Pre-release evaluation of Semielacher petiolatus (Hymenoptera: Eulophidae) in quarantine for the control of citrus leafminer: Host discrimination,relative humidity tolerance,and alternative hosts

To evaluate the potential benefits and risks associated with releasing Semielacher petiolatus Girault (Hymenoptera: Eulophidae) in a classical biological control project directed against the citrus leafminer Phyllocnistis citrella Stainton (Lepidoptera: Gracillariidae) in Florida, we evaluated the ability of S. petiolatus females to discriminate between hosts previously parasitized by Ageniaspis citricola Logvinovskaya (Hymenoptera: Encyrtidae) and unparasitized hosts. In laboratory trials, S. petiolatus females did not discriminate between hosts previously parasitized and oviposited and fed on each host category equally. Hatch rate of S. petiolatus eggs on hosts previously parasitized by A. citricola was normal and development time was not different. However, mortality of immatures was significantly higher on previously parasitized hosts when compared to hosts that were not parasitized by A. citricola, and size of adult females reared on hosts previously parasitized was reduced. The relative survivorship of S. petiolatus adults compared with adults of P. citrella and another parasitoid, A. citricola, under three relative humidities (RHs) (55, 76, and 95% at 28 °C) indicated that S. petiolatus survives longer than A. citricola at all RHs tested, but did not survive as long as the citrus leafminer. Finally, the leafminer Liriomyza trifolii Burgess (Diptera: Agromyzidae) was evaluated as a possible host for S. petiolatus but no progeny were produced in choice and no-choice tests. The lack of discrimination raises the concern that S. petiolatus could disrupt the efficacy of A. citricola, which is already established in Florida, without providing substantial reduction of citrus leafminer populations during early spring.     

Contrasting genetic structuring between colonies of the World’s smallest penguin, <Emphasis Type="Italic">Eudyptula minor</Emphasis> (Aves: Spheniscidae)

The Little Penguin, Eudyptula minor, is a seabird that nests in colonies throughout New Zealand and southern Australia. Individuals from different colonies in southeast Australia differ significantly in morphology and ecology, suggesting that some genetic structuring may exist among colonies. In contrast, the marking of individuals with flipper bands has revealed some, albeit infrequent, movement between colonies. To determine the extent of genetic structuring, we tested the null hypothesis of substantial gene flow within southeast Australia by examining patterns of genetic variation across seven colonies separated by up to 1,500 km. Phylogeographic structuring was absent for mitochondrial control region sequences (2–3 individuals per colony). Microsatellite allele frequencies at five loci and mitochondrial haplotype frequencies (50 individuals per colony) were also homogenous among the majority of colonies sampled, although two colonies at the western periphery of the sampling range were distinct from those to the east. The genetic homogeneity among the majority of colonies can be explained by low but consistent contemporary gene flow among them, or a recent founder event in Bass Strait following the last marine transgression. The genetic break towards the western end of the sampling distribution appears best explained by differences in sea surface temperature and, consequentially breeding phenology, the latter hindering genetically effective migration.     

Genetic Diversity of Candidatus Liberibacter asiaticus Based on Two Hypervariable Effector Genes in Thailand

Huanglongbing (HLB), also known as citrus greening, is one of the most destructive diseases of citrus worldwide. HLB is associated with three species of ‘Candidatus Liberibacter’ with ‘Ca. L. asiaticus’ (Las) being the most widely distributed around the world, and the only species detected in Thailand. To understand the genetic diversity of Las bacteria in Thailand, we evaluated two closely-related effector genes, lasA _I and lasA _II, found within the Las prophages from 239 infected citrus and 55 infected psyllid samples collected from different provinces in Thailand. The results indicated that most of the Las-infected samples collected from Thailand contained at least one prophage sequence with 48.29% containing prophage 1 (FP1), 63.26% containing prophage 2 (FP2), and 19.38% containing both prophages. Interestingly, FP2 was found to be the predominant population in Las-infected citrus samples while Las-infected psyllids contained primarily FP1. The multiple banding patterns that resulted from amplification of lasA _I imply extensive variation exists within the full and partial repeat sequence while the single band from lasA _II indicates a low amount of variation within the repeat sequence. Phylogenetic analysis of Las-infected samples from 22 provinces in Thailand suggested that the bacterial pathogen may have been introduced to Thailand from China and the Philippines. This is the first report evaluating the genetic variation of a large population of Ca. L. asiaticus infected samples in Thailand using the two effector genes from Las prophage regions.     

Mitochondrial DNA and RAPD polymorphisms in the haploid mite Brevipalpus phoenicis (Acari: Tenuipalpidae)

Brevipalpus phoenicis (Geijskes) (Acari: Tenuipalpidae) is recognized as the vector of citrus leprosis virus that is a significant problem in several South American countries. Citrus leprosis has been reported from Florida in the past but no longer occurs on citrus in North America. The disease was recently reported in Central America, suggesting that B. phoenicis constitutes a potential threat to the citrus industries of North America and the Caribbean. Besides B. phoenicis, B. obovatus Donnadieu, and B. californicus (Banks) have been incriminated as vectors of citrus leprosis virus and each species has hundreds of host plants. In this study, Brevipalpus mite specimens were collected from different plants, especially citrus, in the States of Florida (USA) and São Paulo (Brazil), and reared on citrus fruit under standard laboratory conditions. Mites were taken from these colonies for DNA extraction and for morphological species identification. One hundred and two Random Amplified Polymorphic DNA (RAPD) markers were scored along with amplification and sequencing of a mitochondrial cytochrome oxidase subunit I gene fragment (374 bp). Variability among the colonies was detected with consistent congruence between both molecular data sets. The mites from the Florida and Brazilian colonies were morphologically identified as belonging to B. phoenicis, and comprise a monophyletic group. These colonies could be further diagnosed and subdivided geographically by mitochondrial DNA analysis.     

Differences in water relations among the citrus leafminer and two different populations of its parasitoid inhabiting the same apparent microhabitat

The water balance relationships of adult females and pupae of two populations (Australian and Taiwan) of the parasitic wasp Ageniaspis citricola, a biological control agent of the citrus leafminer Phyllocnistis citrella, were compared to determine whether habitat preferences might differ between them. No differences were detected between Australian and Taiwan strains of the wasp during the pupal stage, but adult females from the Australian strain had lower net water loss rates than the parasitoid adults from Taiwan. This implies a greater moisture requirement by adults of the Taiwan strain, which should be considered before they are released into a new environment in classical biological control programs.     

Establishment of Citrostichus phyllocnistoides (Hymenoptera: Eulophidae) as a biological control agent for the citrus leafminer Phyllocnistis citrella (Lepidoptera: Gracillariidae) in Spain

A program of introduction of exotic parasitoids for the biological control of the citrus leafminer Phyllocnistis citrella Stainton (Lepidoptera: Gracillariidae) was carried out in València (Spain) between 1996 and 1999. Eleven species of parasitoids were imported through a total of 37 shipments coming from nine countries. Six species were released in the field, the encyrtid Ageniaspis citricola (Logvinosvskaya), and the eulophids Quadrastichus sp., Semielacher petiolatus (Girault), Galeopsomyia fausta LaSalle, Cirrospilus ingenuus (Gahan), and Citrostichus phyllocnistoides (Narayanan). A. citricola was recovered in summer in many release points, reaching nearly 50% of parasitism and dispersing more than 300 m, but it was not able to overwinter. Quadrastichus sp. and S. petiolatus established temporarily in some sites, but produced little differences in parasitism or density of host population. In 1999, a substantial expansion of C. phyllocnistoides of more than 30 km in all directions was observed from one release point. In 2000 and 2001 this parasitoid expanded to all citrus grown in continental Spain and the Balear Islands, becoming the most abundant parasitoid in all the orchards, and displacing native and other introduced parasitoids. As a consequence, parasitism on second instars increased from less than 16% to 65% and on third instars from 35–38% to 59%. Overall, the mean percentage of parasitism increased from 20–25% to near 60%. Apparently, C. phyllocnistoides reduced by 34% the number of eggs and early instars of the host, and by 72% the number of adults. Damage to citrus foliage decreased by 56%.     

Natural variation in colony inbreeding does not influence susceptibility to a fungal pathogen in a termite

Reduced genetic diversity through inbreeding can negatively affect pathogen resistance. This relationship becomes more complicated in social species, such as social insects, since the chance of disease transmission increases with the frequency of interactions among individuals. However, social insects may benefit from social immunity, whereby individual physiological defenses may be bolstered by collective‐level immune responses, such as grooming or sharing of antimicrobial substance through trophallaxis. We set out to determine whether differences in genetic diversity between colonies of the subterranean termite, Reticulitermes flavipes, accounts for colony survival against pathogens. We sampled colonies throughout the United States (Texas, North Carolina, Maryland, and Massachusetts) and determined the level of inbreeding of each colony. To assess whether genetically diverse colonies were better able to survive exposure to diverse pathogens, we challenged groups of termite workers with two strains of a pathogenic fungus, one local strain present in the soil surrounding sampled colonies and another naïve strain, collected outside the range of this species. We found natural variation in the level of inbreeding between colonies, but this variation did not explain differences in susceptibility to either pathogen. Although the naïve strain was found to be more hazardous than the local strain, colony resistance was correlated between two strains, meaning that colonies had either relatively high or low susceptibility to both strains regardless of their inbreeding coefficient. Overall, our findings may reflect differential virulence between the strains, immune priming of the colonies via prior exposure to the local strain, or a coevolved resistance toward this strain. They also suggest that colony survival may rely more upon additional factors, such as different behavioral response thresholds or the influence of a specific genetic background, rather than the overall genetic diversity of the colony.     

Allozyme,chloroplast DNA and RAPD markers for determining genetic relationships between Abies alba and the relic population of Abies nebrodensis

Allozyme, chloroplast (cpDNA) and random amplified polymorphic DNA (RAPD) markers have been used to estimate genetic and taxonomic relationships among different populations of Abies alba and the relic population of A. nebrodensis. Twelve isozyme gene loci, as well as restriction fragment length polymorphism (RFLP) at cpDNA spacer regions between t-RNA genes were analysed. Moreover, a set of 60 random sequence 10-mer primers were tested. Over all isozyme loci, evident differences in allele frequencies among A. nebrodensis and A. alba populations were found, particularly at 2 loci, phosphoglucose isomerase (Pgi-a) and shikimate dehydrogenase (Skd-a). More than 10% of the total genetic diversity was due to differences among populations. High values of genetic distances among populations were also found. Out of the 60 primers tested, 12 resulted in a polymorphic banding pattern both within and among populations. A total of 84 RAPD fragments were produced by the 12 selected primers. A phenogram of relationships among populations was constructed based on RAPD band sharing: the differentiation of the A. nebrodensis population was evident. The analysis of molecular variance (AMOVA) was used to apportion the variation among individuals within populations and among populations. There was considerable variation within each population: even so, genetic divergence was found among populations. This pattern of genetic variation was very different from that reported for inbred species. Identical cpDNA amplification and restriction patterns were observed among all the individuals sampled from the populations. Taken together, the results of allozyme and RAPDs show a clear differentiation among A. nebrodensis and A. alba populations and provide support for their classification into two different taxonomic groups.     

Contribution of predation and parasitism to mortality of citrus leafminer Phyllocnistis citrella Stainton (Lepidoptera: Gracillariidae) populations in Florida

The citrus leafminer (CLM), Phyllocnistis citrella Stainton, is native to southern Asia and regarded as an important pest in nurseries and young or top-worked citrus trees in Florida. Damage to the leaf cuticle increases susceptibility to citrus canker disease and further aids spread by increasing inoculum loads. Biological control agents are known to play an important role in regulating pest populations. We evaluated individual contributions of predation and parasitisation to cohorts of P. citrella by exclusion and by direct observation of leaf mines in the field. Predation, particularly by ants, was the largest single cause of P. citrella mortality, accounting for more than 30% of all deaths by natural enemies, and 60% of all deaths by predators. First and second instars of P. citrella were most subject to ant predation. Ageniaspis citricola was the most important parasitoid of P. citrella and caused 8.2–28.6% mortality compared to 9.6–14.7% from indigenous parasitoids. The total biotic mortality of P. citrella observed in exclusion experiments was 52–85%. These results were in basic agreement with 89% mortality, predominantly by predation, obtained by reconstructing a cohort from observations of recently mined leaves. A partial life table based on these data predicted an innate rate of increase (R_o) of 2.8 and thus an almost 3-fold increase per generation. These results indicate that, while biotic mortality takes considerable toll of P. citrella populations, the predominance of predation suggests that the parasitoid complex on this exotic pest in Florida is depauperate and would likely be improved by additional introductions.     

Genotyping of Ganoderma species by improved random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) analysis

Species of Ganoderma are used in traditional medicines. An improved random amplified polymorphic DNA (RAPD) analysis, where the RAMP time is prolonged, has been used to characterize the genetic variation in some well known species of Ganoderma. The DNA materials were collected from ten Ganoderma strains, amplified with randomly selected 24 RAPD primers and evaluated by agarose gel electrophoresis. A cluster dendrogram was constructed for genetic analysis on the basis of amplification results. The improved RAPD amplified DNA with consistent and clear banding patterns. A total of 316 bands were found with 93% polymorphism. There was a significant genetic distance between the different strains of Ganoderma, with an index of similarity coefficient in the range of 0.52–0.74. The inter-simple sequence repeat (ISSR) analysis of the Ganoderma DNA samples showed similar trend results to the RAPD analysis with 0.49–0.81 similarity coefficients. This study reports the high level of genetic differences between different species or strains of a single species of Ganoderma and confirms the significance of the improved RAPD method in genetic characterization of organisms. Therefore, the improved RAPD combined with ISSR techniques might be used for the genetic characterization of organisms.     

High levels of genetic variability in the moss Ceratodon purpureus from continental Antarctica, subantarctic Heard and Macquarie Islands, and Australasia

The random amplified polymorphic DNA (RAPD) technique, and DNA sequencing of the conserved nuclear ribosomal DNA internal transcribed spacer region (ITS1-5.8S-ITS2), have been used to assess levels of genetic diversity in the moss Ceratodon purpureus from several locations in Australasia, subantarctic Heard and Macquarie Islands, and continental Antarctica. Populations from Heard and Macquarie Islands and from Antarctica maintain high levels of genetic variation. Both within- and among-colony variation were observed at these locations. DNA sequence analysis showed that samples from the Ross Sea region of Antarctica were most closely related to colonies from Casey and Macquarie Island, and that one colony from Heard Island was most closely related to one from Europe. DNA sequence data separated two Australian populations from the Antarctic and subantarctic group on a dendrogram. Detailed RAPD analysis of a single colony from continental Antarctica demonstrated that mutation probably causes the high variability observed in this moss. DNA sequencing and RAPD analysis are complementary techniques for genetic investigation of Antarctic moss populations.Jenny Ninham was an integral part of this research team for several years. Unfortunately she did not live to see these results published.     

A localized linkage map of the citrus tristeza virus resistance gene region

A localized genetic linkage map was developed of the region surrounding the citrus tristeza virus (CTV) resistance gene (designated Ctv) from Poncirus trifoliate L., a sexually compatible Citrus relative. Bulked segregant analysis (BSA) was used to identify potential resistance-associated RAPD fragment markers in four intergeneric backcross families that were segregating for CTV resistance. Eight RAPD fragments were found that were consistently linked to Ctv in the four families. Map distances and locus order were determined with MAPMAKER 3.0, using the results obtained from 59 individuals in the largest family. Also, a consensus map was constructed with JOINMAP 1.3, using pooled results from the four backcross families. Marker orders were identical, except for 1 marker, on these independently developed maps. Family-specific resistance-associated markers were also identified, as were numerous susceptibility-associated markers. The identification of markers tightly linked to Ctv will enable citrus breeders to identify plants likely to be CTV-resistant by indirect, marker-assisted selection, rather than by labor-intensive direct challenge with the pathogen. These markers also provide a basis for future efforts to isolate Ctv for subsequent genetic manipulation.Florida Agricultural Experimental Station Journal Series No. R-04491     

Sources of mortality in colonies of brown citrus aphid, Toxoptera citricida

Sixteen cohorts of the brown citrus aphid, Toxoptera citricida, were followed from colony initiation to maturation/extinction in citrus groves at two sites in Puerto Rico and nine sites in Florida, USA. Infested citrus terminals were sampled repeatedly in a non-destructive manner and data recorded on the recruitment of natural enemies and the fate of aphid colonies. Coccinellidae were the most efficient predators of T. citricida, primarily Cycloneda sanguinea, and Coelophora inaequalis (Puerto Rico), C. sanguinea, Harmonia axyridis (Florida). Coccinellids were abundant in citrus year-round in Puerto Rico, but in Florida citrus their abundance peaked in spring and declined thereafter. Syrphid flies were other important predators, especially Pseudodorus clavatus (Florida and Puerto Rico) and Ocyptamus fuscipennis (Puerto Rico). Syrphids increased in abundance from spring to fall in both regions. Other predators included Cereaochrysa lineaticornis, Chrysoperla rufilabris (Chrysopidae) and Micromus posticus (Hemerobiidae). The parasitoid Lysiphlebus testaceipes was ubiquitous in T. citricida cohorts, but its impact on colony survival was usually low. The fungal pathogen Verticillium lecanii was a significant source of colony mortality at one site in Puerto Rico, but was not observed on T. citricida in Florida.     

Agonistic behavior among colonies of the Formosan subterranean termite,Coptotermes formosanus Shiraki (Isoptera: Rhinotermitidae), from Florida and Hawaii: Lack of correlation with cuticular hydrocarbon composition

Cuticular hydrocarbon patterns of the Formosan subterranean termite, Coptotermes formosanus Shiraki, were similar among colonies from the same geographical location. Hydrocarbon patterns of Florida colonies were easily distinguished from those of Hawaii colonies by using canonical discriminant analysis. Groups of termites from the same colony did not fight one another when placed in an arena. Intercolonial aggression was not recorded among C. formosanuspopulations from Florida but three colonies from Hawaii fought with the other Hawaiian and three Florida colonies. Of the 12 colonies (six each from Florida and Hawaii) tested, 3 Florida colonies did not direct or receive aggression from any other colony. Cuticular hydrocarbon patterns were not correlated with agonistic behavior.     

Genetic divergence between colonies of Flesh-footed Shearwater <Emphasis Type="Italic">Ardenna carneipes</Emphasis> exhibiting different foraging strategies

Increasing evidence suggests foraging segregation as a key mechanism promoting genetic divergence within seabird species. However, testing for a relationship between population genetic structure and foraging movements among seabird colonies can be challenging. Telemetry studies suggest that Flesh-footed Shearwater Ardenna carneipes that breed at Lord Howe Island or New Zealand, versus southwestern Australia or Saint-Paul Island in the Indian Ocean, migrate to different regions (North Pacific Ocean and northern Indian Ocean, respectively) during the non-breeding season, which may inhibit gene flow among colonies. In this study, we sequenced a 858-base pair mitochondrial region and seven nuclear DNA fragments (352–654 bp) for 148 individuals to test genetic differentiation among colonies of Flesh-footed Shearwaters. Strong genetic divergence was detected between Pacific colonies relative to those further West. Molecular analysis of fisheries’ bycatch individuals sampled in the Sea of Japan indicated that individuals from both western and eastern colonies were migrating through this area, and hence the apparent segregation of the non-breeding distribution based on telemetry is invalid and cannot contribute to the population genetic structure among colonies. The genetic divergence among colonies is better explained by philopatry and evidence of differences in foraging strategies during the breeding season, as supported by the observed genetic divergence between Lord Howe Island and New Zealand colonies. We suggest molecular analysis of fisheries’ bycatch individuals as a rigorous method to identify foraging segregation, and we recommend the eastern and western A. carneipes colonies be regarded as different Management Units.     

Inferring the invasion history of coral berry Ardisia crenata from China to the USA using molecular markers

Genetic comparisons between native and invasive populations of a species can provide insights into its invasion history information, which is useful for guiding management and control strategies. The coral berry Ardisia crenata was introduced to Florida last century as a cultivated ornament plant, and has since spread widely throughout the southern regions of the USA. Previously, the genetic variation among 20 natural populations of A. crenata across its distribution center in southern China was quantified using seven microsatellite markers. Here we expand on that work by additionally sampling individuals from four other native populations in Taiwan and Japan, and from five invasive populations in the USA. We also examined the results from one chloroplast intergenic spacer region (trnF-trnL) in all 29 populations. Our aim is to identify the invasion source and subsequent history of the species?? spread throughout the southern USA. We observed lower genetic diversity in the invasive populations based on both microsatellite and chloroplast markers. Our data show that the invasive populations can be clustered with native populations in southeastern China, inferring this region as the geographic origin of A. crenata cultivars invading the USA. We further classified invasive individuals into invasive I and invasive II clusters. Nantou in Taiwan and Xihu in mainland China are the most closely related populations to those, which identify the former as potential sources for host-specific control agents. Our results, combined with the known introduction records, suggest that A. crenata was first multiply introduced into Florida and then secondarily colonized Louisiana and Texas from Florida.     

Developmental Instability in Incipient Colonies of Social Insects

Social insect colonies can provide homeostatic conditions that buffer the incidence of environmental fluctuations on individuals, which have contributed to their ecological success. Coptotermes (Isoptera: Rhinotermitidae) is a highly invasive termite genus and several species have important economic impact in many areas of the world. Mature Coptotermes colonies with millions of individuals can provide optimal environmental condition and nurturing capacity for the developing brood. However, it was previously suggested that contrary to mature colonies, incipient colonies may be exposed to critical stress, which may explain for the low success rate of establishment within the first year of the life of a termite colony. We here investigated the stress imposed on individuals of incipient colonies by comparing the developmental instability of individuals between incipient and mature colonies of two Coptotermes species, C. formosanus Shiraki and C. gestroi (Wasmann). We assessed the developmental instability by measuring the asymmetry of morphological traits from the head capsule of the soldier caste. Soldiers from incipient colonies of both species displayed strong asymmetrical traits in comparison to soldiers from mature colonies. We suggest that homeostatic conditions for optimal development are reached as the colony matures, and confirmed that the incipient colony remains a critical bottleneck where individuals are exposed to high developmental stress.     

Randomly Amplified Polymorphic DNA Differs with Burrowing Nematode Collection Site,but not with Host Range

The genetic variability of 12 burrowing nematode (Radopholus sp.) isolates from Central America, the Caribbean, and Florida, and one isolate from Ivory Coast were compared with RAPD analysis. A high degree of genetic similarity (>0.82) was determined for isolates from the Western Hemisphere. Genome similarity was greatest among isolates collected within a country. Among isolates collected in Central America and the Caribbean, burrowing nematodes from Belize and Guatemala were genetically more distant. However, the genome of the isolate from Ivory Coast was most dissimilar (>0.30). These results suggest that African and American burrowing-nematode isolates may have had different origins or that they have been geographically isolated for a sufficient amount of time to have accumulated genetic changes detectable by RAPD analysis. No relationship was found between the genomic similarity and extent of reproduction or damage to banana or citrus roots. Morphometric analysis involving eight of the isolates indicated that they were morphologically identical and values for morphometric parameters were well within the range previously published for banana and citrusparasitic burrowing nematodes.     

RAPD fingerprinting Xylella fastidiosa Pierce's disease strains isolated from a vineyard in North Florida

Pierce's disease (PD) strains of Xylella fastidiosa were identified by random amplified polymorphic DNA (RAPD) fingerprinting. Two random primers including OPA-03 (agtcagccac) and OPA-11 (caatcgccgt) were found to be efficient for differentiating PD strains isolated from a vineyard in North Florida in 1996 (129 strains) and 1997 (29 strains) from non-PD strains of X. fastidiosa (citrus variegated chlorosis, mulberry leaf scorch, periwinkle wilt, plum leaf scald, and phony peach) and strains from Xanthomonas campestris pv. vesicatoria and Escherichia coli. This study shows that RAPD fingerprinting is a useful tool to supplement the conventional symptoms-colony morphology-slow growth identification procedure routinely used to identify the PD pathogen.     

Genotype-Based Recognition Among Individuals of the Social Insect Pristomyrmex Punctatus (Japanese Queenless Ant) from Geographically Divergent Populations

The queenless ant, Pristomyrmex punctatus (F. Smith) reproduces parthenogenetically. The workers lay unfertilized eggs, which develop into female workers. This mode of reproduction generates hereditary clones. A previous research shows that when genetically monomorphic colonies were split, the workers tended to reassemble after being split into two groups, but when genetically polymorphic colonies split, they remained as two separate colonies. However, it remains unclear whether the workers can recognize individual genotype. Here, it was investigated whether individuals from geographically divergent, genetically monomorphic colonies would assemble with individuals of the same genotype. Two artificially fused colonies were prepared, A and B, comprising 200 individuals and 100 individuals, respectively. Each half of the artificially fused colony was composed of workers from two different genetically monomorphic source colonies. The workers assembled as a single colony when the genotype of the source colonies was identical. However, when the genotypes of source colonies were different, the workers did not assemble into one colony, but split into two groups according to genotype. These results suggest that P. punctatus can potentially recognize individual genotype, and select colony members based on an individual’s genotype.