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Objective: To construct recombinant plasmid with isocitrate lyase (ICL) gene of Mycobacterium tuberculosis H37Rv for stable and high level expression of ICL in prokaryotic expression system., Methods: The recombinant plasmid with ICL gene (pET30 (a)-Rv0467) was constructed by polymerase chain reaction and cloning. The fusion protein was expressed in E. coli host strain BL21 (DE3). Activity of the fusion protein was studied after it was purified with metal chelating chromatography., Results: We constructed the plasmid which could highly express Mycobacterium tuberculosis H37Rv ICL. The plasmid was highly expressed in E. coli BL21 (DE3), in which the fusion protein accounted for 30% of total protein content. After having been purified by metal chelating chromatography, the purity of the soluble fusion protein was 90%. The fusion protein had activity of ICL., Conclusion: Using the prokaryotic expression system, the ICL gene of Mycobacterium tuberculosis H37Rv was successfully cloned and expressed, which build the basis for screening new anti-tuberculosis drugs with ICL as the target point.

Drug-resistant (including multidrug-resistant) bacteria increase continuously with the wide use of antibiotics, which have seriously threatened the human health. It is an important way to fight against drug-resistance by screening and developing novel drugs based on the various mechanisms of the bacterial drug tolerances. Meanwhile, the basic research related to the new drug R. & D. and studies on the new screening methods for the antimicrobial agents should be taken seriously and strengthened, so as to accelerate the process of finding new drugs and meet the challenge of new pathogens and new drug-resistant strains.

ABSTRACT Colletotrichum crown rot of strawberry in Florida is caused primarily by Colletotrichum gloeosporioides. To determine potential inoculum sources, isolates of Colletotrichum spp. from strawberry and various noncultivated plants growing in the areas adjacent to strawberry fields were collected from different sites. Species-specific internal transcribed spacer primers for C. gloeosporioides and C. acutatum were used to identify isolates to species. Random amplified polymorphic DNA (RAPD) markers were used to determine genetic relationships among isolates recovered from noncultivated hosts and diseased strawberry plants. Selected isolates also were tested for pathogenicity on strawberry plants in the greenhouse. In all, 39 C. gloeosporioides and 3 C. acutatum isolates were recovered from diseased strawberry crowns, and 52 C. gloeosporioides and 1 C. acutatum isolate were recovered from noncultivated hosts. In crown inoculation tests, 18 of the 52 C. gloeosporioides isolates recovered from noncultivated hosts were pathogenic to strawberry. Phylogenetic analysis using RAPD marker data divided isolates of C. gloeosporioides from noncultivated hosts into two separate clusters. One cluster contained 50 of the 52 isolates and a second cluster contained 2 isolates that were homothallic in culture. Isolates from strawberry were interspersed within the cluster containing the 50 isolates that were recovered from noncultivated hosts. The results are not inconsistent with the hypothesis that C. gloeosporioides isolates obtained from strawberry and noncultivated hosts adjacent to strawberry fields are from the same population and that noncultivated hosts can serve as potential inoculum sources for Colletotrichum crown rot of strawberry.

Phacidiopycnis rot, caused by Phacidiopycnis piri, is a newly recognized postharvest disease in pear fruit (Pyrus communis cv. d'Anjou) in the United States. To determine the prevalence and incidence of this disease, decayed fruit were sampled during packing and repacking operations from four packinghouses in 2001 and 2002. During March to May (repacking) in 2001, Phacidiopycnis rot was found in packed fruit that were stored in cardboard boxes from 22 of 26 grower lots (orchards), and accounted for 5 to 71% of the total decay. Phacidiopycnis rot, gray mold caused by Botrytis cinerea, and blue mold caused by Penicillium spp. accounted for an average of 34.1, 10.3, and 33.6% of decayed fruit from conventional orchards, respectively; and 22.8, 35.7, and 23.5% of decayed fruit from organic orchards, respectively. During November 2001 to January 2002 (packing), Phacidiopycnis rot was observed in fruit that were stored in field bins before packing from 30 of 33 grower lots, accounting for 18.4% of decayed fruit sampled. During March to May in 2002, Phacidiopycnis rot was responsible for 2 to 68% of decayed fruit sampled from 36 of 39 grower lots. Phacidiopycnis rot, gray mold, and blue mold accounted for an average of 19.6, 26.8, and 37.4% of decayed fruit from conventional orchards, respectively; and 42.2, 25.7, and 8.2% of decayed fruit from organic orchards, respectively. Most Phacidiopycnis rot that occurred in field bins before packing appeared to originate from wound infections; whereas after packing, approximately 60 and 30% of Phacidiopycnis rot originated from stem and calyx infections, respectively. This study indicates that Phacidiopycnis rot should be considered one of the targets for control of postharvest diseases in d'Anjou pears in the region.

During a survey of postharvest diseases in stored pears conducted in the 2001-02 storage season, a new postharvest fruit rot in d'Anjou pears was discovered in Washington State. Symptoms of this disease were stem-end rot, calyx-end rot, and wound-associated rot, which presumably originated from infections of stem, calyx, and wounds on the fruit surfaces, respectively. The decayed area on the fruit was firm or spongy and appeared brown. During the late storage period from March to May 2002, this disease was observed in 19 of 39 lots and accounted for 2 to 21% of all decayed fruit. The causal agent, Sphaeropsis sp., was consistently recovered from decayed fruit with the symptoms described above. Two isolates of the fungus were used for pathogenicity tests on pear fruit. Decay symptoms developed on fruit inoculated with spore suspensions of the fungus on the stem, calyx, and wounds on the fruit surface. The fungus was reisolated from these decayed fruit. The fungus, Sphaeropsis pyriputrescens sp. nov., was characterized and described. On potato dextrose agar (PDA), oatmeal agar, and pear juice agar at 20°C, the fungus grew at mean rates of 21, 15, and 24 mm day -1 in colony diameter, respectively. On PDA, the fungus formed a circular colony with dense, hyaline hyphae and a few or some aerial mycelia. Colonies appeared light yellow to yellow on 2-week-old PDA cultures. The fungus grew at temperatures from 0 to 25°C, with optimum growth between 15 and 20°C, little or no growth at 30°C, and no growth at 35°C. This is a low-temperature species.

During March to July 2003, a postharvest fruit rot was observed on 'Golden Delicious', 'Granny Smith', and 'Red Delicious' apples (Malus × domestica Borkh.) sampled from commercial packinghouses in Washington State. Losses as high as 24% in storage bins were observed in July on 'Red Delicious'. The disease started at the stem bowl area or the calyx end of the fruit. Decayed fruit was apparently not wounded. Decayed areas were brown and firm. Internal decayed flesh appeared yellowish brown. On 'Red Delicious' apples, decayed fruit was apparently discolored from red to brown. As the disease advanced, pycnidia of a fungus might form on the stem, sepals, or the surface of decayed fruit. Pycnidia were 0.3 to 0.7 mm in diameter, black, and partially immersed in decayed tissues. To isolate the causal agent, decayed fruit was lightly sprayed with 70% ethanol and air dried. Fragments of diseased tissue were removed from the margin of diseased and healthy tissue and plated on acidified potato dextrose agar (PDA). A fungus was consistently isolated from decayed fruit with the symptoms described above. On PDA, the colonies of the fungus first appeared with dense hyaline mycelium and later turned light yellow to yellow. Black pycnidia of the fungus formed on 2- to 3-week-old oatmeal agar cultures at 20°C under 12-h alternating cycles of fluorescent light and dark. The fungus was identified as Sphaeropsis pyriputrescens Xiao & J. D. Rogers, based on the description of the fungus (1). Voucher specimens were deposited at the WSU Mycological Herbarium. Two isolates of the fungus recovered from decayed apples were tested for pathogenicity on apple. Fruit of 'Golden Delicious' and 'Gala' were surface-disinfested for 5 min in 0.5% NaOCl, rinsed, and air dried. Fruit was wounded with a sterile 4-mm-diameter nail head. A 4-mm-diameter plug from the leading edge of a 3-day-old PDA culture or plain PDA (control) was placed in the wound of each of 10 replicate fruit for each isolate or control. Fruit was tray packed with polyethylene liners and stored in cardboard boxes in air at 3°C, and decay was evaluated 2 weeks after inoculation. Five decayed fruits from each treatment were selected for reisolation of the causal agent. The experiment was conducted twice. In a separate pathogenicity test, two isolates (one each from apple and pear) were included in the test. Fruit of 'Red Delicious' apple was prepared and inoculated as the same manner described above, but fruit was stored in air at 0°C. The experiment was conducted twice. All fruit that were inoculated with the fungus developed decay symptoms. No decay developed on fruit in the controls. The same fungus was reisolated from decayed fruit. This indicates that isolates from apple and pear were pathogenic to apple. S. pyriputrescens is the causal agent of a newly reported postharvest disease on 'd'Anjou' pears (1). To our knowledge, this is the first report of this fungus causing postharvest fruit rot on apple. We propose 'Sphaeropsis rot' as the name of this new disease on apple and pear. Preliminary evidence suggests that infection of fruit by this fungus occurred in the orchard prior to storage. Reference: (1) C. L. Xiao and J. D. Rogers. Plant Dis. 88:114, 2004.

A plant yield and disease incidence uniformity trial was conducted to provide information concerning the efficiency and precision of field trials used to evaluate Botrytis fruit rot control methods on strawberry. Fruit yield and Botrytis fruit rot incidence were recorded for individual strawberry plants of cultivars Sweet Charlie and Camarosa grown in an annual production system over two growing seasons. A nested analysis of variance model was used to measure plot edge effects and to obtain variance components to describe the relationship between plot size and plot variance. Mean seasonal yield for Sweet Charlie was 599 g/plant and for Camarosa 972 g/plant. Mean seasonal fruit rot incidence was 10.3% for Sweet Charlie and 3.0% for Camarosa. Plants growing on the edge of plots next to aisles had higher yields (637 versus 577 g/plant for Sweet Charlie and 1047 versus 923 g/plant for Camarosa), but there was no edge effect for disease incidence. Smith's equation was used to determine the relationship between plot size and plot variance within mulched beds. Smith's index was relatively high for yield (0.92 and 0.95) and Botrytis fruit rot incidence (0.91 and 0.69) for Sweet Charlie and Camarosa, respectively. This suggests a relatively uniform distribution of seasonal yield and disease incidence among plants within beds. Using plot variances estimated from Smith's equation, the power of hypothesis tests to discriminate hypothetical treatment effects of different magnitudes was examined for each cultivar. Power analysis suggests that treatment effects ranging from 11.3 to 21.3% of the mean seasonal yield observed for Sweet Charlie and 8.6 to 16.5% of the mean seasonal yield observed for Camarosa can be detected with a power of 0.80 using four replicates of each treatment group and plot sizes ranging from 32 to eight plants/plot. For Botrytis fruit rot incidence, a power of 0.80 is achieved with treatment differences ranging from 24.9 to 44.1% of mean seasonal Botrytis fruit rot incidence for Sweet Charlie and from 44.1 to 64.9% of the mean observed for Camarosa using plot sizes ranging from 32 to 8 plants/plot. Before planting, the crown diameter and leaf number of transplants were recorded. There was no consistent significant correlation between any of these growth traits and seasonal yields or Botrytis fruit rot incidence. However, there was a consistent positive correlation for both cultivars between yield and crown diameter and also yield and leaf number during the months of December and January in the early part of the season (0.1513 ≤ r ≤ 0.314 for crown diameter and 0.106 ≤ r ≤ 0.264 for leaf number).

Epidemics of Botrytis fruit rot (Botrytis cinerea) and powdery mildew (Sphaerotheca macularis f. sp. fragariae) in annual strawberry were compared in large plastic tunnel and field production systems during the 1998-99 and 1999-2000 seasons. Treatments were factorial combinations of two main plots (field and tunnel) and four subplots, including combinations of two cultivars (Camarosa and Sweet Charlie) and two captan schedules arranged in a split-plot design with three replications. The mean incidence of Botrytis fruit rot was 88 to 94% lower in tunnels than in the field. The incidence of Botrytis fruit rot for the untreated control in tunnels was less than 2%, which was 89% lower than that of the 7-day captan schedule in the field. This indicates that Botrytis fruit rot can be controlled effectively without fungicides in a tunnel cultural system. Powdery mildew was severe on susceptible cultivar Camarosa in tunnels. Early season yields of cultivar Sweet Charlie were significantly higher in tunnels than in the field. Shorter periods of leaf wetness and higher temperatures in tunnels may have contributed to a lower incidence of Botrytis fruit rot and a higher incidence of powdery mildew on fruit in tunnels compared with open field plots.

The control of postharvest Botrytis fruit rot was evaluated during 1997-98 and 1998-99. Weekly applications of captan and thiram were examined at two or three different rates, respectively. Iprodione applications were combined with the captan and thiram treatments and also applied alone for two peak bloom periods. Strawberry fruit were harvested and graded twice weekly for marketable yield and preharvest incidence of Botrytis fruit rot. For postharvest evaluations, fruit from four harvests were selected and stored at 4°C, and Botrytis fruit rot incidence was recorded over 14 days of storage. Fungicide treatments reduced the incidence of preharvest Botrytis fruit rot and increased marketable yield. The incidence of postharvest Botrytis fruit rot was significantly affected by harvest date, length of time in storage, and fungicide treatment. The highest rate captan and thiram treatments had the least Botrytis fruit rot and the longest storage life. Reduced-rate captan and thiram treatments generally did not provide the same control as their respective high-rate treatments. Iprodione added to either the captan or thiram treatments did not consistently reduce the preharvest or postharvest incidence of Botrytis fruit rot or increase yield. Regular, full-rate fungicide treatments appear to be necessary to control Botrytis fruit rot in Florida and to provide the storage life necessary to reach distant markets.

The management of Botrytis fruit rot on annual strawberry by fungicides was evaluated in Florida during the 1995-96, 1996-97, and 1997-98 seasons. Weekly applications of captan or thiram, bloom applications of iprodione applied twice during each of two peak flowering periods, and weekly applications of captan combined with iprodione bloom applications were evaluated. Significant treatment effects (P ≤ 0.05) on the incidence of Botrytis fruit rot were detected for the early, late, and whole-season periods each season. Weekly applications of captan or thiram controlled Botrytis fruit rot, reducing disease incidence by more than 41% compared to the untreated control. These treatments also affected marketable yield during two seasons, with a 42 to 127% increase in yield compared to the control. Weekly fungicide applications did not reduce the incidence of Botrytis fruit rot until at least the fourth week of harvest, 9 to 10 weeks after applications began. Bloom applications of iprodione alone reduced the incidence of Botrytis fruit rot during the second peak bloom period, and the reductions in incidence occurred 1 to 3 weeks after the start of bloom applications. This suggests that iprodione applications control infections at flowering or early stages of fruit development. However, early-season bloom applications did not reduce the incidence of Botrytis fruit rot. The control of Botrytis fruit rot by weekly captan applications was not improved by the addition of iprodione bloom applications. These data suggest that early-season fungicide applications for the control of Botrytis fruit rot in annual winter strawberry are of limited efficacy, and that bloom applications of Botryticides such as iprodione should be focused on the second peak bloom period.

To determine the effects of sanitation on yield and incidence of Botrytis fruit rot (Botrytis cinerea) in annual strawberry, replicated experiments were conducted during the 1995-96, 1996-97, and 1998-99 seasons. Leaf sanitation (removal of senescent and necrotic leaves) and fruit sanitation (removal of unmarketable fruit from alleys between beds) were compared to a standard fungicide control program (weekly applications of captan plus four bloom applications of iprodione) and combined sanitation and fungicide treatments. Leaf sanitation reduced Botrytis fruit rot incidence from 12.6 to 8.2% over the entire 1996-97 season, and from 17.6 to 11.8% during the latter half of the 1998-99 season, compared to untreated controls. However, sanitation did not increase marketable yield. Supplementing fungicides with leaf sanitation or leaf and fruit sanitation did not improve disease control and frequently reduced yield. Fruit sanitation had no significant effect on Botrytis incidence or yield. Losses to Botrytis fruit rot in the sanitation treatments were significantly higher (P ≤ 0.05) than in the fungicide treatments each season; marketable yields were significantly lower in 1996-97 and 1998-99. Under Florida conditions, fungicides control Botrytis fruit more effectively and economically than does sanitation.

ABSTRACT Cauliflower root and plant growth and Verticillium wilt development were evaluated under different moisture regimes in the presence or absence of V. dahliae. Treatments included two main plots (V. dahliae-infested and fumigated), two subplots (furrow and subsurface drip irrigation), and three sub-subplots (deficit, moderate, and excessive regimes) that were arranged in a split-split-plot design in the field. Soil cores with roots were periodically sampled at 5 and 25 cm distance from plants. Total roots in each soil core were extracted with a hydropneumatic root elutriator, and root length from each sample was determined with a digital image analysis system. Incidence and severity of Verticillium wilt, plant height, number of leaves, and dry weights of leaves and roots were determined on 10 plants sampled at 7- to 10-day intervals 1 month after cauliflower transplanting and continued until harvest. To evaluate the effects of Verticillium wilt-induced stress on cauliflower plants, stomatal resistance was measured in upper healthy and lower (or diseased) leaves. Root length density at 5 and 25 cm from plant was significantly (P < 0.05) higher in subsurface drip than in furrow irrigation. Root length density was significantly higher in excessive irrigation regime than in the other regimes. Concomitantly, there was higher wilt incidence and severity in excessive and moderate regimes than deficit regime regardless of the irrigation method. Plant height was affected by irrigation methods and deficit regime. Neither the method of irrigation nor the quantity of water affected the other variables. Stomatal resistance in lower diseased leaves was significantly higher in infested than in fumigated plots but it was not in the upper healthy leaves. In this study, cauliflower yield was not affected by V. dahliae and irrigation method, but the deficit irrigation regime resulted in reduced yield even though it suppressed wilt in cauliflower. Thus, higher moisture levels resulted in higher root length density in V. dahliae-infested plots that in turn lead to greater incidence of Verticillium wilt and severity. The pathogen also affected physiological processes such as hydraulic conductance of cauliflower leaves, but not shoot growth or yield under these experimental conditions.

Evaluating fungicide efficacy in annual strawberry production systems can be labor intensive due to continuous harvesting over a relatively long season. The effect of reduced harvest number on the accuracy of least significant difference (LSD) separations for Botrytis fruit rot (Botrytis cinerea) incidence and marketable yield in fungicide efficacy studies was evaluated over three seasons. Fruit were harvested and graded twice a week for a total of 23 to 32 harvests each season. Data from each season were divided into different sample sets. Data from three different harvest periods (early, late, and whole season) and different harvesting frequencies (twice weekly, once weekly, every second, third, or fourth week) were compared with the complete data set (twice weekly for the whole season). Spearman's rank correlation and Pearson's product moment correlation coefficients were used to evaluate the correlation of the complete data sets with data sets from other sampling plans. Harvesting once a week for either the late- or whole-season periods accurately estimated LSD groupings for Botrytis fruit rot incidence among fungicide treatments. The precision of marketable yield estimates using once-a-week harvesting for the late or whole-season periods were relatively lower than for the incidence of Botrytis.

The effects of within-row plant spacing and cultivar on the incidence of Botrytis fruit rot (Botrytis cinerea) and marketable yield of annual strawberry were evaluated during the 1997-98 and 1998-99 seasons. Three cultivars (Camarosa, Rosa Linda, and Sweet Charlie) and four plant spacings (23, 30, 38, and 46 cm) were evaluated. Marketable yield and the incidence of Botrytis fruit rot were determined twice weekly. Cultivar and spacing effects were analyzed for three periods each season (early, late, and whole season). In 1997-98, spacing effects were observed on weekly incidence of Botrytis rot for the late period (P = 0.0925) and on cumulative incidence for the whole season period (P = 0.0795). Further analysis of the late and whole season periods revealed a spacing effect for Camarosa (P = 0.0102). Spacing also had a dramatic effect on cumulative and weekly Botrytis incidence for the late and whole season periods during the 1998-99 season (P ≤ 0.0014), when narrower spacings had higher incidence of Botrytis than wider spacings. Marketable yields were higher at narrower spacings during the early period for both seasons. Whole season marketable yields were also higher at the narrower spacings despite higher incidence of Botrytis. There were significant differences in susceptibility among cultivars.

Peptides purified by HPLC are often in the form of a trifluoroacetate (TFA) salt, because trifluoroacetic acid is used as a solvent in reversed-phase HPLC separation. However, the potential effects of this contaminant in culture systems have not been addressed previously. TFA (10(-8) to 10(-7) M) reduced cell numbers and thymidine incorporation into fetal rat osteoblast cultures after 24 h. Similar effects were found in cultures of articular chondrocytes and neonatal mouse calvariae, indicating that the effect is not specific to one cell type or to one species of origin. When the activities of the TFA and hydrochloride salts of amylin, amylin-(1-8), and calcitonin were compared in osteoblasts, cell proliferation was consistently less with the TFA salts of these peptides, resulting in failure to detect a proliferative effect or wrongly attributing an antiproliferative effect. This finding is likely to be relevant to all studies of purified peptides in concentrations above 10(-9) M in whatever cell or tissue type. Such peptides should be converted to a hydrochloride or biologically equivalent salt before assessment of their biological effects is undertaken.

Calcitonin gene-related peptide (CGRP) and amylin are homologous 37-amino-acid peptides which have been demonstrated to have anabolic effects on bone. It is not clear whether these effects are mediated by a common receptor, nor is it known which ligand is the more potent. These questions are addressed in the present study using cultures of fetal rat osteoblasts. CGRP increased cell number when present in a concentration >/=10-9 M, but 10-8 M CGRP was required to stimulate thymidine and phenylalanine incorporation. Amylin was effective on these indices at 100-fold lower concentrations, and its maximal effects were about twice as great as those of CGRP. ED50's for the effects of amylin and CGRP on cell number were 10-12 M and 10-10 M, respectively. There was no additivity between maximal doses of the peptides on these indices. The effects of specific receptor blockers on the maximal stimulation of cell number by these peptides were also studied. The CGRP receptor-blocker, CGRP-(8-37), completely blocked the effect of CGRP at blocker concentrations >/=10-9 M. In contrast, the amylin receptor blocker, amylin-(8-37), completely blocked the effects of CGRP when the blocker was present in concentrations as low as 10-11 M. The KI of CGRP-(8-37) was 2 x 10-10 M and that of amylin-(8-37) was 7 x 10-12 M. In converse experiments studying the blockade of maximal doses of amylin, amylin-(8-37) 10-10 M was effective (KI 1 x 10-10 M), whereas a 100-fold greater concentration of CGRP-(8-37) was necessary to achieve the same effect (KI 6 x 10-9 M). It is concluded that amylin and CGRP probably act through a common receptor to stimulate osteoblast growth, and that this receptor has a higher affinity for amylin than for CGRP.

ABSTRACT Experiments were conducted in field plots to evaluate the effects of broccoli residue on population dynamics of Verticillium dahliae in soil and on Verticillium wilt development on cauliflower under furrow and subsurface-drip irrigation and three irrigation regimes in 1994 and 1995. Treatments were a factorial combination of three main plots (broccoli crop grown, harvested, and residue incorporated in V.dahliae-infested plots; no broccoli crop or residue in infested plots; and fumigated control plots), two subplots (furrow and subsurface-drip irrigation), and three sub-subplots (deficit, moderate, and excessive irrigation regimes) arranged in a split-split-plot design with three replications. Soil samples collected at various times were assayed for V. dahliae propagules using the modified Anderson sampler technique. Incidence and severity of Verticillium wilt on cauliflower were assessed at 7- to 10-day intervals beginning a month after cauliflower transplanting and continuing until harvest. Number of propagules in all broccoli plots declined significantly (P < 0.05) after residue incorporation and continued to decline throughout the cauliflower season. The overall reduction in the number of propagules after two broccoli crops was approximately 94%, in contrast to the fivefold increase in the number of propagules in infested main plots without broccoli after two cauliflower crops. Disease incidence and severity were both reduced approximately 50% (P < 0.05) in broccoli treatments compared with no broccoli treatments. Differences between furrow and subsurface-drip irrigation were not significant, but incidence and severity were significantly (P < 0.05) lower in the deficit irrigation regime compared with the other two regimes. Abundance of microsclerotia of V. dahliae on cauliflower roots about 8 weeks after cauliflower harvest was significantly (P < 0.05) lower in treatments with broccoli compared with treatments without broccoli. Rotating broccoli with cauliflower and incorporating broccoli residues into the soils is a novel means of managing Verticillium wilt on cauliflower and perhaps on other susceptible crops. This practice would be successful regardless of the irrigation methods or regimes followed on the susceptible crops.

ABSTRACT Microplot and field experiments were conducted to evaluate the effects of inoculum density on Verticillium wilt and cauliflower growth. Soil containing Verticillium dahliae microsclerotia was mixed with various proportions of fumigated soil to establish different inoculum densities (fumigated soil was used as the noninfested control). Seven inoculum density treatments replicated four times were established, and the treatments were arranged in a randomized complete block design. Soil was collected from each microplot immediately after soil infestation for V. dahliae assay by plating onto sodium polypectate agar (NP-10) selective medium using the Anderson sampler technique. Five-week-old cauliflower was transplanted into two beds within each 1.2- by 1.2-m microplot. At the same time, several extra plants were also transplanted at the edge of each bed for destructive sampling to examine the disease onset (vascular discoloration) after planting. Cauliflower plants were monitored for Verticillium wilt development. Stomatal resistance in two visually healthy upper and two lower, diseased leaves in each microplot was measured three times at weekly intervals after initial wilt symptoms occurred. At maturity, all plants were uprooted, washed free of soil, and wilt incidence and severity, plant height, number of leaves, and dry weights of leaves and roots were determined. The higher the inoculum density, the earlier was disease onset. A density of 4 microsclerotia per g of dry soil caused 16% wilt incidence, but about 10 microsclerotia per g of soil caused 50% wilt incidence. Both wilt incidence and severity increased with increasing inoculum density up to about 20 microsclerotia per g of soil, and additional inoculum did not result in significantly higher disease incidence and severity. A negative exponential model described the disease relationships to inoculum levels under both microplot and field conditions. Stomatal resistance of diseased leaves was significantly higher at higher inoculum densities; in healthy leaves, however, no treatment differences occurred. The height, number of leaves, and dry weights of leaves and roots of plants in the fumigated control were significantly higher than in infested treatments, but the effects of inoculum density treatments were variable between years. Timing of cauliflower infection, crop physiological processes related to hydraulic conductance, and wilt intensity (incidence and severity) were thus affected by the inoculum density. Verticillium wilt management methods used in cauliflower should reduce inoculum density to less than four micro-sclerotia per g of soil to produce crops with the fewest number of infected plants.

This study assesses the structure-activity relationships of the actions of amylin on bone. In fetal rat osteoblasts, only intact amylin and amylin-(1-8) stimulated cell proliferation (half-maximal concentrations 2.0 x 10(-11) and 2.4 x 10(-10) M, respectively). Amylin-(8-37), COOH terminally deamidated amylin, reduced amylin, and reduced amylin-(1-8) (reduction results in cleavage of the disulfide bond) were without agonist effect but acted as antagonists to the effects of both amylin and amylin-(1-8). Calcitonin gene-related peptide-(8-37) also antagonized the effects of amylin and amylin-(1-8) on osteoblasts but was substantially less potent in this regard than amylin-(8-37). In contrast, inhibition of bone resorption in neonatal mouse calvariae only occurred with the intact amylin molecule and was not antagonized by any of these peptides. The rate of catabolism of the peptides in calvarial cultures was not accelerated in comparison with that of intact amylin. This dissociation of the actions of amylin suggests that it acts through two separate receptors, one on the osteoclast (possibly the calcitonin receptor) and a second on the osteoblast.

ABSTRACT The spatial patterns of microsclerotia of Verticillium dahliae in soil and wilt symptoms on cauliflower were determined at three sites in each of two fields in 1994 and 1995. Each site was an 8 x 8 grid divided into 64 contiguous quadrats (2 by 2 m each). Soil samples were collected to a depth of 15 cm with a probe (2.5 cm in diameter), and samples from four sites in each quadrat were bulked. Plants in each quadrat were cut transversely, and the number of plants with vascular discoloration and the number without discoloration were recorded. The soil was assayed for microsclerotia by the modified Anderson sampler technique. Lloyd's index of patchiness (LIP) was used as an indicator to evaluate the aggregation of microsclerotia in the field. Spatial autocorrelation and geostatistical analyses were also used to assess the autocorrelation of microsclerotia among quadrats. The LIP for microsclerotia was greater than 1, indicating aggregation of propagules; however, the degree of aggregation at most sites was not high. Significant autocorrelation within or across rows was detected in some spatial autocorrelograms of propagules, and anisotropic patterns were also detected in some oriented semivariograms from geostatistical analyses for microsclerotia, indicating the influence of bed preparation in the fields on pathogen distribution. The parameter estimates p and theta in the beta-binomial distribution and the index of dispersion (D) associated with the distribution were used to assess the aggregation of diseased plants at each site. A random pattern of wilt incidence was detected at 7 of 12 sites, and an aggregated pattern was detected at 5 of 12 sites. The degree of aggregation was not high. A regular pattern of wilt severity was detected at all sites. The high disease incidence (77 to 98%) observed at 11 of the 12 sites could be explained by high inoculum density.