Subsection Truncata and subsection Emarginata are genetically isolated from one another, as shown by sterility of hybrids between them. On the other hand, hybrids between Emarginata species are fertile or partially fertile. The F1 hybrid between Ae. longissima and Ae. sharonensis and the reciprocal hybrid had an 81–85% seed set (Tanaka 1955a; Ankori and Zohary 1962), that between Ae. sharonensis and Ae. bicornis had a 62% seed set (Tanaka 1955a), that between Ae. bicornis and Ae. longissima had a 40% seed set (Waines G, personal communication), and that between Ae. longissima and Ae. searsii had a 35% seed set in one hybrid combination and 7% in another (Feldman et al. 1979).

Ae. speltoides differs from the four species of subsection Emarginata in several additional important features: (i) Ae. speltoides is the only predominantly cross-pollinated species in the section. (ii) Its distribution area is in the central part of the genus area, i.e., in and around the Fertile Crescent, on terra rossa or alluvial soils. In contrast, the species of subsection Emarginata reside in the southern part of the western wing of the Fertile Crescent, with Ae. sharonensis and Ae. bicornis growing on sandy soils, and Ae. longissima on red sandy loam in the coastal plain, on sand derived from Nubian sandstone and on grey calcareous steppe soil or loess in the inland steppe and desert. Ae. searsii grows on terra rossa or basalt soil (Feldman and Kislev 1977; Kimber and Feldman 1987).

The five Sitopsis species differ in their genome size (Eilam et al. 2007), that is primarily due to independent post-speciation amplification of transposons (Yaakov et al. 2013). The sum of repetitive sequences in the different species ranges from 85.99 to 89.81% (Li et al. 2022). Ae. speltoides has the smallest genome, Ae. bicornis and Ae. searsii have a larger genome, and Ae. sharonensis and Ae. longissima have the largest genome.

Ae. speltoides is the only Sitopsis species found in the entire Fertile Crescent and in most parts of Turkey, with both varieties (speltoides and ligustica) often occurring sympatrically and with similar ecological tendencies (Hammer 1980; Kimber and Feldman 1987). Notably, the area of the other four species of the section (Ae. bicornis, sharonensis, longissima, and searsii) occurs within a very limited region in the southeastern corner of the Mediterranean, which marginally overlaps with the southwestern part of the area of Ae. speltoides. It seems as if these species replace Ae. speltoides in the southern climatically and edaphically special environments. Of these, the closely related Ae. sharonensis and Ae. bicornis are an example of bioregional vicariance (Zohary 1962), with the former confined to the coastal plain of the Mediterranean region in Israel and southern Lebanon, and the latter replacing it southwardly up to Lybia and also spreading towards the Saharo-Arabian region in desert sands of the Israeli Negev and Sinai Peninsula. Ae. longissima occurs in two different phytogeographical regions, the Israeli and Egyptian coastal plain (Mediterranean region) and the Israeli Negev, southern Jordan and Sinai Peninsula (Irano-Turanian and Saharo-Arabian regions). Ae. searsii grows in sub-Mediterranean areas in Israel, Jordan, southwestern Syria and southeastern Lebanon and also occurs in the high elevations of the Israeli Negev and southern Jordan. It should also be noted that the distribution of Ae. searsii, similar to Ae. vavilovii, has only recently been defined, and may be larger than hitherto supposed.

### 9.4.2 Aegilops speltoides Tausch

<IP_ADDRESS> Morphological and Geographical Notes Ae. speltoides Tausch [Syn.: Triticum speltoides (Tausch) Gren. ex Richter; Sitopsis speltoides (Tausch) Á. Löve] is a predominantly allogamous, annual plant, with 40–70-cm high culms (excluding spikes), and leaves that are usually hairy, and sometimes pendant. Its spike is linear, narrow, tapers to the tip, either two- or one-rowed, and is (6-) 7-11(- 15)-cm long (excluding awns). At maturity, its spike disarticulates above the basal rudimentary spikelet, either into single spikelets, with the rachis internode immediately below them (wedge-type dispersal unit), or falling entire as a unit (umbrella-type dispersal unit). Spikelets 7–11, 11–15 mm long, lanceolate or linear, sessile in hollows of rachis, and longer or shorter than the adjacent rachis internode. Each contains 4–8 (usually 4–6) florets, with the upper 1–3 being sterile. There is usually 1 rudimentary spikelet at the base of the spike. Glumes are 5–7 mm long, truncate at the apex, asymmetrical, somewhat keeled, and about 2/3 the length of the lemma. Lemmas are 7–10 mm long and boat-shaped, those of lateral spikelets have either short triangular awns or are not awned at all, while those of terminal spikelets are always have longer awns than those on the lateral spikelets. Anthers are 5–6 mm long. The caryopsis is adherent to lemma and palea (Fig. 9.1a, b).

Ae. speltoides has a median-sized distribution in the central region of the distribution of the genus. It is an eastern Mediterranean element, extending into the steppical (Irano-Turanian) region. It occupies primary and secondary habitats. Ae. speltoides grows in Israel (in the Coastal Plain and Esdraelon Plain on humid, alluvial soil; in Mt. Carmel and western slopes of the Samaria Mts., on terra rossa and Rendzina soils), Jordan, Lebanon, Syria, Turkey, northern Iraq and northwestern Iran. It is a weed in central and western Turkey, Greece, Bulgaria and possibly also Crimea. The center of variation of this species is in north Syriasoutheastern Turkey, which is likely the center of its origin.

Ae. speltoides grows sympatrically with Ae. caudata, Ae. umbellulata, Ae. triuncialis, Ae. geniculata, Ae. neglecta, Ae. biuncialis, Ae. peregrina, Ae. cylindrica, wild T. monococcum (ssp. aegilopoides), T. urartu, wild T. timopheevii and wild T. turgidum, and allopatrically with Ae. sharonensis, Ae. longissima and Ae. searsii in its southern distribution area and with Ae. tauschii in its northeastern distribution area.

Ae. speltoides has limited morphological variation, mainly in spike characteristics. It contains two main morphological types that differ markedly in the structure of their fruiting spike and consequently in their mode of seed dispersal. Taxonomists (see van Slageren 1994) regard these two types as varieties, var. speltoides Tausch and var. ligustica (Savign.) Fiori in Fiori & Paoletii.

Var. speltoides [Syn.: Ae. speltoides ssp. speltoides Hammer; Ae. speltoides var. aucheri (boiss.) Fiori in Fiori & Paoletii; Ae. aucheri Boiss.] is characterized by a one-rowed, long and cylindrical spike, with relatively widely interspersed spikelets. Only the apical spikelet is awned and rarely has awns on some lateral spikelets and then shorter and thinner than the awns on the lateral spikelet. The rachis of the spike is tough, except for a brittle node at its base, above a rudimentary spikelet. The spike falls at maturity as a unit (umbrella-type dispersal unit) (Fig. 9.1b).

Var. ligustica [Syn.: Ae. ligustica (Savign.) Coss.; Ae. speltoides ssp ligustica (savign,) Zhuk.; Agropyron ligusticum Savign.] is characterized by a denser, two-rowed spike, in which the lateral spikelets are also awned; awns of terminal spikelet are somewhat sturdier than those of lateral spikelets. The rachis is brittle at every joint, so that the mature spike disarticulates into single spikelets, each with the rachis internode immediately below it (wedge-type dispersal unit). Each individual spikelet thus serves as an independent dissemination unit; it is mechanically adapted to insert itself into the ground and usually buries itself soon after detachment (Fig. 9.1a). The two varieties grow, in most sites, in mixed stands and are cross-fertile; few intermediates are usually found in mixed populations. Presumably, the ligustica form, having the wedge-type dispersal unit typical of many Triticineae species, is the ancestral form. Alternatively, the speltoides form, having one-rowed spike similar to that of several Elymus species, may be the ancestral type.

The morphological differences between var. speltoides and var. ligustica are restricted to the fruiting spike alone. In all other traits, such as vegetative characters and phenological behavior, the types are strikingly similar to each other (Miczynski 1926; Schiemann 1928; Eig 1929a). In addition, F1 hybrids between the two types are fully fertile and show complete pairing of chromosomes at meiosis (Kihara and Lilienfeld 1932; Sears 1941b; Zohary and Imber 1963). Further indications of the close relationship between speltoides and ligustica were obtained from the usual occurrence of mixed stands of the two types (Eig I929a). There are wide fluctuations in the proportions between ligustica and speltoides in different sites and in different plant formations, but both types were almost universally present in the sites examined. In many stands, ligustica and speltoides plants were, more or less, evenly mixed. Yet, ligustica plants predominated the stands in higher altitudes and speltoides in warmer sites (Zohary and Imber 1963). These observations led to the conclusion that mixed populations are the rule; speltoides and ligustica are apparently spatially interconnected almost throughout their distribution area (Zohary and Imber 1963).

Rare intermediate types, e.g., plants with a laterally awned but tough rachis (var. polyathera according to Eig 1929a) or also, very occasionally, plants with brittle rachis but laterally awnless spikes, occur in very low frequencies (less than 1%) in mixed speltoides-ligustica populations (Zohary and Imber 1963). Progeny of speltoides x ligustica crosses revealed that the main differences in spike morphology are inherited in an almost monohybrid Mendelian fashion (dominance of ligustica over speltoides in F1, and 3:1 segregation in F2) (Miczynsky 1926; Schieman 1928; Kihara and Lilienfeld 1932). These data were first interpreted as indicating the presence of only a single pleiotropic gene, but Sears (1941b), based on occasional occurrence of recombinants or intermediate plants, proposed the operation of a block of closely linked genes. This block should possess at least three genes: one determining the length of rachis internodes (short in ligustica bringing about two-rowed spikes and long in speltoides leading to one-rowed spikes); a second gene that determines awns on the lateral spikelets; and a third gene that determines the type of dispersal unit. Luo et al. (2005) mapped the closely linked genes controlling the ligustica/speltoides spike dimorphism to the centromeric region of chromosome 3S. (the genome symbol of Ae. speltoides is S). The location near the centromere, being a chromosome region with rare recombination events, may explain the rare recombination between these three genes.

Progeny tests performed in plants sampled from natural mixed populations, indicated that speltoides and ligustica plants are genetically interconnected by virtue of their mating system of predominantly cross-pollination. Several ligustica plants, that apparently were F1, segregated into 3 ligustica: 1 speltoides and several speltoides plants, that were presumably pollinated by ligustica, yielded ligustica progeny (Zohary and Imber 1963). Accordingly, Zohary and Imber (1963) suggested not to regard speltoides and ligustica types as two independent varieties, but as two constituents of dimorphic populations. The mixed stands of the two types are looked upon as a case of genetically determined fruit dimorphism, where two types of seed dispersal apparatus function in the same population in a complementary way (Zohary and Imber 1963). In the speltoidesligustica pair, the two kinds of dispersal units are borne by different individuals: two types of plants, which differ with regard to their fruit structure, form a common population. This is an instance of genetically determined fruit dimorphism, with a population as its unit of operation.

A model of fruit dimorphism can account for the peculiar mode of inheritance of the many differences between the two types: they are inherited in an almost monohybrid fashion, with only rare cases of recombinants. As already proposed by Sears (1941b), this mode of inheritance suggests the presence of a single block of closely linked genes. A system based on such a single block is essential for the establishment of population dimorphism. The parallel evolvement of fruit dimorphism in two Sitopsis groups, ligustica vs. speltoides on the one hand, and bicornis-sharonensis versus longissima-searsii on the other hand, is an interesting evolutionary phenomenon. The genetic change(s) determining the speltoides and the longissima-spike morphology occurred independently in the two groups, or alternatively, happened once and genes of this system were transferred from one group to the other via interspecific introgressive hybridization. However, it seems there is a difference between the gene system in the two groups; while the genes in ligusticaspeltoides are closely linked, those in sharonensis-longissima are not linked, and segregate independently from one another, as is indicated in the scatter diagram of F2 artificial generation in Ankori and Zohary (1962).

<IP_ADDRESS> Cytology, Cytogenetics, and Evolution Aegilops speltoides is a diploid species containing seven pairs of homologous chromosomes (2n = 2x = 14), all of which have a median or sub-median centromere and lack distinctive morphological features, except for the SAT chromosomes (Riley et al. 1958; Chennaveeraiah 1960). In contrast, Dong et al. (2017), studying karyotypic polymorphism of Ae. speltoides, found intraspecific variation in the centromere position. In Ae. speltoides var. ligustica, there is one pair with a median centromere, while the rest of the pairs in both taxa have submedian centromeres (Chennaveeraiah 1960). The difference between the karyotypes of the two taxa seems to be very small. However, in var. ligustica, the short arms in the satellite pairs are more or less of the same length. In var. speltoides, there seems to be a difference in the lengths of the short arms of the satellite pairs (Chennaveeraiah 1960). The karyotype of both varieties of Ae. speltoides contains two pairs of SAT chromosomes with fairly large satellites (Pathak 1940; Riley et al. 1958; Chennaveeraiah 1960).

In Ae. speltoides, the genes coding for the 18S and 26S ribosomal RNA (rDNA genes) are located in the NOR regions on the short arms of chromosomes of groups 1 and 6 (Dvorak et al. 1984). There is only one 5S rDNA locus on the short arm of group 5 chromosomes that codes for 5S rRNA (Badaeva et al. 1996b). In contrast, Ae. longissima, Ae. sharonensis, Ae. bicornis and Ae. searsii, exhibit a different distribution pattern of NORs and 5S DNA loci. They contain major NOR loci on the short arms of chromosomes of groups 5 and 6 and two 5S rDNA loci on the short arms of chromosomes of groups 1 and 5 (Badaeva et al. 1996b). Thus, the distribution pattern of the NORs and 5S loci in the five Sitopsis species support the subdivision of this section into the two subsections.

The genome of Ae. speltoides is the smallest among the five Sitopsis species; 1C DNA is 5.81 pg (Furuta et al. 1977; Eilam et al. 2007) and 5.1 pg (The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. speltoides genome after genome sequencing and assembly to be somewhat smaller amount, i.e., 4.6 Gb (=4.5 pg), and Avni et al. (2022), also after sequencing and assembly, reported 5.13 Gb (=5.02 pg). No significant difference was noted in the genome size of the two taxa (Furuta et al. 1977; Eilam et al. 2007). Teoh and Hutchinson (1983), used an improved C-banding technique, found that all five Sitopsis species have telomeric, interstitial, and centromeric bands, varying in size and staining intensity. However, Ae. speltoides has exceptionally distinctive and extensive centromeric bands. The extent of polymorphic variation was found to vary between chromosomes of Ae. speltoides and plants were found to be heterozygous for the banding pattern (Teoh and Hutchinson 1983). Yet, the intraspecific variation in C-banding patterns neither differentiated between var. ligustica and var. speltoides, nor corresponded with impaired meiotic chromosomal pairing, since hybrids between the two varieties exhibited complete pairing and full fertility (Teoh and Hutchinson 1983). Badaeva et al. (1996a, b) and Ruban and Badaeva (2018) also found that Ae. speltoides have a different C-banding pattern than those of the other species of section Sitopsis.

Friebe and Gill (1996) established the homoeology between chromosomes of Ae. speltoides and those of T. aestivum, on the basis of similarities in chromosome morphology and C-banding patterns. Maestra and Naranjo (1998) determined the homoeologous chromosome relationship by analyzing homoeologous pairing at meiotic first metaphase of F1 hybrids between Triticum aestivum and Ae. speltoides carrying the homoeologous–pairing suppressor gene of common wheat, Ph1, or deficiencies ph1b for Ph1, and ph2b for Ph2. The chromosomes and their arms were identified, in both species, by C-banding. Data from relative pairing affinities were used to determine homoeologous relationships between Ae. speltoides chromosomes and bread wheat. All arms of the seven chromosomes of the speltoides genome showed normal homoeologous pairing with the wheat chromosomes, implying that no apparent chromosome rearrangements occurred in the evolution of Ae. speltoides relative to chromosomal structure in wheat. There was agreement between Friebe and Gill (1996) and Maestra and Naranjo (1998) in the assignment of five speltoides chromosomes to homoeologous groups 1, 4, 5, 6 and 7, but discrepancy with regards to chromosomes 2S and 3S.

Using in situ hybridization (ISH), Badaeva et al. (1996a) found that all Sitopsis species were similar to each other and to Amblyopyrum muticum in the distribution of the hybridization sites of two highly repetitive DNA sequences, pSc119 and pAsl. Yet, chromosomes of Ae. speltoides contain rich repetitive DNA sequences highly homologous to the pSc119 probe and, on the other hand, lack such homology to probe pTa53 (Dong et al. 2017). The distribution of pSc119 on chromosomes of Ae. speltoides show differences between accessions, between plants of one accession and even between homologous chromosomes in one plant.

Ae. speltoides comprises many of TEs, particularly long terminal repeat (LTR) retrotransposons, of which Ty1-copia superfamilies (Angela, Barbara, and Wis-A) and Ty3-gypsy superfamilies (Fatima and Erika) make up the main fraction (Middleton et al. 2013; Yaakov et al. 2013). Non-LTR retrotransposons (Ramona and Paula) and DNA transposons (Baldwin, Rong, and Charon) also exist in Ae. speltoides. All these TEs families vary greatly in copy number among different accessions of Ae. speltoides (Yaakov et al. 2013). Likewise, Hosid et al. (2012) analyzed intraspecific variation of four LTR retrotransposons (WIS2, Wilma, Daniela, and Fatima) in 13 different populations of Ae. speltoides from all over the distribution area of this species and found significant diversity in retrotransposon distribution. The various genotypes significantly differ with respect to the patterns of the four explored LTR retrotransposons, indicating a constant ongoing process of LTR retrotransposon fraction restructuring among and within populations of Ae. speltoides. Maximum changes were recorded in genotypes from small, stressed populations. The data of Hosid et al. (2012) revealed dynamic changes in LTR retrotransposon fractions in the Ae. speltoides genome, that are continually reshaping the genome of Ae. speltoides, particularly in stressful environments (Raskina et al. 2004a, b, 2008).

In Ae. speltoides euchromatin, widely interspersed TEs are clustered non-randomly, and may affect chromosomal structure. For instance, Raskina et al. (2004a) reported on the involvement of the transposable element system En/Spm [Enhancer (En) and Suppressor-Mutator (Spm)] in ongoing chromosomal repatterning in a small, isolated, peripheral population of Ae. speltoides. Cytogenetic analysis of the dynamics of En/Spm transposons in meiosis indicated that this transposon is active during male gametogenesis, changing the position of the rDNA sites (Raskina et al. 2004a). Such findings may indicate the importance of TEs in genome architecture (Belyayev et al. 2001, 2005; Altinkut et al. 2006), as well as their most important role in intraspecific divergence (Belyayev et al. 2010). The variability of TE content in Ae. speltoides, as well as in other Aegilops and Triticum species, might has a great impact on the dynamic, ongoing evolution of their genomes (Charles et al. 2008; Yaakov and Kashkush 2012).

Repetitive DNA sequences form chromosome-specific heterochromatin patterns in Ae. speltoides (Salina et al. 2006; Raskina et al. 2008). The species-specific tandem-repeat Spelt1 forms distinct clusters almost exclusively in subtelomeric chromosome positions, while Spelt52 clustering was most prominent at interstitial sites (Salina et al. 2006; Raskina et al. 2011). These two repetitive DNA sequences exhibit high intraspecific variation in both number and size (Salina et al. 2006; Raskina et al. 2011). Using fluorescence in situ hybridization, Salina et al. (2006) observed considerable polymorphisms in the hybridization patterns of Spelt1 between and within the studied lines of Ae. speltoides. There was a distinct ecogeographical gradient in the abundance of Spelt1 and Spelt52 blocks in Ae. speltoides; in marginal populations, the number of Spelt 1 chromosomal blocks was, at times, 12–14 times lower than in the center of the species distribution (Raskina et al. 2011). Likewise, Ae. speltoides had distinct Spelt52 hybridization patterns in the studied lines, but several distal Spelt52 sites, namely, on chromosome arms 3SL, 6SS, and 7SS, were common to all.

Aegilops speltoides contains a genetic mechanism that, in hybrids with allopolyploid wheats, suppresses the activity of the homoeologous-pairing suppressor (Ph1) gene of allopolyploid wheats, Triticum aestivum and T. turgidum. Riley and Law (1965) and Dover and Riley (1977) pointed out that increased homoeologous chromosome pairing at meiosis of common wheat x Ae. speltoides occurs only if Ph1 of common wheat is present and therefore proposed that the Ae. speltoides genome can promote homoeologous chromosome pairing by overpowering the activity of Ph1. Chen and Dvorak (1984) and Dvorak et al. (2006) found that the speltoides genes were ineffective in diploid hybrids lacking a gene like Ph1. Consequently, in accord with the above conclusion of Riley and Law (1965) and Dover and Riley (1977), they concluded that Ae. speltoides genes act on the Ph1 locus by suppressing its expression in common wheat x Ae. speltoides hybrids. Dvorak et al. (2006) mapped these, Ae. speltoides suppressor genes and demonstrated their significant effects as Mendelian loci on the long arms of chromosomes 3S and 7S. The chromosome 3S locus was designated Su1-Ph1 and the chromosome-7S locus was designated Su2-Ph1 (Dvorak et al. 2006). A QTL with a minor effect was mapped on the short arm of chromosome 5S and was designated QPh.ucd-5S (Dvorak et al. 2006). The expression of Su1-Ph1 and Su2-Ph1 increased homoeologous chromosome pairing in common wheat Ae. speltoides hybrids by 8.4 and 5.8 chiasmata/cell, respectively. Su1-Ph1 was fully epistatic to Su2-Ph1, and, when acting together, the two genes increased homoeologous chromosome pairing in common wheat Ae. speltoides hybrids to the same level when Su1-Ph1 acting alone. QPh. ucd-5S expression increased homoeologous chromosome pairing by 1.6 chiasmata/cell in common wheat Ae. speltoides hybrids and was additive to the effect of Su2-Ph1. It is hypothesized that the products of Su1-Ph1 and Su2-Ph1 affect pairing between homoeologous chromosomes by regulating the expression of Ph1, while the product of QPh. ucd-5S primarily regulates recombination between homologous chromosomes.

The Ae. speltoides genes that suppress Ph1 activity were transferred from a high-pairing accession of Ae. speltoides to common wheat (Chen et al. 1994). It appeared that two genes, presumably Su1-Ph1 and Su2-Ph1, were transferred. These suppressor genes were transferred to several lines of common wheat and used to induce homoeologous pairing in hybrids between these lines and wild relatives of wheat and, consequently, enhanced the gene transfer from wild relatives to common wheat (Li et al. 2011).

Yet, it is puzzling to assume that the homoeologouspairing promoters of Ae. speltoides (and also of Amblyopyrum muticum) evolved at the diploid level as a preadaptation, to repress the expression of the pairing suppressors at the polyploid level. These promoters may also promote homologous and homoeologous pairing per se or counteract the activity of suppressors that exist at the diploid level. Meiotic chromosome pairing data, collected by Sears (1941b), from hybrids of various diploid Triticineae species were interpreted by Waines (1976) to indicate that intraspecific genotypic differences control the amount of homoeologous chromosome pairing in diploid inter-specific hybrids. He suggested that homoeologous-pairing suppressors analogous to the Ph1 allele in bread wheat, are already present in genotypes of diploid species and do not necessarily have to arise by mutation or translocation after allopolyploid formation. He suggested that these homoeologous-pairing suppressors reinforce the isolating mechanism among the diploid species. The existence of Ph1-like genes in diploid species of the wheat group was also suggested by Okamoto and Inomata (1974) and by Maan (1977a). Feldman (1978) presented evidence for the existence of homoeologous-pairing suppressors in a low-pairing line of Ae. longissima.

There is also evidence that the promoters of the diploid species promote homoeologous pairing per se, not necessarily via the repression of Ph1 expression. Feldman and Mello-Sampayo (1967) concluded that, Ae. speltoides carries a promoter(s) that directly promotes homoeologous pairing in bread wheat x Ae. speltoides hybrids lacking the Ph1 gene. Similarly, all the promoters that exist in bread wheat promote pairing rather than suppress the expression of Ph1 (Feldman 1966). Indication that Ae. speltoides (and also A. muticum) contain genes that promote homoeologous pairing in the absence of Ph1 may be deduced from the following pairing data. The F1 hybrids between Ae. speltoides and T. monococcum and between Ae. speltoides and Ae. comosa analyzed by Sears (1941b), showed a similar amount of chromosome pairing to that observed by Ohta (1990, 1991) in the F1 hybrids between T. monococcum and A. muticum, between Ae. comosa and A. muticum, and between Ae. speltoides and A. muticum.

Massive chromosome restructuring was observed during the substitution of chromosome 6B of cultivar Chinese Spring of bread wheat by chromosome 6S from Ae. speltoides (Kota and Dvorak 1988). The chromosome rearrangements including deletions, translocations, ring chromosomes, dicentric chromosomes and a paracentric inversion in both euchromatic and heterochromatic regions of both wheat and Ae. speltoides chromosomes. The frequency of chromosome rearrangements was high among the B-subgenome chromosomes, moderate among the A-subgenome chromosomes, and low among the D-subgenome chromosomes (Kota and Dvorak 1988). In the B subgenome, the rearrangements were nonrandom, and occurred most frequently in chromosomes 1B and 5B. These observations indicate that wheat genomes can be subject to uneven rates of structural chromosome differentiation, even when within the same nucleus. Other examples of massive chromosome aberrations in wheat and Aegilops species are rare but were observed in root-tip cell of the Brazilian semi-dwarf cultivar IAS-54 of bread wheat (Dos Santos Guerra et al. 1977), in mitotic and meiotic cells of an individual plant of Elymus farctus (=Agropyron junceum) (Heneen 1963), and in first meiotic metaphase of an Ae. longissima line (TL02) (Feldman and Strauss 1983). Chromosome instability was also associated with gametocidal effects caused by several alien chromosomes of homoeologous group 4 in the monosomic state (Maan 1975; Finch et al. 1984; Tsujimoto and Tsunewaki 1984; Endo 1985).

Aegilops speltoides and Amblyopyrum muticum, the two-primitive species of the wheat group, are predominantly cross-pollinated, and contain genes promoting homoeologous pairing. On the other hand, these two species contain B chromosomes that suppress such pairing (Simchen et al. 1971; Mendelson and Zohary 1972; Zarchi et al. 1974; Mochizuki 1957, 1960, 1964; Ohta 1995a). B chromosomes are only known to occur spontaneously in cross-pollinating taxa (Müntzing et al. 1969) and are fully absent in selfpollinators (Jones et al. 2008). In both species, Ae. speltoides and A. muticum, the B chromosomes are absent in the roots but stably present in the aerial tissue (Mendelson and Zohary 1972; Ohta 1995a); a maximum of eight Bs per cell has been reported in Ae. speltoides (Raskina et al. 2004b). A comparable situation of tissue type-specific B chromosome distribution is also known for Agropyron cristatum (Baenziger 1962). The B chromosomes of Ae. speltoides and A. muticum are not homologous since they do not pair in F1 hybrids between these two species (Vardi and Dover 1972). The presence of one to three Bs has a positive effect on the plant, whereas a higher number of Bs reduces fertility and vigor (Mendelson and Zohary 1972; Belyayev et al. 2010).

The B chromosomes of Ae. speltoides are also characterized by a few A-chromosome-localized repeats, like Spelt1, pSc119.2 tandem repeats, 5S rDNA and Ty3-gypsy retroelements (Friebe et al. 1995c; Raskina et al. 2011; Hosid et al. 2012; Belyayev and Raskina 2013). It is assumed that these B chromosomes originated from the standard set of A chromosomes as a consequence of interspecific hybridization or, more likely, from trisomic (2n + 1) plants. Several lines of Ae. speltoides form a low frequency of unreduced gametes as a result of meiotic disorders or spontaneous non-disjunction of the entire chromosome complement, leading to the formation of triploid plants upon the fusion of the 2n gamete with 1n gamete (Belyayev and Raskina 2013; Feldman M, unpublished). Self-pollination of such triploids yields aneuploids and trisomic plants. Several trisomic Ae. speltoides plants were occasionally found in plants that originated from different populations (Feldman M, unpublished). Potential donors of the B chromosome of Ae. speltoides are the A chromosomes 1S, 4S and 5S of the Ae. speltoides genome (Friebe et al. 1995c; Belyayev and Raskina 2013). All Bs in Ae. speltoides have a single intercalary Spelt1 tandem repeat cluster and a 5S rDNA cluster in both arms (Raskina et al. 2011). In addition, a large intercalary cluster of Ty3-gypsy elements was found in close proximity to the 5S rDNA and Spelt1 blocks (Belyayev and Raskina 2013). Since chromosome 4S is the only Ae. speltoides chromosome that carries the intercalary Spelt1 cluster, and chromosome 5S, which is an exclusive source of 5S rDNA in the Ae. speltoides genome, it may be involved in the heterologous synapses and recombination resulting in the formation of Bs in this species (Raskina et al. 2011). Support of the idea that B derived from A chromosome(s) also comes from the observation that Bs pair at meiosis, although very rarely, with A chromosome(s) (Belyayev and Raskina 2013). The similarity in the B chromosome structures throughout the species range indicated that they were generated from a similar heterologous recombination of certain A chromosomes.

Ae. speltoides B chromosomes suppress homoeologous meiotic pairing of A chromosomes in intergeneric hybrids with bread wheat lacking the ph1 gene (Vardi and Dover 1972; Dover and Riley 1972). The effect of B chromosomes is like that of wheat Ph1 on homoeologous pairing in wheat hybrids with alien species. If Ae. speltoides B chromosomes derived from one of the A chromosomes, then one of the first essential prerequisites for the establishment of B chromosome as an independent entity should be to prevent pairing and recombination between the newly formed B and the ancestral A chromosome(s). Thus, the development of a genetic system suppressing homoeologous pairing was necessary for the independent existence and evolution of B chromosomes.

B chromosomes suppress homoeologous pairing in interspecific hybrids and thus, counteract the activity of the pairing promoters of Ae. speltoides. In a predominantly cross-pollinated species, the presence of genes that promote pairing is important, either to assure complete pairing in intraspecific hybrids or to increase pairing and recombination in interspecific ones. Yet, the presence of B chromosomes in some individuals of this species helps to conserve the integrity of their genomes.

# <IP_ADDRESS> Crosses with Other Species of the Wheat

Group Riley (1966a), Vardi and Dover (1972), and Ohta and Tanaka (1983) determined mean chromosome pairing at first meiotic metaphase in F1 hybrids of Ae. speltoides x Amblyopyrum muticum with and without B chromosomes (Tables 8.1 and 8.2). The hybrid speltoides x muticum without B chromosomes had mean pairing much higher than hybrid with four B chromosomes. Seed fertility of the F1 hybrids was very low (about 2.0%). The hybrids with B chromosomes, that suppress homoeologous pairing, indicating that the high pairing in hybrids without B chromosomes results mainly from homoeologous pairing promoted by the homoeologous-pairing promoters that exist in these diploids. Hence, the above pairing data show that the genomes of these two species have substantially diverged from one another.

In general, chromosome pairing at meiotic first metaphase in F1 hybrids between species of the Sitopsis section has been found to be high (Table 9.4) and consequently, Kihara (1949, 1954) grouped the Sitopsis species all together with the same primary genome symbol, S. The species of subsection Emarginata, namely, Ae. sharonensis, Ae. longissima, Ae. bicornis and Ae. searsii, form fertile F1 hybrids with one another, whereas F1 hybrids of any of these species with high- or low-pairing types of Ae. speltoides, the only species of subsection Truncata, show slightly reduced pairing and are sterile (Sears 1941b; Kihara 1949; Tanaka 1955a; Roy 1959; Riley et al. 1961; Kimber 1961; Ankori and Zohary 1962; Feldman et al. 1979). Thus, it is clear that the Ae. sharonensis-longissima-bicornis-searsii are of close affinity, while Ae. speltoides is somewhat more distant.

Hybrids between high-pairing lines of Ae. speltoides and other diploid species of Aegilops, i.e., Ae. caudata, Ae. comosa, Ae. uniaristata, and Ae. umbellulata exhibit about 10.70–11.26 (0.76–0.80%) paired chromosomes, while hybrids between low-pairing type of Ae. speltoides and Ae. caudata, Ae uniaristata, and Ae. umbellulata show only

Chromosome pairing between the allotetraploid species Ae. kotschyi and high-pairing and low-pairing types of Ae. speltoides exhibit somewhat reduced pairing (Table 9.5). The F1 hybrid between Ae. kotschyi and high pairing line of Ae. speltoides show 14.65 paired chromosomes (69.8%), whereas hybrids between Ae. kotschyi and low-pairing type of Ae. speltoides show only 13.8 (65.7%) paired chromosomes (Rubenstein and Sallee 1973). Hybrid between Ae. peregrina with low pairing type of Ae. speltoides showed 8.02 (38.2%) paired chromosomes (Yu and Jahier 1992). Also, F1 hybrid between other allotetraploid species of section Aegilops, namely, Ae. biuncialis, Ae. columnaris, Ae. neglecta and Ae. triuncialis, with Ae. speltoides, presumably a high-pairing type, showed similar levels of reduced pairing (Kihara 1949). In all the above diploid and triploid hybrids, the speltoides high-pairing genotypes promoted higher pairing between the homoeologous chromosomes of the studied species than the low-pairing genotypes. Data of chromosome pairing between allohexaploids of

Table 9.4 Chromosome pairing at first meiotic metaphase of F1 hybrids between diploid Aegilops species

<sup>a</sup> High pairing type of Ae. speltoides <sup>b</sup> Intermediate pairing type of Ae. speltoides

Table 9.5 Chromosome pairing at first meiotic metaphase of F1 hybrids between allotetraploids and diploids of the genus Aegilops

Table 9.5 (continued)

<sup>a</sup> LP—Low paring type of Ae. speltoides <sup>b</sup> HP—High paring type of Ae. speltoides <sup>c</sup> IP—Intermediate paring type of Ae. speltoides

Aegilops section Vertebrata, Ae. crassa 6 x and Ae juvenalis, and Ae. speltoides are presented in Table 9.6. These hybrids had higher pairing because of the effect of the homoeologous-pairing promoters in Ae. speltoides.

Data of chromosome pairing between allohexaploids of Aegilops section Vertebrata, Ae. crassa 6x, Ae juvenalis, and Ae. vavilovii, and diploid species of Aegilops are presented in Table 9.6. The hybrid Ae. crassa 6x x Ae. tauschii had

Table 9.6 Chromosome pairing at first meiotic metaphase of F1 hybrids between allohexaploids and diploids of the genus Aegilops

somewhat higher pairing than most other hybrids indicating high homology between the second D subgenome of the hexaploid and the D genome of the diploid. The remaining hybrid exhibited much less pairing. Chromosome pairing in F1 hybrids between allopolyploid species of Aegilops indicates that also several chromosomes of the differential subgenomes are involved in pairing in addition to that of he shared subgenome (Table 9.7).

Mean chromosomal pairing at meiosis in F1 hybrids between high-pairing type of Ae. speltoides and domesticated T. monococcum, ssp. monococcum, was very high, almost complete (Table 9.8). Similar high pairing was observed in the F1 hybrid between Ae. speltoides (presumably a high-pairing line) and wild T. monococcum, ssp. aegilopoides, was reported by Shang et al. (1989). However, mean chromosomal pairing at meiosis of F1 hybrids between the low-pairing type of Ae. speltoides and T. monococcum ssp. monococcum was low (Table 9.8) These results indicate that the speltoides homoeologouspairing promoter genes bring about a high degree of homoeologous pairing in speltoides x T. monococcum hybrids, and, when these genes are absent, as in the low-pairing genotype, the low level of pairing reveals excessive divergence of the genomes of these two species.

Rod Ring Total III IV V Above V Ae. ventricosa x Ae. crassa 4x DND<sup>c</sup> Xc 14.08 3.97 0.76 4.73 1.16 0.22 0.01 – Kimber and Zhao (1983) Ae. geniculata x Ae. biuncialis Mo UUM<sup>b</sup> 10.25 4.20 2.30 6.50 1.25 0.25 – – Kimber et al. (1988) Ae. geniculata x Ae. columnaris Mo UUX<sup>n</sup> 9.20 4.45 2.70 7.15 1.10 0.30 – Kimber et al. (1988) Ae. geniculata x Ae. kotschyi Mo UUS<sup>v</sup> 10.65 3.20 3.25 6.45 1.35 0.10 – – Kimber et al. (1988) Ae. geniculata x Ae. kotschyi Mo UUS<sup>v</sup> 11.70 1.60 4.42 6.02 1.42 ––– Feldman (1963) Ae. kotschyi x Ae. peregrina US<sup>v</sup> US<sup>v</sup> 0.94 1.94 9.04 10.98 0.20 0.54 0.18 0.24 VI Feldman (1963) Ae. geniculata x Ae. peregrina Mo UUS<sup>v</sup> 11.44 2.02 5.00 7.02 0.84 ––– Feldman (1963) Ae. biuncialis x Ae. peregrina UMb US<sup>v</sup> 12.74 1.22 5.14 6.36 0.74 0.08 – – Feldman (1963)

Table 9.7 Chromosome pairing at first meiotic metaphase of F1 hybrids between allopolyploids of the genus Aegilops

Hybrid combination Genome Univalents Bivalents Multivalents References

Table 9.8 Chromosome pairing at first meiotic metaphase of F1 hybrids between diploid species of Triticum and Aegilops

<sup>a</sup> High pairing type of Ae. speltoides <sup>b</sup> Intermediate pairing type of Ae. speltoides <sup>c</sup> Intermediate pairing type of Ae. longissima

McFadden and Sears (1947) reported that mean chromosomal pairing in the triploid hybrid between wild tetraploid wheat T. turgidum ssp. dicoccoides x Ae. speltoides was high, about two-third of the chromosomes paired (Table 9.9). Similar results were obtained by Riley et al. (1958) in hybrids between several subspecies of tetraploid wheat T. turgidum and Ae. speltoides. Yet, F1 hybrids between T. turgidum ssp. durum x low-pairing type of Ae. speltoides showed much lower pairing, involving very few paired chromosomes (Shands and Kimber 1973).

The triploid hybrid between the second tetraploid species of wheat, T. timopheevii and Ae. speltoides showed somewhat higher pairing than that between T. turgidum and Ae. speltoides (Table 9.9). Hybrids between T. timopheevii with the high-pairing Ae. speltoides genotype showed pairing of more than ten chromosomes, while hybrids with the lowpairing Ae. speltoides genotype showed pairing of seven bivalents only (Shands and Kimber 1973). These data indicate greater homology between Ae. speltoides and one of the subgenomes of T. timopheevii than in T. turgidum x Ae. speltoides hybrids.

Mean chromosome pairing at first meiotic metaphase in hybrids between hexaploid wheat, T. aestivum ssp. aestivum (bread wheat) and the high-pairing line of Ae. speltoides is very high (Table 9.10). On the other hand, hybrids with the low-pairing Ae. speltoides genotype, showed much reduced chromosomal pairing (Kimber and Athwal (1972).

Maestra and Naranjo (1998) analyzed homoeologous chromosome pairing at first meiotic metaphase in F1 hybrids derived from crosses of bread wheat, either carrying the homoeologous-pairing suppressor, Ph1, or the deficiency ph1b, lacking this gene, with a high-pairing genotype of Aegilops speltoides. Data from relative pairing affinities were used to predict homoeologous relationships of Ae. speltoides chromosomes to wheat. Chromosomes of both species, and their arms, were identified by C-banding. The Ae. speltoides genotype carried genes that induced a high level of homoeologous pairing in the two types of hybrids analyzed. All arms of the seven chromosomes of the speltoides S genome showed normal homoeologous pairing that implies that no apparent chromosome rearrangements occurred in the evolution of the genome of Ae. speltoides relative to wheat evolution. A pattern of preferential pairing of two types, A-D and B-S, confirmed that the S genome is very closely related to the B subgenome of wheat. Although this pairing pattern was also reported in hybrids of wheat with Ae. longissima and Ae. sharonensis, a different behavior was found in group 5 chromosomes. In the hybrids

Table 9.9 Chromosome pairing at first meiotic metaphase of F1 hybrids between allotetraploid species of Triticum and diploid species of Aegilops

<sup>a</sup> Triticum turgidum ssp. georgicum is currently T. turgidum ssp. paleocolchicum (Menabde) <sup>Á</sup>. löve & D. Löve; <sup>b</sup> HP-High paring type of Ae. speltoides; <sup>c</sup> IP-Intermediate paring type of Ae. speltoides; <sup>d</sup> LP-Low paring type of Ae. speltoides

of bread wheat with Ae. speltoides, chromosome 5B-5S pairing was much more frequent than 5D-5S, while these chromosome associations reached similar frequencies in the hybrids of Ae. longissima and Ae. sharonensis. These results are in agreement with the hypothesis that the B subgenome of wheat is closely related to the B subgenome of Ae. speltoides.

### 9.4.3 Aegilops bicornis (Forssk.) Jaub. & Spach

<IP_ADDRESS> Morphological and Geographical Notes Aegilops bicornis (Forssk.) Jaub. & Spach [Syn.: Triticum bicorne Forssk.; Sitopsis bicornis (forssk.) Á. Löve] is predominantly an autogamous annual plant. Culms are slender, erect, 15–45-cm-high. Spikes are 4–5(-8)-cm-long (excluding awns), two-rowed, mostly awned, with 8–15 (- 19) spikelets, disarticulating at maturity into spikelets, each spikelet falls with the rachis internode immediately below it (Fig. 2.3; wedge-type dispersal unit). Several lower spikelets often remain attached at the tip of culm. Spikelets are 5.5– 8.5-mm-long (excluding awns), mostly 3-flowered (2 lower florets are fertile). The attached rachis internode is half the length of the spikelet. Rudimentary spikelets at the base of the spike are absent. Glumes are 4.5–5.5-mm-long, emarginated or with an angle between teeth, asymmetrical and keeled. Lemmas are 4–6-mm-long, boat-shaped, mostly ending in a slender, 4.5–6.0-cm-long awn (in lower

Table 9.10 Chromosome pairing at first meiotic metaphase of F1 hybrids between allohexaploid species of Triticum and diploid species of Aegilops

a LP-Low paring type of Ae. speltoides; <sup>b</sup> IP-Intermediate paring type of Ae. speltoides; <sup>c</sup> HP-High paring type of Ae. speltoides. spikelets, awns are short or absent); the awn is usually not flanked by lateral teeth. The caryopsis adheres to lemmas and palea (Fig. 9.1c).

The species includes two varieties: var. bicornis (var. typica Eig) and var. anathera [var. mutica (Aschers.) Eig; Triticum bicorne muticum (Aschers.) Eig]. In var. bicornis, all spikelets are awned, sometimes, the lowest is awnless, whereas, in var. anathera, all spikelets, except for the uppermost 1–3, are awnless. Var. anathera is rare and sporadic, grows together with var. bicornis.

Ae. bicornis exhibits limited morphological variation, involving mainly spike and spikelet size, plant size, and degree of awn development. It is close in spike morphology to Ae. sharonensis, Ae. speltoides var. ligustica, Triticum monococcum ssp. aegilopoides and T. urartu.

Ae. bicornis is a semi-desert element, distributing in southeastern Mediterranean and in Saharo-Arabian regions. It occurs in the coastal regions of Libya (Cyrenaica)), Egypt (lower Egypt and Sinai) and southern Israel (western Negev), as well as in inland desert area in Israeli Negev and in Southern Jordan (Edom). Few populations were found in northeastern Cyprus. It grows from sea level to 200 m above sea level (in southern Jordan, it is found at 200–900 m), in areas with an annual rainfall of 75–275 mm (the driest part of the Mediterranean), usually on stable sandy soils, in open dwarf shrubs or herbaceous steppe-like or desert-like formations, xeric coastal and desert plains, in plantations, edges of cultivation and roadsides. It is common in coastal regions, sometimes in dense populations in the coastal plain of southern Israel, Sinai, and lower Egypt, and sporadic in inner sandy deserts. It is a very early maturing type. Ae. bicornis may have originated in the southern part of the Fertile Crescent. Currently, its center of variation is lower Egypt and the Sinai Peninsula. In the Israeli Negev and south Jordan (and possibly also in Egypt), Ae. bicornis grows sympatrically with Ae. longissima, and Ae. kotschyi. In south Israel, it grows allopatrically with Ae. sharonensis and Ae. peregrina.

<IP_ADDRESS> Cytology, Cytogenetics, and Evolution Ae. bicornis is a diploid (2n = 2x = 14) whose nuclear genome is a modified S genome (designated S<sup>b</sup> by Kihara 1954, Kimber and Tsunewaki 1988; Dvorak 1998) and its organellar genome is unique (designated S<sup>b</sup> by Ogihara and Tsunewaki 1988). Its nuclear 1C DNA content is relatively high (6.84 ± 0.097 pg; Eilam et al. 2007; 7.1 pg; The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. bicornis genome after genome sequencing and assembly to be 5.73 Gb (=5.6 pg), a value that is smaller than those above that were obtained by flow cytometry measurement.

Ae. bicornis has a symmetric karyotype (one chromosome pair has an almost median centromere and the rest of the pairs have submedian centromeres), with one pair with large and another with small satellites on the short arms (Senyaninova-Korchagina 1932; Riley et al. 1958; Chennaveeraiah 1960). The two SAT-chromosomes belong to homoeologous groups 5 and 6 (Friebe and Gill 1996). According to Riley et al. (1958), the karyotype of Ae. bicornis is similar to that of Ae. sharonensis and Ae. longissima.

Ae. bicornis, like the other species of subsection Emarginata, namely, Ae. longissima, Ae. sharonensis, and Ae. searsii, has major NOR loci on chromosomes of groups 5 and 6 and a variable number of minor loci on chromosomes of homoeologous groups 1, 3, 5 and 6 (Badaeva et al. 1996b). The 5S rDNA loci were observed in these species on chromosomes of groups 1 and 5, distal to minor NOR loci in the short arm of chromosome 1S<sup>b</sup> and proximal to a major NOR locus in the short arm of chromosome 5S<sup>b</sup> . Ae. bicornis differs from the other species of subsection Emarginats, with the exception of Ae. searsii, by having a lower heterochromatin content. In addition, Ae. bicornis differs from the other three Emarginata species in the size of the two 5S rDNA sites, and by the presence of a minor polymorphic NOR locus in a distal part of the long arm of chromosome 5S<sup>b</sup> (Badaeva et al. 1996b).

The karyotype of Ae. bicornis differs from that of Ae. longissima, Ae. sharonensis, and Ae. searsii, in both its C-banding pattern and by the size of C-bands detected in the karyotype, all of which are small (Teho and Hutchinson 1983; Teho et al. 1983a; Badaeva et al. 1996a), and most resembled that of Amblyopyrum muticum (Friebe et al. 1996). Ae. longissima and Ae. sharonensis are highly heterochromatic species; C-bands are present in intercalary, telomeric, and proximal regions of the chromosomes, while Ae. searsii and Ae. bicornis have much less C-heterochromatin and have relatively smaller centromeric bands (Teoh and Hutchinson 1983; Badaeva et al. 1996a).

Strong hybridization with the noncoding, highly repetitive DNA sequence, pSc119, derived from Secale cereale, was observed at the telomeres and at some subtelomeric regions of several chromosomes (Badaeva et al. 1996a). A strong signal with the pScll9 probe was detected at both telomeres of chromosomes 1S<sup>b</sup> and one polymorphic site was found in the subterminal region of the long arm of 1S<sup>b</sup> . Chromosomes 2S<sup>b</sup> , 3S<sup>b</sup> , and 6S<sup>b</sup> (a SAT chromosome) had similar labeling patterns with pSc119 but differed in chromosome morphology. The hybridization pattern of chromosome 4S<sup>b</sup> included one telomeric site in the short arm and a double site in the long arm. An additional site was detected in the subterminal region of the long arm. Chromosome 5S<sup>b</sup> , the second SAT chromosome, had one pSc119 site at the telomere of the short arm. Two telomeric pScll9 sites and one subterminal site were detected in the short arm of chromosome 7S<sup>b</sup> . The distribution of pScll9 hybridization sites in the Ae. bicornis genome was like that of Ae. longissima, Ae. sharonensis, Ae. searsii and A. muticum.

Using FISH in Ae. bicornis, Salina et al. (2006) did not find any detectable FISH signal with the two tandem repeated sequences Spelt1 and Spelt52 probes. This confirms earlier studies that concluded that Spelt52 is absent in Ae. bicornis (and in Ae. searsii), while it occurs in Ae. sharonensis and Ae. longissima (Anamthawat-Jonsson and Heslop-Harrison 1993; Zhang et al. 2002; Salina et al. 2004a; Belyayev and Raskina 1998; Raskina et al. 2011). Ae. sharonensis and Ae. longissima have similar pSc119 labeling patterns as those seen in Ae. bicornis and Ae. searsii but differ from them in having more heterochromatic chromosomes (Badaeva et al. 1996a; Friebe and Gill 1996).

# <IP_ADDRESS> Crosses with Other Species of the Wheat

Group The chromosomes of Ae. bicornis pair relatively well with those of the other Sitopsis species, but only the F1 hybrid with Ae. sharonensis was fertile (Tanaka 1955a). Tanaka (1955a) reported that in the F1 hybrid of a biotype of Ae. sharonensis with one of Ae. bicornis, meiotic chromosome pairing was fairly regular, with six to seven bivalents. Pollen and seed fertility in this hybrid was high, even though the plants were dwarf-like and weak. Similarly, high chromosomal pairing was observed in meiosis of a hybrid between a biotype of Ae. longissima and Ae. bicornis (Table 9.4) indicated that these species differed by a reciprocal translocation and the hybrid was more or less self-sterile (Kimber 1961). However, a backcross to either parent set seed when the hybrid was used as the female parent (Waines and Johnson 1972). Kihara (1949) reported 5–7 bivalents (seven bivalents were the mode) in first meiotic metaphase of F1 hybrids between Ae. speltoides and Ae. bicornis, however, the hybrids were sterile.

Few studies have been performed on crosses with other Aegilops species. Meiocytes of the F1 hybrid between Ae. bicornis and Ae. tauschii exhibited from zero to six bivalents and a rare one trivalent (Kihara 1949). Those between Ae. bicornis and tetraploid Ae. crassa as well as with Ae. uniaristata, had similar levels of chromosomal pairing, i.e., one to six bivalents and few multivalents (Kihara 1949). The F1 hybrid between Ae. bicornis and Ae. umbellulata, Ae. biuncialis and Ae. columnaris had an even lower level of pairing (0–5 with 3 bivalents as mode and few trivalents; Kihara 1949), while that with Ae. peregrina had higher pairing (Table 9.5), indicating that the genome of Ae. bicornis is related to one of the subgenomes of Ae. peregrina, presumably subgenome S<sup>v</sup> .

Ohta (1990) produced F1 hybrids between Ae. bicornis and Amblyopyrum muticum with and without B chromosomes (Tables 8.1 and 8.2). In studies of chromosomal pairing at first meiotic metaphase of the hybrids that did not have B chromosomes, he observed very little pairing (0.003 mean arm pairing per cell) whereas in hybrid with B chromosomes the pairing was significantly higher (0.77 mean arm pairing per cell; 82% of the cells had seven bivalents). No cells had multivalents and the frequency of univalents was very low. This high pairing in this hybrid presumably resulted from the activity of the homoeologous pairing promoters that exist in A. muticum rather than from the high degree of homology between the chromosomes of the two species.

Crosses between Ae. bicornis and Triticum monococcum ssp. monococcum was studied by Sears (1941b) (Table 9.8). At meiosis of this F1 hybrid, 5.38 chromosomes paired on average, most of which in the form of rod bivalent. In other crosses between Ae. bicornis and T. monococcum ssp. aegilopoides, the offspring were inviable or poorly viable, whereas hybrid with another variety of ssp. aegilopoides was viable (Sears 1944a). This lack of viability is apparently mono-factorially determined (Sears 1944a). Poor pairing was observed in the triploid hybrids between wild tetraploid wheat, T. turgidum subsp. dicoccoides x Ae. bicornis, and T. timopheevii ssp. timopheevii and Ae. bicornis (Table 9.9), and between hexaploid wheat, T. aestivum subsp. aestivum and Ae. bicornis (Table 9.10). Evidently, very little homology exists between the chromosomes of diploid wheat and Ae. bicornis; the triploid and tetraploid hybrids had even less pairing, due to the presence of the Ph1 gene of the allopolyploid wheats.

### 9.4.4 Aegilops sharonensis Eig

<IP_ADDRESS> Morphological and Geographical Notes Aegilops sharonensis Eig, common name: Sharon goat grass, [Syn.: Aegilops bicornis (Forssk.) Jaub. & Spach var. major Eig; Triticum sharonnse (Eig) Feldman & Sears; T. longissimum (Schweinf. & Muschl.) Bowden ssp. sharonensis (Eig) Chennav.; Aegilops longissima Schweinf. & Muschl. ssp. sharonensis (Eig) Chennav.; Ae. longissima Schweinf. & Muschl. var. major (Eig) Hammer; Sitopsis sharonensis (Eig) Á. Löve] is a predominantly autogamous annual plant. Its culm is (40-) 50–70 (-100)-cm-high (excluding spikes). Spikes are more or less broad, linear, two-rowed, usually awned (except in var. mutica), and 7–13-cm-long (excluding awns). Rachis is zig-zagged, with each segment being bow-shaped, disarticulating to individual spikelets at maturity, each spikelet with the rachis internode immediately below it (wedge-type dispersal unit; Fig. 2.3). The lower-most spikelet or the few lowest spikelets remain attached at the tip of the culm. Spikelets are 8–13-mm-long (excluding awns), linear, elliptical, grow smaller toward the tip, and are more or less flattened. There are 3–5 florets per spikelet, with the upper 1–3 being sterile. The attached rachis internode is shorter than the spikelets and curved. Glumes are 6–7-mm-long, with two small points, one of which is sometimes elongated into a small awn. Lemmas are 8–11-mm-long, with a 40–60-mm-long awn, there are two short broad teeth at the base. Awns increase in length toward the tip of the ear. The 1–3 lowest spikelets are usually almost awnless. Anthers are 5–6-mm-long. The caryopsis adheres to the lemma and palea (Fig. 9.1d).

Since its discovery, the status of Ae. sharonensis as an independent species has been in dispute and has been changed on several occasions. Zhukovsky (1928) considered Ae. sharonensis as a subspecies of Ae. longissima, while Eig (1928a) described it as a variety of Ae. bicornis and named it Ae. bicornis (forssk.) Jaub. & Spach. var. major Eig. Later, Eig (1928b), based on a re-examination of the plant morphology, his advanced knowledge of the various habitats, and the absence of hybrid swarms or intermediate forms in the contact zones of Ae. sharonensis and Ae. bicornis, elevated the former to the species rank and named it Ae. sharonensis Eig. However, its validity as a biological species had since been doubted. Based on complete chromosome pairing of the F1 hybrids at meiosis and their high fertility, Kihara (1937, 1940a, b, 1949, 1954), and Lilienfeld 1951) treated it as a subspecies or variety of Ae. longissima and assigned the two taxa the same genome formula (S<sup>l</sup> ). This treatment was supported by studies of the fertility and chromosome pairing in pollen mother cells (PMCs) of hybrids between these two taxa (Tanaka 1955a; Roy 1959; Kimber 1961). Also, Bowden (1959), while integrating the genus Aegilops into Triticum, considered Ae. sharonensis a subspecies of T. longissimum. Chennaveeraiah (1960), based on karyomorphology and plant morphology, also considered Ae. sharonensis to be a subspecies of Ae. longissima. The merging of Aegilops into Triticum was adopted by several cytogeneticists (e.g., Morris and Sears 1967; Mac Key 1968; Kimber and Sears 1983; Kimber and Feldman 1987), but these scientists considered T. sharonense as a separate species, named T. sharonense (Eig) Kimber and Feldman. While reorganizing the tribe Triticeae in genera based on genome homology, Löve (1984), recognized section Sitopsis as a genus including five diploid species bearing the S genome. Under this taxonomic treatment, Aegilops sharonensis was named Sitopsis sharonensis (Eig) A. löve. However, taxonomists (e.g., Baum 1977, 1978a, b; Hammer 1980; Feinbrun-Dothan 1986a; van Slageren 1994) kept Aegilops as a separate genus. Ankory and Zohary (1962) reported that the two species, Ae. sharonensis and Ae. longissima, are well-differentiated in nature and were apparently separated by seasonal and ecological factors for natural hybridization and, therefore, treated them as different species. Using the same arguments, Kimber and Feldman (1987) also treated them as separate species. Using C-banding pattern of mitotic metaphase chromosomes, Teoh and Hutchinson (1983) found that the differences between Ae. sharonensis and Ae. longissima are large enough to justify their treatment as two separate species. Likewise, studies of electrophoretic mobility of isoenzymes (Brody and Mendlinger 1980; Nakai and Tsuji 1984) and analysis of water-soluble leaf proteins (Mendlinger and Zohary 1995) showed Ae. sharonensis to be a valid species. Waines and Johnson (1972), studying the electrophoretic pattern of ethanol-extracted seed proteins, found that Ae. sharonensis was intermediate between Ae. longissima and Ae. bicornis and concluded that, Ae. sharonensis is genetically different from these two species and should be considered a separate species. Finally, Yen and Kimber (1990b) hybridized Ae. sharonensis with autotetraploid lines of Ae. speltoides, Ae. longissima and Ae. bicornis and analyzed chromosomal pairing at meiotic first metaphase of these hybrids. They found that Ae. sharonensis is almost equally related to Ae. speltoides and Ae. longissima, while it is distant from Ae. bicornis, and concluded, therefore, that Ae. sharonensis should not be treated as a subspecies or variety of Ae. longissima.

Ae. sharonensis includes two varieties: var. sharonensis (var. typica Eig), in which lemmas are awned in all spikelets except in the lowest, and var. mutica (Post) Eig, where lemmas are awnless in all spikelets or awned in the upper-most spikelets only. The latter is rare and occurs in mixed populations in Acre Plain and Sharon Plain of Israel.

Morphological variation involves differences in spike color and size, spikelet number and size, degree of expression of two-rowed nature and awn development. Studies on genetic and phenotypic diversity indicated that Ae. sharonensis is a diverse species, in spite of its limited geographic distribution and not highly variable environments (Olivera and Steffenson 2009). Morphologically, Ae. sharonensis is like to Ae. bicornis by having two-rowed spikes, awns in all the spikelets, spike disarticulation into single spikelets, and the shape of the lemmas (Eig 1929a), but have larger plant and spikelet sizes, larger grains and a somewhat laxer rachis. Edaphically, Ae. sharonensis is closer to Ae. bicornis than to Ae. longissima, though sympatric with the latter in several localities and allopatric with the former.

Olivera et al. (2010) used microsatellites to study genetic diversity and population structure of Ae. sharonensis from different sites in Israel and identified the Sharon Plain as the region exhibiting the highest level of allele richness and average gene diversity. Because it is located in the center of the geographic distribution of the species and includes sites with the highest level of diversity, the authors suggested that the Sharon Plain might be the center of origin and center of variation of Ae. sharonensis. There are two possibilities to explain the origin of Ae. sharonensis. First, it derived from Ae. bicornis that penetrated the coastal plain of southern Israel and absorbed genes from Ae. longissima via introgressive hybridization. Indeed, Waines and Johnson (1972), based on studies of seed protein patterns, proposed that, Ae. sharonensis derived from a hybridization between Ae. bicornis and Ae. longissima. Alternatively, Raskina et al. (2004b), based on studies of intrapopulation variability of rDNA in marginal populations of Ae. speltoides and Ae. sharonensis, suggested that Ae. sharonensis derived from Ae. speltoides.

Phenologically, Ae. sharonensis and Ae. bicornis are the earliest heading species of the genus Aegilops. Both species flower from March to May, albeit Ae. sharonensis starts flowering two weeks later than Ae. bicornis (Eig 1929a). Ae. speltoides, Ae. longissima and Ae. searsii head later; these species flower from April to June and in mountainous areas, even up to July (Feinbrun-Dothan 1986a; van Slageren 1994).

Ae. sharonensis is an east Mediterranean element. It has a very limited distribution in the south-central distribution region of the genus. It is endemic to the coastal plain of Israel (the Acre Plain, Sharon Plain, and Philistean Plain) and south Lebanon (Eig 1928b, 1929a, 1936; Millet 2006; Olivera and Steffenson 2009). The name Ae. sharonensis refers to the Sharon Pain in Israel, where this taxon was first described (Eig 1928a). It grows at 0–100 m above sea level, locally common, often in dense stands, on well-drained sandy soils, consolidated sand dunes, and on marine dilluvial rocks (kurkar rocks), in open park-, shrub-, and herbaceousplant formations, abandoned fields, disturbed habitats and roadsides (Eig 1928b, 1929a; Post 1933; Ankori and Zohary 1962; Witcombe 1983; Kimber and Feldman 1987; van Slageren 1994). It grows sympatrically with Ae. longissima (in few sites in the Israeli Coastal Plain), where intermediate types between these two species in mixed populations, mainly in disturbed habitats in the Sharon Plain, were reported by Eig (1929a) and Ankori and Zohary (1962), and with Ae. peregrina And Ae. geniculata, and allopatrically with Ae. longissima (in most sites in the Sharon and Philistean Plains), Ae. bicornis, and Ae. speltoides. Aegilops sharonensis has a rich source of genes providing resistance to important wheat diseases and abiotic stresses (Olivera and Steffenson 2009 and references therein). Some forms grow in salt-marshes (e.g., Na'aman salt-marsh, north of Haifa) and may contain genes for salt tolerance. With the most limited distribution of any species in the genus Aegilops, Ae. sharonensis is rapidly losing its habitats, owing to the combined effects of modern agricultural intensification and expansion of urban and industrial areas (Millet 2006; Olivera and Steffenson 2009).

<IP_ADDRESS> Cytology, Cytogenetics, and Evolution Ae. sharonensis is diploid (2n = 2x = 14), having a modified S genome, designated S<sup>l</sup> by Kihara (1954) and Dvorak (1998), or Ssh by Theo and Hutchinson (1983) (from here on, the genome symbol of Ae. sharonensis will be Ssh). Its organellar genome is quite similar to that of Ae. longissima, and consequently, is designated S<sup>l</sup> (Ogihara and Tsunewaki 1988). According to Eilam et al. (2007), this species has the largest genome in comparison to all other diploid Aegilops species (1C DNA = 7.52 ± 0.100 pg; 7.1 pg; The Angiosperm C-value database at Kew Botanic gardens (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. sharonensis genome after genome sequencing and assembly to be 6.07 Gb (=5.93 pg), and Avni et al. (2022), also after sequencing and assembly, reported 6.71 Gb (=6.56 pg). Furuta et al. (1986) described the genome size of Ae. sharonensis as equal to that of Ae. longissima and larger than those of Ae. searsii and Ae. speltoides. The karyotype of Ae. sharonensis is symmetric; all chromosome pairs have submedian centromere (Chennaveeraiah 1960). Riley et al. (1958) and Chennaveeraiah (1960) found that the karyotype of Ae. sharonensis is similar to that of Ae. longissima and, in both species, there is one pair with large and another pair with distinctly small satellites. The smaller satellites are slightly less than half the size of the larger ones. The karyotypes of Ae. sharonensis and Ae. longissima are quite similar to each other except for minor differences in the arm ratios of the non-satellited pairs (Chennaveeraiah 1960).

Ae. sharonensis, like the other three species of subsection Emarginata, has major NOR loci on chromosomes of groups 5 and 6 and a variable number of minor loci on chromosomes of groups 1, 3, 5, and 6 (Badaeva et al. 1996b). The 5S rDNA loci were observed on chromosomes of groups 1 and 5, distal to minor NOR loci in the short arm of chromosome 1Ssh and proximal to a major NOR locus in the short arm of chromosome 5Ssh (Badaeva et al. 1996b).

Teoh and Hutchinson (1983) and Teoh et al. (1983) studied the C-banding patterns of chromosomes at mitotic metaphase in all 10 diploid Aegilops species and found that all diploid species exhibit characteristically different patterns that enable the chromosomes of any complement to be individually identified. Ae. sharonensis has a unique C-banding pattern with telomeric, interstitial and exceptionally distinctive centromeric bands, varying in size and staining intensity. Hence, from the specific C-banding pattern, Teoh and Hutchinson (1983) concluded that the genome of Ae. sharonensis is different from that of Ae. longissima, and, consequently, gave it the symbol Ssh. The C-banding pattern of Ae. sharonensis indicates that this species is much more closely related to Ae. speltoides and Ae. longissima than to Ae. bicornis and Ae. searsii.

A repetitive DNA sequence thought to be a noncoding, highly repeated, 260-bp DNA fragment derived from the B subgenome of bread wheat (Hutchinson and Lonsdale 1982), was used in FISH experiments on genomes of all the diploid species of Aegilops (Teoh et al. 1983). This repetitive sequence was found in variable amounts in all diploid species but was restricted to specific regions of the chromosomes. Ae. sharonensis, Ae. speltoides and Ae. longissima possess many copies of the sequence and its distribution is correlated with their respective C-banding patterns. The strongest labelling was observed in Ae. longissima and Ae. sharonensis, both showing localization near the centromeres and also interstitially along the chromosomes.

Salina et al. (2006), using two telomere-associated tandem repeat sequences of Ae. speltoides, Spelt1 and Spelt52, found that the in-situ hybridization patterns of these probes in Ae. speltoides, Ae. longissima and Ae. sharonensis are different. The FISH signal with Spelt1 was only observed in Ae. speltoides, while the FISH signal with Spelt52 was found, in addition to Ae. speltoides, in Ae. sharonensis and Ae. longissima as well. Similar results were obtained by Anamthawat-Jonsson and Heslop-Harrison (1993), Zhang et al. (2002), Salina et al. (2004b) and Raskina et al. (2011). Thus, Spelt1 was completely absent from the genomes of the four Emarginata species, Ae. sharonensis, Ae. longissima, Ae. searsii and Ae. bicornis.

Badaeva et al. (1996a), using C-banding and in situ hybridization with two highly repetitive DNA sequences, pSc119, and pAsl, found that Ae. sharonensis was, together with Ae. speltoides and Ae. longissima, the very heterochromatic species of the diploid Aegilops species. While they did not observed hybridization with the pAsl probe on Ae. sharonensis, Ae. speltoides or Ae. longissima chromosomes, Ae. sharonensis, as well as the other Emarginata species (Ae. longissima, Ae. searsii and Ae. bicornis), showed strong labeling with the pSc119 probe in the telomeric chromosomal regions. Yet, the karyotypes of these species exhibited different distributions of C-heterochromatin (Badaeva et al. 1996b). Ae. sharonensis, like Ae. longissima, was highly heterochromatic; C-bands were present in intercalary, telomeric, and proximal regions of the chromosomes. Ae. searsii and Ae. bicornis had much fewer C-bands in these regions.

Yaakov et al. (2013) utilized quantitative real-time PCR to assess the relative copy numbers of 16 TE element families in various Triticum and Aegilops species. They found that the Latidu family of TEs showed specific proliferation in Ae. sharonensis, with more Latidu than in Ae. speltoides and Ae. longissima, and much more than in Ae. searsii. The Rong family was also observed in relatively high quantities in Ae. sharonensis.

# <IP_ADDRESS> Crosses with Other Species of the Wheat

Group Crosses between Ae. sharonensis and three other Sitopsis species, i.e., Ae. longissima, Ae. bicornis and Ae. speltoides, produced F1 hybrids that exhibited complete or almost complete chromosomal pairing at first meiotic metaphase. Thus, F1 hybrids between Ae. sharonensis and Ae. longissima and their reciprocal hybrid, had 5 bivalents (mostly ring bivalents) and a quadrivalent (or trivalent and univalent), indicating complete homology between the genomes of these two species and the existence of a reciprocal translocation between them (Tanaka 1955a; Kimber 1961; Ankori and Zohary 1962). Only Roy (1959) observed 6.90 bivalent and no multivalents in these hybrids. However, when assessing the photographs presented by Roy, Ankori and Zohary (1962) noticed that the longissima line used was already highly introgressed with Ae. sharonensis, which explains why Roy did not encounter a translocation configuration in his hybrids. Hybrid between Ae. sharonensis and Ae. bicornis exhibited 6–7 bivalents (Tanaka 1955a) and the hybrid between Ae. sharonensis and Ae. speltoides had similar levels of pairing (6.85 bivalents per cell of which 5.35 were ring bivalents; Kihara 1949; Sears 1941b; Kimber 1961), whereas Tanaka (1955a) reported on somewhat less pairing in this hybrid (4–7 bivalents). The F1 hybrids between Ae. sharonensis and Ae. longissima was almost fully fertile (more than 80% seed set; Tanaka 1955a; Ankori and Zohary 1962) and the hybrid Ae. sharonensis x Ae. bicornis had an approximate 62% seed set (Tanaka 1955a), while the hybrid between Ae. sharonensis and Ae. speltoides was sterile (Sears 1941b).

Data of chromosome pairing in F1 hybrids between Ae. sharonensis and other diploid Aegilops species are presented in Table 9.4. These hybrids exhibited low pairing, indicating that the genome of Ae. sharonensis is more distant from the genomes of these species than from those of Sitopsis species. The F1 hybrid between the tetraploid species Ae. kotschyi (genome S<sup>v</sup> Sv UU) and Ae. sharonensis exhibited 4.28 bivalents (of which 2 were ring), 1.50 trivalents, 0.46 quadrivalents and 0.09 pentavalents (Rubenstein and Sallee 1973), indicating high homology between the genome of Ae. sharonensis and one of the subgenomes of Ae. kotschyi, most probably genome S<sup>v</sup> .

In analyzing F1 hybrids between Ae. sharonensis and Ambliopyrum muticum without B chromosomes, Ohta (1990) found chromosome pairing at first meiotic metaphase ranging from 6.22 to 6.50 bivalents (of which 3.10 to 4.48 were ring bivalents) (Table 8.1). Several hybrids had few multivalent configurations. These hybrids showed very good pairing, presumably due to the promotion of homoeologous pairing by the genome of A. muticum but were completely sterile (Ohta 1990).

Ae. sharonensis was crossed with diploid, tetraploid, and hexaploid Triticum species (Tables 9.8, 9.9 and 9.10). The F1 hybrid between Ae. sharonensis and T. monococcum ssp. monococcum had very little pairing and were completely sterile (Kushnir amd Halloran 1981), indicating little homology between the Ssh and A<sup>m</sup> genomes. The F1 hybrid between Ae. sharonensis and tetraploid wild wheat T. turgidum ssp. dicoccoides had 2.23 bivalents (of which 0.03 were ring bivalents) (Kushnir and Halloran 1981), whereas McFadden and Sears (1947) reported 5.18 bivalents and 1.05 trivalents in the reciprocal hybrid. Hybrids between domesticated tetraploid wheat T. turgidum subsp. turgidum and Ae. sharonensis exhibited 2.22 bivalents and 0.02 trivalents (Riley et al. 1958). A similar level of chromosomal pairing in this hybrid combination, i.e., 2.50 bivalents per cell, was reported by Roy (1959). These F1 triploid hybrids had very little chromosome pairing at meiosis, indicating that, in the presence of the homoeologous-pairing suppressor, Ph1, of allopolyploid wheats, Ae. sharonensis chromosomes rarely pair with the wheat chromosomes and, likewise, the wheat chromosomes rarely pair with each other. Similar low chromosomal pairing exists in F1 hybrids between hexaploid wheat T. aestivum ssp. aestivum and Ae. sharonensis (Table 9.10). Interestingly, the amount of chromosome pairing in hybrids with hexaploid wheat is lower than that measured in hybrids with tetraploid wheat.

The homoeologous relationship between Ae. sharonensis and chromosomes of T. aestivum was determined by Friebe and Gill (1996), by comparing the C-banding pattern of Ae. sharonensis chromosomes, described by Teoh and Hutchinson (1983), with those of Ae. longissima chromosomes, whose homoeologous relationships to T. aestivum chromosomes were previously established by Friebe et al. (1993). Then, Friebe and Gill (1996) assigned the seven chromosomes of Ae. sharonensis to each homoeologous group of T. aestivum. Later, Maestra and Naranjo (1997) confirmed the homoeologous relationship between Ae. sharonensis and T. aestivum chromosomes, by C-banding analysis of specific pairing at first meiotic metaphase in F1 hybrids between Ae. sharonensis and T. aestivum ssp. aestivum. They analyzed chromosomal pairing in hybrids between three different genotypes of Triticum aestivum, each carrying (Ph1), or lacking it (ph2b mutant), and Ae. sharonensis, in order to establish the homoeologous relationships of Ae. sharonensis chromosomes to those of hexaploid wheat. Since Ae. sharonensis chromosomes show a distinctive C-banding pattern (Teoh and Hutchinson 1983), and thus, could be distinguished from those of wheat, C-banding was used by Maestra and Naranjo (1997) to identify the chromosomes of both species and their arms. Normal homoeologous relationships for the seven chromosomes of the Ssh genome of Ae sharonensis, and their arms, were revealed in this study. The pattern of pairing between chromosomes of Ae. sharonensis and ssp. aestivum indicated that no apparent chromosome rearrangement occurred during the evolution of the Ae. sharonensis genome relative to the subgenomes of hexaploid wheat. Thus, the chromosome structure of the ancestral genome from which subgenomes A, B, and D of allohexaploid wheat derived, was also preserved in genome Ssh. All three types of hybrids with Ph1, demonstrating low pairing level, however, with ph2b, demonstrating intermediate pairing level was observed, showing preferential pairing between A-D and B-Ssh. A close relationship between the Ssh genome and the B subgenome of bread wheat was confirmed, but the results provided no evidence that the B subgenome was derived from Ae. sharonensis. Similar results were obtained by Fernández-Calvín and Orellana (1993).

On the basis of the results of homoeologous pairing between chromosomes of Ae. sharonensis and T. aestivum, Maestra and Naranjo (1997) were able to identify the homoeologous relationships between all of the chromosomes of the Ssh genome and bread wheat. Their homoeologous pairing findings confirmed the homoeology of chromosome 4Ssh to wheat chromosomes of group 4, reported by Miller et al. (1982), as well as the assignment of the Ssh genome chromosomes to the seven homoeologous groups of wheat, and the arm designation, suggested by Friebe and Gill (1996), based on chromosomal morphology and C-banding.

### 9.4.5 Aegilops longissima Schweinf. & Muschl.

<IP_ADDRESS> Morphological and Geographical Notes Aegilops longissima Schweinf. & Muschl. [Syn.: Aegilops longissima (Schweinf. & Muschl.) ssp. longissima Hammer; Triticum longissimum (Schweinf. & Muschl.) Bowden; Sitopsis longissima (Schweinf. & Muschl.) Á. Löve] is a predominantly autogamous annual plant. Its culm is 40– 110-cm-high (excluding spikes). Spike is narrow, linear, one-rowed, tapering slightly to the tip, and 10–20-cm-long (excluding awns). At maturity, they are fragile near, but usually not at, the base, with the greater part of the spike falling entire, while lower spikelets remain on the culm or fall later singly or in 2–3 pieces. Spikelets 8–15 are 12– 14-mm-long (excluding awns), become thinner and shorter toward the tip of the spike, and are appressed to the rachis segment. There are 3–5 florets in each spikelet, the upper 1– 3 being sterile. The rachis internodes are nearly as long as spikelets in the middle of the spike. Glumes are 6– 8-mm-long, tough, and usually with two teeth separated by a membranous edge. Glumes of the terminal spikelet sometimes have three teeth, with the center one sometimes elongated into a very short awn. Lemmas of the lateral spikelets are canoe-shaped and awnless. Lemmas of the terminal spikelet have a 7–12-cm-long, broad, convex awn, often with a small unequal tooth on each side. Anthers are 5– 6-mm-long. The caryopsis adheres to both lemma and palea (Fig. 9.1e).

Limited morphological variation is mainly seen in spike length, spikelet number, rachis form (zig-zagged or straight) and awn length. Like Ae. speltoides and Ae. searsii, Ae. longissima ripens later than Ae. sharonensis and Ae. bicornis. The center of variation is in steppic regions of Israel and Jordan.

Morphologically, Ae. longissima is taller than Ae. sharonensis and differs from it by its long, narrow, linear, and one-rowed spike, by the absence of awns on the lemmas of the lateral spikelets, and by the presence of long and broad awns on the terminal spikelet. At maturity, the spike breaks near the base and falls almost entire and, at the end of the summer, the spike may disarticulate into single spikelets with the rachis internode beside them (barrel-type disarticulation). Ankori and Zohary (1962) reported that the two species are well-differentiated in nature and apparently separated by seasonal and ecological factors for natural hybridization. Cytologically, the two taxa have minor differences in their karyotype, and even though the hybrid between them exhibits complete chromosomal pairing at meiosis, the two taxa differ by a reciprocal translocation. The occurrence of a reciprocal translocation between the distal regions of chromosome arms 4S<sup>l</sup> L and 7S<sup>l</sup> L in Ae. longissima was later confirmed by Friebe et al. (1993). This translocation is not present in other Sitopsis species (Kihara 1949; Tanaka 1955a; Riley et al. 1961; Kimber 1961; Feldman et al. 1979).

Ae. longissima has a relatively limited distribution in the south-central region of the distribution area of the genus. A steppical (Irano-Turanian) element extends into sub-Mediterranean and desert (Saharo-Arabian) regions. It grows in Egypt (lower Egypt and Sinai), Israel (coastal plain, northern, and central Negev and eastern Judea and Samarian Mountains up to the east to Ein Gev, the eastern shore of the Lake of Galilee), Jordan, Lebanon (southern coastal plain), and Syria. Alt: 0–900 m above sea level (Table 9.1). It grows on sandy loams derived from the sandstone of the coastal plain, rarely on somewhat heavier soil, in the coastal plain of Egypt, Israel and Lebanon, and on sand derived from Nubian sandstone, in grey calcareous steppe soil or loess in the inland steppe or desert regions, in open dwarf shrub or herbaceous steppe-like or desert-like formations, plains, abandoned fields, edges of cultivation and roadsides. It is common, and often abundant in the coastal plain and in several steppic habitats bordering on the Mediterranean region in Israel, Jordan, and Syria.

Like Ae. bicornis, it grows in the hot and dry parts of the south-east Mediterranean, but its distribution area is closer to the center of the distribution area of the genus. The center of variation is in the steppic regions of Israel and Jordan, where it presumably originated and from which it invaded the coastal plain. It is sympatric with Ae. sharonensis [hybrid swarms with Ae. sharonensis can be found in the coastal plain (Ankori and Zohary 1962)] and Ae. peregrina in the coastal plain and with Ae., bicornis, Ae. kotschyi and Ae. vavilovii in the steppical region. It is allopatric with Ae. speltoides in the coastal plain, with Ae. searsii in southern Judea Mts., with Ae. umbellulata, Ae. geniculata and Ae. biuncialis in the sub-Mediterranean area in Lebanon and Syria and with Ae. crassa in the steppical regions of Syria.

<IP_ADDRESS> Cytology, Cytogenetics, and Evolution Ae. longissima is diploid (2n = 2x = 14) with a modified S genome, designated S<sup>l</sup> by Kihara (1949, 1954, 1963, 1970). Its organellar genome was designated Sl2, as a subtype of the Sl plasmon (Wang et al. 1997; Table 9.3. This species has the second largest genome among diploid Aegilops species (1C DNA = 7.48 ± 0.082 pg) (Eilam et al. 2007), 6.0 pg (The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. longissima genome after genome sequencing and assembly to be somewhat smaller amount, i.e., 6.22 Gb (=6.07 pg), and Avni et al. (2022), also after sequencing and assembly, reported 6.70 Gb (=6.55 pg). Furuta et al. (1986) reported that the genome size of Ae. longissima is equal to that of Ae. sharonensis but larger than those of Ae. searsii and Ae. speltoides.

The karyotype of Ae. longissima is symmetric; the satellite chromosomes have median centromeres, while all the non-satellited chromosomes have sub-median centromeres (Senyaninova-Korchagina 1932; Chennaveeraiah 1960). Riley et al. (1958) reported that the karyotypes in Ae. longissima and Ae. sharonensis are similar, and that in both species, there is one pair with large and another with distinctly small satellites. These findings were confirmed by Chennaveeraiah (1960), who stated that the smaller satellites are slightly less than half the size of the larger ones, and that there are minor differences in in the karyotypes of Ae. longissima and Ae. sharonensis in the arm ratios of the non-satellited pairs.

Based on the presence of secondary constrictions (Chennaveeraiah 1960; Chen and Gill 1983; Teoh and Hutchinson 1983), in situ hybridization analysis using a radioactive rDNA probe (Miller et al. 1983; Teoh et al. 1983), and analysis of nucleolar activity by Ag-NOR banding (Cermeño et al. 1984a, b), two pairs of NORs were identified in the secondary constrictions of Ae. longissima. Yet, Friebe et al. (1993) and Badaeva et al. (1996b), using in situ hybridization with a 18S–26S rDNA probe, which enabled high sensitivity, detected, in addition to the two major-active NORs on the short arms of chromosomes 5S<sup>1</sup> and 6S<sup>1</sup> , an additional minor NOR on chromosomes 1S<sup>l</sup> , and a polymorphic minor NOR on chromosome 3S<sup>l</sup> . The minor NORs usually do not form secondary constriction, indicating that these NORs are not transcribed. The 5S rDNA loci were detected in Ae. longissima on chromosomes 1S<sup>l</sup> and 5S<sup>l</sup> , distal to minor NOR loci in the short arm of chromosome 1S<sup>l</sup> and proximal to a major NOR locus in the short arm of chromosomes 5S<sup>l</sup> (Badaeva et al. 1996b).

In situ hybridization (ISH) patterns with both pTa7l (rDNA) and pTa794 (5S rDNA) probes revealed similar distribution of the two probes in Ae. longissima and Ae. sharonensis (Badaeva et al. 1996a). However, while ISH patterns were not polymorphic in Ae. sharonensis, intraspecific polymorphism was found in Ae. longissima. On the other hand, Ae. longissima chromosomes were similar to those of Ae. searsii in hybridization patterns with both probes (Badaeva et al. 1996a). This confirms the high degree of similarity of their genomes, as first observed by the distribution of highly repetitive DNA sequences (Teoh et al. 1983; Badaeva et al. 1996b). Aegilops bicornis differed from Ae. longissima by its lower heterochromatin content, differences in size between the two 5S rDNA sites, and the presence of a minor polymorphic NOR locus in a distal part of the long arm of chromosome 5S<sup>b</sup> (Badaeva et al. 1996b).

The C-banding patterns of Ae. longissima chromosomes were determined at mitotic metaphase by Teoh and Hutchinson (1983), who showed that all seven pairs of Ae. longissima chromosomes can be identified by their characteristic C-banding patterns. Like all other Sitopsis species, Ae. longissima also had telomeric, interstitial and centromeric bands, that varied in size and staining intensity. The proximal and telomeric bands were smaller than the centromeric ones (Teoh and Hutchinson 1983).

The karyotypes of Ae. longissima, Ae. sharonensis, Ae. searsii and Ae. bicornis have different C-banding patterns (Badaeva et al. 1996a). Ae. 1ongissima and Ae. sharonensis are highly heterochromatic species; C-bands are present in intercalary, telomeric, and proximal regions of the chromosomes. Ae. searsii and Ae. bicornis have much less C-heterochromatin compared with the two other Emarginata species.

Teoh et al. (1983) used a noncoding repetitive sequence, derived from T. aestivum, as a probe in in situ hybridization experiments on the genome of diploid Aegilops species. Ae. longissima, like Ae. speltoides and Ae. sharonensis, possesses many more copies of the noncoding sequence than Ae. bicornis and Ae. searsii. The distribution of the copies in each species correlated with their respective C-banding patterns.

Friebe et al. (1993) analyzed C-banding polymorphism in 17 accessions of Ae. longissima from Israel and Jordan and established a generalized idiogram of this species. Polymorphism for C-band size and C-band location was observed between different accessions but did not prevent chromosome identification. The C-banding patterns of Ae. longissima chromosomes they reported were similar to the N- and C-banding patterns reported earlier for this species (Chen and Gill 1983; Jewell and Driscoll 1983; Teoh and Hutchinson 1983; Kota and Dvorak 1985; Hueros et al. 1991). However, in most earlier reports, only one accession was analyzed and, therefore, no data were available on C-band polymorphisms in this species. Moreover, not all Ae. longissima chromosomes assigned correctly according to their homoeologous groups (Friebe et al. 1993).

A complete series of Ae. longissima addition and substitution lines were produced in the T. aestivum background (Hart and Tullen 1983). Sporophytic and gametophytic compensation tests were used to determine the homoeologous relationships of Ae. longissima chromosomes (Friebe et al. 1993). All Ae. longissima chromosomes compensated rather well and fertility was restored even in substitution lines involving wheat chromosomes 2A, 4B and 6B that contain major fertility genes.

The homoeologous relationships between Ae. longissima and wheat were determined for several chromosomes by their ability to compensate for the absence of wheat chromosomes in substitution lines (Jewell and Driscoll 1983; Kota and Dvorak 1985) and by all the chromosomes by C-banding analysis of Ae. longissima chromosomes in disomic and ditelosomic addition lines of Ae. longissima to T. aestivum and in substitution lines of Ae. longissima chromosomes for their T. aestivum homoeologues (Friebe et al. 1993).

The homoeologous relationships between Ae. longissima and wheat chromosomes have been established earlier by isozyme, storage protein, and morphological markers, as well as by analyzing their compensating ability in substitution lines (Hart and Tuleen 1983; Jewell and Driscoll 1983; Netzle and Zeller 1984; Kota and Dvorak 1985; Levy et al 1985; Millet et al. 1988; Hueros et al. 1991). The results obtained by Friebe et al. (1993) largely agreed with those reported previously. However, there were some discrepancies caused by chromosome misidentification in earlier studies.

C-banding patterns and morphology of Ae. longissima chromosomes 1S<sup>l</sup> , 3S<sup>l</sup> and 5S<sup>l</sup> are very similar to those of chromosomes 1B, 3B and 5B of T. aestivum (Gill et al. 1991a). The C-banding pattern of the short arm of 4S<sup>l</sup> is almost identical to that of the short arm of wheat chromosome 4B. The remaining Ae. longissima chromosomes showed differences in C-banding patterns and arm ratio with their wheat homoeologues.

Salina et al. (2006) and Raskina et al. (2001), using fluorescence in situ hybridization (FISH), observed considerable polymorphisms in the hybridization patterns of two tandemly repeated sequences, Spelt1 and Spelt52, among Aegilops species. While there was no detectable Spelt1 FISH signal in any species of section Sitopsis, except for Ae. speltoides, hybridization patterns of Spelt52 were speciesspecific in Ae. speltoides, Ae. longissima and Ae. sharonensis (Salina et al. 2006). Two very small and dim Spelt52 blocks were detected in Ae. searsii, whereas Ae. bicornis did not contain any Spelt52 repeats in its genome (Raskina et al. 2011). On the other hand, Ae. longissima and Ae. sharonensis showed that probe pSc119ƒffƒ (from Secale cereale) labeling patterns similar to Ae. bicornis and Ae. searsii but differed from them by having more heterochromatic chromosomes (Badaeva et al. 1996a; Friebe and Gill 1996).

Badaeva et al. (1996a), using both in situ hybridization (ISH) with the highly repetitive DNA sequences pSc119, from Secale cereale, and pAsl, from Ae. tauschii, as probes, and C-banding, analyzed genome differentiation in all diploid Aegilops species. The level of hybridization and labeling patterns differed among genomes. All five Sitopsis species had different C-banding pattern but they were similar to each other and to Amblyopyrum muticum in the distribution of pSc119 hybridization sites. On the other hand, no hybridization was observed with the pAsl probe on Ae. speltoides, Ae. sharonensis and Ae. longissima chromosomes, whereas a few minor pAsl sites were observed in Ae. searsii and Amblyopyrum muticum (Badaeva et al. 1996a). The ISH Patterns showed only minor intraspecific variations. Ae. bicornis and A. muticum had a low amount of C-heterochromatin, whereas Ae. longissima and Ae. sharonensis were the most heterochromatic species, and Ae. searsii possessed intermediate amounts of C-heterochromatin (Badaeva et al. 1996a). Ae. longissima, like the other three species of subsection Emarginata, Ae. sharonensis, Ae. searsii, and Ae. bicornis, had strong labeling in the telomeric chromosomal regions with the pSc119 probe (Badaeva et al. 1996a).

The C-banding pattern of the A. muticum genome chromosomes was different from that of the chromosomes of Ae. speltoides, and most resembled those of Ae. bicornis (Friebe et al. 1996). Distribution of pScll9 hybridization sites in the A. muticum genome also differed from that of Ae. speltoides but was similar to those of Ae. longissima, Ae. sharonensis, Ae. searsii and Ae. bicornis. Yet, the A. muticum genome was easily morphologically differentiated from the genome of the Sitopsis species.

During its evolution, Ae. longissima suffered a translocation involving the long arm of chromosome 4S<sup>l</sup> and the long arm of chromosome 7S<sup>l</sup> (Hart and Tuleen 1983; Friebe et al. 1993; Naranjo 1995). Analysis of chromosomal pairing at first meiotic metaphase showed a quadrivalent or a trivalent plus univalent in F1 hybrids between Ae. longissima and all other members of Sitopsis, indicating that Ae. longissima differs from the other four species by a reciprocal translocation (Feldman et al. 1979 and references therein). Since F1 hybrids between these four species form only bivalents at meiosis, it is reasonable to conclude that all four Sitopsis species, except Ae. longissima, have the ancestral chromosome structure.

Ae. longissima is probably involved in the parentage of the allotetraploid species Ae. peregrina (genome S<sup>l</sup> Sl UU) and Ae. kotschyi (genome S<sup>l</sup> Sl UU) (Kihara 1954; Feldman 1963; Kimber and Feldman 1987; Kimber and Sears 1987; Yen and Kimber 1989; Zhang et al. 1992; Dvorak 1998). Ae. longissima is a good source for genes conferring resistance to mildew, leaf and stem rust, heat and drought tolerance, and determination of high grain protein.

Some forms of Ae. longissima have genetic mechanisms capable of partially suppressing the Ph1 gene of polyploid wheat in hybrids with T. aestivum and T. turgidum. Mello-Sampayo (1971b) reported that a line of Ae. longissima promoted homoeologous pairing in meiosis of the F1 hybrid with T. aestivum, thereby, partially counteracting the suppressive effect of wheat-Ph1. This line was designated an intermediate-pairing (IP) line to distinguish it from the more common low-pairing (LP) lines. Upadhya and Swaminathan (1967) reported on another IP line of Ae. longissima that promoted pairing in hybrids with common wheat. In studies of chromosome pairing in F1 hybrids between ditelosomic lines of T. aestivum cv. Chinese Spring and the IP line of Ae. longissima, discovered by Mello-Sampayo (1971b), an intermediate amount of pairing was observed (5.49 bivalents, 0.39 trivalents and 0.06 quadrivalents per pollen mother cells (PMCs), most of which was between chromosomes of the wheat B subgenome and those of Ae. longissima (Feldman 1978).

To assess the effect of LP and IP genotypes on homologous pairing, Avivi (1976) produced autotetraploids from both lines of Ae. longissima and followed chromosomal pairing at first metaphase of meiosis. While the two induced autotetraploids did not differ in chiasma frequency or in the number of paired chromosomal arms, they differed significantly in multivalent frequency; the IP autotetraploid exhibited the same multivalent frequency as that expected on the basis of random pairing between the four homologues, namely, 4.7 multivalents per/cell. In contrast, the LP autotetraploid exhibited a significantly lower than expected frequency of multivalents. Avivi (1976) assumed that the LP genotype in the autotetraploid does not affect meiotic pairing per se but modifies the pattern of homologous association by separating the four sets of homologous chromosomes in somatic and premeiotic cells into two groups of two. In contrast, the IP genotype does not affect the spatial arrangement of the chromosomes. Accordingly, the gene Ph1 of common wheat suppresses homoeologous pairing in the aestivum x longissima hybrid by separating the wheat genomes from that of the longissima genome, thereby preventing pairing of distantly located chromosomes, the IP genotype does not do it and, consequently, enables some pairing between common wheat and longissima chromosomes (Avivi 1976).

# <IP_ADDRESS> Crosses with Other Species of the Wheat

Group Several intraspecific crosses were performed between lines of Ae. longissima from different habitats and geographical regions (Feldman et al. 1979 and Feldman M, unpublished). All the hybrids showed seven bivalents, most of which were ring bivalents. Seed set was normal except in one combination between a line from the Israeli Negev (steppe area) and a line from the Israeli coastal plain (Mediterranean region), in which seed fertility was low (53%) (Feldman et al. 1979).

Crosses between Ae. longissima and all of the other four Sitopsis species, i.e., Ae. sharonensis, Ae. bicornis, Ae. searsii and Ae. speltoides, produced F1 hybrids that exhibited complete or almost complete chromosomal pairing at first meiotic metaphase (Table 9.4). Thus, F1 hybrids between Ae. longissima and Ae. sharonensis and the reciprocal hybrids exhibited almost complete chromosome pairing at first meiotic metaphase, with 5 bivalents, one quadrivalent or trivalent and univalent, indicating the existence of high homology and the presence of a reciprocal translocation between these two species (Tanaka 1955a; Kimber 1961; Ankori and Zohary 1962). The F1 hybrids are almost fully fertile, with an 82–86% seed set (Tanaka 1955a). Similarly, high pairing indicating great chromosomal homology and the presence of a reciprocal translocation was observed in all the hybrids of Ae. longissima and the other four Sitopsis species (Table 9.4). Nevertheless, the F1 hybrids involving Ae. longissima and the Emarginata species, namely, Ae. sharonensis, Ae. bicornis and Ae. searsii, were highly or only partially fertile. In contrast, the F1 hybrid Ae. longissima x Ae. speltoides was completely sterile in contrast to the high pairing (Riley et al. (1961), implying that the chromosomes of the two species differ by cryptic structural hybridity or by genetic barriers. Thus, Ae. longissima diverges from the other Sitopsis species by a translocation and is totally isolated from Ae. speltoides by partial internal chromosomal or genetic barriers which are superimposed upon predominant self-fertilization and ecological specialization of these two species. Since all intraspecific hybrids of Ae. longissima studied had seven bivalents, i.e., they were homozygous for the translocation, it is reasonable to assume that the translocation originated during the formation of this species.

Kihara (1949) crossed Ae. longissima with Ae. caudata, Ae. comosa, Ae. uniaristata and Ae. umbellulata analyzed chromosome pairing in the F1 hybrids and in the reciprocal combinations. In longissima x caudata Kihara observed 3–6 bivalents and 0–2 trivalents, in longissima x comosa up to 6 bivalents and 0–1 trivalents, in longissima x uniaristata 2–5 bivalents and in Ae. longissima x Ae. umbellulata 0–4 bivalents (with mode of 2) and 0–1 trivalent These hybrids were completely sterile. These findings indicate that these four-specie are not close to Ae. longissima and have diverged quite considerably from it.

Chromosome pairing in F1 hybrids between Ae. longissima and Amblyopyrum muticum (without B chromosomes) was relatively high (Tables 8.1 and 8.2). These hybrids showed very good pairing, presumably due to the promotion of homoeologous pairing by the genome of A. muticum, but were completely sterile (Ohta 1990). The presence of multivalents indicated that the two species differ by a reciprocal translocation. There was a slight difference in the amount of pairing between these two hybrids. Since the same accession of A. muticum was used in the two hybrids, Ohta (1990) suggested that the frequency and configuration of chromosome pairing observed in these two hybrid plants might be caused by small differences in the chromosomal structure or in the genotypes affecting homoeologous chromosome pairing between the two accessions of Ae. longissima. In hybrids Ae. longissima x A. muticum with B chromosome pairing was considerably reduced (Table 8.2).

Data from analyses of chromosomal pairing at first meiotic metaphase in F1 triploid hybrids between Ae. longissima and a number of allotetraploid species of section Aegilops are presented below. Kihara (1949) reported the presence of 6–8 bivalents (with mode of 7) and 0–1 trivalents in the F1 Ae. peregrina x Ae. longissima. Similar data were also reported by other researchers (Table 9.5) indicating that one of the two subgenomes of Ae. peregrina is homologous to the genome of Ae. longissima. Similar results were observed in the F1 triploid Ae. kotschyi x Ae. longissima (Table 9.5). Chromosomal pairing in the F1 Ae. biuncialis x Ae. longissima exhibited 0–5 bivalents and 0–2 trivalents, and the F1 Ae. geniculata x Ae. longissima hybrid had 2–6 bivalents (with mode of 4), and 0–1 trivalents (Kihara 1949). Evidently, there is reduced homology between the genomes of these species.

Several crosses were performed between Ae. longissima and diploid, tetraploid and hexaploid Triticum species (9.8, 9.9 and 9.10). Kihara (1949) studied chromosome pairing in the F1 Ae. longissima x T. monococcum ssp. monococcum and found 0–3 bivalents at meiosis. Feldman (1978) reported 1.49 bivalents, of which 0.02 were ring bivalents, and 0.07 trivalents in F1 longissima x ssp. monococcum hybrid. These data indicate that the two species have diverged quite considerably from one another.

Chromosome pairing was analyzed in several Ae. longissima x T. turgidum hybrids (Table 9.9). In all these hybrids pairing was very low, indicating that very little homology exists between the chromosomes of Ae. longissima and those of T. turgidum. Kihara (1949) also crossed Ae. longissima with the second tetraploid species of wheat, T. timopheevii, and also found very little pairing in this hybrid (0–7 bivalents and 0.01 trivalents), indicating that the chromosomes of this tetraploid species are not homologous to those of Ae. longissima.

Fig. 9.2 Aegilops species carrying the D genome; <sup>a</sup> A. spike of Ae. tauschii Coss. ssp. tauschii (formerly ssp. eusquarrosa Eig); b A spike of Ae. tauschii Coss. ssp. strangulata (Eig) Tzvelev; c A spike of Ae.

Table 9.1 in McFadden and Sears (1947) includes data on chromosomal pairing in the F1 hybrid between Triticum turgidum subsp. dicoccoides and Aegilops sharonensis, which showed the formation of 5.18 bivalents 0–3 multivalents. But, on page 1016 of their article Fig. 9.2i is showing Ae. longissima and not Ae. sharonensis. Therefore, the pairing data in Table 9.1 is between subsp. dicoccoides and Ae. longissima, and, judging from the level of pairing in this hybrid, as well as in hybrids between T. aestivum and intermediate-pairing (IP) type Ae. longissima (Mello-Sampayo 1971b; Feldman 1978), it is most likely that the longissima parent in the McFadden and Sears (1947) paper is an IP type of Ae. longissima, and not of Ae. sharonensis.

Several crosses were performed between low-pairing (LP) type and IP type Ae. longissima and hexaploid wheat, T. aestivum. Hybrids involving LP types of Ae. longissima yielded very low pairing, with 1.45–1.96 bivalents and 0.01– 0.08 trivalents (Riley et al. 1961; Riley and Chapman 1963; Riley 1966a; Ceoloni et al. 1986; Yu and Jahier 1992; Naranjo and Maestra 1995). Chromosomal pairing in these hybrids also showed little homology between the genomes of Ae. longissima and those of T. aestivum. On the other hand, hybrids involving IP types of Ae. longissima, that contains homoeologous pairing promoter(s), exhibited higher chromosomal pairing, namely, 4.79–5.49, 0.19–0.39

ventricosa Tausch; d A plant and a spike of 4x Ae. crassa Boiss.; e A spike of Ae. vavilovii (Zhuk.) Chennav.; f A spike of Ae. juvenalis (Thell.) Eig; g A spike of Ae. cylindrica Host

trivalents and 0.04–0.07 quadrivalents (Mello-Sampayo 1971; Feldman 1978; Ceoloni et al. 1986).

### 9.4.6 Aegilops searsii Feldman & Kislev Ex Hammer

<IP_ADDRESS> Morphological and Geographical Notes Aegilops searsii Feldman & Kislev ex Hammer [Syn.: Triticum searsii Feldman & Kislev; Sitopsis searsii (Feldman & Kislev ex Hammer) Á. Löve] is a predominantly autogamous annual plant. Its culm is 20–40 (-50)-cm-high (excluding spikes). The spike is narrow, linear, 5–11-cm-long (excluding awns), one-rowed, and tapers slightly towards the tip. At maturity, it disarticulates above the lowest spikelet and falls as a unit. There are 8–12 spikelets, which are linear, and generally adorned with 3 florets per spikelet in middle of the spike, of which 2 are fertile, 1 in the lower and one in the upper part of spike. The uppermost floret in the middle of spike is sterile. The rachis internode is more or less as long as the spikelets. Glumes feature two teeth, separated by a membranous edge. Glumes are ¾ the length of the spikelet. Lemmas of the terminal spikelet are awned. One floret has a short awn, triangular in cross section, while the other floret has a very long (equal to or longer than the spike), flat awn. Awns are flanked at the base by 1–2 unequal, short–articulate teeth; the middle tooth at glumes in terminal spikelet sometimes lengthened into an up to 1-cm-long awn. Anthers are short (2–3 mm). The caryopsis is more or less free at maturity (Fig. 9.1f).

A unique character of Ae. searsii is the unequal (one very long and the second short or sometimes absent) awns in the terminal spikelet, which undoubtedly results from the presence of only one fertile floret in the terminal spikelet. Generally, in the related species of the section, two developed awns form at the top of the ear, due to the occurrence of two fertile florets.

It is assumed (Feldman and Kislev 1977) that Ae. searsii is a young species which returned from the relatively dry habitats of the other Emarginata species to more mesophilic habitats. The very low stature of the plant, as well as its short ear, the almost equal length between the glumes and the florets, as well as the free caryopsis, are considered characteristics of advanced members of the genus (Eig 1929a).

Ae. searsii has limited morphological variation, involving mainly spike size, spikelet number and awn length. Israel and Jordan are the center of variation, where it probably originated. Morphologically, Ae. searsii is close to Ae. longissima, but differs from it in habitat and in the following characteristics: shorter plant, shorter spike, fewer spikelets, fewer fertile florets, larger glumes to spikelet ratio, awns of lemma in the terminal spikelet are very unequal, shorter anthers and free caryopsis.

Analysis of electrophoretically discernible water-soluble leaf proteins (Mendlinger and Zohary 1995) and random amplified polymorphic DNA (RAPD) (Goryunova et al. 2008), showed that Ae. searsii (and Ae. bicornis) displays the lowest diversity among the Sitopsis species. This lies in agreement with earlier studies (Dvorak and Zhang 1992b; Sasanuma et al. 1996; Giorgi et al. 2002; Goryunova et al. 2008), which have shown that Ae. searsii appears to be remote from the other three Emarginata species, equally distant from Ae. longissima, Ae. bicornis and Ae. sharonensis (Mendlinger and Zohary 1995). Likewise, meiotic pairing analyses in triploid hybrids of Ae. searsii with autotetraploid Ae. longissima, autotetraploid Ae. speltoides and autotetraploid Ae. bicornis, indicated the Ss genome of Ae. searsii is equally distant to the S genome of Ae. speltoides, the Sl genome of Ae. longissima and the S<sup>b</sup> genome of Ae. bicornis (Yen and Kimber 1990a).

Ae. searsii has a relatively limited distribution region in the south-central area of the distribution of the genus. It occupies east Mediterranean primary and secondary habitats. It is a sub-Mediterranean element, extending into the Irano-Turanian region. Alt: 200–1000 m above sea level. Ae. searsii is limited to Israel (Judea, Samaria, the Golan Heights, and the higher Negev), Jordan (Gilead, Ammon, Moav, and Edom), southeast Lebanon and southwest Syria. It grows on terra rossa, sometimes mixed with loess, or on basalt soil in the destroyed sub-Mediterranean habitat of Sarcopoterium spinosum, in open-park herbaceous formations, in the degraded deciduous steppe-maquis, in small shrub (Batha) formations, abandoned fields and edges of cultivation. Locally common, sometimes dense stands in Judea.

Ae. searsii grows sympatrically with Ae. peregrina, Ae. geniculata, Ae. biuncialis and T. turgidum subsp. dicoccoides (in Judea and Samaria), with Ae. triuncialis, T. monococcum subsp. aegilopoides and T. urartu (in Lebanon and Syria), and with Ae. kotschyi and Ae. vavilovii (in southern Jordan and Israeli Negev). It grows allopatrically with Ae. speltoides, Ae. longissima, Ae. caudata, Ae. umbellulata and Ae. neglecta. Ae. searsii is involved in the parentage of Ae. vavilovii (mixed populations of these two species occur in the higher central Negev in Israel and in southern Jordan), and it presumably introgressed with wild tetraploid wheat, T. turgidum subsp. dicoccoides, Ae. peregrina and Ae. kotschyi. This is the only diploid Aegilops species that grows in mixed populations with wild T. turgidum in the southern Fertile Crescent. It also has massive contact with T. urartu in southeastern Lebanon and southwestern Syria. Variation in the ability to suppress the Ph1 gene of polyploid wheat has not yet been demonstrated in this species.

<IP_ADDRESS> Cytology, Cytogenetics, and Evolution Ae. searsii is a diploid (2n = 2x = 14), with a modified S genome, designated S<sup>s</sup> by Feldman et al. (1979). Its organellar genome was designated Sv (Ogihara and Tsunewaki 1988; Wang et al. 1997) since it differs from those of the other Sitopsis species (Wang et al. 1997). The genome of Ae. searsii is the smallest genome in the Emarginata subsection (1C DNA = 6.65 ± 0.091 pg; Eilam et al. 2007), 5.9 pg (The Angiosperm C-value database at Kew Botanic gardens) (Tables 2.4 and 9.3). Li et al. (2022) assessed the size of Ae. searsii genome after genome sequencing and assembly to be 5.55 Gb (=5.42 pg). This genome size is somewhat smaller than that of Ae. bicornis and considerately smaller than those of Ae. sharonensis and Ae. longissima, but larger than that of Ae. speltoides. The karyotype of Ae. searsii is symmetric, with two metacentric chromosome pairs and five sub-metacentric pairs (Feldman and Kislev 1977; Feldman et al. 1979). One of the metacentric pairs and one of the submedian pairs are satellited chromosomes; the metacentric pair has large satellites and the submetacentric one has a pair of medium-size satellites. Ae. searsii differs from the other three Emarginata species in that it has a pair of the medium-size satellites, whereas the other species contain a pair of smaller satellites.

Karyotype analysis showed the presence of two secondary constrictions in Ae. searsii (Feldman and Kislev 1977; Feldman et al. 1979; Teoh and Hutchinson 1983). In situ hybridization analysis, using a radioactive rDNA probe (Teoh et al. 1983), revealed that two pairs of NORs are located in the secondary constriction of Ae. searsii. Yet, hybridization with a 18S–26S rDNA probe, which enabled high sensitivity, revealed that the two-major active NORs are located on the short arms of chromosomes 5S<sup>s</sup> and 6S<sup>s</sup> (Badaeva et al. 1996b). The numbers and chromosomal locations of the NORs corresponded to the number of satellited (SAT) chromosomes (Feldman and Kislev 1977; Feldman et al. 1979; Teoh and Hutchinson 1983), indicating that these regions are actively transcribed. An additional pair of minor NORs was detected on the short arm of chromosome 1S<sup>s</sup> (Badaeva et al. 1996b). The minor NORs do not usually form secondary constriction, indicating that these NORs are not transcribed. In situ hybridization with the pTa794 (5S rDNA) probe showed that the 5S rDNA loci are located in Ae. searsii on the short arms of chromosomes 1S<sup>s</sup> and 5S<sup>s</sup> (Badaeva et al. 1996b).

Ae. longissima chromosomes were similar to Ae. searsii chromosomes in hybridization patterns with both the pTa7l and pTa794 DNA probes, confirming the high degree of similarity between their genomes (Teoh et al. 1983; Badaeva et al. 1996b).

Teoh and Hutchinson (1983), using an improved C-banding technique, found that the C-banding patterns of mitotic metaphase chromosomes of Ae. searsii exhibited a characteristically different pattern from those of the Sitopsis species. Moreover, the C-banding pattern enables the chromosomes of this species to be identified individually. Ae. searsii, like other Sitopsis species, has telomeric, interstitial, and centromeric bands. However, Ae. searsii and Ae. bicornis have relatively smaller centromeric bands (Teoh and Hutchinson 1983).

Teoh and Hutchinson (1983) analyzed only one accession per species. C-banding polymorphism in a larger number of accessions of the diploid Aegilops species was studied by Friebe et al. (1992b, 1993, 1995b) and Friebe and Gill (1996). C-banding polymorphism was analyzed in 14 accessions of Ae. searsii from Israel, enabling establishment of a generalized idiogram of the species (Friebe et al. 1995b). All seven Ae. searsii chromosome pairs were individually identified on the basis of their C-banding patterns. No variation was observed within the accessions, but C-band polymorphism was detected between the different accessions. Ae. searsii was easily distinguished from Ae. longissima by their distinct C-banding patterns; Ae. searsii had fewer and smaller C-bands, and, in this respect, was similar to Ae. bicornis (Friebe et al. 1995b). Isozyme studies (Pietro et al. 1988) showed that there is no translocation 4/7 in Ae. searsii, also differing in this respect, from Ae. longissima.

C-banding analysis was also used to identify the seven disomic addition lines of Ae. searsii chromosomes to T. aestivum cv. Chinese Spring, 14 ditelosomic chromosome addition lines, 21 disomic substitution of whole chromosome and 31 ditelosomic chromosome substitution lines, produced by NA Tuleen. The identity of these lines was further confirmed by meiotic pairing analysis. Sporophytic and gametophytic compensation tests were used to determine the homoeologous relationships of the Ae. searsii chromosomes. The results show that the Ae. searsii chromosomes do not compensate well for their wheat homoeologues. The short arm of Ae. searsii, 3S<sup>s</sup> S, causes spike fragility; the spike breaks between the first and the sixth spikelets. Chromosome arm 2S<sup>s</sup> S causes tenacious glumes.

In situ hybridization with the highly repetitive DNA sequences pSc119 and pAsl, and C-banding analysis, enabled the genome differentiation in all the diploid species of Aegilops. The ISH patterns of chromosomes with pSc119 showed only minor intraspecific variations in the subsection Emarginata species. Chromosomes of all these diploid species hybridized with the pSc119 probe; however, the level of hybridization and labeling patterns differed among genomes (Badaeva et al. 1996a). All these species had strong pSc119 hybridization sites located mainly in the telomeres. Ae. longissima and Ae. sharonensis are highly heterochromatic species; C-bands were present in intercalary, telomeric and proximal regions of the chromosomes. Ae. searsii and Ae. bicornis have much less C-heterochromatin compared with the other two Emarginata species and hence, these species form a third subgroup (Badaeva et al 1996a).

On the other hand, the Sitopsis species differ in their ability to hybridize with the pAs1 probe (Badaeva et al. 1996a). No hybridization was observed with the pAs1 probe on Ae. speltoides, Ae. sharonensis and Ae. longissima chromosomes. A few minor pAs1 sites were observed in Ae. searsii and Ae. bicornis.

The evolution of two tandemly repeated sequences, Spelt1 and Spelt52, was studied in species of Aegilops sect. Sitopsis (Salina et al. 2006). Fluorescence in situ hybridization showed considerable polymorphisms in the hybridization patterns of Spelt1 and Spelt52 repeats between and within Aegilops species. Hybridization patterns of Spelt52 in Ae. speltoides, Ae. longissima and Ae. sharonensis were species-specific. There was no detectable Spelt1-associated FISH signal in Section Sitopsis, with the exception of Ae. speltoides. Spelt52 and its analogues pGc1R-1 and pAesKB52, were found in Ae. speltoides, Ae. longissima and Ae. sharonensis, but not in Ae. bicornis or Ae. searsii (Anamthawat-Jonsson and Heslop-Harrison 1993; Zhang et al. 2002; Salina et al. 2004a).

Yaakov et al. (2013) assessed the relative copy numbers of a number of TE families in five accessions of Ae. searsii and in several other Sitopsis species. The analysis of six Gypsy families revealed that Fatima is very abundant in Ae. searsii and in Ae. speltoides. On the other hand, several elements, i.e., Latidu, Sabrina, BAGY2 (all Gypsy retrotransposon) and Charon (Mutator DNA transposon), had specific proliferation in the Ae. searsii genome, in comparison to the other Sitopsis species.

# <IP_ADDRESS> Crosses with Other Species of the Wheat

Group Very few hybrids between Ae. searsii and other Aegilops and Triticum species were produced and analyzed. Feldman et al. (1979) noticed that the crossability between Ae. longissima with Ae. searsii is apparently very low. Over 380 pollinated florets (about 15 spikes) in both directions yielded about 10 seeds, and then, only when Ae. longissima was the female parent. Most of the seeds were shriveled and their embryos poorly developed; they either did not germinate, even after placing the embryo on culture medium, or the seedlings died very young. Only three seeds, representing two hybrid combinations, germinated and survived the seedling stage. When studying chromosome pairing at first meiotic metaphase of these F1 hybrids, Feldman et al. (1979) found that meiosis was irregular and chromosomal pairing was somewhat reduced (Table 9.4). One of the rod bivalents was heteromorphic, indicating that the total length and the arm ratio of the two pairing partners were not the same. Moreover, most meiocytes exhibited one asymmetrical multivalent, indicating that the two species differ in one asymmetric reciprocal translocation. The asymmetrical configuration implies that the translocated segments were unequal. Further indication of chromosomal differentiation between Ae. searsii and Ae. longissima was the occurrence of a chromatid bridge and an acentric fragment at first anaphase of the two hybrids, indicating heterozygosity for a paracentric inversion. A comparison of the karyotypes of the two species confirms the existence of several structural differences between the chromosomes of the two species (Feldman et al. 1979). Pollen and seed fertility of the F1 hybrids was low; pollen fertility was 30–37% and seed fertility 6.7–35%. The significant reduction in meiotic pairing in these hybrids, as manifested by reduced numbers of chiasmata, shows the incomplete homology between the Ae. searsii and Ae. longissima genomes. Clearly, in one or both species, several chromosomes have undergone independent re-patterning, as indicated by the asymmetrical translocation, heteromorphic bivalent and the paracentric inversion.

Ae. searsii and Ae. longissima are presumably isolated via several different mechanisms that include low crossability, partial hybrid sterility and differences in ecological requirements and geographical distribution. In addition to these, hybridization between the two species in nature is also greatly reduced because of the predominance of selfpollination. The multiplicity of isolating mechanisms is probably responsible for the rarity of hybrids between these species in the contact zone between them.

Yu and Jahier (1992) produced F1 hybrid between the allotetraploid Ae. peregrina and Ae. searsii. Analysis of chromosome pairing at first meiotic metaphase (Table 9.5) revealed that the Sv subgenome of Ae. peregrina is not completely homologous to that of Ae. searsii. From the level of pairing in the triploid hybrid, Yu and Jahier (1992) concluded that subgenome S of Ae. peregrina is very close to the genome of Ae. longissima, and relatively more distant from that of Ae. searsii.

Hybrids between hexaploid wheat, Triticum aestivum ssp. aestivum and Ae. searsii were produced and analyzed by Feldman (1978) and Yu and Jahier (1992) (Table 9.10). The low pairing observed in these hybrids indicated that very little homology exists between the Ss genome of Ae. searsii and all of the three subgenomes of T. aestivum. Hence, Ae. searsii is not the source of the B subgenome of allopolyploid wheat.

## 9.5 Section Vertebrata Zhuk. Emend. Kihara

### 9.5.1 General Description

Section Vertebrata Zhuk. emened Kihara [Syn.: Pachystachys Eig, Gastropyrum (Jaub. & Spach) Zhuk.; Polyploids Zhuk.; Aegilonearum Á. Löve] consists of annual, predominantly autogamous species. The plants are robust, with a more or less long spike, (3-) 6–10 (-15)-cm-long, mostly thick, 3–7 mm wide, cylindrical or moniliform, mostly awned and disarticulating at maturity into spikelets, each with the laterally adjacent rachis-internode (barrel-type dispersal unit). Spikelets are more or less ventricose or linear and glumes are mostly awnless. Lemmas of apical spikelets, sometimes of several upper spikelets, with one welldeveloped broad and mostly flat awn. The caryopsis adheres to lemmas and palea (Fig. 9.2).

Section Vertebrata contains five species, one diploid (Ae. tauschii Coss., genome DD) and two tetraploids (Ae. ventricosa Tausch (genome DDNN) and Ae. crassa Boiss. (genome D<sup>c</sup> Dc Xc Xc ). The latter also has a hexaploid cytotype (genome Dc Dc Xc Xc DD)), and two hexaploids (Ae. juvenalis (Thell.) Eig (genome Dc Dc Xc Xc UU) and Ae. vavilovii (Zhuk.) Chennav. (Genome D<sup>c</sup> Dc Xc Xc Ss Ss ). Kimber and Zhao (1983) and Zhao and Kimber (1984), based on analysis of meiotic chromosome pairing, decided that the allopolyploid species containing the D subgenome can be divided into three clusters: (1) T. aestivum, Ae. cylindrica and Ae. ventricosa, in which the D subgenome has undergone little modification from the genome of the diploid progenitor, Ae. tauschii; (2) tetraploid and hexaploid Ae. crassa, in which the D subgenome is somewhat modified; (3) Ae. juvenalis and Ae. vavilovii, in which the D subgenome is substantially modified. Dubcovsky and Dvorak (1995a), Dvorak (1998) and Dvorak et al. (2012) arrived at a similar conclusion but pointed out that the D subgenome of Ae. crassa differentiated considerably from that of the diploid parent and consequently, it was designated Dc (Dvorak 1998). Analyses of the plasmon of these species clearly showed that Aegilops tauschii is the maternal parent of the two allotetraploid species, Ae. crassa and Ae. ventricosa, and that the tetraploid cytotype of Ae. crassa is the maternal parent of three hexaploids, Ae. juvenalis, Ae. vavilovii and 6x Ae. crassa (Tsunewaki 1993, 2009; Ogihara and Tsunewaki 1988; Wang et al. 1997). These studies suggested that the D-genome containing allotetraploids originated at three different times from Ae. tauschii in the following order: Ae. crassa, Ae. cylindrica, and Ae. ventricosa. Ae. crassa is the oldest allotetraploid containing the D genome, while Ae. cylindrica and Ae. ventricosa originated later. (Tsunewaki 1993, 2009; Ogihara and Tsunewaki 1988; Wang et al. 1997). Thus, 4x Ae. crassa appears to be an ancient allotetraploid that originated from hybridization of primitive Ae. tauschii with an ancient species in the evolutionary lineage leading to the section Sitopsis (Dubcovsky and Dvorak 1995a; Badaeva et al. 2002).

The Vertebrata species are characterized by, more or less, high morphological resemblance. All the allopolyploid species contain the D subgenome or a modified D subgenome that derived from the genome of the diploid species, and the impact of this genome is clearly apparent in the morphology and dispersal unit type of all the allopolyploid species. In addition, the organellar genome of all the allopolyploid species derived from that of the diploid species.

The central and northeastern part of the Fertile Crescent (i.e., eastern Turkey, Syria, Iraq and Iran) is the region with the greatest diversity for the Vertebrata species (Table 9.1). Only one species, Ae. ventricosa, occurs exclusively west of the Fertile Crescent, while other three, Ae. tauschii, Ae. crassa and Ae. juvenalis, have spread eastward, and Ae. vavilovii grows in the southeastern part of the genus distribution area. With the exception of Ae. juvenalis and, to a lesser extent, Ae. vavilovii (both hexaploids), all Vertebrata species are widespread.

The distribution of Ae. ventricosa is the most difficult to explain in an evolutionary sense, as it distinctly does not overlap with the areas of any of its putative parents. Ae. tauschii is the donor of one subgenome to Ae. ventricosa (Kihara 1949) and the second genome is derived from Ae. uniaristata (Yen and Kimber 1992b). There seems only a theoretical overlap on the Istrian peninsula of Croatia of uniaristata and ventricosa, but Ae. tauschii is still far away.

Some Vertebrata species are also found at higher altitudes and may show better adaptation to cold than most species of Aegilops. Several species grow in areas with relatively little annual rainfall and may be drought-tolerant. On the other

hand, the D-genome of these species may be responsible for the overall poor performance of species with this genome in resisting rust infection (Hammer 1987).

### 9.5.2 Aegilops tauschii Coss.