Outcome of twin-to-twin transfusion syndrome in monochorionic monoamniotic twin pregnancy: systematic review and meta-analysis.

OBJECTIVES: To explore the outcome of monochorionic monoamniotic (MCMA) twin pregnancies affected by twin-to-twin transfusion syndrome (TTTS). METHODS: MEDLINE and EMBASE databases were searched for studies reporting the outcome of MCMA twin pregnancies complicated by TTTS. The primary outcome was intrauterine death (IUD); secondary outcomes were miscarriage, single IUD, double IUD, neonatal death (NND), perinatal death (PND), survival of at least one twin, survival of both twins and preterm birth (PTB) before 32 weeks' gestation. Outcomes were assessed in MCMA twins affected by TTTS not undergoing intervention and in those treated with amniodrainage, laser therapy or cord occlusion. Subgroup analysis was performed including cases diagnosed before 24 weeks. Random-effects meta-analysis of proportions was used to analyze the data. RESULTS: Fifteen cohort studies, including 888 MCMA twin pregnancies, of which 44 were affected by TTTS, were included in the review. There was no randomized trial comparing the different management options in MCMA twin pregnancies complicated by TTTS. In cases not undergoing intervention, miscarriage occurred in 11.0% of fetuses, while the incidence of IUD, NND and PND was 25.2%, 12.2% and 31.2%, respectively. PTB complicated 50.5% of these pregnancies. In cases treated by laser surgery, the incidence of miscarriage, IUD, NND and PND was 19.6%, 27.4%, 7.4% and 35.9%, respectively, and the incidence of PTB before 32 weeks' gestation was 64.9%. In cases treated with amniodrainage, the incidence of IUD, NND and PND was 31.3%, 13.5% and 45.7% respectively, and PTB complicated 76.2% of these pregnancies. Analysis of cases undergoing cord occlusion was affected by the very small number of included cases. Miscarriage occurred in 19.2%, while there was no case of IUD or NND of the surviving twin. PTB before 32 weeks occurred in 50.0% of these cases. CONCLUSIONS: MCMA twin pregnancies complicated by TTTS are at high risk of perinatal mortality and PTB. Further studies are needed in order to elucidate the optimal type of prenatal treatment in these pregnancies. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Time-Reversal Measurement of the p-Wave Cross Sections of the

The cross sections of the ^{7}Be(n,α)^{4}He reaction for p-wave neutrons were experimentally determined at E_{c.m.}=0.20-0.81 MeV slightly above the big bang nucleosynthesis (BBN) energy window for the first time on the basis of the detailed balance principle by measuring the time-reverse reaction. The obtained cross sections are much larger than the cross sections for s-wave neutrons inferred from the recent measurement at the n_TOF facility in CERN, but significantly smaller than the theoretical estimation widely used in the BBN calculations. The present results suggest the ^{7}Be(n,α)^{4}He reaction rate is not large enough to solve the cosmological lithium problem, and this conclusion agrees with the recent result from the direct measurement of the s-wave cross sections using a low-energy neutron beam and the evaluated nuclear data library ENDF/B-VII.1.

Prediction of time to delivery by transperineal ultrasound in second stage of labor.

OBJECTIVE: To investigate whether the transperineal sonographic (TPS) parameters angle of progression (AoP) and midline angle (MLA) can predict the time remaining in the second stage of labor. METHODS: We evaluated prospectively women with a singleton pregnancy in cephalic presentation at term between October 2013 and September 2014. TPS volumes were obtained immediately after confirmation by digital vaginal examination of a fully dilated cervix. AoP and MLA were measured offline by analyzing the ultrasound volumes. Progression of labor was evaluated every hour during the second stage. The associations of AoP and MLA with the interval between TPS assessment and delivery were evaluated using multivariable Cox proportional hazards analyses in nulliparous and parous women separately. RESULTS: A total of 557 women were evaluated. An AoP ≥ 160° (adjusted hazard ratio (aHR), 2.52 (95% CI, 1.98-3.19)) and MLA ≤ 10° (aHR, 1.79 (95% CI, 1.35-2.34)) in nulliparous women and an AoP ≥ 150° (aHR, 1.86 (95% CI, 1.34-2.57)) and MLA ≤ 20° (aHR, 1.69 (95% CI, 1.21-2.34)) in parous women were significantly associated with the remaining time in labor. The positive/negative likelihood ratios of AoP, MLA, clinical station (fetal head descent as observed by digital examination) and clinical rotation (fetal head rotation as observed by digital examination) at these cut-off points were 3.6/0.6, 2.0/0.6, 1.6/0.6 and 1.6/0.8, respectively, in nulliparous women, and 2.4/0.6, 1.3/0.7, 7.6/0.5 and 5.2/0.7, respectively, in parous women. CONCLUSION: TPS assessment of AoP and MLA in the second stage of labor was useful for predicting the time remaining in labor and had higher predictive value than did digital vaginal examination in nulliparous women. Copyright © 2016 ISUOG. Published by John Wiley & Sons Ltd.

Effect of an impaired oral stage on swallowing in patients with Parkinson's disease.

We investigated the swallowing function in patients with Parkinson's disease (PD) using deteriorated tongue control because patients with PD frequently exhibit an impaired oral stage of swallowing and the tongue movement affects oral and pharyngeal stage. In total, 201 patients with PD (106 men, 95 women; mean age 70·6 ± 8·0 years; median Hoehn-Yahr Stage III) were studied. The patients swallowed 10 mL of liquid barium under videofluorography, and their oral transit time (OTT) was measured. Based on 20 healthy controls (mean age 70·3 ± 7·8 years) with an OTT + 2 standard deviation (0·89 + 2 × 0·46) of 1·81 s, the patients with PD were divided into 167 patients with an OTT < 1·81 s and 34 patients with an OTT ≥ 1·81 s. Swallowing function was compared between the groups and assessed using logistic regression analysis. The following factors were significantly associated with oral stage impairment in both groups: tongue-to-palate contact, tongue root-to-posterior pharyngeal wall contact, premature spillage into the pharynx, aspiration and onset of swallowing reflex. Logistic regression analysis showed that tongue root-to-posterior pharyngeal wall contact, onset of swallowing reflex and aspiration were independent factors. PD patients with prolonged OTT displayed poor lingual control and decreased range of motion of the tongue due to bradykinesia and rigidity. Such problems in the oral stage affected the subsequent pharyngeal stage of swallowing with aspiration. Lingual movement in the oral stage thus appears to play an important role in the sequential movement of swallowing in PD.

Distinct X chromosomal rearrangements in four haemophilia B patients with entire F9 deletion.

INTRODUCTION: Haemophilia B is an X-linked bleeding disorder caused by a coagulation factor IX gene (F9) abnormality. Numerous F9 defects have been identified to date; however, only a few with an entire F9 deletion have been reported in detail. AIM: To elucidate the cause of severe haemophilia B, we investigated the precise X chromosome abnormalities in four Japanese patients who did not show all amplifications in F9-specific PCR. METHODS: We analysed the patient's genomic DNA using Multiplex ligation-dependent probe amplification (MLPA). To assess the extent of any deletions, we further performed mapping PCRs, inverse PCRs or long-range PCRs and direct sequencing analyses of the X chromosome. RESULTS: We detected entire F9 deletions in four haemophilia B patients and identified the precise deleted regions of the X chromosome including F9. Patient 1 had a 149-kb deletion with breakpoints 90-kb upstream and 30-kb downstream from F9. Patients 2 and 3 showed 273-kb and 1.19-Mb deletions respectively. Patient 4 had two deleted regions: a 1663-bp deletion 1.34-Mb upstream from F9 and a 7.2-Mb deletion including F9. These distinct breakpoints found in four different patients suggest that the mechanism of X chromosome deletion may be different between individuals. Non-allelic homologous recombination (NAHR), microhomology-mediated break-induced replication (MMBIR) or fork stalling and template switching (FoSTeS) may occur in respective X chromosomes of the four haemophilia B patients analysed. CONCLUSIONS: We identified diverse X chromosomal rearrangements in four haemophilia B patients, which might be caused by distinct mechanisms of genomic rearrangement.

Species and sex differences in propofol glucuronidation in liver microsomes of humans, monkeys, rats and mice.

Propofol (2,6-diisopropylphenol) is a short-acting anesthetic commonly used in clinical practice, and is rapidly metabolized into glucuronide by UDP-glucuronosyltransferase (UGT). In the present study, propofol glucuronidation was examined in the liver microsomes of male and female humans, monkeys, rats, and mice. The kinetics of propofol glucuronidation by liver microsomes fit the substrate inhibition model for humans and mice, the Hill model for monkeys, and the isoenzyme (biphasic) model for rats. The K(m), V(max), and CL(int) values of human liver microsomes were 50 µM, 5.6 nmol/min/mg protein, and 110 µL/min/mg protein, respectively, for males, and 46 µM, 6.0 nmol/min/mg protein, and 130 µL/min/mg protein, respectively, for females. The rank order of the CL(int) or CL(max) (in vitro clearance) values of liver microsomes was mice humans > monkeys > rats (high-affinity phase) rats (low-affinity phase) in both males and females. Although no significant sex differences were observed in the values of kinetic parameters in any animal species, the in vitro clearance values of liver microsomes were males < females in humans, males = females in rats (low-affinity phase), and males > females in monkeys, rats (high-affinity phase), and mice. These results demonstrated that the kinetic profile of propofol glucuronidation by liver microsomes markedly differed among humans, monkeys, rats, and mice, and suggest that species and sex differences exist in the roles of UGT isoform(s), including UGT1A9, involved in its metabolism.

Reversible phospho-Smad3 signalling between tumour suppression and fibrocarcinogenesis in chronic hepatitis B infection.

Transforming growth factor (TGF)-ß, type I receptor (TßRI) and c-Jun N-terminal kinases (JNK) phosphorylate Smad3 differentially to create 2 isoforms phosphorylated (p) at the COOH-terminus (C) or at the linker region (L) and regulate hepatocytic fibrocarcinogenesis. This study aimed to compare the differences between how hepatitis B virus (HBV) infection affected hepatocytic Smad3 phosphorylated isoforms before and after anti-viral therapy. To clarify the relationship between Smad3 phosphorylation and liver disease progression, we studied 10 random patients in each stage of HBV-related fibrotic liver disease (F1-4) and also 10 patients with HBV-associated HCC. To examine changes in phosphorylated Smad3 signalling before and after anti-HBV therapies, we chose 27 patients with chronic hepatitis B who underwent baseline and follow-up biopsies at 52 weeks from the start of nucleoside analogue treatments (Lamivudine 100 mg daily or Telbivudine 600 mg daily). Fibrosis stage, inflammatory activity and phosphorylated Smad3 positivity in the paired biopsy samples were compared. Hepatocytic pSmad3C signalling shifted to fibrocarcinogenic pSmad3L signalling as the livers progressed from chronic hepatitis B infection to HCC. After nucleoside analogue treatment, serum alanine aminotransferase (ALT) and HBV-DNA levels in 27 patients with HBV-related chronic liver diseases were decreased dramatically. Decrease in HBV-DNA restored pSmad3C signalling in hepatocytes, while eliminating prior fibrocarcinogenic pSmad3L signalling. Oral nucleoside analogue therapies can suppress fibrosis and reduce HCC incidence by successfully reversing phosphorylated Smad3 signalling; even liver disease progressed to cirrhosis in chronic hepatitis B patients.

Clinical characteristics of monochorionic twins with large hemoglobin level discordance at birth.

OBJECTIVES: To evaluate neonatal outcomes and clinical characteristics of monochorionic diamniotic (MCDA) twins with a large intertwin hemoglobin (Hb) difference at birth. METHODS: This was a retrospective cohort study of MCDA twin gestations delivered at Osaka Medical Center and Research Institute for Maternal and Child Health between 2003 and 2012. Cases of pregnancy termination, acardiac twins or intrauterine death were excluded. A large intertwin Hb difference at birth was defined as > 8.0 g/dL according to the postnatal criteria for twin anemia-polycythemia sequence (TAPS). The intertwin reticulocyte count ratio (RCR) was calculated by dividing the reticulocyte count of the anemic twin by that of the polycythemic twin. Cases with Hb differences were divided into two groups according to the RCR, TAPS when the RCR was > 1.7 and acute fetofetal hemorrhage (AFFH) when the RCR was ≤ 1.7. Neonatal outcomes were compared between the TAPS and AFFH groups. RESULTS: During the study period, 432 MCDA twin pregnancies of a total of 532 born at our hospital were analyzed. There were 12 (2.8%) cases of a large intertwin Hb difference. The median gestational age at birth of these cases was 34 (range, 23-38) weeks, and all were delivered by Cesarean section. There were seven (1.6%) cases of TAPS and five (1.2%) of AFFH. The neonatal survival rate was 91.7%; in one pair of twins with TAPS neonatal death occurred. All (100%) cases with TAPS and two (40%) with AFFH required blood transfusion or partial-exchange transfusion for at least one infant. CONCLUSIONS: Although the incidence of TAPS and AFFH may be low in MCDA twins, many affected neonates required treatment for hematological abnormalities. Delivery of MCDA twins via Cesarean section does not appear to prevent AFFH, despite the absence of labor.

Oral administration of bovine lactoferrin attenuates ultraviolet B-induced skin photodamage in hairless mice.

Lactoferrin (LF) is recognized as a host defensive glycoprotein, especially for newborn infants. The aim of this study was to investigate whether orally administered LF had protective activity against UV-induced skin damage in hairless mice. Transepidermal water loss and skin hydration were evaluated in nonirradiated mice, UVB-irradiated mice, and UVB-irradiated and LF-administered mice. Supplementation with LF (1,600 mg/kg per day) effectively suppressed the increase in transepidermal water loss, reduction in skin hydration, aberrant epidermal hyperplasia, and cell apoptosis induced by UV irradiation. Although no significant changes in superoxide dismutase-like activity or malondialdehyde levels were observed in the skin with both UV irradiation and LF administration, UV-stimulated IL-1ß levels in the skin were significantly suppressed by the administration of LF. Oral supplementation with LF has the potential to reduce IL-1ß levels and prevent UV-induced skin damage. Further studies are needed to elucidate the relationships between the antiinflammatory effects and skin protective function of LF.

In vitro glucuronidation of propofol in microsomal fractions from human liver, intestine and kidney: tissue distribution and physiological role of UGT1A9.

Propofol (2,6-diisopropylphenol) is intravenously administered for anesthetic induction and maintenance, and is rapidly metabolized into its glucuronide, mainly by UDP-glucuronosyltransferase 1A9 (UGT1A9). In this study, propofol glucuronidation by liver microsomes (HLM), intestinal microsomes (HIM) and kidney microsomes (HKM) of humans were examined. The expression of UGT1A9 protein in HLM, HIM and HKM was analyzed by immunoblotting. The staining band intensities for UGT1A9 of HIM and HKM were 12% and 119% those of HLM, respectively. The kinetics of propofol glucuronidation by HLM and HKM exhibited substrate inhibition, whereas the kinetics by HIM followed the Michaelis-Menten model. The K(m), V(max) and CL(int) values of HLM were 41.8 µM, 5.21 nmol/min/mg protein and 126 µl/min/mg protein, respectively. The K(m) value of HIM was significantly higher (6.7-fold) than that of HLM, and the V(max) and CL(int) values were significantly lower (56% and 8.3%, respectively) than those of HLM. The K(m) value of HKM was comparable to that of HLM, and the V(max) and CL(int) values were significantly higher (2.1- and 3.7-fold, respectively) than those of HLM, respectively. These findings suggest that UGT1A9 expressed in the kidney as well as in the liver plays an important role in propofol glucuronidation. The information gained in this study should contribute to an appropriate use of drugs metabolized by UGT1A9.

Valuable antiviral therapeutic options for the treatment of chronic hepatitis C patients with thrombocytopenia.

Thrombocytopenia in patients with chronic hepatitis C may represent an obstacle for the initiation of antiviral treatment. The aim of this study was to evaluate factors predictive of successful pegylated interferon (PEG-IFN) α2b and ribavirin (RBV) treatment for patients with thrombocytopenia with no history of splenectomy or partial splenic embolization. One hundred and fifty-one chronic hepatitis C patients (genotype 1: n = 110, genotype 2: n = 41) with TCP (<100 × 10(9) /L) at baseline were enrolled. Pretreatment variables included interleukin 28B (IL28B) genotype (rs8099917) and homoeostasis model assessment of insulin resistance score (HOMA-IR). The kinetics of haemoglobin and platelets according to the inosine triphosphatase (ITPA) genotype (rs1127354) were investigated. Sustained virological response (SVR) was significantly more frequent in hepatitis C virus (HCV) genotype 2 (65.9%) than in genotype 1 (34.5%) patients (P < 0.0001). Multiple logistic regression analysis of HCV genotype 1 extracted IL28B TT genotype [odds ratio (OR) 5.97, P = 0.006] and HOMA-IR <2.5 (OR 7.14, P = 0.0016) as significant independent pretreatment predictors of SVR. The analyses of HCV genotype 2 showed that HOMA-IR was significantly related to SVR, but IL28B genotype was not. Patients with ITPA CC genotype showed a significant haemoglobin reduction and lower degree of platelets decrease than those with ITPA CA/AA genotypes. The most common reason for premature discontinuation of treatment was the development of hepatocellular carcinoma (n = 8, 5.3%). In conclusion, HOMA-IR is a useful predictor of SVR for patients with thrombocytopenia infected with HCV genotype 1 or 2 treated with PEG-IFNα2b and RBV. The inclusion of IL28B, ITPA genotypes and HOMA-IR adds valuable therapeutic information.

Persistent parvovirus B19 infection resulting in red cell aplasia after allogeneic hematopoietic stem cell transplantation.

Persistent parvovirus B19 (PVB) infection has been reported sporadically in immunocompromised patients including hematopoietic stem cell and solid organ transplant recipients. However, the pathogenesis of persistent infection has yet to be fully elucidated. We report here a patient with multiple myeloma developing red cell aplasia during the hematopoietic recovery after allogeneic hematopoietic stem cell transplantation (HSCT) caused by PVB. The patient had already had PVB viremia before transplantation and remained asymptomatic. The route of PVB transmission was considered to be direct contact with the patient's family member with primary PVB infection 1 month before transplantation. Treatment with intravenous immunoglobulin resulted in prompt resolution of anemia. These findings suggest that monitoring of PVB DNA is recommended for patients undergoing HSCT and having contact with individuals with documented PVB infection, even if they are asymptomatic.

Identification of milk proteins enhancing the antimicrobial activity of lactoferrin and lactoferricin.

Lactoferrin (LF) is known as an iron-binding antimicrobial protein present in exocrine secretions such as milk and releases the potent antimicrobial peptide lactoferricin (LFcin) by hydrolysis with pepsin. The antimicrobial activity of LF and LFcin has been studied well; however, their cooperative action with other milk proteins remains to be elucidated. In this study, we identified milk proteins enhancing the antimicrobial activity of bovine LF and LFcin against gram-negative bacteria, gram-positive bacteria, and fungi. As the target fraction, we isolated a minor milk protein fraction around 15 kDa, which was identified as bovine RNase 5 (angiogenin-1), RNase 4, and angiogenin-2 by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry. As these proteins are collectively known as the RNase A family, we referred to the target protein fraction as milk RNase of 15 kDa (MR15). The number of colony-forming units of Escherichia coli and other pathogenic microorganisms with the addition of MR15 to LF (MR15:LF ratio=16:1,000) was dramatically lowered than that with LF alone. On the other hand, MR15 itself did not show any reductions in the number of colony-forming units at the concentrations tested. Similarly, the antimicrobial activities of LFcin against various microorganisms were significantly enhanced by the addition of MR15. These results suggest that LF and MR15 may be concomitantly acting antimicrobial agents in milk.

AXIN1 mutations in hepatocellular carcinomas, and growth suppression in cancer cells by virus-mediated transfer of AXIN1.

The Wnt signaling pathway is essential for development and organogenesis. Wnt signaling stabilizes beta-catenin, which accumulates in the cytoplasm, binds to 1-cell factor (TCF; also known as lymphocyte enhancer-binding factor, LEF) and then upregulates downstream genes. Mutations in CTNNB1 (encoding beta-catenin) or APC (adenomatous polyposis coli) have been reported in human neoplasms including colon cancers and hepatocellular carcinomas (HCCs). Because HCC5 tend to show accumulation of beta-catenin more often than mutations in CTNNB1, we looked for mutations in AXIN1, encoding a key factor for Wnt signaling, in 6 HCC cell lines and 100 primary HCC5. Among the 4 cell lines and 87 HCC5 in which we did not detect CTNNB1 mutations, we identified AXIN1 mutations in 3 cell lines and 6 mutations in 5 of the primary HCCs. In cell lines containing mutations in either gene, we observed increased DNA binding of TCF associated with beta-catenin in nuclei. Adenovirus mediated gene transfer of wild-type AXINI induced apoptosis in hepatocellular and colorectal cancer cells that had accumulated beta-catenin as a consequence of either APC, CTNNB1 or AXIN1 mutation, suggesting that axin may be an effective therapeutic molecule for suppressing growth of hepatocellular and colorectal cancers.

Fungemia due to Rhodotorula mucilaginosa after allogeneic hematopoietic stem cell transplantation.

Rhodotorula species have been increasingly recognized as emerging pathogens, particularly in immunocompromised patients. We herein report on a patient with myelodysplastic syndrome who developed fungemia due to Rhodotorula mucilaginosa after allogeneic hematopoietic stem cell transplantation (HSCT) from an unrelated donor. He developed severe acute graft-versus-host disease requiring high-dose steroids, and had serially been administered fluconazole and micafungin for the prophylaxis of fungal infection. Although several cases of Rhodotorula infection after HSCT have been reported, all of them were recipients of autologous HSCT, not allogeneic HSCT. A review of all the reported cases of Rhodotorula infection after HSCT revealed that all patients had received fluconazole or echinocandins before the onset of infection. The findings suggest that Rhodotorula species could be causative yeasts, particularly in patients receiving fluconazole or echinocandins, both of which are inactive against the species.

[Intraoperative direct angiography for the diminutive central pulmonary artery in a patient with major aortopulmonary collateral arteries].

It can be difficult to judge the degree of arborization of diminutive central pulmonary arteries (cPA) in patients with major aortopulmonary collateral arteries (MAPCA). Even through preoperative cardiac catheterization may not give adequate information. We introduce intra-operative direct angiography of diminutive cPA for patients with MAPCA. This would be one of the good options to judge the degree of arborization of the diminutive cPA, and to decide an initial surgical treatment. In this case, unifocalization of MAPCA without patch augmentation of pulmonary arteries, and an aortopulmonary shunt were performed at the 1st procedure. As enough growth of the cPA was obtained, this patient did not require additional patch augmentation of the pulmonary artery at the time of complete repair.

A prospective dose-finding trial using a modified continual reassessment method for optimization of fludarabine plus melphalan conditioning for marrow transplantation from unrelated donors in patients with hematopoietic malignancies.

BACKGROUND: Because of the less graft-facilitating effect by bone marrow (BM), we need to assess a dosage of conditioning more accurately particularly in combination with reduced-intensity conditioning. Thus we examined that modified continual reassessment method (mCRM) is applicable for deciding appropriate conditioning of allogeneic BM transplantation. PATIENTS AND METHODS: The conditioning regimen consisted of i.v. fludarabine (125 mg/m2) plus an examination dose of i.v. melphalan. The primary endpoint was a donor-type T-cell chimerism at day 28 with successful engraftment defined as >90% donor cells. Five patients per dose level were planned to be accrued and chimerism data were used to determine the next dose. RESULTS: Seventeen patients were enrolled at doses between 130 and 160 mg/m2. The dose was changed from 160 to 130 mg/m(2) (second level) after five full-donor chimerisms. With one patient of 0% chimera in the second level, the dose was increased to 135 mg/m2 (third level). Following five full-donor chimerisms in the third level, the study was complete as projected. CONCLUSIONS: mCRM was shown to be a relevant method for dose-finding of conditioning regimen. The melphalan dose of 135 mg/m2 was determined as the recommended phase II dose to induce initial full-donor chimerism.

Holocentric chromosomes of Luzula elegans are characterized by a longitudinal centromere groove, chromosome bending, and a terminal nucleolus organizer region.

The structure of holocentric chromosomes was analyzed in mitotic cells of Luzula elegans. Light and scanning electron microscopy observations provided evidence for the existence of a longitudinal groove along each sister chromatid. The centromere-specific histone H3 variant, CENH3, colocalized with this groove and with microtubule attachment sites. The terminal chromosomal regions were CENH3-negative. During metaphase to anaphase transition, L. elegans chromosomes typically curved to a sickle-like shape, a process that is likely to be influenced by the pulling forces of microtubules along the holocentric axis towards the corresponding microtubule organizing regions. A single pair of 45S rDNA sites, situated distal to Arabidopsis-telomere repeats, was observed at the terminal region of one chromosome pair. We suggest that the 45S rDNA position in distal centromere-free regions could be required to ensure chromosome stability.

A large inversion in the linear chromosome of Streptomyces griseus caused by replicative transposition of a new Tn3 family transposon.

We have comprehensively analyzed the linear chromosomes of Streptomyces griseus mutants constructed and kept in our laboratory. During this study, macrorestriction analysis of AseI and DraI fragments of mutant 402-2 suggested a large chromosomal inversion. The junctions of chromosomal inversion were cloned and sequenced and compared with the corresponding target sequences in the parent strain 2247. Consequently, a transposon-involved mechanism was revealed. Namely, a transposon originally located at the left target site was replicatively transposed to the right target site in an inverted direction, which generated a second copy and at the same time caused a 2.5-Mb chromosomal inversion. The involved transposon named TnSGR was grouped into a new subfamily of the resolvase-encoding Tn3 family transposons based on its gene organization. At the end, terminal diversity of S. griseus chromosomes is discussed by comparing the sequences of strains 2247 and IFO13350.

Structural features of membrane fusion between influenza virus and liposome as revealed by quick-freezing electron microscopy.

The structure of membrane fusion intermediates between the A/PR/8(H1N1) strain of influenza virus and a liposome composed of egg phosphatidylcholine, cholesterol, and glycophorin was studied using quick-freezing electron microscopy. Fusion by viral hemagglutinin protein was induced at pH 5.0 and 23 degrees C. After a 19-s incubation under these conditions, small protrusions with a diameter of 10-20 nm were found on the fractured convex faces of the liposomal membranes, and small pits complementary to the protrusions were found on the concave faces. The protrusions and pits corresponded to fractured parts of outward bendings of the lipid bilayer or "microprotrusions of the lipid bilayer." At the loci of the protrusions and pits, liposomal membranes had local contacts with viral membranes. In many cases both the protrusions and the pits were aligned in regular polygonal arrangements, which were thought to reflect the array of hemagglutinin spikes on the viral surface. These structures were induced only when the medium was acidic with the virus present. Based on these observations, it was concluded that the microprotrusions of the lipid bilayer are induced by hemagglutinin protein. Furthermore, morphological evidence for the formation of the "initial fusion pore" at the microprotrusion was obtained. The protrusion on the convex face sometimes had a tiny hole with a diameter of <4 nm in the center. The pits transformed into narrow membrane connections <10 nm in width, bridging viruses and liposomes. The structures of the fusion pore and fusion neck with larger sizes were also observed, indicating growth of the protrusions and pits to distinct fusion sites. We propose that the microprotrusion of the lipid bilayer is a fusion intermediate induced by hemagglutinin protein, and suggest that the extraordinarily high curvature of this membrane structure is a clue to the onset of fusion. The possible architecture of the fusion intermediate is discussed with regard to the localization of intramembrane particles at the microprotrusion.