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The purpose of this experiment was to investigate whether honey bees (Apis mellifera) are able to use social discriminative stimuli in a spatial aversive conditioning paradigm. We tested bees’ ability to avoid shock in a shuttle box apparatus across multiple groups when either shock, or the absence of shock, was associated with a live hive mate, a dead hive mate, a live Polistes exclamans wasp or a dead wasp. Additionally, we used several control groups common to bee shuttle box research where shock was only associated with spatial cues, or where shock was associated with a blue or yellow color. While bees were able to learn the aversive task in a simple spatial discrimination, the presence of any other stimuli (color, another bee, or a wasp) reduced initial performance. While the color biases we discovered are in line with other experiments, the finding that the presence of another animal reduces performance is novel. Generally, it appears that the use of bees or wasps as stimuli initially causes an increase in overall activity that interferes with early performance in the spatial task. During the course of the experiment, the bees habituate to the insect stimuli (bee or wasp), and begin learning the aversive task. Additionally, we found that experimental subject bees did not discriminate between bees or wasps used as stimulus animals, nor did they discriminate between live or dead stimulus animals. This may occur, in part, due to the specialized nature of the worker honey bee. Results are discussed with implications for continual research on honey bees as models of aversive learning, as well as research on insect social learning in general.

A combination of arena,Y-tube olfactometer, and flight tunnel assays were used to determine responses of male and reproductive female Polistes exclamans Vierick (Hymenoptera: Vespidae) to odors and extracts of conspecific males and females, as potential pheromone-mediated sexual behaviors. Males rubbed the sternites of the gaster on filter paper in response to extracts of the female mesosoma and gaster, indicating possible scent-marking. In a Y-tube olfactometer, females oriented to extracts of the male head, gaster, legs, seventh sternite, and mandibular glands. In the olfactometer, males oriented to extracts of the female whole body, but were repelled by extracts of the female head. Both females and males oriented to solvent washes of glass jars that held wasps of the opposite sex, as well as volatile collections from the opposite sex. In a flight tunnel, males and females both exhibited chemoanemotaxis that consisted of upwind flight and close range casting in response to odor piped into the tunnel from live wasps of the opposite sex. Males mounted a female wasp model treated with an extract of the female mesosoma. Together, these experiments suggest pheromones stimulate male scent-marking, attraction of females to males and males to females, and mounting, in P. exclamans. Results indicate the mesosoma as a source of female pheromones, and the legs, gastral sternal glands, and mandibular glands, as male pheromone sources.

Some animals minimize the high costs of aggressive conflict by using conventional signals of agonistic ability to assess rivals prior to interacting. Conventional signals are more controversial than other signals of agonistic ability because they lack an inherent physical or physiological link with their bearer’s agonistic ability. Here, we test whether the variable brown facial stripes in Polistes exclamans paper wasps function as a conventional signal. Polistes exclamans were given the option of challenging or avoiding a rival with an experimentally altered facial pattern. Our results show that rival assessment is based on the facial patterns of rivals, as well as an individual’s own size, facial patterns, and nesting strategy. Individuals with larger body size and larger brown facial stripes were more likely to challenge rivals than individuals with smaller body size and smaller brown facial stripes. In addition, large individuals were more likely to challenge rivals with large brown facial stripes than small individuals, while an individual’s own body size did not influence whether or not they challenged rivals with small brown stripes. Individuals who previously nested in multiple queen groups approached rivals more rapidly than individuals who previously nested alone, suggesting that social experience also plays a role in rival assessment. Finally, rivals with small facial stripes were challenged more rapidly than those with large facial stripes. These results demonstrate that P. exclamans facial patterns function as a signal used to minimize the cost of conflict. However, individuals do not make simple decisions based on their rival’s signal alone, as an individual’s own social experience and agonistic abilities also influence rival assessment decisions.

and Summary In the social wasp, Polistes exclamans, relatedness of workers to the brood they rear is quite low. In a 4 year study relatedness of workers to brood was determined on 338 nests by marking all females including the queen and censusing the nests throughout the season. Workers were rearing siblings on 26% of all nests and nieces and nephews on 50% of all nests. Siblings and nieces and nephews were reared together on 10% of nests, and nieces and nephews that were the progeny of two different sisters were reared on 14% of nests. Multiple mating by the egg-layer would reduce relatedness of workers to brood still further. Workers rear siblings on so few nests that high levels of relatedness between workers and brood may not be a critical factor for the maintenance of group living. Differences in the probability that a female can successfully nest alone between early spring and mid summer may be the reason workers usually rear distant relatives on the nest they emerged from rather than beginning their own nest where they could rear progeny. Zusammenfassung Vier Jahre lang wurden in 338 Nestern der sozialen Wespe Polistes exclamans alle Arbeiterinnen und die Koniginnen markiert und uber den Sommer die Individuen gezahlt. Der Verwandtschaftsgrad zwischen den Arbeiterinnen und ihren Pfleglingen ist gering: In 26% aller Nester ziehen die Arbeiterinnen Geschwister auf, in 50% aller Nester Nichten und Neffen, in 10% aller Nester beides, und in 14% aller Nester Nichten und Neffen, die von zwei Schwestern stammten. Waren die Koniginnen mehrfach von verschiedenen Mannchen begattet, dann waren die Verwandtschaftsgrade noch geringer anzusetzen. Besonders enge Verwandtschaft zwischen Arbeiterinnen und Brut kann kaum die Ursache fur die Staatenbildung sein. Eher ist entscheidend, das ein Weibchen in der Zeit zwischen Fruhling und Sommermitte kaum allein ein Nest betreuen kann.

The objective of this paper is to identify common species of paper wasps (Polistes spp.) attacking fall armyworm (Noctuidae: Spodoptera frugiperda) larvae and to investigate foraging activity in turfgrass. Three species — Polistes exclamans, P. dorsalis, and P. metricus — were collected while they foraged in bermudagrass turf. Polistes wasps forage in turf for about 10 h a day beginning a few hours after sunrise (0830 h), waning near midday, and increasing again in the late afternoon. Wasps were more abundant in plots infested with fall armyworms although these differences were statistically significant in one of three trials. The benefits of Polistes wasps may be conserved by scheduling insecticide applications in the early evening (0700 h or later) or by using reduced-risk insecticides.

Polistes exclamans Vier. introduced to Europe from USA together with its parasite hitherto unknown. Hidden in the inside of a special motorcar-bumper a full grown nest of the American Polistes exclamans Vier. had been transported in Sept. 1969 from Pittsburg (Pennsylvania) to Gros Bieberau/Odenwald (Western Germany). There could be caught 19 emerged exclamans-specimens and 25 specimens of a Chalcidide-parasite unknown even in U.S.A. This species has been described in 1971 by Dr. Burks, Washington, as Elasmus polistis nov. spec.

Summary Although the different vespid species of the genus Polistes are found all over the world, studies about allergy sensitization to these species have been made mainly in North America. We studied the specific IgE in a large group of sera from patients sensitized to Polistes dominulus (PD) and determined the crossreactivity with another important European Polistes (P. gallicus [PG]) and with five American relevant species of this genus: P. exclamans (PE), P. fuscalus (PF), P. metricus (PM), P. annularis (PAN) and P. apachus (PAP). Studies indicated that 96.7% of sera with IgE positive to PD were positive to PG. No case positive to just PG was found. The comparison of these two European species with the American relatives indicated that most cases were positive to all the insects although there were significant differences in the radioallergosorbent test (RAST) value. These results were confirmed by RAST inhibition studies, and indicate that although the European and American species are closely related, species specific allergenic differences exist. These data suggest that in vivo studies should be carried out in order to determine the sensitivity and specificity of the European venoms compared with the American species.

Measuring the total carbohydrate concentration in the haemolymph (anthrone-positive material), heterologous injections of honeybee ( Apis mellifera ) and paper wasp ( Polistes exclamans ) corpus cardiacum (CC) extracts resulted in hyperglycemia in Periplaneta , but homologous injection caused no increase in total blood carbohydrates. However, when individual sugars like glucose, fructose and trehalose were quantified using a high pressure liquid chromatography method, then homologous injections of CC extracts showed a distinct hypertrehalosemic effect in both hymenopteran species. There was no effect of the extracts on glucose or fructose titers. The decrease of glucose, fructose and trehalose titers during the time between taking blood samples in honeybees (because of starvation) had to be carefully monitored in order to detect the hypertrehalosemic effect of the extracts. Trehalose titers in unfed wasps on the other hand did not diminish during the time between blood sampling. In spite of large lipid stores in the fat body and a high blood lipid titer (15 mg/ml) in Polistes , there was a complete lack of hyperlipemic response to homologous injections of CC extracts. There was also no evidence of proline utilization as an energy source in either species.

Six variable protein loci and one variable ribosomal DNA restriction site were used for an analysis of population structure in five species of Polistes from Texas. A sample-reuse algorithm was developed that estimated FST, FIS, and 0 (the coefficient of kinship) from probabilities of identity. Of the four species analyzed in detail only one, Polistes exclamans, had statistically significant values of FST. These values may reflect natural constraints on successful nesting for migrants of this species. Three of the four species had significant values of F1s and three of the four species had significant values of 0. In many cases 0 also differed from the expected value under haplodiploidy and random mating. Values of 0 did not differ from expectations under haplodiploidy and local inbreeding. These results emphasize that theories of social behavior and evolution based on coefficients of kinship should include some explicit consideration of population structure.

A wind-tunnel bioassay was developed to test for the presence of sex attractants in the paper wasp,Polistes exclamans Viereck (Hymenoptera: Vespidae). Males exhibited significant upwind flight and attraction (chemoanemotaxis) in response to airflow passed over unmated females, and to hexane extracts of whole bodies and thoraces of unmated females. Unmated females were attracted to hexane extracts of males and to hexane extracts of each body tagma of males, suggesting distribution of the pheromone over the cuticle by grooming. The ectal mandibular and seventh sternal glands are the likely sources of the male-produced sex attractant(s) since extracts of each elicited female attraction in the flight tunnel. These glands are associated with gastral and mandibular rubbing of perch sites by territorial males ofPolistes species.

I examined nestmate and nest recognition among worker paper wasps Polistes exclamans on neighboring colonies in a dense nesting aggregation. In the laboratory, there was a significant positive correlation between the difference in preference toward nestmates and non-nestmates and internest distance (i.e., distance in the field between the nestmates' and non-nestmates' nests). This is the first demonstration of a distance effect in nestmate recognition ability among Polistes. The field significance of the laboratory bioassay of nestmate preference was confirmed by releasing in the field workers whose nests had been removed (i.e., orphaned workers) and finding that they joined colonies near the site of their original nest. Orphaned workers also tended to join smaller colonies, perhaps because these workers had a greater opportunity for reproduction on smaller colonies. This pattern may also reflect a greater tendency on the part of inhabitants of smaller nests to admit joiners, because the addition of a worker may have had a greater impact on colony success on a smaller colony than on a larger nest. Finally, workers prefer fragments of nests located closer in the field to the workers' colony than fragments of more distant nests. This ability may aid workers in choosing which nests to join and which to rob.

In social insect colonies the sterile individuals that assume the worker role realize their reproductive potential through working and rearing the queen's brood. We have examined the role of the presence of brood in the nest as a determinant of caste in the social wasp Polistes exclamans Viereck. We predicted that wasps in colonies where there was no brood would behave like future queens, while wasps in colonies in which brood care was an option would behave like workers. Brood presence was manipulated by removing eggs and larvae or the whole nest from experimental colonies and leaving brood in control colonies. Ability to tolerate cold temperatures was used as an indicator of caste when comparing females emerging in experimental colonies to females emerging in control colonies. Females emerging in experimental colonies survived longer at cold temperatures than those emerging in control colonies. This indicates that females developed characteristics more typical of future queens in response to deprivation of brood. P. exclamans is characterized by small colonies whose nests are destroyed frequently. Therefore caste plasticity in adult females of P. exclamans is advantageous. Key-words: Caste determination, nesting cycle, Polistes exclamans, production schedules

1. Polistes exclamans populations in central Texas have a unique colony cycle. Of all P. exclamans nests 20%–38% produce males with the first brood of workers. Nests producing early males have significantly more females, cells, pupae, and emergences at the time of early male production than nests not producing early males. 2. Early male production may have originated as an adaptation caused by the large number of queens that die before autumn reproductive males and females are produced. It allows egg laying by workers with whom early males mate. 3. Queens or workers sometimes leave their original nest and start new nests nearby that must be joined by workers from the original nest if they are to succeed. Such ‘satellite nests’ were initiated from May to July by 16%–39% of all nests, depending on the population. Although nests initiating satellites have significantly more females and pupae than nests not initiating satellites, satellite nest production by a given nest is not dependent on prior early male production, and is independent of the number of foundresses. 4. More P. exclamans nests 1976–1979 were lost to birds than to any other cause. In the period 1976–1979, 66.7% of the 12 nests with satellites succeeded in producing reproductives after being knocked down while only 5.7% of the 87 nests without satellites that were knocked down were subsequently successful. Concealed nests are less vulnerable to bird predation. 5. The brood parasitoid, C. iphitalis, a pyralid moth, avoids previously infested nests, therefore satellite nesting does not reduce the impact of this parasitoid. However, the brood parasitoid E. polistis, a chalcid wasp, lays eggs in the nest it emerges from, hence a satellite nest provides an escape when this parasitoid is already in the main nest.

Workers of the primitively social wasp,Polistes exclamans, had very short average lifespans, averaging 14 days (S.D.=11, N=497) in 1977, 17 days (S.D.=14, N=503) in 1978 and 14 days (S.D.=10, N=222) in 1979; the oldest worker observed on a natural colony was 102 days old. Colony of origin explained more of the variance in worker longevity than did any other variable. Worker longevity decreased over the season, accompanied by an increase in larval development times and an increase in colony failure rates. In 1978 workers emerging in the presence of greater numbers of other workers lived longer which indicates that they may have performed risky tasks less often when many other females were present. Compared to 1977 and 1979, 1978 was characterized by greater numbers of workers per colony, shorter larva development times, longer average lifespans for workers, much higher probabilities of nest survival and greater average production of reproductives per colony.

The venoms ofPolistes exclamans andP. fuscatus elicit alarm behavior and attract attacking wasps. The response is not species specific, for both hetero- and conspecific venoms elicit similar responses in both species. A test in a wind tunnel provided no support for the hypothesis that alarmed wasps release an alarm pheromone on the nest.

RAST disks were prepared with purified proteins from the venoms of Vespula squamosa, Dolichovespula maculata , and Polistes exlamans and tested against a panel of sera obtained from patients allergic to each of the venoms. By the use of several criteria, IgE binding was strongest to antigen 5 of V squamosa and phospholipase of P. exclamans . Hyaluronidase from Polistes had little IgE binding activity. All five proteins tested from D. maculata bound IgE antibodies strongly. Linear correlation analysis suggested that all of the proteins were distinct allergens in each of the venoms. A variety of patterns of response were observed for individual sera with each of the venoms.

The paper wasp Polistes exclamans loses part of its brood to the parasitoids Chalcoela iphitalis (Lepidoptera:Pyralidae) and Elasmus polistis (Hymenoptera:Chalcidoidea:Eulophidae) and loses entire nests to bird predation, Crematogaster laeviuscula (Hymenoptera:Formicidae) predation, and orphanage (loss of all adult wasps). More nests fail because of bird predation than from all other causes combined. Larger nests are neither better protected nor more vulnerable to nest predators, though larger nests are less likely to fail because of orphanage than are smaller nests. Larger nests are more heavily attacked by Elasmus polistis, which reinfests its natal nest. Polistes exclamans may subdivide larger groups of females by forming satellite nests because of the lack of an overall advantage to large nests. Satellite nests increase the probability that the brood in either the original nest or satellite nest will avoid bird predation. See full-text article at JSTOR

Elevated fructose, glucose and trehalose levels were found inPolistes exclamans andP. annularis after exposure to cold. Glycerol was found only after extensive exposure to cold or in mid-winter. InP. exclamans mean supercooling points (SCP) decreased to a low of −12.9° C in February. InP. annularis SCP were never below −6.8° C in the field, but laboratory acclimation at +5° C resulted in SCP of −10.7° C. Freezing was lethal to adultPolistes under all circumstances. Workers and future queens were separated on the basis of the appearance of their fat bodies. ForP. exclamans, only 17 % of the future queens died during 15 days at +5° C (N=29) while 76 % of the workers died during identical cold exposure (N=45). The surviving future queens had elevated cryoprotectant levels consisting mainly of fructose in their hemolymph, while the few workers that survived did not. These experiments indicate that there are physiological differences between workers and future queens inPolistes collected in autumn and that fat body appearance determined using non-invasive techniques is a reliable indicator of caste. Males were similar to future queens in their response to cold indicating that autumn mating activity may extend into winter.

1. Autumnal nest populations ofPolistes exclamans include externally indistinguishable workers and future queens. Qualitative as well as quantitative caste differences were found in the parietal fat body. 2. An interesting correlation exists between the type of fat body and amount of eye pigmentation in females of this species. 3. The frequency of transfer of marked males between nests was found to be very low. 4. Polistes exclamans is unusual among North American members of its genus in that it appears to be almost completely haplometrotic in parts of its range. This may be evolutionarily connected with the species' comparatively greater vagility and tendency to disperse. Nest dispersal may reduce the rate of parasitism.

Three year of laboratory and field testing of materials being screened for control of tobacco hornworms ( Protoparce sexta (Johan.)) with reduced insecticide residue and reported. Comparative contact toxicity tests indicated the following descending order of effectiveness: Guthion® ( O,O -dimethyl S -(4-oxo- 3H -1,2,3-benzotriazine-3-methyl) phosphorodithioate) =DDVP>endrin=Dipterex® ( O,O -dimethyl 2,2,2-trichloro-l-hydroxyethyl-phosphonate)=Shell SD 4402(1,3,4,5,6,7,8-octachloro-3a, 4,7,7a-Tretrahydro-4-7-methanophthalan)> Thiodan®(6,7,8,9,10,10-hexachloro-1,5,5a,6,9,9a-hexahydro-6,9-methano-2,4,3, benzodioxathiepin-3-oxide) = Sevin® (1-naphthyl- N -methyl carbamate) >Chlorthion® ( O , (3-chloro-4-nitrophenyl) O,O -dimethyl phosphorothioate, Delnave® (2.3,- p -dioxancdithiol S,S -bis ( O,O -did ethy; phosphorodithioate), ethion, and Trithiion® ( S-( p -chlorophenylthio)methyl O,O -diethy; phosphorodithioate. In field tests Thiodam, Sevin, SD 4402, and Dipterex gave larval control generally comparable to endrin at the concentrations tested. Guthion gave poorer results than at comparable concenlrations But was effective at the higher dosages. All materials Were about equal in redueing hornworm dosages estimations Were made 9 days after treatment, but Guthion and Dipterex permitted more damage than the other materials 22 days after treatment. The residues from the compounds dissipated to the extent of R5 % or more during flue-euring. None of the materials affected taste of cured tobaceo except Sevin, which caused a slight off-flavor. Bacillus thuringiensis Berliner was found to he equal to endrin in preventing hornworm damage although not so effective initially in reducing larval populations. The Barilllis organism had no detrimental effect on colonies of Polistes exclamans Vier. Two commereial preparations of Bacillus Were found to be equal to insecticides in preventing hornworm damage.

It is generally difficult to document temporal changes in the geographic distribution o insect species because o the sporadic nature o collections made primarily or taxonomic study and specimen identification. Therefore .some new distribution data providing an unusual amount ot evidence or range extension in the social wasp Polistes exclamans Viereck seem o interest. Several sta.tes have been added to the known range o P. exclamans in the last twenty years; and P. exclamans is now abundant in east-central Missouri, where it was almost certainly absent two decades ago. Solitary nest ounding by P. exclamans queens may enhance the ability of this species to colonize new areas. Porter (1963) first suggested that P. exclamans may be extending its range. He reported new records rom Maryland and New Jersey, noting that these states are considerably north of the northernmost previous records or P. exclamans in the Atlantic Coast states. A collecting trip to St. Louis and Jefferson Counties, Missouri, in July, 967, provided the first evidence of range extension by Polistes exclamans in the midwestern United States. The purpose ot: the trip was to collect specimens o Polistes species observed by the Missouri naturalist Phil Rau prior to his death in 1948. Rau repeatedly mentioned (e.g., in Ra.u I929, 942a, 942b) that only our species of Polistes existed in Missouri: P. pallipes (-- metricus), P. rubiginosus, P. annularis, and P. variatus (-- f,uscatus) (Rau’s determinations by J. Bequaert). However, I ound P. exclamans nests common in barns and abandoned buildings in southern St. Louis County, and collected specimens o P. exclamans in six ot the eight localities where Polistes wasps were t:ound (other species collected were P. m’etricus, P. fuscatus, and P. rubiginosus). Three of the I967 P. exclamans localities were requented by Rau during his studies ot Polistes: "Meramac Highlands", an area near the Meramac River a.bout eight miles southwest of St. Louis (first mentioned in Rau and Rau, I918) Kirkwood, Rau’s home town or 24 years (first mentioned as a study locality in Rau, 94) and a arm near

Potent allergenic extracts of three different species of stinging insects were prepared by a simple procedure. The types of Hymenoptera studied were the bee ( Apis mellifera ), the wasp ( Polistes exclamans ), and the yellow jacket ( Vespula pennsylvanicus ), and extracts of each were prepared from the whole body, from the sacless body, and from the venom sac. In order to facilitate the routine determination of protein concentration, light absorption methods were developed and an extinction coefficient for each type of extract was determined for both ultraviolet absorption, at 280 mμ, and for the biuret reaction at 540 mμ. The allergenicity of these preparations was determined by direct skin tests. Positive intradermal skin tests were elicited in sensitive patients with as little as a 1:10,000 dilution of the preparation of insect antigen. Most of the patients reacted to more than one type of insect. Commercial preparations gave similar results. Zone electrophoresis, by filter paper strip and by starch block, revealed two or three separate components. It was noted that the best pH for optimal resolution, over the range of 8.6 to 4.5, was 6.5 for bee and wasp and 4.5 for yellow jacket. This method seemed to have limited value for the possibilities of fractionation of these extracts. The extensive heterogeneity of these extracts was better shown by analytical ultracentrifugation. All three types of whole body extracts had characteristic patterns of five homologous sedimenting boundaries, although the proportions varied for each insect. The venom sac extracts showed varying patterns of minor fast components and major slow ones. At least some of the venom constituents seemed to be absent from the whole body extracts. It will require further study to identify any of these components with the allergenic substance.

SummaryKin-selection theory underlies our basic understanding of social evolution [1, 2]. Nest drifting in eusocial insects (where workers move between nests) presents a challenge to this paradigm, since a worker should remain as a helper on her natal colony, rather than visit other colonies to which she is less closely related. Here we reveal nest drifting as a strategy by which workers may maximize their indirect fitness by helping on several related nests, preferring those where the marginal return from their help is greatest. By using a novel monitoring technique, radio frequency identification (RFID) tagging, we provide the first accurate estimate of drifting in a eusocial insect: 56% of females drifted in a natural population of the eusocial paper wasp Polistes canadensis, exceeding previous records of drifting in natural populations by more than 30-fold. We demonstrate that drifting cannot be explained through social parasitism, queen succession, mistakes in nest identity, or methodological bias. Instead, workers appear to gain indirect fitness benefits by helping on several related colonies in a viscous population structure. The potential importance of this strategy as a component of the kin-selected benefits for a social insect worker has previously been overlooked because of methodological difficulties in quantifying and studying drifting.

RAST-inhibition studies were performed by use of whole venom sac extracts to inhibit binding to purified venom allergens from various vespid wasps. Immunodiffusion studies were also performed with rabbit antisera raised against purified venom proteins. Immunologic cross-reactivity was demonstrated among the hyaluronidases, phospholipases, and antigen 5 s from both subgenera of yellow jackets, white-faced hornets, and paper wasps. The paper wasp hyaluronidase and antigen 5 were less related to those of the other three species than those of yellow jackets and hornets. It appears that immunologic cross-reactivity is the major mechanism of multiple allergic sensitivity to vespid venoms. Therapy with only the primary venom should be sufficient to protect against reactions from cross-reacting venoms.

Polistes exclamans arizonensis Snelling was successfully overwintered in a cage provided with stacks of cardboard interspersed with spacers. When the wasps emerged in the spring they readily nested in a 1.2×1.8×1.2-m shelter with screen on 3 sides. The wasps were fed 4th- to 6th-stage larvae of the tobacco budworm, Heliothis virescens (F.), and provided with kraft paper torn into small bits for nesting material. A mean of 5.3 budworm larvae was rcquired for nutrition of each wasp.

Pure venoms were collected from individual insects of the species Dolichovespula maculata, white-faced hornet, Vespula squamosa, southern yellow jacket, and Polistes exclamans, paper wasp (one species). The venoms were first fractionated by high-resolution gel filtration on a 1.6 m column of Sephadex G-75 superfine, and the components were then purified by high-performance, ion-exchange chromatography on a Mono-S cation exchange column followed by a further gel filtration step. The isolated components were evaluated for purity by sodium dodecyl sulfate polyacrylamide gel electrophoresis by use of two different types of silver stains, by assays for enzyme activities, and by immunodiffusion with the use of rabbit antisera. The protein components were isolated in highly purified states by these techniques. Only three significant proteins were found in V. squamous venom: phospholipase (PL) A and B, hyaluronidase (HYAL), and antigen 5 (Ag 5). D. maculata venom contained HYAL, Ag 5, two isozymes of PL A and B, a high-molecular-weight protein, and several trace proteins. No significant amounts of proteases were found in D. maculata venom. P. exclamans venom contained HYAL, PL A and B, Ag 5, a high-molecular-weight protein, and several minor proteins. In all three venoms the PL A and B activities were found to be in the same molecule and did not separate. Trace components with apparent PL A activity were observed in the venoms. The venoms were screened for a variety of esterases, proteases, peptidases, glucosidases, and phosphatases, and none were detected in more than trace amounts. Vespid venoms do not appear to contain significant amounts of acid phosphatases as bee venoms do.

Extracts of whole body and of venom sac of wasps (Polistes exclamans) were fractionated by DEAE-cellulose chromatography. Nine fractions were separated from whole body, and eight from venom sac preparations. Fractions 6 of venom sac and 8 of whole body were the active components in direct skin testing. The antigenic heterogeneity of each of the fractionated products was examined by means of appropriate rabbit antisera to wasp extracts. All nine of the whole body fractions gave precipitation, as did four of the eight venom sac fractions. They variously gave between one and three lines of precipitation. In addition to this, fractions 6 and 7 of whole body showed, by testing with rabbit antiserum, common reactions, respectively, with bee and yellow jacket, and were presumed to be the components of wasp that were responsible for cross-sensitizations in patients. A final discussion, comparing the findings for bee, wasp, and yellow jacket, is given in the third paper of this series.

Certain low-molecular weight, basic proteins of Polistes exclamans, P. annularis, P. metricus, Vespula maculata (bald-faced hornet), and Apis mellifera (honey-bee) venoms are capable of complexing with normal (base line) rabbit serum. Precipitin lines form in agar (Ouchterlony double diffusion and immunoelectrophoresis) which have somewhat different qualities than true immunoprecipitin lines. They are less distinct and stain not only with protein stains but with lipophilic stains as well. The non-immunologic complexing involves a serum component (or components) other than Immunoglobulin G, perhaps lipoprotein.