Pollen differentiation as well as pollen tube guidance and discharge are independent of the presence of gametes.

After meiosis, an unequal cell division generates the male gamete lineage in flowering plants. The generative cell will undergo a second division, giving rise to the two gametes, i.e. the sperm cells. The other cell will develop into the vegetative cell that plays a crucial role in pollen tube formation and sperm delivery. Recently, the vegetative cell has been suggested to be important for programming of the chromatin state in sperm cells and/or the resulting fertilization products. Blocking the initial unequal division genetically, we first highlight that the default differentiation state after male meiosis is a vegetative fate, which is consistent with earlier work. We find that uni-nucleated mutant microspores differentiated as wild-type vegetative cells, including chromatin remodeling and the transcriptional activation of transposable elements. Moreover, live-cell imaging revealed that this vegetative cell is sufficient for the correct guidance of the pollen tube to the female gametes. Hence, we conclude that vegetative cell differentiation and function does not depend on the formation or presence of the actual gametes but rather on external signals or a cell-autonomous pace keeper.

Genome-wide transcript profiling of endosperm without paternal contribution identifies parent-of-origin-dependent regulation of AGAMOUS-LIKE36.

Seed development in angiosperms is dependent on the interplay among different transcriptional programs operating in the embryo, the endosperm, and the maternally-derived seed coat. In angiosperms, the embryo and the endosperm are products of double fertilization during which the two pollen sperm cells fuse with the egg cell and the central cell of the female gametophyte. In Arabidopsis, analyses of mutants in the cell-cycle regulator CYCLIN DEPENDENT KINASE A;1 (CKDA;1) have revealed the importance of a paternal genome for the effective development of the endosperm and ultimately the seed. Here we have exploited cdka;1 fertilization as a novel tool for the identification of seed regulators and factors involved in parent-of-origin-specific regulation during seed development. We have generated genome-wide transcription profiles of cdka;1 fertilized seeds and identified approximately 600 genes that are downregulated in the absence of a paternal genome. Among those, AGAMOUS-LIKE (AGL) genes encoding Type-I MADS-box transcription factors were significantly overrepresented. Here, AGL36 was chosen for an in-depth study and shown to be imprinted. We demonstrate that AGL36 parent-of-origin-dependent expression is controlled by the activity of METHYLTRANSFERASE1 (MET1) maintenance DNA methyltransferase and DEMETER (DME) DNA glycosylase. Interestingly, our data also show that the active maternal allele of AGL36 is regulated throughout endosperm development by components of the FIS Polycomb Repressive Complex 2 (PRC2), revealing a new type of dual epigenetic regulation in seeds.

Polycomb repressive complex 2 controls the embryo-to-seedling phase transition.

Polycomb repressive complex 2 (PRC2) is a key regulator of epigenetic states catalyzing histone H3 lysine 27 trimethylation (H3K27me3), a repressive chromatin mark. PRC2 composition is conserved from humans to plants, but the function of PRC2 during the early stage of plant life is unclear beyond the fact that it is required for the development of endosperm, a nutritive tissue that supports embryo growth. Circumventing the requirement of PRC2 in endosperm allowed us to generate viable homozygous null mutants for FERTILIZATION INDEPENDENT ENDOSPERM (FIE), which is the single Arabidopsis homolog of Extra Sex Combs, an indispensable component of Drosophila and mammalian PRC2. Here we show that H3K27me3 deposition is abolished genome-wide in fie mutants demonstrating the essential function of PRC2 in placing this mark in plants as in animals. In contrast to animals, we find that PRC2 function is not required for initial body plan formation in Arabidopsis. Rather, our results show that fie mutant seeds exhibit enhanced dormancy and germination defects, indicating a deficiency in terminating the embryonic phase. After germination, fie mutant seedlings switch to generative development that is not sustained, giving rise to neoplastic, callus-like structures. Further genome-wide studies showed that only a fraction of PRC2 targets are transcriptionally activated in fie seedlings and that this activation is accompanied in only a few cases with deposition of H3K4me3, a mark associated with gene activity and considered to act antagonistically to H3K27me3. Up-regulated PRC2 target genes were found to act at different hierarchical levels from transcriptional master regulators to a wide range of downstream targets. Collectively, our findings demonstrate that PRC2-mediated regulation represents a robust system controlling developmental phase transitions, not only from vegetative phase to flowering but also especially from embryonic phase to the seedling stage.

Low liver stiffness among cirrhotic patients with hepatitis B after prolonged treatment with nucleoside analogs.

OBJECTIVE: Case reports and short-term clinical trials have suggested that treatment for chronic hepatitis B (CHB) may lead to improvement of cirrhosis. The aim of the present study was to measure liver stiffness in patients diagnosed with advanced fibrosis or cirrhosis prior to prolonged treatment with nucleoside or nucleotide analogs (NUCs) for CHB. MATERIALS AND METHODS: Patients with CHB and advanced fibrosis or cirrhosis prior to treatment with NUCs for at least 1 year were offered inclusion in the study. We measured liver stiffness using transient elastography (TE) at follow-up. TE cut-off levels to Metavir classification for fibrosis stage F2, F3 and F4 were ≥7.2 kPa, ≥8.1, and ≥11.0 kPa, respectively. RESULTS: Among 66 patients with a successful TE examination at follow-up, 53 patients (80%) had cirrhosis and 13 had (20%) advanced fibrosis (F3) prior to treatment. Median treatment duration was 50.5 months. Among patients with cirrhosis prior to treatment, 26 (49%) had liver stiffness below 11.0 kPa at follow-up, suggesting regression of cirrhosis. Among patients with advanced fibrosis (F3) prior to treatment, 10 (77%) had liver stiffness below 8.1 kPa after treatment, suggesting improvement of fibrosis. CONCLUSION: Transient elastography examinations demonstrate that prolonged treatment with NUCs in patients with CHB results in low liver stiffness, suggesting regression of fibrosis in a majority of patients with advanced fibrosis or cirrhosis.

Prevalence of hepatitis B and C at a major tuberculosis centre in Denmark.

BACKGROUND: Denmark is a low-prevalence country for tuberculosis (TB), hepatitis B (HBV), and hepatitis C (HCV) but the three diseases have similar sociodemographic risk factors. We estimated the prevalence and possible risk factors of HBV and HCV among TB-patients in a large TB clinic in Denmark. METHODS: All patients starting anti-TB-treatment at Copenhagen University Hospital, Herlev-Gentofte from April 1st 2018 through June 1st 2019 were included. Results from HBV and HCV testing as well as sociodemographic information were collected. Risk factor analyses were carried out using descriptive statistics. RESULTS: Of 82 patients tested for HBV, one (1.2%) had chronic HBV and 16 (19.5%) had serocleared HBV. Of 91 patients tested for HCV, three (3.3%) had chronic HCV and one (1.1%) had serocleared HCV. Country of origin other than Denmark was significantly associated with HBV-seropositivity among TB-patients, particularly patients from Greenland, Asia, Africa and Eastern Europe. No other significantly associated risk factors were found. CONCLUSION: The prevalences of chronic and prior HBV and HCV among TB-patients were lower compared to studies in TB high endemic areas but higher than those found in the Danish background population. We calculated the number needed to test to find one patient with HBV ranged between 27 and 83 and we suggest continuing screening of chronic HBV and HCV in TB-patients in Denmark.

Evaluation of four non-invasive methods for examination and characterization of pressure ulcers.

BACKGROUND: Pressure ulcers are globally of major concern and there is need for research in the pathogenesis for early intervention. Early studies have suggested existence of a hypo-echogenic subepidermal layer at the location of pressure ulcers, visualized by ultrasound scans. As a continuation, we here report on usability of four non-invasive techniques for evaluation of pressure ulcers. METHODS: Fifteen pressure ulcers in stage 0-IV were examined using four different non-invasive techniques [redness index, skin temperature, skin elasticity (i.e. retraction time), and ultrasound scanning]. Measurements were made at the ulcer, 5 cm from the ulcer, and at a reference skin location without ulcers. RESULTS: The redness index was, in all cases, higher at the ulcers than at the reference skin. Temperature measurements were rather scattered. Ultrasound scans showed a hypo-echogenic subepidermal layer at all pressure ulcers, but none at the reference points. The skin retraction time was often higher at the location of a pressure ulcer than at the reference location. We found no correlation between the stage of the ulcers and temperature, redness index, subepidermal layer thickness, or retraction time. CONCLUSION: We conclude that temperature and elasticity measurements do not alone characterize ulceration severity, although redness index in some cases provides a useful indication. We assume that a subepidermal layer found on ultrasound images may be a measure of the pressure that the skin has been subjected to, rather than of the severity of the pressure ulceration. This method may be useful for predicting whether the skin is at risk of developing pressure ulcers. More studies are needed.

Outcome and adverse events in patients with chronic hepatitis C treated with direct-acting antivirals: a clinical randomized study.

OBJECTIVE: New potent direct-acting antiviral (DAA) regimens against hepatitis C virus have been approved in recent years. However, information about the rate of adverse events (AEs) across different DAA regimens is limited. We aimed to evaluate differences in AEs and treatment efficacy in patients with chronic hepatitis C (CHC), genotype (GT) 1 or 3, randomized to two different treatment arms, correspondingly. PATIENTS AND METHODS: We randomly assigned 96 patients in a 1 : 1 ratio, to treatment for 12 weeks with either paritaprevir/ombitasvir/ritonavir/dasabuvir/ribavirin (RBV) or ledipasvir/sofosbuvir (SOF)/RBV if infected with GT1 (72 patients) or to daclatasvir/SOF/RBV for 12 weeks or SOF/RBV for 24 weeks, if infected with GT3 (24 patients). Data on AEs were collected throughout the entire study period. RESULTS: A total of 70 (97%) patients with CHC with GT1 and 20 (83%) patients with GT3 achieved cure. The GT3 treatment arm was prematurely terminated, owing to change in national treatment guidelines. Thus, only AEs for GT1 patients are described. AEs occurred in 70 (97%) GT1 patients, and most common AEs were anemia (n=56/78%), fatigue (n=53/74%), and headache (n=33/46%). No difference was observed in relation to treatment group (P=1.0), anemia (P=1.0), or liver cirrhosis (P=0.53). In seven (11%) patients, AEs assessed by the investigator to be possibly related to the DAA regimen were still present 12 weeks after treatment. CONCLUSIONS: We found no difference in AEs possibly related to the DAA regimen in patients with CHC, but surprisingly, AEs possibly related to the DAA regimen persisted in a significant number of patients after treatment. This finding can be of importance for clinicians in relation to patient information concerning AEs possibly related to DAA treatment.

Comparison of meglumine antimoniate and pentamidine for peruvian cutaneous leishmaniasis.

Pentamidine was compared with meglumine antimoniate (Glucantime) for 80 patients with cutaneous leishmaniasis due to Leishmania braziliensis in Peru. Of the 40 patients administered Glucantime (20 mg of antimony [Sb]/kg/day intravenously for 20 days), 31 cured (78%), 6 failed (15%), of which 5 were due to relapse, and 3 were lost to follow-up (7%). Of the 40 patients administered pentamidine (2 mg/kg every other day for seven injections), 14 were cured (35%), 23 failed (58%), and 3 were lost to follow-up (7%). Five pentamidine failures were due to relapse, and 14 failures were due to the presence of parasites two weeks after therapy. Both regimens were well tolerated. Gastrointestinal, musculoskeletal, and total adverse events were not statistically different in either group. Elevations in levels of liver enzymes and pancreatic enzymes were statistically higher in the Glucantime group, but no patient terminated therapy prematurely. In this study, Glucantime was more effective than pentamidine for treatment of L. braziliensis cutaneous leishmaniasis in Peru based on parasitologic as well as clinical criteria.

Chloroquine-resistant Plasmodium vivax malaria in Peru.

Reports from several sites in South America suggest the presence of isolated cases of chloroquine-resistant Plasmodium vivax malaria. To investigate the possibility of chloroquine-resistant P. vivax in Peru, we conducted 28-day in vivo drug efficacy trials at three sites in the Amazon region and one site on the northern Pacific Coast between 1998 and 2001. A total of 242 patients between the ages of 2 and 60 years were enrolled (177 from the Amazon region and 65 from the northern coast). All subjects received directly observed therapy with chloroquine, 25 mg/kg, over a three-day period. On enrollment, 49% had a documented fever and 96% had a history of fever; their geometric mean parasite density was 5,129 parasites/microL. A total of 212 (88%) of the 242 subjects completed their 28-day follow-up. Four of the 177 patients from the Amazon region had a recurrence of P. vivax parasitemia on days 21 and 28 after treatment was initiated. Two of these patients had chloroquine-resistant infections, based on polymerase chain reaction-single-stranded conformational polymorphism genotyping and chloroquine-desethylchloroquine blood levels, which were > or = 97 ng/mL at the time of the reappearance of parasitemia. None of the subjects studied on the northern Pacific Coast had recurrent parasitemia.

Anemia in parasite- and recombinant protein-immunized aotus monkeys infected with Plasmodium falciparum.

Plasmodium falciparum-induced anemia was characterized in Aotus monkeys repeatedly immunized by infection with P. falciparum (FVO strain) parasites, then cross-challenged with CAMP strain, or in monkeys receiving blood stage challenges as part of malaria vaccine trials. In 4 studies, 25 (30.5%) of 82 monkeys had at least a 50% reduction in hematocrit; mean day of maximum parasitemia was 12.5, whereas the mean day of minimum hematocrit was 18.8 (P < 0.0009). Decreased hematocrit levels were not associated with reticulocytosis until parasite densities decreased significantly from peak levels. Direct antibody tests to detect IgG and C3d on the surface of erythrocytes were negative. Nonantibody/noncomplement-mediated lysis of uninfected erythrocytes seems to be the principal cause of the anemia, and it also seems that bone marrow suppression and lysis of infected erythrocytes contributed to the anemia. Partial immunity-whether induced by repeated immunization with whole parasites or with vaccine-seems important to the development of anemia.

IP-10 can be measured in dried plasma spots in patients with chronic hepatitis C infection.

The chemokine IP-10 (CXCL10) is a candidate marker for hepatitis C virus (HCV) fibrosis monitoring. The aim of this proof-of-concept study is to assess if IP-10 measurements from dried plasma spots (DPS) are accurate in HCV-infected patients with either minimal or significant fibrosis. We measured IP-10 levels in plasma and DPS of 21 HCV-infected patients with cirrhosis and 19 patients with no/little fibrosis (determined with FibroScan). Cirrhotic patients had significantly higher levels of IP-10 compared to patients with minimal fibrosis. DPS and plasma measurements of IP-10 are comparable and the correlation was excellent (r(2) = 0.97, p<0.0001). The DPS based method for IP-10 detection performs well in HCV-infected patients with either minimal or significant fibrosis.

Lower liver stiffness in patients with sustained virological response 4 years after treatment for chronic hepatitis C.

OBJECTIVE: Transient elastography (TE) is a noninvasive and well validated method for measurement of liver stiffness. The aim of this study was to use TE to evaluate whether patients with sustained virological response (SVR) have lower liver stiffness than patients with non-SVR after treatment for chronic hepatitis C (CHC). METHODS: Patients with CHC, who had undergone liver biopsy before treatment with pegylated interferon and ribavirin, were included from four clinical centres in Denmark. All patients were examined with TE and had a blood test taken for hepatitis C virus-virus detection and analysis of alanine aminotransferase, platelet counts and hyaluronic acid. RESULTS: For 110 (92%) of the 120 patients included, it was possible to obtain a successful measurement of liver stiffness. Of these, 71 (64.5%) had achieved SVR. Median follow-up time was 47 months. Patients with pretreatment minimal fibrosis (F0/F1) in their liver biopsy had median liver stiffness of 5.3 kPa for SVR versus 6.1 kPa for non-SVR (P=0.56). Patients with pretreatment moderate fibrosis (F2/F3) had median liver stiffness of 5.4 kPa for SVR versus 9.4 kPa for non-SVR (P<0.001). Median liver stiffness for patients with pretreatment cirrhosis (F4) was 6.8 kPa for SVR versus 24 kPa for non-SVR (P<0.001). CONCLUSIONS: Examination with TE 4 years after treatment shows that patients with CHC, who have achieved SVR, have significantly lower liver stiffness than patients with non-SVR. This indicates that histological liver outcome improves during the first year after the treatment for CHC.

Reactivation of hepatitis D virus after chemotherapy for diffuse large B cell lymphoma despite lamivudine prophylaxis.

We describe a case of reactivation of hepatitis D virus (HDV) in a patient treated with chemotherapy for a diffuse large B cell lymphoma despite lamivudine prophylaxis. This case suggests that previously cleared HDV should be considered when administering chemotherapy to patients with lymphoma.

Transient elastography for liver fibrosis diagnosis.

Liver biopsy is considered the "golden standard" for assessment of hepatic fibrosis. However, the procedure has limitations because of inconvenience and rare but serious complications as bleeding. Furthermore, sampling errors are frequent, and interobserver variability often poses problems. Recently, a modified ultrasound scanner (transient elastography) has been developed to assess fibrosis. The device measures liver elasticity, which correlates well with the degree of fibrosis. Studies have shown that transient elastography is more accurate in diagnosing cirrhosis than minor to moderate fibrosis. Most of the studies have been conducted on patients with chronic hepatitis but a few studies have also covered fibrosis and cirrhosis due to other etiologies, and they also demonstrate the high sensitivity and specificity. Transient elastography for assessment of fibrosis may turn out to be a valuable diagnostic procedure and follow-up of patients with chronic liver diseases.

[New treatment of chronic hepatitis B]. / Ny behandling af kronisk hepatitis B.

Worldwide, 350 million people are infected with chronic hepatitis B. Over the last few years, it has been possible to treat chronic hepatitis B. Treatment very often consists of nucleos(t)ide analogs and in a few cases of pegylated alpha-interferon. In 2007, a new nucleoside analog, Telbivudine, was approved to treat chronic hepatitis B. In phase II and ongoing phase III studies, Telbivudine has proven more effective than the nucleoside analog, Lamivudine, which was very often used up until recently.

[Uveitis--a manifestation of syphilis]. / Uveitis--manifestation af syfilis.

We describe two cases of neurosyphilis whose diagnosis was based on ocular symptoms. Both patients were treated in the Eye Department for bilateral uveitis, and they were tested serologically positive for syphilis. The diagnoses of neurosyphilis were confirmed by demonstration of CSF pleocytosis and specific intrathecal antibody production. Both patients were treated with 21 million units of penicillin IV daily for 15 days. Ocular syphilis is an unusual manifestation of the disease but should be considered in patients with uveitis of uncertain origin, especially if the patient has a rash and/or headache.

Devices for rapid diagnosis of Malaria: evaluation of prototype assays that detect Plasmodium falciparum histidine-rich protein 2 and a Plasmodium vivax-specific antigen.

The ParaSight F test was developed as a pioneer industry effort in the large-scale, process-controlled production of a device for the rapid diagnosis of malaria. This device performed well in field settings but was limited to the detection of a single malaria species, Plasmodium falciparum. The ParaSight F+V assay advanced upon the ParaSight F test format by incorporating a monoclonal antibody directed against a proprietary Plasmodium vivax-specific antigen, in addition to the antibody directed against P. falciparum histidine-rich protein 2, which was used in the ParaSight F assay. The modified assay was developed to add the capability to detect P. falciparum and P. vivax in a single-test-strip format. The present study evaluated three distinct ParaSight F+V prototypes with samples from symptomatic patients in regions of Thailand and Peru where malaria is endemic. Over a 2-year enrollment period (1998 and 1999), a total of 4,894 patients consented to participation in the study. Compared with the results for duplicate microscopic examinations of Giemsa-stained blood smears as the reference diagnostic standard, each successive prototype showed substantial improvement in performance. The final ParaSight F+V prototype, evaluated in 1999, had an overall sensitivity for detection of asexual P. falciparum parasites of 98%. The sensitivity of the device was 100% for P. falciparum densities of >500 parasites/ micro l, with a sensitivity of 83% for parasite densities of 5,000/ micro l, 92% for parasite densities of 1,001 to 5,000/ micro l, 94% for parasite densities of 501 to 1,000/ micro l, and 55% for parasite densities of 1 to 500/ micro l. The specificity for the exclusion of P. vivax was 87%. The areas under the receiver operating characteristic curves for the diagnostic performance of the assay for the detection of P. falciparum and P. vivax were 0.8907 and 0.8522, respectively. These findings indicate that assays for rapid diagnosis have the potential to enhance diagnostic capabilities in those instances in which skilled microscopy is not readily available.