Disease in the Society: Infectious Cadavers Result in Collapse of Ant Sub-Colonies

<div><p>Despite the growing number of experimental studies on mechanisms of social immunity in ant societies, little is known about how social behavior relates to disease progression within the nests of ants. In fact, when empirically studying disease in ant societies, it is common to remove dead ants from experiments to confirm infection by the studied parasite. This unfortunately does not allow disease to progress within the nest as it may be assumed would happen under natural conditions. Therefore, the approach taken so far has resulted in a limited knowledge of diseases dynamics within the nest environment. Here we introduced a single infectious cadaver killed by the fungus <i>Beauveria bassiana</i> into small nests of the ant <i>Camponotus castaneus</i>. We then observed the natural progression of the disease by not removing the corpses of the ants that died following the first entry of the disease. Because some behaviors such as social isolation of sick individuals or the removal of cadavers by nestmates are considered social immune functions and thus adaptations at the colony level that reduce disease spread, we also experimentally confined some sub-colonies to one or two chamber nests to prevent the expression of such behaviors. Based on 51 small nests and survival studies in 1,003 ants we found that a single introduced infectious cadaver was able to transmit within the nest, and social immunity did not prevent the collapse of the small sub-colonies here tested. This was true whether ants did or did not have the option to remove the infectious cadaver. Therefore, we found no evidence that the typically studied social immunity behaviors can reduce disease spread in the conditions here tested.</p></div

Fungal disease progression within small sub-colonies of ants.

<p>The percentage of dead ants across 28 days of experiment. White means all ants are alive and red means all ants have died.</p

Estimation of percentage of ants infected by the introduced infectious cadavers.

<p>Blue bar corresponds to the percentage of ants already dead when we observed <i>B</i>. <i>bassiana</i> growth from secondary cadavers. These must have been infected by the introduced infectious cadavers. Gray bar corresponds to the percentage of ants that died after <i>B</i>. <i>bassiana</i> growth on the secondary cadavers, but before the minimal time required for the fungus to kill (time that the first ant died after exposure to the introduced cadaver). Red bar corresponds to the percentage of ants that could have been infected by either the introduced cadaver or, potentially, secondary cadavers. Note that, in this graphic, we only show the nests where fungus grew from secondary cadavers before 100% of the ants died.</p

<i>Ophiocordyceps camponoti-rufipedis</i>.

<p><b>a</b>) Section of upper part of stroma showing anamorph (<i>Hirsutella</i> A-type), with a palisade of phialides, subulate at the base and tapering to a needle-like neck producing narrowly limoniform conidia (bar  = 10 µm); <b>b</b>) Ascospores newly released onto agar, distinctly vermiform in shape (bar  = 15 µm); <b>c</b>) Ascospore germinating after 24 h on agar, with a needle-like outgrowth (capilliconidiophore) producing a terminal conidium with prominent cap (bar  = 10 µm).</p

<i>Ophiocordyceps camponoti-melanotici</i>.

<p><b>a</b>) <i>Camponotus melanoticus</i> biting into a leaf, the upper part of the stroma is forked, probably due to damage during growth (bar  = 1 mm); <b>b</b>) Detail of ascomatal cushion showing semi-erumpent ascomata with prominent ostioles (bar  = 0.25 mm); <b>c</b>) Section through cushion (bar  = 200 µm), with detail of ostiolar region (<b>d</b>, bar  = 75 µm); <b>e</b>) Ascospores released from ascomata (bar  = 25 µm), and <b>f</b>) Ascospore after one month on agar showing solitary phialide (<i>Hirsutella</i> A-type, bar  = 10 µm).</p

<i>Ophiocordyceps camponoti-novogranadensis</i>.

<p><b>a</b>) Infected <i>Camponotus novogranadensis</i> biting into a lichen epiphyte with anamorph synnemata (<i>Hirsutella</i> B-type) arising from feet (bar  = 0.3 mm), and highlighted in <b>b</b>) and <b>c</b>) (bar  = 0.4 mm); <b>d</b>) Section through ascomatal cushion (bar  = 100 µm); <b>e</b>) Ascomatal cushions with white masses of discharged ascospores (bar  = 0.3 mm).</p

<i>Ophiocordyceps camponoti-rufipedis</i>.

<p><b>a</b>) Single stroma, characteristic of <i>Ophiocordyceps unilateralis sensu lato</i>, with two lateral ascomatal cushions or plates arising from the dorsal pronotum of <i>Camponotus rufipes</i> (the red-legged ant), firmly attached to a leaf vein (bar  = 0.8 mm); <b>b</b>) Detail of fertile region showing the immersed to partially erumpent ascomata within the cushions, with the short necks or ostioles visible (bar  = 0.4 mm); <b>c</b>) Section through an ascomatal cushion showing the mainly immersed arrangement of ascomata (bar  = 150 µm), and detail of asci within chamber (<b>d</b>, bar  = 25 µm); <b>e</b>) Asci, clavate in shape and with prominent refractive cap (arrow, bar  = 7.5 µm).</p

<i>Ophiocordyceps camponoti-balzani</i>.

<p><b>a</b>) Stroma of <i>Ophiocordyceps</i> on <i>Camponotus balzani</i> (bar  = 1 mm); <b>b</b>) Detail of semi-erumpent ascomata with prominent ostioles (bar  = 0.4 mm); <b>c</b>) Section through cushion showing arrangement and semi-erumpent nature of ascomata (bar  = 150 µm); <b>d</b>) Close-ups from <b>a</b>) of mycelial cushions (sporodochia) on legs and antennae (bar  = 0.2 mm), showing <i>Hirsutella</i> C-type phialides (<b>e</b>, bar  = 10 µm) and apical conidia (<b>f</b>, bar  = 5.0 µm); <b>g</b>) Section through ascoma showing prominent ostiole and ascus tips with refractive caps (bar  = 30 µm); <b>h</b>) Ascospore, broadly cylindrical, large and multiseptate (bar  = 12.5 µm, compare with <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0017024#pone-0017024-g003" target="_blank"><b>Fig. 3b</b></a>); <b>i</b>) Ascospore after one month on agar, slightly swollen and producing a lateral swelling, probably a vestigial appressorium (bar  = 12.5 µm).</p

<i>Ophiocordyceps unilateralis</i>.

<p><b>a</b>) Original plate from the 1865 <i>Selecta Fungorum Carpologia</i> of the Tulasne brothers <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0017024#pone.0017024-Tulasne1" target="_blank">[4]</a>, illustrating the holotype of <i>Ophiocordyceps</i> (<i>Torrubia</i>) <i>unilateralis</i> and said to be on the leaf-cutting ant, <i>Atta cephalotes</i>; <b>b</b>) Detail from plate showing the distinctive pronotal plate of <i>Camponotus sericeiventris</i>, as well as a side view of the host which is clearly a carpenter ant and not a leaf-cutter; compare with <b>c</b>) Live worker of <i>C. sericeiventris</i> showing the spines on the pronotal plate (arrow).</p

<i>Ophiocordyceps camponoti-novogranadensis</i>.

<p><b>a</b>) Anamorph (<i>Hirsutella</i> B-type) showing detail of conidiogenesis (bar  = 15 µm), and biguttulate conidia (<b>b</b>, bar  = 10 µm); <b>c</b>) Asci with small but prominent caps (bar  = 10 µm). <b>d</b>) Ascospore after 48 h on agar producing four capilliconidiophores (bar  = 10 µm); <b>e</b>) Filiform ascospore (bar  = 5 µm).</p