Transfer of Non-Dioxin-Like Polychlorinated Biphenyls (ndl-PCBs) from Feed and Soil into Hen Eggs.

Understanding the transfer of non-dioxin-like polychlorinated biphenyls (ndl-PCBs) into foods of animal origin is crucial for human health risk assessment. In two experiments, we investigated the transfer of ndl-PCBs from contaminated feed and soil into eggs and meat of laying hens. The transfer from the feed was investigated with 30 laying hens. The treated hens were divided into two groups fed a contaminated diet (12.8 µg/kg sum of indicator ndl-PCBs; 88% dry matter (DM)) for 28 and 63 days, respectively, and then experienced a depuration period of 100 days with control feed. The transfer from soil was investigated with 72 laying hens kept in three separate outdoor pens (with three levels of ndl-PCB soil contamination) for 168 days. In both experiments, eggs were collected and analyzed for ndl-PCBs. In the second experiment, animals (n = 3 at the beginning, n = 6 per group after 42, 84, and 168 days) were slaughtered to determine ndl-PCBs in meat (breast muscle tissue) fat. The transfer of ndl-PCB from both feed and soil was clearly measurable and concentrations in eggs quickly exceeded maximum levels. Clear differences between individual congeners were observed. In particular, the low-chlorinated ndl-PCBs 52 and 101 are hardly found in eggs, despite their relatively high concentration in feed and soil. PCBs 138, 153, and 180, on the other hand, were found in large proportions in eggs and meat.

Cusp observation at Saturn's high-latitude magnetosphere by the Cassini spacecraft.

We report on the first analysis of magnetospheric cusp observations at Saturn by multiple in situ instruments onboard the Cassini spacecraft. Using this we infer the process of reconnection was occurring at Saturn's magnetopause. This agrees with remote observations that showed the associated auroral signatures of reconnection. Cassini crossed the northern cusp around noon local time along a poleward trajectory. The spacecraft observed ion energy-latitude dispersions-a characteristic signature of the terrestrial cusp. This ion dispersion is "stepped," which shows that the reconnection is pulsed. The ion energy-pitch angle dispersions suggest that the field-aligned distance from the cusp to the reconnection site varies between ∼27 and 51 RS . An intensification of lower frequencies of the Saturn kilometric radiation emissions suggests the prior arrival of a solar wind shock front, compressing the magnetosphere and providing more favorable conditions for magnetopause reconnection. KEY POINTS: We observe evidence for reconnection in the cusp plasma at SaturnWe present evidence that the reconnection process can be pulsed at SaturnSaturn's cusp shows similar characteristics to the terrestrial cusp.

Detection of a strongly negative surface potential at Saturn's moon Hyperion.

On 26 September 2005, Cassini conducted its only close targeted flyby of Saturn's small, irregularly shaped moon Hyperion. Approximately 6 min before the closest approach, the electron spectrometer (ELS), part of the Cassini Plasma Spectrometer (CAPS) detected a field-aligned electron population originating from the direction of the moon's surface. Plasma wave activity detected by the Radio and Plasma Wave instrument suggests electron beam activity. A dropout in energetic electrons was observed by both CAPS-ELS and the Magnetospheric Imaging Instrument Low-Energy Magnetospheric Measurement System, indicating that the moon and the spacecraft were magnetically connected when the field-aligned electron population was observed. We show that this constitutes a remote detection of a strongly negative (∼ -200 V) surface potential on Hyperion, consistent with the predicted surface potential in regions near the solar terminator.

Science potential from a Europa lander.

The prospect of a future soft landing on the surface of Europa is enticing, as it would create science opportunities that could not be achieved through flyby or orbital remote sensing, with direct relevance to Europa's potential habitability. Here, we summarize the science of a Europa lander concept, as developed by our NASA-commissioned Science Definition Team. The science concept concentrates on observations that can best be achieved by in situ examination of Europa from its surface. We discuss the suggested science objectives and investigations for a Europa lander mission, along with a model planning payload of instruments that could address these objectives. The highest priority is active sampling of Europa's non-ice material from at least two different depths (0.5-2 cm and 5-10 cm) to understand its detailed composition and chemistry and the specific nature of salts, any organic materials, and other contaminants. A secondary focus is geophysical prospecting of Europa, through seismology and magnetometry, to probe the satellite's ice shell and ocean. Finally, the surface geology can be characterized in situ at a human scale. A Europa lander could take advantage of the complex radiation environment of the satellite, landing where modeling suggests that radiation is about an order of magnitude less intense than in other regions. However, to choose a landing site that is safe and would yield the maximum science return, thorough reconnaissance of Europa would be required prior to selecting a scientifically optimized landing site.

Intraguild predation provides a selection mechanism for bacterial antagonistic compounds.

Bacteriocins are bacterial proteinaceous toxins with bacteriostatic or bacteriocidal activity towards other bacteria. The current theory on their biological role concerns especially colicins, with underlying social interactions described as an example of spite. This leads to a rock-paper-scissors game between colicin producers and sensitive and resistant variants. The generality of this type of selection mechanism has previously been challenged with lactic acid bacterial (LAB) bacteriocins as an example. In the natural environment of LAB, batch cultures are the norm opposed to the natural habitats of Escherichia coli where continuous cultures are prevailing. This implies that fitness for LAB, to a large degree, is related to survival rates (bottleneck situations) rather than to growth rates. We suggest that the biological role of LAB bacteriocins is to enhance survival in the stationary growth phase by securing a supply of nutrients from lysed target cells. Thus, this social interaction is an example of selfishness rather than of spite. Specifically, it fits into an ecological model known as intraguild predation (IGP), which is a combination of competition and predation where the predator (LAB bacteriocin producer) and prey (bacteriocin susceptible bacteria) share similar and often limited resources. We hypothesize that IGP may be a common phenomenon promoting microbial production of antagonistic compounds.

The genome sequence of the lactic acid bacterium, Carnobacterium maltaromaticum ATCC 35586 encodes potential virulence factors.

The genus Carnobacterium belongs to the lactic acid bacteria and Carnobacterium maltaromaticum is commonly found in modified atmosphere packed and vacuum packed fish and meat products as well as in live fish. This species has been described as a fish pathogenic organism but human clinical isolates have only been obtained at one occasion. To investigate the virulence potential we sequenced the entire genome of strain ATCC 35586, isolated from a diseased salmon. When comparing the translated gene products of ATCC 35586 to those of Gram positive bacterial pathogens and probiotics as well as the related Carnobacterium sp. AT7 we identified a range of putative virulence genes including genes encoding products involved in adhesion to fibronectin and collagen, capsule synthesis, cell wall modification, iron scavenging mechanisms, haemolysis, invasion and resistance to toxic compounds. Of particular interest was the presence of internalin encoding gene homologues to some of those found in Listeria spp. and Lactobacillus plantarum. Furthermore, the ATCC 35586 strain possesses a gene encoding a product similar to the central Listeria monocytogenes transcriptional regulator PrfA, that in this organism controls virulence gene expression by binding to conserved DNA binding sites. Based on the consensus DNA sequence of this binding site, we identified a total of 65 genes in the ATCC 35586 genome that in the upstream region carry a PrfA binding motif. Among these is one of the internalin encoding genes; two genes encoding products involved in capsule biosynthesis as well as various genes encoding products with metabolic functions. In contrast to L. monocytogenes, the ATCC 35586 strain did not encode other PrfA dependent virulence factors such as listeriolysin O, phospholipases A and B, ActA, listeriolysin O, zinc metallo protease and internalins A and B. In conclusion, C. maltaromaticum ATCC 35586 carries putative virulence genes that may explain its reported ability to infect fish. The findings of this study give no reason for concern regarding human health by the presence of this species in food products.

The chitinolytic activity of Listeria monocytogenes EGD is regulated by carbohydrates but also by the virulence regulator PrfA.

Chitin, an insoluble polymer of N-acetyl-D-glucosamine (GlcNAc), is one of the most abundant carbohydrate polymers in marine and terrestrial environments. Chitin hydrolysis by Listeria monocytogenes depends on two chitinase-encoding genes, chiA and chiB, and the aim of this study was to investigate their regulation. Chitin induces the expression of both chitinases in late exponential growth phase, and chiA but not chiB is furthermore induced by the monomer GlcNAc. Furthermore, their expression is subjected to catabolite control. Chitinases expressed by bacterial pathogens have proven to be important not only for nutrient acquisition and environmental survival but also for infecting animals and humans. Interestingly, the central L. monocytogenes virulence gene regulator, PrfA, is required for the chitinolytic phenotype, as chitinase activity was significantly reduced in prfA mutant cells compared to its level in wild-type cells. In agreement with this, Northern blot analysis showed that the amounts of chiA and chiB transcripts upon induction by chitin were significantly lower in the prfA mutant than in the wild type. The chitinolytic activity and chiA and chiB expression were reduced in the absence of the sigB gene, indicating that σ(B) is also important for the production of chitinases. The chiA, chiB, and chiA chiB mutants were not impaired for in vitro adhesion and invasion in epithelial cell lines, but the chiA chiB double mutant showed less survival ability in a chitin-enriched medium. The regulation of chitinolytic activity in L. monocytogenes is complex, and taken together, the results indicate that the biological role of this activity may not be limited to the external environment.

Cloning and comparison of phylogenetically related chitinases from Listeria monocytogenes EGD and Enterococcus faecalis V583.

AIMS: To compare enzymatic activities of two related chitinases, ChiA and EF0361, encoded by Listeria monocytogenes and Enterococcus faecalis, respectively. METHODS AND RESULTS: The chiA and EF0361 genes were amplified by PCR, cloned and expressed with histidine tags, allowing easy purification of the gene products. ChiA had a molecular weight as predicted from the amino acid sequence, whereas EF0361 was 1840 Da lower than expected because of C-terminal truncation. The ChiA and EF0361 enzymes showed activity towards 4-nitrophenyl N,N'-diacetyl-beta-D-chitobioside with K(m) values of 1.6 and 2.1 mmol l(-1), respectively, and k(cat) values of 21.6 and 6.5 s(-1). The enzymes also showed activity towards 4-nitrophenyl beta-D-N, N', N''-triacetylchitotriose and carboxy-methyl-chitin-Remazol Brilliant Violet but not towards 4-nitrophenyl N-acetyl-beta-D-glucosaminide. Chitinolytic specificities of the enzymes were supported by their inactivity towards the substrates 4-nitrophenyl beta-D-cellobioside and peptidoglycan. The pH and temperature profiles for catalytic activities were relatively similar for both the enzymes. CONCLUSION: The ChiA and EF0361 enzymes show a high degree of similarity in their catalytic activities although their hosts share environmental preferences only to some extent. SIGNIFICANCE AND IMPACT OF THE STUDY: This study contributes to an understanding of the chitinolytic activities by L. monocytogenes and Ent. faecalis. Detailed information on their chitinolytic systems will help define potential reservoirs in the natural environment and possible transmission routes into food-manufacturing plants.

Lactic acid bacteria associated with a heat-processed pork product and sources of variation affecting chemical indices of spoilage and sensory characteristics.

AIMS: To evaluate the potential for developing a quality index for a Danish modified atmosphere packaged (MAP) heat-processed and naturally contaminated pork meat product stored at 5 degrees C. METHODS AND RESULTS: The composition of the predominating microflora and changes in contents of tyramine, arginine, organic acids and sensory characteristics were analysed. The microflora was predominated by Lactobacillus sakei, Leuconostoc carnosum and Carnobacterium divergens. The presence of each species varied between products and batches resulting in limited usefulness of the concentrations of these bacteria or their metabolites as indices of quality. Furthermore, the three species differed in their metabolic activities as shown by use of a model meat extract. However, when MAP storage of the processed pork product was followed by aerobic storage then acetic acid showed some potential as a chemical indicator of sensory quality. CONCLUSION: Variation in processing parameters and spoilage microbiota limited the usefulness of concentrations of micro-organisms and their metabolites as indices of spoilage for the studied processed MAP pork product. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study contributes to an understanding of the difficulties experienced in developing quality indices to be used in the control of microbial spoilage of processed MAP meat products.

Chitin hydrolysis by Listeria spp., including L. monocytogenes.

Listeria spp., including the food-borne pathogen Listeria monocytogenes, are ubiquitous microorganisms in the environment and thus are difficult to exclude from food processing plants. The factors that contribute to their multiplication and survival in nature are not well understood, but the ability to catabolize various carbohydrates is likely to be very important. One major source of carbon and nitrogen in nature is chitin, an insoluble linear beta-1,4-linked polymer of N-acetylglucosamine (GlcNAc). Chitin is found in cell walls of fungi and certain algae, in the cuticles of arthropods, and in shells and radulae of molluscs. In the present study, we demonstrated that L. monocytogenes and other Listeria spp. are able to hydrolyze alpha-chitin. The chitinolytic activity is repressed by the presence of glucose in the medium, suggesting that chitinolytic activity is subjected to catabolite repression. Activity is also regulated by temperature and is higher at 30 degrees C than at 37 degrees C. In L. monocytogenes EGD, chitin hydrolysis depends on genes encoding two chitinases, lmo0105 (chiB) and lmo1883 (chiA), but not on a gene encoding a putative chitin binding protein (lmo2467). The chiB and chiA genes are phylogenetically related to various well-characterized chitinases. The potential biological implications of chitinolytic activity of Listeria are discussed.

Limitations in the use of Drosophila melanogaster as a model host for gram-positive bacterial infection.

AIMS: To examine sensitivities of various Drosophila melanogaster strains towards human pathogenic and nonpathogenic gram-positive bacteria. METHODS AND RESULTS: The D. melanogaster Oregon R strain was infected by injecting the thorax with a needle containing Escherichia coli (negative control), Listeria monocytogenes, Staphylococcus aureus (both food-borne pathogens), Listeria innocua, Bacillus subtilis, Carnobacterium maltaromaticum, Lactobacillus plantarum or Pediococcus acidilactici (all nonpathogenic bacteria). Listeria monocytogenes and S. aureus killed the host rapidly compared with the negative control. Infection with L. innocua, B. subtilis or C. maltaromaticum also resulted in a high fly mortality, whereas Lact. plantarum and P. acidilactici resulted in a slightly increased mortality. Four additional D. melanogaster lines, three of which had been selected for heat, cold and desiccation resistance respectively, were subjected to infection by L. monocytogenes, S. aureus and E. coli. Mortality rates were comparable with that of the Oregon R strain. CONCLUSIONS: Use of the injection method shows the limitation of D. melanogaster as a model host for gram-positive bacteria as opportunistic infection by nonpathogenic gram-positive bacteria results in partial or high mortality. In addition, lines of fruit flies resistant to various stress exposures did not show an increased resistance to infection by gram-positive pathogens under the conditions tested. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates the inadequacy of D. melanogaster infected by the injection method in order to distinguish between virulent and nonvirulent gram-positive bacteria.

Identification of a dynamic atmosphere at Enceladus with the Cassini magnetometer.

The Cassini magnetometer has detected the interaction of the magnetospheric plasma of Saturn with an atmospheric plume at the icy moon Enceladus. This unanticipated finding, made on a distant flyby, was subsequently confirmed during two follow-on flybys, one very close to Enceladus. The magnetometer data are consistent with local outgassing activity via a plume from the surface of the moon near its south pole, as confirmed by other Cassini instruments.

Leuconostoc durionis sp. nov., a heterofermenter with no detectable gas production from glucose.

Three lactic acid bacterial (LAB) strains obtained from a Malaysian acid-fermented condiment, tempoyak (made from pulp of the durian fruit), showed analogous but distinct patterns after screening by SDS-PAGE of whole-cell proteins and comparison with profiles of all recognized LAB species. 16S rRNA gene sequencing of one representative strain showed that the taxon belongs phylogenetically to the genus Leuconostoc, with its nearest neighbour being Leuconostoc fructosum (98 % sequence similarity). Biochemical characteristics and DNA-DNA hybridization experiments demonstrated that the strains differ from Leuconostoc fructosum and represent a single, novel Leuconostoc species for which the name Leuconostoc durionis sp. nov. is proposed. The type strain is LMG 22556(T) (= LAB 1679(T) = D-24(T) = CCUG 49949(T)).

Cassini magnetometer observations during Saturn orbit insertion.

Cassini's successful orbit insertion has provided the first examination of Saturn's magnetosphere in 23 years, revealing a dynamic plasma and magnetic environment on short and long time scales. There has been no noticeable change in the internal magnetic field, either in its strength or its near-alignment with the rotation axis. However, the external magnetic field is different compared with past spacecraft observations. The current sheet within the magnetosphere is thinner and more extended, and we observed small diamagnetic cavities and ion cyclotron waves of types that were not reported before.

Detection of resistance of lactic acid bacteria to a mixture of the hop analogue compounds tetrahydroiso-alpha-acids by noninvasive measurement of intracellular pH.

AIMS: To examine the resistance of beer isolates of lactic acid bacteria (LAB) towards a mixture of tetrahydroiso-alpha-acids (Tetra) by growth experiments as well as by measurement of intracellular pH. METHODS AND RESULTS: Beer LAB isolates were identified to species level by SDS-PAGE of whole-cell proteins. Beer isolates of Lactobacillus brevis showed better ability for growth in the presence of Tetra than nonbeer isolates of the L. brevis or other species of LAB including beer and nonbeer isolates. The antimicrobial effect of Tetra was also examined by noninvasive measurement of intracellular pH by fluorescence ratio imaging microscopy for selected beer isolates of L. brevis and Pediococcus inopinatus. Strains of L. brevis showing limited decrease of intracellular pH during exposure to Tetra also showed better ability for growth in the presence of these compounds as well as in commercial beer products. CONCLUSIONS: It was possible to apply a method for noninvasive measurement of intracellular pH to predict the resistance of beer spoilage LAB towards the Tetra hop analogue compounds. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated the usability of a new rapid method for detecting hop-resistant variants of known beer spoilage LAB species.

Intra-specific variation of Lactobacillus plantarum and Lactobacillus pentosus in sensitivity towards various bacteriocins.

Fifty-two strains belonging to the Lactobacillus plantarum species group were identified and typed. They represented 32 clones of Lactobacillus plantarum and 7 clones of Lactobacillus pentosus. Sensitivity of all strains towards bacteriocins of four different producer strains was investigated using a deferred inhibition test (DIT). Substantial intra-specific variation in sensitivity of clones was observed towards bacteriocinogenic lactic acid bacteria producing nisin ( Lactococcus lactis ATCC 11454) or pediocin PA-1 ( Pediococcus acidilactici PAC-1.0), while none of the strains were sensitive towards the two remaining bacteriocin producers. The minimum inhibitory concentration (MIC) of nisin towards selected strains confirmed the DIT results. No correlation between the susceptibility of fourteen selected strains towards nisin and an array of antibiotics was found. The present study indicates that the variation in bacteriocin-sensitivity within target species might be a potential limitation for the application of bacteriocins as biopreservatives.

Identification of lactic acid bacteria from spoilage associations of cooked and brined shrimps stored under modified atmosphere between 0 degrees C and 25 degrees C.

AIMS: To evaluate spoilage and identify lactic acid bacteria (LAB) from spoilage associations of cooked and brined shrimps stored under modified atmosphere packaging (MAP) at 0, 5, 8, 15 and 25 degrees C. METHODS AND RESULTS: Bacterial isolates (102) from spoilage associations of cooked and brined MAP shrimps were characterized by phenotypic tests and identified as lactic acid bacteria (78 isolates), other Gram-positive bacteria (13 isolates) and Gram-negative bacteria (11 isolates). A selection of 48 LAB isolates were further characterized and identified by phenotypic tests and SDS-PAGE electrophoresis of whole cell proteins. Selected clusters of LAB isolates were analysed by plasmid profiling, pulsed field gel electrophoresis and 16S rRNA gene sequencing. Enterococcus faecalis was identified in spoilage associations at 15 degrees C and 25 degrees C, and its metabolic activity corresponded to chemical changes in spoiled products. Carnobacterium divergens, a non-motile Carnobacterium sp. nov. and Lactobacillus curvatus were the LAB species observed in spoilage associations of products stored at 0 degrees C, 5 degrees C and 8 degrees C. CONCLUSIONS: Enterococcus spp. and Carnobacterium spp. were the dominant parts of spoilage associations of cooked and brined MAP shrimps stored at high and low temperatures, respectively. SIGNIFICANCE AND IMPACT OF THE STUDY: The SDS-PAGE technique and simple biochemical keys allowed the majority of LAB isolates from spoilage associations of cooked and brined MAP shrimps to be identified at the species level.

Lactobacillus durianis sp. nov., isolated from an acid-fermented condiment (tempoyak) in Malaysia.

Lactic acid bacteria (LAB) are the predominant micro-organisms in tempoyak, a Malaysian acid-fermented condiment. In a study on the diversity of LAB in this product, three isolates could not be identified using SDS-PAGE of whole-cell proteins or API 50 CH. The taxonomic position of the three isolates was clarified in the present study. 16S rDNA sequencing classified a representative strain in the genus Lactobacillus, clearly separated from all known species, and most closely related to the Lactobacillus reuteri phylogenetic group. DNA-DNA hybridization experiments and an extensive phenotypic description confirm that the strains represent a single and separate novel species among the obligately heterofermentative lactobacilli. The three isolates are distinguished at the intra-species level by plasmid profiling, pulsed-field gel electrophoresis of macro-restriction fragments and biochemical features. The name Lactobacillus durianis sp. nov. is proposed for the novel taxon and the type strain is LMG 19193T (= CCUG 45405T).

Microbial analysis of Malaysian tempeh, and characterization of two bacteriocins produced by isolates of Enterococcus faecium.

AIMS: Isolation of bacteriocinogenic lactic acid bacteria (LAB) from the Malaysian mould-fermented product tempeh and characterization of the produced bacteriocin(s). METHODS AND RESULTS: LAB were present in high numbers in final products as well as during processing. Isolates, Enterococcus faecium B1 and E. faecium B2 (E. faecium LMG 19827 and E. faecium LMG 19828, respectively) inhibited Gram-positive indicators, including Listeria monocytogenes. Partially purified bacteriocins showed a proteinaceous nature. Activity was stable after heat-treatment except at alkaline pH values. Both strains displayed a bacteriostatic mode of action. Bacteriocin production was associated with late exponential/early stationary growth. Molecular mass, calculated by SDS-PAGE, was 3.4 kDa for B1 bacteriocin, and 3.4 kDa and 5.8 kDa for B2 bacteriocins. PCR screening of enterocin-coding genes revealed three amplified fragments in total genomic DNA that may correspond with PCR signals for enterocin P, enterocin L50A and enterocin L50B. Both B1 and B2 contained a 42-kb plasmid. No differences in bacteriocinogenic capacity were found between wild type strains and plasmid-cured strains. CONCLUSIONS: It was possible to isolate bacteriocinogenic E. faecium active against various Gram-positive bacteria from final products of tempeh. SIGNIFICANCE AND IMPACT OF THE STUDY: A first step in applying biopreservation to fermented South-east Asian foods is to obtain bacteriocinogenic LAB from this source. Such isolates may also be used for biopreservation of mould-fermented foods in general, including various types of mould-ripened cheese.