bluuebunny/crossref_metadata_2025 huggingface dataset:
Alu elements mediate MYB gene tandem duplication in human T-ALL
Authors: O'Neil Jennifer , Tchinda Joelle , Gutierrez Alejandro , Moreau Lisa , Maser Richard S. , Wong Kwok-Kin , Li Wei , McKenna Keith , Liu X. Shirley , Feng Bin , Neuberg Donna , Silverman Lewis , DeAngelo Daniel J. , Kutok Jeffery L. , Rothstein Rodney , DePinho Ronald A. , Chin Lynda , Lee Charles , Look A. Thomas
Published: December 2007

Recent studies have demonstrated that the MYB oncogene is frequently duplicated in human T cell acute lymphoblastic leukemia (T-ALL). We find that the human MYB locus is flanked by 257-bp Alu repeats and that the duplication is mediated somatically by homologous recombination between the flanking Alu elements on sister chromatids. Nested long-range PCR analysis indicated a low frequency of homologous recombination leading to MYB tandem duplication in the peripheral blood mononuclear cells of ∼50% of healthy individuals, none of whom had a MYB duplication in the germline. We conclude that Alu-mediated MYB tandem duplication occurs at low frequency during normal thymocyte development and is clonally selected during the molecular pathogenesis of human T-ALL.
Dual GPCR and GAG mimicry by the M3 chemokine decoy receptor
Authors: Alexander-Brett Jennifer M. , Fremont Daved H.
Published: December 2007

Viruses have evolved a myriad of evasion strategies focused on undermining chemokine-mediated immune surveillance, exemplified by the mouse γ-herpesvirus 68 M3 decoy receptor. Crystal structures of M3 in complex with C chemokine ligand 1/lymphotactin and CC chemokine ligand 2/monocyte chemoattractant protein 1 reveal that invariant chemokine features associated with G protein–coupled receptor binding are primarily recognized by the decoy C-terminal domain, whereas the N-terminal domain (NTD) reconfigures to engage divergent basic residue clusters on the surface of chemokines. Favorable electrostatic forces dramatically enhance the association kinetics of chemokine binding by M3, with a primary role ascribed to acidic NTD regions that effectively mimic glycosaminoglycan interactions. Thus, M3 employs two distinct mechanisms of chemical imitation to potently sequester chemokines, thereby inhibiting chemokine receptor binding events as well as the formation of chemotactic gradients necessary for directed leukocyte trafficking.
The Association of Tobacco mosaic virus with Green Spot of Cured Wrapper Tobacco Leaves
Authors: LaMondia J. A.
Published: December 2007

Near-isogenic lines of cigar wrapper tobacco resistant or susceptible to Tobacco mosaic virus (TMV) were used to evaluate the association of TMV infection with green spot symptoms in cured leaves. TMV infection, as determined by double-antibody sandwich enzyme-linked immunosorbent assay (ELISA), was detected on susceptible but not resistant plants in field experiments. Green spot severity on cured leaves was greater for susceptible than resistant plants, even when symptoms of TMV were not evident in the field. Some green spots were present on resistant leaves despite the fact that the virus was not detected by ELISA. Resistant and susceptible plants had similar responses to virus infection and similar ELISA detection of TMV when plants were held at continuous temperatures over 28°C in growth chambers. Plant resistance was not compromised in the field in cloth-covered shade tents even when 33.5 of the 96 h immediately following inoculation were above 28°C. Green spot of cured leaves was strongly associated with TMV infection in susceptible plants, even when plants were infected after leaf expansion and mosaic symptoms were not present. Green spot also occurred to a lesser extent and for a limited time in inoculated resistant plants. The development of green spot symptoms on cured leaves may be the result of either systemic infection of TMV-susceptible plants or associated with the systemic resistance response to TMV inoculation of resistant plants.
A Logistic Regression Model for Predicting Risk of White Mold Incidence on Dry Bean in North Dakota
Authors: Harikrishnan R. , Río L. E. del
Published: December 2007

White mold, caused by Sclerotinia sclerotiorum, is the most important disease affecting dry bean production in North Dakota. This disease currently is managed mainly through fungicides applied during the flowering stage. A disease-forecasting model was developed to help growers with their decision to apply these fungicides. The model was built using weather variables collected during eight consecutive half-month periods between 1 May and 31 August 2003 to 2005 and white mold incidence data obtained from 150 fields. The model was produced using logistic regression analysis, and includes total rainfall, average minimum temperature, and number of rainy days in the first half of June, July, and August, respectively, as predictors and explained 85% of the variability. The model was validated using an independent disease data set collected from 100 fields during the same years. The model exhibited high true positive ratio (0.79) and very high accuracy (0.91) between observed and predicted probabilities of white mold incidence. Results from this study suggest that in-season macro-weather variables could be used to predict the risk of white mold, which in-turn could help growers make better-informed decisions on whether or not to apply fungicides for white mold control.
Characterization of Verticillium dahliae and V. tricorpus Isolates from Lettuce and Artichoke
Authors: Qin Qing-Ming , Vallad Gary E. , Subbarao Krishna V.
Published: December 2007

Verticillium isolates collected from lettuce and artichoke were characterized for morphology, growth and pathogenicity. Several isolates were identified as Verticillium tricorpus based on morphological and cultural characteristics, including the production of dark resting mycelia, chlamydospores, microsclerotia, and yellow to orange pigmentation in culture. Compared with isolates of V. dahliae, these isolates also produced microsclerotia and conidia that were significantly larger and exhibited a distinct growth pattern at varying temperatures. Using database sequence information, primers were developed from the internal transcribed spacer region to produce a diagnostic 337-bp product specific to V. tricorpus and used to confirm the identification of isolates. Pathogenicity tests indicated that isolates of V. tricorpus were weak pathogens, causing a median disease severity (DS) of <1 (0-to-5 scale) on lettuce and artichoke. In contrast, isolates of V. dahliae consistently caused severe wilt with a median DS of >3.5 on lettuce and 5.0 on artichoke. Although lettuce and artichoke inoculated with isolates of V. tricorpus exhibited reduced height and fresh foliar and root weight, the reductions were not statistically significant, unlike in plants inoculated with isolates of V. dahliae. Lettuce co-inoculated with isolates of V. tricorpus and V. dahliae exhibited reduced symptoms of Verticillium wilt and improved growth relative to those inoculated with V. dahliae alone. The early introduction of V. tricorpus in soil-drench inoculations appeared to provide better relief from subsequent V. dahliae inoculation than when the two species were co-inoculated simultaneously using the root-dip method, suggesting competitive exclusion as a plausible mechanism. A spore-polymerase chain reaction assay developed using cultured spores directly as template and primers specific to V. tricorpus confirmed the presence of V. tricorpus on inoculated roots. This work demonstrates the potential use of V. tricorpus to directly reduce the effect of V. dahliae on lettuce and artichoke and, to our knowledge, is the first reported characterization of V. tricorpus isolates collected from lettuce and artichoke.
Genetic Analysis of Fungicide-Resistant Sclerotinia homoeocarpa Isolates from Tennessee and Northern Mississippi
Authors: DeVries R. E. , Trigiano R. N. , Windham M. T. , Windham A. S. , Sorochan J. C. , Rinehart T. A. , Vargas J. M.
Published: December 2007

Sclerotinia homoeocarpa is the causal agent of dollar spot disease that reduces the uniformity and aesthetic value of golf putting greens. Fungicide-resistant isolates of S. homoeocarpa were collected from putting greens at 10 locations across Tennessee and northern Mississippi. Genetic diversity among the 60 isolates was investigated using vegetative compatibility, conserved gene sequences, and amplified fragment length polymorphism (AFLP). Six tester strains were paired with Tennessee and northern Mississippi isolates on potato dextrose agar. Some of the 60 isolates were delineated into vegetative compatibility groups, but fungicide resistance could not be associated with a particular vegetative compatibility group. Genetic similarities of isolates at the vegetative compatibility level could be attributed to founder effects. Sequencing the regions of CAD, EF1-α, β-tubulin, and internal transcribed spacers revealed 100% homology among isolates. Capillary gel electrophoresis and analysis of AFLP fragments indicated 86 to 100% similarity between the isolates. Vegetative compatibility and molecular data indicate that the populations of the pathogen are clonal. Isolates did not cluster according to fungicide resistance during unweighted pair group with arithmetic means analysis, but did appear to cluster according to vegetative compatibility group and location. Although associations could not be made between molecular markers and fungicide resistance, links between vegetative compatibility and AFLP markers may provide a foundation from which other studies could be performed.
Adult Plant Evaluation of Soybean Accessions for Resistance to Phakopsora pachyrhizi in the Field and Greenhouse in Paraguay
Authors: Miles M. R. , Morel W. , Ray J. D. , Smith J. R. , Frederick R. D. , Hartman G. L.
Published: December 2007

Five hundred thirty soybean accessions from maturity groups (MG) III through IX were evaluated for resistance to Phakopsora pachyrhizi in a replicated field trial at Centro Regional de Investigación Agrícola in Capitán Miranda, Itapúa, Paraguay during the 2005–06 season. Soybean rust severities of individual accessions ranged from 0% (resistant) to 30.0% (susceptible). In MG III and IV, the most resistant accessions were PI 506863, PI 567341, and PI 567351B, with severities less than 1.2%. In MG V, the most resistant accessions were PI 181456, PI 398288, PI 404134B, and PI 507305, with severities less than 0.3%. In MG VI, the most resistant accessions were PI 587886, PI 587880A, and PI 587880B, with severities less than 0.3%. In MG VII and VIII, the most resistant were PI 587905 and PI 605779E, with severities less than 1.0%. In MG IX, the most resistant accessions were PI 594754, PI 605833, PI 576102B, and PI 567104B, with severities less than 1.0%. The resistance in 10 selected accessions from MG VI, VII, VIII, and XI was confirmed in subsequent greenhouse and field experiments where severities of 0.4% or less and reddish-brown lesions with sporulation levels less than 3.0 were observed. These accessions, with low severities in the adult plant field evaluation, may be new sources of resistance to P. pachyrhizi.
Detached-Leaf Bioassay for Evaluating Taro Resistance to Phytophthora colocasiae
Authors: Brooks F. E.
Published: December 2007

Taro leaf blight disease, caused by Phytophthora colocasiae, is a major limiting factor in taro production worldwide. P. colocasiae is an aerial pathogen similar to P. infestans, causal agent of potato late blight disease, but occurs in warmer climates. In the year-round subsistence cropping systems of the Pacific Islands, resistant cultivars are essential. Breeding lines from Southeast Asia and Oceania were tested in American Samoa for resistance to taro leaf blight using a detached-leaf bioassay and field trials. Mean lesion diameters from bioassays were highly correlated with field estimates of the number of healthy leaves per plant and yield (corm weight). However, the bioassay did not adequately assess infection efficiency. Additional experiments revealed that attached leaves had smaller lesion diameters than detached leaves incubated in closed containers, but both were very highly correlated. Taro resistance increased with plant age and the second-oldest leaf was more resistant than the third-oldest leaf. The bioassay was a fast, space-saving, effective method of screening taro lines for post-penetration resistance to P. colocasiae. It also provided an easily standardized method of evaluating host–pathogen interactions under controlled conditions.
Relation of Apple Flower Age to Infection of Hypanthium by Erwinia amylovora
Authors: Pusey P. L. , Smith T. J.
Published: December 2007

Blossom age as related to hypanthial susceptibility to Erwinia amylovora is not well established, but is relevant to disease risk assessment. To test this, detached crab apple blossoms were maintained for various periods and at different temperatures before applying inoculum to hypanthia. Inoculum potential on hypanthia due to wetting was evaluated by subjecting detached stigma-inoculated blossoms (~106 CFU per flower) to varying amounts and durations of simulated rain (or dew) at 14°C. Blossoms of varying age on mature ‘Gala’ apple trees were inoculated on hypanthia with 102, 104, or 106 CFU per flower. In the laboratory, susceptibility decreased with flower age at rates that increased with temperature. Wetness periods up to 12 h resulted in populations on hypanthia of <103 CFU per flower; 24 h of wetness resulted in ~104 or ~105 CFU. A dose response was shown in the orchard, and regression curves indicated steepest decline of susceptibility during initial days after petal expansion. Disease models incorporating a blossom-age component may be effective because they indicate the potential for infection when temperatures favor rapid bacterial growth on stigmas within a window of high hypanthial susceptibility. Further investigation of these relationships could lead to advancements in determining fire blight risk.
Resistance to Tan Spot and Stagonospora nodorum Blotch in Wheat-Alien Species Derivatives
Authors: Oliver R. E. , Cai X. , Wang R.-C. , Xu S. S. , Friesen T. L.
Published: December 2007

Tan spot (caused by Pyrenophora tritici-repentis) and Stagonospora nodorum blotch (SNB) (caused by Stagonospora nodorum) are destructive fungal diseases of wheat (Triticum aestivum) throughout the world. Host plant resistance is thought to be an efficient and economical method of control. The objective of the present study was to identify novel sources of tan spot and SNB resistance in wheat genotypes derived from the crosses between wheat and alien species. Evaluations were conducted at the seedling stage in a growth chamber with 100% relative humidity. For each genotype, three replications were used for each disease. Among the 199 wheat-alien species derivatives evaluated, 65 exhibited resistance to tan spot and 30 showed resistance to SNB similar to BR34, a Brazilian wheat line used as the resistant control. Eleven derivatives were resistant to both diseases. Reactions of the derivatives and their respective wheat parents to tan spot and SNB suggest that resistance genes in the derivatives are derived from alien species. These derivatives can serve as desirable bridges for introgression of resistance genes from alien species to cultivated wheat, and could contribute novel and effective tan spot and SNB resistance to wheat breeding.
Comparative Epidemiology of Monosporascus Root Rot and Vine Decline in Muskmelon, Watermelon, and Grafted Watermelon Crops
Authors: Beltrán R. , Vicent A. , García-Jiménez J. , Armengol J.
Published: December 2007

Disease progression and soil population dynamics of Monosporascus cannonballus ascospores were studied and compared through the entire growing season in muskmelon, watermelon, and watermelon grafted onto Cucurbita rootstock crops. In muskmelon and watermelon, there was a decrease of ascospore counts in soil from transplanting until first symptoms of vine decline were observed. This decrease probably was related to ascospore germination and subsequent root infection. Later, there was a remarkable increase of ascospore counts in a parallel way to the quick development of vine decline symptoms, which lead to plant death. In these crops, root colonization adjusted to first-order linear equations and vine decline progression best correlated with exponential and logistic models. In contrast, there were no symptoms of vine decline on grafted watermelon. Furthermore, the isolation of M. cannonballus from Cucurbita roots was considerably lower, perithecia were never observed, and ascospore soil populations remained stable. These results suggest that disease control by grafting onto genus Cucurbita seems to be related primarily by the increased resistance of its root system to infection by M. canonballus, and provide further evidence of the use of grafting as a disease management measure for this disease.
First Report of Cucumber mosaic virus on Jatropha curcas in India
Authors: Raj S. K. , Kumar S. , Snehi S. K. , Pathre U.
Published: December 2007

Jatropha curcas L. is a major commercial biodiesel fuel crop grown on 98 million acres (39.66 million ha) in India. Severe mosaic disease accompanied by yellow spots was noticed on 15% of J. curcas growing in the experimental plots of NBRI, Lucknow, India, during October of 2006. Inoculations with sap from symptomatic plants resulted in systemic mosaic on three of seven J. curcas seedlings. Gel diffusion tests were performed with antiserum to Cucumber mosaic virus (CMV), Tobacco ringspot virus, and Chrysanthemum virus B (PVAS-242a, PVAS-157, and PVAS-349, respectively; ATCC, Manassas, VA). Leaf sap of infected plants reacted only with PVAS-242a, indicating the presence of CMV. Reverse transcription (RT)-PCR assays with CMV coat protein gene specific primers (Genbank Accession Nos. AM180922 and AM180923) and total nucleic acid extracted from symptomatic J. curcas leaf tissue yielded the expected ~650-bp amplicon, which was cloned and sequenced (GenBank Accession No. EF153739). BLAST analysis indicated 98 to 99% nucleotide identity with CMV isolates (GenBank Accession Nos. DQ914877, DQ640743, AF350450, AF281864, X89652, AF198622, DQ152254, DQ141675, and DQ028777). Phylogenetic analysis showed that the J. curcas isolate was more closely related to Indian isolates of CMV belonging to subgroup Ib. Literature surveys revealed records of Jatropha mosaic virus on J. gossypiifolia in Puerto Rico (1) and on J. curcas in India (2). To our knowledge, this is the first report of CMV on J. curcas.

References: (1) J. K. Brown et al. Arch. Virol. 146:1581, 2001. (2) D. S. A. Narayana et al. Curr. Sci. 91:584, 2006.
First Report of Anthracnose Fruit Rot Caused by Colletotrichum acutatum on Pepper and Tomato in Bulgaria
Authors: Jelev Z. J. , Bobev S. G. , Minz D. , Maymon M. , Freeman S.
Published: December 2007

In the late summer of 2005, sporadic and unusual damage was observed on pepper (Capsicum annuum cv. Kurtovska kapia and local cv. Ribka) on two farms and tomato (Lycopersicon esculentum cv. Florida 47) fruits on one farm in the Plovdiv Region of Bulgaria. Dry, round, sunken zones (10 to 20 mm) were observed on pepper fruits that preserved their natural skin color even after black acervuli containing orange masses of conidia appeared. Eventually, the lesions turned brown, coalesced, and the fruits mummified on the plants. Tomato fruits developed similar symptoms, with less prominent discoloration and fewer acervuli. The pathogen was easily isolated from both hosts on potato dextrose agar where it formed white-to-gray colonies with salmon orange pigmentation on the reverse side of the plates. Conidia that formed were hyaline, fusiform, aseptate, and measured 13.3 to 17.4 × 3.5 to 5.5 μm and 11.6 to 15.5 × 4.1 to 5.0 μm for pepper and tomato isolates, respectively. Both cultural and morphological characteristics of the isolates were similar to those described for Colletotrichum acutatum (3). Koch's postulates were performed with two representative isolates from each host by artificial inoculation of healthy, green pepper and ripe tomato fruits from the respective cultivars. Fruits were wound inoculated with a sterile scalpel, and small agar plugs (3 to 4 mm) containing 7-day-old sporulating cultures were placed on each wound (five fruits per isolate), or by pipette tip-pricking and pipetting a 5-μl droplet of a conidial suspension (5 × 106 conidia ml–1) on each wound. The same number of wounded, noninoculated fruits was used as a control. Fruits were maintained in a humidity chamber at 22 to 25°C, and 4 days later, sunken necrotic zones were observed around the wounds of inoculated fruit, whereas control fruits remained symptomless. The pathogen was subsequently reisolated from the inoculated diseased tissues but not from the control fruits. Species-specific PCR (using primer pair CaInt2/ITS4) (2,4) of genomic DNA from three representative isolates (two from pepper and one from tomato) resulted in an amplification product of 490 bp, specific for C. acutatum, further confirming the identity of the pathogen. To our knowledge, this is the second report of C. acutatum in Bulgaria (1), and the first occurrence of that agent on tomato and pepper in this country.

References: (1) S. G. Bobev et al. Plant Dis. 86:1178, 2002. (2) S. Freeman et al. Phytopathology 91:586, 2001. (3) P. S. Gunnell and W. D. Gubler. Mycologia 84:157, 1992. (4) M. L. Lewis Ivey et al. Plant Dis. 88:1198, 2004.
First Report of Root and Crown Necrosis of Bean Caused by Pythium aphanidermatum in Spain
Authors: Serrano Y. , Guirado M. L. , Carmona M. P. , Gómez J. , Melero-Vara J. M.
Published: December 2007

Adult bean plants (Phaseolus vulgaris cv. Festival) growing in a commercial greenhouse in southeastern Spain developed symptoms of root necrosis, necrotic streaks on the basal stems, and plant wilt. A Pythium sp. was isolated consistently from roots and basal stems on selective agar (P5ARP). Single-hyphal transfers produced intercalary antheridia, oogonia (23 to 26 μm in diameter), oospores (18 to 20 μm in diameter), and zoospores in toruloid sporangia. Cardinal temperatures were a minimum of 10°C, an optimum of 28 to 34°C, and a maximum of 40°C. Daily growth rate on corn meal agar at 25°C was 15 mm. The internal transcribed spacer 1 (ITS 1) rDNA sequence of the isolate matched the sequences of Pythium aphanidermatum in GenBank. The sequence of isolate Py-294 was deposited in GenBank, Accession No. AM396563. This isolate was identified as P. aphanidermatum on the basis of morphological and cultural characteristics (1) and the ITS rDNA sequence. To fulfill Koch's postulates, 50 ml of inoculum of isolate Py-294 was used to inoculate bean plants (cvs. Donna and Emerite) at the five-leaf stage. The inoculum was prepared by homogenizing 2-week-old potato dextrose agar-petri plate cultures in 300 ml of distilled water. The plants were maintained in a greenhouse at temperatures of 18.8 to 30.3°C. Irrigation water had an electric conductivity of 0.5 to 0.6 dS·m–1 while the nutrient solution had 1.9 to 2.1 dS·m–1. Two months after sowing, 35.4 and 100% of cvs. Donna and Emerite, respectively, developed root necrosis, while control plants grown in bags containing noninoculated perlite remained healthy. The pathogen was reisolated from roots and basal stems of symptomatic plants. The test was repeated with similar results. To our knowledge, this is the first report of P. aphanidermatum as the causal agent of root and crown necrosis of adult bean plants in Spain.

Reference: (1) A. J. Van der Plaats-Niterink. Stud. Mycol. 21:242, 1981.
First Report of Xylella fastidiosa in Avocado in Costa Rica
Authors: Montero-Astúa M. , Saborío-R. G. , Chacón-Díaz C. , Garita L. , Villalobos W. , Moreira L. , Hartung J. S. , Rivera C.
Published: December 2007

Since the late 1990s, chlorotic mottling, marginal scorch, deformation of leaves, defoliation, shortening of internodes, and branch dieback have been observed in avocado trees (Persea americana Mill.) in Costa Rica. The symptoms are not uniformly distributed in the tree, so some branches are symptomatic while others are not. These symptoms are similar to several leaf scorch diseases caused by the bacterium Xylella fastidiosa Wells (2,4). This bacterium has been detected in coffee and citrus plants in Costa Rica. Of 227 avocado trees tested by double-antibody sandwich (DAS)-ELISA with X. fastidiosa specific antiserum (Agdia Inc., Elkhart, IN) from 2000–2004, 188 were positive. Results of ELISA tests of individual trees varied with the season and branches tested. Fifteen greenhouse-grown, ELISA-negative avocado seedlings were grafted with budwood from an ELISA-positive tree. Eight of these developed scorch symptoms and one also showed chlorotic mottling and deformation, showing that the disease is graft transmitted. All of these features are characteristic of diseases caused by X. fastidiosa (2,4). Transmission electron microscopy of leaf petioles from three field trees positive by ELISA, revealed rod-shaped bacilli approximately 1.6 to 2.0 μm long and 0.3 μm in diameter with a rippled cell wall inside xylem vessels and embedded in a matrix; morphology and measurements that are consistent with those reported for X. fastidiosa (2). DNA extraction and PCR attempts have been limited by mucilaginous sap from avocado. Positive PCR results (approximately 472-bp band) were obtained from two of the grafted seedlings and seven field trees from two distinct geographical locations (Alajuela and San José provinces) with DNA extractions from the plant sap using DNeasy Plant Mini Kit (Qiagen GmbH, Hilden, Germany) following a modified protocol (1) and nested PCR (3). Four of the PCR products, including one from the grafted seedlings, were cloned and sequenced in duplicate. GenBank sequences EU021997 to EU022000 present 99 to 100% sequence identity to a Pierce's disease strain from California (Temecula1) and 94 to 95% to a citrus variegated chlorosis strain from Brazil (Found-5). Several attempts have been made to isolate the bacterium in ‘periwinkle wilt’ and buffered cysteine-yeast extract media with negative results, probably because of the rapid production of mucilaginous sap when the avocado tissues were sampled. To our knowledge, this is the first report of X. fastidiosa in avocado trees.

References: (1) M. J. Green et al. Plant Dis. 83:482, 1999. (2) S. S. Hearon et al. Can. J. Bot. 58:1986, 1980. (3) M. R. Pooler and J. S. Hartung. Curr. Microbiol. 31:377, 1995. (4) A. H. Purcell et al. Phytopathology 89:53, 1999.
First Report of Pestalotiopsis dyospiri Causing Calyx Blight of Sweet Persimmon in Huelva Province (Southwestern Spain)
Authors: Blanco C. , Romero F. , De los Santos B.
Published: December 2007

In November 2006, necrotic calyxes were observed in more than 90% of harvested sweet persimmon fruits (Diospyros kaki L. fil.), cv. Triumph, in orchards located in Huelva Province. Symptoms appeared on sepals as brown-to-black necrotic areas; one to four sepals per fruit could be affected. Necrotic sepal tissues were surface disinfected and placed on potato dextrose agar medium (PDA). A fungus was consistently isolated from affected tissue on 60% of fruits sampled. Isolates formed acervular conidiomata (75.7 ± 4.69 μm in diameter) with fusiform conidia that were straight or rarely curved. Conidia were five celled, including three umbercolored medial cells and hyaline apical and basal cells. All conidia possessed two to three apical appendages and a single basal one. Conidia (N = 60) were 17.03 ± 0.25 μm long × 4.55 ± 0.1 μm wide; the three medial cells were 11.78 ± 0.15 μm long and apical cells were 2.34 ± 0.11 μm, with apical appendages measuring 12.45 ± 0.36 μm. Basal cells were 2.98 ± 0.11 μm and basal appendages were 3.93 ± 0.14 μm long. On the basis of these characteristics, the causal agent was identified as Pestalotiopsis diospyri Sidow et P. Sidow (2), that was previously described as a persimmon pathogen on leaves in Japan (3). To confirm pathogenicity, calyxes of healthy unwounded fruits were sprayed with 3 ml of a conidia suspension (1.5 × 105 conidia per ml) and maintained in moist chambers at 24°C for 10 days. All fruits presented necrotic calyxes, and the pathogen was reisolated from affected tissue in 100% of the samples. P. diospyri has been reported to be present on calyxes of unhealthy persimmon fruits in different productive regions in New Zealand (1), but to our knowledge, this is the first time that pathogenicity of this fungus was demonstrated on sweet persimmon fruits in Spain.

References: (1) H. H. Goh et al. Proc N Z Weed Pest Control Conf. 44:265, 1991. (2) E. F. Guba. Page 184 in: Monograph on Monochaetia and Pestalotia. Harvard University Press, Cambridge, MA. 1961. (3) F. Yasuda et al. J. Gen. Plant Pathol. 69:29, 2003.
First Outbreak of Bacterial Black Rot on Cabbage, Broccoli, and Brussels Sprouts Caused by Xanthomonas campestris pv. campestris in the Mediterranean Region of Turkey
Authors: Mirik M. , Selcuk F. , Aysan Y. , Sahin F.
Published: December 2007

During warm and humid periods in the winters of 2004 to 2006, severe leaf necrosis and vein rot symptoms were observed on cabbage (Brassica oleracea var. capitata L.), broccoli (Brassica oleracea var. italica Plenck.), and Brussels sprouts (Brassica oleracea var. gemmifera D.C.) in the Mediterranean Region of Turkey. Symptoms were characterized by yellow, V-shaped areas of the leaf margin, with the internal tissue turning from brown to black. Infected seedlings were also observed in commercial nurseries in Adana with a disease incidence of nearly 10 to 25%. Isolations made from leaves and veins of the affected plants on yeast dextrose calcium carbonate agar yielded yellow, mucoid, and convex colonies. Twenty isolates recovered from diseased leaf samples were selected at random to identify the causal organism. All isolates were nonspore forming, gram negative, rod shaped, motile, aerobic, oxidase-negative, catalase-positive, and amylolytic-positive (3). All isolates induced hypersensitive responses on tobacco (Nicotiana tabacum cv. Samsun). The isolates were identified as Xanthomonas campestris pv. campestris on the basis of fatty acid methyl ester (FAME) profiles determined by Sherlock Microbial Identification System software (Microbial ID, Newark, DE) and indirect ELISA. The similarity indices for the FAME analysis ranged from 80 to 94% (2). Indirect ELISA with a polyclonal antibody (Agdia, Elkhart IN; BRA 97000/0500) further confirmed the identity of the pathogen in both pure culture and infected plant. The mean absorbance values for three replications of indirect ELISA tests ranged from 1.411 to 3.508 at a wavelength of A405 (1). Pathogenicity of the isolates was tested on 5-week-old cabbage plants by spray inoculation using bacterial suspensions (107 CFU/ml) prepared in saline buffer (0.85% NaCl). Sterile saline buffer was sprayed on negative control plants. Inoculated and control plants were maintained for 5 days at 25°C and 70% relative humidity to observe symptom development. No symptoms developed on negative control plants. The bacterium was reisolated from inoculated cabbage plants and identified as X. campestris pv. campestris by FAME and an ELISA test. To our knowledge, this is the first report of the occurrence and outbreak of X. campestris pv. campestris in the Mediterranean Region of Turkey.

References: (1) A. M. Alvarez et al. Phytopathology 84:1449, 1994. (2) A. R. Chase et al. Phytopathology 82:754, 1992. (3) N. W Schaad et al. Xanthomonas. Page 175 in: Laboratory Guide for Identification of Plant Pathogenic Bacteria. 3rd ed. N. W. Schaad et al., eds. American Phytopathological Society. St. Paul, MN, 2001.
Resolving the structural features of genomic islands: A machine learning approach
Authors: Vernikos Georgios S. , Parkhill Julian
Published: December 2007

Large inserts of horizontally acquired DNA that contain functionally related genes with limited phylogenetic distribution are often referred to as genomic islands (GIs), and structural definitions of these islands, based on common features, have been proposed. Although a large number of mobile elements fall well within the GI definition, there are several concerns about the structural consensus for GIs: The current GI definition was put forward 10 yr ago when only 12 complete bacterial genomes were available, a large number of GIs deviate from that definition, and in silico predictions assuming a full/partial GI structural model bias the sampling of the GI structural space toward “well-structured” GIs. In this study, the structural features of genomic regions are sampled by a hypothesis-free, bottom-up search, and these are exploited in a machine learning approach with the aim of explicitly quantifying and modeling the contribution of each feature to the GI structure. Performing a whole-genome-based comparative analysis between 37 strains of three different genera and 12 outgroup genomes, 668 genomic regions were sampled and used to train structural GI models. The data show that, overall, GIs from the three different genera fall into distinct, genus-specific structural families. However, decreasing the taxa resolution, by studying GI structures across different genus boundaries, provides models that converge on a fairly similar GI structure, further suggesting that GIs can be seen as a superfamily of mobile elements, with core and variable structural features, rather than a well-defined family.
Replication of Colonic Crohn's Disease Mucosal
Escherichia coli
Isolates within Macrophages and Their Susceptibility to Antibiotics
Authors: Subramanian Sreedhar , Roberts Carol L. , Hart C. Anthony , Martin Helen M. , Edwards Steve W. , Rhodes Jonathan M. , Campbell Barry J.
Published: December 2007

There is increasing evidence that
Escherichia coli
organisms are important in Crohn's disease (CD) pathogenesis. In CD tissue they are found within macrophages, and the adherent-invasive CD ileal
E. coli
isolate LF82 can replicate inside macrophage phagolysosomes. This study investigates replication and antibiotic susceptibility of CD colonic
E. coli
isolates inside macrophages. Replication of CD colonic
E. coli
within J774-A1 murine macrophages and human monocyte-derived macrophages (HMDM) was assessed by culture and lysis after gentamicin killing of noninternalized bacteria and verified by electron microscopy (EM). All seven CD colonic isolates tested replicated within J774-A1 macrophages by 3 h (6.36-fold ± 0.7-fold increase;
n
= 7 isolates) to a similar extent to CD ileal
E. coli
LF82 (6.8-fold ± 0.8-fold) but significantly more than control patient isolates (5.2-fold ± 0.25-fold;
n
= 6;
P
= 0.006) and
E. coli
K-12 (1.0-fold ± 0.1-fold;
P
< 0.0001). Replication of CD
E. coli
HM605 within HMDM (3.9-fold ± 0.7-fold) exceeded that for K-12 (1.4-fold ± 0.2-fold;
P
= 0.03). EM showed replicating
E. coli
within macrophage vacuoles. Killing of HM605 within J774-A1 macrophages following a 3-h incubation with antibiotics at published peak serum concentrations (
C
max
) was as follows: for ciprofloxacin, 99.5% ± 0.2%; rifampin, 85.1% ± 6.6%; tetracycline, 62.8% ± 6.1%; clarithromycin, 62.1% ± 5.6% (all
P
< 0.0001); sulfamethoxazole, 61.3% ± 7.0% (
P
= 0.0007); trimethoprim, 56.3% ± 3.4% (
P
< 0.0001); and azithromycin, 41.0% ± 10.5% (
P
= 0.03). Ampicillin was not effective against intracellular
E. coli
. Triple antibiotic combinations were assessed at 10%
C
max
, with ciprofloxacin, tetracycline, and trimethoprim causing 97% ± 0.0% killing versus 86% ± 2.0% for ciprofloxacin alone. Colonic mucosa-associated
E. coli
, particularly CD isolates, replicate within macrophages. Clinical trials are indicated to assess the efficacy of a combination antibiotic therapy targeting intramacrophage
E. coli
.
Staphylococcus aureus
Clinical Isolate with High-Level Methicillin Resistance with an
lytH
Mutation Caused by IS
1182
Insertion
Authors: Fujimura Takaji , Murakami Kazuhisa
Published: December 2007

We previously reported that deficiency of the
lytH
gene, whose product is homologous to lytic enzymes, caused the elevation of methicillin resistance in
Staphylococcus aureus
strain SR17238, a strain of
S. aureus
with a low level of resistance to methicillin (low-level MRSA) (J. Bacteriol. 179:6294-6301, 1997). In this study, we demonstrated that deficiency of
lytH
caused the same phenomenon in four other clinical isolates of low-level MRSA, suggesting this deficiency to exist in clinical isolates. We therefore searched the region including
lytH
in 127 clinical isolates of MRSA by PCR and found one strain, SR17164 (methicillin MIC, 1,600 μg/ml), in which the
lytH
gene was inactivated by insertion sequence IS
1182. lytH
::IS
1182
was replaced with intact
lytH
in this strain by integration and excision of the plasmid carrying the
lytH
region. Recombinants with intact
lytH
genes showed methicillin MICs of 800 μg/ml, twofold lower than those of the recombinants with
lytH
::IS
1182
and the parent. In addition,
S. aureus
SR17164, which has a high level of methicillin resistance, had properties similar to those caused by
lytH
deficiency; that is, the resistance levels of strain SR17164 and
lytH
-deficient variants from strain SR17238 were not significantly affected by
llm
inactivation, which greatly lowered resistance levels in most other high-level MRSA strains. These findings suggest that
lytH
inactivation contributed, to some extent, to the resistance level of
S. aureus
SR17164. To the best of our knowledge, this strain is the first clinical isolate of MRSA for which the genetic base for high-level resistance has been clarified.
Biosynthetic Investigations of Lactonamycin and Lactonamycin Z: Cloning of the Biosynthetic Gene Clusters and Discovery of an Unusual Starter Unit
Authors: Zhang Xiujun , Alemany Lawrence B. , Fiedler Hans-Peter , Goodfellow Michael , Parry Ronald J.
Published: December 2007

The antibiotics lactonamycin and lactonamycin Z provide attractive leads for antibacterial drug development. Both antibiotics contain a novel aglycone core called lactonamycinone. To gain insight into lactonamycinone biosynthesis, cloning and precursor incorporation experiments were undertaken. The lactonamycin gene cluster was initially cloned from
Streptomyces rishiriensis
. Sequencing of ca. 61 kb of
S. rishiriensis
DNA revealed the presence of 57 open reading frames. These included genes coding for the biosynthesis of
l
-rhodinose, the sugar found in lactonamycin, and genes similar to those in the tetracenomycin biosynthetic gene cluster. Since lactonamycin production by
S. rishiriensis
could not be sustained, additional proof for the identity of the
S. rishiriensis
cluster was obtained by cloning the lactonamycin Z gene cluster from
Streptomyces sanglieri
. Partial sequencing of the
S. sanglieri
cluster revealed 15 genes that exhibited a very high degree of similarity to genes within the lactonamycin cluster, as well as an identical organization. Double-crossover disruption of one gene in the
S. sanglieri
cluster abolished lactonamycin Z production, and production was restored by complementation. These results confirm the identity of the genetic locus cloned from
S. sanglieri
and indicate that the highly similar locus in
S. rishiriensis
encodes lactonamycin biosynthetic genes. Precursor incorporation experiments with
S. sanglieri
revealed that lactonamycinone is biosynthesized in an unusual manner whereby glycine or a glycine derivative serves as a starter unit that is extended by nine acetate units. Analysis of the gene clusters and of the precursor incorporation data suggested a hypothetical scheme for lactonamycinone biosynthesis.
In Vitro Resistance Study of AG-021541, a Novel Nonnucleoside Inhibitor of the Hepatitis C Virus RNA-Dependent RNA Polymerase
Authors: Shi Stephanie T. , Herlihy Koleen J. , Graham Joanne P. , Fuhrman Shella A. , Doan Chau , Parge Hans , Hickey Michael , Gao Jingjin , Yu Xiu , Chau Fannie , Gonzalez Javier , Li Hui , Lewis Cristina , Patick Amy K. , Duggal Rohit
Published: December 2007

A novel class of nonnucleoside hepatitis C virus (HCV) polymerase inhibitors characterized by a dihydropyrone core was identified by high-throughput screening. Crystallographic studies of these compounds in complex with the polymerase identified an allosteric binding site close to the junction of the thumb and finger domains, approximately 30 Å away from the catalytic center. AG-021541, a representative compound from this series, displayed measurable in vitro antiviral activity against the HCV genotype 1b subgenomic replicon with a mean 50% effective concentration of 2.9 μM. To identify mutations conferring in vitro resistance to AG-021541, resistance selection was carried out using HCV replicon cells either by serial passages in increasing concentrations of AG-021541 or by direct colony formation at fixed concentrations of the compound. We identified several amino acid substitutions in the AG-021541-binding region of the polymerase, including M423(T/V/I), M426T, I482(S/T), and V494A, with M423T as the predominant change observed. These mutants conferred various levels of resistance to AG-021541 and structurally related compounds but remained sensitive to interferon and HCV polymerase inhibitors known to interact with the active site or other allosteric sites of the protein. In addition, dihydropyrone polymerase inhibitors retained activity against replicons that contain signature resistance changes to other polymerase inhibitors, including S282T, C316N, M414T, and P495(S/L), indicating their potential to be used in combination therapies with these polymerase inhibitors. AG-021541-resistant replicon cell lines provide a valuable tool for mechanism-of-action studies of dihydropyrone polymerase inhibitors. The clinical relevance of in vitro resistance to HCV polymerase inhibitors remains to be investigated.
Deactivation of Human Immunodeficiency Virus Type 1 in Medium by Copper Oxide-Containing Filters
Authors: Borkow Gadi , Lara Humberto H. , Covington Chandice Y. , Nyamathi Adeline , Gabbay Jeffrey
Published: December 2007

Human immunodeficiency virus type 1 (HIV-1) can be transmitted through breast-feeding and through contaminated blood donations. Copper has potent biocidal properties and has been found to inactivate HIV-1 infectivity. The objective of this study was to determine the capacity of copper-based filters to inactivate HIV-1 in culture media. Medium spiked with high titers of HIV-1 was exposed to copper oxide powder or copper oxide-impregnated fibers or passed through copper-based filters, and the infectious viral titers before and after treatment were determined. Cell-free and cell-associated HIV-1 infectivity was inhibited when exposed to copper oxide in a dose-dependent manner, without cytotoxicity at the active antiviral copper concentrations. Similar dose-dependent inhibition occurred when HIV-1 was exposed to copper-impregnated fibers. Filtration of HIV-1 through filters containing the copper powder or copper-impregnated fibers resulted in viral deactivation of all 12 wild-type or drug-resistant laboratory or clinical, macrophage-tropic and T-cell-tropic, clade A, B, or C, HIV-1 isolates tested. Viral inactivation was not strain specific. Thus, a novel means to inactivate HIV-1 in medium has been developed. This inexpensive methodology may significantly reduce HIV-1 transmission from “mother to child” and/or through blood donations if proven to be effective in breast milk or plasma and safe for use. The successful application of this technology may impact HIV-1 transmission, especially in developing countries where HIV-1 is rampant.
New Genetic Element Carrying the Erythromycin Resistance Determinant
erm
(TR) in
Streptococcus pneumoniae
Authors: Camilli Romina , Del Grosso Maria , Iannelli Francesco , Pantosti Annalisa
Published: December 2007

erm
(A) subclass
erm
(TR), a common macrolide resistance determinant in
Streptococcus pyogenes
but quite rare in
Streptococcus pneumoniae
, was found in a clinical
S. pneumoniae
isolate (AP200) from Italy. In this isolate,
erm
(TR) was found included in a genetic element approximately 56 kb in size that did not appear to be conjugative but could be transferred by transformation. An
erm
(TR)-containing DNA fragment of approximately 10 kb was sequenced and 12 open reading frames (ORFs) were identified. Upstream of
erm
(TR), a regulatory protein of the TetR family and the two components of an efflux pump of the ABC type were found. Downstream of
erm
(TR), there were ORFs homologous to a spectinomycin phosphotransferase, transposases, and a relaxase. Since the genomic sequence of
S. pyogenes
MGAS10750 carrying
erm
(TR) became available, comparison between the
erm
(TR)-containing genetic elements in AP200 and in MGAS10750 was performed. The region flanking
erm
(TR) in MGAS10750 showed identity with AP200 for 10 ORFs out of 12. PCR mapping using primers designed on the sequence of MGAS10750 confirmed that AP200 carries a genetic element similar to that of MGAS10750. In AP200 the genetic element was inserted inside an ORF homologous to spr0790 of
S. pneumoniae
R6, coding for a type I restriction modification system. Homologies between the insertion sites in AP200 and MGAS10750 consisted of eight conserved nucleotides, of which three were duplicated, likely representing target site duplication. The structure of the
erm
(TR)-carrying genetic element shows characteristics of a transposon/prophage remnant chimera. In AP200 this genetic element was designated Tn
1806
.
Antimicrobial Activities of Tigecycline and Other Broad-Spectrum Antimicrobials Tested against Serine Carbapenemase- and Metallo-β-Lactamase-Producing
Enterobacteriaceae
: Report from the SENTRY Antimicrobial Surveillance Program
Authors: Castanheira Mariana , Sader Hélio S. , Deshpande Lalitagauri M. , Fritsche Thomas R. , Jones Ronald N.
Published: December 2007

A total of 104 carbapenemase (serine- and metallo-β-lactamase [MβL])-producing strains of the
Enterobacteriaceae
family collected from 2000 to 2005 in medical centers distributed worldwide were tested against tigecycline and 25 comparators by reference broth microdilution methods. The most frequent carbapenemase was KPC-2 or -3 (73 strains), followed by VIM-1 (14), IMP-1 (11), SME-2 (5), and NMC-A (1). All serine carbapenemases were detected in the United States, while MβL-producing strains were isolated in Europe. Carbapenemase-producing
Enterobacteriaceae
showed high rates of resistance to most antimicrobial agents tested. The rank order of in vitro activity against these strains was as follows: tigecycline (100.0% susceptible) > polymyxin B (88.1%) > amikacin (73.0%) > imipenem (37.5%). Tigecycline was very active (MIC
90
, 1 μg/ml) against this significant, contemporary collection of well-characterized strains and appears to be an excellent option compared to the polymyxins for treatment of infections caused by these multidrug-resistant
Enterobacteriaceae
.
Complete DNA Sequence and Analysis of the Transferable Multiple-Drug Resistance Plasmids (R Plasmids) from
Photobacterium damselae
subsp.
piscicida
Isolates Collected in Japan and the United States
Authors: Kim Mi-Jung , Hirono Ikuo , Kurokawa Ken , Maki Takeshi , Hawke John , Kondo Hidehiro , Santos Mudjekeewis D. , Aoki Takashi
Published: December 2007

Photobacterium damselae
subsp.
piscicida
is a bacterial fish pathogen that causes a disease known as pasteurellosis. Two transferable multiple-drug resistance (R) plasmids, pP99-018 (carrying resistance to kanamycin, chloramphenicol, tetracycline, and sulfonamide) and pP91278 (carrying resistance to tetracycline, trimethoprim, and sulfonamide), isolated from
P. damselae
subsp.
piscicida
strains from Japan (P99-018) and the United States (P91278), respectively, were completely sequenced and analyzed, along with the multiple-drug resistance regions of three other R plasmids also from
P. damselae
subsp.
piscicida
strains from Japan. The sequence structures of pP99-018 (150,057 bp) and pP91278 (131,520 bp) were highly conserved, with differences due to variation in the drug resistance and conjugative transfer regions. These plasmids, shown to be closely related to the IncJ element R391 (a conjugative, self-transmitting, integrating element, or constin), were divided into the conjugative transfer, replication, partition, and multiple-drug resistance regions. Each of the five multiple-drug resistance regions sequenced exhibited unique drug resistance marker composition and arrangement.