Sequential MyD88-Independent and -Dependent Activation of Innate Immune Responses to Intracellular Bacterial Infection

AbstractMicrobial infections induce chemokine and cytokine cascades that coordinate innate immune defenses. Infection with the intracellular bacterial pathogen Listeria monocytogenes induces CCR2-dependent monocyte recruitment and activation, an essential response for host survival. Herein we show that invasive L. monocytogenes, but not killed or noninvasive bacteria, induce secretion of MCP-1, the requisite chemokine for monocyte recruitment. Induction of MCP-1, but not TNF or IL-12, following L. monocytogenes infection is MyD88 independent. Consistent with these results, MyD88 deficiency does not impair monocyte recruitment to L. monocytogenes infected spleens, but prevents monocyte activation. Our results indicate that distinct microbial signals activate innate immune responses in an ordered, step-wise fashion, providing a mechanism to specify and modulate antimicrobial effector functions

Available cloned genes and markers for genetic improvement of biotic stress resistance in rice

Biotic stress is one of the major threats to stable rice production. Climate change affects the shifting of pest outbreaks in time and space. Genetic improvement of biotic stress resistance in rice is a cost-effective and environment-friendly way to control diseases and pests compared to other methods such as chemical spraying. Fast deployment of the available and suitable genes/alleles in local elite varieties through marker-assisted selection (MAS) is crucial for stable high-yield rice production. In this review, we focused on consolidating all the available cloned genes/alleles conferring resistance against rice pathogens (virus, bacteria, and fungus) and insect pests, the corresponding donor materials, and the DNA markers linked to the identified genes. To date, 48 genes (independent loci) have been cloned for only major biotic stresses: seven genes for brown planthopper (BPH), 23 for blast, 13 for bacterial blight, and five for viruses. Physical locations of the 48 genes were graphically mapped on the 12 rice chromosomes so that breeders can easily find the locations of the target genes and distances among all the biotic stress resistance genes and any other target trait genes. For efficient use of the cloned genes, we collected all the publically available DNA markers (~500 markers) linked to the identified genes. In case of no available cloned genes yet for the other biotic stresses, we provided brief information such as donor germplasm, quantitative trait loci (QTLs), and the related papers. All the information described in this review can contribute to the fast genetic improvement of biotic stress resistance in rice for stable high-yield rice production

Available cloned genes and markers for genetic improvement of biotic stress resistance in rice

Biotic stress is one of the major threats to stable rice production. Climate change affects the shifting of pest outbreaks in time and space. Genetic improvement of biotic stress resistance in rice is a cost-effective and environment-friendly way to control diseases and pests compared to other methods such as chemical spraying. Fast deployment of the available and suitable genes/alleles in local elite varieties through marker-assisted selection (MAS) is crucial for stable high-yield rice production. In this review, we focused on consolidating all the available cloned genes/alleles conferring resistance against rice pathogens (virus, bacteria, and fungus) and insect pests, the corresponding donor materials, and the DNA markers linked to the identified genes. To date, 48 genes (independent loci) have been cloned for only major biotic stresses: seven genes for brown planthopper (BPH), 23 for blast, 13 for bacterial blight, and five for viruses. Physical locations of the 48 genes were graphically mapped on the 12 rice chromosomes so that breeders can easily find the locations of the target genes and distances among all the biotic stress resistance genes and any other target trait genes. For efficient use of the cloned genes, we collected all the publically available DNA markers (~500 markers) linked to the identified genes. In case of no available cloned genes yet for the other biotic stresses, we provided brief information such as donor germplasm, quantitative trait loci (QTLs), and the related papers. All the information described in this review can contribute to the fast genetic improvement of biotic stress resistance in rice for stable high-yield rice production

Development and evaluation of introgression lines with yield enhancing genes of the Indian mega-variety of rice, MTU1010

MTU 1010 is an early maturing and high-yielding mega rice variety widely grown in an area of 3 Mha. It is characterised by limited grain number and panicle branching. To improve the grain number in MTU 1010, an IRRI breeding line, IR121055-2-10-5 was utilized as donor to transfer yield-enhancing genes Gn1a and OsSPL14 (associated with increased grain number and better panicle branching, respectively) into MTU1010 by Marker-Assisted Backcross Breeding (MABB). At each backcross generation, foreground selection was carried out with Gn1a and OsSPL14- specific molecular markers, whilst background selection was done with a set of SSR markers polymorphic between the IR121055-2-10-5 and MTU1010. With the use of a gene-specific marker, homozygous BC2 F2 plants carrying the yield-enhancing gene were identified and advanced through pedigree-method of selection till BC2 F6 and best performing ten lines were selected and evaluated in replicated station trials for yield contributing traits, where grain number and brancing per panicle exhibited high significant and positive correlation with single plant yield. Three promising lines namely RP6353-5-8-13-24, RP6353-26-13-39-5 and RP6353-32-12-8-16 with higher grain number and yield than MTU1010 were identified and nominated for evaluation in Initial Varietal Trial-Aerobic (IVT-Aerobic) of All India Crop Improvement Programme on Rice (AICRP), of which RP6353-26-13-39-5 (IET28674), was promoted for further testing

Fine mapping and sequence analysis reveal a promising candidate gene encoding a novel NB-ARC domain derived from wild rice (Oryza officinalis) that confers bacterial blight resistance

Bacterial blight disease of rice caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the most serious constraints in rice production. The most sustainable strategy to combat the disease is the deployment of host plant resistance. Earlier, we identified an introgression line, IR 75084-15-3-B-B, derived from Oryza officinalis possessing broad-spectrum resistance against Xoo. In order to understand the inheritance of resistance in the O. officinalis accession and identify genomic region(s) associated with resistance, a recombinant inbred line (RIL) mapping population was developed from the cross Samba Mahsuri (susceptible to bacterial blight) × IR 75084-15-3-B-B (resistant to bacterial blight). The F2 population derived from the cross segregated in a phenotypic ratio of 3: 1 (resistant susceptible) implying that resistance in IR 75084-15-3-B-B is controlled by a single dominant gene/quantitative trait locus (QTL). In the F7 generation, a set of 47 homozygous resistant lines and 47 homozygous susceptible lines was used to study the association between phenotypic data obtained through screening with Xoo and genotypic data obtained through analysis of 7K rice single-nucleotide polymorphism (SNP) chip. Through composite interval mapping, a major locus was detected in the midst of two flanking SNP markers, viz., Chr11.27817978 and Chr11.27994133, on chromosome 11L with a logarithm of the odds (LOD) score of 10.21 and 35.93% of phenotypic variation, and the locus has been named Xa48t. In silico search in the genomic region between the two markers flanking Xa48t identified 10 putatively expressed genes located in the region of interest. The quantitative expression and DNA sequence analysis of these genes from contrasting parents identified the Os11g0687900 encoding an NB-ARC domain-containing protein as the most promising gene associated with resistance. Interestingly, a 16-bp insertion was noticed in the untranslated region (UTR) of the gene in the resistant parent, IR 75084-15-3-B-B, which was absent in Samba Mahsuri. The association of Os11g0687900 with resistance phenotype was further established by sequence-based DNA marker analysis in the RIL population. A co-segregating PCR-based INDEL marker, Marker_Xa48, has been developed for use in the marker-assisted breeding of Xa48t

Development of Oryza sativa L. by Oryza punctata Kotschy ex Steud. monosomic addition lines with high value traits by interspecific hybridization

Oryza punctata\ua0is a distantly related wild\ua0Oryza\ua0species having BB genome with untapped genetic resources for rice improvement. Low crossability between the cultivated\ua0O. sativa\ua0and\ua0O. punctata\ua0restricts the success of transferring many desirable traits into cultivated rice. Artificially induced autotetraploids of an elite breeding line, IR31917-45-3-2, were produced and crossed with\ua0O. punctata. Allotriploid F1\ua0plants were backcrossed to IR31917-45-3-2 and generated progenies with extra chromosomes from\ua0O. punctata. Twenty BC1F1\ua0and 59 BC2F1\ua0plants were produced with chromosome numbers ranging from 24 (2n) to 29 (2n\ua0+\ua05) and 2n\ua0(24) to 26 (2n\ua0+\ua02), respectively. Eleven monosomic alien addition lines (MAALs) were characterized morphologically and cytologically and designated as MAAL 1–12. MAALs were genotyped using\ua0O. punctata\ua0genome-specific molecular markers and detected chromosome segments inherited from\ua0O. punctata.\ua0O. punctata\ua0introgressions across all the chromosomes of\ua0O. sativa\ua0were identified except for chromosome 8. The most frequent introgressions were observed in chromosomes 4, 6, 10, and 11, which could be the recombination hotspots between A and B genomes. Some of the qualitative traits such as black hull, purple coleoptile base, purple stigma, long awn, and short grain size from\ua0O. punctata\ua0were inherited in some disomic introgression lines (DILs). Several DILs inherited genes from\ua0O. punctata\ua0conferring resistance to brown planthopper, green leafhopper, and diseases such as bacterial blight and blast. This is the first report on successful gene transfer from\ua0O. punctata\ua0into\ua0O. sativa

Exploring Key Blast and Bacterial Blight Resistance Genes in Genetically Diverse Rice Accessions through Molecular and Phenotypic Evaluation

Blast and bacterial blight (BB) are the most dangerous rice (Oryza sativa\ua0L.) diseases that limit rice production significantly.\ua0Pib,\ua0Piz‐t, and\ua0Pi9\ua0are reported as key resistance genes for blast whereas\ua0Xa21,\ua0Xa4,\ua0Xa7, and\ua0xa13\ua0are considered as important resistance genes for BB. Using gene‐specific DNA markers, the presence of these resistance genes was screened in 211 diverse rice accessions originating from 26 countries. In molecular marker analyses, specific amplification patterns for the\ua0Pib\ua0and\ua0Piz‐t\ua0resistance alleles were observed in 56 and 23 accessions, respectively, whereas the\ua0Pi9\ua0resistance allele was not observed at all in these accessions. For BB, at least one BB resistance gene was present in 148 of the 211 evaluated accessions. All 211 accessions were evaluated for blast resistance using natural isolates and for BB resistance using Race 4 (PX071) and Race 6 (PX099). Among 211 accessions, 89 exhibited hypersensitive blast resistance reactions, whereas 85 and 37 accessions were rated as resistant or moderately resistance to BB Races 4 and 6, respectively. The combined analysis of molecular and phenotypic reactions (marker‐trait association assay) revealed that landraces possessed rare and several desirable genes compared with breeding lines with a narrow genetic base, hence these landraces serve as the valuable source for exploring new resistance genes for crop improvement. An interesting similarity in gene distribution pattern was observed in\ua0Pib\ua0with\ua0Xa21\ua0and in\ua0Piz‐t\ua0with\ua0Xa7. The analyzed blast and BB resistance genes were in a range of combinations in different landraces and breeding lines, which can be used in gene introgression and pyramiding programs as alternative resistance sources

Adapting Genotyping-by-Sequencing for Rice F2 Populations

Rapid and cost-effective genotyping of large mapping populations can be achieved by sequencing a reduced representation of the genome of every individual in a given population, and using that information to generate genetic markers. A customized genotyping-by-sequencing (GBS) pipeline was developed to genotype a rice F2 population from a cross of Oryza sativa ssp. japonica cv. Nipponbare and the African wild rice species O. longistaminata. While most GBS pipelines aim to analyze mainly homozygous populations, we attempted to genotype a highly heterozygous F2 population. We show how species- and population-specific improvements of established protocols can drastically increase sample throughput and genotype quality. Using as few as 50,000 reads for some individuals (134,000 reads on average), we were able to generate up to 8154 informative SNP markers in 1081 F2 individuals. Additionally, the effects of enzyme choice, read coverage, and data postprocessing are evaluated. Using GBS-derived markers, we were able to assemble a genetic map of 1536 cM. To demonstrate the usefulness of our GBS pipeline, we determined quantitative trait loci (QTL) for the number of tillers. We were able to map four QTL to chromosomes 1, 3, 4, and 8, and partially confirm their effects using introgression lines. We provide an example of how to successfully use GBS with heterozygous F2 populations. By using the comparatively low-cost MiSeq platform, we show that the GBS method is flexible and cost-effective, even for smaller laboratories

Development of an intergeneric hybrid between Oryza sativa L. and Leersia perrieri (A. Camus) Launert

An intergeneric hybrid was successfully developed between\ua0Oryza sativa\ua0L. (IRRI 154) and\ua0Leersia perrieri\ua0(A. Camus) Launert using embryo rescue technique in this study. A low crossability value (0.07%) implied that there was high incompatibility between the two species of the hybrid. The F1\ua0hybrid showed intermediate phenotypic characteristics between the parents but the plant height was very short. The erect plant type resembled the female parent IRRI 154 but the leaves were similar to\ua0L. perrieri. Cytological analysis revealed highly non-homology between chromosomes of the two species as the F1\ua0plants showed 24 univalents without any chromosome pairing. The F1\ua0hybrid plant was further confirmed by PCR analysis using the newly designed 11 indel markers showing polymorphism between\ua0O. sativa\ua0and\ua0L. perrieri. This intergeneric hybrid will open up opportunities to transfer novel valuable traits from\ua0L. perrieri\ua0into cultivated rice