021</xref>). Bisabolene&#x2019;s low plant abundance makes its isolation uneconomical. Therefore, industrial Bisabolene production using synthetic biology and metabolic engineering to create microbial cell factories is more competitive and environmentally sustainable. In this proof-of-principle engineering, the oleaginous yeast <italic>Yarrowia lipolytica</italic> was explored to produce <italic>&#x3b1;</italic>- and <italic>&#x3b2;</italic>-Bisabolene by heterologously expressing the synthase genes from <italic>Abies grandis</italic>, <italic>Zingiber officinale</italic>, and <italic>Helianthus annuus</italic>.</p>
<p>The current research findings suggest that <italic>C. wightii</italic> could be beneficial as a biological agent in the fight against <italic>A. aegypti</italic>. It was found that Bisabolene could be one of the key larvicidal compounds that employ a multitarget strategy and act on proteins associated with the nervous system of <italic>A. aegypti</italic> larvae. Even in cases where <italic>A. aegypti</italic> larva has developed resistance to commercially used pesticides, being a natural compound, Bisabolene can be effective against them.</p>
</sec>
<sec id="s5" sec-type="conclusions">
<title>Conclusion</title>
<p>
<italic>A. aegypti</italic> mosquito, also known as the yellow fever mosquito, is a potent transmitter of several vector-borne diseases such as dengue fever, chikungunya, Zika fever, the Mayaro virus, and yellow fever, in addition to other pathogens. Guggulu, also known as <italic>C. wightii</italic>, has recently been recognized as a reliable source of traditional medicines for the treatment of a variety of conditions, including inflammation, arthritis, obesity, microbial infection, wounds, pain, fractures, tumors, and gastrointestinal diseases. The current investigation posed an additional application of <italic>C. wightii</italic> gum resin for the environment-friendly biological control of <italic>A. aegypti</italic> larva (fourth instar) due to its multitarget action. A total of 15 volatile compounds were identified belonging to diterpene, monoterpene, monoterpene alcohol, and sesquiterpene groups. A total of 230 phytoconstituents have been identified from <italic>Commiphora</italic> species, out of which five unique compounds were identified from <italic>C. wightii</italic>, which were Limonene, <italic>&#x3b1;</italic>-Terpineol, <italic>&#x3b1;</italic>-Pinene, <italic>&#x3b1;</italic>-Myrcene, and Cembrene. The molecular docking study of acetylcholinesterase, GABA receptor, and octopamine receptor revealed certain compounds of the <italic>C. wightii</italic> gum resin having very low binding energy, revealing their role in larvicidal properties. These compounds are known to act upon the nervous system leading to the ultimate death of the larvae.</p>
</sec>
<sec id="s6" sec-type="data-availability">
<title>Data availability statement</title>
<p>The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/<xref ref-type="supplementary-material" rid="SM1">
<bold>Supplementary Material</bold>
</xref>.</p>
</sec>
<sec id="s7" sec-type="ethics-statement">
<title>Ethics statement</title>
<p>The article presents research on animals that do not require ethical approval for their study.</p>
</sec>
<sec id="s8" sec-type="author-contributions">
<title>Author contributions</title>
<p>Conceptualization and supervision: AP, JGT, VY, and DS. Investigation and methodology: KP, DA, RVP, VM, and SW. Original draft preparation: AP, VY, KP, and JGT. Review and final editing: RP, AP, JGT, and JT. All authors contributed to the article and approved the submitted version.</p>
</sec>
</body>
<back>
<ack>
<title>Acknowledgments</title>
<p>The authors are grateful to the Department of Life Sciences, Hemchandracharya North Gujarat University, Patan, Gujarat, India, for providing the laboratory facilities to carry out the research work.</p>
</ack>
<sec id="s9" sec-type="COI-statement">
<title>Conflict of interest</title>
<p>The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.</p>
</sec>
<sec id="s10" sec-type="disclaimer">
<title>Publisher&#x2019;s note</title>
<p>All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.</p>
</sec>
<sec id="s11" sec-type="supplementary-material">
<title>Supplementary material</title>
<p>The Supplementary Material for this article can be found online at: <ext-link ext-link-type="uri" xlink:href="https://www.frontiersin.org/articles/10.3389/fpls.2023.1220339/full#supplementary-material">https://www.frontiersin.org/articles/10.3389/fpls.2023.1220339/full#supplementary-material</ext-link>
</p>
<supplementary-material xlink:href="Image_1.tif" id="SM1" mimetype="image/tiff"/>
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