[Successful bridging therapy with alectinib prior to allogeneic stem cell transplantation for refractory ALK-positive anaplastic large cell lymphoma].

Although alectinib is effective for relapsed or refractory ALK-positive anaplastic large cell lymphoma (ALCL) and has a favorable safety profile, its role as a bridging therapy for allogeneic hematopoietic stem cell transplantation (allo-HSCT) and the role of allo-HSCT itself in this setting are unknown. A 35-year-old man with ALK-positive ALCL experienced relapse after first-line therapy with CHOP. Brentuximab vedotin led to partial response and high-dose chemotherapy combined with autologous HSCT was performed. However, disease progressed 15 months after transplantation, and alectinib was initiated. Complete response (CR) was achieved after three months of treatment, and alectinib was continued for 5 months. After cessation of alectinib, allogeneic bone marrow transplantation from an HLA 1-locus mismatched unrelated donor was performed after conditioning with fludarabine, busulfan, and total body irradiation. GVHD prophylaxis consisted of tacrolimus and short-term methotrexate. The post-transplant course was unremarkable except for grade I acute GVHD. The lymphoma has not recurred for 2 years after allo-HSCT without resuming alectinib. The clinical course of our case suggests that alectinib bridging therapy and allo-HSCT are effective in relapsed/refractory ALK-positive ALCL.

Floral pigments and their perception by avian pollinators in three Chilean Puya species.

The Chilean Puya species, Puya coerulea var. violacea and P. chilensis bear blue and pale-yellow flowers, respectively, while P. alpestris considered to be their hybrid-derived species has unique turquoise flowers. In this study, the chemical basis underlying the different coloration of the three Puya species was explored. We first isolated and identified three anthocyanins: delphinidin 3,3',5'-tri-O-glucoside, delphinidin 3,3'-di-O-glucoside and delphinidin 3-O-glucoside; seven flavonols: quercetin 3-O-rutinoside-3'-O-glucoside, quercetin 3,3'-di-O-glucoside, quercetin 3-O-rutinoside, isorhamnetin 3-O-rutinoside, myricetin 3,3',5'-tri-O-glucoside, myricetin 3,3'-di-O-glucoside and laricitrin 3,5'-di-O-glucoside; and six flavones: luteolin 4'-O-glucoside, apigenin 4'-O-glucoside, tricetin 4'-O-glucoside, tricetin 3',5'-di-O-glucoside, tricetin 3'-O-glucoside and selagin 5'-O-glucoside, which is a previously undescribed flavone, from their petals. We also compared compositions of floral flavonoid and their aglycone among these species, which suggested that the turquoise species P. alpestris has an essentially intermediate composition between the blue and pale-yellow species. The vacuolar pH was relatively higher in the turquoise (pH 6.2) and pale-yellow (pH 6.2) flower species, while that of blue flower species was usual (pH 5.2). The flower color was reconstructed in vitro using isolated anthocyanin, flavonol and flavone at neutral and acidic pH, and its color was analyzed by reflectance spectra and the visual modeling of their avian pollinators. The modeling demonstrated that the higher pH of the turquoise and pale-yellow species enhances the chromatic contrast and spectral purity. The precise regulation of flower color by flavonoid composition and vacuolar pH may be adapted to the visual perception of their avian pollinator vision.

[FLT3-ITD mutation-positive acute myeloid leukemia undergoing clonal transition with PTPN11 mutation at relapse].

A 28-year-old man was diagnosed with acute myelomonocytic leukemia. He achieved complete remission (CR) after two cycles of induction therapy. However, after consolidation therapy, bone marrow aspiration performed to prepare for allogeneic hematopoietic stem cell transplantation revealed disease relapse. Companion diagnostics confirmed the presence of the FLT3-ITD mutation. The patient received gilteritinib monotherapy and achieved CR. Subsequently, he underwent unrelated allogeneic bone marrow transplantation. One year after transplantation, the patient relapsed, and gilteritinib was resumed. However, the leukemia progressed, and panel sequencing using a next-generation sequencer showed that the FLT3-ITD mutation disappeared. A mutation in PTPN11, which regulates the RAS/MAPK signaling pathway, was also detected. Gilteritinib was discontinued, and the patient achieved CR with salvage chemotherapy. He underwent related haploidentical peripheral blood stem cell transplantation but died of relapse. This was a case in which genetic analysis revealed clonal transition and acquisition of resistance to treatment.

[Ruptured mycotic cerebral aneurysm in an adult T-cell leukemia/lymphoma patient undergoing allogeneic stem cell transplantation].

A 63-year-old man with adult T-cell leukemia-lymphoma underwent allogeneic bone marrow transplantation from an HLA-matched unrelated donor. On day 17 after transplantation, chest computed tomography (CT) showed nodules in the lower lobes of both lungs, and invasive pulmonary aspergillosis (IPA) was suspected. Treatment with liposomal amphotericin B was started, and improvement of infectious lesions was confirmed with CT on day 28. The antifungal agent was changed to voriconazole on day 52 because of progressive renal dysfunction. Disorders of consciousness and paralysis of the left upper and lower extremities developed on day 61. Brain CT showed subcortical hemorrhage in the right parietal and occipital lobes, and the patient died on day 62. An autopsy revealed filamentous fungi, suspected to be Aspergillus, in the pulmonary nodules and a ruptured cerebral aneurysm. Although IPA occurs in 10% of transplant recipients, vigilant monitoring for mycotic cerebral aneurysms is required to prevent hematogenous dissemination of Aspergillus, which is associated with a high mortality rate.

Comparison of fludarabine-based conditioning regimens in adult cord blood transplantation for myeloid malignancy: A retrospective, registry-based study.

Fludarabine/busulfan and fludarabine/melphalan are viable options as conditioning regimens. However, the optimal fludarabine-based conditioning in cord blood transplantation (CBT) remains unclear. Therefore, this retrospective, registry-based study aimed to analyze the impact of five fludarabine-containing conditioning regimens on 1395 adult patients (median age, 61 years) with acute myeloid leukemia, myelodysplastic syndrome, and chronic myeloid leukemia who underwent their first CBT. Treatment outcomes of fludarabine combined with melphalan (100-140 mg/m2 ) and low-dose total body irradiation (TBI; FM140T); melphalan (80-99 mg/m2 ) and TBI (FM80T); busulfan (12.8 mg/kg) and melphalan (FB4M); busulfan (12.8 mg/kg) and TBI (FB4T); and busulfan (6.4 mg/kg) and TBI (FB2T) were compared. The 3-year survival rate was 67%, 53%, 44%, 36%, and 39%, respectively (p < .0001). The FM140T survival rate was the most favorable after adjusting for confounders, and the hazard ratios (vs. FM140T) for overall mortality were as follows: FM80T, 1.6 (95% confidence interval [CI], 1.2-2.2); FB4M, 2.1 (95% CI, 1.6-2.8); FB4T, 2.7 (95% CI, 2.0-3.7); and FB2T, 2.2 (95% CI, 1.6-3.1). The better survival observed with FM140T, regardless of the disease, disease risk, age, or transplant year, was attributed to the lower relapse rate and lower non-relapse mortality (NRM) associated with fewer infectious deaths. Conversely, FB4T was associated with a higher relapse rate and higher NRM. The findings indicate that the outcomes of CBT in myeloid malignancies were highly dependent on both the alkylating agent and its dose in combination with fludarabine. Therefore, compared with fludarabine/busulfan-based conditioning, FM140T may be the preferred regimen.

[Acute promyelocytic leukemia with asymptomatic cerebral hemorrhage].

A 43-year-old man presenting with oral bleeding was diagnosed with acute promyelocytic leukemia (APL). Induction chemotherapy consisting of all-trans retinoic acid and idarubicin was initiated, and disseminated intravascular coagulation (DIC) was treated with fresh frozen plasma and recombinant thrombomodulin infusions. The patient was free from neurological symptoms throughout the clinical course. However, cerebral hemorrhagic lesions were detected incidentally on magnetic resonance imaging performed to screen for leukemic central nervous system invasion at 2 weeks after treatment initiation. Imaging findings suggested subacute or later-phase cerebral hemorrhage. Platelet transfusions and other supportive care was provided. Serial imaging evaluations confirmed reduction of the hemorrhagic lesions. Hematological remission was achieved after induction chemotherapy, and no symptoms due to cerebral hemorrhage developed during the subsequent consolidation therapy. As patients with APL characteristically experience hemorrhagic events due to bleeding tendency caused by DIC, physicians should be aware of the possibility of asymptomatic cerebral hemorrhage in these patients.

Impact of anti-thymocyte globulin on survival outcomes in female-to-male allogeneic hematopoietic stem cell transplantation.

Allogeneic hematopoietic cell transplantation between female donors and male recipients (female-to-male allo-HCT) is a well-established risk factor for inferior survival outcomes due to a higher incidence of graft-versus-host disease (GVHD). However, a clinical significance of anti-thymocyte globulin (ATG) in the female-to-male allo-HCT has not been elucidated. In this study, we retrospectively evaluated male patients who underwent allo-HCT between 2012 and 2019 in Japan. In the female-to-male allo-HCT cohort (n = 828), the use of ATG was not associated with a decreased risk of GVHD (HR of acute GVHD 0.691 [95% CI: 0.461-1.04], P = 0.074; HR of chronic GVHD 1.06 [95% CI: 0.738-1.52], P = 0.76), but was associated with favorable overall survival (OS) and a decreased risk of non-relapse mortality (NRM) (HR of OS 0.603 [95% CI: 0.400-0.909], P = 0.016; HR of NRM 0.506 [95% CI: 0.300-0.856], P = 0.011). The use of ATG in female-to-male allo-HCT resulted in survival outcomes that were almost equivalent to those in the male-to-male allo-HCT group. Therefore, GVHD prophylaxis with ATG might overcome the inferiority of survival outcomes in female-to-male allo-HCT.

Effect of Graft-versus-Host Disease on Post-Transplantation Outcomes following Single Cord Blood Transplantation Compared with Haploidentical Transplantation with Post-Transplantation Cyclophosphamide for Adult Acute Myeloid Leukemia.

The possibility that HLA mismatches could reduce relapse after alternative HLA-mismatched allogeneic hematopoietic cell transplantation (HCT) is an attractive concept for treating acute myeloid leukemia (AML). However, it remains unclear whether the prognostic effect of graft-versus-host disease (GVHD) on survival differs between recipients of single-unit cord blood transplantation (CBT) and recipients of haploidentical HCT using post-transplantation cyclophosphamide (PTCy-haplo-HCT) for AML. The objective of this retrospective study was to compare the effect of acute GVHD and chronic GVHD on post-transplantation outcomes between recipients of CBT and recipients of PTCy-haplo-HCT. We retrospectively evaluated the effect of acute and chronic GVHD on post-transplantation outcomes following CBT and PTCy-haplo-HCT in adults with AML (n = 1981) between 2014 and 2020 using a Japanese registry database. In univariate analysis, the probability of overall survival was significantly greater in patients who developed grade I-II acute GVHD (P < .001, log-rank test) and limited chronic GVHD (P < .001, log-rank test) among CBT recipients, but these effects were not significant among PTCy-haplo-HCT recipients. In multivariate analysis, in which the development of GVHD was treated as a time-dependent covariate, the effect of grade I-II acute GVHD on reducing overall mortality differed significantly between CBT and PTCy-haplo-HCT (adjusted hazard ratio [HR] for CBT, .73, 95% confidence interval [CI], .60 to .87; adjusted HR for PTCy-haplo-HCT, 1.07; 95% CI, .70 to 1.64; P for interaction = .038). Our data demonstrate that grade I-II acute GVHD was associated with a significant improvement in overall mortality in adults with AML receiving CBT but not in recipients of PTCy-haplo-HCT.

Risk factors for fatal cardiac complications after allogeneic hematopoietic cell transplantation: Japanese Society for Transplantation and Cellular Therapy transplant complications working group.

Fatal cardiac complications can occur from the early to late phases after hematopoietic cell transplantation (HCT). Herein, the Japanese transplant registry database was used to retrospectively analyze health records of 33,791 allogeneic HCT recipients to elucidate the pathogenesis and risk factors involved. Overall, 527 patients died of cardiac complications at a median of 130 (range 0-3924) days after HCT. The cumulative incidence of fatal cardiac complications was 1.2% (95% confidence interval [CI]: 1.0-1.3) and 1.6% (95% CI: 1.5-1.8) at 1 and 5 years after HCT, respectively. Fatal cardiovascular events were significantly associated with an HCT-specific comorbidity index (HCT-CI) score of ≥1 specific to the three cardiovascular items, lower performance status, conditioning regimen cyclophosphamide dose of >120 mg/kg, and female sex. Cardiovascular death risk within 60 days after HCT was associated with the type of conditioning regimen, presence of bacterial or fungal infections at HCT, and number of blood transfusions. Contrastingly, late cardiovascular death beyond 1 year after HCT was associated with female sex and older age. Lower performance status and positive cardiovascular disease-related HCT-CI were risk factors for cardiac complications in all phases after HCT. Systematic follow-up may be necessary according to the patients' risk factors and conditions.

Elevated mutation rates underlie the evolution of the aquatic plant family Podostemaceae.

Molecular evolutionary rates vary among lineages and influence the evolutionary process. Here, we report elevated genome-wide mutation rates in Podostemaceae, a family of aquatic plants with a unique body plan that allows members to live on submerged rocks in fast-flowing rivers. Molecular evolutionary analyses using 1640 orthologous gene groups revealed two historical increases in evolutionary rates: the first at the emergence of the family and the second at the emergence of Podostemoideae, which is the most diversified subfamily. In both branches, synonymous substitution rates were elevated, indicating higher mutation rates. On early branches, mutations were biased in favour of AT content, which is consistent with a role for ultraviolet light-induced mutation and habitat shift. In ancestors of Podostemoideae, DNA-repair genes were enriched in genes under positive selection, which may have responded to the meristem architectural changes.

Identification of anthocyanin and other flavonoids from the green-blue petals of Puya alpestris (Bromeliaceae) and a clarification of their coloration mechanism.

To understand the unique green-blue color of Puya alpestris (Bromeliaceae) flowers, we investigated their constituent anthocyanin and related compounds. An anthocyanin, two undescribed flavonols, and two flavones were isolated and identified as delphinidin 3,3',5'-tri-O-ß-glucopyranoside, myricetin 3-O-[α-rhamnopyranosyl-(1 â†’ 6)-ß-glucopyranoside]-3',5'-di-O-ß-glucopyranoside, myricetin 3,3',5'-tri-O-ß-glucopyranoside, luteolin 4'-O-glucoside, and apigenin 4'-O-glucoside. Furthermore, the presence of chlorophyll has also been revealed. P. alpestris petals show a gradient color appearance: Green-blue at the tip and blue at the base. This color difference between the tip and base was used to analyze the pigment components underlying the green-blue color expression. It was found that the petal tip contains the anthocyanin, flavonols, flavones, and chlorophyll in high quantities. Furthermore, the pH of petal juice was 6.2 and 5.6 at the tip and base, respectively. In vitro reconstruction revealed the blue color expression occurred via an intermolecular copigmentation between the anthocyanin and flavones, as well as yellow color expression, which was due to an increase in the absorption at 400-450 nm of the flavonols under the higher pH conditions. Furthermore, we found that the petal extract obtaining using 50% acetone containing chlorophyll showed the same absorption spectrum as that observed for the raw petal. These results indicate that the green-blue color of P. alpestris flowers is developed via an intermolecular co-pigmentation of the anthocyanin (delphinidin 3,3',5'-tri-O-ß-glucopyranoside) with flavones, such as luteolin 4'-O-glucoside, the yellow color expression of flavonols, such as myricetin 3,3',5'-tri-O-glucoside under relatively high pH conditions in the cell sap, and the presence of chlorophyll.

[Impaired hematopoiesis due to copper deficiency in a hemodialysis patient supplemented with zinc].

This is a case of a 75-year-old man who was on maintenance hemodialysis for 10 years due to diabetic nephropathy and was prescribed polaprezinc due to a low serum zinc level (55 µg/dl) and dysgeusia. Three months after the polaprezinc treatment was initiated, the patient developed pancytopenia, which persisted even after the serum zinc level was normalized and medication was discontinued. He was referred to our institute so that the progression of pancytopenia could be assessed. A blood biochemical examination revealed a WBC count of 1,700/µl, Hb level of 8.9 g/dl, and Plt count of 9.5×104/µl. A bone marrow aspirate smear showed slight megaloblastic changes and ringed sideroblasts in addition to an elevated WT1 mRNA level (76 copies/µg RNA) in the peripheral blood. Although these findings mimicked those of myelodysplasia, low serum copper (<2 µg/dl) and ceruloplasmin levels (3 mg/dl) were suggestive of hematopoietic abnormalities due to zinc-induced copper deficiency. Treatment with cocoa, a compound generally known to be rich in copper, gradually improved the pancytopenia and dysplastic bone marrow histology. This case indicates that clinicians should consider the risk of zinc-induced copper deficiency and its complications when zinc supplementation is administered to patients with chronic kidney disease, particularly those undergoing hemodialysis.

Nivolumab for Methotrexate-associated Classic Hodgkin's Lymphoma in a Rheumatoid Arthritis Patient.

Nivolumab exerts therapeutic activity in patients with classic Hodgkin's lymphoma (CHL) but may cause several types of immune-related adverse events. Some rheumatoid arthritis (RA) patients develop CHL during methotrexate therapy (MTX-CHL); however, the efficacy and safety of nivolumab for these patients remain unclear. A 68-year-old woman was diagnosed with CHL after six years of MTX therapy for RA. The disease did not respond to any type of chemotherapy. Nivolumab was then initiated, and the patient was successfully treated without the reactivation of RA. The reactivation of RA always needs to be considered with the administration of nivolumab.

Two new genera of Podostemaceae from northern Central Laos: saltational evolution and enigmatic morphology.

Podostemaceae, the river-weeds, are characterized by remarkable differences between species and genera, which resulted from saltational evolution. This paper presents additional cases of such two genera, which are described here from the Phou Khao Khouay National Protected Area in northern Central Laos. Molecular phylogenetic data show that Ctenobryum mangkonense (gen. & sp. nov.) is sister to Hydrodiscus koyamae, while Laosia ramosa (gen. & sp. nov.) is isolated from all Asian genera of subfamily Podostemoideae. Ctenobryum mangkonense is distinct from Hydrodiscus koyamae in the crustose roots (versus rootless in the latter), scattered flowers on the root (versus alternate on the shoot), and pectinate bracts (versus simple, sheath-like). Laosia ramosa is distinct from all the genera in the columnar, endogenously branched axes and single style-stigma complex. The axis is an enigmatic organ with combined characteristics of root, stem and leaf, pending further study. Laos, together with Thailand, is a center of diversity of the Southeast and East Asian Podostemoideae. The three monotypic genera, i.e., Ctenobryum, Hydrodiscus and Laosia, occur in neighboring and closely similar aquatic habitats within the Area. The new taxa are formally described.

An Evolutionarily Conserved Plant RKD Factor Controls Germ Cell Differentiation.

In contrast to animals, in which the germ cell lineage is established during embryogenesis, plant germ cells are generated in reproductive organs via reprogramming of somatic cells. The factors that control germ cell differentiation and reprogramming in plants are poorly understood. Members of the RKD subfamily of plant-specific RWP-RK transcription factors have been implicated in egg cell formation in Arabidopsis based on their expression patterns and ability to cause an egg-like transcriptome upon ectopic expression [1]; however, genetic evidence of their involvement is lacking, due to possible genetic redundancy, haploid lethality, and the technical difficulty of analyzing egg cell differentiation in angiosperms. Here we analyzed the factors that govern germ cell formation in the liverwort Marchantia polymorpha. This recently revived model bryophyte has several characteristics that make it ideal for studies of germ cell formation, such as low levels of genetic redundancy, readily accessible germ cells, and the ability to propagate asexually via gemma formation [2, 3]. Our analyses revealed that MpRKD, a single RWP-RK factor closely related to angiosperm RKDs, is preferentially expressed in developing eggs and sperm precursors in M. polymorpha. Targeted disruption of MpRKD had no effect on the gross morphology of the vegetative and reproductive organs but led to striking defects in egg and sperm cell differentiation, demonstrating that MpRKD is an essential regulator of germ cell differentiation. Together with previous findings [1, 4-6], our results suggest that RKD factors are evolutionarily conserved regulators of germ cell differentiation in land plants.

Fate of HIV-1 cDNA intermediates during reverse transcription is dictated by transcription initiation site of virus genomic RNA.

Retroviral reverse transcription is accomplished by sequential strand-transfers of partial cDNA intermediates copied from viral genomic RNA. Here, we revealed an unprecedented role of 5'-end guanosine (G) of HIV-1 genomic RNA for reverse transcription. Based on current consensus for HIV-1 transcription initiation site, HIV-1 transcripts possess a single G at 5'-ends (G1-form). However, we found that HIV-1 transcripts with additional Gs at 5'-ends (G2- and G3-forms) were abundantly expressed in infected cells by using alternative transcription initiation sites. The G2- and G3-forms were also detected in the virus particle, although the G1-form predominated. To address biological impact of the 5'-G number, we generated HIV clone DNA to express the G1-form exclusively by deleting the alternative initiation sites. Virus produced from the clone showed significantly higher strand-transfer of minus strong-stop cDNA (-sscDNA). The in vitro assay using synthetic HIV-1 RNAs revealed that the abortive forms of -sscDNA were abundantly generated from the G3-form RNA, but dramatically reduced from the G1-form. Moreover, the strand-transfer of -sscDNA from the G1-form was prominently stimulated by HIV-1 nucleocapsid. Taken together, our results demonstrated that the 5'-G number that corresponds to HIV-1 transcription initiation site was critical for successful strand-transfer of -sscDNA during reverse transcription.

Gene expression analysis of aquatic angiosperms podostemaceae to gain insight into the evolution of their enigmatic morphology.

Podostemaceae is a family of aquatic angiosperms growing submerged on rocks in fast-flowing water and called moss-like or alga-like riverweeds. It evolved remarkable innovations to adapt to such an extreme environment, one of which is reduced shoots borne on roots adhering to rock surface. Recent observations revealed that the basal subfamily Tristichoideae, like most other angiosperms, has typical shoot apical meristems (SAMs). In species of the subfamily Podostemoideae, however, shoot apical meristems (SAMs) are not formed during development and new leaves arise from the meristematic basal region of preexisting leaves. The genetic basis of this shoot organogenesis process, e.g., the expression patterns of genes homologous to transcription factors regulating shoot development, is essential to better understand the evolution of Podostemaceae. A gene expression analysis found that the SAM-less Podostemoideae leaf has mixed identity of SAM and leaf, and provided insight into the evolution of the shoot in Podostemaceae.

Non-cell-autonomous microRNA165 acts in a dose-dependent manner to regulate multiple differentiation status in the Arabidopsis root.

In the development of multicellular organisms, cell fate is usually determined by exchanging positional information. Animals employ a class of intercellular signaling molecules that specify different cell fates by their dosage, but the existence of an equivalent system has not been demonstrated in plants, except that the growth regulator auxin has been proposed to act in a similar manner in certain developmental contexts. Recently, it has been reported that, in the Arabidopsis root meristem, endodermis-derived microRNA (miR) 165/166 non-cell-autonomously suppress the expression of the Class III HD-ZIP transcription factor PHABULOSA (PHB) in the peripheral stele, thereby specifying xylem differentiation. Here, we show that the miR165/166-dependent suppression of PHB is required not only for xylem specification, but also for differentiation of the pericycle, as well as for ground tissue patterning. Furthermore, using a plant system that allows quantitative control of miR165 production in the ground tissue, we show that endodermis-derived miR165 acts in a dose-dependent manner to form a graded distribution of PHB transcripts across the stele. These results reveal a previously unidentified role of miR165 in the differentiation of a broad range of root cell types and suggest that endodermis-derived miR165 acts in a dose-dependent manner to control multiple differentiation status in the Arabidopsis root.

Expression of SHOOT MERISTEMLESS, WUSCHEL, and ASYMMETRIC LEAVES1 homologs in the shoots of Podostemaceae: implications for the evolution of novel shoot organogenesis.

Podostemaceae (the river weeds) are ecologically and morphologically unusual angiosperms. The subfamily Tristichoideae has typical shoot apical meristems (SAMs) that produce leaves, but Podostemoideae is devoid of SAMs and new leaves arise below the base of older leaves. To reveal the genetic basis for the evolution of novel shoot organogenesis in Podostemaceae, we examined the expression patterns of key regulatory genes for shoot development (i.e., SHOOT MERISTEMLESS (STM), WUSCHEL (WUS), and ASYMMETRIC LEAVES1/ROUGH SHEATH2/PHANTASTICA (ARP) orthologs) in Tristichoideae and Podostemoideae. In the SAM-mediated shoots of Tristichoideae, like in model plants, STM and WUS orthologs were expressed in the SAM. In the SAM-less shoots of Podostemoideae, STM and WUS orthologs were expressed in the initiating leaf/bract primordium. In older leaf/bract primordia, WUS expression disappeared and STM expression became restricted to the basal part, whereas ARP was expressed in the distal part in a complementary pattern to STM expression. In the reproductive shoots of Podostemoideae with a normal mode of flower development, STM and WUS were expressed in the floral meristem, but not in the floral organs, similar to the pattern in model plants. These results suggest that the leaf/bract of Podostemoideae is initiated as a SAM and differentiates into a single apical leaf/bract, resulting in the evolution of novel shoot-leaf mixed organs in Podostemaceae.

Developmental morphology of seedling and shoot and phylogenetic relationship of Diplobryum koyamae (Podostemaceae).

We demonstrated that saltational evolution was recurrent in the body plans of seedlings of the aquatic angiosperm Podostemaceae, in contrast to other angiosperms with seedlings having almost common body plans. Diplobryum koyamae, transferred to the genus Hydrodiscus described in this paper, has long-floating shoots with an anchoring disk-like base and is rootless. Such a body plan is distinct from other members of Asian Podostemoideae comprising reduced or moderate shoots borne on the root. Here, our molecular phylogenetic analysis revealed that H. koyamae is sister to a crustose-rooted group of Hanseniella, Hydrobryum and Thawatchaia within Asian Podostemoideae. The germinating embryo was devoid of plumule and radicle, and comprised a single cotyledon and a short hypocotyl, which produced an adventitious shoot endogenously. The leaves are formed in the absence of the shoot apical meristem, accompanying the separation of lightly stained cells. Comparison with other species of Asian Podostemoideae having the plumule and the adventitious root in the seedling, along with their phylogenetic relationship, suggests that saltational evolution occurred in the seedling body plan of H. koyamae leading to the extraordinary adult body plan, as in the separate clade of Dalzellia, Indodalzellia, and Indotristicha of the subfamily Tristichoideae.