Sr31-Virulent Races (TTKSK, TTKST, and TTTSK) of the Wheat Stem Rust Pathogen Puccinia graminis f. sp. tritici are Present in Tanzania.

Since the first detection of race TTKSK (syn. Ug99) in Uganda in 1999 (2), the migration and evolution of Sr31-virulent races of Puccinia graminis f. sp. tritici [Pgt] have been closely monitored, particularly in Kenya and countries north, along the likely trajectory of migration to major wheat-producing regions of Asia. More recently, surveillance efforts have been undertaken to the south as well, and Ug99-related races have been detected in South Africa and Zimbabwe (3,4). Here we report for the first time results of a survey conducted in Tanzania. Systematic race surveillance provides data not only on the current distribution of the Ug99 race group, but also on the possible points of origin as well as the pace and probable paths of dispersal of future races from the region. In this context, the presence or absence of the Ug99 group of wheat stem rust races in adjacent countries like Tanzania assumes regional, and possibly global, relevance. A preliminary survey conducted in September 2006 indicated the presence of Sr31-virulent races of Pgt outside Slahhamo Village (3°15'S, 35°48'E) in the Ngorongoro highlands of northern Tanzania, based on compatible reactions with cv. K-Mamba (a.k.a. Mwamba), a cultivar whose pedigree indicates the presence of Sr31. A broader survey was conducted in August 2009, during which infected tissue was collected from currently-grown cultivars in research plots and on large estates, as well as from the mixes of older cultivars common on smallholder farms. In all, Pgt-infected samples were collected from one site in the Arusha region [Monduli (3°16' S, 36°24'E)], three sites in the Ngorongoro highlands [Karatu (3°20' S, 35°40' E), Upper Kitete (3°14' S, 35°53' E), and Slahhamo], one site in the Manyara region [Hanang (4°43' S, 35°40' E)], and one site in the Mbeya region [southern highlands (8°87' S, 33°40' E)], thereby giving representation to all four major wheat growing areas in the country. Sample storage, inoculation, incubation, disease assessment, and derivation of single-pustule cultures were all performed according to the methods described by Jin et al. (1). In addition to the 20 differentials in the expanded Pgt differential set of North America, we included two supplemental tester lines: Siouxland (Sr24 + Sr31) and Sisson (Sr31 + Sr36). Each single-pustule-derived isolate was evaluated for virulence on the differential and supplemental lines at least twice. A total of 39 single-pustule isolates were derived from the six collection sites. All 39 isolates were identified as belonging to the Ug99 race group, with six identified as TTKSK (all four regions), 30 identified as TTKST (Sr31 + Sr24 virulence; Arusha region and the Ngorongoro highlands), and three identified as TTTSK (Sr31 + Sr36 virulence; Manyara region and the Ngorongoro highlands). The results of this study suggest that, to more precisely locate the "hot spots" and thereby gain a better understanding of the mechanisms of novel race emergence in East Africa, it would be prudent to include Tanzania, heretofore a blank area on the wheat rust surveillance map, in future systematic race monitoring efforts. References: (1) Y. Jin et al. Plant Dis. 92:923, 2008. (2) Z. A. Pretorius et al. Plant Dis. 84:203, 2000. (3) Z. A. Pretorius et al. Plant Dis. 94:784, 2010. (4) Z. A. Pretorius et al. Plant Dis. 96:590, 2012.

Registration of wheat lines carrying the partial stripe rust resistance gene Yr36 without the Gpc-B1 high grain protein content allele.

While the high-temperature adult plant resistance gene Yr36 represents a promising source of quantitative and potentially race non-specific resistance to wheat stripe rust (causal organism Puccinia striiformis Westend. f. sp. tritici), its tight linkage (0.3 cM) with the high-grain protein content gene Gpc-B1 may hinder its introgression in certain cases, such as in soft wheat varieties requiring low grain protein content or in lines where the Gpc-B1 allele may be associated with a yield penalty. The development and registration of two donor lines, one tetraploid (Triticum turgidum L. ssp. durum; PI 656793) and one hexaploid (T. aestivum L. ssp. aestivum; PI 664549), each carrying the resistant wild emmer (T. turgidum ssp. dicoccoides) allele for Yr36 linked with the non-functional Gpc-B1 allele, are intended to overcome this potential limitation. Meiotic recombination events breaking the linkage between these two genes were discovered during the systematic screening of a population of 4,500 F2 durum plants (cv. Langdon background) used to fine map Yr36. One of the critical recombination events was selected for fixation by self-pollination and transferred to a California adapted spring hexaploid background (breeding line UC11105+10) through five generations of backcrossing. Genotypic and phenotypic data confirm the presence of Yr36 and the non-functional Gpc-B1 allele in both registered lines.

Ectopic expression of alpha B-crystallin in Chinese hamster ovary cells suggests a nuclear role for this protein.

alpha B-crystallin (alpha B) is known to be a cytosolic, small heat shock-like multimeric protein that has anti-aggregation, chaperone-like properties. The expression of the alpha B-crystallin gene is developmentally regulated and is induced by a variety of stress stimuli. Importantly, alpha B-crystallin expression is enhanced during oncogenic transformation of cells, in a number of tumors, and most notably, in many neurodegenerative disorders, including Alzheimer's disease and multiple sclerosis. Other than its perceived role as a structural protein in the ocular lens, the actual function of alpha B-crystallin in cellular physiology remains unknown. We have stably transfected CHO cells with an inducible alpha B-cDNA-MMTV-promoter construct that allows the synthesis of recombinant alpha B-crystallin only upon exposure of these cells to dexamethasone. Using immunostaining and conventional and confocal microscopy, we have examined the subcellular distribution of the ectopically expressed alpha B-crystallin. We find that in addition to being in the cytoplasm, the protein resides in the nuclear interior in the interphase nucleus. Double labeling with anti alpha B-crystallin and anti-tubulin, concanavallin, and wheat germ agglutinin, respectively, revealed that during cell division alpha B-crystallin is excluded from condensed chromatin and the nascent nuclei. However, the protein again appears in the newly formed nuclei after the completion of cytokinesis suggesting a conditional, regulatory role for alpha B-crystallin in the nucleus.

The actin network in the ciliary stalk of photoreceptors functions in the generation of new outer segment discs.

Cytochalasin D (CD) interferes with the morphogenesis of outer segment disc membrane in photoreceptors. Disruption of either the actin network in the ciliary stalk, where membrane evagination is initiated, or the actin core of the calycal processes, whose position could define the disc perimeter, could be responsible. We have attempted to determine which of these local F-actin populations is involved in membrane morphogenesis and what step in the process is actin-dependent. Biocytin accumulation in nascent discs, detected by fluorescent avidin and laser scanning confocal microscopy (LSCM), provided a means of labeling abnormal discs and a measure of disc membrane addition. F-actin content and distribution were assessed using fluorescent phalloidin and LSCM. First, we examined the effects of a range of CD dosages (0.1, 1.0, or 10.0 microM) on rod photoreceptors in Xenopus laevis eyecup cultures. Ectopic outgrowth of discs, evaluated by LSCM and transmission electron microscopy (TEM), occurred at each concentration. Phalloidin labeling intensified in the ciliary stalk with increasing CD concentration, indicating F-actin aggregation. In contrast, it diminished in the calycal processes, indicating dispersal; TEM showed that calycal process collapse ensued. Disruption was evident at a lower concentration in the ciliary stalk (0.1 microM) than in the calycal processes (1.0 microM). TEM confirmed that the calycal processes remained intact at 0.1 microM. Thus, CD's action on the ciliary stalk network is sufficient to disrupt disc morphogenesis. Second, we examined the effect of CD on temperature-induced acceleration of the rate of disc formation. In the absence of CD, a 10 degrees C temperature shift increased the disc formation rate nearly three-fold. CD (5 microM) caused a 94% inhibition (P < 0.025) of this response; yet, the rate of membrane addition to ectopically growing discs exhibited the expected three-fold increase. Thus, CD's action interferes with the generation of new discs.

Effects of retinal detachment on rod disc membrane assembly in cultured frog retinas.

The authors compared rod outer segment (ROS) disc membrane assembly rates in detached and attached frog retinas to determine if there was a rapid impairment of membrane assembly in response to retinal detachment. Membrane assembly was quantified in vitro by incubating retinas in medium containing Lucifer yellow, which is entrapped by nascent discs. Video microscopy was used to detect incorporation of the dye. During the first 10 hr after separation of the retina from the retinal pigment epithelium (RPE), ROS-disc membrane assembly in isolated Xenopus laevis neural retinas continued at a near normal rate, 0.81 microns/10 hr, a 13% reduction (P less than .01), compared with the 0.93 microns/10 hr observed in attached control retinas. The morphology of the OS appeared normal in most rod photoreceptors by transmission electron microscopy, although vesiculation of the most basal OS membranes was seen in a small population (25%) of rods. Approximately 90% of rod photoreceptors continued to assemble OS membranes for more than 10 hr after detachment, but by the end of 2 days, only 55% were still making new discs. The percentage of rods with normal basal OS membranes also decreased (to approximately 50%). Therefore, only 25% were assembling morphologically normal discs 2 days after detachment. In attached control regions, rod photoreceptors showed a comparatively minor response to culture conditions; assembly of morphologically normal discs continued for 2 days in about 85% and ceased in only 10%. These results indicate that the effects on disc membrane assembly of disrupting photoreceptor-RPE interaction in vitro initially are slight but become progressively severe with time.

Evidence that microtubules do not mediate opsin vesicle transport in photoreceptors.

The organization of the rod photoreceptor cytoskeleton suggests that microtubules (MTs) and F actin are important in outer segment (OS) membrane renewal. We studied the role of the cytoskeleton in this process by first quantifying OS membrane assembly in rods from explanted Xenopus eyecups with a video assay for disc morphogenesis and then determining if the rate of assembly was reduced after drug disassembly of either MTs or F actin. Membrane assembly was quantified by continuously labeling newly forming rod OS membranes with Lucifer Yellow VS (LY) and following the tagged membranes' distal displacement along the OS. LY band displacement displayed a linear increase over 16 h in culture. These cells possessed a longitudinally oriented network of ellipsoid MTs between the sites of OS protein synthesis and OS membrane assembly. Incubation of eyecups in nocodazole, colchicine, vinblastine, or podophyllotoxin disassembled the ellipsoid MTs. Despite their absence, photoreceptors maintained a normal rate of OS assembly. In contrast, photoreceptors displayed a reduced distal displacement of LY-labeled membranes in eyecups treated with cytochalasin D, showing that our technique can detect drug-induced changes in basal rod outer segment assembly. The reduction noted in the cytochalasin-treated cells was due to the abnormal lateral displacement of newly added OS disc membranes that occurs with this drug (Williams, D. S., K. A. Linberg, D. K. Vaughan, R. N. Fariss, and S. K. Fisher. 1988. J. Comp. Neurol. 272:161-176). Together, our results indicate that the vectorial transport of OS membrane constituents through the ellipsoid and their assembly into OS disc membranes are not dependent on elliposid MT integrity.