Recombination in the B Chromosome of Maize to Produce a-B-a Chromosomes

The translocations between the supernumerary B chromosomes and the normal A chromosomes of maize provide a valuable tool for gene localizations, dosage studies and characterization of mutants as null, leaky or gain-of-function. A procedure is described, that relies on recombination in the B chromosome, for marking each of the various B-A translocations with a single dominant marker that will allow dosage classifications of individuals at the mature kernel stage. This marker is R-scm3, which conditions anthocyanin pigment in the aleurone of the endosperm and the scutellum of the embryo. A test for recombination in the B chromosome was conducted by crossing together two translocations, that were broken on opposite sides of the B centromere, and in different A chromosome arms, namely TB-1La and TB-10L18. An example was recovered that linked genetic markers on 1L and 10L to the B centromere. Cytological examination at pachytene of meiosis confirmed the new chromosomal linkage. The use of this procedure to produce a comprehensive set of uniformly marked B-A translocations is discussed.     

Hepatitis B Infection,Viral Load and Resistance in HIV-Infected Patients in Mozambique and Zambia

Background

Few data on the virological determinants of hepatitis B virus (HBV) infection are available from southern Africa.

Methods

We enrolled consecutive HIV-infected adult patients initiating antiretroviral therapy (ART) at two urban clinics in Zambia and four rural clinics in Northern Mozambique between May 2013 and August 2014. HBsAg screening was performed using the Determine^® rapid test. Quantitative real-time PCR and HBV sequencing were performed in HBsAg-positive patients. Risk factors for HBV infection were evaluated using Chi-square and Mann-Whitney tests and associations between baseline characteristics and high level HBV replication explored in multivariable logistic regression.

Results

Seventy-eight of 1,032 participants in Mozambique (7.6%, 95% confidence interval [CI]: 6.1–9.3) and 90 of 797 in Zambia (11.3%, 95% CI: 9.3–13.4) were HBsAg-positive. HBsAg-positive individuals were less likely to be female compared to HBsAg-negative ones (52.3% vs. 66.1%, p<0.001). Among 156 (92.9%) HBsAg-positive patients with an available measurement, median HBV viral load was 13,645 IU/mL (interquartile range: 192–8,617,488 IU/mL) and 77 (49.4%) had high values (>20,000 UI/mL). HBsAg-positive individuals had higher levels of ALT and AST compared to HBsAg-negative ones (both p<0.001). In multivariable analyses, male sex (adjusted odds ratio: 2.59, 95% CI: 1.22–5.53) and CD4 cell count below 200/μl (2.58, 1.20–5.54) were associated with high HBV DNA. HBV genotypes A1 (58.8%) and E (38.2%) were most prevalent. Four patients had probable resistance to lamivudine and/or entecavir.

Conclusion

One half of HBsAg-positive patients demonstrated high HBV viremia, supporting the early initiation of tenofovir-containing ART in HIV/HBV-coinfected adults.     

The Size of the Viral Inoculum Contributes to the Outcome of Hepatitis B Virus Infection

The impact of virus dose on the outcome of infection is poorly understood. In this study we show that, for hepatitis B virus (HBV), the size of the inoculum contributes to the kinetics of viral spread and immunological priming, which then determine the outcome of infection. Adult chimpanzees were infected with a serially diluted monoclonal HBV inoculum. Unexpectedly, despite vastly different viral kinetics, both high-dose inocula (10¹⁰ genome equivalents [GE] per animal) and low-dose inocula (10° GE per animal) primed the CD4 T-cell response after logarithmic spread was detectable, allowing infection of 100% of hepatocytes and requiring prolonged immunopathology before clearance occurred. In contrast, intermediate (10⁷ and 10⁴ GE) inocula primed the T-cell response before detectable logarithmic spread and were abruptly terminated with minimal immunopathology before 0.1% of hepatocytes were infected. Surprisingly, a dosage of 10¹ GE primed the T-cell response after all hepatocytes were infected and caused either prolonged or persistent infection with severe immunopathology. Finally, CD4 T-cell depletion before inoculation of a normally rapidly controlled inoculum precluded T-cell priming and caused persistent infection with minimal immunopathology. These results suggest that the relationship between the kinetics of viral spread and CD4 T-cell priming determines the outcome of HBV infection.The hepatitis B virus (HBV) is a noncytopathic DNA virus that causes acute and chronic hepatitis and hepatocellular carcinoma (5). Viral clearance and disease pathogenesis during acute HBV infection require the induction of a vigorous CD8⁺ T-cell response and the induction of significant hepatic immunopathology (12, 28). In contrast, chronic HBV infection is associated with a markedly diminished CD8⁺ T-cell response to HBV (23, 24) for reasons that are not well defined.We have previously studied the immunobiology and pathogenesis of HBV infection in chimpanzees that we inoculated with a single (10⁸ genome equivalents [GE]) dose of a monoclonal inoculum of HBV (12, 28, 33). In all of these animals, the infection pursued a reproducible, almost stereotypical course irrespective of the age, size, sex, and genetics of the animals, and it spread to 100% of the hepatocytes before it was terminated by the CD8 T-cell response. The reproducibility of these results suggested that the course and outcome of infection were dominated by the impact of the virus on the kinetics and magnitude of the infection and on the kinetics and magnitude of the immune response that it elicited.Because a high viral load has a negative impact on the outcome of other virus infections (reviewed in references 19 and 32), we examined in the present study the impact of the size of the viral inoculum on the outcome of HBV infection in HBV-naive, immunocompetent adult chimpanzees using a wide dose range of the same monoclonal inoculum that we used in our earlier studies.In contrast to the highly reproducible outcome to the 10⁸ GE dose in our previous experiments, we observed a wide range of outcomes to the various dosages used here, including the development of chronic HBV infection, that we could relate to the kinetics of the CD4 T-cell response in each animal. Furthermore, depletion of CD4⁺ cells before infection with a dose of virus that is otherwise rapidly cleared led to persistent infection. These results suggested that the size of the viral inoculum may contribute to the outcome of infection by altering the balance between the kinetics and magnitude of infection versus the kinetics and magnitude of the immune response. Similar results have been recently published based on in situ analysis of the ratio of virus-infected cells to immune effector cells in the tissues of simian immunodeficiency virus-infected macaques and lymphocytic choriomeningitis virus-infected mice (20).Collectively, these results suggest that the kinetics of T-cell priming relative to the kinetics of viral spread determines the outcome of HBV infection. Specifically, they suggest that early priming of the CD4⁺ T-cell response before or during viral spread initiates a vigorous, synchronized, and functionally efficient CD8⁺ T-cell response and the accompanying immunopathology that ultimately terminates HBV infection. In contrast, the virus persists when CD4⁺ T-cell priming is delayed until after all of the hepatocytes are infected.     

UNC93B1 Mediates Host Resistance to Infection with Toxoplasma gondii

UNC93B1 associates with Toll-Like Receptor (TLR) 3, TLR7 and TLR9, mediating their translocation from the endoplasmic reticulum to the endolysosome, hence allowing proper activation by nucleic acid ligands. We found that the triple deficient ‘3d’ mice, which lack functional UNC93B1, are hyper-susceptible to infection with Toxoplasma gondii. We established that while mounting a normal systemic pro-inflammatory response, i.e. producing abundant MCP-1, IL-6, TNFα and IFNγ, the 3d mice were unable to control parasite replication. Nevertheless, infection of reciprocal bone marrow chimeras between wild-type and 3d mice with T. gondii demonstrated a primary role of hemopoietic cell lineages in the enhanced susceptibility of UNC93B1 mutant mice. The protective role mediated by UNC93B1 to T. gondii infection was associated with impaired IL-12 responses and delayed IFNγ by spleen cells. Notably, in macrophages infected with T. gondii, UNC93B1 accumulates on the parasitophorous vacuole. Furthermore, upon in vitro infection the rate of tachyzoite replication was enhanced in non-activated macrophages carrying mutant UNC93B1 as compared to wild type gene. Strikingly, the role of UNC93B1 on intracellular parasite growth appears to be independent of TLR function. Altogether, our results reveal a critical role for UNC93B1 on induction of IL-12/IFNγ production as well as autonomous control of Toxoplasma replication by macrophages.     

Persistent Hz-1 Virus Infection in Insect Cells: Evidence for Insertion of Viral DNA into Host Chromosomes and Viral Infection in a Latent Status

Persistent/latent viral infections of insect cells are a prominent though poorly understood phenomenon. In this study, the long-term association between the Hz-1 virus and insect host cells, conventionally referred to as persistent viral infection, is described. With the aid of a newly developed fluorescent cell-labeling system, we found that productive viral replication occurs by spontaneous viral reactivation in fewer than 0.2% of persistently infected cell lines over a 5-day period. Once viral reactivation takes place, the host cell dies. The persistently infected cells contain various amounts of viral DNA, and, in an extreme case, up to 16% of the total DNA isolated from infected cells could be of viral origin. Both pulsed-field gel electrophoresis and in situ hybridization experiments showed that some of these viral DNA molecules are inserted into the host chromosomes but that the rest of viral DNA copies are free from host chromosomes. Thus, Hz-1 virus is the first nonretroviral insect virus known to insert its genome into the host chromosome during the infection process. These data also suggest that the previously described persistent infection of Hz-1 virus in insect cells should be more accurately referred to as latent viral infection.     

Association Between IL-10 Gene Promoter Polymorphism and Hepatitis B Viral Infection in an Egyptian Population

Cytokines play critical roles in the pathogenesis of hepatitis B virus infection (HBV). This work was designed to study the effect of IL-10 gene polymorphisms (?1082G/A and ?819C/T) on susceptibility of Egyptians to HBV. Genotyping was performed using single-stranded polymorphism-polymerase chain reaction in 118 Egyptian hepatitis B patients and 119 healthy controls, and IL-10 serum levels were measured using ELISA. The frequency of IL-10 ?1082G/G was significantly higher in HBV patients than in healthy controls, and G/A and A/A were not significantly different between groups. The distribution of IL-10 ?819 genotypes was not significantly different between the HBV and healthy control groups. Although AT was significantly different between controls and patients, the distribution of the other haplotypes was not. IL-10 levels were significantly lower among hepatitis B patients. Our data stress the importance of IL-10 gene polymorphism in HBV infection. Depending on our preliminary work, IL-10 ?1082G/G may act as a host genetic factor in the susceptibility to HBV infection in Egyptians.     

The Effect of Arachidonic Acid and Viral Infection on the Phytohemagglutinin Activity during the Development of Tobacco Acquired Resistance

The effects of arachidonic acid (AA) on the development of viral infection and the activity of phytohemagglutinins in Nicotiana tabacum L. plants were studied. Cv. Samsun NN was used, which displayed a genotypically determined hypersensitive response to tobacco mosaic virus (TMV) infection. When tobacco leaf disks were treated with 10^–9 to –10^–7 M AA, viral reproduction was suppressed by 90–100%. The AA concentration of 10^–8 M was optimal for the improvement of plant virus resistance. Tobacco leaves maintained virus resistance for at least two weeks. Both AA treatment and TMV inoculation were accompanied by an enhanced lectin activity, which may indicate the involvement of lectins in the development of plant defense responses. Lectin accumulation was observed in the intact plants developing systemic resistance and in the detached leaves characterized by local resistance.     

Single Origin of Sex Chromosomes and Multiple Origins of B Chromosomes in Fish Genus Characidium

Chromosome painting with DNA probes obtained from supernumerary (B) and sex chromosomes in three species of fish genus Characidium (C. gomesi, C. pterostictum and C. oiticicai) showed a close resemblance in repetitive DNA content between B and sex chromosomes in C. gomesi and C. pterostictum. This suggests an intraspecific origin for B chromosomes in these two species, probably deriving from sex chromosomes. In C. oiticicai, however, a DNA probe obtained from its B chromosome hybridized with the B but not with the A chromosomes, suggesting that the B chromosome in this species could have arisen interspecifically, although this hypothesis needs further investigation. A molecular phylogenetic analysis performed on nine Characidium species, with two mtDNA genes, showed that the presence of heteromorphic sex chromosomes in these species is a derived condition, and that their origin could have been unique, a conclusion also supported by interspecific chromosome painting with a CgW probe derived from the W chromosome in C. gomesi. Summing up, our results indicate that whereas heteromorphic sex chromosomes in the genus Characidium appear to have had a common and unique origin, B chromosomes may have had independent origins in different species. Our results also show that molecular phylogenetic analysis is an excellent complement for cytogenetic studies by unveiling the direction of evolutionary chromosome changes.     

Protective Effects of Specific Immunity to Viral Neuraminidase on Influenza Virus Infection of Mice

Antibody specific for viral neuraminidase can be demonstrated in mice following (i) pulmonary infection with influenza virus, (ii) immunization with ultraviolet-in-activated influenza virus, (iii) immunization with isolated neuraminidase of influenza A(2) virus, and (iv) passive immunization with sera of rabbits immunized with isolated A(2) neuraminidase. Neuraminidase antibody produced by any of these methods exerts a profound inhibiting effect on virus replication in the lungs of mice challenged with strains of virus having homologous neuraminidase protein, even in the absence of hemagglutinating inhibiting antibody to the challenge virus, and results in markedly decreased pulmonary virus titers and diminished lung lesions. These observations suggest that antineuraminidase immunity may play a significant role in the protection against influenza virus challenge observed in mice after infection or artificial immunization.     

内生真菌对植物生长发育及抗逆性的影响

本文从内生真菌的涵义和基本生物学特征出发,以国内外近二十年来关于内生真菌与植物关系的研究情况作了综述。主要包括内生真菌的检测、鉴定和转染,内生真菌对植物生长发育与抗逆性的影响,以及内生真菌在农业及其他相关领域的应用前景。文中提及作者近期开展的关于内生真菌对黑麦草抗旱性影响的研究课题及其初步进展。     

Disease Interactions in a Shared Host Plant: Effects of Pre-Existing Viral Infection on Cucurbit Plant Defense Responses and Resistance to Bacterial Wilt Disease

Both biotic and abiotic stressors can elicit broad-spectrum plant resistance against subsequent pathogen challenges. However, we currently have little understanding of how such effects influence broader aspects of disease ecology and epidemiology in natural environments where plants interact with multiple antagonists simultaneously. In previous work, we have shown that healthy wild gourd plants (Cucurbita pepo ssp. texana) contract a fatal bacterial wilt infection (caused by Erwinia tracheiphila) at significantly higher rates than plants infected with Zucchini yellow mosaic virus (ZYMV). We recently reported evidence that this pattern is explained, at least in part, by reduced visitation of ZYMV-infected plants by the cucumber beetle vectors of E. tracheiphila. Here we examine whether ZYMV-infection may also directly elicit plant resistance to subsequent E. tracheiphila infection. In laboratory studies, we assayed the induction of key phytohormones (SA and JA) in single and mixed infections of these pathogens, as well as in response to the feeding of A. vittatum cucumber beetles on healthy and infected plants. We also tracked the incidence and progression of wilt disease symptoms in plants with prior ZYMV infections. Our results indicate that ZYMV-infection slightly delays the progression of wilt symptoms, but does not significantly reduce E. tracheiphila infection success. This observation supports the hypothesis that reduced rates of wilt disease in ZYMV-infected plants reflect reduced visitation by beetle vectors. We also documented consistently strong SA responses to ZYMV infection, but limited responses to E. tracheiphila in the absence of ZYMV, suggesting that the latter pathogen may effectively evade or suppress plant defenses, although we observed no evidence of antagonistic cross-talk between SA and JA signaling pathways. We did, however, document effects of E. tracheiphila on induced responses to herbivory that may influence host-plant quality for (and hence pathogen acquisition by) cucumber beetles.     

Influence of the Presence of B Chromosomes on DNA Damage in Crepis capillaris

The sensitivity of different plant species to mutagenic agents is related to the DNA content and organization of the chromatin, which have been described in ABCW and bodyguard hypotheses, respectively. Plant species that have B chromosomes are good models for the study of these hypotheses. This study presents an analysis of the correlation between the occurrence of B chromosomes and the DNA damage that is induced by the chemical mutagen, maleic hydrazide (MH), in Crepis capillaris plants using comet assay. The presence of B chromosomes has a detectable impact on the level of DNA damage. The level of DNA damage after MH treatment was correlated with the number of B chromosomes and it was observed that it increased significantly in plants with 3B chromosomes. We did not find evidence of the protective role from chemical mutagens of the constitutive heterochromatin for euchromatin in relation to DNA damage. The DNA damage involving the 25S rDNA sequences was analyzed using the comet-FISH technique. Fragmentation within or near the 25S rDNA involved the loci on the A and B chromosomes. The presence of B chromosomes in C. capillaris cells had an influence on the level of DNA damage that involves the 25S rDNA region.     

Genomic Modifiers of Natural Killer Cells,Immune Responsiveness and Lymphoid Tissue Remodeling Together Increase Host Resistance to Viral Infection

The MHC class I D^k molecule supplies vital host resistance during murine cytomegalovirus (MCMV) infection. Natural killer (NK) cells expressing the Ly49G2 inhibitory receptor, which specifically binds D^k, are required to control viral spread. The extent of D^k-dependent host resistance, however, differs significantly amongst related strains of mice, C57L and MA/My. As a result, we predicted that relatively small-effect modifier genetic loci might together shape immune cell features, NK cell reactivity, and the host immune response to MCMV. A robust D^k-dependent genetic effect, however, has so far hindered attempts to identify additional host resistance factors. Thus, we applied genomic mapping strategies and multicolor flow cytometric analysis of immune cells in naive and virus-infected hosts to identify genetic modifiers of the host immune response to MCMV. We discovered and validated many quantitative trait loci (QTL); these were mapped to at least 19 positions on 16 chromosomes. Intriguingly, one newly discovered non-MHC locus (Cmv5) controlled splenic NK cell accrual, secondary lymphoid organ structure, and lymphoid follicle development during MCMV infection. We infer that Cmv5 aids host resistance to MCMV infection by expanding NK cells needed to preserve and protect essential tissue structural elements, to enhance lymphoid remodeling and to increase viral clearance in spleen.