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ILARIA GIOVANNINI
Ricercatore t.d. art. 24 c. 3 lett. A
Dipartimento di Scienze della Vita sede ex-Biologia
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Pubblicazioni
2023
- Increasing temperature-driven changes in life history traits and gene expression of an Antarctic tardigrade species
[Articolo su rivista]
Giovannini, I.; Manfrin, C.; Greco, S.; Vincenzi, J.; Altiero, T.; Guidetti, R.; Giulianini, P.; Rebecchi, L.
abstract
The Antarctic region has been experiencing some of the planet's strongest climatic changes, including an expected increase of the land temperature. The potential effects of this warming trend will lead ecosystems to a risk of losing biodiversity. Antarctic mosses and lichens host different microbial groups, micro-arthropods and meiofaunal organisms (e.g., tardigrades, rotifers). The eutardigrade Acutuncus antarcticus is considered a model animal to study the effect of increasing temperature due to global warming on Antarctic terrestrial communities. In this study, life history traits and fitness of this species are analyzed by rearing specimens at two different and increasing temperatures (5 & DEG;C vs. 15 & DEG;C). Moreover, the first transcriptome analysis on A. antarcticus is performed, exposing adult animals to a gradual increase of temperature (5 & DEG;C, 10 & DEG;C, 15 & DEG;C, and 20 & DEG;C) to find differentially expressed genes under short- (1 day) and long-term (15 days) heat stress. Acutuncus antarcticus specimens reared at 5 & DEG;C live longer (maximum life span: 686 days), reach sexual maturity later, lay more eggs (which hatch in longer time and in lower percentage) compared with animals reared at 15 & DEG;C. The fitness decreases in animals belonging to the second generation at both rearing temperatures. The short-term heat exposure leads to significant changes at transcriptomic level, with 67 differentially expressed genes. Of these, 23 upregulated genes suggest alterations of mitochondrial activity and oxido-reductive processes, and two intrinsically disordered protein genes confirm their role to cope with heat stress. The long-term exposure induces alterations limited to 14 genes, and only one annotated gene is upregulated in response to both heat stresses. The decline in transcriptomic response after a long-term exposure indicates that the changes observed in the short-term are likely due to an acclimation response. Therefore, A. antarcticus could be able to cope with increasing temperature over time, including the future conditions imposed by global climate change.
2022
- Antioxidant Response during the Kinetics of Anhydrobiosis in Two Eutardigrade Species
[Articolo su rivista]
Giovannini, Ilaria; Corsetto, Paola Antonia; Altiero, Tiziana; Montorfano, Gigliola; Guidetti, Roberto; Rizzo, Angela Maria; Rebecchi, Lorena
abstract
Anhydrobiosis, a peculiar adaptive strategy existing in nature, is a reversible capability of organisms to tolerate a severe loss of their body water when their surrounding habitat is drying out. In the anhydrobiotic state, an organism lacks all dynamic features of living beings since an ongoing metabolism is absent. The depletion of water in the anhydrobiotic state increases the ionic concentration and the production of reactive oxygen species (ROS). An imbalance between the increased production of ROS and the limited action of antioxidant defences is a source of biomolecular damage and can lead to oxidative stress. The deleterious effects of oxidative stress were demonstrated in anhydrobiotic unicellular and multicellular organisms, which counteract the effects using efficient antioxidant machinery, mainly represented by ROS scavenger enzymes. To gain insights into the dynamics of antioxidant patterns during the kinetics of the anhydrobiosis of two tardigrade species, Paramacrobiotus spatialis and Acutuncus antarcticus, we investigated the activity of enzymatic antioxidants (catalase, superoxide dismutase, glutathione peroxidase, and glutathione reductase) and the amount of non-enzymatic antioxidants (glutathione) in the course of rehydration. In P. spatialis, the activity of catalase increases during dehydration and decreases during rehydration, whereas in A. antarcticus, the activity of superoxide dismutase decreases during desiccation and increases during rehydration. Genomic varieties, different habitats and geographical regions, different diets, and diverse evolutionary lineages may have led to the specialization of antioxidant strategies in the two species
2022
- Morphology and taxonomy of the genus Ramazzottius (Eutardigrada; Ramazzottiidae) with the integrative description of Ramazzottius kretschmanni sp. nov
[Articolo su rivista]
Guidetti, R.; Cesari, M.; Giovannini, I.; Ebel, C.; Forschler, M. I.; Rebecchi, L.; Schill, R. O.
abstract
The species of the genus Ramazzottius (Ramazzottiidae, Eutardigrada) are among the most common and widespread tardigrade species in the world. Most of the 28 Ramazzottius species have been described only with morphological characters which were most of the time represented only with drawings. The discovery of a new species of this genus in the Black Forest (Germany) provided the opportunity to compare this species with the type specimens of ten Ramazzottius species, to propose the status of species dubia for Ramazzottius edmondabouti, and through new photographs to elucidate the anatomy of animals and eggs (in particular of the head sensory regions, eye spots, buccal tube, ornamentations of the dorsal posterior cuticle, and morphology of egg processes). These thorough observations led to a better understanding of the diversity and evolution, not only of this cosmopolitan genus, but also of other eutardigrade genera. The new species Ramazzottius kretschmanni is described with an integrative approach integrating morphological (light and electron microscopy observations and morphometric data) and molecular (cox1 and ITS2 genes) data. The PTP and ASAP analyses confirmed the validity of the new species from a molecular point of view. The new species is morphologically similar to Ramazzottius oberhaeuseri, but is distinguishable by the smooth cuticle, the presence of a “cheek-like” area on the head, and the size of egg processes as well as different sequences of the molecular markers.
2022
- Phylogeny of the asexual lineage Murrayidae (Macrobiotoidea, Eutardigrada) with the description of Paramurrayon gen. nov. and Paramurrayon meieri sp. nov
[Articolo su rivista]
Guidetti, Roberto; Giovannini, Ilaria; Del Papa, Valeria; Ekrem, Torbjørn; Nelson, Diane R.; Rebecchi, Lorena; Cesari, Michele
abstract
The peculiar family Murrayidae, comprising the genera Murrayon, Dactylobiotus and Macroversum, contains relatively rare species living in hydrophilic and freshwater habitats on all continents, and contains two of the six exclusively freshwater tardigrade genera. This family probably represents an example of the evolution and persistence of an asexual lineage that differentiated into several taxa without sexual reproduction. Analyses of nuclear and mitochondrial genes (18S, 28S, ITS2 and cox1), and the increase of five taxa to the phylogenetic analyses of Murrayidae led us to infer that Murrayon is polyphyletic, being composed of two ‘species groups’ that also find morphological supports: the ‘dianeae group’ characterised by peculiar egg processes (rod-shaped and covered with a cuticular layer), animals with large, evident epicuticular pillars and small claws; and the ‘pullari group’ characterised by conical egg processes, animals with very small epicuticular pillars, and proportionally larger and longer claws. This latter group is a sister group to Dactylobiotus. Murrayon hastatus is the only species within the genus that has an uncertain position with eggs of the ‘dianeae group’ and animals of the ‘pullari group’. We propose the erection of Paramurrayon gen. nov. (for the ‘dianeae group’ of species), the emendation of Murrayon, and new taxonomic keys for both genera. Possible scenarios of the evolution of taxa within Murrayidae are hypothesised based on synapomorphic characters. Paramurrayon meieri sp. nov. from Norway is described with an integrative approach. Photographs of type material of Murrayon stellatus, Murrayon nocentiniae, Murrayon ovoglabellus and Macroversum mirum are shown for the first time, together with descriptions of new characters. Murrayon hibernicus is considered as nomen dubium and Murrayon hyperoncus is transferred to Macrobiotus pending further analyses.
2022
- Production of reactive oxygen species and involvement of bioprotectants during anhydrobiosis in the tardigrade Paramacrobiotus spatialis
[Articolo su rivista]
Giovannini, Ilaria; Boothby, Thomas C.; Cesari, Michele; Goldstein, Bob; Guidetti, Roberto; Rebecchi, Lorena
abstract
Water unavailability is an abiotic stress causing unfavourable conditions for life. Nevertheless, some
animals evolved anhydrobiosis, a strategy allowing for the reversible organism dehydration and
suspension of metabolism as a direct response to habitat desiccation. Anhydrobiotic animals undergo
biochemical changes synthesizing bioprotectants to help combat desiccation stresses. One stress
is the generation of reactive oxygen species (ROS). In this study, the eutardigrade Paramacrobiotus
spatialis was used to investigate the occurrence of ROS associated with the desiccation process.
We observed that the production of ROS signifcantly increases as a function of time spent in
anhydrobiosis and represents a direct demonstration of oxidative stress in tardigrades. The degree
of involvement of bioprotectants, including those combating ROS, in the P. spatialis was evaluated
by perturbing their gene functions using RNA interference and assessing the successful recovery
of animals after desiccation/rehydration. Targeting the glutathione peroxidase gene compromised
survival during drying and rehydration, providing evidence for the role of the gene in desiccation
tolerance. Targeting genes encoding glutathione reductase and catalase indicated that these
molecules play roles during rehydration. Our study also confrms the involvement of aquaporins 3 and
10 during rehydration. Therefore, desiccation tolerance depends on the synergistic action of many
diferent molecules working together.
2022
- Resistance to Extreme Stresses by a Newly Discovered Japanese Tardigrade Species, Macrobiotus kyoukenus (Eutardigrada, Macrobiotidae)
[Articolo su rivista]
Cesari, Michele; Giovannini, Ilaria; Altiero, Tiziana; Guidetti, Roberto; Cornette, Richard; Kikawada, Takahiro; Rebecchi, Lorena
abstract
Tardigrades are small micrometazoans able to resist several environmental stresses in any
stage of their life cycle. An integrated analysis of tardigrade specimens collected in Tsukuba (Japan)
revealed a peculiar morphology and a new sensory field in the cloaca. Molecular taxonomy and
phylogenetic analysis on different genes (COI, ITS2, 18S and 28S) confirmed that this population is
a new species, Macrobiotus kyoukenus sp. nov., belonging to the widespread Macrobiotus hufelandi
group. The stress resistance capabilities of M. kyoukenus sp. nov. have been tested by submitting
animals to extreme desiccation, rapid freezing, and high levels of ultraviolet radiations (UVB and
UVC). Animals were able to survive desiccation (survivorship 95.71 ± 7.07%) and freezing up to -80
°C (82.33 ± 17.11%). Both hydrated and desiccated animals showed a high tolerance to increasing
UV radiations: hydrated animals survived to doses up to 152.22 kJ m−2 (UVB) and up to 15.00 kJ m−2
(UVC), while desiccated specimens persisted to radiations up to 165.12 kJ m−2 (UVB) and up to 35.00
kJ m−2 (UVC). Present data contribute to the discovery of a larger tardigrade biodiversity in Japan,
and the tolerance capabilities of M.
2021
- When DNA sequence data and morphological results fit together: Phylogenetic position of Crenubiotus within Macrobiotoidea (Eutardigrada) with description of Crenubiotus ruhesteini sp. nov
[Articolo su rivista]
Guidetti, R.; Schill, R. O.; Giovannini, I.; Massa, E.; Goldoni, S. E.; Ebel, C.; Forschler, M. I.; Rebecchi, L.; Cesari, M.
abstract
The integration of morphological data and data from molecular genetic markers is important for examining the taxonomy of meiofaunal animals, especially for eutardigrades, which have a reduced number of morphological characters. This integrative approach has been used more frequently, but several tardigrade taxa lack molecular confirmation. Here, we describe Crenubiotus ruhesteini sp. nov. from the Black Forest (Germany) integratively, with light and electron microscopy and with sequences of four molecular markers (18S, 28S, ITS2, cox1 genes). Molecular genetic markers were also used to confirm the recently described Crenubiotus genus and to establish its phylogenetic position within the Macrobiotoidea (Eutardigrada). The erection of Crenubiotus and its place in the family Richtersiidae are confirmed. Richtersiidae is redescribed as Richtersiusidae fam. nov. because its former name was a junior homonym of a nematode family.
2020
- The species identification problem in mirids (Hemiptera: Heteroptera) highlighted by DNA barcoding and species delimitation studies
[Articolo su rivista]
Piemontese, L.; Giovannini, I.; Guidetti, R.; Pellegri, G.; Dioli, P.; Maistrello, L.; Rebecchi, L.; Cesari, M.
abstract
Due to the difficulties associated with detecting and correctly identifying mirids, developing an accurate species identification approach is crucial, especially for potential harmful species. Accurate identification is often hampered by inadequate morphological key characters, invalid and/or outdated systematics, and biases in the molecular data available in public databases. This study aimed to verify whether molecular characterization (i.e. DNA barcoding) is able to identify mirid species of economic relevance and if species delimitation approaches are reliable tools for species discrimination. Cytochrome c oxydase 1 (cox1) data from public genetic databases were compared with new data obtained from mirids sampled in different Italian localities, including an old specimen from private collection, showing contrasting results. Based on the DNA barcoding approach, for the genus Orthops, all sequences were unambiguously assigned to the same species, while in Adelphocoris, Lygus and Trigonotylus there were over-descriptions and/or misidentifications of species. On the other hand, in Polymerus and Deraeocoris there was an underestimation of the taxonomic diversity. The present study highlighted an important methodological problem: DNA barcoding can be a good tool for pest identification and discrimination, but the taxonomic unreliability of public DNA databases can make this method useless or even misleading.
2019
- A closer look at the antennae of the invasive Halyomorpha halys: fine structure of the sensilla
[Articolo su rivista]
Ibrahim, Aya; Giovannini, Ilaria; Anfora, Gianfranco; Valerio ROSSI STACCONI, Marco; Malek, Robert; Maistrello, Lara; Guidetti, Roberto; Romani, Roberto
abstract
The brown marmorated stink bug, Halyomorpha halys, is an invasive agricultural and urban pest capable of feeding on over 100 species of host plants. The antennae of this bug play an important role not only in detecting food and mates but also in short-range location of conspecifics when aggregating for diapause. The morphology and distribution of antennal sensilla of H. halys were investigated at an ultrastructural level using scanning and transmission electron microscopy approaches. Adults have 5-segmented antennae, made up of a scape, a 2-segmented pedicel and two flagellomeres, while 5th instar nymphs have shorter, 4-segmented antennae, with only one pedicel segment. Five types of sensilla are distinguished, based on their shape, length and basal width and the presence of basal socket and pores: sensilla basiconica (types A, B, C, D and E), sensilla coeloconica, sensilla trichoidea and sensilla chaetica (types A and B). Sexual dimorphism was not observed in this species, with respect to the morphological features and abundance of sensilla. The most abundant sensory structures are sensilla trichoidea showing characteristics typical of olfactory sensilla, whereas the least abundant are sensilla coeloconica which may be involved in thermo- and hygro-reception. Basiconic sensilla type A are solely identified on the antennae of 5th instar nymphs, where they presumably have a dual mechanosensorygustatory role. The putative function of the remaining sensilla are herein discussed.
2018
- Will the Antarctic tardigrade Acutuncus antarcticus be able to withstand environmental stresses related to global climate change?
[Articolo su rivista]
Giovannini, Ilaria; Altiero, Tiziana; Guidetti, Roberto; Rebecchi, Lorena
abstract
Because conditions in continental Antarctica are highly selective and extremely hostile to life, its biota is depauperate, but well adapted to live in this region. Global climate change has the potential to impact continental Antarctic organisms because of increasing temperatures and ultraviolet radiation. This research evaluates how ongoing climate changes will affect Antarctic species, and whether Antarctic organisms will be able to adapt to the new environmental conditions. Tardigrades represent one of the main terrestrial components of Antarctic meiofauna; therefore, the pan-Antarctic tardigrade Acutuncus antarcticus was used as model to predict the fate of Antarctic meiofauna threatened by climate change. Acutuncus antarcticus individuals tolerate events of desiccation, increased emperature and UV radiation. Both hydrated and desiccated animals tolerate increases in UV radiation, even though the desiccated animals are more resistant. Nevertheless, the survivorship of hydrated and desiccated animals is negatively affected by the combination of temperature and UV radiation, with the hydrated animals being more tolerant than desiccated animals. Finally, UV radiation has a negative impact on the life history traits of successive generations of A. antarcticus, causing an increase in egg reabsorption and teratological events. In the long run, A. antarcticus could be at risk of population reductions or even extinction. Nevertheless, because the changes in global climate will proceed gradually and an overlapping of temperature and UV increase could be limited in time, A. antarcticus, as well as many other Antarctic organisms, could have the potential to overcome global warming stresses, and/or the time and capability to adapt to the new environmental conditions.
2017
- Tardigrades Use Intrinsically Disordered Proteins to Survive Desiccation
[Articolo su rivista]
Boothby, Thomas C.; Tapia, Hugo; Brozena, Alexandra H.; Piszkiewicz, Samantha; Smith, Austin E; Giovannini, Ilaria; Rebecchi, Lorena; Pielak, Gary J.; Koshland, Dough; Goldstein, Bob
abstract
Tardigrades are microscopic animals that survive a
remarkable array of stresses, including desiccation.
How tardigrades survive desiccation has remained
a mystery for more than 250 years. Trehalose, a
disaccharide essential for several organisms to survive
drying, is detected at low levels or not at all in
some tardigrade species, indicating that tardigrades
possess potentially novel mechanisms for surviving
desiccation. Here we show that tardigrade-specific
intrinsically disordered proteins (TDPs) are essential
for desiccation tolerance. TDP genes are constitutively
expressed at high levels or induced during
desiccation in multiple tardigrade species. TDPs
are required for tardigrade desiccation tolerance,
and these genes are sufficient to increase desiccation
tolerance when expressed in heterologous systems.
TDPs form non-crystalline amorphous solids
(vitrify) upon desiccation, and this vitrified state mirrors
their protective capabilities. Our study identifies
TDPs as functional mediators of tardigrade desiccation
tolerance, expanding our knowledge of the roles
and diversity of disordered proteins involved in
stress tolerance.
2016
- Comparative analysis of fatty acid profile in three eutardigrade species
[Poster]
Giovannini, I; Mantovani, V; Galeotti, F; Chersoni, L; Guidetti, R; Volpi, N; Rebecchi, L.
abstract
Tardigrades colonize a wide range of habitats in which they can be predators, prey or primary consumers in food webs. Most species are herbivorous, feeding on cell fluid of algae and mosses, while others feed on bacteria, or prey on micrometazoans. Despite the wide range of food sources, details on food preference and on consequent lipid composition of tardigrade species are in practice unknown. Aiming to fill the gap of knowledge, we investigated the fatty acid composition of three eutardigrade species, since fatty acids are the main component of lipids and they play an important role in the function of cell membranes and in the physiological responses of organisms. The species, differing in colonized habitat and probably in diet, were: Acutuncus antarcticus (Hypsibiidae), a freshwater Antarctic species cultured using Chlorococcum sp. as food source, and the moss-dwelling species Macrobiotus macrocalix and Richtersius coronifer (Macrobiotidae). For each species, lipids were extracted from ten replicates of 150-250 animals with chloroform/methanol and the total extracts were used to obtain the fatty acid metylesters that were injected into a gas chromatograph. In all species, the same 21 fatty acids belonging to saturated, monounsaturated (MUFA) and polyunsaturated (PUFA) groups were identified. In A. antarcticus the most represented fatty acids were: palmitic (C16:0), stearic (C18:0), oleic (C18:1n-9), and myristic (C14:0) acids; saturated fatty acids (56.6%) were the most abundant with respect to MUFA (22.3%) and PUFA (21.1%). In M. macrocalix the most represented were: oleic (C18:1n-9), palmitic (C16:0), stearic (C18:0), and linoleic (C18:2n-6) acids; the saturated fatty acids (38.4%), MUFA (28.8%) and PUFA (32.8%) were uniformly distributed. In R. coronifer, alpha-linolenic (C18:3n-3), palmitic (C16:0), stearic (C18:0), and arachidonic (C20:4n-6) acids are the most represented; the percentage of PUFA (52.8%) was higher than that of MUFA (8.2%) and saturated fatty acids (38.9%). These data indicate clear differences in the fatty acid composition and amount among species. The fatty acid profiles reflect the food source and can be used as indicator to assess the feeding diet of tardigrades. Interestingly, species inhabiting the same substrate and eating the same food (moss cell content) use/transform the fatty acids in different way indicating different biochemical needs.
2016
- The Compact Body Plan of Tardigrades Evolved by the Loss of a Large Body Region
[Articolo su rivista]
Smith, Frank W; Boothby, Thomas C; Giovannini, Ilaria; Rebecchi, Lorena; Jockusch, Elizabeth L; Goldstein, Bob
abstract
The superphylum Panarthropoda (Arthropoda, Onychophora,
and Tardigrada) exhibits a remarkable
diversity of segment morphologies, enabling these
animals to occupy diverse ecological niches. The
molecular identities of these segments are specified
by Hox genes and other axis patterning genes
during development [1, 2]. Comparisons of molecular
segment identities between arthropodandonychophoran
species have yielded important insights into the
origins and diversification of their body plans [3–9].
However, the relationship of the segments of tardigrades
to those of arthropods and onychophorans
has remained enigmatic [10, 11], limiting our understanding
of early panarthropod body plan diversification.
Here, we reveal molecular identities for all of the
segments of a tardigrade. Based on our analysis, we
conclude that tardigrades have lost a large intermediate
region of thebody axis—aregion correspondingto
the entire thorax and most of the abdomen of insects—
and that they have lost the Hox genes that
originally specified this region. Our data suggest that
nearly the entire tardigrade body axis is homologous
to just the head region of arthropods. Based on our results,
we reconstruct a last common ancestor of Panarthropoda
that had a relatively elongate body plan
like most arthropods and onychophorans, rather
than a compact, tardigrade-like body plan. These results
demonstrate that the body plan of an animal
phylum can originate by the loss of a large part of
the body.
2016
- They are among us: the European invasion of the alien brown marmorated stinkbugs Halyomorpha halys (Heteroptera, Pentatomidae)
[Poster]
Piemontese, L.; Cesari, Michele; Maistrello, Lara; Giovannini, Ilaria; Dioli, P.; Partsinevelos, G. K.; Rebecchi, Lorena; Guidetti, Roberto
abstract
The brown marmorated stinkbug (BMSB), Halyomorpha halys is an invasive alien species native to
eastern Asia. Its presence outside the original area of distribution has been recorded for the first time
in North America (Canada and U.S.A.) and, more recently, in Europe (Switzerland, France, Hungary,
Romania, Austria, Serbia and Italy), where H. halys is spreading rapidly. Other than being a
household pest all over its introduced range, this stinkbug is causing great economic losses in the
U.S.A and Italian orchards/crops due to its highly polyphagous nature and bivoltinism. Tracing back
the pattern of introduction and monitoring the spread of BMSB in the European territory will be
useful to implement better pest control strategies. The present study aimed to identify the potential
pathways of entry of H. halys in Europe by detecting the genetic diversity of specimens collected all
over Italy, and in Switzerland, Romania and Greece. The analyses of 1,175 bp of mitochondrial DNA
cytochrome c oxidase I and II genes (cox1 and cox2) of over 200 specimens led to the identification
of 12 haplotypes never observed before (10 for cox1 and 2 for cox2). Present data indicate a higher
haplotype diversity of European specimens compared with the American ones; instead, the diversity
is lower with regard to the Asian samples, except for the cox2 marker. A clear-cut difference in
haplotype distribution was found between North and South Europe: Switzerland and France share a
similar haplotype pattern, whereas Italian, Hungarian, Romanian and Greek samples are more similar,
with the Italian and Greek specimens showing the higher genetic diversity. In Italy, genetic diversity
for both markers is higher in Piedmont, Lombardy and Veneto, while in the remaining regions of
Northern Italy in which the species is spread, it is fairly low. Haplotype similarity with both Chinese
and Korean samples led to hypothesize that the introduction of H. halys in Europe, and in Italy in
particular, has occurred by means of multiple events from Asia and that the BMSB is currently
expanding its range in the European continent.
2015
- Life history traits and reproductive mode of the tardigrade Acutuncus antarcticus under laboratory conditions: strategies to colonize the Antarctic environment
[Articolo su rivista]
Altiero, T.; Giovannini, I.; Guidetti, R.; Rebecchi, L.
abstract
Global climate change has become an important issue, particularly for organisms living in the Antarctic region, as the predicted temperature increase can affect their life history traits. The reproductive mode and life history traits of one of the most widespread species of tardigrades in Antarctica were analyzed. Specimens of the eutardigrade Acutuncus antarcticus from a temporary freshwater pond at Victoria Land (Antarctica) were individually cultured. This species reproduced continuously by thelytokous meiotic parthenogenesis. Its life cycle was short (60–90 days) and the reproductive output was low, with a short generation time (25–26 days). The maternal effect can be responsible of the phenotypic plasticity observed in life history traits of the three analyzed generations that may be seen as a bet-hedging strategy, as also observed in other animals inhabiting stochastic environments. These traits, along with the cryptobiotic capability of A. antarcticus, are advantageous for exploiting the conditions suitable for growth and reproduction during the short Antarctic summer, and can explain its wide distribution on the Antarctic continent. These results open new avenues of research for determining the role of bet-hedging strategy in organisms living in unpredictable environments.
2015
- Superoxide Anion Radical Production in the Tardigrade Paramacrobiotus richtersi, the First Electron Paramagnetic Resonance Spin-Trapping Study
[Articolo su rivista]
Savic, Aleksandar G; Guidetti, Roberto; Turi, Ana; Pavicevic, Aleksandra; Giovannini, Ilaria; Rebecchi, Lorena; Mojovic, Milos
abstract
Anhydrobiosis is an adaptive strategy that allows withstanding almost complete body water loss. It has been developed independently by many organisms belonging to different evolutionary lines, including tardigrades. The loss of water during anhydrobiotic processes leads to oxidative stress. To date, the metabolism of free radicals in tardigrades remained unclear. We present a method for in vivo monitoring of free radical production in tardigrades, based on electron paramagnetic resonance and spin-trap DEPMPO, which provides simultaneous identification of various spin adducts (i.e., different types of free radicals). The spin trap can be easily absorbed in animals, and tardigrades stay alive during the measurements and during 24-h monitoring after the treatment. The results show that hydrated specimens of the tardigrade Paramacrobiotus richtersi produce the pure superoxide anion radical ((•)O2(-)). This is an unexpected result, as all previously examined animals and plants produce both superoxide anion radical and hydroxyl radical ((•)OH) or exclusively hydroxyl radical.
2014
- Biodiversity and adaptive strategy to Antarctica: the tardigrades
[Abstract in Atti di Convegno]
Vecchi, Matteo; Mcinnes, Michele Cesari Sandra; Giovannini, Ilaria; Altiero, Tiziana; Rebecchi, Lorena; Bertolani, Roberto; Guidetti, Roberto
abstract
Tardigrades are important members of the Antarctic biota in terms of abundance, distribution, and colonized substrates. Despite their importance and regular occurrence in the harsh Antarctic terrestrial ecosystems, there have been few ecological or taxonomic studies. We carried out an extensive sampling campaign along Victoria Land coastal line, collecting 180+ samples of lichens, mosses and freshwater sediments over a c. 600 km North–South transect. These samples revealed an unexpectedly high diversity: 14 species, four of which new for science.
Our results have underlined the level of undiscovered biodiversity in Antarctica. Some species had very localized distribution and occurred in specific substrates (e.g. moss or lichen). Other species were more scattered, and one (the endemic Acutuncus antarcticus) was present in almost all sampling areas and substrate types. This situation indicates that, as in temperate areas, Antarctica has tardigrade species with low dispersal capabilities, and a few species with high dispersal capabilities. Surprisingly, the genetic variability of A. antarcticus (COI gene) was extremely low even between very distant populations.
Laboratory experiments on the adaptive strategy of A. antarcticus indicated a very short life cycle (c. 3-4 months), shorter than in species from temperate regions. Moreover, A. antarcticus exhibits thelytoky and has cryptobiotic capabilities. These attributes allowed this species to colonize almost all viable habitats in Antarctica
2014
- Desiccation tolerance and production of Reactive Oxygen Species (ROS) in the anhydrobiotic water bear Paramacrobiotus richters
[Abstract in Atti di Convegno]
Giovannini, Ilaria; Guidetti, Roberto; Rebecchi, Lorena
abstract
The production of Reactive Oxygen Species (ROS) during desiccation processes is documented in bacteria and plants, whereas studies on animals are in practice lacking. In this study we investigated the response to dehydration with respect to the ROS production during the kinetic of the desiccation process and the short and long-time permanence in the anhydrobiotic state. We hypothesized that ROS production, and thus oxidative damage, may be greater in animals maintained desiccated for long-time than in animals maintained dry just for one day. The tardigrade Paramacrobiotus richtersi was used as a target animal. In parallel, oxidative stress was assessed in hydrated P. richtersi as control. Adults were experimentally dehydrated in laboratory using an optimal desiccation protocol to achieve a 100% survival rate of P. richtersi. Animals were maintained desiccated (at 3% RH and 20°C) from 1 to 40 days. Intracellular ROS production during rehydration was evaluated in the “storage cells” (free-floating cells in the tardigrade body cavity), after treatment of the tardigrades with the probe 2,7-dichlorodihydrofluorescein-diacetate (DCFH2-DA). The amount of green fluorescent oxidation product (DCF), which reflects the reaction of the probe with intracellular free radicals, was measure by a laser scanner confocal microscope. The desiccation process does not produce a high amount of ROS, even though previous studies on the same species, P. richtersi, demonstrated an increase of antioxidant enzyme activity in desiccated specimens with respect to hydrated ones. Instead, the long-time permanence in anhydrobiosis (20 days) produces a significant increase of ROS, as evidenced after three and twelve hours from rehydration. The collection of data about the permanence in anhydrobiosis for 40 days is in progress. The high amount of ROS detected in animals maintained dry for long-time, and consequently the oxidative molecular damages, justifies that tardigrade survival decreases with the time spent in a dry state.
2014
- The short life cycle of Acutuncus antarcticus (Tardigrada) as adaptation to Antarctic environment
[Abstract in Rivista]
Giovannini, Ilaria; Rebecchi, Lorena; Guidetti, Roberto; Altiero, Tiziana
abstract
The climate global change is altering the tropospheric ozone budget increasing harmful ultraviolet (UV) radiation with consequences on human and ecosystem health. Antarctic micrometazoans are particularly vulnerable to the synergic effects of increasing temperature and UV radiation as their growing season matches with the spring period of ozone depletion. The roles that phenotypic plasticity and genotypic evolution play in whether Antarctic organisms can cope with these environmental changes are not fully understood. Tardigrades represent one of the main terrestrial components of Antarctica fauna in terms of distribution, number of specimens and colonized substrates. Studies on adults of the eutardigrade Acutuncus antarcticus, one of the most abundant species in Antarctic bryophytes and freshwater sediments, showed that they survived to experimental exposition to increased temperature and UV radiation. These results suggest that A. antarcticus could survive to possible environmental changes. To test this hypothesis we firstly needed to know its life history traits, never studied before, in order to carry out further experiments on stress resistance of eggs and juveniles.
Cultures were started using single specimens of A. antarcticus collected from a temporary freshwater pond at the Italian Antarctic base at Victoria Land. Animals were fed with the algae Chlorococcus sp., and reared at 14°C and 12h/12h L/D.
A. antarcticus lays freely (rarely within exuvium) eggs hatching in 7-9 days. Newborns molt 2-3 times before their first oviposition that occurs at the age of 12-15 days. Successive ovipositions occur once a week (for 2-3 months) and are always preceded from a molting. The number of laid eggs (1-4 per oviposition) is a function of the female age. Throughout lifespan (3-4 months) each female lays up to 20 eggs. Individually reared newborns reached sexual maturity and, maintained isolated, laid eggs able to hatch. Males were never seen. Therefore this population reproduces via thelytokous parthenogenesis.
The life cycle evidenced in A. antarcticus, characterized by a short generation time with respects to that of species of temperate regions, represents an adaptive strategy which allows animals to be active and reproduce only during the short-time (ca. 2 months) in which water is available. This knowledge will allow further experiments to verify the eventual effects of temperature and UV radiation on the life history traits and fitness of Antarctic organisms.
2014
- The toughest animals on the Earth: desiccation tolerance and oxidative stress in tardigrades.
[Abstract in Atti di Convegno]
Rebecchi, Lorena; Altiero, Tiziana; Giovannini, Ilaria; Guidetti, Roberto
abstract
Tardigrades are microscopic aquatic ecdysozoans with remarkable abilities to withstand harsh physical conditions, such as dehydration or exposure to harmful highly-energetic radiations, including both solar and cosmic ionizing radiations and the vacuum of the space. They can persist in dehydrated state even for years at any life stage. Yet once external conditions become favorable they resume an unaffected active life. Nevertheless, survival of desiccated tardigrades decreases with the dehydration rate and time spent dry. On the other hand, experimental studies on Paramacrobiotus richtersi provide evidence that exposure to high temperatures, high humidity, and high oxygen partial pressure negatively affect long-term survival of anhydrobiotic tardigrades, and directly influence the time required to reactivate their metabolism. These abiotic factors produce molecular damages, which are accumulated in proportion to the time spent in the desiccated state, potentially leading to tardigrade death. Oxidative stress seems to be one of the most deleterious causes of damages due to water depletion. Experimental studies on P. richtersi indicate that the production of Reactive Oxygen Species (ROS) could occur during anhydrobiosis, being the accumulation of ROS higher in tardigrades maintained desiccated for long-time than in those maintained desiccated for one day only. Therefore anhydrobiosis needs a stringent control of oxidation processes including ROS production. Experimental studies evidence that glutathione and ROS scavenging enzymes represent a key group of molecules for desiccation tolerance in P. richtersi, where the activity of these enzymes is significantly higher in desiccated specimens than in hydrated specimens. We also suggest a role of tardigrade pigments (e.g. carotenoids) as scavengers for ROS forming during dehydration processes and/or exposition to solar radiations.
These data exhibit further evidence on the role of antioxidant defenses in tolerant desiccation organisms, and the role of tardigrades as animal model to discover the secret of life without water.
2013
- A DNA barcoding approach in the study of tardigrades
[Articolo su rivista]
Cesari, Michele; Guidetti, Roberto; Rebecchi, Lorena; Giovannini, Ilaria; Bertolani, Roberto
abstract
DNA barcoding is a technique proposed by Hebert and coworkers in 2003 for discriminating species through analysis of a single gene barcode locus. It aims to obtain a better taxonomic resolution than that achieved through morphological studies, and to avoid the decline in taxonomic knowledge. Today DNA barcoding is a global enterprise, and the implementation of the idea has seen a rapid rise (more than 1900 papers published to date on different organisms). Nonetheless, controversy still arises regarding barcoding and taxonomy. It is important to note that DNA barcoding does not focus on building a tree-of-life or on doing DNA taxonomy, even though sometimes it has been used for these purposes. DNA barcoding rather focuses on producing a universal molecular identification key based on strong taxonomic knowledge that should be included in the barcode reference library. In the phylum Tardigrada, DNA barcoding represents a recent approach to species identification and to help in solving taxonomic problems, especially considering the diminutive size of these animals and the paucity of morphological characters useful for taxonomy. In the framework of the MoDNA Project (Morphology and DNA), carried out by our research group in collaboration with several colleagues, we are combining the study of a fragment of the mitochondrial cytochrome c oxidase subunit I gene (cox1) with morphological data, in a wide sense (cuticular structures, chromosomes, data on sex ratio and reproduction), to form an integrative taxonomy approach for tardigrade species identification. We believe that without verified reference sequences from voucher specimens that have been authenticated by qualified taxonomists, there is no reliable library for newly generated sequences with which to be compared. Methods and protocols for standardized results are focused on obtaining tight correspondence between tardigrade morphology (and egg shell morphology, when useful), possibly both light and scanning electron microscopy images, and molecular sequence. This approach is particularly useful in describing new species, and important when applied on material collected in species type localities. Results using this approach are presented, primarily focusing on a number of species from the so-called “Macrobiotus hufelandi group”.
2013
- Effetti delle radiazioni ultraviolette e della temperatura su un organismo della meiofauna antartica: Acutuncus antarcticus (Eutardigrada).
[Abstract in Rivista]
Altiero, Tiziana; Guidetti, Roberto; Giovannini, Ilaria; Cesari, Michele; Montorfano, G.; Rizzo, A. M.; Bertolani, Roberto; Rebecchi, Lorena
abstract
I cambiamenti climatici in atto, oltre ad aumentare la temperatura, riducono la quantità di ozono troposferico. Questo porta ad un aumento delle radiazioni ultraviolette (UV), con potenziali conseguenze negative sulla salute umana e sugli ecosistemi. Tardigradi e rotiferi, principali componenti della meiofauna terrestre antartica, sono molto vulnerabili agli effetti sinergici di temperatura e UV, in quanto la loro stagione di sviluppo coincide con il periodo primaverile antartico, in cui normalmente si ha una diminuzione dell’ozono.
Sono state quindi analizzate le risposte fisiologiche e biochimiche all’incremento di temperatura e UV nell’eutardigrado Acutuncus antarcticus, una delle specie più abbondanti nelle briofite e nei piccoli invasi di acqua dolce dell’Antartide.
Esperimenti di essiccamento in condizioni controllate hanno evidenziato che A. antarcticus è in grado di attuare l’anidrobiosi, mostrando un’elevata sopravvivenza (92,8%), insolita per una specie di tardigrado generalmente dulciacquicola. Gli antiossidanti non sembrano però essere responsabili di tale sopravvivenza in quanto, ad eccezione della catalasi, non sono emerse differenze significative nell’attività/quantità di antiossidanti tra esemplari idratati e secchi, a differenza di quanto osservato in Paramacrobiotus richtersi, una specie di tardigrado prettamente “terrestre” che vive in zone temperate. Esperimenti tesi a valutare la resistenza di esemplari attivi di A. antarcticus all’incremento della temperatura (da 8°C a 41°C) hanno dimostrato che la specie è in grado di tollerare temperature elevate (a 33°C = 100% vivi; a 37°C = 35% vivi), anche se per breve tempo. Entrambi gli stati fisiologici (idratato ed essiccato) di A. antarcticus hanno dimostrato una buona resistenza alle radiazioni UV. Gli esemplari idratati hanno resistito fino alla dose di 61,9 kJ m-2 (5% vivi), mentre quelli in stato essiccato fino alla dose di 74,8 kJ m-2(7,5% vivi). Negli animali attivi, alla LD50 di 28,6 kJ m-2, l’effetto negativo degli UV aumenta in combinazione con l’incremento della temperatura (8°C: = 42,6% vivi; 15°C: = 1,7% vivi), dimostrando l’esistenza di un effetto sinergico. Pur restando in attesa di maggiori informazioni sul ciclo vitale, si può formulare l’ipotesi che questa specie antartica possieda ampie potenzialità di sopravvivenza ad eventuali cambiamenti ambientali.
2013
- Essiccamento e produzione di specie reattive dell’ossigeno (ROS) in tardigradi anidrobionti
[Abstract in Rivista]
Giovannini, Ilaria; Guidetti, Roberto; Rebecchi, Lorena
abstract
Anche se l’acqua è essenziale per la vita, organismi di linee evolutive diverse hanno sviluppato la capacità di tollerare un essiccamento estremo entrando in un particolare stato fisiologico reversibile, detto anidrobiosi. Con questo adattamento, rotiferi bdelloidei, nematodi e tardigradi perdono fino al 97% dell’acqua corporea, con sospensione del metabolismo e cambiamenti nell’organizzazione e composizione molecolare delle membrane cellulari. Nei tardigradi essiccati, la sopravvivenza a lungo termine è inversamente proporzionale a temperatura, umidità relativa dell’aria e pressione parziale di ossigeno, fattori abiotici che contribuiscono a danneggiare le molecole biologiche. Una delle principali cause di danno durante l’anidrobiosi sembra essere lo stress ossidativo, dovuto al disequilibrio fra l’eccessiva produzione di specie reattive dell’ossigeno (ROS) e la limitata attività degli antiossidanti. Tuttavia, la produzione di ROS durante l’essiccamento è ben documentata solo in pochi organismi, soprattutto autotrofi, mentre mancano praticamente dati sugli animali anidrobionti e, soprattutto, sull’effettivo accumulo di ROS durante la permanenza in anidrobiosi per lunghi periodi di tempo.
È stata quindi valutata la produzione di ROS nell’eutardigrado anidrobionte Paramacrobiotus richtersi, analizzando animali essiccati sperimentalmente in laboratorio [4 h a 18°C e 80% di umidità relativa dell’aria (RH); 4 h a 18°C e 50% RH; 12 h in gel di silice] e mantenuti essiccati per vari periodi di tempo (da 1 a 30 giorni) o sottoposti a temperature elevate (37°C e 60°C). Come controllo sono stati utilizzati animali mantenuti idratati. La produzione di ROS è stata valutata nei globuli cavitari, cellule libere nella cavità corporea dei tardigradi, dopo trattamento degli animali con il marcatore 2,7 diclorodiidrofluoresceina diacetato (DCFH2-DA) e successiva rilevazione della quantità del prodotto di ossidazione fluorescente 2,7 diclorofluoresceina (DFC), mediante microscopia confocale a scansione laser.
I primi dati indicano che in P. richtersi l’entrata in anidrobiosi di per sé non determina la produzione di elevate quantità di ROS, sebbene sia stato dimostrato un incremento dell’attività degli enzimi antiossidanti negli esemplari essiccati rispetto a quelli idratati. La produzione di ROS sembra aumentare quando gli animali essiccati sono mantenuti in anidrobiosi per lunghi periodi di tempo ed esposti ad alti valori di temperatura.
2013
- Physiological and biochemical adaptive responses and defense mechanisms to withstand increasing ultraviolet radiation and temperatures in an Antarctic meiofauna organism.
[Abstract in Atti di Convegno]
Giovannini, Ilaria; Guidetti, Roberto; Altiero, Tiziana; Cesari, Michele; Montorfano, G.; Rizzo, A. M.; Bertolani, Roberto; Rebecchi, Lorena
abstract
Physiological and biochemical adaptive responses and defense mechanisms to withstand increasing ultraviolet radiation and temperatures in an Antarctic meiofauna organism.
2012
- A DNA barcoding approach in the study of tardigrades.
[Abstract in Atti di Convegno]
Cesari, Michele; Rebecchi, Lorena; Guidetti, Roberto; Giovannini, Ilaria; Mori, L.; Bertolani, Roberto
abstract
DNA barcoding is a technique proposed by Hebert and coworkers in 2003 and it aims to discriminate biological entities through analysis of a single gene barcode locus. The DNA barcoding system promised a better taxonomic resolution than that achieved through morphological studies, with a partial solution to the decline in taxonomic knowledge. Today DNA barcoding is a global enterprise, and the implementation of the idea has seen a rapid rise (more than 450 papers published to date on different organisms). Nonetheless, controversy still arises regarding barcoding and taxonomy. It is important to note that DNA barcoding does not focus on building a tree-of-life or on doing DNA taxonomy, even though sometimes it has been used for these purposes. DNA barcoding rather focuses on producing a universal molecular identification key based on strong taxonomic knowledge that should be included in the barcode reference library.
In Phylum Tardigrada, DNA barcoding represents a recent approach to species identification and for helping to solve taxonomic problems, especially considering the diminutive size of these animals and the paucity of morphological characters useful for taxonomy. In the framework of the MoDNA Project (Morphology and DNA), carried out by our research group in collaboration with several colleagues, we are combining the study of a fragment of the mitochondrial cytochrome c oxidase subunit I gene (cox1) with morphological data, in a wide sense, to form an integrative taxonomy of tardigrades.
Building of a database of reference sequences is of paramount importance for a correct application of DNA barcoding in tardigrades. Without verified reference sequences from voucher specimens that have been authenticated by qualified taxonomists, there is no reliable library for newly generated sequences with which to be compared. Methods and protocols for standardized results are focused on obtaining tight correspondence between molecular sequence and animal morphology, possibly both LM and SEM images (and egg shell morphology, when useful). This approach is particularly useful in describing new species, and important when applied on material collected in type localities.
Results using this approach are presented, focusing primarily on a number of species from the so-called “Macrobiotus hufelandi group”.
2012
- L'approccio di DNA barcoding nello studio dei tardigradi
[Abstract in Atti di Convegno]
Bertolani, Roberto; Rebecchi, Lorena; Guidetti, Roberto; Altiero, Tiziana; Giovannini, Ilaria; Mori, L.; Cesari, Michele
abstract
Il DNA barcoding è stato proposto da Hebert e coll. nel 2003 con lo scopo di discriminare entità biologiche attraverso l’analisi di un singolo locus genico. Oggi il suo utilizzo appare in rapida ascesa (più di 450 lavori pubblicati riguardanti organismi anche molto differenti). Ciononostante, esistono ancora molte controversie sull’utilizzo di questo approccio in tassonomia. È importante sottolineare che solitamente il DNA barcoding non dovrebbe essere impiegato per la filogenesi, anche se a volte ciò è accaduto, ma piuttosto focalizzarsi sulla costruzione di una chiave di identificazione molecolare universale, basata però su consistenti informazioni tassonomiche che devono essere incluse nella barcode reference library.
Nel Phylum Tardigrada, il DNA barcoding rappresenta un approccio recente per l’identificazione delle specie e per la risoluzione di problemi tassonomici, anche alla luce delle ridotte dimensioni degli animali e della limitatezza dei caratteri morfologici disponibili. Da queste premesse è stato sviluppato il progetto MoDNA (Morfologia e DNA), combinando lo studio di un frammento del gene mitocondriale citocromo c ossidasi I (cox1) con dati morfologici anche a livello fine, allo scopo di sviluppare e validare l’integrative taxonomy sul modello tardigradi. L’indagine è stata condotta su gruppi di specie affini e criptiche in più famiglie di Eutardigrada.
La costruzione di un database di sequenze di riferimento è di importanza capitale per una corretta applicazione di questa metodica. Tuttavia, non è meno importante che queste sequenze siano strettamente correlate a specifici voucher specimens validati da esperti tassonomi. In mancanza di questo, il database di riferimento non può essere del tutto affidabile.
Per raggiungere questo importante obiettivo sono stati sviluppati metodi e protocolli per ottenere risultati standardizzati ed una stretta corrispondenza tra sequenza di DNA e morfologia, possibilmente con documentazione sia al microscopio ottico che elettronico a scansione. Un valore aggiunto è rappresentato dalla possibilità di utilizzo di materiale proveniente dal locus typicus di una specie già descritta e ancor più dall’utilizzo del DNA barcoding nella descrizione di nuove specie.
2012
- Morphology, DNA barcoding and phylogeny of Macrobiotus persimilis and Macrobiotus polonicus.
[Abstract in Atti di Convegno]
Bertolani, Roberto; Cesari, Michele; Lisi, O.; Rebecchi, Lorena; Giovannini, Ilaria; Pilato, G.
abstract
Macrobiotus polonicus and Macrobiotus persimilis are two species of the “Macrobiotus hufelandi group” which are distinguished one from the other for the presence in M. polonicus of one lateral gibbosity on each hind leg, the size of cuticular pores (smaller and less evident in M. polonicus), the buccal tube size (larger in M. persimilis) and the egg processes (larger terminal discs and with more evident indentation in M. polonicus). However, the two taxa share peculiar characteristics: similar values of the pt index relative to the insertion point of the stylet supports on the buccal tube, first macroplacoid clearly longer than the second and with a central constriction, lunules of the hind legs clearly larger than those of the first three pairs and with an indented margin, egg shell smooth or with faint dots but without a reticular design. Therefore, it could be hypothesized that they belong to a single peculiar group, namely the “Macrobiotus polonicus/persimilis group”.
In order to verify this hypothesis, a morphological and DNA barcoding analysis (cox1) was carried out on tardigrades attributable to these two taxa and coming from five different localities in Italy and France.
This study revealed differences, both morphological and molecular, between some populations apparently belonging to the same species, which in some cases turned out to be cryptic species. A phylogenetic analysis using rDNA 18S was performed in order to verify the relationships among the species of this group and more in general among those of the “M. hufelandi group”.
The DNA barcoding analysis revealed a high divergence, with very high values of genetic distance among some populations (more than 18%). In one case (Enna, Sicily) at least two different and very distinct entities are present together. The morphological and morphometrical investigation, also based on type material, confirms this variety, showing that at least four species related to M. polonicus and three related to M. persimilis should be distinct. In several cases the relationship between morphology and cox1 sequence has been ascertained.
The dendrogram computed on 18S showed that there are two different sequences pertaining to the “M. polonicus/persimilis group” clustered in a single evolutionary line inside the main M. hufelandi cluster.
2011
- An example of problems associated with DNA barcoding in tardigrades: a novel method for obtaining voucher specimens
[Articolo su rivista]
Cesari, Michele; Giovannini, Ilaria; Bertolani, Roberto; Rebecchi, Lorena
abstract
We have in recent papers revealed that an integrative taxonomy approach helps to solve taxonomic problems in tardigrades. However, whole tardigrades are required for DNA work, which leaves no hologenophore voucher specimens with adult morphology. Using a novel methodology for the Tardigrada, we introduce the practice of collecting high quality maximum magnification light microscopy images of recently thawed animals to act as hologenophore voucher specimens of animals later used for DNA barcode sequencing. Within the framework of a DNA barcoding project on tardigrades, we collected a moss sample from the type locality of Macrobiotus terminalis Bertolani & Rebecchi, 1993 (Castelsantangelo, Central Apennines, Italy), a species of the “Macrobiotus hufelandi group”. Within the moss sample we found several animals and eggs with a morphology that corresponded to the original description of M. terminalis, while others were attributable to Macrobiotus macrocalix Bertolani & Rebecchi, 1993. In this study, molecular (cox1 mtDNA) analyses demonstrated no intraspecific variability in M. terminalis from the type locality but very large interspecific differences when compared with M. macrocalix and GenBank data for other species within the M. “hufelandi group”. There was also a large difference between our M. terminalis sequences and the GenBank data of a specimen attributed to the same species. The GenBank sequence originated from a population in the Northern Apennines, whose morphology appeared to be like that of the specimens of the locus typicus. This confirmed the importance in utilising material from the type locality for linking molecular data to the species’ morphological characters. Our paper underlines the importance of an integrative taxonomy in species diagnoses and demonstrates a scenario where morphological observations alone are not always sufficient. Lastly, this work adds reliable information to the sequence reference library that provides a useful building block for further studies on similar and related tardigrade taxa.
2011
- DNA barcoding and integrative taxonomy of Macrobiotus hufelandi C.A.S. Schultze 1834, the first tardigrade species to be described, and some related species.
[Articolo su rivista]
Bertolani, Roberto; Rebecchi, Lorena; Giovannini, Ilaria; Cesari, Michele
abstract
Within the framework of a DNA barcoding project on tardigrade species, a study was carried out on Macrobiotus hufelandi C.A.S. Schultze 1834, the first formally described tardigrade species. We used samples collected from the type locality and additional material from other European sites containing species of the “M. hufelandi group”. The study was performed by integrating morphological, karyological and molecular (mt-DNA cox1) information and comparing these data with morphological data from the type material. Several species from this group were found in the type locality of M. hufelandi (near Freiburg, Black Forest, Germany) and these were all barcoded. One was M. hufelandi, the other two were: Macrobiotus sandrae Bertolani & Rebecchi 1993 (originally described from the same locality), and Macrobiotus vladimiri Bertolani, Biserov, Rebecchi & Cesari in press (type locality Andalo, Italy), all with interspecific genetic distances of more than 19%. A fourth cryptic species, which had the same morphology as M. hufelandi but a genetic distance of 6.7%, was not described as a new taxon but named M. cf. hufelandi sp.1 for this study. Macrobiotus sandrae and M. vladimiri were also present (and barcoded) in Italy (Alps). Additional individuals (animals and eggs) were also found, and barcoded, in Italy (Apennines) and Switzerland that belonged to the haplogroup Macrobiotus cf. hufelandi sp. 1. These data together with other recent studies on tardigrade DNA barcoding represent a starting point for further studies on tardigrade biogeography, phylogeography and diversity.
2011
- Filogenesi e integrative taxonomy nei Macrobiotus del “gruppo hufelandi” (Tardigrada, Eutardigrada)
[Abstract in Atti di Convegno]
Cesari, Michele; Rebecchi, Lorena; Giovannini, Ilaria; Pilato, G.; Lisi, O.; Vicente, F.; Kiosya, Y.; Bertolani, Roberto
abstract
Analiisi della filogenesi dei Macriobiotus del gruppo hufelandi medainte un approccio integrativo