Lethal effects and ultrastructure of cellular uptake of ingested gold nanoparticles in the freshwater rotifer Brachionus calyciflorus (Monogononta: Brachionidae).

Much of the recent literature concerning the threat posed by anthropogenic microscopic pollution has focussed on marine organisms although freshwater environments face the same degree of pollution and therefore risk. Although several studies have documented the ingestion of nanoparticles (NPs) in species of the pelagic freshwater rotifer genus Brachionus, unambiguous evidence for its cellular uptake in this group remains lacking. We therefore used transmission electron microscopy (TEM) of ultrathin sections through the digestive tract of individuals of Brachionus calyciflorus exposed in vitro to citrate stabilized gold nanoparticles (AuCit NPs) in their culture medium to provide the first concrete evidence for the cellular uptake of NPs in rotifers, a group of organisms that comprise an important part of the zooplankton community. Using this method, AuCit NPs with average diameters of 8.5 ± 1.4 nm and 12.5 ± 1.5 nm could be localized clearly within large vacuoles within the stomach cells. Moreover, the occasional presence of pits containing AuCit NPs in the outer membranes of these cells hints that the particles are taken up by some form of endocytosis. In all cases, the ingestion of AuCit NPs showed lethal effects after only one day with virtually no individuals surviving more than two days of exposure. Combined with the TEM evidence above, we hypothesize that death might derive from some form of lysosomal overload. In total, our results document the potential threat that microscopic pollution also poses for freshwater organisms. Through this, we hope that additional emphasis in this context will be directed toward freshwater environments and the potential for such pollution both to enter as well as to move up the food chain via trophic transfer events.

A weighted taxonomic matrix key for species of the rotifer genus Synchaeta (Rotifera, Monogononta, Synchaetidae).

A new, weighted matrix identification key for 34 largely undisputed species of Synchaeta was created with the aim of providing comparable, detailed and diagnostic character sets for each species that can be applied to live and/or preserved specimens. As part of this process, 14 species of Synchaeta were intensively re-investigated with respect to their habitus and trophi morphology using binocular, light, and scanning electron microscopy, which, together with behavioural observations, revealed several new discriminating characters. Whenever possible, missing information for any character was added for the remaining species from the literature, with the two recently described species Synchaeta arcifera and Synchaeta squamadigitata being considered for the first time in an identification key. Beyond its completeness, our key has two distinct advantages. First, the characters are supported by detailed illustrations of their respective character states whenever possible to both simplify identification and minimize any uncertainty in the descriptions themselves. Second, the new approach of weighting the characters according to their reliability, robustness and/or ease of determination was employed. This latter approach is especially advantageous for soft-bodied rotifers such as species of Synchaeta, where, for example, several external characters can be influenced by preservation and are therefore less diagnostic or reliable. Although the key is as comprehensive as possible, information for many species remains missing for many characters, thereby highlighting the need for additional comprehensive and detailed species (re-)investigations within Synchaeta.

Evidence Supporting the Uptake and Genomic Incorporation of Environmental DNA in the "Ancient Asexual" Bdelloid Rotifer Philodina roseola.

Increasing evidence suggests that bdelloid rotifers regularly undergo horizontal gene transfer, apparently as a surrogate mechanism of genetic exchange in the absence of true sexual reproduction, in part because of their ability to withstand desiccation. We provide empirical support for this latter hypothesis using the bdelloid Philodina roseola, which we demonstrate to readily internalize environmental DNA in contrast to a representative monogonont rotifer (Brachionus rubens), which, like other monogononts, is facultative sexual and cannot withstand desiccation. In addition, environmental DNA that was more similar to the host DNA was retained more often and for a longer period of time. Indirect evidence (increased variance in the reproductive output of the untreated F1 generation) suggests that environmental DNA can be incorporated into the genome during desiccation and is thus heritable. Our observed fitness effects agree with sexual theory and also occurred when the animals were desiccated in groups (thereby acting as DNA donors), but not individually, indicating the mechanism could occur in nature. Thus, although DNA uptake and its genomic incorporation appears proximally related to anhydrobiosis in bdelloids, it might also facilitate accidental genetic exchange with closely related taxa, thereby maintaining higher levels of genetic diversity than is otherwise expected for this group of "ancient asexuals".

How does the 'ancient' asexual Philodina roseola (Rotifera: Bdelloidea) handle potential UVB-induced mutations?

Like other obligate asexuals, bdelloid rotifers are expected to suffer from degradation of their genomes through processes including the accumulation of deleterious mutations. However, sequence-based analyses in this regard remain inconclusive. Instead of looking for historical footprints of mutations in these ancient asexuals, we directly examined the susceptibility and ability to repair point mutations by the bdelloid Philodina roseola by inducing cyclobutane-pyrimidine dimers (CPDs) via exposure to UVB radiation (280-320 nm). For comparison, we performed analogous experiments with the facultative asexual monogonont rotifer Brachionus rubens. Different strategies were found for the two species. Philodina roseola appeared to shield itself from CPD induction through uncharacterized UV-absorbing compounds and, except for the genome reconstruction that occurs after desiccation, was largely unable to repair UVB-induced damage. By contrast, B. rubens was more susceptible to UVB irradiation, but could repair all induced damage in ~2 h. In addition, whereas UV irradiation had a significant negative impact on the reproductive output of P. roseola, and especially so after desiccation, that of B. rubens was unaffected. Although the strategy of P. roseola might suffice under natural conditions where UVB irradiation is less intense, the lack of any immediate CPD repair mechanisms in this species remains perplexing. It remains to be investigated how typical these results are for bdelloids as a group and therefore how reliant these animals are on desiccation-dependent genome repair to correct potential DNA damage given their obligate asexual lifestyle.

Ultrastructure and function of the mastax in Dicranophorus forcipatus (Rotifera: Monogononta).

Rotifers are characterized by a complex set of cuticularized jaw elements in the pharynx. The fine structure of the jaw elements has been the subject of SEM studies for some time, but only very limited information exists on the ultrastructure of the jaw elements and their function beyond taxonomic considerations. Drawing on SEM and TEM techniques, the present study presents a detailed analysis of the mastax in Dicranophorus forcipatus, a carnivorous monogonont rotifer species from freshwater habitats characterized by an extrusible, grasping jaw apparatus. Based on ultrathin serial sections, the jaw elements are reconstructed and, in total, nine paired and two unpaired muscles identified. Possibly homologous muscles in other rotifer species are discussed and functional considerations of the forcipate mastax are suggested.

Ultrastructure of the protonephridial system in Neodasys chaetonotoideus (Gastrotricha: Chaetonotida) and in the ground pattern of Gastrotricha.

The taxon Neodasys has a basal position within Gastrotricha. This makes it very interesting for phylogenetic considerations in this group. To complete the reconstruction of the nephridial system in the stem species of Gastrotricha started earlier, we have studied the whole protonephridial system of Neodasys chaetonotoideus by means of complete sets of ultrathin sections and TEM. In many characters, protonephridia of N. chaetonotoideus resemble those of macrodasyidan gastrotrich species. For example, each of the six protonephridia, arranged in three pairs, consists of three distinct cells that constitute the continuous protonephridial lumen. Especially, the terminal cell of the protonephridia of N. chaetonotoideus shows a striking pattern: The perforation of the filter region is a meandering cleft that is continuous with the seam of the enfolded lumen of that cell. With the results presented here and that of former TEM studies, we give a comprehensive idea of the excretory organs in the ground pattern of Gastrotricha. Moreover, we can elaborate on the hypothesized protonephridial system in the stem species of Bilateria. We suggest that a meandering filtration cleft is a feature of the ground pattern of the Bilateria.