Increasing adult density compromises survival following bacterial infections in Drosophila melanogaster

The density-dependent prophylaxis hypothesis predicts that risk of pathogen transmission increases with increase in population density, and in response to this, organisms mount a prophylactic immune response when exposed to high density. This prophylactic response is expected to help organisms improve their chances of survival when exposed to pathogens. Alternatively, organisms living at high densities can exhibit compromised defense against pathogens due to lack of resources and density associated physiological stress; the crowding stress hypothesis. We housed adult Drosophila melanogaster flies at different densities and measured the effect this has on their post-infection survival and resistance to starvation. We find that flies housed at higher densities show greater mortality after being infected with bacterial pathogens, while also exhibiting increased resistance to starvation. Our results are more in line with the density-stress hypothesis that postulates a compromised immune system when hosts are subjected to high densities.

Methods
This file ("Adult_density_experiment.xlsx") was generated in 2019-20 by Paresh Nath Das and others at the Evolutionary Biology Lab, IISER Mohali.

GENERAL INFORMATION

1. Title of Dataset: "Increasing adult density compromises anti-bacterial defense in Drosophila melanogaster"

2. Author Information

A. Principal Investigator Contact Information

Name: Prof. N. G. Prasad

Institution: Indian Institute of Science Education and Research, Mohali

Address: IISER Mohali, Sector 81, Knowledge City, SAS Nagar, Punjab - 140306, India.

Email: [e-mail]

B. Associate or Co-investigator Contact Information

Name: Paresh Nath Das

Institution: Indian Institute of Science Education and Research, Mohali

Address: IISER Mohali, Sector 81, Knowledge City, SAS Nagar, Punjab - 140306, India.

Email: [e-mail]

C. Associate or Co-investigator Contact Information

Name: Aabeer Kumar Basu

Institution: Indian Institute of Science Education and Research, Mohali

Address: IISER Mohali, Sector 81, Knowledge City, SAS Nagar, Punjab - 140306, India.

Email: [e-mail]

3. Duration of data collection: September 2019 - March 2020

4. Geographic location of data collection: Mohali, Punjab, India 

5. Information about funding sources that supported the collection of the data: IISER Mohali, MHRD, Govt. of India.

SHARING/ACCESS INFORMATION

Links to publications that cite or use the data: 

bioRxiv: https://doi.org/10.1101/2022.01.02.474745

Journal of Insect Physiology (in press version): https://doi.org/10.1016/j.jinsphys.2022.104415

METHODOLOGICAL INFORMATION

A. Details of fly populations

Blue Ridge Baseline (BRB) population: BRB2 is a lab-adapted, large, outbred, wild-type population of Drosophila melanogaster, maintained on a 14-day discrete generation cycle, on standard banana-jaggery-yeast medium. 

The BRB population was originally derived by hybridising 19 iso-female lines caught from the wild population at Blue Ridge Mountains, USA. The experiments reported were conducted after 200 generations of lab-adaptation.

B. Effect of density, 8 adults vs. 32 adults, on immune function and starvation resistance.

# Experimental treatments: 2-3 day old adult flies were randomly distributed into two density treatments.

a. 8 adults per vial (1:1 sex ratio)

b. 32 adults per vial (1:1 sex ratio)

Vilas of both treatments had equal amout of standard fly food (1.5-2 ml). Flies were housed like this for 48 hours, and thereafter assayed for immune function and starvation resistance.

# Immune function assay: Flies of both treatments (described above) were assayed separately for resistance against two entomopathogenic bacteria, Enterococcus faecalis and Erwinia c. carotovora, with two independent replicates per pathogen.

Within each replicate experiment, 80 males and 80 females from each treatment (described above) were subjected to infection, and 40 males and 40 females were subjected to sham-infections.

Post-infection mortality was recorded for 120 hours; during this period, flies of both treatments were housed at equal density (4 males and 4 females per vial).  

#Starvation resistance assay: Flies of both treatments (described above) were assayed for starvation resistance; experiment independently replicated twice.

Within each replicate experiment, 80 males and 80 females from each treatment (described above) were subjected to starvation in vials with non-nutritive agar gel only.

Post-starvation mortality was recorded till the last fly died; during this period, flies of both treatments were housed at equal density (4 males and 4 females per vial). 

C. Effect of density, 50 adults vs. 200 adults, on immune function and starvation resistance.

# Experimental treatments: 2-3 day old adult flies were randomly distributed into two density treatments.

a. 50 adults per vial (1:1 sex ratio)

b. 200 adults per vial (1:1 sex ratio)

Vilas of both treatments had equal amout of standard fly food (1.5-2 ml). Flies were housed like this for 48 hours, and thereafter assayed for immune function and starvation resistance.

# Immune function assay: Flies of both treatments (described above) were assayed separately for resistance against two entomopathogenic bacteria, Enterococcus faecalis and Erwinia c. carotovora, with two independent replicates per pathogen.

Within each replicate experiment, 80 males and 80 females from each treatment (described above) were subjected to infection, and 40 males and 40 females were subjected to sham-infections.

Post-infection mortality was recorded for 120 hours; during this period, flies of both treatments were housed at equal density (4 males and 4 females per vial).  

#Starvation resistance assay: Flies of both treatments (described above) were assayed for starvation resistance; experiment independently replicated twice.

Within each replicate experiment, 80 males and 80 females from each treatment (described above) were subjected to starvation in vials with non-nutritive agar gel only.

Post-starvation mortality was recorded till the last fly died; during this period, flies of both treatments were housed at equal density (4 males and 4 females per vial).