A Straightforward Method for the Isolation and Cultivation of Galleria mellonella Hemocytes

Admella, Joana; Torrents, Eduard

doi:10.3390/ijms232113483

Cited by 7 publications

(7 citation statements)

References 42 publications

(53 reference statements)

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“…The filter diameter of 50 µm used in the method exceeds that of the largest described haemocyte sub-population by 2.5-fold (oenocytoids at 19 µm) [ 46 , 47 ]. Similarly, whilst some hemoyctes, such as plasmatocytes and granulocytes are known to be adherent [ 48 ] and may therefore stick to the filter, the loss of these cells would be identifiable following flow cytometry of centrifuged, unfiltered samples ( Figure 2 ). Therefore, the simplest explanation for this discrepancy is that the previously described subpopulations of haemocytes [ 32 ] are artefacts from sample processing.…”

Section: Discussionmentioning

confidence: 99%

Characterising phagocytes and measuring phagocytosis from live Galleria mellonella larvae

Campbell,

Pearce,

Bebes

et al. 2024

Virulence

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Over the last 20 years, the larva of the greater waxmoth, Galleria mellonella , has rapidly increased in popularity as an in vivo mammalian replacement model organism for the study of human pathogens. Experimental readouts of response to infection are most often limited to observing the melanization cascade and quantifying larval death and, whilst transcriptomic and proteomic approaches, and methods to determine microbial load are also used, a more comprehensive toolkit of profiling infection over time could transform the applicability of this model. As an invertebrate, Galleria harbour an innate immune system comprised of both humoral components and a repertoire of innate immune cells – termed haemocytes. Although information on subtypes of haemocytes exists, there are conflicting reports on their exact number and function. Flow cytometry has previously been used to assay Galleria haemocytes, but protocols include both centrifugation and fixation – physical methods which have the potential to affect haemocyte morphology prior to analysis. Here, we present a method for live haemocyte analysis by flow cytometry, revealing that Galleria haemocytes constitute only a single resolvable population, based on relative size or internal complexity. Using fluorescent zymosan particles, we extend our method to show that up to 80% of the Galleria haemocyte population display phagocytic capability. Finally, we demonstrate that the developed assay reliably replicates in vitro data, showing that cell wall β-1,3-glucan masking by Candida albicans subverts phagocytic responses. As such, our method provides a new tool with which to rapidly assess phagocytosis and understand live infection dynamics in Galleria .

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Section: Discussionmentioning

confidence: 99%

Characterising phagocytes and measuring phagocytosis from live Galleria mellonella larvae

Campbell,

Pearce,

Bebes

et al. 2024

Virulence

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“…[41], McCloskey et al . [42] and Admella & Torrents [43]. We sterilized the blade with 70% ethanol between different larvae.…”

Section: Methodsmentioning

confidence: 99%

“…Then, we opened each larva with a surgical blade (Aesculap AG, Germany) behind the posterior prolegs, with the cut length spanning half of the body width. This procedure is similar to the one used by Harding et al [41], McCloskey et al [42] and Admella & Torrents [43]. We sterilized the blade with 70% ethanol between different larvae.…”

Section: (D) Bacterial Load In the Haemolymphmentioning

confidence: 99%

Negative interactions and virulence differences drive the dynamics in multispecies bacterial infections

2023

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Bacterial infections are often polymicrobial, leading to intricate pathogen–pathogen and pathogen–host interactions. There is increasing interest in studying the molecular basis of pathogen interactions and how such mechanisms impact host morbidity. However, much less is known about the ecological dynamics between pathogens and how they affect virulence and host survival. Here we address these open issues by co-infecting larvae of the insect model host Galleria mellonella with one, two, three or four bacterial species, all of which are opportunistic human pathogens. We found that host mortality was always determined by the most virulent species regardless of the number of species and pathogen combinations injected. In certain combinations, the more virulent pathogen simply outgrew the less virulent pathogen. In other combinations, we found evidence for negative interactions between pathogens inside the host, whereby the more virulent pathogen typically won a competition. Taken together, our findings reveal positive associations between a pathogen's growth inside the host, its competitiveness towards other pathogens and its virulence. Beyond being generalizable across species combinations, our findings predict that treatments against polymicrobial infections should first target the most virulent species to reduce host morbidity, a prediction we validated experimentally.

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“…Then, we opened each larva with a surgical blade (Aesculap AG, Germany) behind the posterior prolegs, with the cut length spanning half of the body width. This procedure is similar to the one used by Harding et al (50), McCloskey et al (51), and Admella & Torrents (52). We sterilized the blade with 70% ethanol between different larvae.…”

Section: Bacterial Load In the Hemolymphmentioning

confidence: 99%

Negative interactions and virulence differences drive the dynamics in multispecies bacterial infections

Schmitz

Allen

Kümmerli

2022

Preprint

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Bacterial infections are often polymicrobial, leading to intricate pathogen-pathogen and pathogen-host interactions. There is increasing interest in studying the molecular basis of pathogen interactions and how such mechanisms impact host morbidity. However, much less is known about the ecological dynamics between pathogens and how they affect virulence and host survival. Here we address these open issues by co-infecting larvae of the insect model host Galleria mellonella with one, two, three or four bacterial species, all of which are opportunistic human pathogens. We found that virulence was always driven by the most virulent species regardless of the number of species and pathogen combinations injected. Moreover, we observed a link between a pathogen's virulence and its growth within the host. In certain cases, the more virulent pathogen simply outgrew the less virulent pathogen. In other cases, we found evidence for negative interactions between pathogens inside the host, whereby the more virulent pathogen typically won a competition. Taken together, our findings reveal positive links between a pathogen's growth inside the host, its competitiveness towards other pathogens, and its virulence. Beyond being generalizable across species combinations, our findings suggest that treatment strategies against polymicrobial infections should target the most virulent species.

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A Straightforward Method for the Isolation and Cultivation of Galleria mellonella Hemocytes

Cited by 7 publications

References 42 publications

Characterising phagocytes and measuring phagocytosis from live Galleria mellonella larvae

Characterising phagocytes and measuring phagocytosis from live Galleria mellonella larvae

Negative interactions and virulence differences drive the dynamics in multispecies bacterial infections

Negative interactions and virulence differences drive the dynamics in multispecies bacterial infections

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