Identification of antiparasitic drug targets using a multi-omics workflow in the acanthocephalan model.

BACKGROUND: With the expansion of animal production, parasitic helminths are gaining increasing economic importance. However, application of several established deworming agents can harm treated hosts and environment due to their low specificity. Furthermore, the number of parasite strains showing resistance is growing, while hardly any new anthelminthics are being developed. Here, we present a bioinformatics workflow designed to reduce the time and cost in the development of new strategies against parasites. The workflow includes quantitative transcriptomics and proteomics, 3D structure modeling, binding site prediction, and virtual ligand screening. Its use is demonstrated for Acanthocephala (thorny-headed worms) which are an emerging pest in fish aquaculture. We included three acanthocephalans (Pomphorhynchus laevis, Neoechinorhynchus agilis, Neoechinorhynchus buttnerae) from four fish species (common barbel, European eel, thinlip mullet, tambaqui). RESULTS: The workflow led to eleven highly specific candidate targets in acanthocephalans. The candidate targets showed constant and elevated transcript abundances across definitive and accidental hosts, suggestive of constitutive expression and functional importance. Hence, the impairment of the corresponding proteins should enable specific and effective killing of acanthocephalans. Candidate targets were also highly abundant in the acanthocephalan body wall, through which these gutless parasites take up nutrients. Thus, the candidate targets are likely to be accessible to compounds that are orally administered to fish. Virtual ligand screening led to ten compounds, of which five appeared to be especially promising according to ADMET, GHS, and RO5 criteria: tadalafil, pranazepide, piketoprofen, heliomycin, and the nematicide derquantel. CONCLUSIONS: The combination of genomics, transcriptomics, and proteomics led to a broadly applicable procedure for the cost- and time-saving identification of candidate target proteins in parasites. The ligands predicted to bind can now be further evaluated for their suitability in the control of acanthocephalans. The workflow has been deposited at the Galaxy workflow server under the URL tinyurl.com/yx72rda7 .

Protein speciation is likely to increase the chance of proteins to be determined in 2-DE/MS.

Multiple spotting due to protein speciation might increase a protein's chance of being captured in a random selection of 2-DE spots. We tested this expectation in new (PXD015649) and previously published 2-DE/MS data of porcine and human tissues. For comparison, we included bottom-up proteomics studies (BU-LC/MS) of corresponding biological materials. Analyses of altogether ten datasets proposed that amino acid modification fosters multispotting in 2-DE. Thus, the number of 2-DE spots containing a particular protein more tightly associated with a peptide diversity measure accounting for amino acid modification than with an alternative one disregarding it. Furthermore, every 11th amino acid was a post-translational modification candidate site in 2-DE/MS proteins, whereas in BU-LC/MS proteins this was merely the case in every 21st amino acid. Alternative splicing might contribute to multispotting, since genes encoding 2-DE/MS proteins were found to have on average about 0.3 more transcript variants than their counterparts from BU-LC/MS studies. Correspondingly, resolution completeness as estimated from the representation of transcript variant-rich genes was higher in 2-DE/MS than BU-LC/MS datasets. These findings suggest that the ability to resolve proteomes down to protein species can lead to enrichment of multispotting proteins in 2-DE/MS. Low sensitivity of stains and MS instruments appears to enhance this effect.

Genome-Wide Association Screening Determines Peripheral Players in Male Fertility Maintenance.

Deciphering the functional relationships of genes resulting from genome-wide screens for polymorphisms that are associated with phenotypic variations can be challenging. However, given the common association with certain phenotypes, a functional link should exist. We have tested this prediction in newly sequenced exomes of altogether 100 men representing different states of fertility. Fertile subjects presented with normal semen parameters and had naturally fathered offspring. In contrast, infertile probands were involuntarily childless and had reduced sperm quantity and quality. Genome-wide association study (GWAS) linked twelve non-synonymous single-nucleotide polymorphisms (SNPs) to fertility variation between both cohorts. The SNPs localized to nine genes for which previous evidence is in line with a role in male fertility maintenance: ANAPC1, CES1, FAM131C, HLA-DRB1, KMT2C, NOMO1, SAA1, SRGAP2, and SUSD2. Most of the SNPs residing in these genes imply amino acid exchanges that should only moderately affect protein functionality. In addition, proteins encoded by genes from present GWAS occupied peripheral positions in a protein-protein interaction network, the backbone of which consisted of genes listed in the Online Mendelian Inheritance in Man (OMIM) database for their implication in male infertility. Suggestive of an indirect impact on male fertility, the genes focused were indeed linked to each other, albeit mediated by other interactants. Thus, the chances of identifying a central player in male infertility by GWAS could be limited in general. Furthermore, the SNPs determined and the genes containing these might prove to have potential as biomarkers in the diagnosis of male fertility.

Genomics and transcriptomics of epizoic Seisonidea (Rotifera, syn. Syndermata) reveal strain formation and gradual gene loss with growing ties to the host.

BACKGROUND: Seisonidea (also Seisonacea or Seisonidae) is a group of small animals living on marine crustaceans (Nebalia spec.) with only four species described so far. Its monophyletic origin with mostly free-living wheel animals (Monogononta, Bdelloidea) and endoparasitic thorny-headed worms (Acanthocephala) is widely accepted. However, the phylogenetic relationships inside the Rotifera-Acanthocephala clade (Rotifera sensu lato or Syndermata) are subject to ongoing debate, with consequences for our understanding of how genomes and lifestyles might have evolved. To gain new insights, we analyzed first drafts of the genome and transcriptome of the key taxon Seisonidea. RESULTS: Analyses of gDNA-Seq and mRNA-Seq data uncovered two genetically distinct lineages in Seison nebaliae Grube, 1861 off the French Channel coast. Their mitochondrial haplotypes shared only 82% sequence identity despite identical gene order. In the nuclear genome, distinct linages were reflected in different gene compactness, GC content and codon usage. The haploid nuclear genome spans ca. 46 Mb, of which 96% were reconstructed. According to ~ 23,000 SuperTranscripts, gene number in S. nebaliae should be within the range published for other members of Rotifera-Acanthocephala. Consistent with this, numbers of metazoan core orthologues and ANTP-type transcriptional regulatory genes in the S. nebaliae genome assembly were between the corresponding numbers in the other assemblies analyzed. We additionally provide evidence that a basal branching of Seisonidea within Rotifera-Acanthocephala could reflect attraction to the outgroup. Accordingly, rooting via a reconstructed ancestral sequence led to monophyletic Pararotatoria (Seisonidea+Acanthocephala) within Hemirotifera (Bdelloidea+Pararotatoria). CONCLUSION: Matching genome/transcriptome metrics with the above phylogenetic hypothesis suggests that a haploid nuclear genome of about 50 Mb represents the plesiomorphic state for Rotifera-Acanthocephala. Smaller genome size in S. nebaliae probably results from subsequent reduction. In contrast, genome size should have increased independently in monogononts as well as bdelloid and acanthocephalan stem lines. The present data additionally indicate a decrease in gene repertoire from free-living to epizoic and endoparasitic lifestyles. Potentially, this reflects corresponding steps from the root of Rotifera-Acanthocephala via the last common ancestors of Hemirotifera and Pararotatoria to the one of Acanthocephala. Lastly, rooting via a reconstructed ancestral sequence may prove useful in phylogenetic analyses of other deep splits.

Intra-Protein Coevolution Is Increasingly Functional with Greater Proximity to Fertilization.

Intramolecular coevolution of amino acid sites has repeatedly been studied to improve predictions on protein structure and function. Thereby, the focus was on bacterial proteins with available crystallographic data. However, intramolecular coevolution has not yet been compared between protein sets along a gradient of functional proximity to fertilization. This is especially true for the potential effect of external selective forces on intraprotein coevolution. In this study, we investigated both aspects in equally sized sets of mammalian proteins representing spermatozoa, testis, entire body, and liver. For coevolutionary analyses, we derived the proportion of covarying sites per protein from amino acid alignments of 10 mammalian orthologues each. In confirmation of the validity of our coevolution proxy, we found positive associations with the nonsynonymous or amino acid substitution rate in all protein sets. However, our coevolution proxy negatively correlated with the number of protein interactants (node degree) in male reproductive protein sets alone. In addition, a negative association of our coevolution proxy with protein hydrophobicity was significant in sperm proteins only. Accordingly, the restrictive effect of protein interactants was most pronounced in male reproductive proteins, and the tendency of sperm proteins to form internal structures decreased the more coevolutionary sites they had. Both aspects illustrate that the share of outward and thus functional coevolution increases with greater proximity to fertilization. We found this conclusion confirmed by additional comparisons within sperm proteins. Thus, sperm proteins with high hydrophobicity had the lowest proportions of covarying sites and, according to gene annotations, localized more frequently to internal cellular structures. They should therefore be less exposed to postcopulatory forms of sexual selection. Their counterparts with low hydrophobicity had larger proportions of covarying sites and more often resided at the cell membrane or were secreted. At the cellular level, they are thus closer to externally induced forces of postcopulatory selection which are known for their potential to increase substitution rates. In addition, we show that the intermediary status of the testicular protein set in correlation analyses is probably due to a special combination of reproductive and somatic involvements.

Fertility Relevance Probability Analysis Shortlists Genetic Markers for Male Fertility Impairment.

Impairment of male fertility is one of the major public health issues worldwide. Nevertheless, genetic causes of male sub- and infertility can often only be suspected due to the lack of reliable and easy-to-use routine tests. Yet, the development of a marker panel is complicated by the large quantity of potentially predictive markers. Actually, hundreds or even thousands of genes could have fertility relevance. Thus, a systematic method enabling a selection of the most predictive markers out of the many candidates is required. As a criterion for marker selection, we derived a gene-specific score, which we refer to as fertility relevance probability (FRP). For this purpose, we first categorized 2,753 testis-expressed genes as either candidate markers or non-candidates, according to phenotypes in male knockout mice. In a parallel approach, 2,502 genes were classified as candidate markers or non-candidates based on phenotypes in men. Subsequently, we conducted logistic regression analyses with evolutionary rates of genes (dN/dS), transcription levels in testis relative to other organs, and connectivity of the encoded proteins in a protein-protein interaction network as covariates. In confirmation of the procedure, FRP values showed the expected pattern, thus being overall higher in genes with known relevance for fertility than in their counterparts without corresponding evidence. In addition, higher FRP values corresponded with an increased dysregulation of protein abundance in spermatozoa of 37 men with normal and 38 men with impaired fertility. Present analyses resulted in a ranking of genes according to their probable predictive power as candidate markers for male fertility impairment. Thus, AKAP4, TNP1, DAZL, BRDT, DMRT1, SPO11, ZPBP, HORMAD1, and SMC1B are prime candidates toward a marker panel for male fertility impairment. Additional candidate markers are DDX4, SHCBP1L, CCDC155, ODF1, DMRTB1, ASZ1, BOLL, FKBP6, SLC25A31, PRSS21, and RNF17. FRP inference additionally provides clues for potential new markers, thereunder TEX37 and POU4F2. The results of our logistic regression analyses are freely available at the PreFer Genes website (https://prefer-genes.uni-mainz.de/).

Impact of biotic interactions on the survival of emerging pathogen Acinetobacter baumannii in aquatic media.

Acinetobacter baumannii is an opportunistic pathogen causing infections in immunocompromised patients. Recent studies recorded its persistence in a variety of abiotic conditions, but data regarding the biotic interactions with other microorganisms are limited. The aim was to assess the interaction of clinically relevant A. baumannii with common faecal bacteria Escherichia coli and Enterococcus faecium. Additionally, the interaction with a bdelloid rotifer Adineta vaga as a potential agent for biological control of A. baumannii was examined. Experiments were conducted in nutrient-poor spring water (SW) and nutrient-rich diluted nutrient broth (DNB) at 22 °C. A. baumannii coexisted with E. coli and E. faecium in both media, suggesting the absence of inter-bacterial competition in long-term survival. No difference in the survival of pandrug-resistant, extensively drug-resistant or antibiotic sensitive isolates of A. baumannii was observed. Rotifers contributed to the removal of all tested bacteria, particularly in SW. Rotifers were able to remove 5.5 ± 1.3 log CFU/mL of A. baumannii in SW and 3.5 ± 1.7 log CFU/mL in DNB. Additionally, no intracellular growth of A. baumannii inside A. vaga was detected. In wastewater treatment plants and drinking water facilities, grazing by rotifers might be useful for the removal of emerging human pathogens such as A. baumannii from water.

Correlates of evolutionary rates in the murine sperm proteome.

BACKGROUND: Protein-coding genes expressed in sperm evolve at different rates. To gain deeper insight into the factors underlying this heterogeneity we examined the relative importance of a diverse set of previously described rate correlates in determining the evolution of murine sperm proteins. RESULTS: Using partial rank correlations we detected several major rate indicators: Phyletic gene age, numbers of protein-protein interactions, and survival essentiality emerged as particularly important rate correlates in murine sperm proteins. Tissue specificity, numbers of paralogs, and untranslated region lengths also correlate significantly with sperm genes' evolutionary rates, albeit to a lesser extent. Multifunctionality, coding sequence or average intron lengths, and mean expression level have insignificant or virtually no independent effects on evolutionary rates in murine sperm genes. Gene ontology enrichment analyses of three equally sized murine sperm protein groups classified based on their evolutionary rates indicate strongest sperm-specific functional specialization in the most quickly evolving gene class. CONCLUSIONS: We propose a model according to which slowly evolving murine sperm proteins tend to be constrained by factors such as survival essentiality, network connectivity, and/or broad expression. In contrast, evolutionary change may arise especially in less constrained sperm proteins, which might, moreover, be prone to specialize to reproduction-related functions. Our results should be taken into account in future studies on rate variations of reproductive genes.

Organization and evolution of the proboscis musculature in avian parasites of the genus Apororhynchus (Acanthocephala: Apororhynchida).

The highly enlarged proboscis in adult thorny-headed worms of the genus Apororhynchus suggests that its inner organization might be specialized as well. However, what kind of changes occurred in the stem line of monogeneric Apororhynchida is widely unknown and there are different conceptions regarding the presence/absence of several muscles. To expand our knowledge on this topic, I examined ethanol-fixed specimens, whole mounts, and semi-thin sections of three Apororhynchus species using the light microscope. Incorporation of previously published data increased the overall sample to five out of six Apororhynchus species known to date. Combined data suggest that Apororhynchida kept the full set of muscles which already evolved in the stem line of Acanthocephala: proboscis receptacle, a receptacle surrounding muscle (receptacle protrusor), retinacula, neck retractor, proboscis and receptacle retractors, circular and longitudinal musculature under the metasomal tegument, and a single muscular layer beneath the proboscis wall. However, especially proboscis receptacle and receptacle protrusor underwent considerable re-organization in the apororhynchid stem line: both muscles are subdivided into sail-like strands extending from the cerebral ganglion to the proboscis wall. This reorganization reflects that the two muscles still suspend the cerebral ganglion but are not implicated in the eversion of the proboscis. Spatially separated subtegumental longitudinal muscle cords and a sphincter at the posterior proboscis margin could be additional apomorphies of Apororhynchida. Finally, lack of a muscle plate, a midventral longitudinal muscle, and of lateral receptacle flexors and the absence of an apical sensory organ indicate a basally branching position of Apororhynchida relative to other Archiacanthocephala.

Evolutionary anatomy of the muscular apparatus involved in the anchoring of Acanthocephala to the intestinal wall of their vertebrate hosts.

Different conceptions exist regarding structure, function, and evolution of the muscles that move the acanthocephalan presoma, including the proboscis, i.e., the usually hooked hold-fast anchoring these endoparasites to the intestinal wall of their vertebrate definitive hosts. In order to clarify the unresolved issues, we carried out a light microscopic analysis of series of semi-thin sections and whole mounts representing the three traditional acanthocephalan classes: Archiacanthocephala (Macracanthorhynchus hirudinaceus), Eoacanthocephala (Paratenuisentis ambiguus, Tenuisentis niloticus), and Palaeacanthocephala (Acanthocephalus anguillae, Echinorhynchus truttae, Pomphorhynchus laevis, Corynosoma sp.). Combining our data with published light, transmission electron, and scanning electron microscopic data, we demonstrate that receptacle protrusor and proboscis receptacle in Archi- and Eoacanthocephala are homologous to the outer and inner wall of the proboscis receptacle in Palaeacanthocephala. Besides the proboscis receptacle and a "surrounding muscle," the last common ancestor of Acanthocephala presumably possessed a proboscis retractor, receptacle retractor, neck retractor (continuous with lemnisci compressors), and retinacula. These muscles most probably evolved in the acanthocephalan stem line. Moreover, the last common ancestor of Acanthocephala presumably possessed only a single layer of muscular cords under the presomal tegument while the metasomal body wall had circular and longitudinal strands. Two lateral receptacle flexors (also lateral receptacle protrusors), an apical muscle plate (surrounding one or two apical sensory organs), a midventral longitudinal muscle, and the differentiation of longitudinal body wall musculature at the base of the proboscis probably emerged within Archiacanthocephala. All muscles have a common organization principle: a peripheral layer of contractile filaments encloses the cytoplasm.

Phylogeny of Syndermata (syn. Rotifera): Mitochondrial gene order verifies epizoic Seisonidea as sister to endoparasitic Acanthocephala within monophyletic Hemirotifera.

A monophyletic origin of endoparasitic thorny-headed worms (Acanthocephala) and wheel-animals (Rotifera) is widely accepted. However, the phylogeny inside the clade, be it called Syndermata or Rotifera, has lacked validation by mitochondrial (mt) data. Herein, we present the first mt genome of the key taxon Seison and report conflicting results of phylogenetic analyses: while mt sequence-based topologies showed monophyletic Lemniscea (Bdelloidea+Acanthocephala), gene order analyses supported monophyly of Pararotatoria (Seisonidea+Acanthocephala) and Hemirotifera (Bdelloidea+Pararotatoria). Sequence-based analyses obviously suffered from substitution saturation, compositional bias, and branch length heterogeneity; however, we observed no compromising effects in gene order analyses. Moreover, gene order-based topologies were robust to changes in coding (genes vs. gene pairs, two-state vs. multistate, aligned vs. non-aligned), tree reconstruction methods, and the treatment of the two monogonont mt genomes. Thus, mt gene order verifies seisonids as sister to acanthocephalans within monophyletic Hemirotifera, while deviating results of sequence-based analyses reflect artificial signal. This conclusion implies that the complex life cycle of extant acanthocephalans evolved from a free-living state, as retained by most monogononts and bdelloids, via an epizoic state with a simple life cycle, as shown by seisonids. Hence, Acanthocephala represent a rare example where ancestral transitional stages have counterparts amongst the closest relatives.

Platyzoan paraphyly based on phylogenomic data supports a noncoelomate ancestry of spiralia.

Based on molecular data three major clades have been recognized within Bilateria: Deuterostomia, Ecdysozoa, and Spiralia. Within Spiralia, small-sized and simply organized animals such as flatworms, gastrotrichs, and gnathostomulids have recently been grouped together as Platyzoa. However, the representation of putative platyzoans was low in the respective molecular phylogenetic studies, in terms of both, taxon number and sequence data. Furthermore, increased substitution rates in platyzoan taxa raised the possibility that monophyletic Platyzoa represents an artifact due to long-branch attraction. In order to overcome such problems, we employed a phylogenomic approach, thereby substantially increasing 1) the number of sampled species within Platyzoa and 2) species-specific sequence coverage in data sets of up to 82,162 amino acid positions. Using established and new measures (long-branch score), we disentangled phylogenetic signal from misleading effects such as long-branch attraction. In doing so, our phylogenomic analyses did not recover a monophyletic origin of platyzoan taxa that, instead, appeared paraphyletic with respect to the other spiralians. Platyhelminthes and Gastrotricha formed a monophylum, which we name Rouphozoa. To the exclusion of Gnathifera, Rouphozoa and all other spiralians represent a monophyletic group, which we name Platytrochozoa. Platyzoan paraphyly suggests that the last common ancestor of Spiralia was a simple-bodied organism lacking coelomic cavities, segmentation, and complex brain structures, and that more complex animals such as annelids evolved from such a simply organized ancestor. This conclusion contradicts alternative evolutionary scenarios proposing an annelid-like ancestor of Bilateria and Spiralia and several independent events of secondary reduction.

Mating systems and protein-protein interactions determine evolutionary rates of primate sperm proteins.

To assess the relative impact of functional constraint and post-mating sexual selection on sequence evolution of reproductive proteins, we examined 169 primate sperm proteins. In order to recognize potential genome-wide trends, we additionally analysed a sample of altogether 318 non-reproductive (brain and postsynaptic) proteins. Based on cDNAs of eight primate species (Anthropoidea), we observed that pre-mating sperm proteins engaged in sperm composition and assembly show significantly lower incidence of site-specific positive selection and overall lower non-synonymous to synonymous substitution rates (dN/dS) across sites as compared with post-mating sperm proteins involved in capacitation, hyperactivation, acrosome reaction and fertilization. Moreover, database screening revealed overall more intracellular protein interaction partners in pre-mating than in post-mating sperm proteins. Finally, post-mating sperm proteins evolved at significantly higher evolutionary rates than pre-mating sperm and non-reproductive proteins on the branches to multi-male breeding species, while no such increase was observed on the branches to unimale and monogamous species. We conclude that less protein-protein interactions of post-mating sperm proteins account for lowered functional constraint, allowing for stronger impact of post-mating sexual selection, while the opposite holds true for pre-mating sperm proteins. This pattern is particularly strong in multi-male breeding species showing high female promiscuity.

New Insights on the Male and Female Reproductive Organs of Centrorhynchus globocaudatus (Acanthocephala), Intestinal Parasite of Birds of Prey.

Acanthocephalans are dioecious parasites that gain sexual maturity in the alimentary canal of their definitive hosts (gnathostome vertebrates). This initial survey by light and transmission electron microscopy was conducted on the functional organization of the ovarian balls and uterine bell in mature females and on Saefftigen's pouch and the copulatory bursa in males. We studied these structures via the example of Centrorhynchus globocaudatus (Palaeacanthocephala) in Falco tinnunculus and Buteo buteo, from the Province of Ferrara (Northern Italy). Our study confirms that the ovarian balls have surface microvilli and consist of a multinucleate supporting syncytium and a cellular region with oogonial syncytium, single germ cells, zygotes, and shelled eggs. Germ cells are embedded in the supporting syncytium. The ultrastructural features of these components and data on fertilization, shell formation, and release from the ovarian ball, alongside insights into the likely egg sorting function of the uterine bell, are provided. We also present light and electron microscopy observations of Saefftigen's pouch and a suggestion regarding its hydrostatic functioning in the eversion of the copulatory bursa.

Evolutionary conservation of mammalian sperm proteins associates with overall, not tyrosine, phosphorylation in human spermatozoa.

We investigated possible associations between sequence evolution of mammalian sperm proteins and their phosphorylation status in humans. As a reference, spermatozoa from three normozoospermic men were analyzed combining two-dimensional gel electrophoresis, immunoblotting, and mass spectrometry. We identified 99 sperm proteins (thereof 42 newly described) and determined the phosphorylation status for most of them. Sequence evolution was studied across six mammalian species using nonsynonymous/synonymous rate ratios (dN/dS) and amino acid distances. Site-specific purifying selection was assessed employing average ratios of evolutionary rates at phosphorylated versus nonphosphorylated amino acids (α). According to our data, mammalian sperm proteins do not show statistically significant sequence conservation difference, no matter if the human ortholog is a phosphoprotein with or without tyrosine (Y) phosphorylation. In contrast, overall phosphorylation of human sperm proteins, i.e., phosphorylation at serine (S), threonine (T), and/or Y residues, associates with above-average conservation of sequences. Complementary investigations suggest that numerous protein-protein interactants constrain sequence evolution of sperm phosphoproteins. Although our findings reject a special relevance of Y phosphorylation for sperm functioning, they still indicate that overall phosphorylation substantially contributes to proper functioning of sperm proteins. Hence, phosphorylated sperm proteins might be considered as prime candidates for diagnosis and treatment of reduced male fertility.

Platyzoan mitochondrial genomes.

Platyzoa is a putative lophotrochozoan (spiralian) subtaxon within the protostome clade of Metazoa, comprising a range of biologically diverse, mostly small worm-shaped animals. The monophyly of Platyzoa, the relationships between the putative subgroups Platyhelminthes, Gastrotricha and Gnathifera (the latter comprising at least Gnathostomulida, "Rotifera" and Acanthocephala) as well as some aspects of the internal phylogenies of these subgroups are highly debated. Here we review how complete mitochondrial (mt) genome data contribute to these debates. We highlight special features of the mt genomes and discuss problems in mtDNA phylogenies of the clade. Mitochondrial genome data seem to be insufficient to resolve the position of the platyzoan clade within the Spiralia but can help to address internal phylogenetic questions. The present review includes a tabular survey of all published platyzoan mt genomes.

Phylogenetic analyses of endoparasitic Acanthocephala based on mitochondrial genomes suggest secondary loss of sensory organs.

The metazoan taxon Syndermata (Monogononta, Bdelloidea, Seisonidea, Acanthocephala) comprises species with vastly different lifestyles. The focus of this study is on the phylogeny within the syndermatan subtaxon Acanthocephala (thorny-headed worms, obligate endoparasites). In order to investigate the controversially discussed phylogenetic relationships of acanthocephalan subtaxa we have sequenced the mitochondrial (mt) genomes of Echinorhynchus truttae (Palaeacanthocephala), Paratenuisentis ambiguus (Eoacanthocephala), Macracanthorhynchus hirudinaceus (Archiacanthocephala), and Philodina citrina (Bdelloidea). In doing so, we present the largest molecular phylogenetic dataset so far for this question comprising all major subgroups of Acanthocephala. Alongside with publicly available mt genome data of four additional syndermatans as well as 18 other lophotrochozoan (spiralian) taxa and one outgroup representative, the derived protein-coding sequences were used for Maximum Likelihood as well as Bayesian phylogenetic analyses. We achieved entirely congruent results, whereupon monophyletic Archiacanthocephala represent the sister taxon of a clade comprising Eoacanthocephala and monophyletic Palaeacanthocephala (Echinorhynchida). This topology suggests the secondary loss of lateral sensory organs (sensory pores) within Palaeacanthocephala and is further in line with the emergence of apical sensory organs in the stem lineage of Archiacanthocephala.

The human sperm proteome-Toward a panel for male fertility testing.

BACKGROUND: Although male factor accounts for 40%-50% of unintended childlessness, we are far from fully understanding the detailed causes. Usually, affected men cannot even be provided with a molecular diagnosis. OBJECTIVES: We aimed at a higher resolution of the human sperm proteome for better understanding of the molecular causes of male infertility. We were particularly interested in why reduced sperm count decreases fertility despite many normal-looking spermatozoa and which proteins might be involved. MATERIAL AND METHODS: Applying mass spectrometry analysis, we qualitatively and quantitatively examined the proteomic profiles of spermatozoa from 76 men differing in fertility. Infertile men had abnormal semen parameters and were involuntarily childless. Fertile subjects exhibited normozoospermia and had fathered children without medical assistance. RESULTS: We discovered proteins from about 7000 coding genes in the human sperm proteome. These were mainly known for involvements in cellular motility, response to stimuli, adhesion, and reproduction. Numbers of sperm proteins showing at least threefold deviating abundances increased from oligozoospermia (N = 153) and oligoasthenozoospermia (N = 154) to oligoasthenoteratozoospermia (N = 368). Deregulated sperm proteins primarily engaged in flagellar assembly and sperm motility, fertilization, and male gametogenesis. Most of these participated in a larger network of male infertility genes and proteins. DISCUSSION: We expose 31 sperm proteins displaying deviant abundances under infertility, which already were known before to have fertility relevance, including ACTL9, CCIN, CFAP47, CFAP65, CFAP251 (WDR66), DNAH1, and SPEM1. We propose 18 additional sperm proteins with at least eightfold differential abundance for further testing of their diagnostic potential, such as C2orf16, CYLC1, SPATA31E1, SPATA31D1, SPATA48, EFHB (CFAP21), and FAM161A. CONCLUSION: Our results shed light on the molecular background of the dysfunctionality of the fewer spermatozoa produced in oligozoospermia and syndromes including it. The male infertility network presented may prove useful in further elucidating the molecular mechanism of male infertility.

Testes-specific hemoglobins in Drosophila evolved by a combination of sub- and neofunctionalization after gene duplication.

BACKGROUND: For a long time the presence of respiratory proteins in most insects has been considered unnecessary. However, in recent years it has become evident that globins belong to the standard repertoire of the insect genome. Like most other insect globins, the glob1 gene of Drosophila melanogaster displays a conserved expression pattern in the tracheae, the fat body and the Malpighian tubules. RESULTS: Here we show that the recently discovered D. melanogaster globin genes glob2 and glob3 both display an unusual male-specific expression in the reproductive tract during spermatogenesis. Both paralogs are transcribed at equivalent mRNA levels and largely overlap in their cellular expression patterns during spermatogenesis. Phylogenetic analyses showed that glob2 and glob3 reflect a gene duplication event that occurred in the ancestor of the Sophophora subgenus at least 40 million years ago. Therefore, flies of the Drosophila subgenus harbor only one glob2/3-like gene. CONCLUSIONS: Phylogenetic and sequence analyses indicate an evolution of the glob2 and glob3 duplicates by a combination of sub- and neofunctionalization. Considering their restricted, testes-specific expression, an involvement of both globins in alleviating oxidative stress during spermatogenesis is conceivable.

Sexual size dimorphism predicts rates of sequence evolution of SPerm Adhesion Molecule 1 (SPAM1, also PH-20) in monkeys, but not in hominoids (apes including humans).

Based on a dataset comprising coding DNA sequences of 23 anthropoid primates, we herein investigate if rates of sequence evolution of SPerm Adhesion Molecule1 (SPAM1, also PH-20), which participates in sperm-egg interaction, is lower in more sexually dimorphic species. For comparison, we analyze sequence evolution of apolipoproteinA-IV (APOA4) and apolipoprotein A-V (APOA5), which should evolve under less or even no sexual selection given their expression in blood, digestive tract, liver, and lungs. Regression analyses provides significant support for a negative dependence of SPAM1 derived branch-specific ratios of non-synonymous to synonymous substitution rates (dN/dS) on sexual size dimorphism (SSD) in a subsample comprising New World and Old World monkeys. We moreover observed a tendency for a positive correlation of substitution rates of SPAM1 with relative testes weight (RTW) and significantly lowered dN/dS estimates in uni-male and uni-male/multi-male breeding monkeys. Importantly, the pattern was not reproduced when analyzing partial APOA4 and APOA5 sequences. These findings illustrate that different levels of sperm competition, probably fueled by female cryptic choice, account for species-specific sequence evolution of SPAM1 in monkeys. Remarkably, present data do not support a correlation of species-specific sequence evolution of SPAM1 with sexual selection levels in hominoids (apes including humans). This can partly be ascribed to a relaxation of functional constraint of SPAM1 in some hominoid species. Additional factors confounding regression analyses specifically in hominoids might be higher levels of sperm competition than reflected by SSD and RTW in some species, a rather strong effect of female mate choice on paternity rates in others, and - in particular in humans - socio-cultural factors not measurable by SSD and RTW.