Cloning,expression and cellular localization of Daphnia pulex senescence-associated protein,DpSAP

Daphnia (water fleas) are small crustaceans that undergo an unusual switch from asexual to sexual reproduction that is dependent on environmental conditions. In this study, a senescence-associated protein (SAP) from the common freshwater species Daphnia pulex was cloned using primers based on homologous sequences and rapid amplification of cDNA ends (RACE). Real-time PCR was employed to quantify the expression of D. pulex SAP (DpSAP) in individual organisms. The role of DpSAP in the reproductive transformation was further investigated in both parthenogenetic and sexual females by using digoxin-labeled SAP RNA probes and RNA whole-mount in situ hybridization. DpSAP was more highly expressed in sexual females, indicating a role in growth and reproduction. Cellular localization studies using RNA whole-mount in situ hybridization showed specific expression in the second tentacle joints. These expression patterns suggest an important role for DpSAP in the reproductive transformation of D. pulex.     

Description of Lentistivalius philippinensis,a new species of flea (Siphonaptera,Pygiosyllomorpha, Stivaliidae), and new records of Ascodipterinae (Streblidae) on bats and other small mammals from Luzon,The Philippines

During May 2009 and July 2011, we collected 357 mammals and examined each for ectoparasites. Among the ectoparasites collected, a new species of flea was discovered. This new species, Lentistivalius philippinensis, is described from the male sex only. Two males were recovered from two specimens of the soricid Crocidura grayi Dobson in Municipality Maria Aurora, Aurora Province, Luzon, Philippines. Additional fleas included Thaumapsylla breviceps orientalis Smit, Thaumapsylla longiforceps Traub, and Ischnopsyllus indicus Jordan. Although the latter species is common in Japan and documented in Guam (as well as mainland Southeast Asia) also on Pipistrellus javanicus (Gray), Ischnopsyllus indicus represents a new record in the Philippine Islands. The ascodipterinae (Streblidae) Maabella stomalata and Ascodipteron speiserianum Muir collected from Rhinolophus inops K. Andersen and Rhinolophus subrufus K. Andersen, respectively, also represent new host records. A key to the species of the flea genus Lentistivalius Traub is provided.     

Temperature Effects on the Attachment of Pasteuria penetrans Endospores to Meloidogyne arenaria Race 1

Pasteuria penetrans is a gram positive bacterium that prevents Meloidogyne spp. from reproducing and diminishes their ability to penetrate roots. The attachment of the endospores to the cuticle of the nematodes is the first step in the life cycle of the bacterium and is essential for its reproduction. As a preliminary study to a field solarization test, the effects of temperature on the attachment of P. penetrans on Meloidogyne arenaria race 1 were investigated. Preexposing second-stage juveniles (J2) of M. arenaria to approximately 30 °C in water before exposing them to endospores increased their receptivity to endospore attachment when compared to treating J2 at 25 °C or 35 °C. In tests with soil, highest attachment occurred when J2 were incubated in soil infested with endospores and maintained at 20 °C to 30 °C for 4 days. Heating J2 in soil to sublethal temperatures (35 °C to 40 °C) decreased endospore attachment. Incubating P. penetrans endospores in soil at 30 °C to 70 °C for 5 hours a day over 10 days resulted in reductions of endospore attachment to nematodes as temperatures of incubation increased to 50 °C and higher.     

DNA transposon dynamics in populations of Daphnia pulex with and without sex

We investigate the role of recombination in transposable element (TE) proliferation in the cyclical parthenogen, Daphnia pulex. Recombination provides a mechanism by which the rate of both TE gain and loss can be accelerated, a duality that has long intrigued many biologists interested in the influence of sex on mutation accumulation. We compared TE loads among populations of D. pulex where sex occurs regularly (cyclical parthenogens or ‘sexuals’) with those in which the ability to reproduce sexually has been completely lost (obligate ‘asexuals’) for six different families of DNA transposons. Transposon display assays showed that sexuals have more TEs than asexuals, contrary to the expectations under Muller''s ratchet but consistent with the idea that sex facilitates TE spread. Sexuals also exhibit higher insertion site polymorphism among lineages, as predicted because recombination accelerates rates of loss and gain. Asexuals, however, have proportionally more singletons (loci occupied in a single isolate), which differs from previous studies where selfing and outcrossing were used as a proxy for high and low recombination. Our multi-element survey reveals that the impact of sex on TE proliferation is consistent among different Class II TE families and we discuss the genomic consequences of different reproductive strategies over long time periods.     

Effects of Monoclonal Antibodies,Cationized Ferritin,and Other Organic Molecules on Adhesion of Pasteuria penetrans Endospores to Meloidogyne incognita

The incidence of adhesion of Pasteuria penetrans endospores to Meloidogyne incognita second-stage juveniles (J2) was studied after pretreatment of the latter with monoclonal antibodies (MAb), cationized ferritin, and other organic molecules in replicated trials. Monoclonal antibodies developed to a cuticular epitope of M. incognita second-stage juveniles gave significant reductions in attachment of P. penetrans endospores to treated nematodes. MAb bound to the entire length of J2 except for the area of the lateral field, where binding was restricted to the incisures. Since reductions in attachment with MAb treatment were modest, it is uncertain if these results implicated a specific surface protein as a factor that interacted in binding of the endospore to the nematode cuticle. Endospore attachment was decreased following treatment of the nematode with the detergents sodium dodecyl sulfate (SDS) and cetyltrimethylammonium bromide (CTAB). Endospore attachment to live nematodes was significantly greater than attachment to dead nematodes. Attachment rates of three P. penetrans isolates to M. incognita race 3 varied between isolates. The effects of neuraminidase, pronase, pepsin, trypsin, lipase, and Na periodate on endospore attachment were inconsistent. The cationic dye alcian blue, which binds sulfate and carboxyl groups on acidic glycans, had no consistent effect on endospore attachment. The incidence of endospore attachment was significantly lower but modest, at best, for nematodes that were treated with cationized ferritin alone or cationized ferritin following monoclonal antibody. The lack of consistency or extreme reduction in most experiments suggests that attachment of P. penetrans spores to M. incognita is not specified by only one physico-chemical factor, but may involve a combination of at least two physico-chemical factors (including surface charge and movement of the J2). This points to a need for analysis of combined or factorial treatment effects.     

Survey of Flea Infestation in Dogs in Different Geographical Regions of Iran

Medically important arthropods, including fleas, play an important role in causing clinical disorders and disease in man and domestic animals. This study was conducted to determine the seasonal flea infestations for domestic dogs from different geographic regions of Iran. A total of 407 fleas, belonging to 5 different species, were recovered from 83 domestic dogs from 3 regions. There was a distinctive pattern of species distribution and infestations with the highest infestation rates observed in a temperate climate and higher rainfall. Additionally, fleas were observed over all seasons, except February and March, with the highest infestation rate observed in August (24.7%) and the lowest rate in January (1.7%). They also parasitize dogs with a different spectrum of species. The cat flea, Ctenocephalides felis (67.5%), exhibited the highest prevalence among all flea species found on dogs. Thus, climatic conditions and seasonal patterns impact on flea infestation and must be considered in developing control programs.     

Mapping of the Interaction Between Agrobacterium tumefaciens and Vanda Kasem’s Delight Orchid Protocorm-Like Bodies

Physical contact between A. tumefaciens and the target plant cell walls is essential to transfer and integrate the transgene to introduce a novel trait. Chemotaxis response and attachment of Agrobacterium towards Vanda Kasem’s Delight (VKD) protocorm-like bodies (PLBs) were studied to analyse the interaction between Agrobacterium and PLB during the transformation event. The study shows that initially A. tumefaciens reversibly attached to PLB surface via polar and lateral mode of adherence followed by the irreversible attachment which involved the production of cellulosic fibril by A. tumefaciens. Cellulosic fibril allows formation of biofilm at the tip of trichome. Contrarily, attachment mutant Escherichia coli strain DH5α was significantly deficient in the attachment process. Spectrophotometric GUS assay showed the mean value of attachment by A. tumefaciens was 8.72 % compared to the negative control E. coli strain DH5α that produced 0.16 %. A. tumefaciens swarmed with sharper and brighter edge when severe wounding was applied to the PLBs producing the highest swarming ratio of 1.46 demonstrating the positive effect of the plant exudates on bacterial movement. The study shows that VKD’s PLBs are the suitable explants for Agrobacterium-mediated transformation since the bacteria expressed higher competency rate.

Electronic supplementary material

The online version of this article (doi:10.1007/s12088-015-0519-7) contains supplementary material, which is available to authorized users.     

Description of Medwayella independencia (Siphonaptera,Stivaliidae), a new species of flea from Mindanao Island,the Philippines and their phoretic mites,and miscellaneous flea records from the Malay Archipelago

Medwayella independencia, a new species of flea, is described from the tupaiid host Urogale everetti (Thomas) from Mindanao Island, Philippines. Several other species of fleas are also recorded from the Philippines including a single male of Lentistivalius philippinensis Hastriter and Bush, 2013 (previously known only from two males), the bat fleas Thaumapsylla breviceps orientalis Smit and Thaumapsylla longiforceps Traub, a single unidentified female species of Macrostylophora Ewing collected from the murid Bullimus bagobos Mearns, and a pair of Medwayella robinsoni ssp. from Sundasciurus hoogstraali (Sanborn) from Busuanga Island, Philippines. Representatives of Medwayella Traub, 1972 and Macrostylophora have not previously been recorded from the Philippines. A key to the male sex of Medwayella is provided. Phoretic mites of the genus Psylloglyphus (family Winterschmidtiidae) were present under the abdominal sclerites of several male and female specimens of M. independencia. This is the second report of a phoretic mite on a species of Medwayella Traub. The co-evolutionary implications between phoretic mites and fleas are discussed.     

UV radiation affects antipredatory defense traits in Daphnia pulex

In aquatic environments, prey perceive predator threats by chemical cues called kairomones, which can induce changes in their morphology, life histories, and behavior. Predator‐induced defenses have allowed for prey, such as Daphnia pulex, to avert capture by common invertebrate predators, such as Chaoborus sp. larvae. However, the influence of additional stressors, such as ultraviolet radiation (UVR), on the Daphnia–Chaoborus interaction is not settled as UVR may for instance deactivate the kairomone. In laboratory experiments, we investigated the combined effect of kairomones and UVR at ecologically relevant levels on induced morphological defenses of two D. pulex clones. We found that kairomones were not deactivated by UVR exposure. Instead, UVR exposure suppressed induced morphological defense traits of D. pulex juveniles under predation threat by generally decreasing the number of neckteeth and especially by decreasing the size of the pedestal beneath the neckteeth. UVR exposure also decreased the body length, body width, and tail spine length of juveniles, likely additionally increasing the vulnerability to Chaoborus predation. Our results suggest potential detrimental effects on fitness and survival of D. pulex subject to UVR stress, with consequences on community composition and food web structure in clear and shallow water bodies.     

The effect of ecological and temporal factors on the composition of Bartonella infection in rodents and their fleas

The composition of Bartonella infection was explored in wild Gerbillus andersoni rodents and their Synosternus cleopatrae fleas. Rodent blood samples and fleas were collected in two periods (two different seasons; 4 months apart) from juveniles and adult hosts, and their bartonellae lineages were identified by a 454-pyrosequencing analysis targeting a specific Bartonella citrate synthase gene (gltA) fragment. The rate of Bartonella spp. co-infection was estimated and the assemblage and distribution of bartonellae lineages across the samples with respect to ecological and phylogenetic distance similarities were analyzed. Moreover, environmental factors that could explain potential differences between samples were investigated. Out of the 91 bartonellae-positive samples, 89% were found to be co-infected with more than two phylogenetically distant Bartonella genotypes and additional closely related (but distinguishable) variants. These bartonellae lineages were distributed in a non-random manner, and a negative interaction between lineages was discovered. Interestingly, the overall composition of those infections greatly varied among samples. This variability was partially explained by factors, such as type of sample (blood versus fleas), flea sex and period of collection. This investigation sheds light on the patterns of Bartonella infection and the organization of Bartonella lineages in fleas and rodents in nature.     

kakapo, a Gene Required for Adhesion Between and Within Cell Layers in Drosophila, Encodes a Large Cytoskeletal Linker Protein Related to Plectin and Dystrophin

Mutations in kakapo were recovered in genetic screens designed to isolate genes required for integrin-mediated adhesion in Drosophila. We cloned the gene and found that it encodes a large protein (>5,000 amino acids) that is highly similar to plectin and BPAG1 over the first 1,000–amino acid region, and contains within this region an α-actinin type actin-binding domain. A central region containing dystrophin-like repeats is followed by a carboxy domain that is distinct from plectin and dystrophin, having neither the intermediate filament-binding domain of plectin nor the dystroglycan/syntrophin-binding domain of dystrophin. Instead, Kakapo has a carboxy terminus similar to the growth arrest–specific protein Gas2. Kakapo is strongly expressed late during embryogenesis at the most prominent site of position-specific integrin adhesion, the muscle attachment sites. It is concentrated at apical and basal surfaces of epidermal muscle attachment cells, at the termini of the prominent microtubule bundles, and is required in these cells for strong attachment to muscles. Kakapo is also expressed more widely at a lower level where it is essential for epidermal cell layer stability. These results suggest that the Kakapo protein forms essential links among integrins, actin, and microtubules.     

The chemical cue tetrabromopyrrole from a biofilm bacterium induces settlement of multiple Caribbean corals

Microbial biofilms induce larval settlement for some invertebrates, including corals; however, the chemical cues involved have rarely been identified. Here, we demonstrate the role of microbial biofilms in inducing larval settlement with the Caribbean coral Porites astreoides and report the first instance of a chemical cue isolated from a marine biofilm bacterium that induces complete settlement (attachment and metamorphosis) of Caribbean coral larvae. Larvae settled in response to natural biofilms, and the response was eliminated when biofilms were treated with antibiotics. A similar settlement response was elicited by monospecific biofilms of a single bacterial strain, Pseudoalteromonas sp. PS5, isolated from the surface biofilm of a crustose coralline alga. The activity of Pseudoalteromonas sp. PS5 was attributed to the production of a single compound, tetrabromopyrrole (TBP), which has been shown previously to induce metamorphosis without attachment in Pacific acroporid corals. In addition to inducing settlement of brooded larvae (P. astreoides), TBP also induced larval settlement for two broadcast-spawning species, Orbicella (formerly Montastraea) franksi and Acropora palmata, indicating that this compound may have widespread importance among Caribbean coral species.     

Effect of Temperature on Attachment,Development, and Interactions of Pasteuria penetrans on Meloidogyne incognita

The effect of temperature (10, 20, 25, 30, and 35 C) on attachment and development of Pasteuria penetrans on Meloidogyne arenaria race 1 was elevated in growth chambers. The greatest attachment rate of endospores of P. penetrans occurred on second-stage juveniles at 30 C. The bacterium developed more quickly within its host at 30 and 35 C than at 25 C or below. The development of the bacterium within the nematode female was divided into nine recognizable life stages, which ranged from early vegetative thalli to mature sporangia. Mature sporangium was the predominant life stage observed after 35, 40, 81, and 116 days at 35, 30, 25, and 20 C, respectively. The body width and length of M. arenaria females infected with P. penetrans were smaller initially than the same dimensions in uninfected females, but became considerably larger over time at 25, 30, and 35 C. This isolate of P. penetrans also parasitized and completed its life cycle in males of M. arenaria.     

In Silico Prediction and In Vivo Validation of Daphnia pulex Micrornas

Daphnia pulex, the crustacean with the first sequenced genome, is an important organism that has been widely used in ecological and toxicological research. MicroRNAs (miRNAs) are 21–25 nucleotide small non-coding RNAs that are involved in a myriad of physiological processes. In this research, we predicted 75 D. pulex miRNAs by sequence homology and secondary structure identification from the full genome sequence. Fourteen predicted miRNAs were selected for quantitative real time polymerase chain reaction (RT-PCR) validation. Out of these, eight (mir-8, mir-9, mir-12, mir-92, mir-100, mir-133, mir-153 and mir-283) were successfully amplified and validated. Next, expression levels were quantified at three different life stages (days 4, 8 and 12 of age) using U6 spliceosomal RNA as a reference gene. The expression of mir-8, mir-9, mir-12, mir-92 and mir-100 significantly differed across time suggesting these microRNAs might play a critical role during D. pulex development. This is the first study to identify and validate miRNAs in D. pulex, which is an important first step in further studies that evaluate their roles in development and response to environmental and ecological stimuli.     

Transcontinental Phylogeography of the Daphnia pulex Species Complex

Daphnia pulex is quickly becoming an attractive model species in the field of ecological genomics due to the recent release of its complete genome sequence, a wide variety of new genomic resources, and a rich history of ecological data. Sequences of the mitochondrial NADH dehydrogenase subunit 5 and cytochrome c oxidase subunit 1 genes were used to assess the global phylogeography of this species, and to further elucidate its phylogenetic relationship to other members of the Daphnia pulex species complex. Using both newly acquired and previously published data, we analyzed 398 individuals from collections spanning five continents. Eleven strongly supported lineages were found within the D. pulex complex, and one lineage in particular, panarctic D. pulex, has very little phylogeographical structure and a near worldwide distribution. Mismatch distribution, haplotype network, and population genetic analyses are compatible with a North American origin for this lineage and subsequent spatial expansion in the Late Pleistocene. In addition, our analyses suggest that dispersal between North and South America of this and other species in the D. pulex complex has occurred multiple times, and is predominantly from north to south. Our results provide additional support for the evolutionary relationships of the eleven main mitochondrial lineages of the D. pulex complex. We found that the well-studied panarctic D. pulex is present on every continent except Australia and Antarctica. Despite being geographically very widespread, there is a lack of strong regionalism in the mitochondrial genomes of panarctic D. pulex – a pattern that differs from that of most studied cladocerans. Moreover, our analyses suggest recent expansion of the panarctic D. pulex lineage, with some continents sharing haplotypes. The hypothesis that hybrid asexuality has contributed to the recent and unusual geographic success of the panarctic D. pulex lineage warrants further study.     

Predators modify the evolutionary response of prey to temperature change

As climate regimes shift in many ecosystems worldwide, evolution may be a critical process allowing persistence in rapidly changing environments. Organisms regularly interact with other species, yet whether climate-mediated evolution can occur in the context of species interactions is not well understood. We tested whether a species interaction could modify evolutionary responses to temperature. We demonstrate that predation pressure by Dipteran larvae (Chaoborus americanus) modified the evolutionary response of a freshwater crustacean (Daphnia pulex) to its thermal environment over approximately seven generations in laboratory conditions. Daphnia kept at 21°C evolved higher population growth rates than those kept at 18°C, but only in those populations that were also reared with predators. Furthermore, predator-mediated selection resulted in the evolution of elevated Daphnia thermal plasticity. This laboratory natural selection experiment demonstrates that biotic interactions can modify evolutionary adaptation to temperature. Understanding the interplay between multiple selective forces can improve predictions of ecological and evolutionary responses of organisms to rapid environmental change.