Agronomic Investigation of Spray Dispersion of Metal-Based Nanoparticles on Sunflowers in Real-World Environments.

In environmental and agronomic settings, even minor imbalances can trigger a range of unpredicted responses. Despite the widespread use of metal-based nanoparticles (NPs) and new bio-nanofertilizers, their impact on crop production is absent in the literature. Therefore, our research is focused on the agronomic effect of spray application of gold nanoparticles anchored to SiO2 mesoporous silica (AuSi-NPs), zinc oxide nanoparticles (ZnO-NPs), and iron oxide nanoparticles (Fe3O4-NPs) on sunflowers under real-world environments. Our findings revealed that the biosynthetically prepared AuSi-NPs and ZnO-NPs were highly effective in enhancing sunflower seasonal physiology, e.g., the value of the NDVI index increased from 0.012 to 0.025 after AuSi-NPs application. The distribution of leaf trichomes improved and the grain yield increased from 2.47 t ha-1 to 3.29 t ha-1 after ZnO-NPs application. AuSi-NPs treatment resulted in a higher content of essential linoleic acid (54.37%) when compared to the NPs-free control (51.57%), which had a higher determined oleic acid. No NPs or residual translocated metals were detected in the fully ripe sunflower seeds, except for slightly higher silica content after the AuSi-NPs treatment. Additionally, AuSi-NPs and NPs-free control showed wide insect biodiversity while ZnO-NPs treatment had the lowest value of phosphorus as anti-nutrient. Contradictory but insignificant effect on physiology, yield, and insect biodiversity was observed in Fe3O4-NPs treatment. Therefore, further studies are needed to fully understand the long-term environmental and agricultural sustainability of NPs applications.

A learning experience elicits sex-dependent neurogenomic responses in Bicyclus anynana butterflies.

Sexually dimorphic behaviour is pervasive across animals, with males and females exhibiting different mate selection, parental care, foraging, dispersal, and territorial strategies. However, the genetic underpinnings of sexually dimorphic behaviours are poorly understood. Here we investigate gene networks and expression patterns associated with sexually dimorphic imprinting-like learning in the butterfly Bicyclus anynana. In this species, both males and females learn visual preferences, but learn preferences for different traits and use different signals as salient, unconditioned cues. To identify genes and gene networks associated with this behaviour, we examined gene expression profiles of the brains and eyes of male and female butterflies immediately post training and compared them to the same tissues of naïve individuals. We found more differentially expressed genes and a greater number of associated gene networks in the eyes, indicating a role of the peripheral nervous system in visual imprinting-like learning. Females had higher chemoreceptor expression levels than males, supporting the hypothesized sexual dimorphic use of chemical cues during the learning process. In addition, genes that influence B. anynana wing patterns (sexual ornaments), such as invected, spalt, and apterous, were also differentially expressed in the brain and eye, suggesting that these genes may influence both sexual ornaments and the preferences for these ornaments. Our results indicate dynamic and sex-specific responses to social scenario in both the peripheral and central nervous systems and highlight the potential role of wing patterning genes in mate preference and learning across the Lepidoptera.

Effects of Foliar Application of ZnO Nanoparticles on Lentil Production, Stress Level and Nutritional Seed Quality under Field Conditions.

Nanotechnology offers new opportunities for the development of novel materials and strategies that improve technology and industry. This applies especially to agriculture, and our previous field studies have indicated that zinc oxide nanoparticles provide promising nano-fertilizer dispersion in sustainable agriculture. However, little is known about the precise ZnO-NP effects on legumes. Herein, 1 mg·L-1 ZnO-NP spray was dispersed on lentil plants to establish the direct NP effects on lentil production, seed nutritional quality, and stress response under field conditions. Although ZnO-NP exposure positively affected yield, thousand-seed weight and the number of pods per plant, there was no statistically significant difference in nutrient and anti-nutrient content in treated and untreated plant seeds. In contrast, the lentil water stress level was affected, and the stress response resulted in statistically significant changes in stomatal conductance, crop water stress index, and plant temperature. Foliar application of low ZnO-NP concentrations therefore proved promising in increasing crop production under field conditions, and this confirms ZnO-NP use as a viable strategy for sustainable agriculture.

Stage- and sex-specific transcriptome analyses reveal distinctive sensory gene expression patterns in a butterfly.

BACKGROUND: Animal behavior is largely driven by the information that animals are able to extract and process from their environment. However, the function and organization of sensory systems often change throughout ontogeny, particularly in animals that undergo indirect development. As an initial step toward investigating these ontogenetic changes at the molecular level, we characterized the sensory gene repertoire and examined the expression profiles of genes linked to vision and chemosensation in two life stages of an insect that goes through metamorphosis, the butterfly Bicyclus anynana. RESULTS: Using RNA-seq, we compared gene expression in the heads of late fifth instar larvae and newly eclosed adults that were reared under identical conditions. Over 50 % of all expressed genes were differentially expressed between the two developmental stages, with 4,036 genes upregulated in larval heads and 4,348 genes upregulated in adult heads. In larvae, upregulated vision-related genes were biased toward those involved with eye development, while phototransduction genes dominated the vision genes that were upregulated in adults. Moreover, the majority of the chemosensory genes we identified in the B. anynana genome were differentially expressed between larvae and adults, several of which share homology with genes linked to pheromone detection, host plant recognition, and foraging in other species of Lepidoptera. CONCLUSIONS: These results revealed promising candidates for furthering our understanding of sensory processing and behavior in the disparate developmental stages of butterflies and other animals that undergo metamorphosis.

Foliar Application of Low Concentrations of Titanium Dioxide and Zinc Oxide Nanoparticles to the Common Sunflower under Field Conditions.

Nano-fertilisers have only recently been introduced to intensify plant production, and there still remains inadequate scientific knowledge on their plant-related effects. This paper therefore compares the effects of two nano-fertilisers on common sunflower production under field conditions. The benefits arising from the foliar application of micronutrient-based zinc oxide fertiliser were compared with those from the titanium dioxide plant-growth enhancer. Both the zinc oxide (ZnO) and titanium dioxide (TiO2) were delivered by foliar application in nano-size at a concentration of 2.6 mg·L-1. The foliar-applied nanoparticles (NPs) had good crystallinity and a mean size distribution under 30 nm. There were significant differences between these two experimental treatments in the leaf surfaces' trichomes diversity, ratio, width, and length at the flower-bud development stage. Somewhat surprisingly, our results established that the ZnO-NPs treatment induced generally better sunflower physiological responses, while the TiO2-NPs primarily affected quantitative and nutritional parameters such as oil content and changed sunflower physiology to early maturation. There were no differences detected in titanium or zinc translocation or accumulation in the fully ripe sunflower seeds compared to the experimental controls, and our positive results therefore encourage further nano-fertiliser research.

Pulse magnetization elicits differential gene expression in the central nervous system of the Caribbean spiny lobster, Panulirus argus.

Diverse animals use Earth's magnetic field to guide their movements, but the neural and molecular mechanisms underlying the magnetic sense remain enigmatic. One hypothesis is that particles of the mineral magnetite (Fe3O4) provide the basis of magnetoreception. Here we examined gene expression in the central nervous system of a magnetically sensitive invertebrate, the Caribbean spiny lobster (Panulirus argus), after applying a magnetic pulse known to alter magnetic orientation behavior. Numerous genes were differentially expressed in response to the pulse, including 647 in the brain, 1256 in the subesophageal ganglion, and 712 in the thoracic ganglia. Many such genes encode proteins linked to iron regulation, oxidative stress, and immune response, consistent with possible impacts of a magnetic pulse on magnetite-based magnetoreceptors. Additionally, however, altered expression also occurred for numerous genes with no apparent link to magnetoreception, including genes encoding proteins linked to photoreception, carbohydrate and hormone metabolism, and other physiological processes. Overall, the results are consistent with the magnetite hypothesis of magnetoreception, yet also reveal that in spiny lobsters, a strong pulse altered expression of > 10% of all expressed genes, including many seemingly unrelated to sensory processes. Thus, caution is required when interpreting the effects of magnetic pulses on animal behavior.

Effect of Foliar Spray Application of Zinc Oxide Nanoparticles on Quantitative, Nutritional, and Physiological Parameters of Foxtail Millet (Setaria italica L.) under Field Conditions.

It has been shown that the foliar application of inorganic nano-materials on cereal plants during their growth cycle enhances the rate of plant productivity by providing a micro-nutrient source. We therefore studied the effects of foliarly applied ZnO nanoparticles (ZnO NPs) on Setaria italica L. foxtail millet's quantitative, nutritional, and physiological parameters. Scanning electron microscopy showed that the ZnO NPs have an average particle size under 20 nm and dominant spherically shaped morphology. Energy dispersive X-ray spectrometry then confirmed ZnO NP homogeneity, and X-ray diffraction verified their high crystalline and wurtzite-structure symmetry. Although plant height, thousand grain weight, and grain yield quantitative parameters did not differ statistically between ZnO NP-treated and untreated plants, the ZnO NP-treated plant grains had significantly higher oil and total nitrogen contents and significantly lower crop water stress index (CWSI). This highlights that the slow-releasing nano-fertilizer improves plant physiological properties and various grain nutritional parameters, and its application is therefore especially beneficial for progressive nanomaterial-based industries.

Near absence of differential gene expression in the retina of rainbow trout after exposure to a magnetic pulse: implications for magnetoreception.

The ability to perceive the Earth's magnetic field, or magnetoreception, exists in numerous animals. Although the mechanism underlying magnetoreception has not been clearly established in any species, in salmonid fish, it is hypothesized to occur by means of crystals of magnetite associated with nervous tissue such as the brain, olfactory organ or retina. In this study, rainbow trout (Oncorhynchus mykiss) were exposed to a brief magnetic pulse known to disrupt magnetic orientation behaviour in several animals. Changes in gene expression induced by the pulse were then examined in the retina. Analyses indicated that the pulse elicited differential expression of only a single gene, gamma-crystallin M3-like (crygm3). The near absence of an effect of the magnetic pulse on gene expression in the retina stands in sharp contrast to a recent study in which 181 genes were differentially expressed in brain tissue of O. mykiss after exposure to the same pulse. Overall, our results suggest either that magnetite-based magnetoreceptors in trout are not located in the retina, or else that they are unaffected by magnetic pulses that can disrupt magnetic orientation behaviour in animals.

Size-dependent avoidance of a strong magnetic anomaly in Caribbean spiny lobsters.

On a global scale, the geomagnetic field varies predictably across the Earth's surface, providing animals that migrate long distances with a reliable source of directional and positional information that can be used to guide their movements. In some locations, however, magnetic minerals in the Earth's crust generate an additional field that enhances or diminishes the overall field, resulting in unusually steep gradients of field intensity within a limited area. How animals respond to such magnetic anomalies is unclear. The Caribbean spiny lobster, Panulirus argus, is a benthic marine invertebrate that possesses a magnetic sense and is likely to encounter magnetic anomalies during migratory movements and homing. As a first step toward investigating whether such anomalies affect the behavior of lobsters, a two-choice preference experiment was conducted in which lobsters were allowed to select one of two artificial dens, one beneath a neodymium magnet and the other beneath a non-magnetic weight of similar size and mass (control). Significantly more lobsters selected the control den, demonstrating avoidance of the magnetic anomaly. In addition, lobster size was found to be a significant predictor of den choice: lobsters that selected the anomaly den were significantly smaller as a group than those that chose the control den. Taken together, these findings provide additional evidence for magnetoreception in spiny lobsters, raise the possibility of an ontogenetic shift in how lobsters respond to magnetic fields, and suggest that magnetic anomalies might influence lobster movement in the natural environment.

Candidate genes mediating magnetoreception in rainbow trout (Oncorhynchus mykiss).

Diverse animals use Earth's magnetic field in orientation and navigation, but little is known about the molecular mechanisms that underlie magnetoreception. Recent studies have focused on two possibilities: (i) magnetite-based receptors; and (ii) biochemical reactions involving radical pairs. We used RNA sequencing to examine gene expression in the brain of rainbow trout (Oncorhynchus mykiss) after exposure to a magnetic pulse known to disrupt magnetic orientation behaviour. We identified 181 differentially expressed genes, including increased expression of six copies of the frim gene, which encodes a subunit of the universal iron-binding and trafficking protein ferritin. Functions linked to the oxidative effects of free iron (e.g. oxidoreductase activity, transition metal ion binding, mitochondrial oxidative phosphorylation) were also affected. These results are consistent with the hypothesis that a magnetic pulse alters or damages magnetite-based receptors and/or other iron-containing structures, which are subsequently repaired or replaced through processes involving ferritin. Additionally, some genes that function in the development and repair of photoreceptive structures (e.g. crggm3, purp, prl, gcip, crabp1 and pax6) were also differentially expressed, raising the possibility that a magnetic pulse might affect structures and processes unrelated to magnetite-based magnetoreceptors.

Effect of magnetic pulses on Caribbean spiny lobsters: implications for magnetoreception.

The Caribbean spiny lobster, Panulirus argus, is a migratory crustacean that uses Earth's magnetic field as a navigational cue, but how these lobsters detect magnetic fields is not known. Magnetic material thought to be magnetite has previously been detected in spiny lobsters, but its role in magnetoreception, if any, remains unclear. As a first step toward investigating whether lobsters might have magnetite-based magnetoreceptors, we subjected lobsters to strong, pulsed magnetic fields capable of reversing the magnetic dipole moment of biogenic magnetite crystals. Lobsters were subjected to a single pulse directed from posterior to anterior and either: (1) parallel to the horizontal component of the geomagnetic field (i.e. toward magnetic north); or (2) antiparallel to the horizontal field (i.e. toward magnetic south). An additional control group was handled but not subjected to a magnetic pulse. After treatment, each lobster was tethered in a water-filled arena located within 200 m of the capture location and allowed to walk in any direction. Control lobsters walked in seemingly random directions and were not significantly oriented as a group. In contrast, the two groups exposed to pulsed fields were significantly oriented in approximately opposite directions. Lobsters subjected to a magnetic pulse applied parallel to the geomagnetic horizontal component walked westward; those subjected to a pulse directed antiparallel to the geomagnetic horizontal component oriented approximately northeast. The finding that a magnetic pulse alters subsequent orientation behavior is consistent with the hypothesis that magnetoreception in spiny lobsters is based at least partly on magnetite-based magnetoreceptors.

Multi-Modal Homing in Sea Turtles: Modeling Dual Use of Geomagnetic and Chemical Cues in Island-Finding.

Sea turtles are capable of navigating across large expanses of ocean to arrive at remote islands for nesting, but how they do so has remained enigmatic. An interesting example involves green turtles (Chelonia mydas) that nest on Ascension Island, a tiny land mass located approximately 2000 km from the turtles' foraging grounds along the coast of Brazil. Sensory cues that turtles are known to detect, and which might hypothetically be used to help locate Ascension Island, include the geomagnetic field, airborne odorants, and waterborne odorants. One possibility is that turtles use magnetic cues to arrive in the vicinity of the island, then use chemical cues to pinpoint its location. As a first step toward investigating this hypothesis, we used oceanic, atmospheric, and geomagnetic models to assess whether magnetic and chemical cues might plausibly be used by turtles to locate Ascension Island. Results suggest that waterborne and airborne odorants alone are insufficient to guide turtles from Brazil to Ascension, but might permit localization of the island once turtles arrive in its vicinity. By contrast, magnetic cues might lead turtles into the vicinity of the island, but would not typically permit its localization because the field shifts gradually over time. Simulations reveal, however, that the sequential use of magnetic and chemical cues can potentially provide a robust navigational strategy for locating Ascension Island. Specifically, one strategy that appears viable is following a magnetic isoline into the vicinity of Ascension Island until an odor plume emanating from the island is encountered, after which turtles might either: (1) initiate a search strategy; or (2) follow the plume to its island source. These findings are consistent with the hypothesis that sea turtles, and perhaps other marine animals, use a multi-modal navigational strategy for locating remote islands.

Multiparameter flow cytometry: advances in high resolution analysis.

Over the past 40 years, flow cytometry has emerged as a leading, application-rich technology that supports high-resolution characterization of individual cells which function in complex cellular networks such as the immune system. This brief overview highlights advances in multiparameter flow cytometric technologies and reagent applications for characterization and functional analysis of cells modulating the immune network. These advances significantly support high-throughput and high-content analyses and enable an integrated understanding of the cellular and molecular interactions that underlie complex biological systems.

Single and coexpression of CXCR4 and CXCR5 identifies CD4 T helper cells in distinct lymph node niches during influenza virus infection.

Influenza virus infection results in strong, mainly T-dependent, extrafollicular and germinal center B cell responses, which provide lifelong humoral immunity against the homotypic virus strain. Follicular T helper cells (T(FH)) are key regulators of humoral immunity. Questions remain regarding the presence, identity, and function of T(FH) subsets regulating early extrafollicular and later germinal center B cell responses. This study demonstrates that ICOS but not CXCR5 marks T cells with B helper activity induced by influenza virus infection and identifies germinal center T cells (T(GC)) as lymph node-resident CD4(+) ICOS(+) CXCR4(+) CXCR5(+) PSGL-1(lo) PD-1(hi) cells. The CXCR4 expression intensity further distinguished their germinal center light and dark zone locations. This population emerged strongly in regional lymph nodes and with kinetics similar to those of germinal center B cells and were the only T(FH) subsets missing in influenza virus-infected, germinal center-deficient SAP(-/-) mice, mice which were shown previously to lack protective memory responses after a secondary influenza virus challenge, thus indicting the nonredundant functions of CXCR4- and CXCR5-coexpressing CD4 helper cells in antiviral B cell immunity. CXCR4-single-positive T cells, present in B cell-mediated autoimmunity and regarded as "extrafollicular" helper T cells, were rare throughout the response, despite prominent extrafollicular B cell responses, revealing fundamental differences in autoimmune- and infection-induced T-dependent B cell responses. While all ICOS(+) subsets induced similar antibody levels in vitro, CXCR5-single-positive T cells were superior in inducing B cell proliferation. The regulation of T cell localization, marked by the single and coexpression of CXCR4 and CXCR5, might be an important determinant of T(FH) function.

Structural characteristics of small intestinal submucosa constructs dictate in vivo incorporation and angiogenic response.

The rate of angiogenesis and cellular infiltration into degradable biomaterials determines scaffold persistence in vivo. The ability to tune the degradation properties of naturally derived biomaterials has been a popular goal in tissue engineering, yet has often depended on chemical crosslinking. Small intestinal submucosa (SIS) is a naturally derived, collagen-based, bioactive scaffold that has broad clinical success in many therapeutic applications. Two methods for producing multilayer, non-crosslinked SIS constructs were compared in vitro and in vivo. Traditional and cryo SEM, mercury intrusion porosimetry, and a novel enzymatic degradation assay determined that lyophilization produced an open, porous scaffold, in contrast to the collapsed, denser structure of SIS constructs produced using a vacuum press process. The angiogenic responses to lyophilized and vacuum-pressed SIS constructs were evaluated in vivo using a subcutaneous implant assay in mice. Explanted samples were compared after 7 and 21 days using fluorescence microangiography and light microscopy. Capacity of the implant neovasculature was also determined. These experiments revealed that the lyophilized SIS was infiltrated and vascularized more rapidly than the vacuum pressed. These data demonstrate the tunable incorporation of a non-crosslinked ECM-based biomaterial, which may have implications for the persistence of this degradable scaffold in tissue engineering.

Absorption of bioactive molecules into OASIS wound matrix.

OBJECTIVE: To examine the ability of OASIS Wound Matrix to absorb, retain, and protect bioactive molecules from solution. DESIGN: Samples of OASIS Wound Matrix were incubated in solutions of bioactive molecules, specifically heparin, albumin, fibronectin, basic fibroblast growth factor 2, and platelet-derived growth factor (PDGF). Half of the samples were then rinsed, and all of the samples were evaluated using enzyme-linked immunosorbent assays (ELISAs) and dye-mediated spectrophotometric methods for absorption and retention of the bioactive molecules. Protection of PDGF was measured by placing PDGF-incubated and control samples into a degradation solution containing plasmin. Intact PDGF levels were then evaluated using a PDGF-specific ELISA. MAIN OUTCOME MEASURES: The main outcome measures were the amount of each bioactive molecule that was absorbed after incubation in solutions and retained after rinses as well as the amount of PDGF remaining after plasmin degradation. MAIN RESULTS: OASIS Wound Matrix absorbed bioactive molecules from solution, selectively absorbed PDGF from serum, and protected PDGF from protease degradation. CONCLUSIONS: Although OASIS Wound Matrix potentially has multiple functions in wound healing, it likely promotes wound healing, in part, by absorbing, retaining, and protecting bioactive molecules from the wound environment.

Human ABCB6 localizes to both the outer mitochondrial membrane and the plasma membrane.

Expression of the ATP-binding cassette transporter ABCB6 has been associated with multiple cellular functions, including resistance to several cytotoxic agents, iron homeostasis, and porphyrin transport. To further elucidate its physiological function and/or role in drug resistance, we determined the subcellular location of ABCB6. Using three novel ABCB6-specific antibodies, Western blot analysis of cells expressing cDNA-derived or endogenous ABCB6 revealed two distinct molecular weight forms. Confocal microscopy indicates that the protein localizes to both mitochondria and the plasma membrane. Differential centrifugation revealed that the lower molecular weight form predominantly resides in the mitochondria, while the larger protein form is more abundant in the plasma membrane. Preliminary studies indicate that ABCB6 is functionally relevant in the plasma membrane, where its expression prevents the accumulation of specific porphyrins in the cell. Digitonin solubilization of mitochondria demonstrated that ABCB6 is present in the outer mitochondrial membrane, while back-titration assays with the ABCB6-specific antibodies reveal that the nucleotide binding domain of ABCB6 is cytoplasmic. These studies are the first to demonstrate that ABCB6 exists in two molecular weight forms, is localized to both the outer mitochondrial membrane and the plasma membrane, and plays a functional role in the plasma membrane.

Effects of sterilization on an extracellular matrix scaffold: part I. Composition and matrix architecture.

The impact of peracetic acid (PAA), lyophilization, and ethylene oxide (EO) sterilization on the composition and three dimensional matrix structure of small intestinal submucosa (SIS), a biologic scaffold used to stimulate the repair of damaged tissues and organs, was examined. Fibronectin and glycosaminoglycans are retained in SIS following oxidation by peracetic acid and alkylation using ethylene oxide gas. Significant amounts of FGF-2 are also retained, but VEGF is susceptible to the effects of PAA and is dramatically reduced following processing. Further, matrix oxidation, lyophilization, and sterilization with EO can be performed without irreversibly collapsing the three dimensional structure of the native SIS. These structural features and growth promoting extracellular matrix constituents are likely to be important variables underlying cellular attachment, infiltration and eventual incorporation of SIS into healing host tissues.

Tensile strength comparison of small intestinal submucosa body wall repair.

BACKGROUND: Porcine small intestinal submucosa (SIS) has been studied for body wall repair. However, the best method to implant the biological material has not been investigated. The objective of this study was to compare tensile strengths achieved after healing when SIS was placed using three implant techniques (onlay, inlay, underlay) in a porcine model of abdominal wall defect. MATERIALS AND METHODS: Twenty female domestic pigs had three abdominal midline sites assigned to one of five test groups: SIS implantation using inlay, onlay, or underlay technique; sham surgery (sutured midline incision) or normal body-wall control. Full-thickness muscle/fascia midline abdominal defects (6 x 4 cm) were surgically created and then repaired using eight-layer SIS. Healing was evaluated at 1 and 4 post-operative months by tensile strength testing and histopathology. RESULTS: Hernias were not observed. Tensile strengths were not statistically different between the five test groups (P = 0.39) or between months 1 and 4 (P = 0.35). The caudal site was stronger than the cranial or middle sites in the 1 month group (P < 0.0001). Histologically, healing appeared to progress over time as the repair site showed remodeling towards an interlacing fibrous connective tissue pattern. CONCLUSIONS: No significant differences in tensile strength were found between implant techniques and were not statistically different from sham surgery and normal control tissue. This study suggested that SIS healing/remodeling provides sufficient tensile strength for the repair of ventral (anterior) abdominal wall defects when implanted using any of three common techniques.

Your alliances are too stable.

A 2004 McKinsey survey of more than 30 companies reveals that at least 70% of them have major alliances that are underperforming and in need of restructuring. Moreover, JVs that broaden or otherwise adjust their scope have a 79% success rate, versus 33% for ventures that remain essentially unchanged. Yet most firms don't routinely evaluate the need to overhaul their alliances or intervene to correct performance problems. That means corporations are missing huge opportunities: By revamping just one large alliance, a company can generate 100 million dololars to 300 million dollars in extra income a year. Here's how to unlock more value from alliances: (1) Launch the process. Don't wait until your venture is in the middle of a crisis; regularly scan your major alliances to determine which need restructuring. Once you've targeted one, designate a restructuring team and find a senior sponsor to push the process along. Then delineate the scope of the team's work. (2) Diagnose performance. Evaluate the venture on the following performance dimensions: ownership and financials, strategy, operations, governance, and organization and talent. Identify the root causes of the venture's problems, not just the symptoms, and estimate how much each problem is costing the company. (3) Generate restructuring options. Based on the diagnosis, decide whether to fix, grow, or exit the alliance. Assuming the answer is fix or grow, determine whether fundamental or incremental changes are needed, using the five performance dimensions above as a framework. Then assemble three or four packages of restructuring options, test them with shareholders, and gain parents' approval. (4) Execute the changes. Embark on a widespread and consistent communication effort, building support among executives in the JV and the parent companies. So the process stays on track, assign accountability to certain groups or individuals.