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|
||
<div id="article-top-actions-bar" class="article-top-actions-bar">
|
||
<div class="inner-wrap">
|
||
<div class="actions-container">
|
||
<div class="return-to-search">
|
||
|
||
|
||
</div>
|
||
|
||
<div class="multiple-results-actions " role="region" aria-label="save, email, send to">
|
||
<button id="save-results-panel-trigger"
|
||
type="button"
|
||
class="save-results save-results-panel-trigger"
|
||
aria-expanded="false"
|
||
aria-controls="save-action-panel"
|
||
data-ga-category="save_share"
|
||
data-ga-action="save"
|
||
data-ga-label="open">
|
||
Save
|
||
</button><button id="email-results-panel-trigger"
|
||
type="button"
|
||
class="email-results email-results-panel-trigger"
|
||
aria-expanded="false"
|
||
aria-controls="email-action-panel"
|
||
data-ga-category="save_share"
|
||
data-ga-action="email"
|
||
data-ga-label="open">
|
||
Email
|
||
</button><div class="more-actions dropdown-block"><button id="more-actions-trigger"
|
||
type="button"
|
||
class="trigger more-actions-trigger"
|
||
ref="linksrc=show_moreactions_btn"
|
||
aria-label="Send to"
|
||
aria-controls="more-actions-dropdown">Send to</button><div id="more-actions-dropdown" class="dropdown dropdown-container" hidden><div class="content"><ul class="more-actions-links"><li><a class="clipboard-panel-trigger dropdown-block-link "
|
||
href="#"
|
||
role="button"
|
||
data-ga-category="save_share"
|
||
data-ga-action="clipboard"
|
||
data-ga-label="send">
|
||
Clipboard
|
||
</a></li><li><a role="button" class="dropdown-block-link bibliography-trigger-target" href="https://account.ncbi.nlm.nih.gov/?back_url=https%3A%2F%2Fpubmed.ncbi.nlm.nih.gov%2F33764297%2F%23open-bibliography-panel">My Bibliography</a></li><li><a role="button" class="dropdown-block-link collections-trigger-target" href="https://account.ncbi.nlm.nih.gov/?back_url=https%3A%2F%2Fpubmed.ncbi.nlm.nih.gov%2F33764297%2F%23open-collections-panel">Collections</a></li><li><a role="button" class="citation-manager-panel-trigger dropdown-block-link citation-manager-trigger-target" href="#">Citation manager</a></li></ul></div></div></div>
|
||
</div>
|
||
|
||
|
||
|
||
<div class="display-options" data-format-key="format"
|
||
data-default-format-value="abstract"
|
||
data-current-format-value="abstract" role="region">
|
||
<button
|
||
class="trigger usa-button"
|
||
title="Change format"
|
||
data-ga-category="display_options"
|
||
data-ga-action="display_options_dialog">
|
||
<span class="button-label">Display options</span>
|
||
</button>
|
||
|
||
<div class="dropdown" hidden>
|
||
<div class="title">
|
||
Display options
|
||
</div>
|
||
|
||
<div class="content">
|
||
|
||
<div class="format-container">
|
||
<span class="option-label">
|
||
Format
|
||
</span>
|
||
<select name="format" data-initial-value="abstract" aria-label="Display format" id="article-display-format">
|
||
<option value="abstract">Abstract</option>
|
||
|
||
<option value="pubmed">PubMed</option>
|
||
|
||
<option value="pmid">PMID</option>
|
||
|
||
</select>
|
||
</div>
|
||
|
||
</div>
|
||
</div>
|
||
</div>
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
</div>
|
||
</div>
|
||
</div>
|
||
|
||
<div id="article-panels-container" class="article-panels-container">
|
||
<div id="save-action-panel" class="save-action-panel action-panel" aria-hidden="true">
|
||
<div class="inner-wrap">
|
||
<h2 class="action-panel-heading">
|
||
Save citation to file
|
||
</h2>
|
||
<form id="save-action-panel-form"
|
||
class="action-panel-content action-form"
|
||
action="/results-export-ids/"
|
||
data-by-search-action="/results-export-search-data/"
|
||
data-by-ids-action="/results-export-ids/"
|
||
method="post"
|
||
data-by-search-method="post"
|
||
data-by-ids-method="post">
|
||
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
|
||
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<label for="save-action-format" class="action-panel-label">
|
||
Format:
|
||
</label>
|
||
<select id="save-action-format" name="results-format" class="action-panel-selector">
|
||
<option value="summary-text">Summary (text)</option>
|
||
<option value="pubmed-text">PubMed</option>
|
||
<option value="pmid">PMID</option>
|
||
<option value="abstract">Abstract (text)</option>
|
||
<option value="csv">CSV</option>
|
||
</select>
|
||
</div>
|
||
|
||
<div class="action-panel-actions">
|
||
<button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Saving..."
|
||
data-ga-category="save_share"
|
||
data-ga-action="save"
|
||
data-ga-label="save">
|
||
Create file
|
||
</button>
|
||
<button class="action-panel-cancel"
|
||
aria-label="Close 'Save citations to file' panel"
|
||
ref="linksrc=close_save_panel"
|
||
aria-controls="save-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="save_share"
|
||
data-ga-action="save"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button>
|
||
</div>
|
||
</form>
|
||
</div>
|
||
</div>
|
||
|
||
|
||
|
||
|
||
<div id="email-action-panel"
|
||
class="email-action-panel action-panel"
|
||
aria-hidden="true"
|
||
data-recaptcha-script-url="https://www.recaptcha.net/recaptcha/enterprise.js?render=explicit&onload=onloadCaptchaCallback"
|
||
data-recaptcha-enabled="true">
|
||
<div class="inner-wrap">
|
||
<h2 class="action-panel-heading">
|
||
Email citation
|
||
</h2>
|
||
<form id="email-action-panel-form"
|
||
class="action-panel-content action-form"
|
||
action="/send-email/"
|
||
data-by-search-action="/results-export-email-by-search-data/"
|
||
data-by-ids-action="/send-email/"
|
||
data-by-search-method="post"
|
||
data-by-ids-method="post"
|
||
method="post"
|
||
>
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<span class="action-panel-label">
|
||
Subject:
|
||
</span>
|
||
<span class="email-subject" id="email-subject" aria-label="Email subject">1 selected item: 33764297 - PubMed</span>
|
||
<input type="hidden" name="email-subject" value="1 selected item: 33764297"/>
|
||
<input type="hidden" name="email-subject-hash" value="23f8431cf60225be6032ce911e6a48655e2f07b62131cb2a91e2cba52a540df4"/>
|
||
</div>
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<label for="email-to" class="action-panel-label required-field-asterisk">
|
||
To:
|
||
</label>
|
||
<input type="email" aria-label="Recipient Email Address" placeholder="email@example.com" id="email-to" class="email-to" pattern="^[a-zA-Z0-9_.+-]+@[a-zA-Z0-9-]+\.[a-zA-Z0-9-.]+$" maxlength="100" required>
|
||
</div>
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<label for="email-from" class="action-panel-label">
|
||
From:
|
||
</label>
|
||
<input type="email" aria-label="Sender Email Address" placeholder="email@example.com" id="email-from" class="email-from" pattern="^[a-zA-Z0-9_.+-]+@[a-zA-Z0-9-]+\.[a-zA-Z0-9-.]+$" maxlength="256">
|
||
</div>
|
||
|
||
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<label for="email-citation-format" class="action-panel-label">
|
||
Format:
|
||
</label>
|
||
<select id="email-citation-format" name="citation-format" class="action-panel-selector email-citation-format">
|
||
<option selected="selected" value="summary">Summary</option>
|
||
<option value="summary-text">Summary (text)</option>
|
||
<option value="abstract">Abstract</option>
|
||
<option value="abstract-text">Abstract (text)</option>
|
||
|
||
</select>
|
||
</div>
|
||
<div class="include-supplemental-container">
|
||
<input type="checkbox" aria-label="Include MeSH and other data" name="include-supplemental" id="email-include-supplemental" class="email-include-supplemental">
|
||
<label for="email-include-supplemental" class="email-include-supplemental-label">MeSH and other data</label>
|
||
</div>
|
||
|
||
|
||
|
||
<div class="form-field recaptcha ">
|
||
|
||
|
||
|
||
<div class="g-recaptcha" id="id-recaptcha" data-sitekey="6LfsWHMdAAAAAClKbtOpjQ2pMjgsGxvv7NdZW9uI"></div>
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
<div id="captcha-error-message" class="usa-input-error-message captcha-validation-message" role="alert"></div>
|
||
|
||
|
||
|
||
<div class="action-panel-actions">
|
||
<button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Sending..."
|
||
data-ga-category="save_share"
|
||
data-ga-action="email"
|
||
data-ga-label="send">
|
||
Send email
|
||
</button>
|
||
<button class="action-panel-cancel"
|
||
aria-label="Close 'Email citations' panel"
|
||
ref="linksrc=close_email_panel"
|
||
aria-controls="email-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="save_share"
|
||
data-ga-action="email"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button>
|
||
</div>
|
||
<input type="hidden" name="email-search-details" value="" />
|
||
<input type="hidden" name="email-search-details-hash" value="0e42663a6c3bd85498fcb88798998fed7bfdc45d457db35281e41afe13cc0524" />
|
||
</form>
|
||
</div>
|
||
</div>
|
||
|
||
<div id="collections-action-panel"
|
||
class="collections-action-panel action-panel in-progress-dots-panel"
|
||
aria-hidden="true"
|
||
data-collections-open-panel-enabled="false"
|
||
data-collections-open-panel-url-hash="#open-collections-panel">
|
||
<div class="inner-wrap">
|
||
<h3 class="action-panel-heading">
|
||
Add to Collections
|
||
</h3>
|
||
|
||
|
||
<form id="collections-action-panel-form"
|
||
class="collections-action-panel-form action-panel-content action-form action-panel-smaller-selectors"
|
||
data-existing-collections-url="/list-existing-collections/"
|
||
data-add-to-existing-collection-url="/add-to-existing-collection/"
|
||
data-create-and-add-to-new-collection-url="/create-and-add-to-new-collection/"
|
||
data-get-article-ids-by-search-url="/get-article-ids-by-search/"
|
||
data-myncbi-max-collection-name-length="100"
|
||
data-add-to-collection-max-amount="1000"
|
||
data-collections-root-url="https://www.ncbi.nlm.nih.gov/myncbi/collections/">
|
||
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
|
||
|
||
|
||
<div class="choice-group" role="radiogroup">
|
||
<ul class="radio-group-items">
|
||
<li>
|
||
<input type="radio"
|
||
id="collections-action-panel-new"
|
||
class="collections-new"
|
||
name="collections"
|
||
value="new"
|
||
data-ga-category="save_share"
|
||
data-ga-action="collections"
|
||
data-ga-label="collections_radio_new">
|
||
<label for="collections-action-panel-new">Create a new collection</label>
|
||
</li>
|
||
<li>
|
||
<input type="radio"
|
||
id="collections-action-panel-existing"
|
||
class="collections-existing"
|
||
name="collections"
|
||
value="existing"
|
||
checked="true"
|
||
data-ga-category="save_share"
|
||
data-ga-action="collections"
|
||
data-ga-label="collections_radio_existing">
|
||
<label for="collections-action-panel-existing">Add to an existing collection</label>
|
||
</li>
|
||
</ul>
|
||
</div>
|
||
|
||
<div class="controls-wrapper">
|
||
<div class="action-panel-control-wrap new-collections-controls">
|
||
<label for="collections-action-panel-add-to-new" class="action-panel-label required-field-asterisk">
|
||
Name your collection:
|
||
</label>
|
||
<input
|
||
type="text"
|
||
name="add-to-new-collection"
|
||
id="collections-action-panel-add-to-new"
|
||
class="collections-action-add-to-new"
|
||
pattern="[^"&=<>\/]*" title="The following characters are not allowed in the Name field: "&=<>/"
|
||
maxlength="100"
|
||
data-ga-category="save_share"
|
||
data-ga-action="create_collection"
|
||
data-ga-label="non_favorties_collection">
|
||
<div class="collections-new-name-too-long usa-input-error-message selection-validation-message">
|
||
Name must be less than 100 characters
|
||
</div>
|
||
</div>
|
||
<div class="action-panel-control-wrap existing-collections-controls">
|
||
<label for="collections-action-panel-add-to-existing" class="action-panel-label">
|
||
Choose a collection:
|
||
</label>
|
||
<select id="collections-action-panel-add-to-existing"
|
||
class="action-panel-selector collections-action-add-to-existing"
|
||
data-ga-category="save_share"
|
||
data-ga-action="select_collection"
|
||
data-ga-label="($('#collections-action-add-to-existing').val() === 'Favorites') ? 'Favorites' : 'non_favorites_collection'">
|
||
</select>
|
||
<div class="collections-retry-load-on-error usa-input-error-message selection-validation-message">
|
||
Unable to load your collection due to an error<br>
|
||
<a href="#">Please try again</a>
|
||
</div>
|
||
</div>
|
||
</div>
|
||
|
||
<div class="action-panel-actions">
|
||
<button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Adding..."
|
||
data-pinger-ignore
|
||
data-ga-category="save_share"
|
||
data-ga-action="collections"
|
||
data-ga-label="add">
|
||
Add
|
||
</button>
|
||
<button class="action-panel-cancel"
|
||
aria-label="Close 'Add to Collections' panel"
|
||
ref="linksrc=close_collections_panel"
|
||
aria-controls="collections-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="save_share"
|
||
data-ga-action="collections"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button>
|
||
</div>
|
||
</form>
|
||
</div>
|
||
</div>
|
||
|
||
<div id="bibliography-action-panel"
|
||
class="bibliography-action-panel action-panel in-progress-dots-panel"
|
||
aria-hidden="true"
|
||
data-bibliography-open-panel-enabled="false"
|
||
data-bibliography-open-panel-url-hash="#open-bibliography-panel">
|
||
<div class="inner-wrap">
|
||
<h3 class="action-panel-heading">
|
||
Add to My Bibliography
|
||
</h3>
|
||
<form id="bibliography-action-panel-form"
|
||
class="bibliography-action-panel-form action-panel-content action-form action-panel-smaller-selectors"
|
||
data-add-to-bibliography-max-amount="100"
|
||
data-add-to-bibliography-batch-size="10"
|
||
data-bibliography-delegates-url="/list-bibliography-delegates/"
|
||
data-add-to-bibliography-url="/add-to-bibliography/"
|
||
data-get-article-ids-by-search-url="/get-article-ids-by-search/"
|
||
data-mybib-root-url="https://www.ncbi.nlm.nih.gov/myncbi/collections/mybibliography/">
|
||
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
|
||
|
||
|
||
<div class="action-panel-control-wrap bibliographies-controls">
|
||
<div class="choice-group">
|
||
<ul class="bibliographies-action-add radio-group-items">
|
||
<li>
|
||
<input name="bibliography" id="my-bibliography" class="my-bibliography" type="radio" checked/>
|
||
<label for="my-bibliography">My Bibliography</label>
|
||
</li>
|
||
</ul>
|
||
</div>
|
||
</div>
|
||
|
||
<div class="bibliographies-retry-load-on-error usa-input-error-message selection-validation-message">
|
||
Unable to load your delegates due to an error<br>
|
||
<a href="#">Please try again</a>
|
||
</div>
|
||
|
||
<div class="action-panel-actions">
|
||
<button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Adding..."
|
||
data-pinger-ignore>
|
||
Add
|
||
</button>
|
||
<button class="action-panel-cancel"
|
||
aria-label="Close 'Add to bibliography' panel"
|
||
ref="linksrc=close_bibliography_panel"
|
||
aria-controls="bibliography-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="save_share"
|
||
data-ga-action="mybib"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button>
|
||
</div>
|
||
</form>
|
||
</div>
|
||
</div>
|
||
|
||
|
||
|
||
|
||
<div id="saved-search-action-panel" class="saved-search-action-panel action-panel " aria-hidden="true"
|
||
data-saved-search-open-panel-enabled="false"
|
||
data-saved-search-open-panel-url-hash="#open-saved-search-panel">
|
||
<div class="inner-wrap">
|
||
<h2 class="action-panel-heading">
|
||
Your saved search
|
||
</h2>
|
||
|
||
<form id="saved-search-action-panel-form"
|
||
class="saved-search-action-panel-form action-panel-content action-form"
|
||
data-create-saved-search-url="/create-saved-search/"
|
||
data-try-search-terms-url="/try-search-term/"
|
||
data-saved-search-root-url="https://www.ncbi.nlm.nih.gov/myncbi/searches/">
|
||
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<label for="saved-search-name" class="action-panel-label saved-search-name-label required-field-asterisk">
|
||
Name of saved search:
|
||
</label>
|
||
<input maxlength="200"
|
||
type="text"
|
||
name="saved-search-name"
|
||
id="saved-search-name"
|
||
class="saved-search-name"
|
||
value=""
|
||
required
|
||
pattern="[^"&=<>\/]*" title="The following characters are not allowed in the Name field: "&=<>/">
|
||
</div>
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<label for="saved-search-term" class="action-panel-label required-field-asterisk">
|
||
Search terms:
|
||
</label>
|
||
<textarea name="saved-search-term" id="saved-search-term" class="saved-search-term" required></textarea>
|
||
</div>
|
||
<div class="test-search-term-wrap">
|
||
<a href="#" class="try-search-term">Test search terms</a>
|
||
</div>
|
||
<div class="choice-group action-panel-extra-margin-top">
|
||
<span class="action-panel-label" id="fieldset-label">
|
||
Would you like email updates of new search results?
|
||
</span>
|
||
<fieldset id="saved-search-alert" aria-describedby="fieldset-label">
|
||
<legend class="usa-sr-only">Saved Search Alert Radio Buttons</legend>
|
||
<ul class="radio-group-items">
|
||
<li>
|
||
<input type="radio" id="saved-search-alert-yes" class="saved-search-alert-yes" name="saved-search-alert" value="yes" checked>
|
||
<label for="saved-search-alert-yes" class="action-panel-label">Yes</label>
|
||
</li>
|
||
<li>
|
||
<input aria-label="No radio input" type="radio" id="saved-search-alert-no" class="saved-search-alert-no" name="saved-search-alert" value="no">
|
||
<label for="saved-search-alert-no" class="action-panel-label">No</label>
|
||
</li>
|
||
</ul>
|
||
</fieldset>
|
||
</div>
|
||
|
||
<div class="alert-schedule-wrap">
|
||
<div class="action-panel-control-wrap">
|
||
<label class="action-panel-label">
|
||
Email:
|
||
</label>
|
||
<span aria-label="Email address" id="saved-search-email" class="action-panel-label"><span class="action-panel-label-bold"></span> (<a class="myncbi-account-settings" href="https://www.ncbi.nlm.nih.gov/account/settings/">change</a>)</span>
|
||
</div>
|
||
<div class="action-panel-control-wrap action-panel-extra-margin-top">
|
||
<label for="saved-search-frequency" class="action-panel-label">
|
||
Frequency:
|
||
</label>
|
||
<select id="saved-search-frequency" class="no-border-panel-selector saved-search-frequency">
|
||
<option value="monthly">Monthly</option>
|
||
<option value="weekly">Weekly</option>
|
||
<option value="daily">Daily</option>
|
||
</select>
|
||
</div>
|
||
<div class="action-panel-control-wrap saved-search-monthly-additional">
|
||
<label for="saved-search-monthly-on-day" class="action-panel-label">
|
||
Which day?
|
||
</label>
|
||
<select id="saved-search-monthly-on-day" class="no-border-panel-selector">
|
||
<option value="Sunday">The first Sunday</option>
|
||
<option value="Monday">The first Monday</option>
|
||
<option value="Tuesday">The first Tuesday</option>
|
||
<option value="Wednesday">The first Wednesday</option>
|
||
<option value="Thursday">The first Thursday</option>
|
||
<option value="Friday">The first Friday</option>
|
||
<option value="Saturday">The first Saturday</option>
|
||
<option value="day">The first day</option>
|
||
<option value="weekday">The first weekday</option>
|
||
</select>
|
||
</div>
|
||
<div class="action-panel-control-wrap saved-search-weekly-additional">
|
||
<label for="saved-search-weekly-on-day" class="action-panel-label">
|
||
Which day?
|
||
</label>
|
||
<select id="saved-search-weekly-on-day" class="no-border-panel-selector saved-search-weekly-on-day">
|
||
<option value="Sunday">Sunday</option>
|
||
<option value="Monday">Monday</option>
|
||
<option value="Tuesday">Tuesday</option>
|
||
<option value="Wednesday">Wednesday</option>
|
||
<option value="Thursday">Thursday</option>
|
||
<option value="Friday">Friday</option>
|
||
<option value="Saturday">Saturday</option>
|
||
</select>
|
||
</div>
|
||
<div class="action-panel-control-wrap">
|
||
<label for="saved-search-report" class="action-panel-label">
|
||
Report format:
|
||
</label>
|
||
<select id="saved-search-report" class="no-border-panel-selector saved-search-report">
|
||
<option value="DocSum">Summary</option>
|
||
<option value="DocSumText">Summary (text)</option>
|
||
<option value="Abstract">Abstract</option>
|
||
<option value="AbstractText">Abstract (text)</option>
|
||
<option value="MEDLINE">PubMed</option>
|
||
</select>
|
||
</div>
|
||
<div class="action-panel-control-wrap">
|
||
<label for="saved-search-amount" class="action-panel-label">
|
||
Send at most:
|
||
</label>
|
||
<select id="saved-search-amount" class="no-border-panel-selector saved-search-amount">
|
||
<option value="1">1 item</option>
|
||
<option value="5" selected>5 items</option>
|
||
<option value="10">10 items</option>
|
||
<option value="20">20 items</option>
|
||
<option value="50">50 items</option>
|
||
<option value="100">100 items</option>
|
||
<option value="200">200 items</option>
|
||
</select>
|
||
</div>
|
||
<div>
|
||
<input type="checkbox" id="saved-search-send-if-no-result" class="saved-search-send-if-no-result" name="saved-search-send-if-no-result">
|
||
<label for="saved-search-send-if-no-result" class="action-panel-label smaller-checkbox">
|
||
Send even when there aren't any new results
|
||
</label>
|
||
</div>
|
||
<div class="action-panel-control-wrap option-text-in-email-wrap">
|
||
<label for="saved-search-email-text" class="action-panel-label">
|
||
Optional text in email:
|
||
</label>
|
||
<textarea name="saved-search-email-text"
|
||
id="saved-search-email-text"
|
||
class="saved-search-email-text"></textarea>
|
||
</div>
|
||
|
||
</div>
|
||
|
||
<div class="action-panel-actions">
|
||
<button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Saving..."
|
||
data-ga-category="save_share"
|
||
data-ga-action="alert"
|
||
data-ga-label="save">
|
||
Save
|
||
</button>
|
||
<button class="action-panel-cancel"
|
||
aria-label="Close 'Your saved search' panel"
|
||
ref="linksrc=close_saved_search_panel"
|
||
aria-controls="saved-search-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="save_share"
|
||
data-ga-action="alert"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button>
|
||
</div>
|
||
</form>
|
||
</div>
|
||
</div>
|
||
|
||
<div id="citation-manager-action-panel" class="citation-manager-action-panel action-panel" aria-hidden="true">
|
||
<div class="inner-wrap">
|
||
<h2 class="action-panel-heading">
|
||
Create a file for external citation management software
|
||
</h2>
|
||
<form id="citation-manager-action-panel-form"
|
||
class="action-panel-content action-form"
|
||
action="/results-export-ids/"
|
||
data-by-search-action="/results-export-search-data/"
|
||
data-by-ids-action="/results-export-ids/"
|
||
method="post"
|
||
data-by-search-method="post"
|
||
data-by-ids-method="post">
|
||
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
|
||
|
||
|
||
<input name="results-format" type="hidden" value="pubmed"/>
|
||
|
||
<div class="action-panel-actions">
|
||
<button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Sending..."
|
||
data-ga-category="save_share"
|
||
data-ga-action="citation_manager"
|
||
data-ga-label="save">
|
||
Create file
|
||
</button>
|
||
<button class="action-panel-cancel"
|
||
aria-label="Close 'Send citations to citation manager' panel"
|
||
ref="linksrc=close_citation_manager_panel"
|
||
aria-controls="citation-manager-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="save_share"
|
||
data-ga-action="citation_manager"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button>
|
||
</div>
|
||
</form>
|
||
</div>
|
||
</div>
|
||
|
||
|
||
|
||
|
||
<div id="rss-action-panel" class="rss-action-panel action-panel " aria-hidden="true">
|
||
<div class="inner-wrap">
|
||
<h2 class="action-panel-heading">
|
||
Your RSS Feed
|
||
</h2>
|
||
|
||
<form id="rss-action-panel-form"
|
||
class="rss-action-panel-form action-panel-content action-form"
|
||
data-create-rss-feed-url="/create-rss-feed-url/"
|
||
data-search-form-term-value="">
|
||
|
||
<input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z">
|
||
|
||
<div class="action-panel-control-wrap">
|
||
<label for="rss-name" class="action-panel-label required-field-asterisk">
|
||
Name of RSS Feed:
|
||
</label>
|
||
<input maxlength="200"
|
||
placeholder="Name"
|
||
type="text"
|
||
name="rss-name"
|
||
id="rss-name"
|
||
class="rss-name"
|
||
value=''
|
||
required
|
||
pattern="[^"&=<>\/]*" title="The following characters are not allowed in the Name field: "&=<>/">
|
||
</div>
|
||
|
||
<div class="rss-limit-wrap">
|
||
<div class="action-panel-control-wrap action-panel-extra-margin-top">
|
||
<label for="rss-limit" class="action-panel-label">
|
||
Number of items displayed:
|
||
</label>
|
||
<select id="rss-limit" class="no-border-panel-selector rss-limit">
|
||
<option value="5">5</option>
|
||
<option value="10">10</option>
|
||
<option value="15" selected="selected">15</option>
|
||
<option value="20">20</option>
|
||
<option value="50">50</option>
|
||
<option value="100">100</option>
|
||
</select>
|
||
</div>
|
||
</div>
|
||
|
||
<div class="action-panel-actions">
|
||
<button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Creating..."
|
||
data-ga-category="save_share"
|
||
data-ga-action="alert"
|
||
data-ga-label="save">
|
||
Create RSS
|
||
</button>
|
||
<button class="action-panel-cancel"
|
||
aria-label="Close 'Your RSS' panel"
|
||
ref="linksrc=close_rss_panel"
|
||
aria-controls="rss-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="save_share"
|
||
data-ga-action="alert"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button>
|
||
</div>
|
||
|
||
<div class="action-panel-control-wrap rss-link-copy-wrap">
|
||
<label for="rss-link" class="usa-sr-only">RSS Link</label>
|
||
<input placeholder="Your RSS Feed Link" type="text" name="rss-link" id="rss-link" class="rss-link" title="RSS Link">
|
||
<button
|
||
type="button"
|
||
disabled
|
||
class="rss-link-copy-button disabled"
|
||
data-ga-category="save_share"
|
||
data-ga-action="rss"
|
||
data-ga-label="copy">
|
||
Copy
|
||
</button>
|
||
</div>
|
||
</form>
|
||
</div>
|
||
</div>
|
||
|
||
</div>
|
||
</div>
|
||
|
||
<div class="article-page" id="article-page" data-article-pmid="33764297">
|
||
<aside class="page-sidebar">
|
||
<div class="inner-wrap">
|
||
|
||
<div class="full-text-links">
|
||
<div class="full-view">
|
||
<h3 class="title">
|
||
Full text links
|
||
</h3>
|
||
<div class="full-text-links-list">
|
||
<a class="link-item
|
||
|
||
dialog-focus"
|
||
href="https://doi.org/10.7554/eLife.64774"
|
||
target="_blank"
|
||
rel="noopener"
|
||
|
||
ref="linksrc=fulltextorjournal_fulltext&is_pmc=False&PrId=8981&itool=Abstract-def&log$=linkouticon&uid=33764297&db=pubmed&nlmid=101579614"
|
||
title="See full text options at eLife Sciences Publications, Ltd"
|
||
|
||
data-ga-category="full_text"
|
||
data-ga-action="eLife Sciences Publications, Ltd"
|
||
data-ga-label="33764297"
|
||
><img src="https://cdn.ncbi.nlm.nih.gov/corehtml/query/egifs/https:--cdn.elifesciences.org-images-elife_oa@1x.png" alt="eLife Sciences Publications, Ltd full text link"><span class="text">
|
||
|
||
eLife Sciences Publications, Ltd
|
||
|
||
</span></a><a class="link-item
|
||
pmc
|
||
"
|
||
href="https://pmc.ncbi.nlm.nih.gov/articles/pmid/33764297/"
|
||
target="_blank"
|
||
rel="noopener"
|
||
|
||
ref="linksrc=fulltextorjournal_fulltext&is_pmc=True&PrId=3494&itool=Abstract-def&log$=linkouticon&uid=33764297&db=pubmed&nlmid=101579614"
|
||
title="Free full text at PubMed Central"
|
||
|
||
data-ga-category="full_text"
|
||
data-ga-action="PMC"
|
||
data-ga-label="33764297"
|
||
><span class="text">
|
||
|
||
Free PMC article
|
||
|
||
</span></a>
|
||
</div>
|
||
|
||
</div>
|
||
|
||
<div class="short-view">
|
||
<a href="#" class="full-text-links-button full-text-links-dialog-trigger">
|
||
Full text links
|
||
</a>
|
||
</div>
|
||
</div>
|
||
|
||
|
||
|
||
|
||
<div class="actions-buttons sidebar"><h3 class="title">Actions</h3><div class="inner-wrap"><button class="citation-button citation-dialog-trigger"
|
||
aria-label="Open dialog with citation text in different styles"
|
||
data-ga-category="save_share"
|
||
data-ga-action="cite"
|
||
data-ga-label="open"
|
||
data-all-citations-url="/33764297/citations/"
|
||
data-citation-style="nlm"
|
||
data-pubmed-format-link="/33764297/export/"><span class="button-label">Cite</span></button><link type="text/css" href="ncbi-overlay-block/src/overlay-block.css"><div class="collections-button-container" data-article-id="33764297" data-article-db="pubmed"><button class="collections-button collections-dialog-trigger"
|
||
aria-label="Save article in MyNCBI collections."
|
||
data-ga-category="collections_button"
|
||
data-ga-action="click"
|
||
data-ga-label="collections_button"
|
||
data-collections-open-dialog-enabled="false"
|
||
data-collections-open-dialog-url="https://account.ncbi.nlm.nih.gov/?back_url=https%3A%2F%2Fpubmed.ncbi.nlm.nih.gov%2F33764297%2F%23open-collections-dialog"
|
||
data-in-collections="false"><span class="button-label">Collections</span></button><div class="overlay" role="dialog"><div id="collections-action-dialog"
|
||
class="dialog collections-dialog"
|
||
aria-hidden="true"><div class="title">Add to Collections</div><div class="collections-action-panel action-panel"><form id="collections-action-dialog-form"
|
||
class="collections-action-panel-form action-panel-content action-form action-panel-smaller-selectors"
|
||
data-existing-collections-url="/list-existing-collections/"
|
||
data-add-to-existing-collection-url="/add-to-existing-collection/"
|
||
data-create-and-add-to-new-collection-url="/create-and-add-to-new-collection/"
|
||
data-myncbi-max-collection-name-length="100"
|
||
data-collections-root-url="https://www.ncbi.nlm.nih.gov/myncbi/collections/"><input type="hidden" name="csrfmiddlewaretoken" value="ch3GntjWYFWcfqvIFRIakzaSXbnbk9yft4NL0jMiygJG9RByialcOHGwvf5ndK6z"><div class="choice-group" role="radiogroup"><ul class="radio-group-items"><li><input type="radio"
|
||
id="collections-action-dialog-new"
|
||
class="collections-new"
|
||
name="collections"
|
||
value="new"
|
||
data-ga-category="collections_button"
|
||
data-ga-action="click"
|
||
data-ga-label="collections_radio_new"><label for="collections-action-dialog-new">Create a new collection</label></li><li><input type="radio"
|
||
id="collections-action-dialog-existing"
|
||
class="collections-existing"
|
||
name="collections"
|
||
value="existing"
|
||
checked="true"
|
||
data-ga-category="collections_button"
|
||
data-ga-action="click"
|
||
data-ga-label="collections_radio_existing"><label for="collections-action-dialog-existing">Add to an existing collection</label></li></ul></div><div class="controls-wrapper"><div class="action-panel-control-wrap new-collections-controls"><label for="collections-action-dialog-add-to-new" class="action-panel-label required-field-asterisk">
|
||
Name your collection:
|
||
</label><input
|
||
type="text"
|
||
name="add-to-new-collection"
|
||
id="collections-action-dialog-add-to-new"
|
||
class="collections-action-add-to-new"
|
||
pattern="[^"&=<>\/]*" title="The following characters are not allowed in the Name field: "&=<>/"
|
||
maxlength="100"
|
||
data-ga-category="collections_button"
|
||
data-ga-action="create_collection"
|
||
data-ga-label="non_favorties_collection"><div class="collections-new-name-too-long usa-input-error-message selection-validation-message">
|
||
Name must be less than 100 characters
|
||
</div></div><div class="action-panel-control-wrap existing-collections-controls"><label for="collections-action-dialog-add-to-existing" class="action-panel-label">
|
||
Choose a collection:
|
||
</label><select id="collections-action-dialog-add-to-existing"
|
||
class="action-panel-selector collections-action-add-to-existing"
|
||
data-ga-category="collections_button"
|
||
data-ga-action="select_collection"
|
||
data-ga-label="($('.collections-action-add-to-existing').val() === 'Favorites') ? 'Favorites' : 'non_favorites_collection'"></select><div class="collections-retry-load-on-error usa-input-error-message selection-validation-message">
|
||
Unable to load your collection due to an error<br><a href="#">Please try again</a></div></div></div><div class="action-panel-actions"><button class="action-panel-submit"
|
||
type="submit"
|
||
data-loading-label="Adding..."
|
||
data-pinger-ignore
|
||
data-ga-category="collections_button"
|
||
data-ga-action="click"
|
||
data-ga-label="add">
|
||
Add
|
||
</button><button class="action-panel-cancel"
|
||
aria-label="Close 'Add to Collections' panel"
|
||
ref="linksrc=close_collections_panel"
|
||
aria-controls="collections-action-panel"
|
||
aria-expanded="false"
|
||
data-ga-category="collections_button"
|
||
data-ga-action="click"
|
||
data-ga-label="cancel">
|
||
Cancel
|
||
</button></div></form></div></div></div></div><button class="more-actions-button more-actions-dialog-trigger"
|
||
title="Open dialog with more actions to take"
|
||
ref="linksrc=more_actions_btn"></button><div class="display-options" data-format-key="format"
|
||
data-default-format-value="abstract"
|
||
data-current-format-value="abstract" role="region"><button
|
||
class="trigger usa-button"
|
||
title="Change format"
|
||
data-ga-category="display_options"
|
||
data-ga-action="display_options_dialog"><span class="button-label">Display options</span></button><div class="dropdown" hidden><div class="title">
|
||
Display options
|
||
</div><div class="content"><div class="format-container"><span class="option-label">
|
||
Format
|
||
</span><select name="format" data-initial-value="abstract" aria-label="Display format" id="sidebar-actions-display-format"><option value="abstract">Abstract</option><option value="pubmed">PubMed</option><option value="pmid">PMID</option></select></div></div></div></div></div></div>
|
||
|
||
<div class="social-sharing">
|
||
<h3 class="title">
|
||
Share
|
||
</h3>
|
||
<ul class="social-sharing-links usa-unstyled-list">
|
||
|
||
|
||
<li>
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</a></li></ul></div></div></div><span class="period">. </span><span class="cit">2021 Mar 25:10:e64774.</span>
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Highly parallelized droplet cultivation and prioritization of antibiotic producers from natural microbial communities
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Highly parallelized droplet cultivation and prioritization of antibiotic producers from natural microbial communities
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</span><select name="format" data-initial-value="abstract" aria-label="Display format" id="inline-actions-display-format"><option value="abstract">Abstract</option><option value="pubmed">PubMed</option><option value="pmid">PMID</option></select></div></div></div></div></div></div>
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<div class="abstract" id="abstract">
|
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<h2 class="title">
|
||
Abstract
|
||
|
||
</h2>
|
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<div class="abstract-content selected"
|
||
id="eng-abstract">
|
||
|
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|
||
<p>
|
||
|
||
Antibiotics from few culturable microorganisms have saved millions of lives since the 20th century. But with resistance formation, these compounds become increasingly ineffective, while the majority of microbial and with that chemical compound diversity remains inaccessible for cultivation and exploration. Culturing recalcitrant bacteria is a stochastic process. But conventional methods are limited to low throughput. By increasing (i) throughput and (ii) sensitivity by miniaturization, we innovate microbiological cultivation to comply with biological stochasticity. Here, we introduce a droplet-based microscale cultivation system, which is directly coupled to a high-throughput screening for antimicrobial activity prior to strain isolation. We demonstrate that highly parallelized in-droplet cultivation starting from single cells results in the cultivation of yet uncultured species and a significantly higher bacterial diversity than standard agar plate cultivation. Strains able to inhibit intact reporter strains were isolated from the system. A variety of antimicrobial compounds were detected for a selected potent antibiotic producer.
|
||
</p>
|
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</div>
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|
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<p>
|
||
|
||
<strong class="sub-title">
|
||
Keywords:
|
||
</strong>
|
||
|
||
antibiotics; antimicrobial compound; high-throughput cultivation; infectious disease; microbiology; strain isolation; uncultured microorganisms.
|
||
</p>
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
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|
||
<div class="abstract" id="plain-language-summary">
|
||
<h2 class="title">
|
||
Plain language summary
|
||
</h2>
|
||
|
||
|
||
|
||
|
||
|
||
|
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<p>
|
||
|
||
Antibiotics are chemicals derived from microorganisms that can kill the bacteria that harm human health. In the 20<sup>th</sup> and 21<sup>st</sup> centuries antibiotics saved millions of lives, but new strains of dangerous bacteria that cannot be killed by antibiotics, known as antibiotic resistant strains, are becoming more frequent. Most antibiotics are produced by only a small group of microorganisms, but many more microorganisms exist in nature. So it is possible that microorganisms outside this small group can produce different antibiotics that are effective against antibiotic resistant strains. Unfortunately, finding the microorganisms that produce these alternative antibiotics is challenging because researchers do not know which bacteria are producing these molecules and how to grow these microorganisms in the laboratory. To solve this problem, Mahler et al. developed a new method for growing a new subset of microorganisms in the laboratory. This would allow researchers to study the new microorganisms under controlled conditions, and determine whether any of the substances they produce have antibiotic properties. Mahler et al. generated tiny droplets that could only contain a single cell of a microorganism, so each microbe could grow alone in its own protected environment. Using this approach, it was possible to grow completely different types of microorganisms than with traditional techniques, and keep them isolated from each other. This allowed each different species of microbe to be screened for antimicrobial activity, allowing the identification of chemicals that could potentially be developed into new antibiotics. This new method is automated and miniaturized, paving the way for growing many more cells in few hours, with very low material and space requirements. These results showcase a way of growing new types of microorganisms in the laboratory, making it easier and faster to study them and determine what chemicals they produce. Understanding a greater variety of microorganisms in detail can help identify new chemicals for industrial applications, including new ways of combating infections.
|
||
</p>
|
||
|
||
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||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
|
||
|
||
|
||
<p class="copyright" id="copyright">
|
||
© 2021, Mahler et al.
|
||
</p>
|
||
|
||
|
||
<p class="disclaimer" id="disclaimer">
|
||
<a href="/disclaimer/" target="_blank" data-ga-category="literature_resources"
|
||
data-ga-action="disclaimer_link" data-ga-label="under_abstract">PubMed Disclaimer</a>
|
||
</p>
|
||
|
||
<div class="conflict-of-interest" id="conflict-of-interest">
|
||
<h2 class="title">
|
||
Conflict of interest statement
|
||
</h2>
|
||
|
||
<div class="statement">
|
||
|
||
|
||
|
||
<p xmlns:xlink="http://www.w3.org/1999/xlink" xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:p1="http://pubmed.gov/pub-one">LM, SN, KM, TW, KS, CH, MR, MR No competing interests declared</p>
|
||
|
||
|
||
</div>
|
||
</div>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<div class="figures" id="figures">
|
||
<h2 class="title">
|
||
Figures
|
||
</h2>
|
||
|
||
<div class="figures-list" id="slides-container"
|
||
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||
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|
||
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||
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||
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|
||
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|
||
data-image-height="412"
|
||
data-image-alt="Figure 1."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig1">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-0"
|
||
src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/e638c18dacb8/elife-64774-fig1.gif"
|
||
alt="Figure 1." />
|
||
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||
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|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 1.. Concept of microbial droplet cultivation… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
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|
||
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|
||
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||
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|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 1.. Concept of microbial droplet cultivation and activity screening. </p>
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents"><p> ( <b> A </b> ) General workflow… </p></div>
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-0" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 1.. Concept of microbial droplet cultivation and activity screening.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">(<b>A</b>) General workflow of in-droplet cultivation with two options for strain isolation. (1) Single cells of the soil community are encapsulated in droplets. (2) Droplets are incubated under oxic conditions in bulk. (3a) Droplets are deposited on agar plates for strain isolation without previous selection. (3b) Droplets are screened for antibiotic producers and only screening hits are deposited for strain isolation. (<b>B</b>) Brightfield microscopy images of droplet samples after droplet generation and 28 days of incubation. Bacterial colonies were not visible right after droplet generation (top). Negative control droplets remained empty throughout the entire incubation period (bottom left). A mixture of five <i>Streptomyces</i> strains (Table 1) was used as a positive control (middle). Co-encapsulated soil particles are visible as dark particles in the images of droplets with soil community (right). A variety of different morphologies were observed in droplets with soil community, ranging from different sizes and densities of cocci- and rod-shaped cells to filamentous growth (bottom right). Droplets with cell colonies are marked with arrows.</div>
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
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|
||
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|
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|
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||
data-image-width="762"
|
||
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|
||
data-image-alt="Figure 1—figure supplement 1."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig1s1">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-1"
|
||
src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/6db7b6969668/elife-64774-fig1-figsupp1.gif"
|
||
alt="Figure 1—figure supplement 1." />
|
||
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|
||
|
||
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||
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|
||
|
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|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 1—figure supplement 1.. Workflow for the… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
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|
||
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|
||
|
||
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|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 1—figure supplement 1.. Workflow for the comparison of cultivation outcome obtained with in-droplet and… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-1" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 1—figure supplement 1.. Workflow for the comparison of cultivation outcome obtained with in-droplet and agar plate cultivation.
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
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|
||
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|
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|
||
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|
||
data-image-alt="Figure 1—figure supplement 2."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig1s2">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-2"
|
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src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/69be8e30cd8c/elife-64774-fig1-figsupp2.gif"
|
||
alt="Figure 1—figure supplement 2." />
|
||
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|
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|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 1—figure supplement 2.. Average colony-forming unit… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
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|
||
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|
||
|
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|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 1—figure supplement 2.. Average colony-forming unit (CFU) concentration over time with different cultivation methods. </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-2" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 1—figure supplement 2.. Average colony-forming unit (CFU) concentration over time with different cultivation methods.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">Displayed is the average CFU concentration over time obtained for the extracted soil community by cultivating the soil community in droplets or on plates using the same media composition and equal incubation time. CFUs for in-droplet cultivation were determined by counting the relative abundance of droplets occupied with bacterial biomass via microscopic image analysis at given time points. We transferred abundance of occupied droplets to CFU under the assumption that one cell was initially encapsulated per occupied droplet. For example, among 1000 droplets in total 224 were occupied by colonies, meaning that in 200 nL droplet liquid (1000*200 pL) at least 224 CFU were contained. Considering the dilution of the soil community in medium to obtain droplet liquid, this amounts to 3 x 10<sup>6</sup> CFU/mL of soil community in droplets. Error bars show the standard error of the mean.</div>
|
||
|
||
|
||
|
||
|
||
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|
||
|
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|
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|
||
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|
||
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|
||
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|
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|
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|
||
<strong class="figure-label">
|
||
<p> Figure 2.. Differences in bacterial diversity due… </p>
|
||
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|
||
|
||
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|
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|
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|
||
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|
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|
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|
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|
||
<strong class="figure-label">
|
||
<p> Figure 2.. Differences in bacterial diversity due to cultivation technique. </p>
|
||
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|
||
|
||
|
||
|
||
<div class="figure-caption-contents"><p> Sample-type abbreviations represent the following:… </p></div>
|
||
|
||
|
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|
||
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|
||
|
||
<figcaption id="figure-caption-3" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 2.. Differences in bacterial diversity due to cultivation technique.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">Sample-type abbreviations represent the following: droplet PC – droplet-positive control of five selected <i>Streptomyces</i> strains (Table 1); droplet sample – droplet population inoculated with soil community; plate PC – plate-positive control of five selected <i>Streptomyces</i> strains; plate sample – agar plates inoculated with soil community; soil – original soil sample; extr. community – extracted soil community. (<b>A</b>) Principal coordinate analysis based on Bray–Curtis distance matrix visualized similarities between microbial communities. (<b>B</b>) Community structure based on Illumina amplicon sequencing of the 16S rRNA gene. The phylogenetic affiliation at the phylum level is displayed for each sample type. All samples for one sample type were merged. The bars depict the relative abundance of the 10 most abundant phyla.</div>
|
||
|
||
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||
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|
||
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|
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|
||
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|
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|
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|
||
<p> Figure 2—figure supplement 1.. Alpha diversity indices… </p>
|
||
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|
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<p> Figure 2—figure supplement 1.. Alpha diversity indices for the subsampled 16S rDNA datasets with (… </p>
|
||
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Figure 2—figure supplement 1.. Alpha diversity indices for the subsampled 16S rDNA datasets with (<b>A</b>) total number of operational taxonimic units; (<b>B</b>) Total number of Singletons; (<b>C</b>) Good's Coverage; (<b>D</b>) Chao value; (<b>E</b>) Abundance-based Coverage Estimator (ACE); (<b>F</b>) Inverse Simpson's index; (<b>G</b>) Shannon index; and (<b>H</b>) Pielou's evenness.
|
||
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|
||
|
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|
||
|
||
<div class="figure-caption-contents">Means between droplet sample and plate sample were tested by Wilcoxon rank-sum test. Cutoff for significance was p<0.05. *** Significant with p<0.001, * significant with p<0.05.</div>
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<p> Figure 3.. Comparing taxonomic classification on genus… </p>
|
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|
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<p> Figure 3.. Comparing taxonomic classification on genus rank for replicating cells in agar plate and… </p>
|
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<strong class="figure-label">
|
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Figure 3.. Comparing taxonomic classification on genus rank for replicating cells in agar plate and in-droplet cultivation.
|
||
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|
||
|
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|
||
|
||
<div class="figure-caption-contents">(<b>A</b>) Triangles depict the relative abundances for genera in eight biological replicates per droplet and plate cultivation method. Underlying boxplots show the distribution of data points. Relative abundances of operational taxonomic units (OTU) (evolutionary distance of 0.03) were agglomerated on the genus level (evolutionary distance of 0.1). Displayed are the 10 most abundant genera of in total 238 assigned genera covering 66.8% of all sequences. The 30 most abundant genera covering a total of almost 90% are shown in Figure 3—figure supplement 5. Means of relative abundances were compared by Wilcoxon rank-sum test for each genus (α = 0.05), applying Holm–Bonferroni correction for multiple comparisons. Adjusted p-values are displayed. As effect size, Cohen’s <i>d</i> was computed and plotted as bars to indicate which differences are practically relevant. The direction and the color of the bars depend on the sample type in which the larger mean was found (blue – larger mean in plate samples, green – larger mean in droplet samples). (<b>B</b>) Venn diagram for the taxa on genus level. Of 238 assigned genera, 142 were found in both cultivation methods while 93 were unique to droplet cultivation and three were only observed in plate cultivation. The area of the Venn elements corresponds to the total number of genera found for the cultivation techniques. (<b>C</b>) Venn diagram for the OTU level. Of 1687 OTUs, 677 were found in both cultivation methods, 983 were unique to droplets, and 27 were unique to plates. The area of the Venn elements corresponds to the total number of OTUs found for the cultivation techniques.</div>
|
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<p> Figure 3—figure supplement 1.. Comparing taxonomic classification… </p>
|
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<p> Figure 3—figure supplement 1.. Comparing taxonomic classification on phylum rank for replicating cells in agar… </p>
|
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<strong class="figure-label">
|
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Figure 3—figure supplement 1.. Comparing taxonomic classification on phylum rank for replicating cells in agar plate and in-droplet cultivation.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">Boxplots depict the distribution of relative abundances for phyla in droplet and plate cultivation samples. Relative abundances of operational taxonomic units were agglomerated on the phylum level. Displayed are 13 assigned phyla covering 100% of all sequences. Means of relative abundances were compared by Wilcoxon rank-sum test for each phylum (α = 0.05), applying Holm–Bonferroni correction for multiple comparisons. ** Significant with p<0.01, * significant with p<0.05. As effect size, Cohen's <i>d</i> was computed and plotted as bars to indicate which differences are practically relevant. The direction and the color of the bars depend on the sample type in which the larger mean was found (blue – larger mean in plate samples, green – larger mean in droplet samples).</div>
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<p> Figure 3—figure supplement 2.. Comparing taxonomic classification… </p>
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<p> Figure 3—figure supplement 2.. Comparing taxonomic classification on class rank for replicating cells in agar… </p>
|
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<strong class="figure-label">
|
||
Figure 3—figure supplement 2.. Comparing taxonomic classification on class rank for replicating cells in agar plate and in-droplet cultivation.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">Boxplots depict the distribution of relative abundances for phyla in droplet and plate cultivation samples. Relative abundances of operational taxonomic units were agglomerated on the class level. Displayed are the 10 most abundant classes of in total 41 assigned classes covering 99.42% of all sequences. Means of relative abundances were compared by Wilcoxon rank-sum test for each class (α = 0.05), applying Holm–Bonferroni correction for multiple comparisons. ** Significant with p<0.01, * significant with p<0.05. As effect size, Cohen's <i>d</i> was computed and plotted as bars to indicate which differences are practically relevant. The direction and the color of the bars depend on the sample type in which the larger mean was found (blue – larger mean in plate samples, green – larger mean in droplet samples).</div>
|
||
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<p> Figure 3—figure supplement 3.. Comparing taxonomic classification… </p>
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<p> Figure 3—figure supplement 3.. Comparing taxonomic classification on order rank for replicating cells in agar… </p>
|
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<strong class="figure-label">
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Figure 3—figure supplement 3.. Comparing taxonomic classification on order rank for replicating cells in agar plate and in-droplet cultivation.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">Boxplots depict the distribution of relative abundances for phyla in droplet and plate cultivation samples. Relative abundances of operational taxonomic units were agglomerated on the order level. Displayed are the 20 most abundant orders of in total 72 assigned orders covering 99.03% of all sequences. Means of relative abundances were compared by Wilcoxon rank-sum test for each order (α = 0.05), applying Holm–Bonferroni correction for multiple comparisons. ** Significant with p<0.01, * significant with p<0.05. As effect size, Cohen's <i>d</i> was computed and plotted as bars to indicate which differences are practically relevant. The direction and the color of the bars depend on the sample type in which the larger mean was found (blue – larger mean in plate samples, green – larger mean in droplet samples).</div>
|
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<p> Figure 3—figure supplement 4.. Comparing taxonomic classification… </p>
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<p> Figure 3—figure supplement 4.. Comparing taxonomic classification on family rank for replicating cells in agar… </p>
|
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Figure 3—figure supplement 4.. Comparing taxonomic classification on family rank for replicating cells in agar plate and in-droplet cultivation.
|
||
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|
||
|
||
|
||
|
||
<div class="figure-caption-contents">Boxplots depict the distribution of relative abundances for phyla in droplet and plate cultivation samples. Relative abundances of operational taxonomic units were agglomerated on the family level. Displayed are the 30 most abundant classes of in total 144 assigned families covering 96.37% of all sequences. Means of relative abundances were compared by Wilcoxon rank-sum test for each family (α = 0.05), applying Holm–Bonferroni correction for multiple comparisons. ** Significant with p<0.01, * significant with p<0.05. As effect size, Cohen's <i>d</i> was computed and plotted as bars to indicate which differences are practically relevant. The direction and the color of the bars depend on the sample type in which the larger mean was found (blue – larger mean in plate samples, green – larger mean in droplet samples).</div>
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Figure 3—figure supplement 5.. Comparing taxonomic classification on genus rank for replicating cells in agar plate and in-droplet cultivation.
|
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<div class="figure-caption-contents">Boxplots depict the distribution of relative abundances for phyla in droplet and plate cultivation samples. Relative abundances of operational taxonomic units were agglomerated on the genus level. Displayed are the 30 most abundant genera of in total 240 assigned genera covering 89.54% of all sequences. Means of relative abundances were compared by Wilcoxon rank-sum test for each genus (α = 0.05), applying Holm–Bonferroni correction for multiple comparisons. ** Significant with p<0.01, * significant with p<0.05. As effect size, Cohen's <i>d</i> was computed and plotted as bars to indicate which differences are practically relevant. The direction and the color of the bars depend on the sample type in which the larger mean was found (blue – larger mean in plate samples, green – larger mean in droplet samples).</div>
|
||
|
||
|
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|
||
|
||
</figcaption>
|
||
|
||
|
||
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|
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|
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data-label-slug="figure-3figure-supplement-6">
|
||
<a class="figure-link"
|
||
href="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/df777f46d649/elife-64774-fig3-figsupp6.jpg"
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aria-describedby="figure-caption-11"
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||
role="button"
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data-image-width="748"
|
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data-image-height="287"
|
||
data-image-alt="Figure 3—figure supplement 6."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig3s6">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-11"
|
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src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/e56ce96bb87b/elife-64774-fig3-figsupp6.gif"
|
||
alt="Figure 3—figure supplement 6." />
|
||
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|
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|
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<strong class="figure-label">
|
||
<p> Figure 3—figure supplement 6.. Detailed view into… </p>
|
||
</strong>
|
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||
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|
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|
||
|
||
|
||
|
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|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 3—figure supplement 6.. Detailed view into exclusive and uncultured taxa found in droplets or… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-11" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 3—figure supplement 6.. Detailed view into exclusive and uncultured taxa found in droplets or on plates.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">(<b>A</b>) Overview of taxa that were exclusively found in droplet samples and/or plate samples with taxonomic classification on the phylum level. (<b>B</b>) Overview of the exclusive taxa that were classified as uncultured found in droplet and/or plate samples. Taxa that were unclassified on the genus level and likely belong to the uncultured category were not considered for this comparison.</div>
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
</figure>
|
||
|
||
<figure class="figure-item tail"
|
||
itemscope itemtype="http://schema.org/ImageObject"
|
||
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|
||
data-slide-index="12"
|
||
data-label-slug="figure-4">
|
||
<a class="figure-link"
|
||
href="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/6e51fe3419b2/elife-64774-fig4.jpg"
|
||
itemprop="contentUrl"
|
||
aria-describedby="figure-caption-12"
|
||
role="button"
|
||
data-image-width="800"
|
||
data-image-height="559"
|
||
data-image-alt="Figure 4."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig4">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-12"
|
||
src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/d28570f2db7d/elife-64774-fig4.gif"
|
||
alt="Figure 4." />
|
||
</a>
|
||
|
||
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|
||
<meta itemprop="height" itemtype="http://schema.org/ImageObject" content="559">
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|
||
|
||
<div class="caption-wrap">
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 4.. Continuous droplet deposition of preincubated… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
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|
||
</div>
|
||
|
||
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 4.. Continuous droplet deposition of preincubated droplets on an agar plate. </p>
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents"><p> ( <b> A </b> )… </p></div>
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-12" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 4.. Continuous droplet deposition of preincubated droplets on an agar plate.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">(<b>A</b>) Sketch of continuous droplet deposition workflow. Droplets inoculated with cells of the soil community were incubated for 1 month before they were injected into a capillary (100 µm ID) positioned above an agar plate. The agar plate moved in x–y direction at a speed of 1 mm/s, while the droplets were injected at 10–50 droplets/s. The droplet frequency was monitored by a custom-developed sensor. (<b>B</b>) Image of the capillary taken while droplets are deposited on an agar plate. The tip of the capillary is highlighted by a white arrow. The capillary tip is close to the agar surface in order to establish a continuous fluid film. (<b>C</b>) An agar plate immediately after droplet deposition with the dried oil film in the spiral pattern of continuous deposition. (<b>D</b>) An agar plate with deposited droplets after incubation for 2 weeks. The dried oil film is still visible, on which the bacterial colonies are now aligned.</div>
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
</figure>
|
||
|
||
<figure class="figure-item tail"
|
||
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|
||
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|
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|
||
data-label-slug="figure-5">
|
||
<a class="figure-link"
|
||
href="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/cdda6a6009ec/elife-64774-fig5.jpg"
|
||
itemprop="contentUrl"
|
||
aria-describedby="figure-caption-13"
|
||
role="button"
|
||
data-image-width="800"
|
||
data-image-height="540"
|
||
data-image-alt="Figure 5."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig5">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-13"
|
||
src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/690212c1c07e/elife-64774-fig5.gif"
|
||
alt="Figure 5." />
|
||
</a>
|
||
|
||
<meta itemprop="width" itemtype="http://schema.org/ImageObject" content="800">
|
||
<meta itemprop="height" itemtype="http://schema.org/ImageObject" content="540">
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|
||
<div class="figure-caption figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="caption" aria-hidden="true">
|
||
|
||
<div class="caption-wrap">
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 5.. Abundance of classified bacterial isolates… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
</div>
|
||
|
||
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 5.. Abundance of classified bacterial isolates obtained from droplet cultivation. </p>
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents"><p> The distribution of abundance… </p></div>
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-13" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 5.. Abundance of classified bacterial isolates obtained from droplet cultivation.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">The distribution of abundance on the genus rank is displayed. Colors of the bars indicate the affiliation to phyla. Each bar is labeled with its exact relative abundance.</div>
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
</figure>
|
||
|
||
<figure class="figure-item tail"
|
||
itemscope itemtype="http://schema.org/ImageObject"
|
||
itemprop="associatedMedia"
|
||
data-slide-index="14"
|
||
data-label-slug="figure-6">
|
||
<a class="figure-link"
|
||
href="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/9f807b7d7a9f/elife-64774-fig6.jpg"
|
||
itemprop="contentUrl"
|
||
aria-describedby="figure-caption-14"
|
||
role="button"
|
||
data-image-width="800"
|
||
data-image-height="440"
|
||
data-image-alt="Figure 6."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig6">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-14"
|
||
src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/4cd59dfddab6/elife-64774-fig6.gif"
|
||
alt="Figure 6." />
|
||
</a>
|
||
|
||
<meta itemprop="width" itemtype="http://schema.org/ImageObject" content="800">
|
||
<meta itemprop="height" itemtype="http://schema.org/ImageObject" content="440">
|
||
|
||
|
||
<div class="figure-caption figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="caption" aria-hidden="true">
|
||
|
||
<div class="caption-wrap">
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 6.. Taxonomic diversity of isolates obtained… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
</div>
|
||
|
||
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 6.. Taxonomic diversity of isolates obtained in the screening for antibiotic producers. </p>
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents"><p> The classification… </p></div>
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-14" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 6.. Taxonomic diversity of isolates obtained in the screening for antibiotic producers.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">The classification is depicted on the level of order and based on the Sanger sequences of the nearly complete 16S rRNA gene. The bars depict the relative abundance of all detected orders for the isolates of all four screening rounds combined and for each screening round separately.</div>
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
</figure>
|
||
|
||
<figure class="figure-item tail"
|
||
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|
||
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|
||
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|
||
data-label-slug="figure-6figure-supplement-1">
|
||
<a class="figure-link"
|
||
href="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/81f4c3841cc3/elife-64774-fig6-figsupp1.jpg"
|
||
itemprop="contentUrl"
|
||
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|
||
role="button"
|
||
data-image-width="768"
|
||
data-image-height="587"
|
||
data-image-alt="Figure 6—figure supplement 1."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig6s1">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-15"
|
||
src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/2844c35598d3/elife-64774-fig6-figsupp1.gif"
|
||
alt="Figure 6—figure supplement 1." />
|
||
</a>
|
||
|
||
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|
||
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|
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|
||
|
||
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|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 6—figure supplement 1.. Maximum Iikelihood tree… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
</div>
|
||
|
||
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 6—figure supplement 1.. Maximum Iikelihood tree based on nearly full-length 16S rRNA gene sequence. </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-15" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 6—figure supplement 1.. Maximum Iikelihood tree based on nearly full-length 16S rRNA gene sequence.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">The tree shows the phylogenetic placement of isolates among type strains belonging to the Cytophagaceae. Bootstrap values (100 resamplings) are indicated at nodes. Isolate names starting with D were obtained in the antimicrobials-targeted isolation, while S-11-23-2/1/1 was isolated in the untargeted approach.</div>
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
</figure>
|
||
|
||
<figure class="figure-item tail"
|
||
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|
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|
||
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|
||
data-label-slug="figure-7">
|
||
<a class="figure-link"
|
||
href="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/44b15138aeb5/elife-64774-fig7.jpg"
|
||
itemprop="contentUrl"
|
||
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|
||
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|
||
data-image-width="774"
|
||
data-image-height="512"
|
||
data-image-alt="Figure 7."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig7">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-16"
|
||
src="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/c357cbe1a82f/elife-64774-fig7.gif"
|
||
alt="Figure 7." />
|
||
</a>
|
||
|
||
<meta itemprop="width" itemtype="http://schema.org/ImageObject" content="774">
|
||
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|
||
|
||
|
||
<div class="figure-caption figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="caption" aria-hidden="true">
|
||
|
||
<div class="caption-wrap">
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 7.. Detected natural product arsenal of… </p>
|
||
</strong>
|
||
|
||
|
||
|
||
|
||
|
||
|
||
</div>
|
||
</div>
|
||
|
||
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
<p> Figure 7.. Detected natural product arsenal of <i> Bacillus </i> sp. strain D121-0906-b3-2-1. </p>
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents"><p> ( <b> A </b> ) Metabolic… </p></div>
|
||
|
||
|
||
|
||
|
||
</div>
|
||
|
||
<figcaption id="figure-caption-16" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
|
||
|
||
|
||
|
||
|
||
|
||
|
||
<strong class="figure-label">
|
||
Figure 7.. Detected natural product arsenal of <i>Bacillus</i> sp. strain D121-0906-b3-2-1.
|
||
</strong>
|
||
|
||
|
||
|
||
<div class="figure-caption-contents">(<b>A</b>) Metabolic profiles of <i>Bacillus</i> sp. extracts under different growth and extraction conditions detected by liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS) (displayed as extracted ion chromatograms in negative mode). (<b>B</b>) Bacillaene A (<b>1</b>) and B (<b>2</b>) and (<b>C</b>) surfactin-like compounds (<b>3</b>–<b>5</b>) detected in extracts of <i>Bacillus</i> sp. isolate D121-0906-b3-2-1. Tandem mass spectromety (MS/MS) spectra for <b>1–5</b> are provided in Figure 7—figure supplements 4 and 5.</div>
|
||
|
||
|
||
|
||
|
||
</figcaption>
|
||
|
||
|
||
</figure>
|
||
|
||
<figure class="figure-item tail"
|
||
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|
||
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|
||
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|
||
data-label-slug="figure-7figure-supplement-1">
|
||
<a class="figure-link"
|
||
href="https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4ce8/8081529/2ec85a125a03/elife-64774-fig7-figsupp1.jpg"
|
||
itemprop="contentUrl"
|
||
aria-describedby="figure-caption-17"
|
||
role="button"
|
||
data-image-width="669"
|
||
data-image-height="712"
|
||
data-image-alt="Figure 7—figure supplement 1."
|
||
data-pmc-id="PMC8081529"
|
||
data-figure-id="fig7s1">
|
||
<img class="figure-thumb" itemprop="thumbnail"
|
||
id="article-image-17"
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<p> Figure 7—figure supplement 1.. Primary screening validation… </p>
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<p> Figure 7—figure supplement 1.. Primary screening validation bioactivity assays with D121-0906-b3-2-1 crude culture supernatant and… </p>
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Figure 7—figure supplement 1.. Primary screening validation bioactivity assays with D121-0906-b3-2-1 crude culture supernatant and selected test strains.
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<div class="figure-caption-contents">Test strains suspended in the agar are indicated in panel captions. Label denotes the medium, in which D121-0906-b3-2-1 was cultivated and the duration of incubation at sampling time point. (<b>A</b>, <b>B</b>) Medium A – MGY; medium B – MGY + Gluc; NC, negative control – MGY only; PC, positive control – amphotericin B antibiotic. (<b>C</b>) Medium A – MGY; medium B – 0.5 SM + Gluc; NC – 0.5S M; PC – chlortetracycline. (<b>D</b>) Medium A – 0.5 SM; NC, negative control – 0.5 SM; PC, positive control – ciprofloxacin antibiotic. The plate was incubated for 18 hr at imaging and is displayed as an example for the rapid growth of the <i>Bacillus</i> isolate when in contact with <i>M. vaccae</i>. For determining the antimicrobial activity of the isolate against <i>M. vaccae</i> further bioactivity plates were prepared with filtered (0.2 µm pore size) culture supernatant.</div>
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<p> Figure 7—figure supplement 2.. Metabolic profile of… </p>
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<p> Figure 7—figure supplement 2.. Metabolic profile of <i> Bacillus </i> sp. D121-0906-b3-2-1 (lower panels, soy mannitol medium… </p>
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Figure 7—figure supplement 2.. Metabolic profile of <i>Bacillus</i> sp. D121-0906-b3-2-1 (lower panels, soy mannitol medium + cold-extracted soil extract [CESE], 5 days cultivation, XAD adsorber extraction) in comparison to medium control (soy mannitol medium + CESE, XAD extraction).
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<div class="figure-caption-contents">Relative intensity according to highest peak in spectrum. Photodiode array signal (PDA) 210–600 nm. Surfactin complex including compounds <b>3</b>–<b>5</b>. TIC: total ion chromatogram.</div>
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<p> Figure 7—figure supplement 3.. Bioactivity assay with diluted fractions of <i> Bacillus </i> sp. extract against <i> B.… </i> </p>
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Figure 7—figure supplement 3.. Bioactivity assay with diluted fractions of <i>Bacillus</i> sp. extract against <i>B. subtilis</i>.
|
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<div class="figure-caption-contents">Fraction D contains bacillaenes, fractions G–H contain surfactins. Cip: ciprofloxacin; MeOH: methanol; A–H: fractions collected from a crude extract from <i>Bacillus</i> sp.</div>
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<p> Figure 7—figure supplement 4.. Characterization for bacillaene-like… </p>
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Figure 7—figure supplement 4.. Characterization for bacillaene-like compounds (<b>1</b> and <b>2</b>) in extracts of <i>Bacillus</i> sp. isolate D121-0906-b3-2-1.
|
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<div class="figure-caption-contents">Physicochemical properties of bacillaene A (<b>1</b>) and B (<b>2</b>): (<b>1</b>) electrospray ionization ESI(–) <i>m/z</i> 579 [M–H]<sup>–</sup>, high resolution electrospray ionization HRESI(–)-MS <i>m/z</i> 579.3454 (calcd. for C<sub>34</sub>H<sub>47</sub> N<sub>2</sub>O<sub>6</sub> 579.3434); (<b>2</b>) ESI(–) <i>m/z</i> 741 [M–H]<sup>–</sup>, HRESI(–)-MS <i>m/z</i> 741.3990 (calcd. for C<sub>40</sub>H<sub>57</sub> N<sub>2</sub>O<sub>11</sub> 741.3962). (<b>A</b>) Extracted ion chromatograms in negative mode of <b>1</b> and <b>2</b>. (<b>B</b>) UV absorption spectrum of <b>1</b>. (<b>C</b>) Tandem mass spectrometry (MS/MS) fragmentation pattern of <i>m/z</i> 579.3454 [M–H]<sup>–</sup> (<b>1</b>) in negative mode. (<b>D</b>) MS/MS fragmentation pattern of <i>m/z</i> 741.3987 [M–H]<sup>–</sup> (<b>2</b>) in negative mode. (<b>E</b>) MS/MS fragmentation pattern of <i>m/z</i> 581.3575 [M+H]<sup>+</sup> (<b>1</b>) in positive mode. (<b>F</b>) MS/MS fragmentation pattern of <i>m/z</i> 743.4119 [M+H]<sup>+</sup> (<b>2</b>) in positive mode.</div>
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Figure 7—figure supplement 5.. Characterization for surfactin-like compounds (<b>3</b>–<b>5</b>) in extracts of <i>Bacillus</i> sp. isolate D121-0906-b3-2-1.
|
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<div class="figure-caption-contents">Physicochemical properties of surfactins (<b>3</b> , <b>4</b>) and (<b>5</b>): (<b>3</b>) ESI(+) <i>m/z</i> 1008 [M+H]<sup>+</sup>, HRESI(+)-MS <i>m/z</i> 1008.6594 (calcd. for C<sub>51</sub>H<sub>90</sub> N<sub>7</sub>O<sub>13</sub> 1008.6597), ESI(–) <i>m/z</i> 1006 [M–H]<sup>–</sup>, HRESI(–)-MS <i>m/z</i> 1006.6480 (calcd. for C<sub>51</sub>H<sub>88</sub> N<sub>7</sub>O<sub>13</sub> 1006.6446); base-hydrolyzed (<b>3</b>) ESI(+) <i>m/z</i> 1026 [M+H]<sup>+</sup>, HRESI(+)-MS <i>m/z</i> 1026.6676 (calcd. for C<sub>51</sub>H<sub>92</sub>N<sub>7</sub>O<sub>14</sub> 1026.6697); (<b>4</b>) ESI(–) <i>m/z</i> 1020 [M–H]<sup>–</sup>, HRESI(–)-MS <i>m/z</i> 1020.6633 (calcd. for C<sub>52</sub>H<sub>90</sub> N<sub>7</sub>O<sub>13</sub> 1020.6602); (<b>5</b>) ESI(–) <i>m/z</i> 1034 [M–H]<sup>–</sup>, HRESI(–)-MS <i>m/z</i> 1034.6791 (calcd. for C<sub>53</sub>H<sub>92</sub> N<sub>7</sub>O<sub>13</sub> 1034.6759). (<b>A</b>) Comparison of surfactin standard (Merck) and compound <b>3</b> as extracted ion chromatograms. (<b>B</b>) MS/MS fragmentation pattern of <i>m/z</i> 1008.6594 [M+H]<sup>+</sup> (<b>3</b>) in positive mode. (<b>C</b>) MS/MS fragmentation pattern of <i>m/z</i> 1026.6683 [M+H]<sup>+</sup> (base-hydrolyzed <b>3</b>) in positive mode. (<b>D</b>) MS/MS fragmentation pattern of <i>m/z</i> 1006.6480 [M–H]<sup>–</sup> (<b>3</b>) in negative mode. (<b>E</b>) MS/MS fragmentation pattern of <i>m/z</i> 1020.6638 [M–H]<sup>–</sup> (<b>4</b>) in negative mode. (<b>F</b>) MS/MS fragmentation pattern of <i>m/z</i> 1034.6791 [M–H]<sup>–</sup> (<b>5</b>) in negative mode.</div>
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Abate AR, Hung T, Mary P, Agresti JJ, Weitz DA. High-throughput injection with microfluidics using picoinjectors. PNAS. 2010;107:19163–19166. doi: 10.1073/pnas.1006888107.
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Akselband Y, Cabral C, Castor TP, Chikarmane HM, McGrath P. Enrichment of slow-growing marine microorganisms from mixed cultures using gel microdrop (GMD) growth assay and fluorescence-activated cell sorting. Journal of Experimental Marine Biology and Ecology. 2006;329:196–205. doi: 10.1016/j.jembe.2005.08.018.
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Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the keio collection. Molecular Systems Biology. 2006;2:1–11. doi: 10.1038/msb4100050.
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Baret JC, Miller OJ, Taly V, Ryckelynck M, El-Harrak A, Frenz L, Rick C, Samuels ML, Hutchison JB, Agresti JJ, Link DR, Weitz DA, Griffiths AD. Fluorescence-activated droplet sorting (FADS): efficient microfluidic cell sorting based on enzymatic activity. Lab on a Chip. 2009;9:1850–1858. doi: 10.1039/b902504a.
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Ben-Dov E, Kramarsky-Winter E, Kushmaro A. An in situ method for cultivating microorganisms using a double encapsulation technique. FEMS Microbiology Ecology. 2009;68:363–371. doi: 10.1111/j.1574-6941.2009.00682.x.
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