Crop production in the USA is frequently limited by a lack of pollinators.

Most of the world's crops depend on pollinators, so declines in both managed and wild bees raise concerns about food security. However, the degree to which insect pollination is actually limiting current crop production is poorly understood, as is the role of wild species (as opposed to managed honeybees) in pollinating crops, particularly in intensive production areas. We established a nationwide study to assess the extent of pollinator limitation in seven crops at 131 locations situated across major crop-producing areas of the USA. We found that five out of seven crops showed evidence of pollinator limitation. Wild bees and honeybees provided comparable amounts of pollination for most crops, even in agriculturally intensive regions. We estimated the nationwide annual production value of wild pollinators to the seven crops we studied at over $1.5 billion; the value of wild bee pollination of all pollinator-dependent crops would be much greater. Our findings show that pollinator declines could translate directly into decreased yields or production for most of the crops studied, and that wild species contribute substantially to pollination of most study crops in major crop-producing regions.

Managed and Wild Bee Flower Visitors and Their Potential Contribution to Pollination Services of Low-Chill Highbush Blueberry (Vaccinium corymbosum L.; Ericales: Ericaceae).

Highbush blueberry (Vaccinium corymbosum L.; Ericales: Ericaceae) is an important crop grown throughout the eastern United States and Canada. Cross-pollination by insects greatly enhances pollination and fruit set in highbush blueberry. In Florida, low-chill cultivars that flower during the winter when most bees are dormant are used, thus, making it difficult to utilize and depend on unmanaged bees. We investigated flower visitation rates by managed and wild bees and the subsequent berry formation, berry weight, and number of seeds/berry in highbush blueberry fields in north-central Florida. Additionally, we tested three pollinator treatments: 1) pollinator-excluded flowers, 2) open-pollinated treatments that were available to managed and wild bees, and 3) flowers that were hand pollinated. Overall, we found seven native bee species that contribute to highbush blueberry pollination in Florida, but managed honey bees and bumble bees were the main flower visitors. Additionally, 14.5 times more blueberries formed in the open treatments than in the pollinator exclusion treatments, thus illustrating the economic impact bees have on blueberry pollination. Most of the wild bees observed visiting blueberry flowers were ground-nesting species that need uncultivated areas for nesting sites. Therefore, leaving field edges uncultivated and some undisturbed habitat may increase native bee numbers within blueberry farms over time.

Managed Bumble Bees (Bombus impatiens) (Hymenoptera: Apidae) Caged With Blueberry Bushes at High Density Did Not Increase Fruit Set or Fruit Weight Compared to Open Pollination.

Highbush blueberry (Vaccinium corymbosum L.) is an important crop grown throughout Florida. Currently, most blueberry growers use honey bees (Apis mellifera L.) to provide pollination services for highbush blueberries even though bumble bees (Bombus spp.) have been shown to be more efficient at pollinating blueberries on a per bee basis. In general, contribution of bumble bees to the pollination of commercial highbush blueberries in Florida is unknown. Herein, we determined if managed bumble bees could contribute to highbush blueberry pollination. There were four treatments in this study: two treatments of caged commercial bumble bee (Bombus impatiens Cresson) colonies (low and high weight hives), a treatment excluding all pollinators, and a final treatment which allowed all pollinators (managed and wild pollinators) in the area have access to the plot. All treatments were located within a highbush blueberry field containing two cultivars of blooming plants, 'Emerald' and 'Millennia', with each cage containing 16 mature blueberry plants. We gathered data on fruit set, berry weight, and number of seeds produced per berry. When pollinators were excluded, fruit set was significantly lower in both cultivars (<8%) compared to that in all of the other treatments (>58%). Berry weight was not significantly different among the treatments, and the number of seeds per berry did not show a clear response. This study emphasizes the importance of bumble bees as an effective pollinator of blueberries and the potential beneficial implications of the addition of bumble bees in commercial blueberry greenhouses or high tunnels.

Experimental determination and system level analysis of essential genes in Escherichia coli MG1655.

Defining the gene products that play an essential role in an organism's functional repertoire is vital to understanding the system level organization of living cells. We used a genetic footprinting technique for a genome-wide assessment of genes required for robust aerobic growth of Escherichia coli in rich media. We identified 620 genes as essential and 3,126 genes as dispensable for growth under these conditions. Functional context analysis of these data allows individual functional assignments to be refined. Evolutionary context analysis demonstrates a significant tendency of essential E. coli genes to be preserved throughout the bacterial kingdom. Projection of these data over metabolic subsystems reveals topologic modules with essential and evolutionarily preserved enzymes with reduced capacity for error tolerance.

Bacterial fatty acid biosynthesis: targets for antibacterial drug discovery.

The increase in drug-resistant pathogenic bacteria has created an urgent demand for new antibiotics. Among the more attractive targets for the development of new antibacterial compounds are the enzymes of fatty acid biosynthesis. Although a number of potent inhibitors of microbial fatty acid biosynthesis have been discovered, few of these are clinically useful drugs. Several of these fatty acid biosynthesis inhibitors have potential as lead compounds in the development of new antibacterials. This review encompasses the known inhibitors and prospective targets for new antibacterials.

Escherichia coli FadR positively regulates transcription of the fabB fatty acid biosynthetic gene.

In Escherichia coli expression of the genes of fatty acid degradation (fad) is negatively regulated at the transcriptional level by FadR protein. In contrast the unsaturated fatty acid biosynthetic gene, fabA, is positively regulated by FadR. We report that fabB, a second unsaturated fatty acid biosynthetic gene, is also positively regulated by FadR. Genomic array studies that compared global transcriptional differences between wild-type and fadR-null mutant strains, as well as in cultures of each strain grown in the presence of exogenous oleic acid, indicated that expression of fabB was regulated in a manner very similar to that of fabA expression. A series of genetic and biochemical tests confirmed these observations. Strains containing both fabB and fadR mutant alleles were constructed and shown to exhibit synthetic lethal phenotypes, similar to those observed in fabA fadR mutants. A fadR strain was hypersensitive to cerulenin, an antibiotic that at low concentrations specifically targets the FabB protein. A transcriptional fusion of chloramphenicol acetyltransferase (CAT) to the fabB promoter produces lower levels of CAT protein in a strain lacking functional FadR. The ability of a putative FadR binding site within the fabB promoter to form a complex with purified FadR protein was determined by a gel mobility shift assay. These experiments demonstrate that expression of fabB is positively regulated by FadR.

Structure of a phosphoglycerate mutase:3-phosphoglyceric acid complex at 1.7 A.

The crystal structure of the tetrameric glycolytic enzyme phosphoglycerate mutase from the yeast Saccharomyces cerevisiae has been determined to 1.7 A resolution in complex with the sugar substrate. The difference map indicates that 3-phosphoglycerate is bound at the base of a 12 A cleft, positioning C2 of the substrate within 3.5 A of the primary catalytic residue, histidine 8.

Evidence for the involvement of carbonic anhydrase and urease in calcium carbonate formation in the gravity-sensing organ of Aplysia californica.

To better understand the mechanisms that could modulate the formation of otoconia, calcium carbonate granules in the inner ear of vertebrate species, we examined statoconia formation in the gravity-sensing organ, the statocyst, of the gastropod mollusk Aplysia californica using an in vitro organ culture model. We determined the type of calcium carbonate present in the statoconia and investigated the role of carbonic anhydrase (CA) and urease in regulating statocyst pH as well as the role of protein synthesis and urease in statoconia production and homeostasis in vitro. The type of mineral present in statoconia was found to be aragonitic calcium carbonate. When the CA inhibitor, acetazolamide (AZ), was added to cultures of statocysts, the pH initially (30 min) increased and then decreased. The urease inhibitor, acetohydroxamic acid (AHA), decreased statocyst pH. Simultaneous addition of AZ and AHA caused a decrease in pH. Inhibition of urease activity also reduced total statoconia number, but had no effect on statoconia volume. Inhibition of protein synthesis reduced statoconia production and increased statoconia volume. In a previous study, inhibition of CA was shown to decrease statoconia production. Taken together, these data show that urease and CA play a role in regulating statocyst pH and the formation and maintenance of statoconia. CA produces carbonate ion for calcium carbonate formation and urease neutralizes the acid formed due to CA action, by production of ammonia.

Application of energy-resolved measurements to Laue diffraction: determination of unit-cell parameters, deconvolution of harmonics and assignment of systematic absences.

The use of energy-resolved area detection of Laue diffraction patterns for the determination of unit-cell parameters and systematic absences is demonstrated. Seven different crystals having previously known unit cells were re-examined using Laue diffraction methods. These crystals included four different crystal systems including cubic, orthorhombic, tetragonal and monoclinic cells. The crystals had cell sizes ranging from 179.4 to 4588.3 A(3). Comparison of known and re-determined cells showed good agreement (ratio of known to measured cells = 0.987 +/- 0.18). A single procedure was suitable for all unit-cell determinations. The accuracy of the method is presently limited by the quality of the available energy measurements. Some of the crystals represent space groups containing systematic absences normally obscured by harmonic overlap when using the Laue method. These include absences due to 2(1) screw axes (h, k or 1 = 2n + 1) and cell centering (h + k = 2n + 1). All systematic absences were identified using a combination of multiple linear regression with either stepwise elimination or stepwise inclusion and an F test for assignment of systematic absence. The methods are discussed in detail and simulations are used to evaluate critical tolerances for future systems.

Proliferation index as a predictor of prognosis in malignant gliomas of childhood.

BACKGROUND: The prognosis for children with high grade gliomas remains somewhat unpredictable because histologic features alone provide an imperfect assessment of the biologic behavior of a given lesion. Whereas some patients experience prolonged disease control after surgery and adjuvant therapy, others with lesions that appear comparable exhibit rapid disease progression and death. METHODS: Because proliferative activity may provide a potential correlate of biologic aggressiveness, the authors examined the relationship between MIB-1 labeling index and outcome in a series of 29 archival pediatric malignant nonbrainstem gliomas from patients treated consecutively at the study institution between 1975 and 1992, in which clinical, histologic, diagnostic, and therapeutic parameters were previously defined. Three patients who died perioperatively were excluded from outcome analyses. All tumors were rereviewed by two neuropathologists and classified as Grade 3 or 4 lesions based on contemporary guidelines. RESULTS: Among the specimens from the 26 patients who survived the perioperative period, a striking difference in outcome was apparent between tumors with MIB-1 indices < 12 (n = 10) and those with indices > 12 (n = 16). Median progression free survival was >48 months in the low MIB-1 group compared with only 6 months in the high MIB-1 group (P = 0.014, rank-sum test). Median overall survival was >48 months in the low MIB-1 group compared with only 16 months in the high MIB-1 group (P = 0.012). MIB-1 index remained associated with survival after taking into account the effect of resection extent, which also correlated strongly with outcome in this cohort. Although MIB-1 index was associated with histopathologic grade (Grade 3: 11.9 +/- 9.7 vs. Grade 4: 27.3 +/- 19.0; P = 0.015, Fisher's exact test), it proved to be a much stronger predictor of outcome than histology. CONCLUSIONS: MIB-1 index may supplement routine histologic classification as a means for improving the accuracy of predicting the biologic behavior of childhood malignant gliomas and may provide a basis for stratifying patients in future malignant glioma studies and refining therapeutic decision-making.

Morbidity and mortality of incontinence surgery in elderly women: an analysis of Medicare data.

OBJECTIVE: Our purpose was to examine length of hospital stay, complications, morbidity, and mortality associated with surgery for urinary incontinence in women > 65 years old. STUDY DESIGN: Data obtained from Medicare billing forms (HCFA form 1450, UB-82, UB-92) in the Medicare Provider Analysis and Review Record database for surgical procedures done for incontinence (international Classification of Diseases, Ninth Revision, codes 59.3 to 59.79) from 1984 to 1991 were reviewed. Comorbidities and reasons for readmission were extracted from secondary diagnosis codes. RESULTS: There wee 66,478 patients in the cohort. The mean and median ages were 71 years. The 30-day surgical mortality was 0.33%. Length of stay and mortality increased linearly with age. Acute events associated with death were myocardial infarction (14.2% of deaths), cerebrovascular accident (stroke) (14.6%), pulmonary embolism or deep vein thrombosis (9.7%), and pneumonia (2.7%). Patients who died had higher rates of diabetes and heart failure but not of hypertension. The 30-day readmission rate was 4.8% with higher rates in patients > 80 years old. Urinary tract infection (12.2%), hypertension (16%), and unspecified complications (9%) were the most frequent diagnoses on readmissions. Myocardial infarction, pulmonary embolism, cerebrovascular accident, deep vein thrombosis, and pneumonia each occurred in only 1%. CONCLUSION: Incontinence surgery is safe in the "young elderly." Those > 80 years old and with certain chronic illness should be counseled about increased risks.

The relationship between TP53 mutations and overexpression of p53 and prognosis in malignant gliomas of childhood.

The prognosis for children with high-grade gliomas remains somewhat unpredictable. Although prolonged disease control is sometimes achieved after surgery, radiotherapy, and chemotherapy, most patients exhibit rapid disease progression. Because p53-dependent apoptosis mechanisms are involved in the cytotoxic effects of irradiation and chemotherapy, we questioned whether p53 status might be associated with outcome in childhood malignant gliomas. Therefore, we examined p53 status, both immunohistochemically and by direct sequencing of exons 5-8, in a series of 29 archival pediatric malignant non-brainstem gliomas treated consecutively at our institution between 1975 and 1992. Eighteen tumors had dense p53 staining in the majority of cells, although only 11 had mutations of the p53 gene (TP53). On univariate analysis, there was a significant association between p53 overexpression and a shorter progression-free survival (PFS) and overall survival (OS; P = 0.019 and 0.013, respectively; rank sum test). In addition, there was a significant association between TP53 mutations and a poorer PFS (P = 0.04), and a strong trend toward a shorter OS among patients with TP53 mutations (P = 0.06). Median PFS and OS for patients with TP53-mutated tumors were 6 months and 16 months, respectively, and for those with p53 overexpression 5.5 months and 14 months, respectively, versus 16 months and 25 months, respectively, for those without TP53 mutations and 25 months and >4 years, respectively, for those without p53 overexpression. The percentage of patients in this series with TP53 mutations (37.9%) was substantially higher than in previous studies of childhood gliomas and comparable to the frequency of mutations noted in adult gliomas. However, both TP53 mutation and p53 overexpression were significantly less frequent in tumors from children younger than 4 than from older children (P = 0.02 and 0.01, respectively). These results indicate that p53 mutation and expression status may be associated with prognosis in childhood malignant gliomas, and thus may provide a basis for stratifying patients biologically in future malignant glioma studies.

Growth factors in gliomas: antisense and dominant negative mutant strategies.

Antisense and dominant negative mutant strategies were developed as 'magic bullets' to suppress the function of a particular gene while preserving the remaining cellular activities. While experience with these techniques has dispelled some of the 'magic', these strategies remain useful for understanding the function of particular gene products. Antisense strategies involve the administration of either a synthetic oligodeoxynucleotide or a plasmid construct which produces a sequence that is complementary to the DNA or mRNA of the gene of interest. Antisense binding should inhibit transcription or translation of the gene, and thus decrease synthesis of the protein for which the gene encodes. Conversely, dominant negative mutations inhibit activity of a gene product by encoding for a second protein which suppresses the function of the gene of interest. For example, a single mutant subunit in a multimeric protein might allow normal assembly of the protein while inhibiting its activity. The use of these techniques for investigating the role of various growth factor pathways in glial neoplasia and their potential therapeutic applications are reviewed below.

Basic fibroblast growth factor expression as a predictor of prognosis in pediatric high-grade gliomas.

Clinical and histopathological factors fail to adequately predict outcomes in children with high-grade gliomas, indicating a need to identify relevant biological markers of tumor behavior to guide therapeutic decision-making. Basic fibroblast growth factor (bFGF) is a mitogenic and angiogenic factor that has been observed to be overexpressed in a significant percentage of malignant gliomas, although the prognostic significance of this expression is unknown. To address this issue, the expression status of bFGF was examined immunohistochemically in a series of 27 archival pediatric malignant non-brainstem gliomas treated consecutively at our institution between 1975 and 1992. Tumors were categorized based on expression levels, and the association between expression status and outcome was examined. Sixteen cases showed high levels of expression of bFGF, and 11 showed low levels. There was no correlation between expression status and either tumor histology, patient age, or tumor location. However, there was a significant difference in outcome between patients with high levels of bFGF immunoreactivity and those with low expression. Median progression-free survival was >66 months in the low bFGF group as compared to 6 months in the high bFGF group (P = 0.006). Median overall survival was >66 months in the low bFGF group as compared to 18 months in the high bFGF group (P = 0.03). Tumor bFGF expression seems to be strongly associated with outcome in children with high-grade gliomas and, consequently, may serve as a biological correlate of patient prognosis in conjunction with other prognostic variables.

Cerebellar astrocytomas in children.

Cerebellar astrocytomas, as a group, carry a more favorable prognosis than most other brain tumors, because these neoplasms generally are histologically benign and amenable to extensive resection. However, it is clear that a number of factors have an impact on prognosis. In particular, resection extent has been strongly associated with progression-free survival: patients undergoing gross total resection appear to have a substantially better prognosis than those undergoing incomplete resection. Brainstem invasion, which is the factor that most often precludes a complete resection, has also been associated with a less favorable prognosis. In addition, histological features indicative of malignancy are clearly associated with a poor outcome. In contrast to the above observations, which have been established convincingly in the literature, a number of issues regarding cerebellar astrocytomas remain unresolved. First, the correlation between histology and prognosis among patients with low-grade cerebellar astrocytomas is uncertain: in some series, pilocytic astrocytomas have been associated with a better prognosis than non-pilocytic tumors, but in other studies, no such relationship has been observed. Second, the role of radiotherapy after incomplete resection of a low-grade cerebellar astrocytoma remains problematic. In view of the lack of convincing data in this regard, many groups, including our own, defer radiotherapy until there is evidence of progressive disease that is surgically unresectable. Finally, the frequency of follow-up in patients with cerebellar astrocytomas remains largely empirical. Although most recurrences are detected within a few years after initial surgery, late recurrences are well known, which raises the question of when and if such patients should be regarded as "cured' of their disease. Long-term multi-institutional natural history studies are in progress to address the above issues.

High-grade astrocytomas in children: radiologically complete resection is associated with an excellent long-term prognosis.

High-grade astrocytomas comprise approximately 10% of intracranial tumors in children. A better prognosis in children than in adults has been reported for patients with these neoplasms, although the reasons for this survival advantage are uncertain. To determine whether any consistent factors were associated with long-term survival, we reviewed the records of 31 children with high-grade non-brain stem gliomas who were treated at our institution between 1975 and 1992. Histology was reviewed and classified according to the World Health Organization scheme, and neuroimaging studies were examined to determine the extent of resection, in both instances by individuals who were unaware of the patients' outcomes. The median overall survival for the 28 patients who survived the perioperative period was 18.5 months; 10 (36%) are currently alive, with a median follow-up of 70.5 months. The median progression-free survival (PFS) was 10.5 months; eight patients (29%) remain progression-free with a median follow-up of 78 months. The extent of resection at initial operation was associated most closely with PFS and overall survival as revealed by multivariate analysis. The 14 patients who underwent subtotal (< 90%) resection and the 7 who underwent near-total (90-99%) resection had median PFS of 5.5 and 11 months, respectively (P = 0.38), and overall survival of 10.5 and 25 months, respectively (P = 0.02). None of the seven patients who underwent gross total removal of tumor as confirmed by postoperative imaging had disease progression, with a median follow-up of 84 months (P < 0.0001). All of the tumors that underwent gross total resection were situated within the cerebral hemispheres; five of the seven patients had seizures as a presenting symptom. Both tumor location and seizures were significantly associated with outcome as determined by univariate analysis, but because of the overwhelming impact of resection extent on outcome, these factors were not independently associated with outcome as revealed by multivariate analysis. Histology was associated with outcome in the subgroup of patients with incompletely resected hemispheric tumors, in which children with anaplastic astrocytoma had a significantly better PFS than those with glioblastoma multiforme (P = 0.009). In summary, our results support the role of cytoreductive surgery in the treatment of cerebral hemispheric high-grade astrocytomas in children, which may encompass a biologically distinct group of tumors that, by virtue of their location and growth characteristics, are amenable to aggressive resection. The prognosis for children with deep-seated lesions and for those with subtotally resected hemispheric lesions is generally poor with conventional therapy.