Effect of Population Structure and Size on Aggregation ... - BioOne

SAMPLING, DISTRIBUTION, DISPERSAL

Effect of Population Structure and Size on Aggregation Behavior of Cimex lectularius (Hemiptera: Cimicidae) MARGIE PFIESTER,1 PHILIP G. KOEHLER,

AND

ROBERTO M. PEREIRA

Department of Entomology, University of Florida, Bldg. 970 Natural Area Drive, Gainesville, FL 32611-0620

J. Med. Entomol. 46(5): 1015Ð1020 (2009)

ABSTRACT The bed bug, Cimex lectularius L. (Hemiptera: Cimicidae), occurs in aggregations until the conditions are no longer beneÞcial, leading to dispersal. Active and passive bed bug dispersal causes migrations from main aggregations either within a room, from room to room within a building, or from building to building. Because bed bug movement is an important factor in the spread of infestations, we wanted to determine how population structure and size affect bed bug aggregations. Engorged bed bugs were placed in glass petri dish arenas at varying densities, sex ratios, and population compositions. Nymphs had a high tendency to aggregate, varying between 94 and 98%, and therefore were not the likely dispersal stage of the bed bug. At densities of 10 and 40 adults at a 1:1 sex ratio, there were signiÞcantly more lone females than lone males. When the population composition was varied, the percentage of lone females was signiÞcantly higher than that of males and nymphs at population compositions of 40 and 80% adults. When the sex ratio of adults was varied, there were signiÞcantly more lone females than males in arenas with 20, 50, and 80% males. Females, being found away from aggregations signiÞcantly more often than any other life stage, are potentially the dispersal stage of the bed bug. Active female dispersal away from main aggregations can potentially lead to treatment failures and should be taken into account when using control methods. KEY WORDS Cimex lectularius, bed bug, dispersal, aggregation

The bed bug, Cimex lectularius L. (Hemiptera: Cimicidae), occurs in aggregations (Usinger 1966), consisting of bed bugs in all life stages, feeding statuses, and mating conditions (Johnson 1942, Reinhardt and Siva-Jothy 2007). Bed bugs may aggregate for multiple reasons. Similarly to Triatoma infestans (Klug) (Lorenzo and Lazzari 1996), bed bug aggregations may occur in places that are considered safe from predators (Pinto et al. 2007). Like Nezara viridula (L.) (Lockwood and Story 1986), aggregated bed bugs display a higher resistance to desiccation than solitary bed bugs (Benoit et al. 2007) and have a better chance in Þnding mates (Pinto et al. 2007). Bed bug aggregation behavior seems to be chemically mediated, in part by airborne aggregation pheromones (Siljander et al. 2008) as well as by contact pheromones emitted by males and nymphs (Siljander et al. 2007). Aggregations also may be initiated by the recognition of neighboring bed bugs using mechanoreceptors on antennae (Levinson et al. 1974), which may reßect the thigomotactic afÞnity of bed bugs (Usinger 1966). The persistence of aggregations depends on a balance between advantages and costs associated with them. There is usually a threshold triggered by different factors that, when reached, causes aggregations to no longer beneÞt the organism (Wertheim et al. 1

Corresponding author, e-mail: insects@uß.edu.

2005). Aggregated individuals tend to have higher competition for food, space, and mates, may be more apparent to natural predators, can cause deterioration of environmental conditions because of overuse, and can be subjected to inbreeding (Wertheim et al. 2005). These factors can lead to individual dispersal, or movement away from aggregations. Bed bug aggregation and dispersal behavior is an important factor in the spread of infestations. Dispersal of bed bugs can be either active, when they walk from one place to another, or passive, when they are carried to new locations (Usinger 1966). These dispersal methods affect the spread of bed bug infestations that occur in three ways: within a room, room to room within a building, and building to building. Bed bug migration within a room is active and seems to occur when bed bugs are searching for a host or leaving an aggregation due to competition for resources (Pinto et al. 2007). Bed bugs actively move from room to room by going through wall voids, such as those used for wires or pipes (Reinhardt and SivaJothy 2007, Pinto et al. 2007). Movement from one building to another occurs passively when bed bugs hitchhike on items such as clothing, backpacks, luggage, and furniture (Usinger 1966, Pinto et al. 2007). However, the factors motivating bed bug passive dispersal are unknown.

0022-2585/09/1015Ð1020$04.00/0 䉷 2009 Entomological Society of America

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The purpose of our study was to determine how population structure and size affect bed bug aggregation and movement. Differences in aggregation and movement tendencies may result in some life stages being found alone more often than others. Better understanding of bed bug behavior may improve our ability to manage bed bug infestations. Materials and Methods Bed Bugs. The Harlan strain (Harold Harlan, Armed Forces Pest Management Board, U.S. Department of Defense, Washington, DC) of the bed bug was reared at the University of FloridaÕs Department of Entomology and Nematology (Gainesville, FL). The insects were reared as described in PÞester et al. (2008). In summary, bed bugs were maintained in glass rearing jars lined with Þlter paper, using pieces of manila folder as harborages. Nylon mesh was placed over the mouth of the rearing jars and secured by a screw-on lid to prevent escape. Bed bugs were fed to engorgement weekly on live chickens. The insects were harvested with a camelÕs-hair paintbrush when needed and were fed ⬍2 h before use. All animals in this study were used in accordance with University of Florida Institutional Animal Care and Use Committee protocol E380. Arena Setup. A Þlter paper circle (14.5 cm in diameter) was placed into an inverted petri dish cover (150 by 20 mm; Pyrex, Corning Inc., Corning NY). After addition of bed bugs (see experiment descriptions below), the petri dish bottom was inverted into the petri dish cover to close the arena and press on the Þlter paper, preventing the bed bugs from crawling underneath. Yellow theatrical gel (⬇22 by 36 cm; SG/Lux 10, Rosco Laboratories Inc., Stamford, CT) was placed around the inverted petri dish because it made bed bugs behave as though they were in the dark. The wrapped petri dish was then placed on an inverted deli cup where bed bugs were left undisturbed and therefore free to move within the arena. The room the arenas were placed in was used daily, so the light cycle varied throughout the experiment, whereas the temperature remained steady at ⬇24⬚C with a relative humidity of ⬇46%. All experiments were performed in the same room. Early Instar Density Experiment. First-instar bed bugs were harvested and placed into the center of each arena at densities of 50, 100, and 150 bed bugs per arena. An experimental unit consisted of an arena with bed bugs, and each nymphal density was replicated Þve times for a total of 15 experimental units. Data were recorded daily at 4 p.m. on each experimental unit for 5 d. Time observations were originally designed to detect changes through time on aggregation tendencies. The data recorded included the percentage of lone nymphs, the percentage of aggregated nymphs, the number of aggregations, and the number of insects in each aggregation. For all experiments, a lone bed bug was deÞned as one separated from any other bed bug by a distance greater than the length of one adult bed bug (⬇5 mm). An aggregation was

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deÞned as two or more bed bugs separated by a distance less than or equal to the length of one adult bed bug. Adult Density Experiment. Adult bed bugs were harvested, sexed, and placed into arenas at densities of 10, 20, and 40 per arena, at a 1:1 sex ratio. Each adult density was replicated Þve times for a total of 15 experimental units. Data were recorded as described above. The data recorded included the percentage of lone males and females, the percentage of aggregated males and females, the number of aggregations, and the number and sex of insects in each aggregation. Life-Stage Experiment. Bed bug nymphs were separated into three life stages: early stage (Þrst or second instar), mid-stage (third or fourth instar), and late stage (Þfth instar). Adult and nymphal bed bugs (30 total bed bugs per arena) were harvested and placed into the center of each arena at population compositions of 20, 40, 60, and 80% adults. Adults were at a 1:1 sex ratio, and nymphs were at a 1:1:1 life-stage ratio. Each population composition was replicated four times for a total of 16 experimental units. Data were recorded as described above. The data recorded included the percentage of lone males, lone females, and lone nymphs, the percentage of aggregated bed bugs, the number of aggregations, and the number of insects in each aggregation. Sex Ratio Experiment. Adult bed bugs were harvested and sexed. Ten adult bed bugs were placed into the center of each arena at proportions of 100, 80, 50, 20, and 0% males. Each sex ratio was replicated nine times for a total of 45 experimental units. Data were recorded as described above. The data recorded included the percentage of lone males and females, the percentage of aggregated males and females, the number of aggregations, and the number of insects in each aggregation. Statistical Analysis. Percentages were arcsine square root transformed before analysis. In each experiment, data were analyzed by analysis of variation using a linear model accounting for autocorrelation among repeated observations on the same arenas over the 5-d observation period. Post hoc comparisons were conducted on values averaged over time for each treatment because effect of time was not signiÞcant at the ␣ ⫽ 0.05 level in any experiment. Treatment means were separated with StudentÐNewmanÐKeuls (P ⬍ 0.05; SAS Institute 2003). Results Early Instar Density Experiment. Released bed bugs proceeded to form aggregations along the edges of the arenas. There were few lone nymphs, ranging from ⬇1 to 3% (Table 1), and the percentage of lone nymphs was not signiÞcantly affected by density (F ⫽ 3.38; df ⫽ 2, 12; P ⫽ 0.0685), nor was the percentage of aggregated nymphs (F ⫽ 0.89; df ⫽ 2, 12; P ⫽ 0.4355). As insect density increased, the percentage of aggregated nymphs was consistent, ranging from 96 to 98%. Also, the number of aggregations remained steady as density increased, ranging from 1 to 3 (F ⫽ 1.82; df ⫽ 2, 12; P ⫽ 0.2044). Furthermore, the number

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Table 1. Effect of density (mean % ⴞ SEM) on aggregated and lone first-instar nymphal bed bugs (C. lectularius), the number of aggregations, and the number of insects in aggregations of nymph only populations Density

% lonea

% aggregatedb

No. aggregations

No. insects in aggregations

50 100 150

3.1 ⫾ 0.46 0.8 ⫾ 0.17 1.2 ⫾ 0.15

95.7 ⫾ 1.07 96.5 ⫾ 2.44 98.1 ⫾ 0.19

1.7 ⫾ 0.18 2.5 ⫾ 0.13 2.6 ⫾ 0.20

35.8 ⫾ 2.92 43.2 ⫾ 2.84 67.7 ⫾ 6.59

There were no signiÞcant differences at all variables (P ⫽ 0.05; StudentÐNewmanÐKeuls; SAS Institute 2003). a A lone bed bug was deÞned as a bed bug separated from any other bed bug by a distance greater than the length of one adult bed bug. b An aggregation was deÞned as two or more bed bugs separated by a distance less than or equal to the length of one adult bed bug.

of insects in each aggregation was not affected by density (F ⫽ 3.34; df ⫽ 2, 12; P ⫽ 0.0701), although the aggregations did grow larger as the density of bed bugs in arenas increased. Aggregations grew from 36 insects at a density of 50 to 68 insects at a density of 150. Adult Density Experiment. As density increased (Table 2), the percentage of lone females signiÞcantly decreased, from 68% at a density of 10 to 27% at a density of 40 (F ⫽ 12.80; df ⫽ 2, 12; P ⫽ 0.0011). The percentage of lone males was not affected by density (F ⫽ 2.58; df ⫽ 2, 12; P ⫽ 0.1167). The percentage of bed bugs that aggregated signiÞcantly increased as density increased, from 52 to 80% (F ⫽ 17.00; df ⫽ 2, 12; P ⫽ 0.0003), as did the number of aggregations from two to seven (F ⫽ 103.54; df ⫽ 2, 12; P ⬍ 0.0001). The number of insects in each aggregation was signiÞcantly higher at a density of 40 (F ⫽ 14.19; df ⫽ 2, 12; P ⫽ 0.0007). With 10 adults in the arenas, the percentage of lone females was signiÞcantly higher than that for lone males (F ⫽ 21.03, P ⫽ 0.0018), which also occurred when there were 40 adults in the arenas (F ⫽ 16.23, P ⫽ 0.0038). There was no signiÞcant difference between the percentage of lone males and females when there were 20 adults in the arenas (F ⫽ 1.15, P ⫽ 0.3144). At densities of 20 (F ⫽ 28.50, P ⫽ 0.0007) and 40 (F ⫽ 52.42, P ⬍ 0.0001), signiÞcantly more females were found in female-biased aggregations than in male-biased aggregations (Fig. 1), but at a density of 10 there was no signiÞcant difference (F ⫽ 4.26, P ⫽ 0.0729). Population Composition Experiment. Of the bed bugs released in the arenas, there were few lone nymphs (Table 3). The percentage of lone nymphs

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(F ⫽ 0.36; df ⫽ 3, 12; P ⫽ 0.7807), the percentage of lone adult males (F ⫽ 0.60; df ⫽ 3, 12; P ⫽ 0.6247), and the percentage of lone adult females (F ⫽ 1.33; df ⫽ 3, 12; P ⫽ 0.3117) were not affected by the population composition. Increasing the proportion of adults in the arenas did not signiÞcantly affect the percentage of bed bugs that aggregated (F ⫽ 0.76; df ⫽ 3, 12; P ⫽ 0.5404), the number of aggregations (F ⫽ 1.32; df ⫽ 3, 12; P ⫽ 0.3126), and the number of bed bugs per aggregation (F ⫽ 0.87; df ⫽ 3, 12; P ⫽ 0.4819). When the percentage of lone females, males, and nymphs were compared at each population composition, the percentage of lone females was signiÞcantly higher than that of males and nymphs at population compositions of 40 (F ⫽ 9.76; df ⫽ 2, 36; P ⫽ 0.0056) and 80% adults (F ⫽ 5.54; df ⫽ 2, 36; P ⫽ 0.0271). There was no signiÞcant difference at 20 (F ⫽ 0.16; df ⫽ 2, 36; P ⫽ 0.8537) or 60% adults (F ⫽ 1.91; df ⫽ 2, 36; P ⫽ 0.2034). Sex Ratio Experiment. Sex ratio (Table 4) signiÞcantly affected the percentage of lone males (F ⫽ 3.90; df ⫽ 3, 32; P ⫽ 0.0176) and lone females (F ⫽ 6.31; df ⫽ 3, 32; P ⫽ 0.0017). As the proportion of males increased in arenas, the proportion of lone males and lone females also increased. Similarly to the adult density experiment, the percentage of lone females was signiÞcantly higher than the percentage of lone males in populations with 20 (F ⫽ 16.61, P ⬍ 0.0015), 50 (F ⫽ 20.83, P ⬍ 0.0003), and 80% males (F ⫽ 22.65, P ⬍ 0.0002). The percentage of adult bed bugs that aggregated was not signiÞcantly affected by sex ratio (F ⫽ 1.61; df ⫽ 4, 40; P ⫽ 0.1908), although females had a greater tendency to aggregate when more females were present. Sex ratio also had no signiÞcant affect on the number of aggregations (F ⫽ 0.20; df ⫽ 4, 40; P ⫽ 0.9347) and the number of insects in each aggregation (F ⫽ 1.23; df ⫽ 4, 40; P ⫽ 0.3148). Discussion Dispersal of nymphs as seen in the German cockroach, Blatella germanica (L.) (Ross et al. 1984), did not occur in our experiments, whereas the nymphal stages of the bed bug had a strong tendency to aggregate. Nymphs, which lose water more rapidly when solitary (Benoit et al. 2007), beneÞt from aggregation behavior, and they are attracted by airborne aggregation pheromone and contact aggregation pheromone produced by other nymphs (Siljander et al. 2007, 2008). The low numbers of nymphs observed alone in

Table 2. Effect of density (mean % ⴞ SEM) on aggregated and lone adult bed bugs (C. lectularius), the number of aggregations, and the number of insects in aggregations of adult only populations with a 1:1 sex ratio Density

Lone femalesa

Lone malesa

Aggregatedb

No. aggregations

No. insects in aggregations

10 20 40

68.0 ⫾ 4.76a* 39.2 ⫾ 4.47b 26.8 ⫾ 1.47b*

27.2 ⫾ 4.14 29.2 ⫾ 4.20 14.2 ⫾ 1.91

52.4 ⫾ 3.57a 65.8 ⫾ 3.04b 79.5 ⫾ 1.35c

1.9 ⫾ 0.16a 4.3 ⫾ 0.29b 7.4 ⫾ 0.34c

3.0 ⫾ 0.18a 3.3 ⫾ 0.17a 4.4 ⫾ 0.20b

Means in a column followed by different lowercase letters are signiÞcantly different (P ⫽ 0.05; StudentÐNewmanÐKeuls; SAS Institute 2003). Lone male and female columns were compared at each density level and signiÞcantly higher percentages are indicated by an asterisk (*). a A lone bed bug was deÞned as a bed bug separated from any other bed bug by a distance greater than the length of one adult bed bug. b An aggregation was deÞned as two or more bed bugs separated by a distance less than or equal to the length of one adult bed bug.

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Fig. 1. Percentage of aggregated females in female- and male-biased aggregations at various densities. Bed bugs used were adults at a 1:1 sex ratio.

both the nymphal density experiment and the population composition experiment suggests that nymphs are better protected around adults and thus are not likely responsible for bed bug dispersal. Although females almost always represented the highest percentage of lone bed bugs, males were found alone in certain situations. As the proportion of males increased in an arena, the proportion of lone males also increased, perhaps due to the need of males to Þnd mates. Bed bug males tend to copulate as often as possible in the Þrst 36 h after feeding (Stutt and SivaJothy 2001) because engorged females display less resistance (Reinhardt et al. 2009). If females are rare, recently fed males may abandon aggregations in search of females. The presence of an aggregation pheromone in males, which arrests or attracts other males and females (Siljander et al. 2007, 2008), may help uninseminated females Þnd a mate-searching male, especially because only virgin females respond to the airborne aggregation pheromone (Siljander et al. 2008). Although our results indicate that male bed bugs may participate in movement away from aggregations, male dispersal alone cannot be responsible for population spread.

In every experiment we performed, females were found alone most often. Unlike bed bug nymphs and males, females do not seem to produce any contact aggregation pheromone (Siljander et al. 2007), contrary to previous reports (Usinger 1966, Levinson and Bar Ilan 1971), and only respond to airborne aggregation pheromone before mating (Siljander et al. 2008). By not producing or responding to aggregation pheromone, females can leave aggregations without attracting other bed bugs in the population. Thus, females can expand the population range, establish new infestation sites, and exploit new resources, whereas other bed bugs continue to use resources available at original infestation sites. The idea that the spread of bed bug infestations partially depends on bed bugs actively seeking new refugia, along with observations that female bed bugs are found away from aggregations most often in Þeld populations (Siljander et al. 2008) supports the Þndings from our experiments. Changes in the density of adults in arenas did not lead to movement away from aggregations, but we did observe some interesting aggregation behaviors. The percentage of aggregated males did not vary as density

Table 3. Effect of population composition (mean % ⴞ SEM) on aggregated and lone adult and nymphal bed bugs (C. lectularius), the number of aggregations, and the number of insects in each aggregation of mixed populations with nymphal and adult bed bugs Popa composition (% adults)

% lone femalesb

% lone malesb

% lone nymphsb

% aggregatedc

No. aggregations

No. insects in each aggregation

20 40 60 80

11.7 ⫾ 3.65 19.2 ⫾ 2.78A 19.4 ⫾ 3.94 19.6 ⫾ 2.85A

15.0 ⫾ 4.51 10.8 ⫾ 2.78B 12.2 ⫾ 3.21 5.0 ⫾ 1.64B

5.6 ⫾ 1.10 3.0 ⫾ 0.62B 5.7 ⫾ 2.57 5.0 ⫾ 1.75B

91.5 ⫾ 1.33 91.0 ⫾ 1.08 88.2 ⫾ 2.20 87.3 ⫾ 1.07

3.2 ⫾ 0.25 3.5 ⫾ 0.31 4.3 ⫾ 0.36 4.5 ⫾ 0.34

9.5 ⫾ 1.14 9.6 ⫾ 1.24 7.5 ⫾ 0.90 6.7 ⫾ 0.62

There were no signiÞcant differences between means in a column (P ⫽ 0.05; StudentÐNewmanÐKeuls; SAS Institute 2003). Lone male, female, and nymphal columns were compared at each density level and signiÞcant differences are indicated by uppercase letters. a Adult bed bugs and nymphs of all life-stages were used at varying compositions of 20, 40, 60, and 80% adults. Adults used were at a 1:1 sex ratio, and nymphal population was at a 1:1:1 life-stage ratio; early stage (Þrst or second instar), mid-stage (third or fourth instar), and late stage (Þfth instar). b A lone bed bug was deÞned as a bed bug separated from any other bed bug by a distance greater than the length of one adult bed bug. c An aggregation was deÞned as two or more bed bugs separated by a distance less than or equal to the length of one adult bed bug.

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Table 4. Effect of sex ratio (mean % ⴞ SEM) on aggregated and lone adult bed bugs (C. lectularius), the number of aggregations, and the number of insects in aggregations of adult only populations with varying sex ratio Sex ratio (% males) 0 20 50 80 100

% lone malesa 21.3 ⫾ 4.87a 20.7 ⫾ 2.64ab 35.4 ⫾ 3.28b 37.4 ⫾ 3.17b

% lone femalesa

% aggregatedb

No. aggregations

No. insects in each aggregation

25.9 ⫾ 2.31a 38.9 ⫾ 3.22ab* 50.4 ⫾ 3.20b* 54.6 ⫾ 4.99b*

74.1 ⫾ 2.31 64.6 ⫾ 2.96 64.4 ⫾ 2.16 60.7 ⫾ 2.95 62.6 ⫾ 3.17

2.2 ⫾ 0.10 2.1 ⫾ 0.13 2.1 ⫾ 0.10 2.1 ⫾ 0.11 2.2 ⫾ 0.10

3.8 ⫾ 0.25 3.6 ⫾ 0.22 3.5 ⫾ 0.20 3.0 ⫾ 0.23 2.8 ⫾ 0.18

Means in a column followed by a different letter are signiÞcantly different (P ⫽ 0.05; StudentÐNewmanÐKeuls; SAS Institute 2003). Lone male and female columns were compared at each density level, and signiÞcantly higher percentages are indicated by an asterisk (*). a A lone bed bug was deÞned as a bed bug separated from any other bed bug by a distance greater than the length of one adult bed bug. b An aggregation was deÞned as two or more bed bugs separated by a distance less than or equal to the length of one adult bed bug.

of adults increased, but the percentage of aggregated females increased with density. This increase in female aggregation occurred mostly in female-biased aggregations. A similar situation is found with the African damselßy Platycypha caligata (Selys) in which females form groups to avoid courting males, leading to a higher oviposition rate (Martens and Rehfeldt 1989, Wertheim et al. 2005). This aggregation behavior allows female bed bugs to beneÞt from aggregations in the same way that nymphs and males do (Benoit et al. 2007) but also provides a protected situation from males. In Þeld population studies, bed bug adults were found to encompass one third of populations, but no sexual bias was observed with adults existing at a 1:1 sex ratio (Johnson 1942, Newberry and Jansen 1986, Stutt and Siva-Jothy 2001). These studies, however, did not look at the composition of individual aggregations. Our results suggest that bed bugs occur in dynamic, changing aggregations, unlike the constant aggregations suggested in the literature (Usinger 1966). The higher tendency of females to be alone and the tendency for males to leave aggregations when females are rare suggest that the composition of individual aggregations may change over time, although the sex ratio of the population remains constant. Based on our studies, the aggregation-level sex ratios in Þeld populations are likely to be quite different from the population-level sex ratio. However, this hypothesis awaits conÞrmation from Þeld-collected data. Bed bug female movement away from aggregations is an important concept for bed bug management and should be taken into account during treatments. Because females are likely to move away from infestation centers, mated females seem more likely than other life stages to move into new locations. This can lead to egg dispersal and nymphal emergence away from typical bed bug aggregation foci. Females, eggs, and emerging nymphs away from aggregations may be at a lower risk of exposure to localized bed bug control actions that target established bed bug aggregation sites. Thus, the aggregation behavior reported here should be considered as a potential cause for bed bug control treatment failures. These behaviors also stress the importance of using adequate detection methods,

such as trained canines that have the ability to detect small, early infestations and viable eggs (PÞester et al. 2008), which would result from females leaving aggregation centers. Although our experiments focused on aggregation behaviors, our results provide limited insight into potential dispersal behavior of bed bugs. Bed bug aggregation and dispersal behaviors may have several effects on population spread, response to control measures, and resistance to insecticides. Acknowledgments We thank Josh Weston for assistance in feeding and maintaining bed bug colonies. We also thank Meghan Brennan for help with statistical analysis.

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Pfiester, M., P. G. Koehler, and R. M. Pereira. 2008. Ability of bed bug-detecting canines to locate live bed bugs and viable bed bug eggs. J. Econ. Entomol. 101: 1389 Ð1396. Pinto, L. J., R. Cooper, and S. K. Kraft. 2007. Bed bug handbook: the complete guide to bed bugs and their control. Pinto & Associates, Mechanicsville, MD. Reinhardt, K., R. Naylor, and M. T. Siva-Jothy. 2009. Situation exploitation: higher male mating success when female resistance is reduced by feeding. Evolution 63: 29Ð39. Reinhart, K., and M. T. Siva-Jothy. 2007. Biology of the bed bugs (Cimicidae). Annu. Rev. Entomol. 52: 352Ð374. Ross, M. H., B. L. Bret, and C. B. Keil. 1984. Population growth and behavior of Blattella germanica (L.) (Orthoptera: Blattellidae) in experimentally established shipboard infestations. Ann. Entomol. Soc. Am. 77: 740 Ð 752. Siljander, E., D. Penman, H. Harlan, and G. Gries. 2007. Evidence for male- and juvenile-speciÞc contact phero-

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mones of the common bed bug Cimex lectularius. Entomol. Exp. Appl. 125: 215Ð219. Siljander, E., R. Gries, G. Khaskin, and G. Gries. 2008. IdentiÞcation of the airborne aggregation pheromone of the common bed bug, Cimex lectularius. J. Chem. Ecol. 34: 708 Ð718. Stutt, A. D., and M. T. Siva-Jothy. 2001. Traumatic insemination and sexual conßict in the bed bug Cimex lectularius. Proc. Natl. Acad. Sci. U.S.A. 98: 5683Ð5687. Usinger, R. L. 1966. Monograph of Cimicidae (HemipteraHeteroptera). Entomological Society of America, College Park, MD. Wertheim, B., E. A. van Baalen, M. Dicke, and L.E.M. Vet. 2005. Pheromone-mediated aggregations in nonsocial arthropods: an evolutionary ecological perspective. Annu. Rev. Entomol. 50: 321Ð346. Received 6 February 2009; accepted 20 May 2009.