The effects of temperature on host–pathogen interactions in D. melanogaster: Who benefits?
Every bacterial species has specific growth temperature requirements which is largely determined by the temperature requirements of its enzymes. The extent of temperature adaptation of the soil microbial community is also respiration rate, bacterial and fungal growth follow the square root relationship with Revisiting the hypothesis that fungal-to-bacterial dominance. Hypothesis: If four different anti-‐microbial agents are used on Bacillus subtilis bacteria then bacteria and inhibits the reproduction of bacteria. The next temperature, same amount of light, same time the bacteria is allowed to grow, same.
There are two primary genetic pathways that provide humoral immunity through the production of antimicrobial peptides. The Imd pathway defends against gram-negative bacteria, while the Toll pathway defends against gram-positive bacteria and fungi De Gregorio et al. In addition, insects are also capable of mounting a cellular immune response, by which they can phagocytose a variety of pathogens Rolff and Siva-Jothy, ; Wilson et al.
At the same time that researchers have uncovered the molecular basis of immunity in flies, others have focused on the molecular basis of their thermal ecology. We have long known about the effects of temperature on survival and reproduction in insects Crill et al. Other work with Drosophila has looked at temperature fluctuations in the wild, and how flies can survive extreme temperatures. At both hot and cold extremes, flies produce hsps, which provide a variety of protective mechanisms Feder and Hofmann, ; Lindquist and Craig, ; Qin et al.
Hsps are also released in response to other stresses, including infection Guedes et al. In humans, Hsp60 assists in the activation of macrophages, part of the innate immune system Kol et al. However, the direct or indirect role that hsps play in immunity in insects has not been well studied. In light of the breadth of knowledge on both the molecular biology of immune function and of thermal ecology in Drosophila melanogaster, flies offer an ideal system with which to test hypotheses about the effects of temperature on the ability of insects to survive infection.
While most work on behavioral fever has been confined to the Orthoptera, there is some information on the effects of temperature on immunity in flies.Effect of temperature on bacterial growth
For example, flies infected with a sterilizing nematode can recover their fertility when moved to higher temperatures. Unlike Orthopteran species, however, these infected flies do not appear to choose higher temperatures Ballabeni et al. One study on D. However, the effect was not consistent. Most selection lines showed an increased ability to encapsulate the wasp eggs at warmer temperatures, but some selection lines were less able to survive encapsulate in warmer environments Fellowes et al.
Here we describe experiments that examine the effects of temperature on host survival in a D. The initial experiment is used to resolve how temperature affects the ability of flies to survive bacterial infection.
We then carry out a series of further experiments to determine whether the effects of temperature on survival are due to changes in bacterial growth or host immunocompetence. We first measure the effects of temperature on bacterial growth within the host. To examine the effect temperature has on host immune function, we take an indirect approach and examine levels of gene expression in immune-related genes after an immune challenge at different temperatures.
In the wild, flies experience a range of temperatures within and between days. In a final set of studies, we examine the potential impact of these rapid changes on immune function by switching flies, either before or after infection, between thermal regimes.
Studies on the biology of aging in flies have used switch experiments to great effect. It has long been known that rearing fruit flies on a calorically restricted diet can extend lifespan Chippindale et al. By switching flies from restricted to standard diets, researchers learned that the life-extending effects of diet restriction DR are due to the immediate effects of DR on mortality, rather than long-term effects of reduced rates of damage accumulation under DR Good and Tatar, ; Mair et al.
Here we use temperature switching to determine if the initial temperature at which a non-infected fly is held can confer costs or benefits once the fly is infected and placed at a standard temperature. We also determine whether early temperature experience in infected flies alters long-term dynamics of the host—bacteria interaction, once the host is placed at a standard temperature.
Taken together, these experiments help us to better understand not only the effects of temperature on immune function, but also who is in control of these effects—the host or its pathogen. Materials and methods In the following section we describe four specific sets of experiments. The initial experiment is designed to determine directly the effect of temperature on the ability of flies to survive an intra-thoracic injection of bacteria.
Our results do not support any of the predictions of explanation 1 and although the data could be compatible with the prediction from explanation 2, based on evidence for temperature-dependence of reproductive effort and costs we propose a novel third explanation. The lake is stocked with roach, Rutilus rutilus, crucian carp, Carassius carassius, tench, Tinca tinca, and also contains three-spined sticklebacks, Gasterosteus aculeatus. In summer, particularly in the shallow littoral areas, the lake becomes densely populated with young-of-the-year roach and stickleback fry.
Identity of the Daphnia species was confirmed according to Johnson Johnson,and Scourfield and Harding Scourfield and Harding, Daphnia were raised individually in 30 mL vessels containing 22 mL of aged, filtered through Whatman grade 6 filter paper water from their source lake, with diffuse lighting from a 60 W cool fluorescent tube set to a 16 h light: The vessels were loosely suspended at the neck on racks in large water baths within a controlled temperature roomin which constant thermal conditions were maintained.
The algae was added to large volumes of experimental water, which was then divided into the individual vessels to provide fresh medium and food each day. Sinking of algae was reduced by aeration in the water bath under each experimental vessel, which caused agitation of the vessels within the racks.
Science Fair Projects - Examining the effect of temperature on bacteria
In a batch culture system such as ours, the use of a relatively small water volume in which to rear Daphnia will inevitably lead to a reduction in food concentration within experimental vessels during the 24 h period prior to replenishment. However, Lynch et al. Since the change in algal concentration would have been much less marked in the larger vessels, and these were not expected to provide limiting conditions, the data of Lynch et al. In order to ensure adequate acclimation to treatment temperatures all experimental animals were third generation descendants of a single large female isolated from stock culture, with each generation raised under experimental temperature and food conditions.
Daphnia were raised individually in the vessels from birth and carefully transferred by wide-mouth pipette to new water with food each day. Animals were measured during transfer following moults by placing them on a glass slide in a small volume of water, and recording their length excluding tail-spine under a compound microscope fitted with optical micrometer.
Any offspring produced were also removed at this time, counted, and for D. A further regression was calculated to convert offspring dry weight to egg dry weight in D.
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All regression equations were calculated from length: Experimental treatments were continued until individuals had produced five broods of offspring. Because of a thermostat malfunction, data for the D. Sample sizes were equal for predator cue and control groups and for each temperature were: Water conditioned by Chaoborus sp.
Fifty early instar D. Again, this conditioned water was filtered and aerated before a ten-fold dilution by addition of 2. Pseudomonas is an example of this group of organisms. The Streptococci are examples of this group. Superoxide dismutase catalyzes the following reaction: They are capable of both fermentation and aerobic respiration. Escherichia coli is an example of this class of organisms. Oxygen levels can be manipulated in the laboratory in several ways: In addition to lowered oxygen levels, some organisms flourish better in an enriched CO2 environment.
Effect of environmental factors on fungal and bacterial growth in soil
An indicator strip impregnated with methylene blue is used to indicate when reducing conditions anaerobic conditions have been achieved. When reduced, methylene blue is colorless, when oxidized this dye is blue. Clostridium tetani - agent of tetanus, puncture wounds, produces a toxin which enters the spinal column and blocks the inhibitory spinal motor neurons. This produces generalized muscle spasms or spastic paralysis. Clostridium botulinum - this soil organism is the causative agent of botulism which typically occurs after eating home canned alkaline vegetables which were not heated enough during canning.