Bacteria in food packaging paper and board

The bacteria of food packaging paper and board were studied. Most of the aerobic strains were spore-formers; members of the genus Bacillus with B. cereus group (B. cereus, B. mycoides, B. thuringiensis), B. polymyxa group (B. polymyxa, B. circulans, B. macerans, B. pabuli), B. brevis and B. licheniformis predominated. The main source of spore-forming bacteria in paper and board was the broke (rejected paper or board, which is repulped and recycled into the process). Gram-negative bacteria were rare in paper and board in spite of their abundance in the stock. A strain of B. pumilus forming clumping, hairy spores may be of significance in aseptic packaging.     

Bacteria in food packaging paper and board

The bacteria of food packaging paper and board were studied. Most of the aerobic strains were spore-formers; members of the genus Bacillus with B. cereus group ( B. cereus, B. mycoides, B. thuringiensis ), B. polymyxa group ( B. polymyxa, B. circulans, B. macerans, B. pabuli ), B. brevis and B. licheniformis predominated. The main source of spore-forming bacteria in paper and board was the broke (rejected paper of board, which is repulped and recycled into the process). Gram-negative bacteria were rare in paper and board in spite of their abundance in the stock. A strain of B. pumilus forming clumping, hairy spores may be of significance in aseptic packaging.     

Developments in Wood and Packaging Materials Life Cycle Inventories in ecoinvent (9 pp)

Goal, Scope and Background This paper gives an overview on how the wood and packaging material production is inventoried in ecoinvent. Packaging materials have been a very important topic in the area of Life Cycle Assessment for more than twenty years. Wood is the most important renewable material and regenerative fuel used worldwide, and an important raw material for paper / board. Several methodological problems arising when inventorying wood for material and energetic uses in a generic database are discussed in more detail. Within the ecoinvent project, the Swiss data base for life cycle inventory data, two reports are dedicated to these two important topics – report No. 9 for wood and report No. 11 for packaging materials. Methods The whole wood chain has been modeled in a consistent way. This allows one to use this data for LCAs of building materials, bioenergy or paper production. The data represent average technologies used in Central Europe in the year 2000. A revenue-based co-product allocation approach is used for the different outputs. Correction factors are introduced for the consistent modeling of mass-based, material inherent wood properties such as solar energy, carbon uptake and land use. For packaging materials, the datasets represent European average data for the most often used materials as well as specific datasets for the production of actual packaging boxes and containers.Results and Discussion For wood, revenue-based allocation and the use of the correction factors for mass-related wood properties are shown and explained. For packaging materials, the importance of the raw material wood to the total load is shown. Furthermore trends in the data inventories for board packaging materials over the last two decades are discussed: mainly due to the increased comprehensiveness of the data, higher cumulative emissions can be observed. Conclusion For wood, the database ecoinvent provides consistent datasets for the entire chain from forestry to intermediate products such as timber, different types of wood-based boards, chips, pellets, etc. For packaging materials, the number of datasets of basic materials has been extended. A modular concept for actual packaging container datasets allows the user an easy modeling of various types of packaging containers/boxes. In the area of paper and board, a comprehensive database for the production of various types of pulp, paper and board is provided, which is representative for the average European production situation. Outlook Since wood is only limited and representative data for Europe is therefore not included, an update in the near future would be reasonable. Possible further extensions in the future could include various, final wooden products. For the data on paper/board, different levels of quality are observed, requiring a selective up-date of these data. Future extensions could include datasets for the import of pulp from overseas – especially from South America and Canada.     

Redesign of retrovirus packaging cell lines to avoid recombination leading to helper virus production.

Retrovirus vectors can be made in the absence of helper virus by using retrovirus packaging cell lines. Helper-free virus is critical for a variety of gene transfer studies. The most useful packaging cell lines contain helper virus DNA from which the signal required for packaging of the viral RNA genome into virions has been deleted. However, we showed that the ability to package virus is conferred at very low frequency to cells infected with virus from these packaging cell lines, presumably by low-frequency transmission of the deleted virus genome. In addition, these packaging cell lines can interact with some retroviral vectors to yield replication-competent virus. We constructed packaging cell lines containing helper virus DNA that had several alterations in addition to deletion of the packaging signal. The new packaging cells retained the useful features of previously available lines but did not yield helper virus after introduction of any of the vectors tested, and transfer of the packaging function was not detected.     

A defined in vitro system for DNA packaging by the bacteriophage SPP1: insights into the headful packaging mechanism

Tailed icosahedral bacteriophages and other viruses package their double-stranded DNA inside a preformed procapsid. In a large number of phages packaging is initiated by recognition and cleavage by a viral packaging ATPase (terminase) of the specific pac sequence (pac cleavage), which generates the first DNA end to be encapsidated. A sequence-independent cleavage (headful cleavage) terminates packaging, generating a new starting point for another round of packaging. The molecular mechanisms underlying headful packaging and its processivity remain poorly understood. A defined in vitro DNA packaging system for the headful double-stranded DNA bacteriophage SPP1 is reported. The in vitro system consists of DNA packaging reactions with highly purified terminase and SPP1 procapsids, coupled to a DNase protection assay. The high yield obtained enabled us to quantify directly the efficiency of DNA entry into the procapsids. We show that in vitro DNA packaging requires the presence of both terminase subunits. The SPP1 in vitro system is able to efficiently package mature SPP1 DNA as well as linear plasmid DNAs. In contrast, no DNA packaging could be detected with circular DNA, signifying that in vitro packaging requires free DNA extremities. Finally, we demonstrate that SPP1 in vitro DNA packaging is independent of the pac signal. These findings suggest that the formation of free DNA ends that are generated by pac cleavage in vivo is the rate-limiting step in processive headful DNA packaging.     

Optimization of processing variables in wood-rubber composite panel manufacturing technology

The feasibility of manufacturing wood-rubber functional composite panels with a polymeric methylene diphenyl diisocyanate (PMDI) and urea-formaldehyde (UF) combination binder system was investigated. Mechanisms of interacted independent variables (board density, pressing time and pressing temperature) for effect on board properties were opened out. The board performance was evaluated by measuring internal bond (IB) strength, modulus of rupture (MOR) and modulus of elasticity (MOE). The test results were statistically analyzed by using response surface method (RSM) of Design-Expert software to determine the significant independent variables that influenced board properties. A mathematical simulation or response surface models were developed to predict the board properties (MOR, MOE and IB). The results showed that board density and some interactions between the experimental variables were significant factors that influenced board mechanical properties. The suggested optimal board manufacturing conditions were about 170 degrees C, for pressing temperature, 300 s for pressing time, and 1000 g cm(-3) for board density.     

The large subunit of bacteriophage lambda's terminase plays a role in DNA translocation and packaging termination

The DNA packaging enzyme of bacteriophage lambda, terminase, is a heteromultimer composed of a small subunit, gpNu1, and a large subunit, gpA, products of the Nu1 and A genes, respectively. The role of terminase in the initial stages of packaging involving the site-specific binding and cutting of the DNA has been well characterized. While it is believed that terminase plays an active role in later post-cleavage stages of packaging, such as the translocation of DNA into the head shell, this has not been demonstrated. Accordingly, we undertook a generalized mutagenesis of lambda's A gene and found ten lethal mutations, nine of which cause post-cleavage packaging defects. All were located in the amino-terminal two-thirds of gpA, separate from the carboxy-terminal region where mutations affecting the protein's endonuclease activity have been found. The mutants fall into five groups according to their packaging phenotypes: (1) two mutants package part of the lambda chromosome, (2) one mutant packages the entire chromosome, but very slowly compared to wild-type, (3) two mutants do not package any DNA, (4) four mutants, though inviable, package the entire lambda chromosome, and (5) one mutant may be defective in both early and late stages of DNA packaging. These results indicate that gpA is actively involved in late stages of packaging, including DNA translocation, and that this enzyme contains separate functional domains for its early and late packaging activities.     

Nonreciprocal Packaging of Human Immunodeficiency Virus Type 1 and Type 2 RNA: a Possible Role for the p2 Domain of Gag in RNA Encapsidation

The ability of human immunodeficiency virus types 1 (HIV-1) and 2 (HIV-2) to cross-package each other’s RNA was investigated by cotransfecting helper virus constructs with vectors derived from both viruses from which the gag and pol sequences had been removed. HIV-1 was able to package both HIV-1 and HIV-2 vector RNA. The unspliced HIV-1 vector RNA was packaged preferentially over spliced RNA; however, unspliced and spliced HIV-2 vector RNA were packaged in proportion to their cytoplasmic concentrations. The HIV-2 helper virus was unable to package the HIV-1 vector RNA, indicating a nonreciprocal RNA packaging relationship between these two lentiviruses. Chimeric proviruses based on HIV-2 were constructed to identify the regions of the HIV-1 Gag protein conferring RNA-packaging specificity for the HIV-1 packaging signal. Two chimeric viruses were constructed in which domains within the HIV-2 gag gene were replaced by the corresponding domains in HIV-1, and the ability of the chimeric proviruses to encapsidate an HIV-1-based vector was studied. Wild-type HIV-2 was unable to package the HIV-1-based vector; however, replacement of the HIV-2 nucleocapsid by that of HIV-1 generated a virus with normal protein processing which could package the HIV-1-based vector. The chimeric viruses retained the ability to package HIV-2 genomic RNA, providing further evidence for a lack of reciprocity in RNA-packaging ability between the HIV-1 and HIV-2 nucleocapsid proteins. Inclusion of the p2 domain of HIV-1 Gag in the chimera significantly enhanced packaging.     

A life cycle assessment of packaging options for contrast media delivery: comparing polymer bottle vs. glass bottle

Purpose

This paper compares environmental impacts of two packaging options for contrast media offered by GE Healthcare: +PLUSPAK? polymer bottle and traditional glass bottle. The study includes all relevant life cycle stages from manufacturing to use and final disposal of the bottles and includes evaluation of a variety of end-of-life disposal scenarios. The study was performed in accordance with the international standards ISO 14040/14044, and a third-party critical review was conducted.

Methods

The functional unit is defined as the packaging of contrast media required to deliver one dose of 96 mL to a patient for an X-ray procedure. Primary data are from GE Healthcare and its suppliers; secondary data are from the ecoinvent database and the literature. A variety of end-of-life disposal scenarios are explored using both cutoff and market-based allocation. Impact assessment includes human health (midpoint) and ecosystems and resources (end point) categories from ReCiPe (H) and cumulative energy demand. Sensitivity analyses include (1) bottle size, (2) secondary packaging, (3) manufacturing electricity, (4) glass recycled content, (5) scrap rate, (6) distribution transport, (7) contrast media, and (8) choice of impact assessment method. Uncertainty analysis is performed to determine how data quality affects the study conclusions.

Results and discussion

This study indicates that the polymer bottle outperforms the glass bottle in every environmental impact category considered. Bottle components are the most significant contributors, and the vial body has the highest impacts among bottle components for both polymer and glass bottles. The polymer bottle exhibits lower impact in all impact categories considered regardless of the following: end-of-life treatment (using either cutoff or market-based allocation), bottle size, manufacturing electricity grid mix, glass recycled content, scrap rate, contrast media, distribution transport (air vs. ocean), and choice of impact assessment method. Secondary packaging can be a major contributor to impact. The polymer bottle has considerably lower impact compared to the glass bottle for all multi-pack configurations, but the comparison is less clear for single-pack configurations due to significantly higher packaging material used per functional dose, resulting in proportionally higher impacts in all impact categories.

Conclusions

The lower impacts of the polymer bottle for this packaging application can be attributed to lower material and manufacturing impacts, lower distribution impacts, and lower end-of-life disposal impacts. The results of this study suggest that using polymer rather than glass bottles provides a means by which to lower environmental impact of contrast media packaging.     

The Nucleocapsid Domain Is Responsible for the Ability of Spleen Necrosis Virus (SNV) Gag Polyprotein To Package both SNV and Murine Leukemia Virus RNA

Murine leukemia virus (MLV)-based vector RNA can be packaged and propagated by the proteins of spleen necrosis virus (SNV). We recently demonstrated that MLV proteins cannot support the replication of an SNV-based vector; RNA analysis revealed that MLV proteins cannot efficiently package SNV-based vector RNA. The domain in Gag responsible for the specificity of RNA packaging was identified using chimeric gag-pol expression constructs. A competitive packaging system was established by generating a cell line that expresses one viral vector RNA containing the MLV packaging signal (Psi) and another viral vector RNA containing the SNV packaging signal (E). The chimeric gag-pol expression constructs were introduced into the cells, and vector titers as well as the efficiency of RNA packaging were examined. Our data confirm that Gag is solely responsible for the selection of viral RNAs. Furthermore, the nucleocapsid (NC) domain in the SNV Gag is responsible for its ability to interact with both SNV E and MLV Psi. Replacement of the SNV NC with the MLV NC generated a chimeric Gag that could not package SNV RNA but retained its ability to package MLV RNA. A construct expressing SNV gag-MLV pol supported the replication of both MLV and SNV vectors, indicating that the gag and pol gene products from two different viruses can functionally cooperate to perform one cycle of retroviral replication. Viral titer data indicated that SNV cis-acting elements are not ideal substrates for MLV pol gene products since infectious viruses were generated at a lower efficiency. These results indicate that the nonreciprocal recognition between SNV and MLV extends beyond the Gag-RNA interaction and also includes interactions between Pol and other cis-acting elements.     

Environmental effects from a recycling rate increase of cardboard of aseptic packaging system for milk using life cycle approach

Goal, Scope and Background  Despite the well-known advantages of recycling materials to reduce solid waste or save natural resources, the recycling stage is an additional process within the life cycle that has its own energy and input requirements, as well as specific emissions. The objective of the present paper is to analyze the life cycle inventory associated with the increase in recycling rate (from 2% up to 22% at present) of the cardboard contained in the aseptic packaging for long-life milk. The main aspects of the manufacturing of the Tetra Pak aseptic package, including the filling of the product, the distribution of the conditioned product, up to the final disposal and recycling rates, were considered. Materials and Methods  This study was conducted in accordance with the general directives of the ISO 14040 series. The packaging material system was assessed using 1000 liters of milk as a functional unit, in a packaging system containing 12 units of 1 L cartons each, placed on a corrugated paperboard tray wrapped in polyethylene shrink film and arranged onto one-way wooden pallets. Brazilian inventories for energy, carton, corrugated paperboard and aluminum, based on site-collected data were employed. The final disposal of used packages was modeled using the Average Brazilian Municipal Solid Waste Management data collected for the purpose of the census of the year 2000. Results  Comparison of the total energy consumption throughout the whole life cycle of two recycling scenarios (i.e. different recycling rates) analyzed shows that the higher recycling rate led to a 6% reduction of the total energy requirement for the long-life milk package material system. The most significant reductions in the consumption of natural resources were: 8% water, 11% wood and 10% land use savings. Greenhouse gases were the main reduced air emissions and contributed with a reduction of 9.7% in GWP. Most water emissions were reduced: 10% COD, 9% BOD and 6% TSS. A unique drawback directly caused by the increase of the recycling rate was an increase of 14.4 g in TDS emissions (57%). Discussion  The reduction in energy requirements are related and limited to the proportionality among the different materials that make up the packaging system. Most emission reductions result from the replacement of virgin materials with recycled materials in the packaging system. Although the average balance of water emissions is positive, the need to improve wastewater treatment processes in the paper recycling plants to reduce TDS is highlighted as a key issue. Conclusions  It may be concluded that the increase in the recycling rate brings about a series of benefits in terms of reduction of energy and natural resource consumption, air pollutants and most water emissions. In this case, the increase of the recycling rate improved the overall environmental performance of the aseptic Tetra Pak system for milk. Recommendations and Perspectives  The authors are currently analyzing alternative recycling scenarios that will enable one to evaluate maximum reduction in GWP. Further studies could include the agriculture stages, livestock and consumer phase to broaden the environmental evaluation. ESS-Submission Editor: Dr. Andreas A. Detzel (andreas.detzel@ifeu.de)     

cis-Acting elements important for retroviral RNA packaging specificity

Spleen necrosis virus (SNV) proteins can package RNA from distantly related murine leukemia virus (MLV), whereas MLV proteins cannot package SNV RNA efficiently. We used this nonreciprocal recognition to investigate regions of packaging signals that influence viral RNA encapsidation specificity. Although the MLV and SNV packaging signals (Psi and E, respectively) do not contain significant sequence homology, they both contain a pair of hairpins. This hairpin pair was previously proposed to be the core element in MLV Psi. In the present study, MLV-based vectors were generated to contain chimeric SNV/MLV packaging signals in which the hairpins were replaced with the heterologous counterpart. The interactions between these chimeras and MLV or SNV proteins were examined by virus replication and RNA analyses. SNV proteins recognized all of the chimeras, indicating that these chimeras were functional. We found that replacing the hairpin pair did not drastically alter the ability of MLV proteins to package these chimeras. These results indicate that, despite the important role of the hairpin pair in RNA packaging, it is not the major motif responsible for the ability of MLV proteins to discriminate between the MLV and SNV packaging signals. To determine the role of sequences flanking the hairpins in RNA packaging specificity, vectors with swapped flanking regions were generated and evaluated. SNV proteins packaged all of these chimeras efficiently. In contrast, MLV proteins strongly favored chimeras with the MLV 5'-flanking regions. These data indicated that MLV Gag recognizes multiple elements in the viral packaging signal, including the hairpin structure and flanking regions.     

The influence of packaging on consumers’ quality perception of carrots

The aim of the present research was to investigate the influence of packaging design on consumers' perception of quality of fresh carrots. We adopted a conjoint analytic approach in which 251 Danish consumers rated the perceived quality and value (expected price) of nine packaging images, obtained by systematically varying packaging type (plastic bag, plastic box, cardboard paper) and label color (blue, brown, grey). The results revealed that the main attribute influencing the perceptions of the consumer was packaging type. Specifically, the box packages (both plastic and cardboard) were associated to carrots of significantly higher perceived value and quality compared to the plastic bag packages. Furthermore, the study identified the most important aspects consumers attend to when purchasing carrots. A transparent packaging, allowing consumers to inspect the produce, was mentioned as the most important aspect. Being organic and local were identified as the second and third most important, respectively.

Practical applications

Packaging is an important extrinsic product attribute that can influence consumer perceptions of fresh produce. The results have implications for retailers and producers with respect to the choice of packaging and label design. Specifically, consumers associated box packages to higher quality produce, suggesting that carrots in this type of package may command higher price and/or be preferred to bagged alternatives at a similar price point. The study further indicated the importance of using a transparent packaging that clearly allow consumers to inspect the produce, and also suggest that “organic” and “local” are important drivers of purchase for this product category.     

Construction of a large phage display antibody library by in vitro package and in vivo recombination

Capacity and diversity are extremely important to the quality of various phage display libraries. In this work, λ phage-based in vitro package was applied to construct a filamentous phage display antibody library so as to enlarge its capacity and introduce more sequence diversity in the final library. In vivo recombination via Cre recombinase/lox sites was also exploited to create V_H/V_L combination diversity based on multivalent package of λ phage packaging extracts on phagemid DNA concatemers. The library constructed with 10 μg concatenated phagemid DNA and ten vials of λ phage packaging extracts was calculated to contain 1.40×10¹⁰ independent clones. Higher capacity can be easily achieved when more materials are consumed. This strategy is somewhat more efficient than prior methods.     

Environmental performance assessment of hardboard manufacture

Background, aim and scope  The forest-based and related industries comprise one of the most important industry sectors in the European Union, representing some 10% of the EU's manufacturing industries. Their activities are based on renewable raw material resources and efficient recycling. The forest-based industries can be broken down into the following sectors: forestry, woodworking, pulp and paper manufacturing, paper and board converting and printing and furniture. The woodworking sector includes many sub-sectors; one of the most important is that of wood panels accounting for 9% of total industry production. Wood panels are used as intermediate products in a wide variety of applications in the furniture and building industries. There are different kinds of panels: particleboard, fibreboard, veneer, plywood and blockboard. The main goal of this study was to assess the environmental impacts during the life cycle of wet-process fibreboard (hardboard) manufacturing to identify the processes with the largest environmental impacts. Methods  The study covers the life cycle of hardboard production from a cradle-to-gate perspective. A hardboard plant was analysed in detail, dividing the process chain into three subsystems: wood preparation, board forming and board finishing. Ancillary activities such as chemicals, wood chips, thermal energy and electricity production and transport were included within the system boundaries. Inventory data came from interviews and surveys (on-site measurements). When necessary, the data were complemented with bibliographic resources. The life cycle assessment procedure followed the ISO14040 series. The life cycle inventory (LCI) and impact assessment database for this study were constructed using SimaPro Version 7.0 software. Results  Abiotic depletion (AD), global warming (GW), ozone layer depletion (OLD), human toxicity (HT), ecotoxicity, photochemical oxidant formation (PO), acidification (AC) and eutrophication (EP) were the impact categories analysed in this study. The wood preparation subsystem contributed more than 50% to all impact categories, followed by board forming and board finishing, which is mainly due to chemicals consumption in the wood preparation subsystem. In addition, thermal energy requirements (for all subsystems) were fulfilled by on-site wood waste burning and, accordingly, biomass energy converters were considered. Several processes were identified as hot spots in this study: phenol-formaldehyde resin production (with large contribution to HT, fresh water aquatic ecotoxicity and PO), electricity production (main contributor to marine aquatic ecotoxicity), wood chips production (AD and OLD) and finally, biomass burning for heat production (identified as the largest contributor to AC and EP due to NO _X emissions). In addition, uncontrolled formaldehyde emissions from manufacturing processes at the plant such as fibre drying should be controlled due to relevant contributions to terrestrial ecotoxicity and PO. A sensitivity analysis of electricity profile generation (strong geographic dependence) was carried out and several European profiles were analysed. Discussion  Novel binding agents for the wood panel industry as a substitute for the currently used formaldehyde-based binders have been extensively investigated. Reductions of toxic emissions during drying, mat forming and binder production are desirable. The improved method would considerably reduce the contributions to all impact categories. Conclusions  The results obtained in this work allow forecasting the importance of the wood preparation subsystem for the environmental burdens associated with hardboard manufacture. Special attention was paid to the inventory analysis stage for each subsystem. It is possible to improve the environmental performance of the hardboard manufacturing process if some alternatives are implemented regarding the use of chemicals, electricity profile and emission sources in the production processes located inside the plant. Recommendations and perspectives  This study provides useful information for forest-based industries related to panel manufacture with the aim of increasing their sustainability. Our research continues to assess the use phase and final disposal of panels to complete the life cycle assessment. Future work will focus on analysing the environmental aspects associated with plywood, another type of commonly used wood panel.     

The Structure of the Herpes Simplex Virus DNA-Packaging Terminase pUL15 Nuclease Domain Suggests an Evolutionary Lineage among Eukaryotic and Prokaryotic Viruses

Herpes simplex virus 1 (HSV-1), the prototypic member of herpesviruses, employs a virally encoded molecular machine called terminase to package the viral double-stranded DNA (dsDNA) genome into a preformed protein shell. The terminase contains a large subunit that is thought to cleave concatemeric viral DNA during the packaging initiation and completion of each packaging cycle and supply energy to the packaging process via ATP hydrolysis. We have determined the X-ray structure of the C-terminal domain of the terminase large-subunit pUL15 (pUL15C) from HSV-1. The structure shows a fold resembling those of bacteriophage terminases, RNase H, integrases, DNA polymerases, and topoisomerases, with an active site clustered with acidic residues. Docking analysis reveals a DNA-binding surface surrounded by flexible loops, indicating considerable conformational changes upon DNA binding. In vitro assay shows that pUL15C possesses non-sequence-specific, Mg²⁺-dependent nuclease activity. These results suggest that pUL15 uses an RNase H-like, metal ion-mediated catalysis mechanism for cleavage of viral concatemeric DNA. The structure reveals extra structural elements in addition to the RNase H-like fold core and variations in local architecture of the nuclease active site, which are conserved in herpesvirus terminases and bear great similarity to the phage T4 gp17 but are distinct from podovirus and siphovirus orthologs and cellular RNase H, delineating a new evolutionary lineage among a large family of eukaryotic viruses and simple and complex prokaryotic viruses.     

Packaging Strategies That Save Food: A Research Agenda for 2030

Thoroughly considering and optimizing packaging systems can avoid food loss and waste. We suggest a number of issues that must be explored and review the associated challenges. Five main issues were recognized through the extensive experience of the authors and engagement of multiple stakeholders. The issues promoted are classified as follows: (1) identify and obtain specific data of packaging functions that influence food waste; (2) understand the total environmental burden of product/package by considering the trade‐off between product protection and preservation and environmental footprint; (3) develop understanding of how these functions should be treated in environmental footprint evaluations; (4) improve packaging design processes to also consider reducing food waste; and (5) analyze stakeholder incentives to reduce food loss and waste. Packaging measures that save food will be important to fulfill the United Nations Sustainable Development goal to halve per capita global food waste at the retail and consumer levels and to reduce food losses along production and supply chains.     

Enzymatic characterization ofBacilli from food packaging paper and board machines

Summary Aerobic spore-forming bacteria were found dominant in the microflora of food packaging paper and board. Twenty-five strains of bacteria belonging to the genusBacillus were isolated from these paper and board machines, papermaking chemicals, and final products of papermaking. Nineteen strains were analyzed for production of -amylase, -glucosidase, glucoamylase, pullulanase, -glucanase, carboxymethyl cellulase, and caseinase, and also for resistance towards industrial biocides. pH and temperature optima for the activity of the enzymes were determined. All strains were found to produce one or more of the enzymes studied. The amylolytic enzymes of most strains had high temperature optima for activity. Vegetative cells of all strains were found very resistant towards the different commercial slimicides used in paper and board mills. This property together with the ability to survive through the dry end of the machine to the final board and paper, and the production of enzymes degrading papermaking chemicals makes these bacteria potentially harmful in paper and board mills.     

Mechanisms of human immunodeficiency virus type 2 RNA packaging: efficient trans packaging and selection of RNA copackaging partners

Human immunodeficiency virus type 2 (HIV-2) has been reported to have a distinct RNA packaging mechanism, referred to as cis packaging, in which Gag proteins package the RNA from which they were translated. We examined the progeny generated from dually infected cell lines that contain two HIV-2 proviruses, one with a wild-type gag/gag-pol and the other with a mutant gag that cannot express functional Gag/Gag-Pol. Viral titers and RNA analyses revealed that mutant viral RNAs can be packaged at efficiencies comparable to that of viral RNA from which wild-type Gag/Gag-Pol is translated. These results do not support the cis-packaging hypothesis but instead indicate that trans packaging is the major mechanism of HIV-2 RNA packaging. To further characterize the mechanisms of HIV-2 RNA packaging, we visualized HIV-2 RNA in individual particles by using fluorescent protein-tagged RNA-binding proteins that specifically recognize stem-loop motifs in the viral genomes, an assay termed single virion analysis. These studies revealed that >90% of the HIV-2 particles contained viral RNAs and that RNAs derived from different viruses were copackaged frequently. Furthermore, the frequencies of heterozygous particles in the viral population could be altered by changing a 6-nucleotide palindromic sequence at the 5'-untranslated region of the HIV-2 genome. This finding indicates that selection of copackaging RNA partners occurs prior to encapsidation and that HIV-2 Gag proteins primarily package one dimeric RNA rather than two monomeric RNAs. Additionally, single virion analyses demonstrated a similar RNA distribution in viral particles regardless of whether both viruses had a functional gag or one of the viruses had a nonfunctional gag, providing further support for the trans-packaging hypothesis. Together, these results revealed mechanisms of HIV-2 RNA packaging that are, contrary to previous studies, in many respects surprisingly similar to those of HIV-1.     

Role for the adenovirus IVa2 protein in packaging of viral DNA

Although it has been demonstrated that the adenovirus IVa2 protein binds to the packaging domains on the viral chromosome and interacts with the viral L1 52/55-kDa protein, which is required for viral DNA packaging, there has been no direct evidence demonstrating that the IVa2 protein is involved in DNA packaging. To understand in greater detail the DNA packaging mechanisms of adenovirus, we have asked whether DNA packaging is serotype or subgroup specific. We found that Ad7 (subgroup B), Ad12 (subgroup A), and Ad17 (subgroup D) cannot complement the defect of an Ad5 (subgroup C) mutant, pm8001, which does not package its DNA due to a mutation in the L1 52/55-kDa gene. This indicates that the DNA packaging systems of different serotypes cannot interact productively with Ad5 DNA. Based on this, a chimeric virus containing the Ad7 genome except for the inverted terminal repeats and packaging sequence from Ad5 was constructed. This chimeric virus replicates its DNA and synthesizes Ad7 proteins, but it cannot package its DNA in 293 cells or 293 cells expressing the Ad5 L1 52/55-kDa protein. However, this chimeric virus packages its DNA in 293 cells expressing the Ad5 IVa2 protein. These results indicate that the IVa2 protein plays a role in viral DNA packaging and that its function is serotype specific. Since this chimeric virus cannot package its own DNA, but produces all the components for packaging Ad7 DNA, it may be a more suitable helper virus for the growth of Ad7 gutted vectors for gene transfer.