The ability to survive changing environmental conditions has enabled certain insects to exist for millions of years. Termites, for example, have inhabited Earth for around 200 million years, and some species have the ability to survive periods of anoxia (Henderson 2001). Termites cost the United States close to 2 billion dollars annually (Kowalsick 2004). The reason for this is because landscape mulches are a source of cellulose, which attracts termites and may lead to house damage (Duryea et al 1999). In areas that flood for days or weeks at a time, termites can still pose problems to wooden structures. Therefore, it is beneficial to understand the physiology of termites and to explore how anoxic conditions affect the consumption rate of termites.
The Eastern subterranean termite, Reticulitermes flavipes, is native to the United States. It is one of the most common and widespread species of termites in the eastern region of North America and ranges from Toronto, Ontario to the gulf coast and eastward from the Great Lakes to the Atlantic coast (Krishna 1970). Along with the southern subterranean termite, R. virginicus, R. flavipes has the ability to forage for food 75 meters from the colony. Ninety-five percent of termite damage to wood comes from these two species (Duryea et al 1999).
The digestive process in the termite gut is extremely complicated but necessary to the survival of the subterranean termite. In the foregut and midgut of the termite, the termite uses its own enzymes to break down some cellulose. The cellulolytic protozoa in the hindgut hydrolyze, or add water to, cellulose, which breaks down this complex carbohydrate into individual glucose molecules. Next, each molecule of glucose ferments into two acetate compounds, two carbon dioxide molecules, and four hydrogen molecules (Brauman et al 1992). There is also some ATP produced during this process. Bacteria referred to as ectobionts or epibionts also live in or on these protozoa (Ohkuma 2001). Some of these bacteria are responsible for converting the hydrogen and carbon dioxide products from the fermentation process into another acetate compound. The three acetate compounds made per glucose molecule are oxidized by the termite to produce carbon dioxide and water (Brauman et al 1992). The water aids in the respiratory system of the termite, and the carbon dioxide is used and reduced by methanogens (Brauman et al 1992).
Subterranean termites need warmth, moisture, and cellulose to survive, and landscape mulches provide all three. Attracting termites in this way could be a common problem, since people often use landscape mulches around their houses to conserve water, control weeds, improve soil content, or simply enhance their yards. Termites have been shown to consume certain mulches like pine bark, cypress, melaleuca, and pine straw (Duryea et al 1999). This may be due to component factors, such as the presence of certain chemicals in the mulches. Some mulches produce carbon dioxide as they decompose, and subterranean termites are more attracted to soil that has a continual supply of carbon dioxide (Bernklau et al 2005). Also, it is thought that termites are attracted to areas with higher concentrations of nitrogen since their natural diet is usually low in nitrogen (Breznak and Potrikus 1981). Some types of mulches, like pine straw, put high amounts of nitrogen into the soil during the decomposition process (Duryea and English 1999).
While some common types of mulches attract termites, other types have been shown to repel them (Duryea et al 1999). This may be due to high concentrations of lignin, which interferes with the breakdown of cellulose in the termite digestive system (Melillo and Muratore 1982). It has also been shown that termites survive just as long while starving than they do while feeding on mulch, which may show that some types of mulches may not contain the essential nutrients for long-term termite survival (Long et al 2001).
Even though termites are attracted to the moisture that mulches provide, too much water can affect the physiology of R. flavipes. Termites have the innate ability to survive hypoxic conditions, like flooding. For example, R. flavipes can survive for approximately 19 hours totally submerged in water (Forschler and Henderson 1995). This research indicates that termites escape drowning in areas prone to flooding by entering a quiescent state instead of seeking higher ground (Forschler and Henderson 1995).
The bacteria and protozoa living in the termite gut have the ability to survive extreme conditions like anoxia. However, they may be stressed when the termite is submerged in water because of the limited food supply. The protozoa flagellates and the bacteria in the hindgut are responsible for degrading cellulose and producing acetate as a source of carbon for the termite to absorb and use for energy (Ohkuma 2001). If the gut symbiotes are stressed, the termite would probably be affected. There has been no known research that has tested whether the feeding behaviors of subterranean termites are affected by their ability to lower their metabolism when in an anoxic environment.
The purpose of our study was to determine which mulches R. flavipes prefer and whether termites that have survived a flooded environment can recover and feed at the same rate as termites that have not experienced a flooded environment. The results could lead to new ways to control termites. The data collected will be used to better understand the physiology of the R. flavipes and how they respond to certain environmental stresses.
Materials and Methods
This experiment was conducted to test the hypothesis that the consumption rate of landscape mulches by post-anoxic Reticulitermes flavipes is equal under normal conditions and post-anoxic conditions. R. flavipes workers were ordered from Carolina Biological Company. The laboratory setup consisted of an incubator, an electronic balance, and 32 Petri dishes. Four different types of mulches were used in both experiments. These included cedar, cypress, pine bark, and mixed hardwood. The mulches were collected from a lawn and garden store in Statesville, NC.
In Experiment 1, each type of mulch was air-dried for two days. The mulches were then weighed to the nearest one-thousandth of a gram. The mulches were placed in their respective Petri dishes, and 5 mL of water was added to the mulch in each Petri dish. The Petri dishes were labeled appropriately. Thirty worker termites were then added to each Petri dish. The dishes were covered but not sealed to allow moisture inside the dishes. All four dishes were then placed in an incubator at 24˚C. Humidity was created by placing a metal pan filled with water at the bottom of the incubator. A digital hygrometer was used to monitor the humidity level, which fluctuated between 85-90%. The termites were monitored for two weeks, and the number of termites that died in each dish was recorded. At the end of the two weeks, the mulch samples were air-dried for two days to standardize the weight. Each type of mulch was then weighed. The percentage of mulch consumed was calculated and recorded. This experiment was repeated four times and the results were recorded. The results from this experiment served as the control group.
The methods for Experiment 2 were the same as for Experiment 1, with a few exceptions. Water was boiled to decrease the oxygen. Thirty termite workers were placed in each of four petri dishes, and the dishes were filled with 20 mL of the deoxygenated water. The termites were submerged in water for two hours before being placed in the dishes with mulch. Lids were used to cover the dishes, and they were again placed in the incubator at 24˚C with 90% humidity. The termites that died at the end of the two weeks were recorded. At the end of two weeks, the mulch was dried and weighed. This experiment was also replicated four times.
The data from these two experiments were subjected to an analysis of variance. The analysis determined if there was a significant difference in percentage consumed among the four types of mulches. It was also used to determine whether there was a significant relationship between post-anoxic and/or normal termites and mulch consumption and/or survival. After determining the significant level for the analysis, a Tukey test was employed for each variable. The average consumption rates of both groups of termites were plotted as a bar graph.
Temperature and humidity levels had to be monitored because both factors are very important to the survival of termites. The wood termites feed on usually contains between 8% to 12% moisture (Duryea et al. 1999). If there is no moisture available in the soil or no soil at all, termites need a minimum of 30% moisture to survive (Anonymous 1997). This is why humidity levels were kept between 85% and 90%. Temperature is also vital to termite survival. In lower temperatures, the protozoa in the hindguts of eastern subterranean termites have been shown to feed at slower rates (Belitz and Waller 1998). Therefore, the temperature was held constant at 24˚C.
In Experiment 1, the termites consumed all four types of mulch under normal conditions. As shown in figure 3, termites consumed on average 10.6% of the pine bark mulch, 43.9% of the cypress, 16.8% of the hardwood, and 11.9% of the cedar. Thus, termites under normal conditions were shown to consume cypress mulch at a significantly higher rate than pine bark mulch, mixed hardwood, or cedar mulch. This result is consistent with other experiments conducted by Duryea et al. (1999). All of the termites used in Experiment 1 survived the two weeks in the Petri dishes.
Experiment 2 showed that termites in distress would also consume landscape mulches. After the water was removed from the dishes, the termites instantly moved towards the mulch. On average, the termites in post-anoxia consumed 12.7% of the pine bark, 21.3% of the cypress, 10.4% of the hardwood, and 6.4% of the cedar. These results are shown in figure 3. The termites all survived the two hours submerged in water and the two weeks in the dishes with the mulch.
The results of the two experiments were compared to determine if there was a significant difference between the consumption rates of termites in normal conditions and those in post-anoxic conditions. All of the termites survived the two weeks, so survival rate was not a factor. Mulch type was used as the factor and the percentage of consumption was used as the variable. The results showed that termites in post-anoxia consumed mulch at a significantly lower rate than termites in normal conditions (p=0.022).
In both experiments, the termites consumed a higher percentage of cypress mulch than any other type of mulch. When the data from both experiments were combined, it was found that termites consumed cypress mulch at a significantly higher rate than the other three mulches. Another study that tested the consumption rates of seven different types of mulches showed that termites consumed a mulch mixture composed of oaks, camphor, cedar, and southern pines at a higher rate than other mulches (Duryea et al. 1999).
Termites in Experiment 2 consumed mulch at a lower rate than the termites in Experiment 1. Hoback and Stanly (2001) stated that many different types of insects are able to survive flooded conditions by entering a quiescent state. Forschler and Henderson (1995) showed that termites could do this and survive. However, the results from this experiment indicate that placing termites in an anoxic environment for a certain amount of time (two hours in this case) does affect their feeding behavior.
Cleveland described the importance of the relationship between the microorganisms in the termite guts and the termites (1924). It has been shown that termites have the ability to live exclusively on a cellulose diet and survive indefinitely (Cleveland 1925). This may explain why all of the termites in Experiments 1 and 2 survived. It has also been shown that when the gut symbiotes of the termite are stressed or killed, the termite cannot function normally, and will not survive in some cases. Cleveland stated that termites cannot survive when the protozoa are removed from their guts (1928). However, Cleveland also showed that termites fed broken-down wood can survive even without the protozoa in their guts (Cleveland 1924). This indicates that the protozoa are necessary for the subterranean termite to break down wood.
The gut symbiotes of the termites in Experiment 2 may have been stressed to a point but were not killed. The termites may have consumed mulch at a slower rate because the microorganisms were not able to breakdown and digest the cellulose at a normal rate. The gut symbiotes of the termites in Experiment 1 did not experience any type of stress. Therefore, the termites consumed mulch at a higher rate.
There are several questions that need to be answered in the future. Will survival rates differ among termites in various types of mulches if the termites are allowed to stay in the dishes longer than two weeks? Will submerging termites in water for a longer period of time actually kill the gut symbiotes and therefore kill the termites? Can flooding be used as a control measure against termites that may infest landscape mulches? Further research is warranted to find answers to these specific questions.
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