Towards a phylogeny and evolution of Acochlidia (Mollusca: Gastropoda: Opisthobranchia)

The Acochlidia are unique among opisthobranch gastropods in combining extremely high morphological and ecological diversity with modest species diversity. The phylogeny of acochlidians has never been addressed by cladistic means, as their evolution has remained unknown. This study gives a first overview on more than 150 biological and morphological characters that are potentially useful for phylogenetic analysis. Based on 107 characters, a parsimony analysis (PAUP) was performed for all 27 valid acochlidian species together with 11 (plus two) outgroup taxa. The resulting strict consensus tree shows a moderate overall resolution, with at least some bootstrap support for most resolved nodes. The Acochlidia are clearly monophyletic, and originate from an unresolved basal opisthobranch level. The Acochlidia split into the Hedylopsacea (Tantulum (Hedylopsis (Pseudunela (Strubellia (‘Acochlidium’, ‘Palliohedyle’))))) and Microhedylacea (Asperspina (Pontohedyle, ‘Parhedyle’, ‘Microhedyle’, (Ganitus, Paraganitus))). The formerly enigmatic Ganitidae, resembling sacoglossan opisthobranchs by having dagger‐like rachidian radular teeth, are likely to be highly derived microhedylids. The paraphyly of some of the traditionally recognized family level taxa induced a preliminary reclassification. From the phylogenetic hypothesis obtained, we conclude that the acochlidian ancestor was marine mesopsammic. The colonization of limnic systems occurred twice, independently: first in the Caribbean (with the development of the small interstitial Tantulum elegans), and second in the Indo‐Pacific, with a radiation of large‐sized benthic acochlidian species. The evolution of extraordinary reproductive features, such as hypodermic impregnation by a complex copulative aparatus in hedylopsaceans, cutaneous insemination via spermatophores in microhedylaceans, and gonochorism in Microhedylidae s.l. (including Ganitidae), is discussed. © 2010 The Linnean Society of London, Zoological Journal of the Linnean Society, 2010, 158 , 124–154.     

Toxic hydrogen sulfide and dark caves: phenotypic and genetic divergence across two abiotic environmental gradients in Poecilia mexicana

Divergent natural selection drives evolutionary diversification. It creates phenotypic diversity by favoring developmental plasticity within populations or genetic differentiation and local adaptation among populations. We investigated phenotypic and genetic divergence in the livebearing fish Poecilia mexicana along two abiotic environmental gradients. These fish typically inhabit nonsulfidic surface rivers, but also colonized sulfidic and cave habitats. We assessed phenotypic variation among a factorial combination of habitat types using geometric and traditional morphometrics, and genetic divergence using quantitative and molecular genetic analyses. Fish in caves (sulfidic or not) exhibited reduced eyes and slender bodies. Fish from sulfidic habitats (surface or cave) exhibited larger heads and longer gill filaments. Common-garden rearing suggested that these morphological differences are partly heritable. Population genetic analyses using microsatellites as well as cytochrome b gene sequences indicate high population differentiation over small spatial scale and very low rates of gene flow, especially among different habitat types. This suggests that divergent environmental conditions constitute barriers to gene flow. Strong molecular divergence over short distances as well as phenotypic and quantitative genetic divergence across habitats in directions classic to fish ecomorphology suggest that divergent selection is structuring phenotypic variation in this system.     

Impairment of mucosal immunity by total parenteral nutrition: requirement for IgA in murine nasotracheal anti-influenza immunity

Secretory IgA (SIgA) is the primary mucosal Ig and has been shown to mediate nasotracheal (NT) mucosal immunity in normal immune BALB/c mice. This finding has been challenged by a report of NT immunity without IgA in knockout mice, suggesting that IgA may not be necessary for the protection of mucosal surfaces. Although other protective mechanisms may become active in the congenital absence of SIgA, these mechanisms are not the primary means of protection in normal mice. In this paper we show that feeding chemically defined total parenteral nutrition (TPN) to genetically normal, immune ICR mice by the i.v. route results in loss of nasal anti-influenza immunity and a significant drop in influenza-specific SIgA in the upper respiratory tract compared with chow-fed mice (p < 0.005), while the serum influenza-specific IgG titer is unaffected. Loss of upper respiratory tract mucosal immunity is not related to serum Ab, because 10 of 13 TPN-fed mice shed virus into their nasal secretions despite adequate serum anti-influenza IgG titers. The number of IgG Ab-secreting cells in the nasal passages and spleens of TPN-fed mice was unaffected, while both the number and the percentage of splenic IgA-secreting cells were decreased relative to those in chow-fed animals. The loss of immunity is due to the route of nutrition, not the composition of the diet, because TPN solution fed orally via gastrostomy instead of i.v. maintains NT anti-influenza mucosal immunity. We hypothesize that delivery of nutrition via the gut triggers the release of gastrointestinal neuropeptides necessary for maintenance of the mucosal immune system.     

Cytosolic free Ca(2+) changes and calpain activation are required for beta integrin-accelerated phagocytosis by human neutrophils

Phagocytosis of microbes coated with opsonins such as the complement component C3bi is the key activity of neutrophils. However, the mechanism by which opsonins enhance the rate of phagocytosis by these cells is unknown and has been difficult to study, partly because of the problem of observing and quantifying the events associated with phagocytosis. In this study, C3bi-opsonized particles were presented to neutrophils with a micromanipulator, so that the events of binding, pseudopod cup formation, engulfment, and completion of phagocytosis were clearly defined and distinguished from those involved with chemotaxis. Using this approach in combination with simultaneous phase contrast and Ca(2+) imaging, the temporal relationship between changes in cytosolic free Ca(2+) concentration and phagocytosis were correlated. Here we show that whereas small, localized Ca(2+) changes occur at the site of particle attachment and cup formation as a result of store release, rapid engulfment of the particle required a global change in cytosolic free Ca(2+) which resulted from Ca(2+) influx. This latter rise in cytosolic free Ca(2+) concentration also liberated a fraction of beta2 integrin receptors which were initially immobile on the neutrophil surface, as demonstrable by both fluorescence recovery after laser bleaching and by visualization of localized beta2 integrin labelling. Inhibitors of calpain activation prevented both the Ca(2+)-induced liberation of beta2 integrin and the rapid stage of phagocytosis, despite the persistence of the global Ca(2+) signal. Therefore, we propose that Ca(2+) activation of calpain causes beta2 integrin liberation, and that this signal plays a key role in the acceleration of beta2 integrin-mediated phagocytosis.     

Phylogenetic analysis using insertion sequence fingerprinting in Escherichia coli

Chromosomal DNA from 23 closely related, pathogenic strains of Escherichia coli was digested and probed for the insertion sequences IS1, IS2, IS4, IS5, and IS30. Under the assumption that elements residing in DNA restriction fragments of the same apparent length are identical by descent, parsimony analysis of these characters yielded a unique phylogenetic tree. This analysis not only distinguished among bacterial strains that were otherwise identical in their biochemical characteristics and enzyme electrophoretic mobilities, but certain aspects of the topology of the tree were consistent across several unrelated insertion elements. The distribution of IS elements was then reexamined in light of the inferred phylogenetic relationships to investigate the biological properties of the elements, such as rates of insertion and deletion, and to discover apparent recombinational events. The analysis shows that the pattern of distribution of insertion elements in the bacterial genome is sufficiently stable for epidemiological studies. Although the rate of recombination by conjugation has been postulated to be low, at least two such events appear to have taken place.      

Microbial Transport, Survival, and Succession in a Sequence of Buried Sediments

Abstract Two chronosequences of unsaturated, buried loess sediments, ranging in age from <10,000 years to >1 million years, were investigated to reconstruct patterns of microbial ecological succession that have occurred since sediment burial. The relative importance of microbial transport and survival to succession was inferred from sediment ages, porewater ages, patterns of abundance (measured by direct counts, counts of culturable cells, and total phospholipid fatty acids), activities (measured by radiotracer and enzyme assays), and community composition (measured by phospholipid fatty acid patterns and Biolog substrate usage). Core samples were collected at two sites 40 km apart in the Palouse region of eastern Washington State, near the towns of Washtucna and Winona. The Washtucna site was flooded multiple times during the Pleistocene by glacial outburst floods; the Winona site elevation is above flood stage. Sediments at the Washtucna site were collected from near surface to 14.9 m depth, where the sediment age was approximately 250 ka and the porewater age was 3700 years; sample intervals at the Winona site ranged from near surface to 38 m (sediment age: approximately 1 Ma; porewater age: 1200 years). Microbial abundance and activities declined with depth at both sites; however, even the deepest, oldest sediments showed evidence of viable microorganisms. Same-age sediments had equal quantities of microorganisms, but different community types. Differences in community makeup between the two sites can be attributed to differences in groundwater recharge and paleoflooding. Estimates of the microbial community age can be constrained by porewater and sediment ages. In the shallower sediments (<9 m at Washtucna, <12 m at Winona), the microbial communities are likely similar in age to the groundwater; thus, microbial succession has been influenced by recent transport of microorganisms from the surface. In the deeper sediments, the populations may be considerably older than the porewater ages, since microbial transport is severely restricted in unsaturated sediments. This is particularly true at the Winona site, which was never flooded.     

Selective neutrality of glucose-6-phosphate dehydrogenase allozymes in Escherichia coli

Six naturally occurring alleles representing four electromorphs of the enzyme glucose-6-phosphate dehydrogenase were transferred by P1- mediated transduction from natural isolates of Escherichia coli into the genetic background of E. coli K12 and were studied in pairwise competition in chemostats limited for glucose in order to estimate differences in growth rate associated with the alleles. Although the level of resolution of such experiments is a growth rate differential of approximately 0.002 h-1, no significant differences among the strains were found. Studies of apparent Km and Vmax in crude enzyme extracts of the strains also failed to reveal any significant differences among the electromorphs. These results support the view that the alleles are selectively neutral or nearly neutral under these conditions.      

Haloperoxidases: Enzymatic Synthesis of alpha,beta-Halohydrins from Gaseous Alkenes

The enzymatic synthesis of alpha,beta-halohydrins from gaseous alkenes is described. The enzymatic reaction required an alkene, a halide ion, dilute hydrogen peroxide, and a haloperoxidase enzyme. A wide range of gaseous alkenes were suitable for this reaction, including those containing isolated, conjugated, and cumulative carbon-carbon double bonds. Chlorohydrins, bromohydrins, and iodohydrins could be formed. The combining of this enzymatic synthesis with a previously described enzymatic synthesis of epoxides from alpha,beta-halohydrins provides an alternate pathway, other than the well-known enzymatic direct epoxidation pathway, from alkene to an epoxide.