A scoping review on tools and methods for trait prioritization in crop breeding programmes.

Trait prioritization studies have guided research, development and investment decisions for public-sector crop breeding programmes since the 1970s, but the research design, methods and tools underpinning these studies are not well understood. We used PRISMA-ScR (Preferred Reporting Items for Systematic review and Meta-Analysis Protocols) to evaluate research on trait ranking for major crops over the past 40 years (1980-2023). Data extraction and descriptive analysis on 657 papers show uneven attention to crops, lack of systematic sex disaggregation and regional bias. The lack of standardized trait data taxonomy across studies, and inconsistent research design and data collection practices make cross-comparison of findings impossible. In addition, network mapping of authors and donors shows patterns of concentration and the presence of silos within research areas. This study contributes to the next generation of innovation in trait preference studies to produce more inclusive, demand-driven varietal design that moves beyond trait prioritization focused on productivity and yield.

p21(WAF1) gene promoter is epigenetically silenced by CTIP2 and SUV39H1.

Mainly regulated at the transcriptional level, the cellular cyclin-dependent kinase inhibitor, CDKN1A/p21(WAF1) (p21), is a major cell cycle regulator of the response to DNA damage, senescence and tumor suppression. Here, we report that COUP-TF-interacting protein 2 (CTIP2), recruited to the p21 gene promoter, silenced p21 gene transcription through interactions with histone deacetylases and methyltransferases. Importantly, treatment with the specific SUV39H1 inhibitor, chaetocin, repressed histone H3 lysine 9 trimethylation at the p21 gene promoter, stimulated p21 gene expression and induced cell cycle arrest. In addition, CTIP2 and SUV39H1 were recruited to the silenced p21 gene promoter to cooperatively inhibit p21 gene transcription. Induction of p21(WAF1) gene upon human immunodeficiency virus 1 (HIV-1) infection benefits viral expression in macrophages. Here, we report that CTIP2 further abolishes Vpr-mediated stimulation of p21, thereby indirectly contributing to HIV-1 latency. Altogether, our results suggest that CTIP2 is a constitutive p21 gene suppressor that cooperates with SUV39H1 and histone methylation to silence the p21 gene transcription.

Molecular mimicry in inducing DNA damage between HIV-1 Vpr and the anticancer agent, cisplatin.

The human immunodeficiency virus type 1 (HIV-1) viral protein R (vpr) gene is an evolutionarily conserved gene among the primate lentiviruses. Several functions are attributed to Vpr including the ability to cause cell death, cell cycle arrest, apoptosis and DNA damage. The Vpr domain responsible for DNA damage as well as the mechanism(s) through which Vpr induces this damage is unknown. Using site-directed mutagenesis, we identified the helical domain II within Vpr (aa 37-50) as the region responsible for causing DNA damage. Interestingly, Vpr Delta(37-50) failed to cause cell cycle arrest or apoptosis, to induce Ku70 or Ku80 and to suppress tumor growth, but maintained its capability to activate the HIV-1 LTR, to localize to the nucleus and to promote nonhomologous end-joining. In addition, our cytogenetic data indicated that helical domain II induced chromosomal aberrations, which mimicked those induced by cisplatin, an anticancer agent. This novel molecular mimicry function of Vpr might lead to its potential therapeutic use as a tumor suppressor.

Production of cytotoxic factor by peripheral blood mononuclear cells (PBMC) in patients with dengue haemorrhagic fever.

A unique cytokine, human cytotoxic factor (hCF), has been shown to occur in the sera of patients with dengue fever (DF) and dengue haemorrhagic fever (DHF). The present study was undertaken to investigate the ability of fresh PBMC of such patients to produce hCF. The PBMC were cultured for 24 h and the culture supernatants (CS) were analysed for the presence of hCF by cytotoxicity assay, competitive ELISA and dot blot tests. In 90% of 246 cases CS were positive for hCF by the three tests. CS were positive for hCF in PBMC collected from days 1-20 of illness but not at later periods. Higher cytotoxic activity was observed in CS of days 1-4 of illness and was highest in cases of DHF grade IV and lowest in cases of DF. Dot blot hybridization of RNA extracted from the PBMC of the patients showed the presence of mRNA for hCF in 94% of cases. A similar number of patients showed the presence of hCF in situ in the PBMC smears by fluorescent antibody technique. hCF was found only in CD4+ T cells. The findings thus present direct evidence of the production of hCF by CD4 T cells of cases of DF/DHF.

ELISA for detection of dengue virus-induced cytokine and its antibody.

ELISA technique has been standardized for detection of a dengue type 2 virus (DV)-induced cytokine, the cytotoxic factor (CF) and CF-specific antibodies. The performed ELISA was found to be sensitive (90.9%), specific (92.5%), accurate (91%) and reproducible and was able to detect a minimum of 7 ng/ml CF in the test samples. The technique was used to detect CF in DV inoculated mouse sera and DV-infected mouse spleen cell culture fluids. Significant utility of the test was detection of a CF-like cytokine in the sera of human cases of dengue haemorrhagic fever.

Release of reactive oxygen intermediates by dengue virus-induced macrophage cytotoxin.

Dengue type 2 virus (DV) induces a subpopulation of T lymphocytes of mice to produce a cytokine, cytotoxic factor (mCF), which induces H-2A positive macrophages to produce macrophage cytotoxin (CF2). The present study was undertaken to investigate the mechanism of cytotoxicity of CF2. It was observed that CF2 induced production of superoxide anion (O2-) and hydrogen peroxide (H2O2) by the spleen cells of mice in vitro and in vivo. The maximum production of O2- (260 +/- 10 nM/4 x 10(6) cells) was at 45 minutes while that of H2O2 was at 90 minutes after inoculation of CF2. Pretreatment of mice or spleen cells with anti-CF2-antisera inhibited O2- and H2O2 production in a dose-dependent manner. Superoxide dismutase (SOD) inhibited O2- production and cytotoxicity while H2O2 production was increased by increasing SOD concentration in the culture. This indicated that O2- production is necessary for the cytotoxic activity of CF2. Pretreatment of the cells with Ca2+ channel blocking drugs, nifedipine or verapamil, inhibited CF2-induced O2- and H2O2 production in a dose-dependent manner. We have shown earlier that the cytotoxic activity of CF2 is known to be Ca2+ dependent and CF2-induced production of nitrite and the cytotoxicity is inhibited by NG-monomethyl-L-arginine. Thus, it is suggested that O2- and nitrite are necessary for cell killing by CF2 in a Ca(2+)-dependent manner and the killing may possibly be by generation of peroxynitrite.

Production of nitrite by dengue virus-induced cytotoxic factor.

Dengue type 2 virus (DV) infection induces production of a cytokine, the cytotoxic factor (CF) in the spleen of mice. The present study was undertaken to investigate the production of nitrite (NO2-) by the spleen cells of mice in vitro and in vivo following inoculation of DV or CF. Maximum NO2- production occurred at 45 min after inoculation of 5 microg CF, both in vitro and in vivo. The NO2- was produced by macrophages and T cells and not by B cells. Pretreatment of CF with anti-CF antisera inhibited production of NO2-. DV-stimulated spleen cell culture supernatants showed peak production of CF and NO2- at 72h. In DV-infected mouse spleen, maximum NO2- production occurred at 8-11 days post-infection, which correlated with peak cytotoxic activity in the spleen. Pretreatment of spleen cells with N(G)-monomethyl L-arginine (NMMA) inhibited NO2- production. NO2- production was abrogated in a dose-dependent manner by treatment of spleen cells with Ca2+ channel blocking drug, Nifedipine. The findings demonstrate that DV-induced CF induces production of NO2- in spleen cells, probably in a Ca2+-dependent manner, and may be a mechanism of target cell killing.

Dengue virus-induced cytotoxin releases nitrite by spleen cells.

Dengue type-2 virus (DV) infection in mice induces T cells to produce a cytokine, the cytotoxic factor (CF), which induces H2-A positive macrophages to produce another cytokine, cyototoxic (CF2), which amplifies its cytotoxic effects on target cells. The present study was undertaken to investigate the production of nitrite (NO2-) by the spleen cells of mice in vitro and in vivo following inoculation of CF2. Maximum NO2- production occurred at 1 hour after inoculation of 100 micrograms CF2. Pretreatment of CF2 with anti-CF2-antisera (CF2-As) inhibited the production of NO2-. Pretreatment of the spleen cells with NG-monomethyl-L-arginine (NMA) or with arginase inhibited NO2- production. The NO2- production was diminished in a dose dependent manner by treatment of spleen cells with the Ca2+ channel blocking drug, nifedipine and Zn2+ as ZnSO4. The findings of the present study thus demonstrate that CF2 induces production of NO2- in the spleen cells in a CA(2+)-dependent manner which may be a mechanism of target cell killing.

Effect of adjuvants on immunization with dengue virus-induced cytotoxic factor.

Specific active immunization with dengue type 2 virus (DV)-induced cytokine, cytotoxic factor (CF), prevents CF-mediated pathology in mice. The present study was undertaken to determine the optimum dose of CF and the effect of different adjuvants on the immune response as assessed by the study of anti-CF antibody titre by ELISA and protection against increase in capillary permeability to challenging dose of 3 micrograms CF. The maximum protection of 94 +/- 4% against increase in capillary permeability was observed at week 4 after immunization with 5 micrograms dose of CF mixed with Freund's incomplete adjuvant (FIA), which gradually decreased to 21 +/- 10% on week 24. With a dose of 10 micrograms the protection obtained was 79 +/- 5%, but persisted for a longer time at a higher level. The response was poor with 1 microgram dose of CF. The mean anti-CF antibody titres gradually decreased after reaching the peak at week 4 after immunization. Mice immunized with different adjuvants emulsified with 5 micrograms CF were challenged at different intervals with 3 micrograms CF. Maximum protection observed with CF + tetanus toxoid (TT) and 84/246 was about 93 +/- 2% and 97 +/- 2%, while that with alhydrogel was 33 +/- 12% and with bacille Calmette-Guérin (BCG) was 67 +/- 4%. At week 24 after immunization, however, the best response was obtained with 10 micrograms of adjuvant 84/246. Intracerebral challenge with 10 or 100 LD50 dose of dengue type 2 virus showed significantly prolonged mean survival time and delayed onset of signs of sickness in immunized mice compared with normal mice. The maximum survival time was with adjuvant 84/246 even at week 24. The findings thus show that the optimum dose of CF is 5 micrograms and the adjuvant of choice is 84/246.

Dengue virus-induced human cytotoxic factor: production by peripheral blood leucocytes in vitro.

During dengue type 2 virus (DV) infection of mice a unique cytokine, the cytotoxic factor (CF), is produced which reproduces the pathological lesions seen in patients of dengue haemorrhagic fever (DHF). Recently we have observed a CF-like protein in the sera of DHF cases. The present study was undertaken to investigate whether DV can stimulate human peripheral blood mononuclear cells (PBMC) in vitro to produce human CF (hCF). Cultures prepared from PBMC or its enriched subpopulations were inoculated with 1000 LD50 of DV and controls with normal mouse brain suspension (NMB). Aliquots of cultures were harvested daily from 24 h to 96 h and their supernatant (CS) and cells were separated. CS were screened for viral titres and for the presence of hCF by cytotoxicity assay, inhibition ELISA, dot blot and Western blot tests using anti-mouse-CF antibodies. The RNA from the cells was screened in Northern blot and dot blot tests for the presence of mRNA for CF. It was observed that hCF appeared in the CS of DV-infected culture of PBMC and T-enriched cells at 48 h and was present until 96 h; no CF was detected in CS of B cells or monocyte cultures. The production of hCF was abrogated by pretreatment of the T cells with anti-CD4 antibodies but not with anti-CD8 antibodies, indicating that hCF was produced by CD4+ T cells. The Northern blot analysis using oligonucleotide probes prepared on the basis of amino-terminal sequence of mouse CF, showed presence of mRNA for hCF in PBMC and T cell cultures. DV replicated in PBMC cultures with peak titres at 48 h. The findings of the present study demonstrate that DV-induced hCF is produced by human T cells.

Flow cytometric analysis of T4:T8 splenic cell during dengue virus infection of mice.

The present study was undertaken to investigate the effect of dengue type 2 virus (DV) and DV-induced cytokines (CF and CF2) on T lymphocyte subpopulations of spleen by flow cytometry. Following DV-ic inoculation in mice the percent number of CD4+ and CD8+ lymphocytes in the spleen was reduced, the peak reduction in both was observed on the 6th day post-inoculation (p.i.). Intravenous inoculation of CF or CF2 in mice also decreased the percent number of CD4+ as well as CD8+ T lymphocytes subpopulation in the spleen, the maximum reduction being observed at 1 and 2 hr, respectively. The reduction in T lymphocyte subpopulation by CF and CF2 was found to be dose dependent. Thus, the alterations of T lymphocyte subpopulations during DV infection are mediated via cytokines.

Isolation of the novel agent from human stool samples that is associated with sporadic non-A, non-B hepatitis.

The agent(s) responsible for sporadic non-A, non-B hepatitis in humans was serially transmitted in rhesus monkeys by intravenous inoculation of the stool extract from a patient. A novel agent called HFV (hepatitis French [origin] virus) was present as 27- to 37-nm particles in the infectious stool extract. Hepatopathic lesions were noticed in infected monkeys during the acute phase of illness. The purified viral 27- to 37-nm particles consist of a double-stranded DNA of approximately 20 kb and are detected in infected monkey liver. Analysis of cell culture detects the approximately 20-kb-long viral DNA in stool samples from infected monkeys and sporadic enteric non-A, non-B hepatitis patients. Furthermore, the 27- to 37-nm viral particles were able to protect monkeys challenged with infectious stool extract. Our results indicate that 27- to 37-nm virus like particles are responsible for sporadic non-A, non-B hepatitis in rhesus monkeys.

Active immunization by a dengue virus-induced cytokine.

Dengue type 2 virus (DV)-induced cytotoxic factor (CF) is capable of reproducing various pathological lesions in mice that are seen in human dengue. The present study was undertaken to investigate the protective effect of active immunization of mice with CF. Mice were immunized with 5 microgram of CF and prevention of CF-induced increase in capillary permeability and damage to the blood-brain barrier were studied at weekly intervals, up to 48 weeks, by challenging with 3 microgram of CF. Maximum protection against increase in capillary permeability and damage to the blood-brain barrier was observed in week 4 after immunization. A breakthrough in the protection occurred with higher doses of CF in a dose-dependent manner. Challenge with a lethal intracerebral (i.c.) dose of DV showed significantly prolonged mean survival time and delayed onset of symptoms of sickness in the immunized mice compared with the normal mice, but the titre of the virus in the brain was similar in the two groups. On i.p. challenge with the virus the protection against damage to the blood-brain barrier was 86 +/- 7% at week 4 and 17 +/- 4% at week 26 after immunization. Sera obtained from the immunized mice showed the presence of CF-specific antibodies by ELISA, Western blot, and by neutralization of the cytotoxic activity of CF in vitro. The present study describes successful prevention of a cytokine-induced pathology by specific active immunization.