Seasonal development of the blowfly <Emphasis Type="Italic">Caliphora vicina</Emphasis> R.-D. (Diptera,Calliphoridae) in the environs of St. Petersburg

Four-year monitoring of phenology of the blowfly Caliphora vicina under quasi-natural conditions in the environs of St. Petersburg demonstrated development of two complete generations and overwintering of the third generation. The duration of preimaginal development (from egg laying to adult emergence) of the 1st and 2nd generations varied from 19 to 32 days; the required sum of effective temperatures (SET) was 326 ± 31 degree-days with a threshold of 5°С. Reproductive maturation of females (the period of time from adult emergence to laying of the first eggs) lasted from 15 to 31 days (SET was 262 ± 39 degree-days with a threshold of 5°С). The duration of development of one complete generation (from egg stage to the beginning of oviposition) during the period of observation varied from 34 to 57 days (SET was about 600 degree-days with a threshold of 5°С). The first diapausing larvae hatched from the eggs laid in the middle of August. In September, at shorter day lengths and at a temperature of 10–11°С, 80–100% of the larvae entered diapause. A small fraction of the larvae of the 1st and 2nd generation and most of the larvae of the 3rd generation overwintered. Successful overwintering of adults in the environs of St. Petersburg is hardly possible.     

Neurogenic and septic inductions of synthesis of peptide antibiotics in larvae of <Emphasis Type="Italic">Calliphora vicina</Emphasis> R.-D. (Diptera: Calliphoridae)

Synthesis of antimicrobial peptides in diapausing larvae of the blowfly Calliphora vicina can be induced by two different pathways. One pathway is well known in insects and includes recognition of microbial particles by the pattern-recognizing receptors. The other pathway includes perception and transduction of stress signal to immunocompetent cells by neuroendocrine system. This phenomenon consists in stimulation of synthesis of defensins, cecropins, and diptericins under effect of chromic stimulation of mechanoreceptors with ligature applied on the larva head end. Formation of immune response in brain is established to need less than 30 s, after which isolation of the neuroendocrine complex does not eliminate activation of immune response. As judging from rate of the neurogenic induction, transduction of the stimulating signal from brain to the immune system cells can be connected with release into hemolymph of biogenic amines or other neurohormones stored preliminarily in the neurohemal organ. The nature of this inductor at present remains unknown, as analysis of role of octopamine, dopamine, and adipokinetic hormone did not reveal stimulating effect on synthesis of bactericidal peptides. Physiological mechanism of this phenomenon is not finally understood, its key links seem to be CNS, hormonal factor of cardiac bodies, and system of antimicrobial peptides. Synthesis of antimicrobial peptides is directly regulated by the neuroendocrine system that can produce both stimulating and stress action by reminding in this aspect the known immunoneuroendocrine interrelations in vertebrates. The existence of similar integrating mechanisms in such polar animal kingdom groups as insects and vertebrates indicate that they are more ancient than this was considered earlier.     

Effect of food and temperature on the reproduction of the blowfly, <Emphasis Type="Italic">Calliphora vicina</Emphasis> R.-D. (Diptera,Calliphoridae), a popular model object in biological research

Egg maturation in Calliphora vicina is known to occur within a wide range of temperatures, from 12°C to nearly 30°C (Vinogradova, 1991). Photoperiodism has no effect on this process. Some females enter diapause already at 20°C; their fraction increases at lower temperatures and reaches 100% at 6°C. Reproducing females with eggs can survive for a long time and even lay eggs at low temperatures (4–5°C). Experiments with C. vicina from Leningrad Province revealed some effects of the diet (liver or fish) and temperature on the fly reproduction. At 20 and 25°C, 7–10-day old females begin to oviposit, but at 20°C egg maturation is observed in 98% of females feeding on liver and in only 5% of females feeding on fish. On the liver diet, the mean daily fecundity is significantly correlated with the day of oviposition but not with the temperature. At 20°C a significant correlation is observed between the mean daily fecundity and both the day of oviposition and food. The total number of eggs laid by flies after feeding on fish is half that produced after feeding on liver. The optimal conditions for Calliphora vicina cultivation are a 16-h light day, temperatures within the range from 20 to 25°C, and liver as food.     

Fat body and hemocyte contribution to the antimicrobial peptide synthesis in <Emphasis Type="Italic">Calliphora vicina</Emphasis> R.-D. (Diptera: Calliphoridae) larvae

Antimicrobial peptides accumulated in the hemolymph in response to infection are a key element of insect innate immunity. The involvement of the fat body and hemocytes in the antimicrobial peptide synthesis is widely acknowledged, although release of the peptides present in the hemolymph from the immune cells was not directly verified so far. Here, we studied the presence of antimicrobial peptides in the culture medium of fat body cells and hemocytes isolated from the blue blowfly Calliphora vicina using complex of liquid chromatography, mass spectrometry, and antimicrobial activity assays. Both fat body and hemocytes are shown to synthesize and release to culture medium defensin, cecropin, diptericins, and proline-rich peptides. The spectra of peptide antibiotics released by the fat body and hemocytes partially overlap. Thus, the results suggest that insect fat body and blood cells are capable of releasing mature antimicrobial peptides to the hemolymph. It is notable that the data obtained demonstrate dramatic difference in the functioning of insect antimicrobial peptides and their mammalian counterparts localized into blood cells’ phagosomes where they exert their antibacterial activity.     

Methods of short-term storage of cultures of the blowfly <Emphasis Type=Italic>Calliphora Vicina</Emphasis> R.-D. (Diptera,Calliphoridae)

Three methods of short-term storage of the blowfly Calliphora vicina strains are considered based on the experimental study of 21 strains originating from different parts of the species range. The colony can be preserved as diapausing adults at 6° and darkness for 2–3 months or more, depending on the geographical origin of the population. During the first five days of adult life the flies should be kept at 12° and short day on a sugar diet, after which they should be transferred into a refrigerator. During artificial hibernation the flies also require periodic sugar feeding every 20 days (3–4 h at 20°C) to maintain their vital functions. The combination of temperatures of 20–23°C and a protein diet terminates reproductive diapause, and oviposition starts in 10–17 days. The fly strain may be preserved as reproductive females at 6°C and darkness with sugar feeding. Flies also require periodic sugar feeding at 20°C (3–4 hours). In this case the flies start laying eggs 2–3 days after being transferred to 20–23°C. The preservation of diapausing larvae is a more reliable method of prolonged strain storage. In this case the flies of maternal generation are maintained at 20–23°C on sugar and protein diet. The egg rafts laid during 5–6 hours are then transferred into 12°C and short day until hatchment. The hatched larvae should be immediately placed into a refrigerator (2–3 or 5–6°C), where they feed during 1–1.5 months and enter diapause. For strain restoration, the diapausing larvae should be transferred into 20–25°C, where they pupariate in 3–5 days and the flies emerge in nearly 10 days.     

Some principles of selecting natural material for rearing and the endogenous processes in laboratory strains of the blowfly <Emphasis Type=Italic>Calliphora vicina</Emphasis> R.-D. (Diptera,Calliphoridae)

The ability to enter reproductive and larval diapauses should be taken into account when natural populations of the blowfly Calliphora vicina are selected for laboratory rearing. The reproductive diapause occurring at 20–25° in some females retards their maturation and decreases the reproductive activity of the laboratory strain. On the other hand, diapausing females conditioned by low temperature may be used for temporary conservation of the strain. However, keeping the diapausing larvae is the most reliable way of storage. The endogenous variation of the incidence of larval (from 0 to 100%) and reproductive (from 0 to 55%) diapauses was observed in successive generations of several laboratory strains developing under constant photoperiodic and temperature conditions. The significance of such endogenous processes for interpretation of experimental data is discussed. For over 20 years of laboratory breeding of C. vicina, cases of strain depression were observed, when the number of the egg rafts decreased and then oviposition ceased, abnormal puparia appeared, the flies hardly emerged, etc. Such depressed strains may be revived by changing the photoperiodic regime.     

Induction of larval diapause in the blowfly, Calliphora vicina R.-D. (Diptera, Calliphoridae) under field and laboratory conditions

Field experiments conducted in the environs of St. Petersburg (Russia) with a local population of Calliphora vicina showed that induction of larval diapause under natural conditions was significantly dependent on day lengths and temperature. The maternal photoperiodic response had a distinct threshold: the first diapausing larvae hatched from the eggs laid in the middle of August when the day length was 16 h; at shorter photoperiods, the fraction of diapausing larvae depended only on temperature. At the mean temperature of 16°C, larval diapause was rarely recorded; at 12–13°C, about 50% of the larvae entered diapause; at 7–9°C, nearly all the larvae entered diapause. These results of the field experiments agree well with the parameters of photoperiodic and thermal responses studied in the laboratory at constant temperatures and photoperiods.     

Induction of antimicrobial peptide synthesis by the fat body cells of maggots of <Emphasis Type=Italic>Calliphora vicina</Emphasis> R.-D. (Diptera,Calliphoridae)

Fat body is the major site of peptide antibiotic synthesis in insects. The goal of this work was to search in Calliphora vicina maggot for humoral factors that induce synthesis of antimicrobial peptides by the fat body cells in trauma. The preliminary analysis has shown that the activation factor revealed in hemolymph is a thermostable hydrophilic compound with a molecular mass lower than 3 kDa. The integument epitheliocytes were also found to release the humoral factors directly stimulating synthesis of antimicrobial peptides by the fat body.     

Regulation of antimicrobial peptide synthesis in larvae of <Emphasis Type="Italic">Calliphora vicina</Emphasis> (Diptera,Calliphoridae): a dose-dependent effect of ecdysteroids

Ecdysteroids are multifunctional hormones regulating virtually all morphogenetic events in insects. Their role in stress and immune response regulation is not well researched. Here we study the effect of 20-hydroxyecdysone on antimicrobial peptide synthesis in the larvae of Calliphora vicina. An inverse correlation was observed between the 20-hydroxyecdysone titer and antimicrobial peptide concentration in the hemolymph of naïve and bacteria challenged maggots. High and low doses of 20-hydroxyecdysone, injected simultaneously with bacterial cells, had an opposite effect on antimicrobial peptide synthesis in the diapausing larvae. Morphogenetically effective doses of 20-hydroxyecdysone demonstrated immunosuppressive activity. A low dose of 20-hydroxyecdysone, on the contrary, moderately stimulated antimicrobial peptide synthesis. The data suggest that ecdysteroids are directly involved in the regulation of the immune system activity and that the final effect is dose-dependant.     

<Emphasis Type="Italic">Ty</Emphasis>1<Emphasis Type="Italic">/copia</Emphasis>- and <Emphasis Type="Italic">Ty</Emphasis>3<Emphasis Type="Italic">/gypsy</Emphasis>-like DNA sequences in <Emphasis Type="Italic">Helianthus </Emphasis>species

Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes.     

Dispersal and Burial Behavior in Larvae of <Emphasis Type="Italic">Chrysomya megacephala</Emphasis> and <Emphasis Type="Italic">Chrysomya albiceps</Emphasis> (Diptera,Calliphoridae)

Blowflies use discrete and ephemeral substrates to feed their larva. After they run out of food, the larvae begin to disperse in order to find adequate places for pupation or additional food sources, a process named post-feeding larval dispersal. Briefly state the aspects and why they are important were studied in a circular arena of 25 cm in diameter and covered with wood shavings to a height of 40 cm allowing post-feeding dispersal from the center of the arena. Larvae of both Chrysomya albiceps and C. megacephala were used in five experiments for each species. For each pupa location, determined as distance from the center, depth, and weight were evaluated. Statistical tests were done to verify the relation between weight, depth and distance for pupation and for larvae of two species shows that the media distance is significantly different for two species and for C. megacephala this distance is greater than the distance for C. albiceps. The depth too is different for each species, as the larvae of C. megacephala buries deeper than C. albiceps. With relation of weight, there is no statistic evidence that have any difference between weights for pupation for each species.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.     

<Emphasis Type="Italic">Quinua</Emphasis> and Relatives (<Emphasis Type="Italic">Chenopodium</Emphasis> sect.<Emphasis Type="Italic">Chenopodium</Emphasis> subsect.<Emphasis Type="Italic">Celluloid</Emphasis>)

Traditionally viewed as an Andean grain crop,Chenopodium quinoa Willd. includes domesticated populations that are not Andean, and Andean populations that are not domesticated. Comparative analysis of leaf morphology and allozyme frequencies have demonstrated that Andean populations, both domesticated(quinua) and free-living(ajara), represent an exceptionally homogeneous unit that is well differentiated from allied domesticates of coastal Chile(quingua) and freeliving populations of the Argentine lowlands(C. hircinum). This pattern of relationships indicates that Andean populations represent a monophyletic crop/weed system that has possibly developed through cyclic differentiation (natural vs. human selection) and introgressive hybridization. Relative levels of variation suggest that this complex originated in the southern Andes, possibly from wild types allied withC. hircinum, with subsequent dispersal north to Colombia and south to the Chilean coast. Coastal populations were apparently isolated from post-dispersal differentiation and homogenization that occurred in the Andes. Other data point toward a center of origin in the northern Andes with secondary centers of genetic diversity subsequently developing in the southern Andes and the plains of Argentina. Comparative linkage of South American taxa, all tetraploid, with North American tetraploids of the subsection will eventually clarify this problem. While the possibility of a direct phyletic connection betweenC. quinoa and the Mexican domesticate(C. berlandieri subsp. nuttalliae,) cannot be excluded, available evidence indicates that the latter represents an autonomous lineage that is associated with the basal tetraploid, C. b. subsp.berlandieri, through var.sinuatum, whereas South American taxa show possible affinities to either var. zschackei or var.berlandieri. An extinct domesticate of eastern North America,C. b. subsp.jonesianum, represents either another instance of independent domestication, possibly from subsp. b. var.zschackei, or a northeastern outlier of subsp.nuttalliae.     

In planta transformation of <Emphasis Type="Italic">Notocactus scopa</Emphasis> cv. <Emphasis Type="Italic">Soonjung</Emphasis> by <Emphasis Type="Italic">Agrobacterium </Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>

Notocactus scopa cv. Soonjung was subjected to in planta Agrobacterium tumefaciens-mediated transformation with vacuum infiltration, pin-pricking, and a combination of the two methods. The pin-pricking combined with vacuum infiltration (20-30 cmHg for 15 min) resulted in a transformation efficiency of 67-100%, and the expression of the uidA and nptII genes was detected in transformed cactus. The established in planta transformation technique generated a transgenic cactus with higher transformation efficiency, shortened selection process, and stable gene expression via asexual reproduction. All of the results showed that the in planta transformation method utilized in the current study provided an efficient and time-saving procedure for the delivery of genes into the cactus genome, and that this technique can be applied to other asexually reproducing succulent plant species.     

Dispersal and Predation Behavior in Larvae of <Emphasis Type="Italic">Chrysomya albiceps</Emphasis> and <Emphasis Type="Italic">Chrysomya megacephala</Emphasis> (Diptera: Calliphoridae)

Chrysomya albiceps and Chrysomya megacephala are exotic blowfly species known by producing myiasis in humans and other animals and by transmitting pathogens mechanically. C. albiceps stand out by being a facultative predator of other dipteran larvae. In this paper we investigated the influence of larval predation on the dispersal of larvae of C. albiceps and C. megacephala single and double species for three photophases. An experimental acrylic channel graduated and covered with wood shavings was used to observe the larval dispersal. The results showed that C. albiceps attacks C. megacephala larvae during dispersal and keeps an aggregated pattern close to the release point, in single and double species, independently of the different photophases. Chrysomya megacephala single species exhibited the same pattern, but in double species this was changed to a random distribution.     

Foraging behaviour by an intraguild predator blowfly, <Emphasis Type="Italic">Chrysomya albiceps</Emphasis> (Diptera: Calliphoridae)

Optimal foraging theory assumes that predators use different prey types to maximize their rate of energetic gain. Studies focusing on prey preference are important sources of information to understand the foraging dynamics of Chrysomya albiceps. The purpose of this investigation is to determine the influence of larval starvation in C. albiceps on the predation rate of different prey blowfly species and instars under laboratory conditions. Our results suggest that C. albiceps prefers Cochliomyia macellaria larvae to Chrysomya megacephala under non-starvation and starvation conditions. Nevertheless, predators gained more weight consuming C. macellaria. This result suggests that C. albiceps profit more in consuming C. macellaria rather than C. megacephala. The foraging behaviour displayed by C. abiceps on their prey and the consequences for the blowfly community are also discussed.     

Both cooperation and nest position improve larval survival in the sweat bee, <Emphasis Type="Italic">Lasioglossum</Emphasis> (<Emphasis Type="Italic">Evylaeus</Emphasis>) <Emphasis Type="Italic">baleicum</Emphasis>

Nesting by multiple females in sweat bees raises the question of the benefits associated with grouping. Adult numbers of the sweat bee, Lasioglossum (Evylaeus) baleicum, exhibited variation within a nest population. The effects of adult number and nest position on larval survival were analyzed in a nest aggregation. Both variables independently affected larval survival, with the presence of multiple adults in a nest markedly improving survival rate. Nests located near the periphery of the aggregation of nests suffered high larval mortalities. Increased frequency of cell-inspection in multiple-female nests seems to be associated with a concomitant increase in larval cell protection from external predation. Such predation pressure was assumed, given the existence of an underground ant colony that employs a recruitment system while foraging in the study area.     

Common genomic features of <Emphasis Type="Italic">Campylobacter jejuni</Emphasis> subsp. <Emphasis Type="Italic">doylei</Emphasis> strains distinguish them from <Emphasis Type="Italic">C. jejuni</Emphasis> subsp. <Emphasis Type="Italic">jejuni</Emphasis>

Background  

Campylobacter jejuni has been divided into two subspecies: C. jejuni subsp. jejuni (Cjj) and C. jejuni subsp. doylei (Cjd). Nearly all of the C. jejuni strains isolated are Cjj; nevertheless, although Cjd strains are isolated infrequently, they differ from Cjj in two key aspects: they are obtained primarily from human clinical samples and are associated often with bacteremia, in addition to gastroenteritis. In this study, we utilized multilocus sequence typing (MLST) and a DNA microarray-based comparative genomic indexing (CGI) approach to examine the genomic diversity and gene content of Cjd strains.