Micro RNA in Semen/Urine from Non-Obstructive Azoospermia Patients as Biomarkers to Predict the Presence of Testicular Spermatozoa and Spermatogonia.

About half of testicular sperm extraction (TESE) procedures in men with non-obstructive azoospermia (NOA), including men with Klinefelter syndrome (KS), are unsuccessful. To avoid unnecessary invasive surgery, biomarkers for spermatozoa were studied. In addition, markers for spermatogonia in testis tissue were explored. This study aimed to find biomarkers in the semen and/or urine of NOA patients to predict the presence of spermatogonia in the testis. Differentially expressed miRNAs were identified (1) between samples from patients with and without a positive TESE procedure as well as (2) between TESE-negative patients with and without spermatogonia. A total of thirteen upregulated miRNAs (ten in seminal plasma and three in urine) were found in the TESE-negative/spermatogonia-positive group compared to the TESE-negative/spermatogonia-negative group. These miRNAs could be potential biomarkers for spermatogonia; however, more research is necessary to validate their predictive power.

On the genus Typhlopolycystis Karling, 1956 (Platyhelminthes, Kalyptorhynchia, Polycystididae), with data on the five known species and the description of eleven new species.

An overview is given of all representatives of the genus Typhlopolycystis Karling, 1956: the five known species (T. coeca Karling, 1956; T. mediterranea Brunet, 1965; T. coomansi Schockaert and Karling, 1975; T. schockaerti Karling, 1978 and T. rubra Noldt and Reise, 1985) and eleven new species: T. microphthalma n. sp. from S. France, T. fonsecai n. sp. and T. maikoni n. sp. from Brazil, T. norenburgi n. sp. and T. leasiae n.sp from the Pacific coast of Panama, T. pluvialiae n. sp. from Lanzarote (Canary Islands, Spain), T. riegeri n. sp. from Tanzania, T. tahitienis n. sp. from French Polynesia, T. australiensis n. sp. from South Australia, T. sarda n. sp. from Sardinia (Italy), the S. of France and from Lanzarote, and T. errata n. sp. from Sardinia (Italy). We consider T. limicola Schilke, 1970 as a species inquirenda of which no type material exists. Finally, some comments are given on the distribution of the genus of which representatives have been found in almost all places that have been sampled world wide.

Molecular characterization and species delimiting of plant-parasitic nematodes of the genus Pratylenchus from the penetrans group (Nematoda: Pratylenchidae).

Root-lesion nematodes of the genus Pratylenchus are an important pest parasitizing a wide range of vascular plants including several economically important crops. However, morphological diagnosis of the more than 100 species is problematic due to the low number of diagnostic features, high morphological plasticity and incomplete taxonomic descriptions. In order to employ barcoding based diagnostics, a link between morphology and species specific sequences has to be established. In this study, we reconstructed a multi-gene phylogeny of the Penetrans group using nuclear ribosomal and mitochondrial gene sequences. A combination of this phylogenetic framework with molecular species delineation analysis, population genetics, morphometric information and sequences from type location material allowed us to establish the species boundaries within the Penetrans group and as such clarify long-standing controversies about the taxonomic status of P. penetrans, P. fallax and P. convallariae. Our study also reveals a remarkable amount of cryptic biodiversity within the genus Pratylenchus confirming that identification on morphology alone can be inconclusive in this taxonomically confusing genus.

Integrative taxonomy of root-knot nematodes reveals multiple independent origins of mitotic parthenogenesis.

During sampling of several Coffea arabica plantations in Tanzania severe root galling, caused by a root-knot nematode was observed. From pure cultures, morphology and morphometrics of juveniles and females matched perfectly with Meloidogyne africana, whereas morphology of the males matched identically with those of Meloidogyne decalineata. Based on their Cox1 sequence, however, the recovered juveniles, females and males were confirmed to belong to the same species, creating a taxonomic conundrum. Adding further to this puzzle, re-examination of M. oteifae type material showed insufficient morphological evidence to maintain its status as a separate species. Consequently, M. decalineata and M. oteifae are synonymized with M. africana, which is herewith redescribed based on results of light and scanning electron microscopy, ribosomal and mitochondrial DNA sequences, isozyme electrophoresis, along with bionomic and cytogenetic features. Multi-gene phylogenetic analysis placed M. africana outside of the three major clades, together with M. coffeicola, M. ichinohei and M. camelliae. This phylogenetic position was confirmed by several morphological features, including cellular structure of the spermatheca, egg mass position, perineal pattern and head shape. Moreover, M. africana was found to be a polyphagous species, demonstrating that "early-branching" Meloidogyne spp. are not as oligophagous as had previously been assumed. Cytogenetic information indicates M. africana (2n = 21) and M. ardenensis (2n = 51-54) to be a triploid mitotic parthenogenetic species, revealing at least four independent origins of mitotic parthenogenesis within the genus Meloidogyne. Furthermore, M. mali (n = 12) was found to reproduce by amphimixis, indicating that amphimictic species with a limited number of chromosomes are widespread in the genus, potentially reflecting the ancestral state of the genus. The wide variation in chromosome numbers and associated changes in reproduction modes indicate that cytogenetic evolution played a crucial role in the speciation of root-knot nematodes and plant-parasitic nematodes in general.

An early record of Meloidogyne fallax from Ireland.

Root-knot nematodes, Meloidogyne spp., cause huge economic losses worldwide. Currently, three Meloidogyne spp. are present on the quarantine A2 list of EPPO, Meloidogyne chitwoodi, Meloidogyne fallax and Meloidogyne enterolobii. As a quarantine organism, Meloidogyne fallax has been detected in England and Northern Ireland on sport turf in 2011, and in England on leek in 2013. However, its presence in Ireland has probably been overlooked since 1965, when Mr. John F. Moore and Dr. Mary T. Franklin had detected a new Meloidogyne species for that time. While the relevant data was recorded and a preliminary manuscript describing the species was prepared but never submitted for publication, and together with the original slides, pictures and drawings, it was restudied recently. We compared the population of Irish Meloidogyne sp. to other similar Meloidogyne spp. Careful observation and comparison shows that it belongs to Meloidogyne fallax. The characters found to be common for Irish Meloidogyne sp. and Meloidogyne fallax are female stylet length (14.6 µm) with oval to rounded basal knobs, oval shaped perineal pattern with moderately high dorsal arch, slender stylet in males (18.5 µm) with set off and rounded basal knobs, slightly set off male head with one post-labial annule and incomplete transverse incisures, and second-stage juveniles with large and rounded stylet basal knobs, and a gradually tapering tail (46.9 µm) with a broadly rounded tip and a clearly delimitated smooth hyaline part sometimes marked by constrictions (12.9 µm). The host test and gall formation also correspond to Meloidogyne fallax. The identification could not be additionally supported by molecular analysis, as we were unable to extract DNA from the old permanent slides. Nevertheless, our study reveals that the Meloidogyne species detected in Ireland in 1965 belongs to Meloidogyne fallax.

Mitochondrial coding genome analysis of tropical root-knot nematodes (Meloidogyne) supports haplotype based diagnostics and reveals evidence of recent reticulate evolution.

The polyphagous parthenogenetic root-knot nematodes of the genus Meloidogyne are considered to be the most significant nematode pest in sub-tropical and tropical agriculture. Despite the crucial need for correct diagnosis, identification of these pathogens remains problematic. The traditionally used diagnostic strategies, including morphometrics, host-range tests, biochemical and molecular techniques, now appear to be unreliable due to the recently-suggested hybrid origin of root-knot nematodes. In order to determine a suitable barcode region for these pathogens nine quickly-evolving mitochondrial coding genes were screened. Resulting haplotype networks revealed closely related lineages indicating a recent speciation, an anthropogenic-aided distribution through agricultural practices, and evidence for reticulate evolution within M. arenaria. Nonetheless, nucleotide polymorphisms harbor enough variation to distinguish these closely-related lineages. Furthermore, completeness of lineage sorting was verified by screening 80 populations from widespread geographical origins and variable hosts. Importantly, our results indicate that mitochondrial haplotypes are strongly linked and consistent with traditional esterase isozyme patterns, suggesting that different parthenogenetic lineages can be reliably identified using mitochondrial haplotypes. The study indicates that the barcode region Nad5 can reliably identify the major lineages of tropical root-knot nematodes.

The first multi-gene phylogeny of the Macrostomorpha sheds light on the evolution of sexual and asexual reproduction in basal Platyhelminthes.

The Macrostomorpha-an early branching and species-rich clade of free-living flatworms-is attracting interest because it contains Macrostomum lignano, a versatile model organism increasingly used in evolutionary, developmental, and molecular biology. We elucidate the macrostomorphan molecular phylogeny inferred from both nuclear (18S and 28S rDNA) and mitochondrial (16S rDNA and COI) marker genes from 40 representatives. Although our phylogeny does not recover the Macrostomorpha as a statistically supported monophyletic grouping, it (i) confirms many taxa previously proposed based on morphological evidence, (ii) permits the first placement of many families and genera, and (iii) reveals a number of unexpected placements. Specifically, Myozona and Bradynectes are outside the three classic families (Macrostomidae, Microstomidae and Dolichomacrostomidae) and the asexually fissioning Myomacrostomum belongs to a new subfamily, the Myozonariinae nov. subfam. (Dolichomacrostomidae), rather than diverging early. While this represents the first evidence for asexuality among the Dolichomacrostomidae, we show that fissioning also occurs in another Myozonariinae, Myozonaria fissipara nov. sp. Together with the placement of the (also fissioning) Microstomidae, namely as the sister taxon of Dolichomacrostomidae, this suggests that fissioning is not basal within the Macrostomorpha, but rather restricted to the new taxon Dolichomicrostomida (Dolichomacrostomidae+Microstomidae). Furthermore, our phylogeny allows new insights into the evolution of the reproductive system, as ancestral state reconstructions reveal convergent evolution of gonads, and male and female genitalia. Finally, the convergent evolution of sperm storage organs in the female genitalia appears to be linked to the widespread occurrence of hypodermic insemination among the Macrostomorpha.

A new genus with six new species of Typhlopolycystidinae Evdonin, 1977 (Platyhelminthes, Kalyptorhynchia, Polycystididae).

Five new species of the new taxon Brunetorhynchus n. gen. are described: B. deconincki n. sp., B. microstylis n. sp. and B. complicatus n. sp. are from the Mediterranean, B. canariensis n. sp. is from the Canary Island Lanzarote, B. cannoni n. sp. is from the Australian East coast and one species from the Galapagos, formerly described as Limipolycystis spec., is transferred to the new genus as B. dubius n. sp.. As in Limipolycystis, these species have a single stylet, (accessory stylet type II), an accessory secretion vesicle (type II) and a prostate vesicle (type III) in the male atrium, although the latter vesicle is absent in some species. Unlike the species of Limipolycystis, where the seminal receptacle is a sclerotized tubule, the species of the new taxon have a pear-shaped seminal receptacle on the oviduct.