Article

ISSN 0006-2979, Biochemistry (Moscow), 2025, Vol. 90, No. 11, pp. 1711-1722 © Pleiades Publishing, Ltd., 2025.

Russian Text © The Author(s), 2025, published in Biokhimiya, 2025, Vol. 90, No. 11, pp. 1830-1842.

1711

Conflicting Phylogenetic Signals between

the Nuclear Ribosomal and Plastome DNA

as Evidence for Hybrid Origin of the Tetraploid

Member of Salicornia (Amaranthaceae s.l.)

Tahir H. Samigullin

1,a

*, Maria D. Logacheva

1,2

, Galina V. Degtjareva

1,3

Dmitry D. Sokoloff

, Svetlana S. Beer

, and Carmen M. Valiejo-Roman

Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia

Center for Molecular and Cellular Biology, Skolkovo Institute of Science and Technology,

121205 Moscow, Russia

Research and Educational Center – Botanical Garden of PeterI, Faculty of Biology,

Lomonosov Moscow State University, 119991 Moscow, Russia

School of Plant Sciences and Food Security, Faculty of Life Sciences, Tel Aviv University,

6997820 Tel Aviv, Israel

Private School “Obninsk Free School”, 249038 Obninsk, Russia

e-mail: samigul@belozersky.msu.ru

Received July 8, 2025

Revised October 6, 2025

Accepted October 8, 2025

Abstract— Species of the genus Salicornia (Amaranthaceaes.l.) are globally distributed and highly salt-tolerant.

They are used as food and for biofuel production. Formation of pure lines through self-pollination, combined

with sporadic cross-pollination, polyploidy, high phenotypic plasticity, and a limited number of diagnostic

characters, significantly complicates taxonomy of the genus. Salicornia (glassworts) is an evolutionarily young

group, where the number of informative substitutions in the traditionally analyzed regions of nuclear and

plastid DNA is insufficient to establish relationships between the species. The very concept of a species in

this genus remains a subject of debate. To clarify relationships among the Eastern European species, we used

high-throughput sequencing to determine sequences and perform phylogenetic analysis of the plastomes of

11 samples representing all major morphotypes of Eastern European glassworts. We also analyzed variability

of the nuclear rDNA external transcribed spacer (nrETS). The sizes of the assembled plastomes ranged from

153,290  bp to 153,504  bp and exhibited a typical architecture with a large single-copy region (84,625-84,797 bp),

a small single-copy region (18,818-18,870 bp), and two inverted repeats (24,898-24,908 bp). Comparison of phy-

logenetic trees reconstructed from all currently available plastome data and nrETS alignments of the same

glasswort accessions revealed a discrepancy in the placement of the tetraploid S.  procumbens subsp. pojar-

kovae and S.  brachiata accessions, which show affinities to different lineages depending on the use of either

plastid or nuclear (nrETS) data. Our results highlight the role of reticulate evolution in the genus Salicornia.

DOI: 10.1134/S0006297925602072

Keywords: plastome, nrETS DNA, phylogenetic signal conflict, polyploidy, hybridization, Salicornia

* To whom correspondence should be addressed.

INTRODUCTION

Hybridization, followed by potential loss of ge-

netic material, is one of the primary mechanisms of

angiosperm evolution. In addition to formation of

auto- or allopolyploids, mitochondrial or plastid intro-

gression is frequently observed [1-7]. These processes

collectively lead to the emergence of groups where

organismal relationships do not fit the standard

model of dichotomous evolution and can be poorly

SAMIGULLIN et al.1712

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

illuminated by classical methods. Additional challeng-

es arise from the often-insufficient informativeness of

the analyzed genome regions of organelles and nu-

cleus, especially in the context of recent evolution-

ary events. This situation is observed in glassworts

(SalicorniaL., Amaranthaceaes.l.) – an evolutionarily

young group where the very concept of a species is a

subject of debate [8-13].

Salicornia is a genus of hygrohalophytic succu-

lent annual plants widely distributed in Eurasia, Af-

rica, North and South America, Australia, and New

Zealand [14-19]. In Europe and America, Salicornia

species are cultivated and consumed as food; oil ex-

tracted from their seeds is similar in fatty acid com-

position to safflower oil [20,  21]. Glassworts are also

used for biofuel production on coastal saline lands

where traditional crops do not thrive and are consid-

ered promising for phytoremediation [22].

Taxonomic boundaries of the genus Salicornia

are highly controversial due to the high phenotypic

plasticity, scarcity of diagnostic morphological traits,

simplified morphology, polyploidy, and formation of

inbred lines with minor but presumably fixed phe-

notypic differences [10, 23]. It is no coincidence that

the traditional division of European Salicornia spe-

cies into sections was based on chromosome num-

ber  [24]. Typically, Salicornia species are either dip-

loid (2n  =  18) or tetraploid (2n  =  36) [25-30], although

a triploid form of Salicornia veneta Pign. et Lausi was

found in Italy [31], and a decaploid species, Salicor-

nia altaica Lomon., was discovered in the southeast-

ern Altai and northwestern Mongolia [32]. Tetraploids

generally differ from diploids by having larger an-

thers, a greater ratio of the length of the central flow-

er in the dichasium to the lateral flower, longer and

thicker inflorescences, and fewer sterile metamers

on the main shoot [24, 33]. However, the ranges of

morphological traits in diploids and tetraploids over-

lap, making delineation of the species within the dip-

loid and tetraploid lineages an extremely challenging

task [10].

In delineation of the Salicornia species, there is

a growing interest in molecular markers. Using var-

ious non-coding regions of nuclear and plastid DNA,

as well as microsatellite loci, it has been shown that

reticulate evolution is common among the Western

European species, tetraploids have an allopolyploid

origin, and there is insufficient information on both

diploids and tetraploids to understand their interre-

lationships [8-13, 33-37].

A comprehensive phylogenetic study of Salicor-

nia, covering all major regions of its geographic range

based on nucleotide sequences of the nuclear ribo-

somal external transcribed spacer (nrETS), revealed

widespread parallel evolution of morphological traits

(combined with strong phenotypic plasticity) [10].

Some species of the genus have been proposed to be

considered cryptic, as they cannot be distinguished

using morphological traits [23].

In this study, we focused on Eastern European

representatives of the genus Salicornia. Compared

to the Western Europe, a relatively small number of

species occur here, simplifying the study of reticulate

evolution. Tzvelev [38] distinguished four Salicornia

species in Eastern Europe: S.  perennans Willd., S.  eu-

ropaea L., S.  borysthenica Tzvel., and S. pojarkovae

N. Semen. Later, Beer and Demina [39] described the

species S. heterantha Beer et Demina from the Ros-

tov Region (Kuma-Manych Depression), which differs

from all other species of the genus by the complete

fusion of the perianth tube of the central flowers of

the dichasium and the main axis of inflorescence. Ac-

cording to Tzvelev, two diploid species, S.  perennans

and S. europaea, grow in the southern and eastern

regions of Eastern Europe (S.  perennans) and on the

coasts of the White and Baltic Seas (S.  europaea); the

tetraploid species S.  pojarkovae is found on the coast

of the White Sea, and S.  borysthenica is morpholog-

ically similar to the Western European tetraploid

S.  dolichostachya Moss, but Tzvelev had no data on

the chromosome number for this species. Analysis of

nrETS sequences [10, 23] placed the diploid species

S.  heterantha and S.  borysthenica in one clade with

the tetraploid S.  pojarkovae and some other tetra-

ploids (including S.  dolichostachya). Sukhorukov and

Akopyan [40] placed S.  heterantha among the syn-

onyms of S. perennans Willd. Sukhorukov [41], using

the treatment suggested by Kadereit etal. [23] regard-

ing the species boundaries, synonymized S.  heteran-

tha with S.  procumbens subsp. procumbens. Chatre-

noor and Akhani [33], using new materials, support

the species status of S.  heterantha; however, their

work does not use all available GenBank data.

Based on the molecular data, the boundaries of

European Salicornia species have been revised, and

new nomenclatural combinations have been proposed

[23]. Most species have been transferred to the rank

of subspecies, while others have been synonymized.

According to the system of Kadereit et al. [23], four

taxa occur in Eastern Europe: Salicornia perennans

Willd. subsp. perennans, S.  procumbens Sm. subsp.

procumbens (incl. S.  borysthenica Tzvel.), S.  procum-

bens subsp. heterantha (S.  S.  Beer & Demina) G. Ka-

dereit & Piirainen, and S. procumbens subsp. pojar-

kovae (Semenova) G. Kadereit & Piirainen. It should

be noted that molecular data correlate well with

geographic distribution but often do not correspond

to any morphological traits. Therefore, S.  perennans

and S.  europaea should be considered cryptic spe-

cies in the new concept [23, 42]. Molecular data sug-

gest that material from the north and northwest of

European Russia, previously identified as S.  europaea

HYBRID ORIGIN OF A TETRAPLOID IN THE GENUS Salicornia 1713

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

(e.g., in Tzvelev’s work [38]), should be attributed

to S. perennans [23].

The aim of this article is to clarify relationships

of the Eastern European species using an expanded

sample and additional molecular data. For the first

time, we used high-throughput sequencing to deter-

mine the plastome sequences of 11 Salicornia sam-

ples from Eastern Europe. We analyzed these data in

comparison with the nrETS data. We studied material

that, according to the system of Kadereit et  al.  [23],

corresponds to S.  perennans, S.  procumbens subsp.

pojarkovae, S.  procumbens subsp. procumbens, and

S.  procumbens subsp. heterantha. For the description

and discussion of the results, we followed the system

of Kadereit etal. [23]. Our results allow us to discuss

the possibility of hybridization among the represen-

tatives of the genus Salicornia.

MATERIALS AND METHODS

Plant material. Samples were collected from

natural habitats in the southern (Kuma-Manych De-

pression, Black Sea and Caspian Sea lowlands) and

northern (White Sea coast) parts of Eastern Eu-

rope. Detailed information on the studied samples,

GenBank identifiers, and references to the samples

already sequenced in previous studies are provided

in the Online Resource 1.

DNA extraction. DNA was extracted using a

NucleoSpin Plant II kit (Macherey-Nagel, Germany) ac-

cording to the manufacturer’s protocol. Quantity and

quality of DNA were assessed using a Qubit dsDNA

HS Assay Kit (Thermo Fisher Scientific, Waltham, MA,

USA) and electrophoresis in 0.8% agarose gel.

Sequencing, plastome assembly, annotation,

and alignment. DNA from each sample was frag-

mented using a Covaris S220 ultrasonicator (Covaris,

USA). End repair, adenylation, and adapter ligation

followed by PCR were performed using an Illumina

TruSeq DNA Sample Prep Kit (Illumina, USA) accord-

ing to the manufacturer’s instructions. Libraries were

sequenced using an Illumina HiSeq 2000 sequencer

with a read length of 101 bp from each end of the

fragment. Read trimming was performed using Trim-

momatic [43]. De  novo plastome assembly was car-

ried out using the CLC Genomics Workbench v5.5

software package. Contigs with extensive similarity

to the known plastid genomes were joined at over-

lapping ends. Correctness of the assembly was ver-

ified by mapping trimmed reads to the assembled

plastomes. Automatic annotation was performed us-

ing the online CpGAVAS program [44], followed by

manual correction. All tRNAs were verified using

tRNAscan-SE v2.0 [45] and ARAGORN [46]. The plas-

tome map was visualized using OGDRAW [47].

Plastome sequences were aligned using MAFFT

v7.017 [48] and then manually corrected in BioEdit [49].

Amplification, sequencing, assembly from

GenBank raw data, and alignment of nrETS. Am-

plification of the 3′ end of nrETS was performed

using combination of primers ETS-Salicornia-5′-GTC-

CCTATTGTGTAGATTTCAT-3′ and 18S-II rev. 5′-CTCTA-

ACTGATTTAATGAGCCATTCGCA-3′ [10, 50]. PCR prod-

ucts were purified using a Cleanup Mini Kit for PCR

product purification (Evrogen, Russia). Direct sequenc-

ing of both strands of the spacer was performed at

the Genome Center for Collective Use (Russian Acad-

emy of Sciences).

To correctly compare topologies of the plastid

and ETS trees, we assembled genomes from GenBank

sequence read archive data for six glasswort samples

with known plastid genomes (as described above, ex-

cept for PacBio HiFi sequencing data, for which we

used the Canu assembler [51]) and obtained nrETS se-

quences (Online Resource1), which were also used for

analysis. DNA sequences were aligned using MUSCLE

[52] and then manually corrected in BioEdit [49].

Intragenomic polymorphism of nrETS. To de-

tect intragenomic polymorphism, the trimmed reads

were mapped to nrETS sequences. Polymorphism in

eleven Salicornia samples was assessed visually using

Tablet (version 1.17.08.17) [53].

Phylogenetic analysis. A set of ten completely

assembled and one partially assembled plastomes

was supplemented with six sequences from the Gen-

Bank database (Online Resource 1). As an outgroup,

we used plastomes of Suaeda malacosperma Hara

(NC_039180), Bienertia sinuspersici Akhani (MT316307),

Kalidium gracile Fenzl (ON149858), and Salicornia fru-

ticosa (L.) L. (NC_066030). Phylogenetic analysis in-

cluded 20 plastome sequences in total; one inverted

repeat was removed. Gap-rich positions (more than

50%) were excluded from the analysis. Phylogenetic

analysis was performed using the Bayesian approach

in MrBayes v3.2.6 [54, 55] with the GTR+Γ model,

which was selected as the best according to the

Akaike Information Criterion (AIC). For 16 Markov

chains (4 parallel runs of 4 chains each), 20,000,000

steps were set, and every thousandth tree was sam-

pled. The first thousand trees were discarded, and

the remaining trees were used to construct a major-

ity-rule consensus tree and estimate posterior proba-

bilities of the internal branches.

The matrix of aligned nrETS sequences was also

analyzed using the Bayesian approach with the same

settings as for the plastome data; in polymorphic

positions, the predominant (major) base was indicat-

ed. As an outgroup, we used sequences of Microcne-

mum coralloides (Loscos & J. Pardo) Buen (EF433589),

Arthrocnemum macrostachyum (Moric.) K.  Koch

(EF433587), and S. fruticosa.

SAMIGULLIN et al.1714

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

RESULTS AND DISCUSSION

Characteristics of Salicornia plastomes. We

performed shallow genomic sequencing of 11 glass-

wort samples representing all major morphotypes of

the genus Salicornia found in Eastern Europe. Com-

plete plastome sequences were assembled for ten

samples: one sample of S.  procumbens subsp. pro-

cumbens (1209); two samples of S.  procumbens subsp.

pojarkovae (1196 and 1194); three samples of S.  pro-

cumbens subsp. heterantha (1213, 1214, and 1219);

and four samples of S.  perennans (1197, 1195, 1211,

and 1226). The assembled and annotated sequences

were deposited in the GenBank database; accession

numbers are provided in the Online Resource  1. For

sample 1208, approximately 98% of the plastome

length was assembled, so we considered it accept-

able to use this incomplete plastome in phylogenet-

ic analysis. Sizes of the fully assembled plastomes

ranged from 153,290 bp to 153,504 bp and exhibited

Fig. 1. Circular map of the Salicornia plastid genome. Genes located inside and outside the circle are transcribed clockwise

and counterclockwise, respectively. Pseudogenes are marked with “ψ.” LSC, large single-copy region; SSC, small single-copy

region; IRa and IRb, inverted repeats. Photo: S. procumbens subsp. pojarkovae, by G. V. Degtjareva.

HYBRID ORIGIN OF A TETRAPLOID IN THE GENUS Salicornia 1715

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

Fig. 2. Position of the boundaries of inverted repeats in the plastomes of glassworts. S.  pr. – S. procumbens; JLB and JSB –

boundaries of IRb with LSC and SSC, respectively; JSA and JLA – boundaries of IRa with SSC and LSC, respectively.

a typical architecture with a large single-copy region

(LSC; 84,625-84,797  bp), a small single-copy region

(SSC; 18,818-18,870  bp), and two inverted repeats (IR;

24,898-24,908 bp) (Fig. 1).

After annotation, 113 unique genes were identi-

fied, including 30 tRNA genes, 4 rRNA genes, and 79

protein-coding genes (listed in the Online Resource2).

The plastomes contained three pseudogenes: rpl23,

ycf1, and rps19, the latter two being truncated frag-

ments of the corresponding genes at the boundaries

of the inverted repeats. Pseudogenization of rpl23 is

observed in all representatives of the Amaranthace-

ae family; multiple cases of independent pseudog-

enization of this gene in plastomes have also been

noted in other families of the order Caryophyllales

[56, 57]. Composition and order of the genes in the

newly assembled and previously known Salicornia

plastomes do not differ from each other, and no any

differences were found in the arrangement of homol-

ogous genes (synteny) compared to other autotrophic

representatives of the order Caryophyllales. In the

plastid genomes of glassworts, as in the overwhelm-

ing majority of the species in the core Caryophyllales,

the intron of the rpl2 gene is absent [57, 58]. Over-

all, it can be stated that the plastomes of glassworts

are highly conserved in structure, and the differenc-

es in genome sizes of Salicornia samples are mainly

determined by the variability of intergenic regions:

indels of various lengths are present in many spacer

sequences, with the most variable being the psbA–

trnH and rpl33–rps18 spacer regions.

Boundaries of the inverted repeats with the large

single-copy region in all glassworts intersected the

rps19 (LSC/IRb) and trnH-GUG (LSC/IRa) genes and

were located identically, while positions of the SSC/

IRb (between the ycf1 gene fragment and the ndhF

gene) and SSC/IRa (within the ycf1 gene) boundaries

differed slightly (Fig. 2).

SAMIGULLIN et al.1716

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

Table 1. Intragenomic polymorphism of nrETS sequences in glassworts

Samples

Alignment positions

171 274 376 414 456 469

S. perennans subsp. perennans

2n = 18

1211

C A T C A G

1226 C A T C A G

1195 C A T C A G

1197 C A T C A G

S. procumbens subsp. pojarkovae

2n = 36

1194

C A T C A T

1196 C A T C A T

S. procumbens subsp. heterantha

2n = 18

1219

A/C A T C A T

1213 C/A G/A T C A T

1214 C/A G/A T C A T

S. procumbens subsp. procumbens

2n = 18

1209

C/A G/A T C A T

1208 A/C A/G T C A T

S. europaea 1 C A T C R G

Salicorniasp. variant 2 C A T C A G

Salicorniasp. variant 1 C AAT A T

S. sinus-persica MW679260 C AAT A T

S. persica subsp. iranica EF433645 C AAT A T

*S.persica subsp. persica MW679241 C A WY A K

*S.persica subsp. iranica MW679243

C A WY A K

Note. In polymorphic positions, the predominant (major) variant is listed first. R = G or A, W = A or T, Y = C or T, K = GorT.

* Sequences not included in phylogenetic analysis.

Intragenomic polymorphism and phylogenetic

analysis of nrETS. Mapping of the reads obtained

directly by us using high-throughput sequencing to

the nrETS sequence revealed intragenomic polymor-

phism at two positions (171 and 274) in five diploid

southern samples: S.  procumbens subsp. heterantha

(samples 1219, 1213, 1214) and S.  procumbens subsp.

procumbens (samples 1208 and 1209). Polymorphic

positions are presented in Table 1 along with other

characteristic substitutions.

Notably, in the assembled contigs of the Salicor-

niasp. sample (data from the GenBank sequence read

archive, PacBio HiFi sequencing, SRR24425728) we

identified two variants of ETS sequences differing at

three alignment positions: 376, 414, and 469 (Table 1).

This sample was deposited in the database as S.  eu-

ropaea, but the detected intragenomic polymorphism

differs from that observed in S.  europaea and is more

consistent with the sequences of S.  persica subsp.

iranica (GenBank ID: MW679243) or S. persica subsp.

persica (GenBank ID: MW679241), which calls into

question the species identification of this sample (see

also Jamdade et al. [59]).

No intragenomic polymorphism was detected

in the diploid northern S.  perennans (samples 1195

and 1197), southern S.  perennans (samples 1211 and

1226), northern tetraploids S.  procumbens subsp.

pojarkovae (samples 1194 and 1196), or the tetra-

ploid S.  bigelovii. Concerted evolution of the nucle-

ar ribosomal operon sequences has long been well

known [60], so heterogeneity of the ribosomal op-

eron sequences is generally considered character-

istic of the recent hybrids. Since there have been

no suggestions about the hybrid nature of these

S. procumbens subsp. heterantha and S.  procumbens

subsp. procumbens samples, it would be premature

HYBRID ORIGIN OF A TETRAPLOID IN THE GENUS Salicornia 1717

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

to explain the detected polymorphism by recent hy-

bridization, especially since potential second paren-

tal form is clearly not represented in our analysis.

Nevertheless, such explanation cannot be ruled out at

this stage, nor can be the alternative hypothesis of in-

complete unification of inherited ancestral polymor-

phism (see also the discussion in Yurtseva etal. [61]).

Phylogenetic analysis. Phylogenetic tree obtained

from the Bayesian analysis of 48 nrETS sequences

is presented in Fig.  3a. The tree shows three main

groups of Eastern European Salicornia, with samples

grouped mainly by ribotypes (nrETS haplotypes).

S.  procumbens forms a well-supported monophyletic

group (posterior probability PP  =  1), within which

two clades of southern diploid S.  procumbens subsp.

heterantha with S.  procumbens subsp. procumbens

are distinguished. One clade consists of six sequenc-

es with ribotype 20 (here and below, ribotypes are

indicated according to Kadereit et al. [23]), and the

other consists of four sequences of two ribotypes dif-

fering by a single nucleotide substitution, with both

clades containing representatives of two subspecies.

In the unresolved part of this monophyletic group,

eight northern tetraploids of S.  procumbens subsp.

pojarkovae (ribotype 21) are located. Sequences of the

S.  perennans representatives do not form a monophy-

letic group: samples from the southern regions with

ribotype 9 formed a weakly supported (PP  =  0.87) un-

resolved clade, while representatives of the northern

populations of S.  perennans with ribotype 2 are locat-

ed in the group combining sequences of S.  brachiata,

S.  europaea, and S.  aff. europaea with variant 2 ETS

of Salicornia sp. (PP = 0.98). Details of relationships

within this group remain unclear, as does the posi-

tion of the S.  perennans 484 sample with ribotype 11

and the trio of S.  bigelovii representatives. In a sep-

arate clade, also of unclear position relative to other

glasswort species, the sequences of S.  sinus-persica,

S.  persica, and variant 1 ETS of Salicornia sp. are

combined (PP  =  0.99). Overall, the arrangement of

glasswort samples does not contradict the previous-

ly presented results obtained from the analysis of a

similar set of samples [10, 12, 23, 62].

In the consensus tree obtained from the Bayes-

ian analysis of plastid data, the plastomes of glass-

worts assembled by us were distributed into several

well-supported clades (Fig.  3b). The plastomes of five

samples of diploid southern S.  procumbens subsp. het-

erantha and S.  procumbens subsp. procumbens (1208,

1209, 1213, 1214, and 1219) formed a well-supported

but poorly resolved monophyletic group (as in the

nrETS tree). The plastome sequences of the northern

diploid S.  perennans (1195 and 1197) and S. euro-

paea 1 formed a group sister to the southern diploid

S.  perennans (1211 and 1226) plus S. brachiata. Two

samples of the tetraploid S.  procumbens subsp. pojar-

kovae (1196 and 1194) cluster with the plastome se-

quence of Salicornia sp. with low support (PP =  0.96)

and are more closely related to the plastomes of S. pe-

rennans than to other plastomes of S. procumbens.

The plastome with the identifier OL449699, deposited

in GenBank as S. europaea [63], turned out to be a

sequence of S.  bigelovii or a very closely related taxon,

which is also confirmed by phylogenetic analysis of

the ETS sequence of this sample.

Notably, the plastid tree shows a good resolution

in the relationships of S.  perennans samples that re-

main unclear in the nrETS tree, and at the same time,

within the species S.  procumbens, it shows slightly

worse resolution, revealing only closer relationship

between the samples S.  procumbens subsp. procum-

bens 1208 and S. procumbens subsp. heterantha 1219.

However, if the lack of resolution in the nrETS tree

is due to the lack of variability (for example, nrETS

sequences of samples 1213, 1214, and 1209 are iden-

tical and belong to the same ribotype), then the res-

olution in the plastid tree is associated with the lack

of informative substitutions, since all analyzed plas-

tomes differ (both in length and sequence), including

those of samples with nrETS of the same ribotype.

The plastome variability we identified is an example

of the widespread non-strict conservation of plas-

tome sequences in the plants within a species [64],

the scale of which remains to be assessed, but it is

already clear that the plastome data may be promis-

ing in resolving questions of relationships at least for

some glasswort species.

Comparison of phylogenetic trees reconstructed

from the plastid and nrETS alignments reveals dis-

crepancy in the placement of a pair of tetraploid

S.  procumbens subsp. pojarkovae samples (1194 and

1196), which show closer relationship with the dip-

loid S.  procumbens in the nrETS tree but cluster with

the diploid S.  perennans, S.  brachiata, and S. euro-

paea in the plastid tree. It can be definitively stated

that the tetraploid glasswort lineage studied in this

work should be interpreted as a hybrid, and hybrid-

ization occurred between the lineage represented in

our study by the southern diploids of S.  procumbens

and the lineage combining S.  perennans, S.  europaea,

S.  brachiata, and possibly S.  persica. Further research

using expanded sampling with involvement of plas-

tome data is required for more accurate determina-

tion of the parental forms of these hybrids. Differenc-

es in the revealed relationships for S.  brachiata in the

plastid (representatives of the southern populations

of S.  perennans) and ETS (S.  europaea and represen-

tatives of the northern populations of S.  perennans)

trees also raise suspicions about the hybrid nature of

this taxon, but additional research is required to con-

firm this assumption and identify the parental forms.

SAMIGULLIN et al.1718

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

Fig. 3. Phylogenetic tree reconstructed using the Bayesian approach for nrETS (a) and plastid (b) sequences. Posterior prob-

abilities exceeding 0.85 are indicated. Branch lengths are proportional to the number of substitutions per site, according

to the corresponding scale. Samples with known chromosome numbers are underlined; samples with plastomes sequenced

by us are in bold. Ribotypes are indicated according to the numbering proposed by Kadereit et al. [23].

HYBRID ORIGIN OF A TETRAPLOID IN THE GENUS Salicornia 1719

BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025

CONCLUSION

Our data indicate the need for further study of

microevolution of the genus Salicornia with broad-

er use of plastome data and further improvement

of glasswort systematics. In particular, the revealed

position of the S.  procumbens subsp. procumbens

samples relative to S.  procumbens subsp. heterantha

samples calls into question the criteria for distin-

guishing subspecies within S.  procumbens. In addi-

tion to solving purely taxonomic issues (which will

remain a subject of debate in such complex group),

the study of genetic regulation and adaptive value of

the main trait underlying the distinction of S.  heter-

antha – fusion of the perianth of the central flower

of the dichasium with the inflorescence axis [65] –

is of interest. The fact that samples possessing this

trait do not form a single clade suggests a relatively

simple genetic regulation of this trait. With the fas-

cinating progress in plant evo-devo and considering

fast life cycle of glassworts, it is possible to identify

the genes associated with this trait, which will con-

tribute to understanding the patterns of organ fusion

regulation in angiosperm flowers [66].

Abbreviations

nrETS

nuclear ribosomal external transcribed

spacer

Supplementary information

The online version contains supplementary material

available at https://doi.org/10.1134/S0006297925602072.

Acknowledgments

We are grateful to G. Kadereit for useful discussions

and to the reviewers for valuable comments.

Contributions

M. D. Logacheva, C. M. Valiejo-Roman, and S. S. Beer –

collection of material and experiments; T. H. Sami-

gullin, D. D. Sokoloff, G. V. Degtjareva, and M. D. Lo-

gacheva – processing and discussion of experimen-

tal results; C. M. Valiejo-Roman, T. H. Samigullin, and

D. D. Sokoloff – writing the text; M. D. Logacheva,

C. M. Valiejo-Roman, G. V. Degtjareva, D. D. Sokoloff,

S. S. Beer, and T. H. Samigullin – editing the article

text.

Funding

The study was conducted under the state assignment

of Lomonosov Moscow State University.

Ethics approval and consent to participate

This work does not contain any studies involving

human and animal subjects performed by any of the

authors.

Conflict of interest

The authors of this work declare that they have

noconflicts of interest.

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