Regulating biocontrol agents: a historical perspective and a critical examination comparing microbial and macrobial agents

Ever since the inclusion of microbial biocontrol agents (MBCAs) within the regulatory frameworks initially designed for chemical pesticides, there has been awareness that these frameworks are not optimal for assessment and registration of new microbial biocontrol products. It is often claimed that the regulatory situation has contributed to a relatively slow uptake of microbial biocontrol in practice. In contrast to the MBCAs, non-indigenous invertebrate biocontrol agents (IBCAs) are regulated in many countries through quarantine and other biosecurity related legislation for prevention of introduction of alien organisms, whereas use of indigenous IBCAs are generally unregulated. In this study, we investigate what scientific support there is for performing evaluations of these two main groups of biocontrol agents (BCAs) within different frameworks. We compare potential risks of MBCAs and IBCAs, present a retrospective analysis of the development and implementation of the regulatory frameworks, and compare current requirements for MBCAs with those for other applications with microorganisms. One conclusion is that the ecological risks are of similar types between the two groups of BCAs, and that for both groups the environmental safety is most pertinently evaluated according to biological and ecological principles. The main difference between MBCAs and IBCAs with respect to human health is that the former may cause infectious disease. However, we found no evidence that this hazard is more serious for microorganisms for biocontrol than for microbes used in other types of applications, which generally have substantially lower regulatory demands than those for MBCAs. Several international initiatives have produced helpful guidelines and recommendations for simplified assessments and authorisations of BCAs. Still, we conclude that as long as MBCAs are evaluated within systems initially developed for chemicals, the risk for inappropriate emphasis of chemical hazards and therefore unnecessarily complicated assessments will be maintained. Therefore, this study supports the idea that development of new systems for the regulatory oversight of MBCAs, possibly a mutual framework covering all living BCAs, should be considered. Research issues that need to be further explored are to what extent utilisation of MBCAs actually results in increased exposure of non-targets to microorganisms, the biogeography and microbial ecology of representative MBCAs, and finally development of better methodology for determining potential human toxicity and pathogenicity of candidate MBCAs.     

Crystal Structures of the Human G3BP1 NTF2-Like Domain Visualize FxFG Nup Repeat Specificity

Ras GTPase Activating Protein SH3 Domain Binding Protein (G3BP) is a potential anti-cancer drug target implicated in several cellular functions. We have used protein crystallography to solve crystal structures of the human G3BP1 NTF2-like domain both alone and in complex with an FxFG Nup repeat peptide. Despite high structural similarity, the FxFG binding site is located between two alpha helices in the G3BP1 NTF2-like domain and not at the dimer interface as observed for nuclear transport factor 2. ITC studies showed specificity towards the FxFG motif but not FG and GLFG motifs. The unliganded form of the G3BP1 NTF2-like domain was solved in two crystal forms to resolutions of 1.6 and 3.3 Å in space groups P2₁2₁2₁ and P6₃22 based on two different constructs, residues 1–139 and 11–139, respectively. Crystal packing of the N-terminal residues against a symmetry related molecule in the P2₁2₁2₁ crystal form might indicate a novel ligand binding site that, however, remains to be validated. The crystal structures give insight into the nuclear transportation mechanisms of G3BP and provide a basis for future structure based drug design.     

Effects of a transient oxic period on mineralization of organic matter to CH4 and CO2 in anoxic peat incubations

Rates of organic matter mineralization in peatlands, and hence production of the greenhouse gases CH₄ and CO₂, are highly dependent on the distribution of oxygen in the peat. Using laboratory incubations of peat, we investigated the sensitivity of the anoxic production of CH₄ and CO₂ to a transient oxic period of a few weeks’ duration. Production rates during 3 successive anoxic periods were compared with rates in samples incubated in the presence of oxygen during the second period. In surface peat (5–10‐cm depth), with an initially high level of CH₄ production, oxic conditions during period 2 did not result in a lower potential CH₄ production rate during period 3, although production was delayed ～1 week. In permanently anoxic, deep peat (50–55‐cm depth) with a comparatively low initial production of CH₄, oxic conditions during period 2 resulted in zero production of CH₄ during period 3. Thus, the methanogens in surface peal—but not in deep peat—remained viable after several weeks of oxic conditions. In contrast to CH₄ production, the oxic period had a negligible effect on anoxic CO₂ production during period 3, in surface as well as deep peat. In both surface and deep peat, CO₂ production was several times higher under oxic than under anoxic conditions. However, for the first 2 weeks of oxic conditions, CO₂ production in the deep peat was very low. Still, deep peat obviously contained facultative microorganisms that, after a relatively short period, were able to maintain a considerably higher rate of organic matter mineralization under oxic than under anoxic conditions.     

Structure motif discovery and mining the PDB

MOTIVATION: Many of the most interesting functional and evolutionary relationships among proteins are so ancient that they cannot be reliably detected through sequence analysis and are apparent only through a comparison of the tertiary structures. The conserved features can often be described as structural motifs consisting of a few single residues or Secondary Structure (SS) elements. Confidence in such motifs is greatly boosted when they are found in more than a pair of proteins. RESULTS: We describe an algorithm for the automatic discovery of recurring patterns in protein structures. The patterns consist of individual residues having a defined order along the protein's backbone that come close together in the structure and whose spatial conformations are similar. The residues in a pattern need not be close in the protein's sequence. The work described in this paper builds on an earlier reported algorithm for motif discovery. This paper describes a significant improvement of the algorithm which makes it very efficient. The improved efficiency allows us to use it for doing unsupervised learning of patterns occurring in small subsets in a large set of structures, a non-redundant subset of the Protein Data Bank (PDB) database of all known protein structures.     

Uptake of foreign ferritin in platy Xiphophorus maculatus (Poeciliidae: Teleostei)

The ability and capacity of various tissues in platy Xiphophorus maculatus L. to take up horse-spleen ferritin injected into the blood stream are described. Ferritin was injected intraperioneally, and the cellular uptake was demonstrated as Prussian blue precipitations in tissues treated with acid ferrocyanide solutions. Ferritin was detected within the heart endocardial cells and macrophages in the trunk kidney and spleen, 1/4 h after the injection, i.e. foreign ferritin was taken up very rapidly by these cells. When the time elapsed between the ferritin injection and sacrifice exceeded 6 h, these cells, and also macrophages in the gill and intestine, were almost completely filled with ferritin. At these stages, however, the amounts of Prussian blue precipitations per volume unit of the tissue were much larger in the heart than in the other organs studied in the present work, i.e. the endocardial tissue seems to play an important role in the clearance of the blood circulation in this species. We suggest that this tissue in platy is specialized to endocytose waste and foreign macromolecules, including pathogenic particles, from the blood stream. The eosinophilic and neutrophilic granulocytes do not appear to take up foreign ferritin, i.e. these cells may play no endocytotic role in the clearance of foreign macromolecules in platy.     

The clinicopathological and gene expression patterns associated with ulceration of primary melanoma

Ulceration of primary melanomas is associated with poor prognosis yet is reported to predict benefit from adjuvant interferon. To better understand the biological processes involved, clinicopathological factors associated with ulceration were determined in 1804 patients. From this cohort, 348 primary tumor blocks were sampled to generate gene expression data using a 502‐gene cancer panel and 195 blocks were used for immunohistochemistry to detect macrophage infiltration and vessel density. Gene expression results were validated using a whole genome array in two independent sample sets. Ulceration of primary melanomas was associated with more proliferative tumors, tumor vessel invasion, and increased microvessel density. Infiltration of tumors with greater number of macrophages and gene expression pathways associated with wound healing and up‐regulation of pro‐inflammatory cytokines suggests that ulceration is associated with tumor‐related inflammation. The relative benefit from interferon reported in patients with ulcerated tumors may reflect modification of signaling pathways involved in inflammation.     

Time Trends in Incidence and Mortality of Acute Myocardial Infarction,and All-Cause Mortality following a Cardiovascular Prevention Program in Sweden

Background

In 1988, a cardiovascular prevention program which combined an individual and a population-based strategy was launched within primary health-care in Sollentuna, a municipality in Stockholm County. The aim of this study was to investigate time trends in the incidence of and mortality from acute myocardial infarction and all-cause mortality in Sollentuna compared with the rest of Stockholm County during a period of two decades following the implementation of a cardiovascular prevention program.

Materials and Methods

The average population in Sollentuna was 56,589 (49% men) and in Stockholm County (Sollentuna included) 1,795,504 (49% men) during the study period of 1987–2010. Cases of hospitalized acute myocardial infarction and death were obtained for the population of Sollentuna and the rest of Stockholm County using national registries of hospital discharges and deaths. Acute myocardial infarction incidence and mortality were estimated using the average population of Sollentuna and Stockholm in 1987–2010.

Results

During the observation period, the incidence of acute myocardial infarction decreased more in Sollentuna compared with the rest of Stockholm County in women (-22% vs. -7%; for difference in slope <0.05). There was a trend towards a greater decline in Sollentuna compared to the rest of Stockholm County in the incidence of acute myocardial infarction (in men), acute myocardial mortality, and all-cause mortality but the differences were not significant.

Conclusion

During a period of steep decline in acute myocardial infarction incidence and mortality in Stockholm County the municipality of Sollentuna showed a stronger trend in women possibly compatible with favorable influence of a cardiovascular prevention program.

Trial Registration

ClinicalTrials.gov NCT02212145     

A Protein Deep Sequencing Evaluation of Metastatic Melanoma Tissues

Malignant melanoma has the highest increase of incidence of malignancies in the western world. In early stages, front line therapy is surgical excision of the primary tumor. Metastatic disease has very limited possibilities for cure. Recently, several protein kinase inhibitors and immune modifiers have shown promising clinical results but drug resistance in metastasized melanoma remains a major problem. The need for routine clinical biomarkers to follow disease progression and treatment efficacy is high. The aim of the present study was to build a protein sequence database in metastatic melanoma, searching for novel, relevant biomarkers. Ten lymph node metastases (South-Swedish Malignant Melanoma Biobank) were subjected to global protein expression analysis using two proteomics approaches (with/without orthogonal fractionation). Fractionation produced higher numbers of protein identifications (4284). Combining both methods, 5326 unique proteins were identified (2641 proteins overlapping). Deep mining proteomics may contribute to the discovery of novel biomarkers for metastatic melanoma, for example dividing the samples into two metastatic melanoma “genomic subtypes”, (“pigmentation” and “high immune”) revealed several proteins showing differential levels of expression. In conclusion, the present study provides an initial version of a metastatic melanoma protein sequence database producing a total of more than 5000 unique protein identifications. The raw data have been deposited to the ProteomeXchange with identifiers PXD001724 and PXD001725.     

Biological preservation of plant derived animal feed with antifungal microorganisms: safety and formulation aspects

During storage of moist animal feed, growth of detrimental fungi causing spoilage, or being mycotoxigenic or pathogenic, is a severe problem. Addition of biopreservative yeasts or lactic acid bacteria can significantly reduce this problem. However, their use requires several careful considerations. One is the safety to the animal, humans and the environment, tightly connected to legal aspects and the need for pre-market authorisation when supplementing feed with microorganisms. Although both yeasts and lactic acid bacteria are considered comparatively safe organisms due to low production of toxic metabolites, it is of great importance to understand the mechanisms behind the biopreservative abilities. Another important issue concerns practical aspects, such as the economic production of large amounts of the organisms and the development of a suitable formulation giving the organisms a long shelf life. These aspects are discussed and a recommendation of this review is that both safety and formulation aspects of a specific microbe should be considered at an early stage in the selection of new organisms with biopreservation potential.