BIOCHEMICAL TAXONOMY OF MARINE PHYTOPLANKTON BY ELECTROPHORESIS OF ENZYMES. II. LOSS OF HETEROZYGOSITY IN CLONAL CULTURES OF THE CENTRIC DIATOMS SKELETONEMA COSTATUM AND THALASSIOSIRA PSEUDONANA1, 2

An electrophoretic survey of 12 new isolates of Thalassiosira pseudonana Hasle & Heimdal and 25 new isolates of Skeletonema costatum (Grev.) Cleve revealed several heterozygote genotypes at malate dehydrogenase (MDH) and phosphohexose isomerase (PHI) loci. The new clones were maintained in culture for 6 mo and then reasayed at these two loci. All MDH heterozygotes and halj of the PHI heterozygotes had become homozygous. This resulted in a collection of clones that are largely homozygous but that are samples of polymorphic species. The physlogical implications of this loss of heterozygosity in clonal cultures has not been analyzed. Hawever, any change in a clone that is the result of culturing conditions reduces the usefulness of that clone as a laboratory test organism for ecological correlations.     

DNA Barcodes indicate members of the Anopheles fluviatilis (Diptera: Culicidae) species complex to be conspecific in India

Anopheles fluviatilis, a major vector of malaria in India has been described as a complex of three sibling species members, named as S, T and U, based on variations in chromosomal inversions. Also, ribosomal DNA markers (repetitive Internal Transcribed Spacer 2 (ITS2) and 28S D3 region) were described to differentiate these three sibling species members. However, controversies prevail on the genetic isolation status of these cryptic species. Hence, we evaluated this taxonomic incongruence employing DNA barcoding, the well established methodology for species identification, using 60 An. fluviatilis sensu lato specimens, collected from two malaria endemic eastern states of India. These specimens were also subjected to sibling species characterization by ITS2 and D3 DNA markers. The former marker identified 31 specimens among these as An. fluviatilis S and 21 as An. fluviatilis T. Eight specimens amplified DNA fragments specific for both S and T. The D3 marker characterized 39 specimens belonging to species S and 21 to species T. Neither marker identified species U. Neighbor Joining analysis of mitochondrial cytochrome c oxidase gene 1 sequences (the DNA barcode) categorized all the 60 specimens into a single operational taxonomic unit, their Kimura 2 parameter (K2P) genetic variability being only 0.8%. The genetic differentiation (F_ST) and gene flow (N_m) estimates were 0.00799 and 62.07, respectively, indicating these two ‘species’ (S & T) as genetically con‐specific intermixing populations with negligible genetic differentiation. Earlier investigations have refuted the existence of species U. Also, this study demonstrated that An. fluviatilis and the closely related An. minimus could be taxonomically differentiated by the DNA Barcode approach (K2P = 5.0%).     

Patterns of allozyme variation and clonal diversity in Carex lasiocarpa and C. pellita (Cyperaceae)

We compared genetic variability and clonal diversity in the closely related sedges Carex lasiocarpa and C. pellita using allozyme markers at 12 loci. Both species produce spreading rhizomes, occur in open habitats, and can form large clones; C. lasiocarpa primarily along lakeshores, or in bogs, fens, or marshes, and C. pellita more commonly along riverbanks and ditches or in wet meadows. Polymorphism was high in both species, similar to that of other rhizomatous Carex and to other species with comparable growth forms and life histories. Total gene diversity was higher for C. lasiocarpa (H_T = 0.266) than for C. pellita (H_T = 0.248), but the coefficient of differentiation among populations was lower. Rare alleles usually occurred as heterozygotes rather than homozygotes, and significant deviations from Hardy-Weinberg equilibrium were found at few loci, suggesting that both species are predominantly outcrossing. Mean clone size, diversity, and evenness were not significantly different between the two species, although populations varied from monoclonal to those where nearly every sampled shoot belonged to a different clone. No significant differences in genetic variation or clone size of C. pellita were found when comparisons were made between groups based on climate (warmer vs. cooler) or successional stage (pioneer vs. established). However, within C. lasiocarpa, genetic variability decreased and clone size increased in cooler climates and at established sites.     

MOLECULAR GENETIC MANIPULATION OF THE DIATOM THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE)1

Here, we describe the first system for genetic transformation of Thalassiosira pseudonana (Hustedt) Hasle et Heimdal, the only diatom for which a complete genome sequence is presently available. This method is based on microparticle bombardment followed by selection of transformants using the antibiotic nourseothricin. It exhibits the highest transformation efficiency compared with transformation systems for other diatom species. To achieve the high transformation efficiency, it is important to allow recovery of the bombarded T. pseudonana cells in non‐selective suspension culture before spreading on nourseothricin containing agar plates. It is demonstrated that T. pseudonana is readily susceptible to co‐transformation allowing for the simultaneous introduction of a non‐selective gene together with the selection marker gene. Both introduced genes are stably inherited even in the absence of the antibiotic selection pressure. We have developed two T. pseudonana‐specific expression vectors that can drive constitutive expression (vector pTpfcp) and inducible expression (vector pTpNR) of introduced genes. In combination with the available genome data the T. pseudonana transformation system is expected to provide a powerful tool for functional genomics in diatoms.     

THE MARINE,PLANKTONIC DIATOMS THALASSIOSIRA OCEANICA SP. NOV. AND T. PARTHENEIA1

The diatom clone 13–1 isolated from the Sargasso Sea by Dr. R. R. L. Guillard as Cyclotella nana Hust. or Thalassiosira pseudonana Hasle and Heimdal is described as a separate species, Thalassiosira oceanica sp. nov. An amplified diagnosis of Thalassiosira partheneia Schrader is given, and a comparison of the two species is made based on light and electron microscopy. Similarities are present in the apparent distribution pattern, cell size, and girdle structure. Differences are present in the shape of the areolae of the valvocopula and the copula, in the structure of the vela of these bands, in the texture of the external valve surface, in the morphology of the strutted processes, in the location of the labiate process, in the distance between the marginal strutted processes, and by the presence of a marginal ridge in T. oceanica. Fifteen nanoplanktonic (maximum diameter = 20 UmUm) Thalassiosira species are listed, among them T. oceanica and T. partheneia.     

SELENIUM: AN ESSENTIAL ELEMENT FOR GROWTH OF THE COASTAL MARINE DIATOM THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE)1,2

An obligate requirement for selenium is demonstrated in axenic culture of the coastal marine diatom Thalassiosira pseudonana (clone 3H) (Hust.) Hasle and Heimdal grown in artificial seawater medium. Selenium deficiency was characterized by a reduction in growth rate and eventually by a cessation of cell division. The addition of 10⁻¹⁰ M Na₂SeO₃ to nutrient enriched artificial seawater resulted in excellent growth of T. pseudonana and only a slight inhibition of growth occurred at Na₂SeO₃ concentrations of 10⁻³ and 10⁻² M. By contrast, Na₂SeO₄ failed to support growth of T. pseudonana when supplied at concentrations less than 10⁻⁷ M and the growth rate at this concentration was only one quarter of the maximum growth rate. The addition of 10⁻³ and 10⁻² M Na₂SeO₄ to the culture medium was toxic and cell growth was completely inhibited. Eleven trace elements were tested for their ability to replace the selenium requirement by this alga find all were without effect. In selenium-deficient and selenium-starved cultures of T. pseudonana cell volume increased as much as 10-fold as a result of an increase in cell length (along the pervalvar axis) but cell width was constant. It is concluded that selenium is an indispensable element for the growth of T. pseudonana and it should be included as a nutrient enrichment to artificial seawater medium when culturing this alga.     

EFFECTS OF VARIATION IN TEMPERATURE. II. ON THE FATTY ACID COMPOSITION OF EIGHT SPECIES OF MARINE PHYTOPLANKTON1

Eight species of marine phytoplankton showed significant variation in the relative amount of some fatty acids (FAs) in response to variation in temperature. Large changes in relative amounts of certain FAs occurred as a result of a 15° C change in growth temperature. For example, 14:0 increased from ?4% of total FAs at 10° C to > 20% at 25° C for Chaetoceros simplex and Isochrysis aff. galbana but decreased for Phaeodactylum tricornutum. The percentage of the polyunsaturated fatty acid (PUFA) 16:ω1 was consistently greater at 10° C than at 25° C, and the converse was usually true for 16: 4ω3. Calculated over all eight species, there was a modest but significant inverse relationship between the percentage of PUFAs and temperature. Only for Thalassiosira pseudonana was the percentage of either of the PUFAs and nutritionally essential fatty acids (EFAs) also an inverse function of temperature. For T. pseudonana, the percentage of the EFA 22:6ω3 decreased linearly with increasing temperature over the range from 10 to 25° C. For three species, the ratio of unsaturated/saturated FAs was correlated with growth rate when growth rate was controlled by variation in irradiance and temperature. Only for Thalassiosira pseudonana was the ratio of unsaturated/saturated FAs also an inverse function of temperature alone.     

Suicide is not the inevitable outcome of “perpetual” selfing in tetrahymenines collected from natural habitats

A significant fraction of the Tetrahymena clones isolated from natural habitats self (mating occurs within a clone). Early attempts to study such clones failed because stable subclones were rarely, if ever, observed, and isolated pairs all died. Isozyme analysis revealed that these wild selfers were a diverse group; some were very similar to T. australis, a species with synclonal mating type determination and to T. elliotti, shown recently to have a karyonidal mating type system. One originally stable clone of T. australis included some selfing clones after a few years in our laboratory. Other clones manifested unique zymograms. Subclones isolated from 18 selfer strains were heterogeneous. All subclones of several selfers mated massively at each transfer through 100 fissions. Selfing among subclones of other selfers was highly variable or not observed. Although 77% of the pairs isolated died, and 9% of the pair cultures selfed, 15 selfers yielded some viable nonselfing “immature” progeny. Additional immature progeny were obtained by isolating pairs from macronuclear retention synclones. Although some “immature” progeny eventually selfed, most remained stable. Giemsa staining revealed macronuclear anlagen in nearly all mating pairs and some anomalies. Crosses among the F1 progeny clones of the T. elliotti selfers yield viability data comparable to those from crosses among normal strains. Perhaps perpetual selfing is a mechanism of getting rid of deleterious combinations of genes and uncovering better combinations in homozygous state by playing genetic roulette. © 1992 Wiley-Liss, Inc.     

Plant regeneration from the protoplasts of Solanum tuberosum, S. nigrum and S. bulbocastanum

The mesophyll protoplasts were isolated from the Solanum tuberosum (S. tbr) clones of different ploidy level (4x Bzura cv., 2x H-8105, and 2x ZEL-1136) as well as from the wild species: S. bulbocastanum (S. blb, 2x) and two accessions of S. nigrum (S. ngr, 6x). Additionally, the protoplasts were isolated from the cell suspensions of Bzura cv. and H-8105 clone. The conditions of protoplast isolation as well as the media for their culturing and regeneration, were selected and optimized for the studied genotypes. For mesophyll protoplasts, the shooting calli were produced by all the cultured protoclones except that of S. bulbocastanum. The shoots excised from the protoplast-derived calli developed into whole plants in all the studied potato clones but only in one accession of S. nigrum, i.e. S. ngr var. gigantea. As for suspension-cell-derived protoplasts, only H-8105 clone produced the regenerative type of calli, though normal shoots could not be obtained. The regenerative capacity of the protoplasts isolated from leaves and cell suspensions is compared and discussed. We regret to report the death of M. Sc. Maria Borkowska after the completion of this work.     

Changes in the species composition of a thermotolerant community of acidophilic chemolithotrophic microorganisms upon switching to the oxidation of a new energy substrate

Construction and analysis of the 16S rDNA clone libraries was used to investigate the species composition of two thermotolerant communities of acidophilic chemolithotrophic microorganisms (ACM) isolated from the pulp of laboratory reactors used for oxidation of different gold-containing ore concentrates. The first community was formed during oxidation of the pyrite-arsenopyrite ore concentrate from the Kyuchus deposit. The clones of the bacterial component of this community belonged to the genera Sulfobacillus (32 clones) and Leptospirillum (33 clones). The Sulfobacillus clones belonged to three groups: Sb. thermosulfidooxidans, Sb. benefaciens, and Sb. thermotolerans. All Leptospirillum clones were closely related to L. ferriphilum. All clones of the archaeal component belonged to Ferroplasma acidiphilum. The microorganisms of this community were used as inoculum for biooxidation of a different mineral concentrate, the pyrrhotite-containing pyrite-arsenopyrite ore concentrate from the Olympiadinskoe deposit, and the structure of the community formed in the process was investigated. The clones of the bacterial component of the second community also belonged to the genera Sulfobacillus (14 clones) and Leptospirillum (48 clones). The Sulfobacillus clones belonged to the species Sb. thermosulfidooxidans (13 clones) and Sb. thermotolerans (1 clone). All Leptospirillum clones were closely related to L. ferriphilum. All clones of the archaeal component belonged to Ferroplasma acidiphilum. During the adaptation of the community to a new oxidized mineral substrate, both the composition and the ratio of the constituent microbial species changed.     

Examination of diatom type material: Nitzschia delicatissima Cleve,Thalassiosira minuscula Krasske,and Cyclotella nana Hustedt

Type material of Thalassiosira minuscula Krasske and material identified by the respective authors as Nitzschia delicatissima Cleve and Cyclotella nana Hustedt (= Thalassiosira pseudonana Hasle et Heimdal) have been examined in light and electron microscopes. It is suggested that the slide labelled “Nitzschia delicatissima Helder 7/5–97” is accepted as neotype of the species since the original material must be regarded as lost, and that Thalassiosira pseudonana is identical to the diatom isolated by Guillard as clone 3-H. Nitzschia actydrophila Hasle and Thalassiosira monoporocyclus Hasle are rejected as separate species, the first being conspecific with N. delicatissima and the second with T. minuscula. The question is raised whether more emphasis should be laid on iconotypes of small diatom species identifiable only in the electron microscope than on real individuals.     

EFFECT OF THE SINKING RATE OF TWO DIATOMS (THALASSIOSIRA SPP.) ON UPTAKE FROM LOW CONCENTRATIONS OF PHOSPHATE1

The effect of the sinking rate, or rate of medium flow (φ) on the rate of phosphate incorporation (V) by the planktonic diatoms Thalassiosira fluviatilis Hust. and T. pseudonana Hasle & Heimdal in batch and chemostat cultures was determined by passing medium at defined flow rates (0.5–25.0 mm·min^?1) over algae on membrane filters. At concentrations from 1 to 100 μg phosphorus·l^?1 V, increases with increasing velocity of flow, approaching a maximum value (V_m) as described by the empirical relationship: where K_φ is the sinking rate value when V = 1/2 V_m+ V_o and V_o is the uptake at 0 rate of flow. By comparing uptake at controlled flow with uptake in a vigorously stirred medium, the phosphate concentration in the cell boundary layer can be determined. The sinking rate that reduces the phosphate concentration in the boundary layer to half of nominal concentration in the medium is much lower for the larger T. fluviatilis than for T. pseudonana. For both diatoms, it is inversely related to the nominal concentration.     

Riboflavin content of six species of microalgae used in mariculture

Concentrations of riboflavin — a vitamin essential for maricultured animals—were measured in six species of microalgae commonly used in mariculture. These were two diatoms (Chaetoceros gracilis, Thalassiosira pseudonana); two prymnesiophytes (Isochrysis sp. (clone T.ISO),Pavlova lutheri); one chlorophyte (Nannochloris atomus) and one eustigmatophyte (Nannochloropsis oculata). Cultures were analysed during both logarithmic and stationary growth phase.The proportions of riboflavin (µg g^-1 dry weight) during logarithmic growth-phase ranged from 20 (T. pseudonana) to 40 µg g^-1 (Isochrysis sp. T.ISO). With the onset of stationary phase, the proportion of riboflavin increased in all species; the increase was most dramatic in cultures ofC. gracilis, T. pseudonana andN. atomus (2- to 3-fold).Chaetoceros gracilis contained more riboflavin (106 µg g^-1) than all other species (48 to 61 µ g^-1).Despite the differences in the composition of the different microalgae, across both logarithmic and stationary growth-phases, all species should provide a rich source of riboflavin for maricultured animals.Author for correspondence     

KINETICS OF SILICIC ACID UPTAKE AND RATES OF SILICA DISSOLUTION IN THE MARINE DIATOM THALASSIOSIRA PSEUDONANA1,2

Tracer techniques using the stable isotope ³⁰Si were used to measure rates of silicic acid uptake and silica dissolution in silicon replete and silicon depleted populations of 2 clones of the marine diatom Thalassiosira pseudonana Hasle & Heimdal. Uptake kinetics were describable using the Michaelis-Menten equation for enzyme kinetics, and no threshold concentration for uptake was evident. The maximum specific uptake rate of the estuarine clone 3H (0.062–0.092 · h^?1) was higher than that of the Sargasso Sea clone 13-1 (0.028–0.031 · h^?1), but half-saturation constants for uptake by the 2 clones were not measurably different (0.8–2.3 μM for 3H; 1.4–1.5 μM for 13-1). There was little or no light dependence of uptake in populations grown under optimal light conditions prior to the experiment. Exponentially growing populations released silicic acid to the medium by dissolution of cellular silica at rates ranging from 6.5 to 15% of the maximum uptake rate.     

Genetic variation in a host-parasite association: potential for coevolution and frequency-dependent selection

Abstract.— Models of host‐parasite coevolution assume the presence of genetic variation for host resistance and parasite infectivity, as well as genotype‐specific interactions. We used the freshwater crustacean Daphnia magna and its bacterial microparasite Pasteuria ramosa to study genetic variation for host susceptibility and parasite infectivity within each of two populations. We sought to answer the following questions: Do host clones differ in their susceptibility to parasite isolates? Do parasite isolates differ in their ability to infect different host clones? Are there host clone‐parasite isolate interactions? The analysis revealed considerable variation in both host resistance and parasite infectivity. There were significant host clone‐parasite isolate interactions, such that there was no single host clone that was superior to all other clones in the resistance to every parasite isolate. Likewise, there was no parasite isolate that was superior to all other isolates in infectivity to every host clone. This form of host clone‐parasite isolate interaction indicates the potential for coevolution based on frequency‐dependent selection. Infection success of original host clone‐parasite isolate combinations (i.e., those combinations that were isolated together) was significantly higher than infection success of novel host clone‐parasite isolate combinations (i.e., those combinations that were created in the laboratory). This finding is consistent with the idea that parasites track specific host genotypes under natural conditions. In addition, correspondence analysis revealed that some host clones, although distinguishable with neutral genetic markers, were susceptible to the same set of parasite isolates and thus probably shared resistance genes.     

A STATISTICAL CHARACTERIZATION OF GROWTH AMONG CLONES OF SYNURA PETERSENII (SYNUROPHYCEAE)1

Each of four clones from the Synura petersenii complex was grown at different pHs (5.5, 6.5, 7.5, 8.5) in batch culture experiments. Growth response curves and exponential growth rates were compared among clones and pH treatments in order to examine growth trend variation among the clonal groups. The clones were isolated from geographically distant North American localities. The clonal groups represented distinct mating types, an isolate and its subisolate, and S. petersenii- and S. glabra-like scale morphologies. No consistent relationship existed between growth response curve, and culture medium pH. Additionally, the trends across time differed according to clone and pH combination. Pairwise comparisons of linear trends from transformed growth response curves indicated two distinct clonal associations. Although the clonal associations corresponded with the final cell density of the cultures, growth response curves did not correspond with mating type, the parent-isolate and subisolate, or scale morphology. Clones with glabra-like scales had greater growth rates than the clone with petersenii-like scales. The conflicting results generated from growth response curve and growth rate analyses support the concept that S. petersenii and S. glabra form a highly variable, homogeneous grouping.     

Triacylglycerol accumulation and profiling in the model diatoms Thalassiosira pseudonana and Phaeodactylum tricornutum (Baccilariophyceae) during starvation

Although substantial economic barriers exist, marine diatoms such as Thalassiosira pseudonana and Phaeodactylum tricornutum hold promise as feedstock for biodiesel because of their ability to manufacture and store triacylglycerols (TAGs). The recent sequencing of these two marine diatom genomes by the United States Department of Energy Joint Genome Institute and the development of improved systems for genetic manipulation should allow a more systematic approach to understanding and maximizing TAG production. However, in order to best utilize these genomes and genetic tools, we must first gain a deeper understanding of the nutrient-mediated regulation of TAG anabolism. By determining both the yield and molecular species distribution of TAGs we will, in the future, be able to fully characterize the effects of genetic manipulation. Here, we lay the groundwork for understanding TAG production in T. pseudonana and P. tricornutum, as a function of nitrate and silicate depletion. Diatoms were starved of either nitrate or silicate, and TAGs were extracted with hexane from lyophilized samples taken at various time intervals following starvation. The timing of TAG production and the relative abundance of TAGs were estimated by fluorescence spectroscopy using Nile red and the total yield per biomass determined by gravimetric assay. TAGs were analyzed using thin layer chromatography, gas chromatography–mass spectrometry, and electrospray ionization mass spectrometry to identify the major TAG species produced during the growth curve. Under our conditions, the TAG yield from T. pseudonana is about 14–18% of total dry weight. The TAG yield from P. tricornutum is about 14% of total dry weight. Silicate-starved T. pseudonana accumulated an average of 24% more TAGs than those starved for nitrate; however, the chemotypes of the TAGs produced were generally similar regardless of the starvation condition employed.     

INTERACTION (ALLELOPATHY) BETWEEN MARINE DIATOMS: THALASSIOSIRA PSEUDONANA AND PHAEODACTYLUM TRICORNUTUM1

Two marine diatoms were studied singly and in mixed culture. Thalassiosira pseudonana (Hust.) Hade & Heimdal was capable of a higher growth rate (μ_max) than Phaeodactylum tricornutum Bohlin. In two-species batch cultures P. tricornutum took over in the latter portion of the exponential phase, possibly due to allelopathy. The filtrate from this species caused an initial lag phase and a reduced terminal population density for T. pseudonana. Two-species continuous cultures showed verification of these interactions. At high dilution rate (i.e., high growth rate) P. tricornutum washed out when added at low density, whereas T. pseudonana maintained constant cell density. However, when sufficient density of P. tricornutum was added as a contaminant, both species washed out. At a lower dilution rate P. tricornutum increased in density when added and eventually reached a stable population; T. pseudonana then washed out.     

Biochemical genetic markers in sugarcane

Summary Isozyme variation was used to identify biochemical markers of potential utility in sugarcane genetics and breeding. Electrophoretic polymorphism was surveyed for nine enzymes among 39 wild and noble sugarcane clones, belonging to the species most closely related to modern varieties. Up to 114 distinct bands showing presence versus absence type of variation were revealed and used for qualitative characterization of the materials. Multivariate analysis of the data isolated the Erianthus clone sampled and separated the Saccharum spontaneum clones from the S. robustum and S. officinarum clones; the latter two were not differentiated from one another. The analysis of self-progenies of a 2n=112 S. spontaneum and of a commercial variety showed examples of mono- and polyfactorial segregations. Within the progeny of the variety, co-segregation of two isozymes frequent in S. spontaneum led to them being assigned to a single chromosome initially contributed by a S. spontaneum donor. This illustrates how combined survey of ancestral species and segregation analysis in modern breeding materials should permit using the lack of interspecific cross-over to establish linkage groups in a sugarcane genome.     

Genome‐wide single‐nucleotide polymorphism data reveal cryptic species within cryptic freshwater snail species—The case of the Ancylus fluviatilis species complex

DNA barcoding utilizes short standardized DNA sequences to identify species and is increasingly used in biodiversity assessments. The technique has unveiled an unforeseeably high number of morphologically cryptic species. However, if speciation has occurred relatively recently and rapidly, the use of single gene markers, and especially the exclusive use of mitochondrial markers, will presumably fail in delimitating species. Therefore, the true number of biological species might be even higher. One mechanism that can result in rapid speciation is hybridization of different species in combination with polyploidization, that is, allopolyploid speciation. In this study, we analyzed the population genetic structure of the polyploid freshwater snail Ancylus fluviatilis, for which allopolyploidization was postulated as a speciation mechanism. DNA barcoding has already revealed four cryptic species within A. fluviatilis (i.e., A. fluviatilis s. str., Ancylus sp. A–C), but early allozyme data even hint at the presence of additional cryptic lineages in Central Europe. We combined COI sequencing with high‐resolution genome‐wide SNP data (ddRAD data) to analyze the genetic structure of A. fluviatilis populations in a Central German low mountain range (Sauerland). The ddRAD data results indicate the presence of three cryptic species within A. fluviatilis s. str. occurring in sympatry and even syntopy, whereas mitochondrial sequence data only support the existence of one species, with shared haplotypes between species. Our study hence points to the limitations of DNA barcoding when dealing with organismal groups where speciation is assumed to have occurred rapidly, for example, through the process of allopolyploidization. We therefore emphasize that single marker DNA barcoding can underestimate the true species diversity and argue in strong favor of using genome‐wide data for species delimitation in such groups.