[A comparative genetic and cytogenetic mapping of wheat chromosome 5B using introgression lines].

The genetic map of chromosome 5B has been constructed by using microsatellite (SSR) analysis of 381 plants from the F2 population produced by cross of the Chinese Spring (CS) and Renan cultivars. Initially, 180 SSR markers for the common wheat 5B chromosome have been used for analysis of these cultivars. The 32 markers able to detect polymorphism between these cultivars have been located on the genetic map of chromosome 5B. Cytogenetic mapping has involved a set of CS 5B chromosome deletion lines. Totally, 51 SSR markers have been located in ten regions (deletion bins) of this chromosome by SSR analysis of these deletion lines. Five genes--TaCBFIIIc-B10, Vrn--B1, Chi--B1, Skr, and Ph1--have been integrated into the cytogenetic map of chromosome 5B using the markers either specific of or tightly linked to the genes in question. Com- parison of the genetic and cytogenetic maps suggests that recombination is suppressed in the pericentromeric region of chromosome 5B, especially in the short arm segment. The 18 markers localized to deletion bins 5BL16-0.79-1.00 and 5BL8-0.66-0.79 have been used to analyze common wheat introgression lines L842, L5366-180, L73/00i, and L21-4, carrying fragments of alien genomes in the terminal region of 5B long arm. L5366-180 and L842 lines carry a fragment of the Triticum timopheevii 5GL chromosome, while L73/00i ? L21-4 lines, a fragment of the Aegilopsspeltoides 5SL chromosome. As has been shown, the translocated fragments in these four lines are of different lengths, allowing bin 5BL18-0.66-0.79 to be divided into three shorter regions. The utility of wheat introgression lines carrying alien translocations for increasing the resolution of cytogenetic mapping is discussed.

Microsatellite mapping of Ae. speltoides and map-based comparative analysis of the S, G, and B genomes of Triticeae species.

The first microsatellite linkage map of Ae. speltoides Tausch (2n = 2x = 14, SS), which is a wild species with a genome closely related to the B and G genomes of polyploid wheats, was developed based on two F(2) mapping populations using microsatellite (SSR) markers from Ae. speltoides, wheat genomic SSRs (g-SSRs) and EST-derived SSRs. A total of 144 different microsatellite loci were mapped in the Ae. speltoides genome. The transferability of the SSRs markers between the related S, B, and G genomes allowed possible integration of new markers into the T. timopheevii G genome chromosomal maps and map-based comparisons. Thirty-one new microsatellite loci assigned to the genetic framework of the T. timopheevii G genome maps were composed of wheat g-SSR (genomic SSR) markers. Most of the used Ae. speltoides SSRs were mapped onto chromosomes of the G genome supporting a close relationship between the G and S genomes. Comparative microsatellite mapping of the S, B, and G genomes demonstrated colinearity between the chromosomes within homoeologous groups, except for intergenomic T6A(t)S.1G, T4AL.5AL.7BS translocations. A translocation between chromosomes 2 and 6 that is present in the T. aestivum B genome was found in neither Ae. speltoides nor in T. timopheevii. Although the marker order was generally conserved among the B, S, and G genomes, the total length of the Ae. speltoides chromosomal maps and the genetic distances between homoeologous loci located in the proximal regions of the S genome chromosomes were reduced compared with the B, and G genome chromosomes.

Genetic diversity and linkage disequilibrium studies on a 3.1-Mb genomic region of chromosome 3B in European and Asian bread wheat (Triticum aestivum L.) populations.

Genetic diversity and linkage disequilibrium (LD) were investigated in 376 Asian and European accessions of bread wheat (Triticum aestivum L.). After a first and rapid screening about diversity and genetic structure at the whole genome scale using 70 simple sequence repeats (SSRs), we focused on a sequenced contig (ctg954) of 3.1 Mb located on the short arm of chromosome 3B of cv. Chinese Spring, using 32 SSRs and 10 single nucleotide polymorphisms. This contig is part of a multiple fungal resistance region. Mean polymorphism information content value on the 32 SSRs was slightly higher in the Asian genepool (0.396) than that for the European (0.329) pool. Compared with results at the whole genome scale, data from this 3.1-Mb region indicated similar trends in genetic diversity indices between both genepools. Population structure and molecular variance analyses demonstrated significant genetic differentiation and geographical subdivision in both groups of accessions. Concerning LD at the contig level, the European population had a significantly higher mean r(2) value (0.23) than the Asian population (0.18), indicating a stronger LD in the European material. With a mean of 1 marker every 74 kb, the resolution reached here allowed to perform a detailed comparative analysis of the LD and genetic diversity along the complete 3.1-Mb region in both genepools. A sliding-window approach revealed some interesting regions of the contig where LD is increasing when genetic diversity is decreasing. This study provides an in-depth understanding of molecular population genetics in European and Asian wheat gene pools, and prospects for association mapping of important sources of fungal disease resistance.

[Chromosome synteny of the A genome of two evolutionary wheat lines].

In order to estimate synteny between A(t) and A polyploid wheat genomes belonging to different evolutionary lines (Timopheevi and Emmer), saturation of chromosome maps of Triticum timopheevii A(t) genome by molecular markers has been conducted. Totally, 179 EST-SSR and 48 genomic SSR-markers have been used with the following integration of 13 and 7 markers correspondingly into chromosome maps of A(t) genome. EST-SSR showed higher transferability and lower polymorphism than genomic SSR markers. The chromosome maps designed were compared to maps of homoeologous chromosome group of the T. aestivum A genome. No disturbances of colinearity, i.e., of the order of markers within the chromosome segments on which they had been previously mapped, were observed. According to the quantity assessment of markers amplifying in homoeologous chromosomes, the maximum divergence was detected in two groups (4A(t)/4A and 3A(t)/3A) among the seven chromosomes examined in the A(t) and A genomes. Comparison of molecular genetic mapping results with the published results of studying meiosis of F1 hybrids and the frequency of chromosomes substitution in introgressive T. aestivum x T. timopheevii lines suggest that individual chromosomes of the At and A genomes evolve differently. Translocations were shown to introduce the major impact on the divergence of 4A(t)/4A and 6A(t)/6A chromosomes, while mutations of the primary DNA structure, on the divergence of homoeologous group 3 chromosomes. The level of reorganization of other chromosomes during the evolution in the A(t) and A genomes was significantly lower.

Evaluation of the genetic variability of homoeologous group 3 SSRs in bread wheat.

Thorough characterization of the genetic variability in bread wheat (Triticum aestivum L.) is important for a better improvement of this key crop and to increase cereal yield in the context of sustainable agriculture to face human needs in the next decades. To study the genetic variability of SSRs on wheat homoeologous group 3 chromosomes, we characterized 38 hexaploid and two tetraploid wheat lines using a set of 165 microsatellites that we cytogenetically assigned to the 17 deletion bins for chromosomes group 3. A comparative analysis of the genetic variability through the PIC value study, allele numbers and SSR lengths indicated that there were no statistically significant differences (p > 0.05) between the three chromosomes of this homoeologous group despite the fact that SSRs from chromosome 3B exhibited slightly more alleles per locus compared to chromosomes 3A and 3D as well as slightly higher PIC values compared to chromosome 3D. However, there was a stronger correlation between SSR length and allele number on the short arms compared to the long arms and the correlation increased from the centromeres toward the telomeres. We did not find statistically significant differences in allele numbers and PIC values for SSRs located in more distal bins on 3A and 3B chromosomes. On the contrary, for chromosome 3D, we observed significant differences (p < 0.05) between the PIC values determined for SSRs assigned to deletion bin 3DL3-0.81-1.00 bin that is located distal compare to the more proximal region (C-3DL3-0.81). These results suggest that recombination which is higher in the telomeric regions does not contribute to increase a lot the variability of the SSRs.

Quantitative trait Loci mapping for adult-plant resistance to powdery mildew in bread wheat.

ABSTRACT Powdery mildew, caused by Blumeria graminis f. sp. tritici, is a major disease to wheat (Triticum aestivum) worldwide. Use of adult-plant resistance (APR) is an effective method to develop wheat cultivars with durable resistance to powdery mildew. In the present study, 432 molecular markers were used to map quantitative trait loci (QTL) for APR to powdery mildew in a doubled haploid (DH) population with 107 lines derived from the cross Fukuho-komugi x Oligoculm. Field trials were conducted in Beijing and Anyang, China during 2003-2004 and 2004-2005 cropping seasons, respectively. The DH lines were planted in a randomized complete block design with three replicates. Artificial inoculation was carried out in Beijing with highly virulent isolate E20 of B. graminis f. sp. tritici and the powdery mildew severity on penultimate leaf was evaluated four times, and the maximum disease severity (MDS) on penultimate leaf was investigated in Anyang under natural inoculation in May 2004 and 2005. The heritability of resistance to powdery mildew for MDS in 2 years and two locations ranged from 0.82 to 0.93, while the heritability for area under the disease progress curve was between 0.84 and 0.91. With the method of composite interval mapping, four QTL for APR to powdery mildew were detected on chromosomes 1AS, 2BL, 4BL, and 7DS, explaining 5.7 to 26.6% of the phenotypic variance. Three QTL on chromosomes 1AS, 2BL, and 7DS were derived from the female, Fukuho-komugi, while the one on chromosome 4BL was from the male, Oligoculm. The QTL on chromosome 1AS showed high genetic effect on powdery mildew resistance, accounting for 19.5 to 26.6% of phenotypic variance across two environments. The QTL on 7DS associated with the locus Lr34/Yr18, flanked by microsatellite Xgwm295.1 and Ltn (leaf tip necrosis). These results will benefit for improving powdery mildew resistance in wheat breeding programs.

Studies of the transferability of microsatellites derived from Triticum tauschii to hexaploid wheat and to diploid related species using amplification, hybridization and sequence comparisons.

Hexaploid wheat ( Triticum aestivum L em Thell) is derived from a complex hybridization procedure involving three diploid species carrying the A, B and D genomes, respectively. We recently isolated microsatellites from a T. tauschii library enriched for various motifs and evaluated the transferability of these markers to several diploid species carrying the A, B or D genomes. All of the primer pairs amplifying more than one locus on bread wheat and half of those giving D-genome-specific loci gave an amplification product on A-and/or B-diploid species. All of the markers giving a single amplification product for T. tauschii and no amplification on the other diploid species were D-genome-specific at the hexaploid level. The non-specific microsatellite markers (which gave an amplification product on diploid species carrying the A, B or D genome) gave either a complex amplification pattern on bread wheat (with several bands) or generated a single band which mapped to the D genome. Southern blot hybridizations with probes corresponding to the microsatellite flanking regions gave a signal on all diploid and hexaploid species, whatever the specificity of the microsatellite. The patterns observed on bread wheat were generally in accordance with those observed for diploid species, with slight rearrangements. This suggests that the specificity of microsatellite markers is probably due to mutations in microsatellite flanking regions rather than sequence elimination during polyploidization events and that genome stringency is higher at the polyploid than at the diploid level.

Characterisation of polymorphic microsatellite markers from Aegilops tauschii and transferability to the D-genome of bread wheat.

Microsatellites were isolated from a Aegilops tauschii (the D-genome donor of bread wheat) library enriched for various motifs. Primers generated from the flanking region of the microsatellites were used successfully to amplify the corresponding loci in the D genome of bread wheat. Additional amplification sometimes also occurred from the A and B genomes. The majority of the microsatellites contained (GA)(n) and (GT)(n) motifs. GA and GT repeats appeared to be both more abundant in this library and more polymorphic than other types of repeats. The allele number for both types of dinucleotide repeats fitted a Poisson distribution. Deviance analysis showed that GA and GT were more polymorphic than other motifs in bread wheat. Within each motif type (di-, tri- and tetra-nucleotide repeats), repeat number has no influence on polymorphism. The microsatellites were mapped using the Triticum aestivum Courtot x Chinese Spring mapping population. A total of 100 markers was developed on this intraspecific map, mainly on the D genome. For polyploid species, isolation of microsatellites from an ancestral diploid donor seems to be an efficient way of developing markers for the corresponding genome in the polyploid plant.

Comparative mapping of the wheat 5B short chromosome arm distal region with rice, relative to a crossability locus.

Colinearity between wheat and rice genomes is quite well established at the chromosome level, but less is known at a finer level. We tried to specify these relationships for the wheat 5BS chromosome-arm distal region, where a major locus for crossability was located. By developing AFLP markers, we succeeded to locate this major QTL more precisely. One cloned AFLP fragment mapped to rice chromosome 11, which was in agreement with a rice chromosome-11 linkage block reported in this region. However a second marker, a RFLP probe, showed a break in synteny because it mapped to rice long-arm chromosomes 1 and 5, while screening a rice BAC library with the same probe identified rice chromosomes 5 and 6. Therefore, we concluded that the syntenic relationships were more complex at the fine level. The observed results might indicate the presence of a linkage block carrying a crossability gene on wheat groups 1, 5 and 7, and also on rice chromosomes 5 and 6.

Confocal microscopic examination of trabecular meshwork removed during ab externo trabeculectomy.

AIMS: The aim of the ab externo trabeculectomy (AET) is to remove the external portion of the trabecular meshwork (ETM) responsible for the main aqueous outflow resistance in glaucoma patients, with no opening of the anterior chamber. ETM characteristics were evaluated with a confocal microscope. METHODS: A prospective comparative observational case series was performed in 60 consecutive medically treated patients with primary open angle glaucoma and eight postmortem normal donors' eyes that underwent AET. Once deroofing the Schlemm' s canal (SC), a deeper dissection led to removal of a coherent membrane (ETM) which allowed satisfactory aqueous egress through the remaining intact internal trabecular meshwork (TM) layers. After fixation with acetone and immunostaining with anti-vimentin antibody, ETM were analysed with a confocal microscope. RESULTS: Glaucomatous ETM (mean thickness: 29.5 (7.6) micro m) were characterised by a severe paucicellularity compared with the controls (respectively 37.3 (9.7) cells/area and 167.5 (24.9) cells/area, p<10(-4)). ETM analysis showed involvement of both cribriform and corneoscleral layers. ETM cell density was significantly decreased in case of preoperative fluorometholone instillation. CONCLUSION: Paucicellularity of glaucomatous TM is confirmed by this original technique. Structural characteristics of the ETM, whose removal allows satisfactory aqueous egress, suggest that aqueous outflow resistance not only involves inner wall of SC and juxtacanalicular meshwork but also corneoscleral trabecular layers.

Optical coherence tomography of macular thickness after cataract surgery.

PURPOSE: To evaluate macular thickness changes after uneventful cataract surgery using optical coherence tomography (OCT) and compare the findings with those of flare and cell measurements of the anterior chamber. SETTING: Clinique Sourdille, Nantes, France. METHODS: In this retrospective study, 41 eyes having uneventful cataract surgery with a clear corneal small incision and intracapsular fixation of a foldable intraocular lens were evaluated by OCT and laser flare and cell measurements preoperatively and 1, 8 to 12, and 30 to 60 days postoperatively. RESULTS: Some postoperative increase in macular thickness was noted in 11 eyes. This was not related to a higher postoperative flare. Visual consequences were proportional to the macular elevation. CONCLUSION: Clinical and subclinical thickness changes, without breakdown of the blood-aqueous barrier, can be detected after cataract surgery. Most of these changes resolve spontaneously, but their mid- and long-term significance is unknown.

Reticulated hyaluronic acid implant in nonperforating trabecular surgery.

PURPOSE: To evaluate experimentally and clinically the tolerance and efficacy of a reticulated hyaluronic acid implant in nonperforating trabecular surgery (NPTS). SETTING: Bascom Palmer Eye Institute, Miami, Florida, USA, and Clinique Sourdille and Clinique Ophtalmologique Universitaire, Nantes, France. METHODS: In experimental surgery, NPTS was performed with and without a hyaluronic acid implant in 25 rabbit eyes. In a pilot study, the results of NPTS with a hyaluronic acid implant in 72 human eyes were retrospectively analyzed in terms of visual acuity, intraocular pressure (IOP), external filtration, postoperative inflammation, and gonioscopy. Mean follow-up was 13.8 months (range 6 to 24 months). RESULTS: In the experimental surgery, the rabbit eyes with the implant showed a different healing process than the eyes without the implant. The implant was slowly bioabsorbed and remnants were seen at the operative site (where the tissue was removed) up to day 56 postoperatively. This site was detectable at all histology study periods. Intraocular pressure reduction was longer in the implant group: greater than 5 months versus 3 weeks (P < .05). In the pilot study, visual acuity remained stable, IOP decreased from a mean preoperative level of 26.3 mm Hg +/- 5.22 (SD) to a mean postoperative level without treatment of 15.4 +/- 3.1 mm Hg (P < .0001). No external filtration was detected in 60 eyes, a slightly elevated conjunctiva was noted in 12 eyes. Postoperative inflammation (laser flare and cell measurements) was low. Gonioscopy consistently demonstrated the persistence of a decompression space behind the trabeculum. CONCLUSION: Comparative experimental surgery results showed excellent tolerance and efficacy in the rabbit eyes with a hyaluronic acid implant. Clinical results, to be confirmed by a randomized comparative study, also showed excellent biocompatibility and encouraging efficacy.

[Lamellar graft in the treatment of herpes simplex ocular (author's transl)]. / La kératoplastie lamellaire dans le traitement de l'herpès oculaire.

72 lamellar grafts for herpes simplex ocular infections have been performed. A retrospective study tries to specify the indications. Penetrating graft is the best for leucomae. The most difficult choice occurs with active herpetic disease. With lamellar grafts good results are obtained in stromal keratitis without deep involvement (visual acuity is 0.2 or more in 75 per cent of cases). When endothelio-uveal involvement, prognosis becomes uncertain in any type of keratoplasty: the authors think that the best method is to perform surgery after a few months of inactive disease.

[Optic disc nodules with haemorrhage (author's transl)]. / Druses de la papille avec hémorragies.

A thirteen year old boy has shown optic disc drusens with haemorrhages in his left eye. Then, his right eye was completely all right. But six years later, the same symptomatology can be seen in this one.

[Penetrating keratoplasties (author's transl)]. / Kératoplastie perforante et cristallin artificiel incidence sur la population cellulaire endothéliale.

The authors relate the specular microscopy study of 19 penetrating keratoplasties. Seven of them were associated with an intra-ocular lens. The mean cell density is 1 100 cells/SQ. MM. The endothelial cell density does not differ in the two series. One graft remains clear with 330 cells/SQ. MM.

[Evaluation of confocal microscopy in the analysis of the external trabecular membrane during deep nonpenetrating sclerectomy]. / Apport de la microscopie confocale dans l'analyse de la membrane trabéculaire prélevée au cours de la chirurgie filtrante non perforante.

PURPOSE: Deep non penetrating sclerectomy (DNPS) is a new filtering surgical procedure whose aim is a selective ablation of the external portion of the trabecular meshwork (TM) which is involved in the aqueous outflow resistance, i.e. the inner wall of Schlemm's canal (SC) and the external trabecular layers, especially the cribriform TM. We evaluated, with a confocal microscope, the structural characteristics of this part of the TM in glaucomatous patients. METHODS: Thirty-six external trabecular membranes (ETM) were obtained from 33 consecutive glaucomatous patients (mean age: 56.5+/-14.5 years) and from four post-mortem normal donors (60.5 +/-7.7 years), which underwent DNPS according to the same surgical procedure. Under conjunctival and scleral flaps, the roof of the SC was opened and removed. A deeper dissection led to the removal of the inner wall of the SC and the adjacent ETM, i.e the TEM, which allowed a satisfactory aqueous flow through the remaining internal TM layers. After fixation with acetone or triton X100 and immunostaining with anti-fibronectin or vimentin antibodies, the samples were analyzed with a confocal microscope (Nikon EZ 2000). RESULTS: The mean thickness of the ETM was 34.4+/-7.3 microm in glaucomatous eyes, not significantly different from the controls (39.0+/-10.7 microm). The main characteristic of the glaucomatous ETM membrane is a paucicellularity as compared to the controls (respectively 21.6+/-12.1 cells/area and 156.1+/-28.8 cells/area). The confocal microscope analysis shows that the ETM involves two different portions of the trabecular meshwork. The architectural characteristics of the outermost portion of the ETM with its star-shaped cells arranged in a homogenous extra-cellular matrix, suggest that it is the trabecular cribriform meshwork. The inner portion of the ETM shows cells arranged regularly in a fibrillar extra-cellular matrix as described in the corneoscleral trabecular meshwork. CONCLUSION: These results confirm the previous reported histopathological changes of the glaucomatous TM. The thickness of the removed ETM and its structural characteristics show that the ablation of the TM layers necessary to obtain a satisfactory aqueous filtration, not only involves the cribriform layers but also one of the inner parts of the TM, i.e. the corneoscleral TM. This result suggests that aqueous humor resistance not only involves the cribriform trabecular meshwork but also a part of the corneoscleral trabecular meshwork.

[Eye controlled microscope used in ophthalmic microsurgery]. / Commande du microscope par le regard appliquée à la microchirurgie ophtalmologique.

This article presents a system of oculomotor control of the microscope used in ophthalmic microsurgery. An eye movement sensor has been integrated within the microscope lens. The information provided by this sensor is used to control the position of the microscope with no alteration of the surgeon's visual functions and no interference with his visual sensori-motor reflexes. Results have been obtained from laboratory experimentations as well as from a series of anterior and posterior segment surgical procedures. They demonstrate the high precision and sensitivity of the gaze control which can be used to compensate for the reduction of the field of vision resulting from the optical magnification of the microscope.

[Fate of artificial lenses: a scanning electron microscopy study]. / Devenir des cristallins artificiels: étude en microscopie électronique à balayage.

Thirty one intraocular lenses removed from human eyes 2 months to 30 years after implantation were examined by scanning electron microscopy. Binkhorst 4 loop lenses (12) and 2 loop lenses (6), and Choyce lenses (6) were the most prevalent styles examined. Most of the lenses produced before 1978 were found to have manufacturing defects (mold disalignment, unfinished gate areas, etc.). More recent lenses showed an improvement in the quality of the surface. The lenticular, nonimpinging portion of the lenses made with polymethylmethacrylate did not demonstrate signs of degradation. However the holes in the edges of the lenses were rough and poorly finished with obvious irregularities. On the other hand, the lens foot of a Choyce lens revealed severe unusual abnormalities: the plastic surface changes may be consistent with degradation. Examination of explanted iris clip lenses showed degradation of the polypropylene loops in two cases: the loops were found to have fractured surfaces.