Possible impacts of the predominant Bacillus bacteria on the Ophiocordyceps unilateralis s. l. in its infected ant cadavers

Animal hosts infected and killed by parasitoid fungi become nutrient-rich cadavers for saprophytes. Bacteria adapted to colonization of parasitoid fungi can be selected and can predominate in the cadavers, actions that consequently impact the fitness of the parasitoid fungi. In Taiwan, the zombie fungus, Ophiocordyceps unilateralis sensu lato (Clavicipitaceae: Hypocreales), was found to parasitize eight ant species, with preference for a principal host, Polyrhachis moesta. In this study, ant cadavers grew a fungal stroma that was predominated by Bacillus cereus/thuringiensis. The bacterial diversity in the principal ant host was found to be lower than the bacterial diversity in alternative hosts, a situation that might enhance the impact of B. cereus/thuringiensis on the sympatric fungus. The B. cereus/thuringiensis isolates from fungal stroma displayed higher resistance to a specific naphthoquinone (plumbagin) than sympatric bacteria from the environment. Naphthoquinones are known to be produced by O. unilateralis s. l., and hence the resistance displayed by B. cereus/thuringiensis isolates to these compounds suggests an advantage to B. cereus/thuringiensis to grow in the ant cadaver. Bacteria proliferating in the ant cadaver inevitably compete for resources with the fungus. However, the B. cereus/thuringiensis isolates displayed in vitro capabilities of hemolysis, production of hydrolytic enzymes, and antagonistic effects to co-cultured nematodes and entomopathogenic fungi. Thus, co-infection with B. cereus/thuringiensis offers potential benefits to the zombie fungus in killing the host under favorable conditions for reproduction, digesting the host tissue, and protecting the cadaver from being taken over by other consumers. With these potential benefits, the synergistic effect of B. cereus/thuringiensis on O. unilateralis infection is noteworthy given the competitive relationship of these two organisms sharing the same resource.

Bacillus Thuringiensis Predominated in the Zombie Fungus Ophiocordyceps Unilateralis S. L.-Infected Ant Cadavers and Investigation of its Biological Properties

Animal hosts infected and killed by parasitoid fungi become a nutrient-rich cadaver for saprophytes. Bacteria adapted to colonization of parasitoid fungi can be selected and can predominate in the cadaver, actions that consequently impact the fitness of the parasitoid fungi. In Taiwan, the zombie fungus, Ophiocordyceps unilateralis sensu lato (Clavicipitaceae: Hypocreales), was found to parasitize eight ant species, with preference for a principal host, Polyrhachis moesta. In this study, ant cadavers grew a fungal stroma that was predominated by Bacillus thuringiensis. The bacterial diversity in the principal ant host was found to be lower than the bacterial diversity in alternative hosts, a situation that might enhance the impact of B. thuringiensis on the sympatric fungus. The B. thuringiensis isolates from fungal stroma displayed higher resistance to a specific naphthoquinone (plumbagin) than B. thuringiensis from the environment. Naphthoquinones are known to be produced by O. unilateralis s. l., and hence the resistance displayed by B. thuringiensis isolates to these compounds suggests an advantage to B. thuringiensis to grow in the ant cadaver. Bacteria proliferating in the ant cadaver inevitably compete for resources with the fungus. However, the B. thuringiensis isolates displayed in vitro capabilities of hemolysis, production of hydrolytic enzymes, and antagonistic effects to co-cultured nematodes and entomopathogenic fungi. Thus, co-infection with B. thuringiensis offers potential benefits to the zombie fungus in killing the host under favorable conditions for reproduction, digesting the host tissue, and protecting the cadaver from being taken over by other consumers. With these potential benefits, B. thuringiensis is not entirely a competitor but could also be a synergist in O. unilateralis s. l. infections under certain conditions.

Zombie ant graveyard dynamics in Gunung Mulu National Park

Ophiocordyceps is a genus of pathogenic fungi, which predominantly parasitise insects. This study investigates the spatial dynamics of zombie ant graveyards, and explores the optimal height for Ophiocordyceps unilateralis spore dispersal in a Bornean rainforest. While there is considerable research derived from alternative tropical regions, there is limited documentation of Ophiocordyceps fungi in Borneo. This paper aims to build on the current body of knowledge, focusing on the spatial dynamics of zombie ant graveyards, and in particular the height at which infected ants are found. In the present study, an area of Gunung Mulu National Park was searched for O. unilateralis-infected ants. Once an infected ant was located, the surrounding area was methodically searched to allow for the height and location of all surrounding ants to be recorded. Infected ants were found at variable heights between the four sites (means of 28.9–57.6 cm) above the expected height laid out in similar studies (approximately 25 cm). It is suggested that these heights may correspond to locations at which temperature and humidity are optimal for spore dispersal and fungal growth and that these heights differ depending on unique features of the environment.

The genus Simplicillium

Simplicillium species have a wide host range and an extensive distribution. Some species are associated with rusts, as well as other plant pathogenic fungi and play an important role in biological control. In this study, two specimens of Simplicillium were collected from Chiang Mai Province, Thailand. Simplicillium formicae sp. nov. was isolated from an infected ant and S. lanosoniveum from Ophiocordyceps unilateralis which is a new host record. Species were initially identified using ITS gene sequences and confirmed using morphology coupled with phylogenetic analyses of a combined nrLSU, nrSSU, TEF and RPB1 dataset. Simplicillium formicae differs from other species in the genus by the presence of flask-shaped synnemata and phialides with intercalary nodes. Simplicillium lanosoniveum resembles other collections of the species by its completely solitary, tapering phialides and globose to ellipsoidal conidia which adhere in a slimly head. A key to species of Simplicillium is also provided.

Specialist and Generalist Fungal Parasites Induce Distinct Biochemical Changes in the Mandible Muscles of Their Host

Some parasites have evolved the ability to adaptively manipulate host behavior. One notable example is the fungus Ophiocordyceps unilateralis sensu lato, which has evolved the ability to alter the behavior of ants in ways that enable fungal transmission and lifecycle completion. Because host mandibles are affected by the fungi, we focused on understanding changes in the metabolites of muscles during behavioral modification. We used High-Performance Liquid Chromatography-Mass/Mass (HPLC-MS/MS) to detect the metabolite difference between controls and O. unilateralis-infected ants. There was a significant difference between the global metabolome of O. unilateralis-infected ants and healthy ants, while there was no significant difference between the Beauveria bassiana treatment ants group compared to the healthy ants. A total of 31 and 16 of metabolites were putatively identified from comparisons of healthy ants with O. unilateralis-infected ants and comparisons of B. bassiana with O. unilateralis-infected samples, respectively. This result indicates that the concentrations of sugars, purines, ergothioneine, and hypoxanthine were significantly increased in O. unilateralis-infected ants in comparison to healthy ants and B. bassiana-infected ants. This study provides a comprehensive metabolic approach for understanding the interactions, at the level of host muscles, between healthy ants and fungal parasites.

Three-dimensional visualization and a deep-learning model reveal complex fungal parasite networks in behaviorally manipulated ants

Some microbes possess the ability to adaptively manipulate host behavior. To better understand how such microbial parasites control animal behavior, we examine the cell-level interactions between the species-specific fungal parasite Ophiocordyceps unilateralis sensu lato and its carpenter ant host (Camponotus castaneus) at a crucial moment in the parasite’s lifecycle: when the manipulated host fixes itself permanently to a substrate by its mandibles. The fungus is known to secrete tissue-specific metabolites and cause changes in host gene expression as well as atrophy in the mandible muscles of its ant host, but it is unknown how the fungus coordinates these effects to manipulate its host’s behavior. In this study, we combine techniques in serial block-face scanning-electron microscopy and deep-learning–based image segmentation algorithms to visualize the distribution, abundance, and interactions of this fungus inside the body of its manipulated host. Fungal cells were found throughout the host body but not in the brain, implying that behavioral control of the animal body by this microbe occurs peripherally. Additionally, fungal cells invaded host muscle fibers and joined together to form networks that encircled the muscles. These networks may represent a collective foraging behavior of this parasite, which may in turn facilitate host manipulation.

Unravelling the diversity behind the Ophiocordyceps unilateralis (Ophiocordycipitaceae) complex: Three new species of zombie-ant fungi from the Brazilian Amazon

In tropical forests, one of the most commonly encountered relationships between parasites and insects is that between the fungus Ophiocordyceps (Ophiocordycipitaceae, Hypocreales, Ascomycota) and ants, especially within the tribe Camponotini. Here, we describe three newly discovered host-specific species, Ophiocordyceps camponoti-atricipis, O. camponoti-bispinosi and O. camponoti-indiani, on Camponotus ants from the central Amazonian region of Brazil, which can readily be separated using morphological traits, in particular the shape and behavior of the ascospores. DNA sequence data support inclusion of these species within the Ophiocordyceps unilateralis complex.