Embryo culture of Medicago scutellata and M. sativa

Resistance to the alfalfa weevil (Hypera postica (Gyllenhal)) and the potato leafhopper (Empoasco fabae (Harris)) is lacking in cultivated alfalfa. However, a closely related annual Medicago, Medicago scutellata, possesses dense glandular stem and leaf hairs which provides a mechanism for resistance. Several attempts have been made at transfering the glandular haired traint from M. scutellata to perennial alfalfa with limited success. Earlier studies have shown that one reason for the lack of success is embryo abortion. Therefore, this study was initiated to observe zygotic embryo-genesis and to develop an embryo rescue technique for M. scutellata and M. sativa. Observations of zygotic embryogenesis showed that the two species are similar in morphology and can be described from youngest to oldest as globular, heart, torpedo, and hook shaped embryos. M. sativa embryos are smaller than M. scutellata embryos and develop three to four days later. Self pollinated M. scutellata (PI 307446) and sib mated M. sativa (Saranac AR) embryos were cultivated on Murashige and (2,4-D), indolacetic acid (IAA), 6-benzylaminopurine (BAP), and kinetic (KIN). Embryos from both species were also cultured on Schenk and Hildebrandt's (SH) basal medium with the addition of L-glutamine and L-proline. The experimental design was a completely randomized factorial for each experiment. Heart and torpedo shaped embryos from M. scutellata grew best (27.5% plantlet recovery) when cultured on MS medium with 0.05 mgl^-1 of both IAA and BAP. After 15 to 30 days on this medium, the embryos had only developed shoots. Therefore, it was necessary to transfer the shoots to MS basal medium without phytohormones for rooting. Rooting occurred in 15 to 30 days and the plantlets could be acclimatized to soil within 2 to 4 weeks. M. sativa embryos grew best (31% plantlet recovery) on SH medium with 50 mM L-glutamine. M. sativa embryos developed both shoots and roots on this medium. This information may now be applied to the development of an embryo culture method for recovering insect resistant hybrids between M. scutellata and M. sativa. Disclaimer statement: Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the USDA, and does not imply its approval to the exclusion of other products that may also be suitable.     

Vector competence of Culicoides bolitinos and C. imicola for South African bluetongue virus serotypes 1, 3 and 4

Abstract .The susceptibility of field-collected Culicoides bolitinos to infection by oral ingestion of bluetongue virus serotypes 1, 3 and 4 (BLU 1, 3 and 4) was compared with that of field-collected C. imicola and laboratory reared C. variipennis sonorensis . The concentration of the virus per millilitre of bloodmeal was 10^5.0 and 10^6.0TCID₅₀ for BLU 4 and 10^7.2TCID₅₀ for BLU 1 and 3. Of 4927 C. bolitinos and 9585 C. imicola fed, 386 and 287 individual midges survived 10 days extrinsic incubation, respectively. Midges were assayed for the presence of virus using a microtitration assay on BHK-21 cells and/or an antigen capture ELISA. Infection prevalences for the different serotypes as determined by virus isolation ranged from 22.7 to 82.0% in C. bolitinos and from 1.9 to 9.8% in C. imicola; infection prevalences were highest for BLU 1, and lowest for BLU 4 in both species. The mean log₁₀ TCID₅₀ titre of the three BLU viruses per single fly was higher in C. bolitinos than in C. imicola . The results suggested that C. bolitinos populations are capable vectors of the BLU viruses in South Africa. A high correlation was found between virus isolation and ELISA results for the detection of BLU 1, and less for BLU 4; the ELISA failed to detect the presence of BLU 3 in infected flies. The C. v. sonorensis colonies had a significantly lower susceptibility to infection with BLU 1, 3 and 4 than C. bolitinos and C. imicola . However, since infection prevalence of C. v. sonorensis was determined only by ELISA, this finding may merely reflect the insensitivity of this assay at low virus titres, compared to virus isolation.     

Infection of the gonads of Glaucosoma hebvaicum by the nematode Philometva lateolabracis: occurrence and host response

The prevalence of infection of the West Australian dhufish Glaucosoma hebraicum off the lower west coast of Australia by Philometra lateolabracis was greater among females than males, which contrasts with the situation with Pagrus auratus in waters off New Zealand. Live P. lateolabracis were represented solely by females and were found only in the gonads of fish of mature size ( L ₅₀ at first maturity = c . 300 mm) caught between December and April and thus during the spawning period of dhufish. Since the gonads were largest during that period, they, and particularly ovaries, would provide an abundant food source in the form of blood. The prevalence of the parasite (live + dead individuals) increased with host body size to reach a maximum of c . 80% in the 700–799 mm length class of females and c . 50% in the largest length class of males. During the spawning period of G hebraicum, P. lateolabracis developed from small, non-gravid females to large gravid females, representing an increase in mass of more than 200 times. The maximum length of P. lateolabracis (470 mm) in G hebraicum is the greatest yet recorded for this species. Gonadosomatic indices provided no evidence that infection by P. lateolabracis leads to a conspicuous atrophy of the ovaries, which contrasts with the situation with the gonads of some of the teleosts infected by Philometra species, and presumably reflects, in part, the small volume of the ovary occupied by P. lateolabracis (c . 3%) and the low intensity of infection (mean = 2.0, maximum = 7). Although the presence of live P. lateolabracis did not stimulate an obvious tissue response by the ovaries during the spawning period of G. hebraicum , both the dead and any live parasites that remain after spawning become encapsulated in fibrous tissue.     

Annual cycle of stored spermatozoa within the ovaries of Helicolenus dactylopterus dactylopterus (Teleostei, Scorpaenidae)

The aim of this work was to analyse the ultrastructure of storage crypts and stored spermatozoa, and to describe changes during the annual reproductive cycle of the bluemouth Helicolenus dactylopterus dactylopterus , which has internal fertilization and a zygoparous mode of reproduction. Spermatozoa had elongated heads and long midpieces, two characteristics which are thought to be fairly advanced and correlated with internal fertilization, as is the case of the bluemouth. A remarkable spermatozoon feature was the retention of a significant quantity of cytoplasm around the head, a condition that appeared to be related to nourishment during the long storage period, up to 10 months in the intraovarian crystal structures of the female. Male sex cells' protection against the female immune system was ensured by junctional complexes between the crypt cells composed of tight junctions and desmosomes.     

Phylogeography of the parasitic fly Batrachomyia in the Wet Tropics of north-east Australia, and susceptibility of host frog lineages in a mosaic contact zone

Parasites could differentially impact intraspecific host lineages due to genetic, phenotypic, ecological, or behavioural differences between the lineages, or the development of reproductive isolation between them. Batrachomyia (Diptera: Chloropidae) are flies that exclusively parasitize Australian frogs, and in the Wet Tropics rainforest of north‐east Australia larvae are largely restricted to the green‐eyed tree frog Litoria genimaculata (Anura: Hylidae). This frog species consists of two highly divergent genetic lineages that overlap in two nearby, but independent, contact zones. At one contact zone there has been extensive phenotypic divergence and speciation between the lineages whereas, at the other contact relatively lower levels of phenotypic divergence and reproductive isolation suggest that speciation has not occurred. In the present study we tested: (1) whether the deep phylogeographic divergence between northern and southern host populations is mirrored by congruent genetic structuring in the parasite populations and (2) whether the host lineages are differentially impacted by parasitism. We found that the two divergent frog lineages are parasitized by a single lineage of Batrachomyia, which exhibits strikingly little phylogeographic structuring. We found a significant difference in Batrachomyia prevalence between the host lineages at mixed lineage sites in both contact zones, with the magnitude and direction of this effect being consistent in both. The pattern did not differ between the two contacts even though recent phenotypic divergence and speciation has occurred between the lineages at one contact but not the other. Taken together, this suggests a fundamental difference in susceptibility between the genetically divergent host lineages. Using weight relative to body length as a measure of body condition, we found no differential impact of parasitism on the body condition of each host lineage, and no evidence that parasitism impacts the body condition of the host in general. © 2007 The Linnean Society of London, Biological Journal of the Linnean Society, 2007, 92 , 593–603.     

Egg surface structure in the annual fishes Simpsonichthys (subgenera Ophthalmolebias and Xenurolebias) and Nematolebias (Teleostei: Cyprinodontiformes: Rivulidae): variability and phylogenetic significance

The surfaces of the egg chorion of nine species of Simpsonichthys , two of Nematolebias and eight other species of rivulids were studied with scanning electron microscopy revealing details of ornamentation that were previously undocumented. Surface structures of eggs of Simpsonichthys of the subgenus Ophthalmolebias are described for the first time: eggs of Simpsonichthys constanciae , Simpsonichthys perpendicularis , Simpsonichthys rosaceus and Simpsonichthys suzarti are ornamented with a very characteristic palm-like structure, that is restricted to these species among the rivulids examined. The surface of the egg chorion of Simpsonichthys bokermanni lacks the well-developed palm-like structure, but has a characteristic conical projection, distinct from the structures of other rivulids examined, that is proposed as homologous to the palm-like structure. Simpsonichthys bokermanni shares with Nematolebias papilliferus and Nematolebias whitei the presence of concentric rows of spiny projections around the micropylar region. Species of Simpsonichthys of the subgenus Ophthalmolebias , Simpsonichthys myersi (of the subgenus Xenurolebias ), and N. whitei share the presence of large rounded protuberances on the surface of the egg chorion. The phylogenetic significance of these features are evaluated and discussed in light of current knowledge about rivulid relationships.     

Particulate organic matter in a mountain stream in the south-western Cape,South Africa

The quality and quantity of allochthonous inputs and of benthic organic matter were investigated in a second-order, perennial mountain stream in the south-west Cape, South Africa, between April 1983 and January 1986. Although the endemic, riparian vegetation is sclerophyllous, low and evergreen, inputs of allochthonous detritus to the stream (434 to 500 g m^–2y^–1) were similar to those recorded for riparian communities worldwide, as were calorific values of these inputs (9548 to 10 032 KJ m^–2y^–1). Leaf fall of the riparian vegetation is seasonal, occurring in spring (November) as discharge decreases, resulting in retention of benthic organic matter (BOM) on the stream bed during summer and early autumn (maximum 224 g m^–2). Early winter rains (May) scoured the stream almost clean of benthic detritus (winter minimum 8 g m^–2). Therefore, BOM was predictably plentiful for about half of each year and predictably scarce for the other half. Coarse BOM (CBOM) and fine BOM (FBOM) constituted 46–64% of BOM standing stock, ultra-fine BOM (UBOM) 16–33% and leaf packs 13–24%. The mean annual calorific value of total BOM standing stock was 1709 KJ m^–2. Both standing stocks and total calorific values of BOM were lower than those reported for streams in other biogeographical regions. Values of C:N ratios decreased with decrease in BOM particle size (CBOM 27–100; FBOM 25–27; UBOM 13–19) with no seasonal trends. The stream is erosive with a poor ability to retain organic detritus. Its character appears to be dictated by abiotic factors, the most important of which is winter spates.     

Phyletic and evolutionary relationships ofBrachyscome lineariloba (Compositae)

A comparison of karyotypes ofBrachyscome breviscapis (2n = 8),B. lineariloba cytodemes E (2n = 10), B (2n = 12) and C (2n = 16) suggests that these species have a homoelogous basic set of four chromosome pairs, two large pairs and two small, and that theB. lineariloba cytodemes E, B and C are related toB. breviscapis by successive additions of small chromosomes. A pronounced asynchrony of chromosome condensation between these large and small chromosomes has been observed. In the artificial hybrids betweenB. dichromosomatica (2n = 4) ×B. breviscapis, and theB. lineariloba cytodemes, theB. dichromosomatica chromosomes are similar in size and condensation behaviour to the small chromosomes ofB. breviscapis and ofB. lineariloba cytodemes E, B and C. Meiotic pairing in these hybrids also demonstrates the strong affinities between these chromosomes. It is suggested thatB. breviscapis may be of amphidiploid origin between a species with two large early condensing chromosome pairs and another,B. dichromosomatica-like species with two small late condensing pairs. It seems most likely that the additional small and late condensing chromosomes inB. lineariloba cytodemes E, B and C are derived from theB. dichromosomatica-like parent, and that each addition increases vigour, fecundity and drought tolerance, allowing these cytodemes to colonize more open and arid environments. Transmission of the univalents in the quasidiploidB. lineariloba cytodeme E was verified as being via the pollen, and not via the embryo sacs.The cytology ofBrachyscome lineariloba (Compositae, Asteroidae), 10.     

Seed Germination Biology of the Narrowly Endemic Species Lesquerella stonensis (Brassicaceae)

Abstract Lesquerella stonensis (Brassicaceae) is an obligate winter annual endemic to a small portion of Rutherford County in the Central Basin of Tennessee, where it grows in disturbed habitats. This species forms a persistent seed bank, and seeds remain viable in the soil for at least 6 years. Seeds are dormant at maturity in May and are dispersed as soon as they ripen. Some of the seeds produced in the current year, as well as some of those in the persistent seed bank, afterripen during late spring and summer; others do not afterripen and thus remain dormant. Seeds require actual or simulated spring/summer temperatures to come out of dormancy. Germination occurs in September and October. Fully afterripened seeds germinate over a wide range of thermoperiods (15/6–35/20°C) and to a much higher percentage in light (14 h photoperiod) than in darkness. The optimum daily thermoperiod for germination was 30/15°C. Nondormant seeds that do not germinate in autumn are induced back into dormancy (secondary dormancy) by low temperatures (e.g., 5°C) during winter, and those that are dormant do not afterripen; thus seeds cannot germinate in spring. These seed dormancy/ germination characteristics of L. stonensis do not differ from those reported for some geographically widespread, weedy species of winter annuals and thus do not help account for the narrow endemism of this species.     

Age and growth rate determinations of an intertidal bivalve, Phacosoma japonicum, using internal shell increments

A venerid bivalve Phacosoma japonicum (Reeve) occurring commonly in the Japanese coastal area preserves periodic growth lines in the shell cross-section. Long-term shell growth patterns of this species have been traced for many individuals on the intertidal flat of the Seto Inland Sea, west Japan. Sclerochronological analysis of these individuals and specimens collected monthly shows that several growth cessation marks within their shells are formed during the winter of each year prior to spawning. Hence the marks were used for age and growth rate determinations. As large individuals showed little shell growth for more than two years after the formation of 7 or 8 annual increments, this species probably has a lifespan of more than ten years. Shell growth patterns of this species based on annual increments can be accurately approximated by a von Bertalanffy curve. The number of microgrowth increments formed during a year tends to decrease with age, although it varies markedly among specimens of the same age. Furthermore, even in summer during rapid shell growth, the microgrowth increments do not represent daily and/or sub-daily tidal rhythms in many specimens. The results of this study and those by several authors strongly suggest that the annual increments are the key for age and growth rate determinations of both living and fossil bivalve species.