Microfabricated silicon leak for sampling planetary atmospheres with a mass spectrometer.

A microfabricated silicon mass spectrometer inlet leak has been designed, fabricated, and tested. This leak achieves a much lower conductance in a smaller volume than is possible with commonly available metal or glass capillary tubing. It will also be shown that it is possible to integrate significant additional functionality, such as inlet heaters and valves, into a silicon microleak with very little additional mass. The fabricated leak is compatible with high temperature (up to 500 degrees C) and high pressure (up to 100 bars) conditions, as would be encountered on a Venus atmospheric probe. These leaks behave in reasonable agreement with their theoretically calculated conductance, although this differs between devices and from the predicted value by as much as a factor of 2. This variation is believed to be the result of nonuniformity in the silicon etching process which is characterized in this work. Future versions of this device can compensate for characterized process variations in order to produce devices in closer agreement with designed conductance values. The integration of an inlet heater into the leak device has also been demonstrated in this work.

Descriptions of the mature spermatozoa of the lizards Crotaphytus bicinctores, Gambelia wislizenii (Crotaphytidae), and Anolis carolinensis (Polychrotidae) (Reptilia, Squamata, Iguania).

The spermatozoa of Crotaphytus bicinctores and Gambelia wislizenii (Crotaphytidae), and Anolis carolinensis (Polychrotidae) exhibit the squamate autapomorphies of a single perforatorium extending anteriorly from the apical tip of the paracrystalline subacrosomal cone, the presence of an epinuclear electron-lucent region, and extension of the fibrous sheath into the midpiece. Crotaphytid sperm differ from those of polychrotids in several respects, including: the structure of the perforatorium, the size of the epinuclear electron-lucent region, aspects of the acrosome complex, the arrangement and structure of intermitochondrial dense bodies, and in the distance the fibrous sheath extends into the midpiece. The sperm of C. bicinctores, G. wislizenii, and A. carolinensis are most similar to those of the agamids and phrynosomatids examined to date. No spermatozoal autapomorphies for Crotaphytidae or Polychrotidae were found. The condition of having the intermitochondrial dense bodies arranged in regular incomplete rings is tentatively defined as a synapomorphy of Iguania (although modified in Chamaeleonidae). Spermatozoal ultrastructure offers no characters that justify the separation of Iguanidae (sensu lato) into several separate families.

Morphology of the spermatozoa of the iguanian lizards Uta stansburiana and Urosaurus ornatus (Squamata, Phrynosomatidae).

The spermatozoa of Uta stansburiana and Urosaurus ornatus show the following squamate autapomorphies: a single perforatorium extending anteriorly from the apical tip of the paracrystalline subacrosomal cone; the presence of an epinuclear electron lucent region; intermitochondrial dense bodies; and the fibrous sheath extending into the midpiece. The acrosome vesicle is flattened and concentrically zoned apically; basally it overlies a subacrosomal cone which invests the nuclear rostrum. A stopper-like perforatorial base plate, rounded nuclear shoulders and a basal nuclear fossa are present. The proximal centriole contains a density within its centre for approximately one half its length and lies at approximately 80 degrees to the distal centriole. The two central singlets of the axoneme extend into the short distal centriole. A peripheral dense fibre is associated with each of the nine triplets of the distal centriole, and the fibre continues posteriorly with each of the nine doublets of the axoneme. A central fibre is associated with the two central singlets. All fibres are absent or vestigial at the level of the annulus. Mitochondria are short sinuous with a maximum of eight seen in transverse section. Uta and Urosaurus sperm differ from each other in their arrangement of intermitochondrial dense bodies in two ways: 1) longitudinally, Uta has five incomplete 'rings' of dense bodies, whereas Urosaurus has only four such rings; 2) in cross section, each individual 'ring' of Uta may contain up to four irregularly spaced dense bodies, whereas Urosaurus contains a maximum of only two dense bodies. The sperm of Uta and Urosaurus show strong similarities to those of the agamids and polychrotids. No spermatozoal autapomorphies for the Phrynosomatidae were found.

Phylogeny of the monopisthocotylea and Polyopisthocotylea (Platyhelminthes) inferred from 28S rDNA sequences.

This study focuses on the phylogenetic relationships within the Polyopisthocotylea and Monopisthocotylea, two groups that are often grouped within the monogeneans, a group of disputed paraphyly. Phylogenetic analyses were conducted with multiple outgroups chosen according to two hypotheses, a paraphyletic Monogenea or a monophyletic Monogenea, and with three methods, namely maximum parsimony, neighbour joining and maximum likelihood. Sequences used were from the partial domain C1, full domain D1, and partial domain C2 (550 nucleotides, 209 unambiguously aligned sites) from the 28S ribosomal RNA gene for 16 species of monopisthocotyleans, 26 polyopisthocotyleans including six polystomatids, and other Platyhelminthes (61 species in total, 27 new sequences). Results were similar with outgroups corresponding to the two hypotheses. Within the Monopisthocotylea, relationships were: ¿[(Udonella, capsalids), monocotylids], (diplectanids, ancyrocephalids)¿; each of these families was found to be monophyletic and their monophyly was supported by high bootstrap values in neighbour joining and maximum parsimony. Within the Polyopisthocotylea, the polystomatids were the sister-group of all others. Among the latter, Hexabothrium, parasite of chondrichthyans, was the most basal, and the mazocraeids, mainly parasites of clupeomorph teleosts, were the sister-groups of all other studied polyopisthocotyleans, these, mainly parasites of euteleosts, being polytomous.

Sperm ultrastructure of six Australian hylid frogs from two genera (Litoria and Cyclorana): phylogenetic implications.

The spermatozoa of four fossorial (Litoria alboguttata, Cyclorana brevipes, Cyclorana novaehollandiae and Cyclorana cryptotis) and two non-fossorial australian hylid frogs (Litoria aurea and Litoria moorei) together with previously examined Litoria (Hylidae: Anura) are compared. In spermatozoal ultrastructure (in particular the structure of the sperm tail) Cyclorana includes species which appear derived (apomorphic) relative to non-fossorial species of Litoria while the fossorial L. alboguttata groups with Cyclorana. All hylid species examined here are united by the bufonoid synapomorphy of a conical subacrosomal cone consisting of separate sheaves and the eubufonoid synapomorphy of a mitochondrial sheath or collar separated by a cytoplasmic canal from the centriolar region and tail. Spermatozoal symplesiomorphies for the Eubufonoidea, seen in Litoria (with the exception of L. alboguttata), are the well developed thin undulating membrane with juxta-axonemal and axial fibre. L. alboguttata, C. novaehollandiae and C. brevipes appear monophyletic in the apomorphic modification of the undulating membrane as a thick, dense structure. In L. alboguttata and C. novaehollandiae this structure retains a swelling, at the free edge, homologized with the axial rod. C. brevipes has a further apomorphy as the undulating membrane forms a parallel-sided dense structure with no separate differentiation of an axial fibre. C. cryptotis, however, retains the plesiomorphic sperm tail with a thin undulating membrane, juxta-axonemal and axial fibre. That these differences in spermatozoal ultrastructure have phylogenetic significance is endorsed by the similarity of the fertilization biology of the species examined. On the basis of sperm ultrastructure three separate lineages are discerned within Cyclorana s. lat.: 1) C. cryptotis; 2) L. alboguttata and C. novaehollandiae; and 3) C. brevipes. The evidence of sperm ultrastructure, supported by previously published molecular, morphological and karyological data, clearly places Litoria alboguttata within the genus Cyclorana.

Phylogenetic analysis of the Monogenea and their relationships with Digenea and Eucestoda inferred from 28S rDNA sequences.

Platyhelminth phylogeny is controversial. Phylogenetic analyses of the partial domain C1 and the full domains D1 and C2 (358 nucleotides) from the 28S ribosomal RNA gene for 21 species from the Monogenea, Digenea, Cestoda, and, as the outgroup, Tricladida reveal major departures from prevailing theory. The Digenea and not the Monogenea (Monopisthocotylea and Polyopisthocotylea) form the sister group of the cestodes; the Monopisthocotylea and Polyopisthocotylea are each monophyletic, but the Monogenea do not form a monophylum; the sister group of the Digenea + Cestoda is the Polyopisthocotylea; and Monopisthocotylea are the sister group of all other parasitic flatworms.

Spermatozoal ultrastructure in three species of parrots (aves, Psittaciformes) and its phylogenetic implications.

BACKGROUND: DNA-DNA hybridization studies suggest that Psittaciformes are highly, but not the most, derived nonpasserines. Multilocus protein electrophoresis indicates that cockatoos (Cacatuinae) form a monophyletic lineage distant from the other Australo-Papuan psittacids (Psittacinae). METHODS: Transmission electron microscope procedures are applied to the spermatozoa of three parrots, in the Cacatuninae and Psittacinae, to investigate these relationships. RESULTS: Psittaciform sperm have the following characteristics: (1) conical acrosome vesicle; rodlike perforatorium; cylindrical, highly condensed nucleus; proximal and distal centriole embedded in dense material; elongate periaxonemal mitochondrial midpiece, (2) nine dense peripheral axonemal fibers (coarse fibers), (3) no fibrous sheath around the axoneme, (4) mitochondria with linear cristae, lacking intra- (or inter-) mitochondrial dense bodies, (5) restriction of the endonuclear perforatorial canal to the anterior region of the nucleus, (6) a short distal centriole, and (7) nucleus abutting on but not penetrating the acrosome. CONCLUSIONS: (1) These features are tetrapod symplesiomorphies, (2) is an amniote synapomorphy; the fibers differ from those of reptiles in being uniform in size, (3) loss of the fibrous sheath is an apomorphy known elsewhere only in columbiforms, (4) are apomorphies relative to basal aminiotes (Chelonia, Sphenodon, and Crocodilia), (5) is an apomorphic condition shared with other nonpasserines (galliforms and the white-naped crane) and crocodilians, (6) the latter taxa differ from parrots in a plesiomorphic elongation of the distal centriole, and (7) is a unique apomorphy of parrot sperm relative to other nonpasserines and reptiles. The short midpiece of N. hollandicus distinguishes this cacatuine from the two psittacines.

Sequence evolution and phylogenetic signal in control-region and cytochrome b sequences of rainbow fishes (Melanotaeniidae).

The nucleotide sequences of segments of the cytochrome b gene (351 bp), the tRNA(Pro) gene (49 bp), and the control region (approximately 313 bp) of mitochondrial DNA were obtained from 26 fish representing different populations and species of Melanotaenia and one species of Glossolepis, freshwater rainbow fishes confined to Australia and New Guinea. The purpose was to investigate relative rates and patterns of sequence evolution. Overall levels of divergence were similar for the cytochrome b and tRNA control-region sequences, both ranging from < 1% within subspecies to 15%-19% between genera. However, the patterns of sequence evolution differed. For the cytochrome b gene, transitions consistently exceeded transversions, the bias ranging from 4.2:1 to 2:1, depending on the level of sequence divergence. However, in the control-region sequence, a bias toward transitions (2:1) was observed only in comparisons between very similar sequences, and transversions outnumbered transitions in comparisons of divergent sequences. Graphic comparisons suggested that the control region was saturated for transitions at relatively low levels of sequence divergence but accumulated transversions at a greater rate than did the cytochrome b sequence. These distinct patterns of base substitution are associated with differences in A+T content, which is 70% for the tRNA control-region segment versus 50% for cytochrome b. A test for skewness in the distribution of lengths of random trees indicated that both segments contained phylogenetic signal. Parsimony analyses of the data from the two regions, with or without weighting schemes appropriate to the respective patterns of sequence evolution, identified the same five groupings of sequences, but the relationships among the groups differed. However, in most cases the branches uniting different combinations of groups were poorly supported, and the differences among topologies were insignificant. Considering the observed patterns of base substitution and the results of the phylogenetic analyses, we deduce that both the control region and cytochrome b are appropriate for population genetic studies but that the control region is less effective than cytochrome b for resolving relationships among divergent lineages of rainbow fishes.

Complex spermatozoon of the live-bearing half-beak, Hemirhamphodon pogonognathus (Bleeker): ultrastructural description (Euteleostei, Atherinomorpha, Beloniformes).

The spermatozoon of Hemirhamphodon pogonognathus shows modifications that are frequent though not obligate in internally fertilizing sperm, notably elongation of the nucleus and extension of the mitochondria of the midpiece as an elongate sheath around the proximal region of the axoneme. These similarities to poecilid and jenynsid sperm are considered homoplasic. As in the mature sperm of all but one investigated teleost, an acrosome is absent. The elongate, blade-shaped, electron-dense nucleus has a mean length of 3.2 microns; its basal implantation fossa, less than one-tenth of the length of the nucleus, houses the anterior half of the distal and only centriole (of triplet construction with satellite rays), a centriolar plug, and a mass connecting the centriole to the wall of the fossa. A unilateral putative centriole adjunct is present. The anterior region of the axoneme is surrounded by a mitochondrial sleeve, and internal to this, separated by a cisterna, by a submitochondrial sleeve. The mitochondrial sleeve unites posteriorly with the submitochondrial sleeve. Between the submitochondrial sleeve and the axoneme is a space, the cytoplasmic canal, that is open to the exterior posteriorly. The discrete, cristate mitochondria, in their sleeve, are unique in investigated atherinomorph sperm in being bilateral, grouped on only two opposing sides of the axoneme, with an arc-shaped "intermitochondrial link" between. The 9 + 2 flagellum is unique for the Animalia in having 23 radial subplasmalemmal rods, repeated longitudinally (periodicity 0.025 microns) in a quasicrystalline array. Internal fertilization is deduced to have arisen in the Exocoetoidei independently of that in the Cyprinidontiformes.

Phylogenetic relations between microbats, megabats and primates (Mammalia: Chiroptera and Primates).

We examine the paraphylectic hypothesis of bat origins, both in the light of previous discussions, and in the light of new evidence from our analyses of neurological traits and wing morphology. Megabats share with primates a variety of complex details in the organization of neural pathways that have not been found in any other mammalian group, particularly not in microbats. The features previously used to link microbats and megabats have been examined and found to be questionable bases for support of a monophyletic origin. In particular, morphological analyses of the musculoskeletal adaptations associated with the flight apparatus are consistent with two separate origins of the mammalian wing. Taken together, these analyses suggest that megabats evolved from an early branch of the primate lineage. This branch was comprised of moderate-sized, phytophagous gliders, of which the other living descendants are the dermopterans. Microbats, in contrast, probably evolved much earlier from small, agile insectivores whose forelimbs had long metacarpals in relation to their phalanges.

The spermatozoa of the polychaeta (Annelida): an ultrastructural review.

1. Polychaete sperm are divisible into ect-aquasperm, ent-aquasperm, and introsperm. 2. Ect-aquasperm are the commonest type of polychaete sperm and are considered plesiomorphic for the Polychaeta. Re-evolution of ect-aquasperm (as neo-aquasperm) is, nevertheless, tentatively hypothesized for some Sabellida. 3. In terms of ultrastructural studies of sperm in the investigated polychaete families, only ect-aquasperm have been demonstrated for 16 families; only ent-aquasperm for 3 families; ect- and ent-aquasperm for 3; ect- and intro-sperm for 2; ect-, ent- and intro-sperm for 1 family; and only introsperm for 11 families but investigations can only be regarded as preliminary. To date no family is known to have ent- and intro-sperm only. Sperm ultrastructure has yet to be examined in the orders Magelonida, Psammodrilida, Cossurida, Spintherida, Sternapsida, Flabelligerida and Fauvelopsida. 4. Much variation occurs in gross morphology, ultrastructure and configuration of the several components of ect-aquasperm: acrosome, nucleus, mitochondria, and centrioles and associated anchoring apparatus. A 9 + 2 axoneme is constant. 5. Group-specific sperm structure has been demonstrated for the Nereidae (chiefly ect-aquasperm), and for introsperm of the families Histriobdellidae, Questidae; Capitellidae, Spionidae and Protodrilidae. Species-specificity of all classes of spermatozoa is well established. 6. The very small size of ect-aquasperm is correlated with production of large numbers of sperm as an adaptation to broadcast spawning. Simplicity of structure may relate more to conservation of materials than to hydrodynamics. 7. Fertilization by ent-aquasperm requires fewer eggs than in external fertilization and is accompanied by a tendency to lecithotrophy. Elongation of the nucleus and development of asymmetry are seen in several of the few known examples of ent-aquasperm. Whether modifications are related to transfer or to other features, such as lecithotrophy, is uncertain. 8. Evident multiple origins of polychaete introsperm contraindicate their value in establishing relationship between families, in contrast with their utility in groups such as decapod crustacea. 9. At the intrafamilial level polychaete introsperm have taxonomic and phylogenetic value, as seen in the Spionidae, Capitellidae, and Histriobdellidae, and are distinctive of each of these and other families. 10. At higher taxonomic levels, the ultrastructure of the sperm of the oligochaetoid Questidae distinguishes this family from euclitellates, each class of which has its own distinctive subtype of the euclitellate introsperm. 11.(ABSTRACT TRUNCATED AT 400 WORDS)

ON THE PHYLOGENY AND HIGHER CLASSIFICATION OF THE OLIGOCHAETA.

Abstract- The 50 oligochaete taxa representing all families of "opisthoporous" oligochaetes (Aliuroididae, earthworms and aquatic "megadriles") together with two representatives of the Haplotaxidac and three examples of "microdiles" were subjected to cladistic analysis using the PAUP program. Sixty-eight characters used in the analyses were derived from a comprehensive range of somatic and genital systems. The optimal result, in terms of maximal number of characters and taxa and of parsimony, produced two trees (consistency index 0.362) differing only in the placement of the monotypic clade for the family Lumbriculidae. From a line originating from the presumed octogonadial ancestor, the following branches were derived, in sequence from the basal to most derived (new taxa asterisked): subclass Randiellata* (order Randiellida*, Randiellidae); subclass Tubificata* (order Tubificida, Tubincidae, Naididae computed and others not computed); subclass Lumbriculata* (order Lumbriculida, Lumbriculidae); superorder Haplotaxidea* (order Haplotaxida, Haplotaxidae); order Moniligastrida (Moniligastridae); suborder Alluroidina (Aliuroididae and Syngenodrilidae); cohort Aquamegadrili* (with, in succession, superfamilice Sparganophiloidea, Sparganophilidae; Biwadriloidea, Biwadrilidac, and Almoidca-Lutodrilidac and Almidae, including Criodrilus); superfamily Eudriloidea*, superfamily Lumbricoidea and, as the adelphotaxon of the latter, the superfamily Megascolecoidea. Intermediate nodes were given the following names, with the adelpholaxon through to the Megascolecoidea, M, in parentheses: subclass Diplotesticulata (Haplotaxidea-M); superorder Metagynopohora* (Moniligastrida-M); order Opisthopora (Alluroidina M); suborder Crassiclitellata* (Aquamegadrili - M); cohort Terrimegadrili* (Ocnerodriloidea M); unnamed (Eudriloidea M); unnamed (Lumbricoidea and Megascolecoidea). Recognition of the Randiellata, which alone were added intuitively and not computed, and the position of the Lumbriculata, are tentative. Location of the Lumbricoidea as the adelphotaxon of a restricted Megascoecloidea is heuristic, but the alternative depiction of lumbricoids in some analyses, as the adelphotaxon of an ocnerodrilid-eudrilid-megascolecoid clade (the conventional Megascolecoidea s. lat.), is not conclusively dismissed.