Incompatible Crosses Research Articles

Warm temperature inhibits cytoplasmic incompatibility induced by endosymbiotic Rickettsiella in spider hosts.

Bacterial endosymbionts manipulate reproduction in arthropods to increase their prevalence in the host population. One such manipulation is cytoplasmic incompatibility (CI), wherein the bacteria sabotage sperm in infected males to reduce the hatch rate when mated with uninfected females, but zygotes are 'rescued' when that male mates with an infected female. In the spider Mermessus fradeorum (Linyphiidae), Rickettsiella symbionts cause variable levels of CI. We hypothesised that temperature affects the strength of CI and its rescue in M. fradeorum, potentially mediated by bacterial titre. We reared Rickettsiella-infected spiders in two temperature conditions (26°C vs. 20°C) and tested CI induction in males and rescue in females. In incompatible crosses between infected males and uninfected females, the hatch rate from warm males was doubled (mean ± standard error = 0.687 ± 0.052) relative to cool males (0.348 ± 0.046), indicating that CI induction is weaker in warm males. In rescue crosses between infected females and infected males, female rearing temperature had a marginal effect on CI rescue, but the hatch rate remained high for both warm (0.960 ± 0.023) and cool females (0.994 ± 0.004). Bacterial titre, as measured by quantitative polymerase chain reaction, was lower in warm than cool spiders, particularly in females, suggesting that bacterial titre may play a role in causing the temperature-mediated changes in CI.

Major chromosome rearrangements in intergeneric wheat × rye hybrids in compatible and incompatible crosses detected by GBS read coverage analysis

The presence of incompatibility alleles in primary amphidiploids constitutes a reproductive barrier in newly synthesized wheat-rye hybrids. To overcome this barrier, the genome stabilization process includes large-scale chromosome rearrangements. In incompatible crosses resulting in fertile amphidiploids, the elimination of one of the incompatible alleles Eml-A1 or Eml-R1b can occur already in the somatic tissue of the wheat × rye hybrid embryo. We observed that the interaction of incompatible loci Eml-A1 of wheat and Eml-R1b of rye after overcoming embryo lethality leads to hybrid sterility in primary triticale. During subsequent seed reproductions (R1, R2 or R3) most of the chromosomes of A, B, D and R subgenomes undergo rearrangement or eliminations to increase the fertility of the amphidiploid by natural selection. Genotyping-by-sequencing (GBS) coverage analysis showed that improved fertility is associated with the elimination of entire and partial chromosomes carrying factors that either cause the disruption of plant development in hybrid plants or lead to the restoration of the euploid number of chromosomes (2n = 56) in the absence of one of the incompatible alleles. Highly fertile offspring obtained in compatible and incompatible crosses can be successfully adapted for the production of triticale pre-breeding stocks.

Pectin methylesterase activities in reproductive tissues of maize plants with different haplotypes of the Ga1 and Ga2 cross incompatibility systems

Key messageTotal PME activity in reproductive tissues was related to haplotypes at maize cross incompatibility loci, suggesting that these loci function by controlling PME activity.In maize, the pollination outcome depends on the haplotypes of the interacting male gametophyte (germinated pollen) and female sporophyte (silk) at several cross-incompatibility loci. Functional alleles (-S haplotypes) of the cross-incompatibility loci Ga1 and Ga2, both encode two pectin methylesterases (PMEs), one that is expressed in silk and the other in pollen. We examined total PME activity in reproductive tissues containing functional and null haplotypes at the Ga1 or Ga2 loci. In pollinated silks, there was a correlation between total PME activity and the -S haplotype pollen in both Ga1 and Ga2 systems. We did not detect a significant relationship between PME activity and pollination outcome of either system. We re-examined previously reported active site amino acid substitutions in PMEs encoded by cross incompatibility loci. We observed that different active site substitutions are present in the pollen and silk PMEs of cross incompatibility loci and these differences are conserved across Ga1, Ga2 and Tcb-1. This work establishes a relationship between total PME activity and the haplotypes of the Ga1 locus in pollinated silks.

Bacterial community and genome analysis of cytoplasmic incompatibility-inducing Wolbachia in American serpentine leafminer, Liriomyza trifolii.

Liriomyza trifolii, an agricultural pest, is occasionally infected by Wolbachia. A Wolbachia strain present in Liriomyza trifolii is associated with cytoplasmic incompatibility (CI) effects, leading to the death of embryos resulting from incompatible crosses between antibiotic-treated or naturally Wolbachia-free strain females and Wolbachia-infected males. In this study, high-throughput sequencing of hypervariable rRNA genes was employed to characterize the bacterial community in Wolbachia-infected L. trifolii without antibiotic treatment. The analysis revealed that Wolbachia dominates the bacterial community in L. trifolii, with minor presence of Acinetobacter, Pseudomonas, and Limnobacter. To elucidate the genetic basis of the CI phenotype, metagenomic sequencing was also conducted to assemble the genome of the Wolbachia strain. The draft-genome of the Wolbachia strain wLtri was 1.35 Mbp with 34% GC content and contained 1,487 predicted genes. Notably, within the wLtri genome, there are three distinct types of cytoplasmic incompatibility factor (cif) genes: Type I, Type III, and Type V cifA;B. These genes are likely responsible for inducing the strong cytoplasmic incompatibility observed in L. trifolii.

Intra-lineage microevolution of Wolbachia leads to the emergence of new cytoplasmic incompatibility patterns.

Mosquitoes of the Culex pipiens complex are worldwide vectors of arbovirus, filarial nematodes, and avian malaria agents. In these hosts, the endosymbiotic bacteria Wolbachia induce cytoplasmic incompatibility (CI), i.e., reduced embryo viability in so-called incompatible crosses. Wolbachia infecting Culex pipiens (wPip) cause CI patterns of unparalleled complexity, associated with the amplification and diversification of cidA and cidB genes, with up to 6 different gene copies described in a single wPip genome. In wPip, CI is thought to function as a toxin-antidote (TA) system where compatibility relies on having the right antidotes (CidA) in the female to bind and neutralize the male's toxins (CidB). By repeating crosses between Culex isofemale lines over a 17 years period, we documented the emergence of a new compatibility type in real time and linked it to a change in cid genes genotype. We showed that loss of specific cidA gene copies in some wPip genomes results in a loss of compatibility. More precisely, we found that this lost antidote had an original sequence at its binding interface, corresponding to the original sequence at the toxin's binding interface. We showed that these original cid variants are recombinant, supporting a role for recombination rather than point mutations in rapid CI evolution. These results strongly support the TA model in natura, adding to all previous data acquired with transgenes expression.

Boron homeostasis affects Longan yield: a study of NIP and BOR boron transporter of two cultivars

BackgroundEssential micronutrient Boron (B) plays crucial roles in plant survival and reproduction but becomes toxic in higher quantities. Although plant cells have different B transport systems, B homeostasis is mainly maintained by two transporter protein families: B exporters (BOR) and nodulin-26-like intrinsic proteins (NIP). Their diversity and differential expression are responsible for varied B tolerance among plant varieties and species. Longan is a highly admired subtropical fruit with a rising market in China and beyond. In the present study, we cultured Shixia (SX) and Yiduo (YD), two differently characterized Longan cultivars, with foliar B spray. We analyzed their leaf physiology, fruit setting, B content, and boron transporter gene expression of various tissue samples. We also traced some of these genes' subcellular localization and overexpression effects.ResultsYD and SX foliage share similar microstructures, except the mesophyll cell wall thickness is double in YD. The B spray differently influenced their cellular constituents and growth regulators. Gene expression analysis showed reduced BOR genes expression and NIP genes differential spatiotemporal expression. Using green fluorescent protein, two high-expressing NIPs, NIP1 and NIP19, were found to translocate in the transformed tobacco leaves' cell membrane. NIPs transformation of SX pollen was confirmed using magnetic beads and quantified using a fluorescence microscope and polymerase chain reaction. An increased seed-setting rate was observed when YD was pollinated using these pollens. Between the DlNIP1 and DlNIP19 transformed SX pollen, the former germinated better with increasing B concentrations and, compared to naturally pollinated plants, had a better seed-setting rate in YD♀ × SX♂.ConclusionSX and YD Longan have different cell wall structures and react differently to foliar B spray, indicating distinct B tolerance and management. Two B transporter NIP genes were traced to localize in the plasma membrane. However, under high B concentrations, their differential expression resulted in differences in Jasmonic acid content, leading to differences in germination rate. Pollination of YD using these NIPs transformed SX pollen also showed NIP1 overexpression might overcome the unilateral cross incompatibility between YD♀ × SX♂ and can be used to increase Longan production.

Breeding Cultivars for Resistance to the African Sweetpotato Weevils, Cylas puncticollis and Cylas brunneus, in Uganda: A Review of the Current Progress.

In sub-Saharan Africa, sweetpotato weevils are the major pests of cultivated sweetpotato, causing estimated losses of between 60% and 100%, primarily during dry spells. The predominantly cryptic feeding behavior of Cylas spp. within their roots makes their control difficult, thus, host plant resistance is one of the most promising lines of protection against these pests. However, limited progress has been made in cultivar breeding for weevil resistance, partly due to the complex hexaploid genome of sweetpotato, which complicates conventional breeding, in addition to the limited number of genotypes with significant levels of resistance for use as sources of resistance. Pollen sterility, cross incompatibility, and poor seed set and germination in sweetpotato are also common challenges in improving weevil resistance. The accurate phenotyping of sweetpotato weevil resistance to enhance the efficiency of selection has been equally difficult. Genomics-assisted breeding, though in its infancy stages in sweetpotato, has a potential application in overcoming some of these barriers. However, it will require the development of more genomic infrastructure, particularly single-nucleotide polymorphism markers (SNPs) and robust next-generation sequencing platforms, together with relevant statistical procedures for analyses. With the recent advances in genomics, we anticipate that genomic breeding for sweetpotato weevil resistance will be expedited in the coming years. This review sheds light on Uganda's efforts, to date, to breed against the Cylas puncticollis (Boheman) and Cylas brunneus (Fabricius) species of African sweetpotato weevil.

The Ga1 locus of the genus Zea is associated with novel genome structures derived from multiple, independent non-homologous recombination events.

The Ga1 locus controls cross incompatibility between field corn and popcorn. The Ga1-S haplotype contains two types of pectin methylesterase (PME) genes, ZmPme3 and several copies of ZmGa1P, that are expressed in silk and pollen, respectively. The ga1 haplotype contains non-functional tandem repeat sequences related to ZmPme3 and ZmGa1P. This haplotype can cross pollinate freely and is widely present in field corn. The primary objective of this study is to characterize the repeat sequences from a diverse collection of maize and teosinte lines and use this information to understand the evolution of the Ga1 locus. First, we characterized the complexity of the Ga1 genome region in high quality maize genome assemblies which led to their categorization into five groups based on the number and type of PME-like sequences found at this region. Second, we studied duplication events that led to the ga1 and Ga1-S repeats using maximum-likelihood phylogenetic reconstruction. Divergence estimates of the ga1 haplotype suggest that the duplication events occurred more than 600 kya whereas those in Ga1-S occurred at three time points i.e., > 600 kya, ∼ 260 kya and ∼ 100 kya. These estimates suggest that the ga1 and Ga1-S tandem duplication events occurred independently. Finally, analysis of ZmPme3 and ZmGa1P homologs in Zea and Tripsacum genomes suggests that ga1 and Ga1-S repeats originated from an ancestral pair of PME genes that duplicated and diverged through two evolutionary branches prior to the domestication of maize.

Incomplete reproductive barriers and genomic differentiation impact the spread of resistance mutations between green- and red-colour morphs of a cosmopolitan mite pest.

Pesticide resistance represents a clear and trackable case of adaptive evolution with a strong societal impact. Understanding the factors associated with the evolution and spread of resistance is imperative to develop sustainable crop management strategies. The two-spotted spider mite Tetranychus urticae, a major crop pest with worldwide distribution and a polyphagous lifestyle, has evolved resistance to most classes of pesticides. Tetranychus urticae exists as either a green- or a red-coloured morph. However, the extent of genetic divergence and reproductive compatibility vary across populations of these colour morphs, complicating their taxonomic resolution at the species level. Here, we studied patterns of genetic differentiation and barriers to gene flow within and between morphs of T. urticae in order to understand the factors that influence the spread of resistance mutations across its populations. We derived multiple iso-female lines from Tetranychus populations collected from agricultural crops. We generated genomic and morphological data, characterized their bacterial communities and performed controlled crosses. Despite morphological similarities, we found large genomic differentiation between the morphs. This pattern was reflected in the incomplete, but strong postzygotic incompatibility in crosses between colour morphs, while crosses within morphs from different geographical locations were largely compatible. In addition, our results suggest recent/on-going gene flow between green-coloured T. urticae and T. turkestani. By screening the sequences of 10 resistance genes, we found evidence for multiple independent origins and for single evolutionary origins of target-site resistance mutations. Our results indicate that target-site mutations mostly evolve independently in populations on different geographical locations, and that these mutations can spread due to incomplete barriers to gene flow within and between populations.

Differences in morphology, mitochondrial genomes, and reproductive compatibility between two clades of parasitic wasps Aphelinus mali (Hymenoptera: Aphelindae) in China.

Aphelinus mali (Haldeman) (Hymenoptera: Aphelinidae) in China is comprised of two clades (termed, the Shandong and Liaoning clades). In order to clarify the genetic relationship between these two clades, we compared and analyzed the morphological characteristics and the mitochondrial genome of each, and performed a hybridization experiment. Morphological results showed that both males and females of the Liaoning clade were larger than Shandong clade, in terms of whole body, abdominal, wing and antennal lengths, however, there were no significant differences between clades for total length of the middle or hind leg of females. The length of the mitochondrial genome of the Shandong clade was 14415 bp and, for the Liaoning clade, it was 14804 bp. Each contained 31 genes, including 13 protein-encoded genes, 16 tRNA genes, and 2 rRNA genes. The highest AT level among the 13 protein-coding genes for the two clades were the same gene (ATP8) (Shandong clade, 91.52%; Liaoning clade, 90.91%). By hybridization and backcrossing, we found that there was no cross incompatibility between these two clades of A. mali. Our results indicate that the historic geographical isolation between these clades has not yet caused reproductive isolation of these populations, and they belong to the same species.

TISSUE CULTURE AND ITS APPLICATION IN MODERN AGRICULTURE

Tissue culture is a technique of growing plant cells, tissues and organs in an artificial prepared medium under aseptic conditions, which was invented by Gottlieb Haberlandt in 1902. Plant tissue culture is based on cellular totipotency, dedifferentiation, and redifferentiation. This paper overviews about procedures and applications of tissue culture by the analysis of numerous published studies. Compared to normal plant breeding methods, which take substantially longer, crops produced through tissue culture is developed through accurate and time-saving methods. It enables the recovery of embryos created by incompatible crosses, eliminates the phenomena of seed dormancy seen in plant species, and the life cycle of some species whose life cycles are relatively long are shorten. Even in situations where the species would typically have poor rates of multiplication, tissue culture enables the quick generation of a large number of plants. Also tissue culture required less space for multiplication of clones. An agriculturally valuable phenotype should come from the genetic diversity recovered from tissue culture regenerated plants. There are numerous issues arises in tissue culture. Researchers are having difficulty propagating plant tissues and acclimating in vitro produced plants in their natural habitat. However, greater understanding and study will lead to more widespread use of tissue culture.

Inheritance of Unilateral Incompatibility in the Genus Capsicum

Unilateral incompatibility is reported to occur between the species of Capsicum pubescens complex (Capsicum cardenasii Capsicum. eximium and Capsicum pubescens) and other species of genus Capsicum when the latter are used as the male parent. The relationship between self-incompatibility and unilateral incompatibility are yet to be resolved even in the most intensively investigated crosses within and between species of Lycopersicon and Solanum. Unilateral incompatibility in Capsicum has been less intensively studied compared to some other genus in family Solanaceae. This study was, therefore, conducted to help us to understand the genetic control of unilateral incompatibility in Capsicum. Experiment results showed that while the pistil behaviour of F1 hybrids agrees with the pistil behaviour of F1 hybrids from unilaterally incompatible crosses in other genera of Solanaceae, the pollen behaviour does not agree with data obtained from other genera of Solanaceae. No segregation ratio was obtained for unilateral incompatibility in the backcross progenies, this result does not agree with data from other genera in the Solanaceae either. Possible reasons for not obtaining segregation for unilateral incompatibility may be the small size of backcross progenies and/or distorted segregation ratios. As a conclusion it seemed more probable that, in Capsicum, unilateral incompatibility has arisen as a by-product of genetic divergence between the C. pubescens complex and the other chile peppers, not as a product of natural selection.

Exploration of incompatible crosses in plants for novel and useful variations

AbstractCrop improvement through pre‐breeding utilizes elite lines, germplasm and species of compatible gene pools, but crossing and characterizing the derivatives between incompatible species, especially distant genera or species, is realized to introduce de novo variations that are heritable, stable and especially economically contributing. This is a novel approach to introduce large‐scale de novo variations in crops that can be applied to widening of the genetic base. It helps to introduce heritable and agronomical variations that can be evolved in a short time for use in trait specific breeding programmes. In crosses between incompatible species many times F1 are delusive appearing like maternal parent. But large variations in F2 are observed due to genetic changes that include partial or complete elimination of the genome from the paternal parent, reshuffling of the maternal genome leading to several changes that are translated to phenotype. Genome, transcriptome and epigenetic reprogramming is responsible for the above genetic changes. Crossing two distant species can be used as an additional tool in plant breeding.

Three types of genes underlying the Gametophyte factor1 locus cause unilateral cross incompatibility in maize

Unilateral cross incompatibility (UCI) occurs between popcorn and dent corn, and represents a critical step towards speciation. It has been reported that ZmGa1P, encoding a pectin methylesterase (PME), is a male determinant of the Ga1 locus. However, the female determinant and the genetic relationship between male and female determinants at this locus are unclear. Here, we report three different types, a total of seven linked genes underlying the Ga1 locus, which control UCI phenotype by independently affecting pollen tube growth in both antagonistic and synergistic manners. These include five pollen-expressed PME genes (ZmGa1Ps-m), a silk-expressed PME gene (ZmPME3), and another silk-expressed gene (ZmPRP3), encoding a pathogenesis-related (PR) proteins. ZmGa1Ps-m confer pollen compatibility. Presence of ZmPME3 causes silk to reject incompatible pollen. ZmPRP3 promotes incompatibility pollen tube growth and thereby breaks the blocking effect of ZmPME3. In addition, evolutionary genomics analyses suggest that the divergence of the Ga1 locus existed before maize domestication and continued during breeding improvement. The knowledge gained here deepen our understanding of the complex regulation of cross incompatibility.

Population Dynamics of Wide Compatibility System and Evaluation of Intersubspecific Hybrids by indica-japonica Hybridization in Rice.

The exploitation of heterosis through intersubspecific hybridisation between indica and japonica has been a major breeding target in rice, but is marred by the cross incompatibility between the genomes. Wide compatibility (WC) is a triallelic system at the S5 locus on chromosome 6 that ensures the specificity of hybridisation within and between indica and japonica. The S5n allele that favours intercrossing is sparsely distributed in the rice gene pool and therefore warrants identification of diverse WC sources to develop superior intersubspecific hybrids. In this study, we have identified several novel WC sources through the marker-assisted screening of a large set of 950 rice genotypes. Seventeen percent of the genotypes carried S5n, which fell into two subpopulations. The WC genotypes showed wide phenotypic and genotypic variability, including both indica and japonica lines. Based on phenotypic performance, the WC varieties were grouped into three clusters. A subset of 41 WC varieties was used to develop 164 hybrids, of which WC/japonica hybrids showed relative superiority over WC/indica hybrids. The multilocation evaluation of hybrids indicated that hybrids derived from WC varieties, such as IRG137, IRG143, OYR128, and IRGC10658, were higher yielding across all the three different locations. Most of the hybrids showed the stability of performance across locations. The identified diverse set of wide compatible varieties (WCVs) can be used in the development of intersubspecific hybrids and also for parental line development in hybrid rice breeding.

Interacting host modifier systems control Wolbachia-induced cytoplasmic incompatibility in a haplodiploid mite.

Reproductive parasites such as Wolbachia spread within host populations by inducing cytoplasmic incompatibility (CI). CI occurs when parasite‐modified sperm fertilizes uninfected eggs and is typified by great variation in strength across biological systems. In haplodiploid hosts, CI has different phenotypic outcomes depending on whether the fertilized eggs die or develop into males. Genetic conflict theories predict the evolution of host modulation of CI, which in turn influences the stability of reproductive parasitism. However, despite the ubiquity of CI‐inducing parasites in nature, there is scarce evidence for intraspecific host modulation of CI strength and phenotype. Here, we tested for intraspecific host modulation of Wolbachia‐induced CI in haplodiploid Tetranychus urticae mites. Using a single CI‐inducing Wolbachia variant and mitochondrion, a nuclear panel was created that consisted of infected and cured near‐isogenic lines. We performed a highly replicated age‐synchronized full diallel cross composed of incompatible and compatible control crosses. We uncovered host modifier systems that cause striking variation in CI strength when carried by infected T. urticae males. We observed a continuum of CI phenotypes in our crosses and identified strong intraspecific female modulation of the CI phenotype. Crosses established a recessive genetic basis for the maternal effect and were consistent with polygenic Mendelian inheritance. Both male and female modulation interacted with the genotype of the mating partner. Our findings identify spermatogenesis as an important target of selection for host modulation of CI strength and underscore the importance of maternal genetic effects for the CI phenotype. Our findings reveal that intraspecific host modulation of CI is underpinned by complex genetic architectures and confirm that the evolution of reproductive parasitism is contingent on host genetics.

Proteomic Analysis of Pollen–Stigma Interaction between Vitis rotundifolia and Vitis vinifera

Muscadine grape (Vitis rotundifolia) is highly resistant to many diseases and insects that attack european grape (Vitis vinifera). However, distant hybridization incompatibility between V. rotundifolia (female) and V. vinifera (male) impedes the utilization of V. rotundifolia in grape breeding. This study used fourth-dimension label-free protein quantitation to detect the key genes and pathways in the V. rotundifolia stigma after self-pollination (V. rotundifolia × V. rotundifolia) and cross-pollination (V. rotundifolia × V. vinifera). A histological analysis showed that pollen tube growth in the stigma of V. rotundifolia was arrested 8 hours after cross-pollination, but not after self-pollination. A proteomic analysis identified 32 differentially expressed proteins (DEPs) in the stigma of V. rotundifolia between self-pollination and cross-pollination. A heatmap analysis grouped these DEPs into four clusters. The top gene ontology terms were ATPase-coupled transmembrane transporter activity, extracellular region, DNA replication, and cellular carbohydrate biosynthetic process. A Kyoto Encyclopedia of Genes and Genomes analysis revealed that these DEPs participated in DNA replication and starch and sucrose metabolism pathways. The downregulated A5AY88, D7TJ35, D7SU26, F6HJI1, and F6GUE7 may have a role in cross incompatibility. This study revealed the cross incompatibility of grapes at histological and proteomic levels.

Integrating Genome-Wide Association Study with Transcriptomic Analysis to Predict Candidate Genes Controlling Storage Root Flesh Color in Sweet Potato

Sweet potato is a hexaploid heterozygote with a complex genetic background, self-pollination infertility, and cross incompatibility, which makes genetic linkage analysis quite difficult. Genome-wide association studies (GWAS) provide a new strategy for gene mapping and cloning in sweet potato. Storage root flesh color (SRFC) is an important sensory evaluation, which correlates with storage root flesh composition, such as starch, anthocyanin, and carotenoid. We performed GWAS using SRFC data of 300 accessions and 567,828 single nucleotide polymorphism (SNP) markers. Furthermore, we analyzed transcriptome data of different SRFC varieties, and conducted real-time quantitative PCR (qRT-PCR) to measure the expression level of the candidate gene in purple and non-purple fleshed sweet potato genotypes. The results showed that five unique SNPs were significantly (−log10P > 7) associated with SRFC. Based on these trait-associated SNPs, four candidate genes, g55964 (IbF3′H), g17506 (IbBAG2-like), g25206 (IbUGT-73D1-like), and g58377 (IbVQ25-isoform X2) were identified. Expression profiles derived from transcriptome data and qRT-PCR analyses showed that the expression of g55964 in purple-fleshed sweet potato was significantly (p < 0.01) higher than that of non-purple fleshed sweet potato. By combining the GWAS, transcriptomic analysis and qRT-PCR, we inferred that g55964 is the key gene related to purple formation of storage root in sweet potato. Our results lay the foundation for accelerating sweet potato genetic improvement of anthocyanin through marker-assisted selection.

Red Cell Antibodies Against High Frequency Antigens, A Two Year Experience At A Regional Transfusion Institute

Objective: To assess the clinicohaematological presentation and outcome of patients with antibodies to high frequency antigen(HFA) in our setup.Study Design: Cross sectional study.
 Place and Duration of Study: Armed Forces Institute of Transfusion, Rawalpindi, from Jan 2016 to Jan 2018.
 Methodology: We diagnosed 29 cases of uncommon red cell antibodies including antibodies to HFA, in a span of two years (2016-2018). Information regarding patient’s age, diagnosis, transfusion history, incompatible cross match and history of pregnancy was obtained. Venous blood sample of 3ml was taken in EDTA for grouping, red cell phenotyping and Direct AntiGlobulin Test (DAT) and 5 ml venous blood was taken for Indirect Antiglobulin Test (IAT), antibody screening and identification. Tube method was used for forward and reverse blood grouping, DAT and IAT. Column Agglutination Technique (CAT) was used for antibody screening, identification and red cell phenotyping.
 Results: Out of a total of 29 cases, 24 (83%) were of anti H antibody (Bombay blood group). Rest of 5 (17%) included one each of anti Rh 17, anti Rh 29 (Rh null), anti P1Pk, anti-ENA/WRB and anti P antibodies. Anti H Lectin (Ulexeuropaeus) was used for confirmation of H antigen absence, while rest of antibodies were confirmed by IBGRL Bristol, UK.
 Conclusion: The sources of antigen negative blood are family members, rare donor registry, autologous donations and frozen blood banks. Finding compatible blood for patients with an antibody to HFA may be a challenge.

Investigating the Mechanism of Unilateral Cross Incompatibility in Longan (Dimocarpus longan Lour.) Cultivars (Yiduo × Shixia).

Longan (Dimocarpus longan Lour.) is an important subtropical fruit tree in China. Nearly 90% of longan fruit imports from Thailand are from the cultivar Yiduo. However, we have observed that there exists a unilateral cross incompatibility (UCI) when Yiduo is used as a female parent and Shixia (a famous Chinese cultivar) as a male parent. Here, we performed a comparative transcriptome analysis coupled with microscopy of pistils from two reciprocal pollination combinations [Shixia♂ × Yiduo♀(SY) and Yiduo♀ × Shixia♂(YS)] 4, 8, 12, and 24 h after pollination. We also explored endogenous jasmonic acid (JA) and jasmonyl isoleucine (JA-Ile) levels in pistils of the crosses. The microscopic observations showed that the UCI was sporophytic. The endogenous JA and JA-Ile levels were higher in YS than in SY at the studied time points. We found 7,251 differentially expressed genes from the transcriptome analysis. Our results highlighted that genes associated with JA biosynthesis and signaling, pollen tube growth, cell wall modification, starch and sucrose biosynthesis, and protein processing in endoplasmic reticulum pathways were differentially regulated between SY and YS. We discussed transcriptomic changes in the above-mentioned pathways regarding the observed microscopic and/or endogenous hormone levels. This is the first report on the elaboration of transcriptomic changes in longan reciprocal pollination combination showing UCI. The results presented here will enable the longan breeding community to better understand the mechanisms of UCI.