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<!DOCTYPE article PUBLIC "-//NLM//DTD JATS (Z39.96) Journal Publishing DTD v1.1d1 20130915//EN" "http://jats.nlm.nih.gov/publishing/1.1d1/JATS-journalpublishing1.dtd">
<article xmlns:xlink="http://www.w3.org/1999/xlink" xmlns:mml="http://www.w3.org/1998/Math/MathML" article-type="research-article" xml:lang="en">
<front>
<journal-meta>
<journal-id journal-id-type="publisher-id">Bothalia</journal-id>
<journal-title-group>
<journal-title>Bothalia African Biodiversity &#x0026; Conservation</journal-title>
</journal-title-group>
<issn pub-type="ppub">0006-8241</issn>
<issn pub-type="epub">2311-9284</issn>
<publisher>
<publisher-name>AOSIS</publisher-name>
</publisher>
</journal-meta>
<article-meta>
<article-id pub-id-type="publisher-id">ABC-49-2429</article-id>
<article-id pub-id-type="doi">10.4102/abc.v49i1.2429</article-id>
<article-categories>
<subj-group subj-group-type="heading">
<subject>New Distribution Record</subject>
</subj-group>
</article-categories>
<title-group>
<article-title>Three new Drosophilidae species records for South Africa</article-title>
</title-group>
<contrib-group>
<contrib contrib-type="author">
<contrib-id contrib-id-type="orcid">https://orcid.org/0000-0002-8468-9778</contrib-id>
<name>
<surname>De Araujo</surname>
<given-names>Liana I.</given-names>
</name>
<xref ref-type="aff" rid="AF0001">1</xref>
</contrib>
<contrib contrib-type="author" corresp="yes">
<contrib-id contrib-id-type="orcid">https://orcid.org/0000-0002-5143-7312</contrib-id>
<name>
<surname>Karsten</surname>
<given-names>Minette</given-names>
</name>
<xref ref-type="aff" rid="AF0001">1</xref>
</contrib>
<contrib contrib-type="author">
<contrib-id contrib-id-type="orcid">https://orcid.org/0000-0001-9665-9405</contrib-id>
<name>
<surname>Terblanche</surname>
<given-names>John S.</given-names>
</name>
<xref ref-type="aff" rid="AF0001">1</xref>
</contrib>
<aff id="AF0001"><label>1</label>Center for Invasion Biology, Department of Conservation Ecology and Entomology, Stellenbosch University, Cape Town, South Africa</aff>
</contrib-group>
<author-notes>
<corresp id="cor1"><bold>Corresponding author:</bold> Minette Karsten, <email xlink:href="minettek@sun.ac.za">minettek@sun.ac.za</email></corresp>
</author-notes>
<pub-date pub-type="epub"><day>26</day><month>06</month><year>2019</year></pub-date>
<pub-date pub-type="collection"><year>2019</year></pub-date>
<volume>49</volume>
<issue>1</issue>
<elocation-id>2429</elocation-id>
<history>
<date date-type="received"><day>12</day><month>12</month><year>2018</year></date>
<date date-type="accepted"><day>07</day><month>03</month><year>2019</year></date>
</history>
<permissions>
<copyright-statement>&#x00A9; 2019. The Authors</copyright-statement>
<copyright-year>2019</copyright-year>
<license license-type="open-access" xlink:href="https://creativecommons.org/licenses/by/4.0/">
<license-p>Licensee: AOSIS. This work is licensed under the Creative Commons Attribution License.</license-p>
</license>
</permissions>
<abstract>
<sec id="st1">
<title>Background</title>
<p>Data on the current species diversity from the Drosophilidae family in South Africa is limited or outdated.</p>
</sec>
<sec id="st2">
<title>Objectives</title>
<p>Using haphazard, limited trapping for a different study, we serendipitously report on and document Drosophilidae species in two distinct regions (representing a sub-tropical and a Mediterranean climate region) of South Africa.</p>
</sec>
<sec id="st3">
<title>Method</title>
<p>Drosophilidae were trapped using mixed fruit and mushroom traps around urban areas in two climatically distinct regions of South Africa. The flies were identified using standard barcoding (Cytochrome c Oxidase Subunit I [COI] gene sequence) and, in some cases, additional identification from a taxonomical expert using morphological traits. Species were checked against literature, online resources and a previously compiled library of South African Drosophilidae to determine whether they were new records.</p>
</sec>
<sec id="st4">
<title>Results</title>
<p>Thirteen species were readily collected and identified. Of these, three species (<italic>Drosophila ananassae, Drosophila nasuta</italic> and <italic>Zaprionus taronus</italic>) have not been reported previously in South Africa. One of the species (<italic>Z. taronus</italic>) was captured in a home garden, while the other two species were captured in an urban-agricultural region with a sub-tropical climate.</p>
</sec>
<sec id="st5">
<title>Conclusions</title>
<p>From our limited serendipitous sampling, three new species records have been found in sub-tropical climates in South Africa. With more comprehensive, systematic sampling, a better understanding of the South African Drosophilidae composition, and thus the detection of alien or invasive species, can be pursued. Baseline data for understanding spatio-temporal patterns of native biodiversity, or for informing management actions in the case of alien or invasive species, are currently inadequate for this group in the region.</p>
</sec>
</abstract>
<kwd-group>
<kwd>Drosophilidae</kwd>
<kwd><italic>Zaprionus</italic></kwd>
<kwd><italic>Drosophila</italic></kwd>
<kwd>COI</kwd>
<kwd>barcoding</kwd>
</kwd-group>
</article-meta>
</front>
<body>
<sec id="s0001">
<title>Background</title>
<p>Insects in the family Drosophilidae, commonly known as vinegar flies, are used as model systems for a diverse range of research fields, especially medicine (Hewitt &#x0026; Whitworth <xref ref-type="bibr" rid="CIT0005">2016</xref>). More recently, there has been growing concern surrounding invasive or pest Drosophilidae species in South Africa, as several invasive Drosophilidae species have been detected in other parts of the world and can target ripe soft-bodied fruit, resulting in economic losses for the agriculture sector (Farnsworth et al. <xref ref-type="bibr" rid="CIT0002">2017</xref>). Consequently, current species composition, and spatial and temporal patterns of Drosophilidae biodiversity are important baseline information. South African Drosophilidae are a case in point: there is surprisingly poor current knowledge of Drosophilidae species diversity, geographic range extent of common or rare species or how dynamic population abundances are in this group (McEvey, Potts &#x0026; Rogers <xref ref-type="bibr" rid="CIT0007">1988</xref>; Tsacas <xref ref-type="bibr" rid="CIT0009">1990</xref>). Furthermore, what little information is available is likely outdated or not necessarily representative of the different regions of the country. The most recent, scattered records show 76 species in the country documented between 1900 and 2013 in an unpublished document updated in 2016 and provided by Dr Shane McEvey (Australian Museum Research Institute), while the most recent accessible published data indicate 70 known species from South Africa (Tsacas <xref ref-type="bibr" rid="CIT0009">1990</xref>). From the unpublished document, more recent records include <italic>D. immigrans</italic> (2010), <italic>D. simulans</italic> (2013), <italic>D. punctatonervosa</italic> (2013) and <italic>Scaptomyza oxyphallus</italic> (2013). Although an invasive drosophilid species, <italic>D. flavohirta</italic> (1983), has been recorded in South Africa previously, no recent information regarding this species&#x2019; abundance or its potential impacts on native biodiversity is available (Tsacas <xref ref-type="bibr" rid="CIT0009">1990</xref>).</p>
</sec>
<sec id="s0002">
<title>Methods</title>
<p>Drosophilidae species were sampled sporadically in two urban-agricultural regions of South Africa using fruit-filled bucket traps (<xref ref-type="table" rid="T0001">Table 1</xref>) for a different project, but also aimed at generating biodiversity knowledge. Species were identified using the Universal cytochrome oxidase I (COI) primers (LCO1490 and HCO2198; Folmer et al. <xref ref-type="bibr" rid="CIT0003">1994</xref>) by InqabaBiotech<sup>TM</sup> and the South African Sugarcane Research Institute&#x2019;s (SASRI) biotechnology department. PCR conditions were an initial denaturation of 5 min at 94&#x00B0;C, followed by 35 cycles of 30 s denaturation at 94&#x00B0;C, 30 s annealing at 50&#x00B0;C and 60 s of extension at 68&#x00B0;C, and a final elongation at 68&#x00B0;C for 10 min. Sequences were aligned and edited in BioEdit (Hall <xref ref-type="bibr" rid="CIT0004">1999</xref>) and closest sequence matches obtained using BLAST (Altschul et al. <xref ref-type="bibr" rid="CIT0001">1997</xref>) in NCBI. Morphology of some species was confirmed against relevant species identification keys (Yassin &#x0026; David <xref ref-type="bibr" rid="CIT0011">2010</xref>). Positive matches were downloaded to include in a phylogenetic tree to confirm identification based on both neighbour-joining (NJ) and maximum likelihood (ML) methods using MEGA X (Kumar et al. <xref ref-type="bibr" rid="CIT0006">2018</xref>). The resulting trees were similar, and therefore we only show the tree based on ML (<xref ref-type="fig" rid="F0001">Figure 1</xref>).</p>
<fig id="F0001">
<label>FIGURE 1</label>
<caption><p>A phylogenetic tree based on maximum likelihood showing the relationships between the 13 Drosophilidae species collected in this study and reference specimens downloaded from National Center for Biotechnology Information (NCBI) (with accession numbers next to species names).The neighbour-joining tree has the same topology. Values on branches indicate bootstrap values (only values above 70&#x0025; shown) calculated over 1000 replicates.</p></caption>
<graphic xmlns:xlink="http://www.w3.org/1999/xlink" xlink:href="ABC-49-2429-g001.tif"/>
</fig>
<table-wrap id="T0001">
<label>TABLE 1</label>
<caption><p>The locations and description of collection sites, date of capture, method of identification (ID; by COI barcode [Sequence], by <italic>Drosophila</italic> expert [expert], identified by author using morphological keys [morphology]), bait type and National Center for Biotechnology Information (NCBI) accession numbers for the 13 Drosophilidae species collected in our study in South Africa.</p></caption>
<table frame="hsides" rules="groups">
<thead>
<tr>
<th valign="top" align="left">Species</th>
<th valign="top" align="left">Bait</th>
<th valign="top" align="center">Latitude (decimal degrees, &#x00B0;S)</th>
<th valign="top" align="center">Longitude (decimal degrees, &#x00B0;E)</th>
<th valign="top" align="left">Site description</th>
<th valign="top" align="left">Climate</th>
<th valign="top" align="center">Date of capture (dd/mm/yy)</th>
<th valign="top" align="left">ID</th>
<th valign="top" align="center">Sub-mission #</th>
<th valign="top" align="center">1st for Republic of South Africa</th>
</tr>
</thead>
<tbody valign="top">
<tr>
<td align="left"><bold><italic>Drosophila ananassae</italic></bold></td>
<td align="left"><bold>orange and lemon mix</bold></td>
<td align="center"><bold>&#x2212;29.7062</bold></td>
<td align="center"><bold>31.0444</bold></td>
<td align="left"><bold>Urban</bold></td>
<td align="left"><bold>Sub-tropical</bold></td>
<td align="center"><bold>22/03/18</bold></td>
<td align="left"><bold>Sequence</bold></td>
<td align="center"><bold>2172994</bold></td>
<td align="center"><bold>yes</bold></td>
</tr>
<tr>
<td align="left"><italic>Drosophila busckii</italic></td>
<td align="left">avocado</td>
<td align="center">&#x2212;33.8956</td>
<td align="center">18.5642</td>
<td align="left">Home garden</td>
<td align="left">Mediterranean</td>
<td align="center">02/11/17</td>
<td align="left">Sequence</td>
<td align="center">2172984</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Drosophila funebris</italic></td>
<td align="left">banana</td>
<td align="center">&#x2212;33.3465</td>
<td align="center">19.6232</td>
<td align="left">Mountain/ Nature Reserve</td>
<td align="left">Mediterranean</td>
<td align="center">09/01/18</td>
<td align="left">Sequence</td>
<td align="center">2172997</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Drosophila hydei</italic></td>
<td align="left">mushroom, banana, orange</td>
<td align="center">&#x2212;33.8956</td>
<td align="center">18.5642</td>
<td align="left">Home garden</td>
<td align="left">Mediterranean</td>
<td align="center">11/10/17</td>
<td align="left">Sequence, expert</td>
<td align="center">2172999</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Drosophila immigrans</italic></td>
<td align="left">banana</td>
<td align="center">&#x2212;33.8956</td>
<td align="center">18.5642</td>
<td align="left">Home garden</td>
<td align="left">Mediterranean</td>
<td align="center">02/11/17</td>
<td align="left">Sequence, expert</td>
<td align="center">2172876</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Drosophila melanogaster</italic></td>
<td align="left">banana, orange</td>
<td align="center">&#x2212;29.7013</td>
<td align="center">31.1009</td>
<td align="left">Coastal forest</td>
<td align="left">Sub-tropical</td>
<td align="center">08/06/18</td>
<td align="left">Sequence</td>
<td align="center">2172890</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Drosophila nasuta</italic></td>
<td align="left"><bold>orange and lemon mix</bold></td>
<td align="center"><bold>&#x2212;29.7062</bold></td>
<td align="center"><bold>31.0444</bold></td>
<td align="left"><bold>Urban</bold></td>
<td align="left"><bold>Sub-tropical</bold></td>
<td align="center"><bold>22/0318</bold></td>
<td align="left"><bold>Sequence</bold></td>
<td align="center"><bold>2173275</bold></td>
<td align="center"><bold>yes</bold></td>
</tr>
<tr>
<td align="left"><italic>Drosophila simulans</italic></td>
<td align="left">orange</td>
<td align="center">&#x2212;33.8956</td>
<td align="center">18.5642</td>
<td align="left">Home garden</td>
<td align="left">Mediterranean</td>
<td align="center">02/11/17</td>
<td align="left">Sequence, expert</td>
<td align="center">2173001</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Drosophila yakuba</italic></td>
<td align="left">orange and lemon mix, banana</td>
<td align="center">&#x2212;29.7062</td>
<td align="center">31.0444</td>
<td align="left">Urban</td>
<td align="left">Sub-tropical</td>
<td align="center">22/03/18</td>
<td align="left">Sequence</td>
<td align="center">2173281</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Zaprionus indianus</italic></td>
<td align="left">banana</td>
<td align="center">&#x2212;29.7062</td>
<td align="center">31.0444</td>
<td align="left">Urban</td>
<td align="left">Sub-tropical</td>
<td align="center">22/03/18</td>
<td align="left">Sequence</td>
<td align="center">2173002</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Zaprionus taronus</italic></td>
<td align="left"><bold>lemon</bold></td>
<td align="center"><bold>&#x2212;33.9353</bold></td>
<td align="center"><bold>18.8907</bold></td>
<td align="left"><bold>Home vegetable garden</bold></td>
<td align="left"><bold>Mediterranean</bold></td>
<td align="center"><bold>16/11/17</bold></td>
<td align="left"><bold>Sequence</bold></td>
<td align="center"><bold>2172892</bold></td>
<td align="center"><bold>yes</bold></td>
</tr>
<tr>
<td align="left"><italic>Zaprionus tuberculatus</italic></td>
<td align="left">banana</td>
<td align="center">&#x2212;29.7062</td>
<td align="center">31.0444</td>
<td align="left">Urban</td>
<td align="left">Sub-tropical</td>
<td align="center">22/05/18</td>
<td align="left">Sequence, morphology</td>
<td align="center">2173286</td>
<td align="center">no</td>
</tr>
<tr>
<td align="left"><italic>Zaprionus vittiger</italic></td>
<td align="left">mango</td>
<td align="center">&#x2212;33.9365</td>
<td align="center">18.8657</td>
<td align="left">Botanical garden</td>
<td align="left">Mediterranean</td>
<td align="center">20/10/17</td>
<td align="left">Sequence</td>
<td align="center">2173266</td>
<td align="center">no</td>
</tr>
</tbody>
</table>
<table-wrap-foot>
<fn><p>Note. The final column indicates whether this species has any previous records in South Africa by indicating a &#x2018;no&#x2019; if it has been recorded before and a &#x2018;yes&#x2019; if there is no previous record. The species with first records for South Africa are indicated in bold.</p></fn>
</table-wrap-foot>
</table-wrap>
</sec>
<sec id="s0003">
<title>Results</title>
<p>Once species were identified, it was evident that some species had no prior published records regarding their occurrence in South Africa (Tsacas <xref ref-type="bibr" rid="CIT0009">1990</xref>). TaxoDros was also consulted and showed no records for some species in South Africa. This was the case for three species, namely <italic>Drosophila ananassae</italic> (<xref ref-type="fig" rid="F0002">Figure 2a</xref>), <italic>D. nasuta</italic> (no image available) and <italic>Zaprionus taronus</italic> (<xref ref-type="fig" rid="F0002">Figure 2h</xref>). <italic>Zaprionus taronus</italic> has never before been reported from South Africa (McEvey, pers. comm.); however, this species is surrounded by ongoing taxonomic uncertainty. <italic>Zaprionus taronus</italic> is probably not invasive and more likely overlooked or previously mis-identified as it has a confirmed Afrotropical distribution (Yassin &#x0026; David <xref ref-type="bibr" rid="CIT0011">2010</xref>). The expertise of the person doing the identification, how the determination was made and what material was used for the determination are key information that needs to be established. Prigent, Suwalski and Veuille (<xref ref-type="bibr" rid="CIT0008">2017</xref>) and Yassin et al. (<xref ref-type="bibr" rid="CIT0012">2010</xref>) have highlighted the importance of sequence data to establish species delineation in the Drosophilidae.</p>
<fig id="F0002">
<label>FIGURE 2</label>
<caption><p>Eleven of the 13 Drosophilidae species sampled across South Africa. (a) <italic>Drosophila ananassae</italic>, female; (b) <italic>D. busckii</italic>, female; (c) <italic>D. melanogaster</italic>, female; (d) <italic>D. funebris</italic>, male; (e) <italic>D. hydei</italic>, male; (f) <italic>D. simulans</italic>, female; (g) <italic>Zaprionus indianus</italic>, female; (h) <italic>Z. taronus</italic>, male; (i) <italic>Z. tuberculatus</italic>, male; (j) <italic>Z. vittiger</italic>, male; and (k) <italic>D. immigrans,</italic> female. Note all species are represented at a shared scale of 1 mm. Black boxes indicate the species that are new records for South Africa; there is no image available for <italic>D. nasuta</italic> and <italic>D. yakuba</italic>.</p></caption>
<graphic xmlns:xlink="http://www.w3.org/1999/xlink" xlink:href="ABC-49-2429-g002.tif"/>
</fig>
<p><italic>Zaprionus taronus</italic> was recorded in a home garden in Stellenbosch, Western Cape province, South Africa, while the other two species were sampled near buildings at the South African Sugarcane Research Institute in Mount Edgecombe, KwaZulu-Natal province. We are unable to determine whether these species should be classified as alien or invasive as there is limited data covering Drosophilidae species&#x2019; historical native distributions in the region. The current distribution of <italic>D. ananassae</italic> is cosmopolitan, while <italic>D. nasuta</italic> largely occurs in the Afrotropical region and south Asia (Brake &#x0026; B&#x00E4;chli 2008 cited in Vilela &#x0026; Go&#x00F1;i <xref ref-type="bibr" rid="CIT0010">2015</xref>). The species collected were bycatch from another research study, and thus the bait variety was quite restrictive and sample sites selective; therefore, this may be the reason for the exclusion of other more commonly reported South African drosophilids (e.g. <italic>Z. capensis</italic>). A single sampling site has been provided per species as the primary project only required a single site per species.</p>
</sec>
<sec id="s0004">
<title>Conclusion</title>
<p>Although there are 70 Drosophilidae species known from South Africa (Tsacas <xref ref-type="bibr" rid="CIT0009">1990</xref>), little is known about the current composition of species nor how this varies across space and through time. From our limited effort of a small number of sites with few traps, three of the 13 species we identified were found to be first records for South Africa. This highlights the importance of surveying Drosophilidae in various biomes, and across seasons or climatic conditions, and using diverse baiting methods to capture the full range of species diversity in the family. With improved knowledge of species composition, species interactions and natural diets (land/habitat use) and other ecological functions, the potential impacts of invasive species in this group can be better understood.</p>
</sec>
</body>
<back>
<ack>
<title>Acknowledgements</title>
<p>The authors are grateful to Matroosberg Private Nature Reserve, South African Sugarcane Research Institute (SASRI), Stellenbosch University Botanical Gardens, Tania Vergnani and Jan Coetzee who allowed sampling on their properties. Their special thanks go to Dr Shane McEvey for assistance with species identifications. L.I.D.A. was supported by a Centre for Invasion Biology (C&#x00B7;I&#x00B7;B) postgraduate bursary and M.K. by HORTGRO and SATI.</p>
<sec id="s20005" sec-type="COI-statement">
<title>Competing interests</title>
<p>The authors declare that they have no financial or personal relationships that may have inappropriately influenced them in writing this article.</p>
</sec>
<sec id="s20006">
<title>Authors&#x2019; contributions</title>
<p>M.K. and J.S.T. conceived the project; L.I.D.A., M.K. and J.S.T. secured funding; L.D.A. performed experimental work and gathered data; L.I.D.A., M.K. and J.S.T. analysed the data and co-wrote the article.</p>
</sec>
<sec id="s20007">
<title>Funding</title>
<p>This research was funded by the Centre for Invasion Biology (C&#x00B7;I&#x00B7;B), Stellenbosch University, South Africa.</p>
</sec>
<sec id="s20008">
<title>Disclaimer</title>
<p>The views and opinions expressed in this article are those of the authors and do not necessarily reflect the official policy or position of any affiliated agency of the authors.</p>
</sec>
</ack>
<ref-list id="references">
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<fn><p><bold>How to cite this article:</bold> De Araujo, L.I., Karsten, M. &#x0026; Terblanche, J.S., 2019, &#x2018;Three new Drosophilidae species records for South Africa&#x2019;, <italic>Bothalia</italic> 49(1), a2429. <ext-link ext-link-type="uri" xlink:href="https://doi.org/10.4102/abc.v49i1.2429">https://doi.org/10.4102/abc.v49i1.2429</ext-link></p></fn>
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