Anionic nanocellulose as competing agent in microbial DNA extraction from mine process samples.

Microorganisms in flotation and minerals processing may significantly affect the grade and yield of metal concentrates. However, studying the phenomena requires working techniques to detach microorganisms and their DNA from mineral particles to which they strongly adhere. We developed a new method utilizing the competitive properties of anionic nanocellulose to block sorption of DNA to and detach microbial cells from mineral particles from ore processing. In general, up to one ng DNA mL-1 sample was obtained with the custom anionic nanocellulose method (CM) compared to DNA amounts below the Qubit assay's detection limit for extractions with a commercial kit (KIT). Similarly, 0.5-4 orders of magnitude more bacterial 16S and fungal 5.8S rRNA gene copies were detected by qPCR from CM treated samples compared to KIT extractions. A clear difference in the detected microbial community structure between CM and KIT extracted samples was also observed. Commercial kits optimized for mineral soils are easy to use and time efficient but may miss a considerable part of the microbial communities. A competing agent such as anionic nanocellulose may decrease the interaction between microorganisms or their DNA and minerals and provide a comprehensive view into the microbial communities in mineral processing environments.

Role of aryl hydrocarbon receptor (AHR) in overall retinoid metabolism: Response comparisons to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure between wild-type and AHR knockout mice.

Young adult wild-type and aryl hydrocarbon receptor knockout (AHRKO) mice of both sexes and the C57BL/6J background were exposed to 10 weekly oral doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; total dose of 200 µg/kg bw) to further characterize the observed impacts of AHR as well as TCDD on the retinoid system. Unexposed AHRKO mice harboured heavier kidneys, lighter livers and lower serum all-trans retinoic acid (ATRA) and retinol (REOH) concentrations than wild-type mice. Results from the present study also point to a role for the murine AHR in the control of circulating REOH and ATRA concentrations. In wild-type mice, TCDD elevated liver weight and reduced thymus weight, and drastically reduced the hepatic concentrations of 9-cis-4-oxo-13,14-dihydro-retinoic acid (CORA) and retinyl palmitate (REPA). In female wild-type mice, TCDD increased the hepatic concentration of ATRA as well as the renal and circulating REOH concentrations. Renal CORA concentrations were substantially diminished in wild-type male mice exclusively following TCDD-exposure, with a similar tendency in serum. In contrast, TCDD did not affect any of these toxicity or retinoid system parameters in AHRKO mice. Finally, a distinct sex difference occurred in kidney concentrations of all the analysed retinoid forms. Together, these results strengthen the evidence of a mandatory role of AHR in TCDD-induced retinoid disruption, and suggest that the previously reported accumulation of several retinoid forms in the liver of AHRKO mice is a line-specific phenomenon. Our data further support participation of AHR in the control of liver and kidney development in mice.

First insights to the microbial communities in the plant process water of the multi-metal Kevitsa mine.

Metallurgical processes demand large quantities of water. However, in many locations, water is becoming scarce and process water recycling is needed. Closing water loops can be challenging due to build-up of flotation chemicals, metal ions and microorganisms in the recycled water affecting the flotation performance. Here, we have characterized the microbial communities over a 2-month period in different locations of the multi-metal Kevitsa mine in Northern Finland, by microbiome sequencing, enumeration of bacteria, archaea and fungi by qPCR, and cultivation. The microbial communities showed high diversity, but were dominated by Alpha- and Gammaproteobacteria. In addition, various fungal taxa were detected, whereas the archaeal taxa were only sparsely detected from the sequence data. The number of bacterial 16S rRNA gene copies in Process water and Ni thickener overflow varied between 0.5-3.3 × 105 mL-1, whereas the Flotation tailings showed two orders of magnitude lower amounts. Fungi were present at 3.0 × 102-8.1 × 104 5.8S rRNA gene copies mL-1 in all samples, while the number of archaea fluctuated between 8.8 × 101-3.2 × 105 16S rRNA gene copies mL-1. The number of all microbial groups were generally lower in September than in August. When tested on 8 different cultivation media, the microorganisms generally responded positively to organic carbon, and were also shown to oxidize thiosulfate, which may indicate that build-up of organic flotation chemicals and sulfur species from the ore may cause the microbial numbers to increase. This study is part of the H2020 ITERAMS project (Grant agreement# 730480), which strives to improve the recycling of water and minimize the environmental impact of mines.

Transgenerational epigenetic and transcriptomic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure in rat.

In rats, direct exposure to TCDD causes myriad toxicities. Exposed rats experience hepatotoxicity, wasting syndrome and immune suppression, amongst others. "Inherited exposure", as occurs in the F3 generation of directly exposed F0 animals, has also been shown to cause toxicity: both male and female F3 rats demonstrate an increased incidence of adult onset disease, females also display reproductive abnormalities and increased incidence of ovarian diseases while males show increased incidence of kidney disease and an altered sperm epigenome. Here, we explore the hepatic transcriptomic profile of male and female F3 Sprague-Dawley rats bred through the paternal germ line from F0 dams exposed to a single dose of TCDD (0, 30, 100, 300 or 1000 ng/kg body weight) by oral gavage. We hypothesize that RNA transcripts with altered abundance in livers of unexposed F3 progeny of treated F0 Sprague-Dawley rats may result from epigenetic modifications to the genome. We further survey patterns of differential methylation within male F3 rat testis. Female F3 rats demonstrated more TCDD-mediated hepatic transcriptomic changes than males, with differences primarily in the lowest dose group. In testis from male F3 rats, multiple olfactory receptors displayed patterns of differential methylation. Hypermethylation of Egfr and Mc5r among testes from TCDD lineage rats was observed, but without corresponding changes in hepatic mRNA abundance. Further studies examining these differences in other tissue types are warranted.

Data on the optimization of an archaea-specific probe-based qPCR assay.

Estimation of archaeal numbers by use of fluorescent DNA binding dyes is challenging, because primers targeting the archaeal 16SrRNA genes readily also bind to bacterial 16S rRNA gene sequences, especially when the relative abundance of bacteria is greater than that of archaea. In order to increase specificity, we optimized a fluorescent probe-based assay using previously published archaeal primers and probe. The assay was tested on genomic DNA of pure bacterial and archaeal cultures and optimized using PCR amplicons of the archaeal pure cultures. The used bacterial strains showed slight amplification using the fluorescent dye assay, whereas all archaeal strains could be amplified with the archaea primers used. Due to differences in genome size and number of 16S rRNA gene copies between the tested archaeal strains, the amplification level varied greatly between the strains. Therefore, we also tested the amplification using PCR amplified fragments of the archaeal 16S rRNA genes. The tests with the archaeal 16S rRNA gene amplicons showed good amplification, although the amplification efficiency still varied between archaeal strains. The qPCR assay was used to estimate the archaeal numbers in process water of a multi-metal mine's metallurgical plant [1] and will be used in similar future microbiological analysis included in the H2020 ITERAMS project (Grant agreement# 730480).

Oil degradation potential of microbial communities in water and sediment of Baltic Sea coastal area.

Two long-term potentially oil exposed Baltic Sea coastal sites near old oil refineries and harbours were compared to nearby less exposed sites in terms of bacterial, archaeal and fungal microbiomes and oil degradation potential. The bacterial, archaeal and fungal diversities were similar in oil exposed and less exposed sampling sites based on bacterial and archaeal 16S rRNA gene and fungal 5.8S rRNA gene amplicon sequencing from both DNA and RNA fractions. The number of genes participating in alkane degradation (alkB) or PAH-ring hydroxylation (PAH-RHDα) were detected by qPCR in all water and sediment samples. These numbers correlated with the number of bacterial 16S rRNA gene copies in sediment samples but not with the concentration of petroleum hydrocarbons or PAHs. This indicates that both the clean and the more polluted sites at the Baltic Sea coastal areas have a potential for petroleum hydrocarbon degradation. The active community (based on RNA) of the coastal Baltic Sea water differed largely from the total community (based on DNA). The most noticeable difference was seen in the bacterial community in the water samples were the active community was dominated by Cyanobacteria and Proteobacteria whereas in total bacterial community Actinobacteria was the most abundant phylum. The abundance, richness and diversity of Fungi present in water and sediment samples was in general lower than that of Bacteria and Archaea. Furthermore, the sampling location influenced the fungal community composition, whereas the bacterial and archaeal communities were not influenced. This may indicate that the fungal species that are adapted to the Baltic Sea environments are few and that Fungi are potentially more vulnerable to or affected by the Baltic Sea conditions than Bacteria and Archaea.

Diversity and functionality of archaeal, bacterial and fungal communities in deep Archaean bedrock groundwater.

The diversity and metabolic functions of deep subsurface ecosystems remain relatively unexplored. Microbial communities in previously studied deep subsurface sites of the Fennoscandian Shield are distinctive to each site. Thus, we hypothesized that the microbial communities of the deep Archaean bedrock fracture aquifer in Romuvaara, northern Finland, differ both in community composition and metabolic functionality from the other sites in the Fennoscandian Shield. We characterized the composition, functionality and substrate preferences of the microbial communities at different depths in a 600 m deep borehole. In contrast to other Fennoscandian deep biosphere communities studied to date, iron-oxidizing Gallionella dominated the bacterial communities, while methanogenic and ammonia-oxidizing archaea were the most prominent archaea, and a diverse fungal community was also detected. Potential for methane cycling and sulfate and nitrate reduction was confirmed by detection of the functional genes of these metabolic pathways. Organotrophs were less abundant, although carbohydrates were the most preferred of the tested substrates. The microbial communities shared features with those detected from other deep groundwaters with similar geochemistry, but the majority of taxa distinctive to Romuvaara are different from the taxa previously detected in saline deep groundwater in the Fennoscandian Shield, most likely because of the differences in water chemistry.

Microbiome composition and geochemical characteristics of deep subsurface high-pressure environment, Pyhäsalmi mine Finland.

Pyhäsalmi mine in central Finland provides an excellent opportunity to study microbial and geochemical processes in a deep subsurface crystalline rock environment through near-vertical drill holes that reach to a depth of more than two kilometers below the surface. However, microbial sampling was challenging in this high-pressure environment. Nucleic acid yields obtained were extremely low when compared to the cell counts detected (1.4 × 10(4) cells mL(-1)) in water. The water for nucleic acid analysis went through high decompression (60-130 bar) during sampling, whereas water samples for detection of cell counts by microscopy could be collected with slow decompression. No clear cells could be identified in water that went through high decompression. The high-pressure decompression may have damaged part of the cells and the nucleic acids escaped through the filter. The microbial diversity was analyzed from two drill holes by pyrosequencing amplicons of the bacterial and archaeal 16S rRNA genes and from the fungal ITS regions from both DNA and RNA fractions. The identified prokaryotic diversity was low, dominated by Firmicute, Beta- and Gammaproteobacteria species that are common in deep subsurface environments. The archaeal diversity consisted mainly of Methanobacteriales. Ascomycota dominated the fungal diversity and fungi were discovered to be active and to produce ribosomes in the deep oligotrophic biosphere. The deep fluids from the Pyhäsalmi mine shared several features with other deep Precambrian continental subsurface environments including saline, Ca-dominated water and stable isotope compositions positioning left from the meteoric water line. The dissolved gas phase was dominated by nitrogen but the gas composition clearly differed from that of atmospheric air. Despite carbon-poor conditions indicated by the lack of carbon-rich fracture fillings and only minor amounts of dissolved carbon detected in formation waters, some methane was found in the drill holes. No dramatic differences in gas compositions were observed between different gas sampling methods tested. For simple characterization of gas composition the most convenient way to collect samples is from free flowing fluid. However, compared to a pressurized method a relative decrease in the least soluble gases may appear.

In utero/lactational and adult exposures to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) show differential effects on craniofacial development and growth in rats.

In a previous study of female Han/Wistar (H/W) and Long-Evans (L-E) rats, we found that adult exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was associated with size decreases in the cranium and especially the face. In this study we compared these crania to those from male and female Sprague-Dawley (S-D) rats with in utero/lactational exposure to TCDD, using morphometric variables of size, shape, and fluctuating asymmetry to quantify the effects of dose on craniofacial development and growth. At the highest levels of exposure, in utero/lactational and adult TCDD exposures both resulted in small but significant reductions in facial size parameters (i.e., 3-5%) in only females and minor effects on facial shape in both sexes. The shape effects of in utero/lactational exposure were most significant at the sutural intersections, whereas adult exposure to TCDD corresponded to dose-dependent changes of decreasing facial length and vault breadth. Fluctuating asymmetry in general explained a relatively small amount of shape variation compared with other effects, and only increased significantly in female L-E rats with high levels of adult exposure to TCDD. These results indicate that TCDD-related changes in cranial development and growth in rats can vary with the timing and duration of exposure, and with sex. Further investigations of other dioxin-like compounds and animal species will broaden our understanding of how chemicals exposure can influence the development and growth of the mammalian skeleton.

Revealing the unexplored fungal communities in deep groundwater of crystalline bedrock fracture zones in Olkiluoto, Finland.

The diversity and functional role of fungi, one of the ecologically most important groups of eukaryotic microorganisms, remains largely unknown in deep biosphere environments. In this study we investigated fungal communities in packer-isolated bedrock fractures in Olkiluoto, Finland at depths ranging from 296 to 798 m below surface level. DNA- and cDNA-based high-throughput amplicon sequencing analysis of the fungal internal transcribed spacer (ITS) gene markers was used to examine the total fungal diversity and to identify the active members in deep fracture zones at different depths. Results showed that fungi were present in fracture zones at all depths and fungal diversity was higher than expected. Most of the observed fungal sequences belonged to the phylum Ascomycota. Phyla Basidiomycota and Chytridiomycota were only represented as a minor part of the fungal community. Dominating fungal classes in the deep bedrock aquifers were Sordariomycetes, Eurotiomycetes, and Dothideomycetes from the Ascomycota phylum and classes Microbotryomycetes and Tremellomycetes from the Basidiomycota phylum, which are the most frequently detected fungal taxa reported also from deep sea environments. In addition some fungal sequences represented potentially novel fungal species. Active fungi were detected in most of the fracture zones, which proves that fungi are able to maintain cellular activity in these oligotrophic conditions. Possible roles of fungi and their origin in deep bedrock groundwater can only be speculated in the light of current knowledge but some species may be specifically adapted to deep subsurface environment and may play important roles in the utilization and recycling of nutrients and thus sustaining the deep subsurface microbial community.

Toxicological profile of ultrapure 2,2',3,4,4',5,5'-heptachlorbiphenyl (PCB 180) in adult rats.

PCB 180 is a persistent non-dioxin-like polychlorinated biphenyl (NDL-PCB) abundantly present in food and the environment. Risk characterization of NDL-PCBs is confounded by the presence of highly potent dioxin-like impurities. We used ultrapure PCB 180 to characterize its toxicity profile in a 28-day repeat dose toxicity study in young adult rats extended to cover endocrine and behavioral effects. Using a loading dose/maintenance dose regimen, groups of 5 males and 5 females were given total doses of 0, 3, 10, 30, 100, 300, 1000 or 1700 mg PCB 180/kg body weight by gavage. Dose-responses were analyzed using benchmark dose modeling based on dose and adipose tissue PCB concentrations. Body weight gain was retarded at 1700 mg/kg during loading dosing, but recovered thereafter. The most sensitive endpoint of toxicity that was used for risk characterization was altered open field behavior in females; i.e. increased activity and distance moved in the inner zone of an open field suggesting altered emotional responses to unfamiliar environment and impaired behavioral inhibition. Other dose-dependent changes included decreased serum thyroid hormones with associated histopathological changes, altered tissue retinoid levels, decreased hematocrit and hemoglobin, decreased follicle stimulating hormone and luteinizing hormone levels in males and increased expression of DNA damage markers in liver of females. Dose-dependent hypertrophy of zona fasciculata cells was observed in adrenals suggesting activation of cortex. There were gender differences in sensitivity and toxicity profiles were partly different in males and females. PCB 180 adipose tissue concentrations were clearly above the general human population levels, but close to the levels in highly exposed populations. The results demonstrate a distinct toxicological profile of PCB 180 with lack of dioxin-like properties required for assignment of WHO toxic equivalency factor. However, PCB 180 shares several toxicological targets with dioxin-like compounds emphasizing the potential for interactions.

New insights to the role of aryl hydrocarbon receptor in bone phenotype and in dioxin-induced modulation of bone microarchitecture and material properties.

Bone is a target for high affinity aryl hydrocarbon receptor (AHR) ligands, such as dioxins. Although bone morphology, mineral density and strength are sensitive endpoints of dioxin toxicity, less is known about effects on bone microarchitecture and material properties. This study characterizes TCDD-induced modulations of bone tissue, and the role of AHR in dioxin-induced bone toxicity and for normal bone phenotype. Six AHR-knockout (Ahr(-/-)) and wild-type (Ahr(+/+)) mice of both genders were exposed to TCDD weekly for 10 weeks, at a total dose of 200µg/kgbw. Bones were examined with micro-computed tomography, nanoindentation and biomechanical testing. Serum levels of bone remodeling markers were analyzed, and the expression of genes related to osteogenic differentiation was profiled using PCR array. In Ahr(+/+) mice, TCDD-exposure resulted in harder bone matrix, thinner and more porous cortical bone, and a more compact trabecular bone compartment. Bone remodeling markers and altered expression of a number of osteogenesis related genes indicated imbalanced bone remodeling. Untreated Ahr(-/-) mice displayed a slightly modified bone phenotype as compared with untreated Ahr(+/+) mice, while TCDD exposure caused only a few changes in bones of Ahr(-/-) mice. Part of the effects of both TCDD-exposure and AHR-deficiency were gender dependent. In conclusion, exposure of adult mice to TCDD resulted in harder bone matrix, thinner cortical bone, mechanically weaker bones and most notably, increased trabecular bone volume fraction in Ahr(+/+) mice. AHR is involved in bone development of a normal bone phenotype, and is crucial for manifestation of TCDD-induced bone alterations.

Unexpected gender difference in sensitivity to the acute toxicity of dioxin in mice.

The acute toxicity of the ubiquitous environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) varies widely among species and strains. Previous studies in rats have established that females are approximately 2-fold more sensitive to TCDD lethality than males. However, there is a surprising gap in the literature regarding possible gender-related sensitivity differences in mice. In the present study, by using three substrains of TCDD-sensitive C57BL/6 mice and transgenic mice on this background, we demonstrated that: 1) in contrast to the situation in rats, female mice are the more resistant gender; 2) the magnitude of the divergence between male and female mice depends on the substrain, but can amount to over 10-fold; 3) AH receptor protein expression levels or mutations in the primary structure of this receptor are not involved in the resistance of female mice of a C57BL/6 substrain, despite their acute LD50 for TCDD being over 5000 µg/kg; 4) transgenic mice that globally express the rat wildtype AH receptor follow the mouse type of gender difference; 5) in gonadectomized mice, ovarian estrogens appear to enhance TCDD resistance, whereas testicular androgens seem to augment TCDD susceptibility; and 6) the gender difference correlates best with the severity of liver damage, which is also reflected in hepatic histopathology and the expression of pro-inflammatory cytokines, especially IL-6. Hence, the two closely related rodent species most often employed in toxicological risk characterization studies, rat and mouse, represent opposite examples of the influence of gender on dioxin sensitivity, further complicating the risk assessment of halogenated aromatic hydrocarbons.

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure on bone material properties.

Dioxins are known to decrease bone strength, architecture and density. However, their detailed effects on bone material properties are unknown. Here we used nanoindentation methods to characterize the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on nanomechanical behaviour of bone matrix. Pregnant rats were treated with a single intragastric dose of TCDD (1 microg/kg) or vehicle on gestational day 11. Tibias of female offspring were sampled on postnatal day (PND) 35 or 70, scanned at mid-diaphysis with pQCT, and evaluated by three-point bending and nanoindentation. TCDD treatment decreased bone mineralization (p<0.05), tibial length (p<0.01), cross-sectional geometry (p<0.05) and bending strength (p<0.05). Controls showed normal maturation pattern between PND 35 and 70 with decreased plasticity by 5.3% and increased dynamic hardness, storage and complex moduli by 26%, 13% and 12% respectively (p<0.05), while similar maturation was not observed in TCDD-exposed pups. In conclusion, for the first time, we demonstrate retardation of bone matrix maturation process in TCDD-exposed animals. In addition, the study confirms that developmental TCDD exposure has adverse effects on bone size, strength and mineralization. The current results in conjunction with macromechanical behaviour suggest that reduced bone strength caused by TCDD is more associated with the mineralization and altered geometry of bones than with changes at the bone matrix level.

Ecology of Listeria spp. in a fish farm and molecular typing of Listeria monocytogenes from fish farming and processing companies.

This study focused on the ecology of Listeria monocytogenes in a fish farm by following the changes in its occurrence in different types of samples for a three year period. In addition, L. monocytogenes isolates from different seafood industry areas were compared with pulsed field gel electrophoresis (PFGE) typing to discover possible associations between primary production, further processing and final products. Weather conditions were found to have a strong influence on the probability of finding Listeria spp. in a fish farm environment. The number of samples contaminated with Listeria spp. was typically bigger after rainy periods. Brook and river waters as well as other runoff waters seemed to be the main contamination source at the farm studied. The farmed fish originally found to carry L. monocytogenes become gradually Listeria free. The time needed for the purification of the fish was several months. The sea bottom soil samples were the ones that preserved the L. monocytogenes contamination the longest time. It can be stated that the fish and fish farm equipment studied did not spread listeria contamination. On the contrary, they were found to suffer from listeria contamination coming from outside sources like the brook water. There was a wide range of different L. monocytogenes PFGE-pulsotypes (30) found at 15 Finnish fish farms and fish processing factories. L. monocytogenes isolates from the final products often belonged to the same pulsotypes as did the isolates from the processing environment as well as from the raw fish. This suggests that, in addition to the fish processing factory environment, the fish raw materials are important sources of L. monocytogenes contamination in final products.

The effect of perinatal TCDD exposure on caries susceptibility in rats.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), the model compound of polychlorinated dibenzo-p-dioxins and furans, is a potent toxicant with the ability to hamper development. Accidental exposure to TCDD has been linked with various developmental dental aberrations in humans, and experimentally it has been shown that TCDD causes, among other defects, hypomineralization of dental hard tissues in rodents. Here, we studied the effect of very low perinatal TCDD exposure on dental caries susceptibility and mineral composition of tooth enamel in rats. Pregnant line C rats (rat line developed in our laboratory) were dosed 0.03-1.0 microg/kg TCDD on gestation day 15 and allowed to give birth and nurse until weaning on postnatal day 21. The offspring were challenged with cariogenic treatment including sugar-rich diet and three inoculations with Streptococcus mutans. Control groups involved animals with or without cariogenic challenge or TCDD treatment. The number of caries lesions in left lower molars was determined by Schiff's staining after 8 weeks of weaning. TCDD treatment increased cariogenic lesions in the enamel at the lowest maternal dose used, 0.03 microg/kg, and at the highest maternal dose, 1 microg/kg, the lesions extended through the enamel to dentin more frequently. Changes in mineral composition measured by electron probe microanalyzer, scanning electron microscopy, and energy-dispersive spectrometry could not explain the increased caries susceptibility. In conclusion, perinatal TCDD exposure can render rat molars more susceptible to caries.

Prevalence and location of Listeria monocytogenes in farmed rainbow trout.

A total of 510 rainbow trout originating from fish farms in lakes and sea areas around Finland were studied for the presence of Listeria monocytogenes. Samples were studied as pools from five fish. Gill, viscera, and skin from the pooled samples were analysed separately. The individual samples were analysed later if the pooled sample was found to be Listeria positive. The prevalence of Listeria spp. and L. monocytogenes in pooled unprocessed fresh rainbow trout was on average 35.0% and 14.6%, respectively. On the other hand, the prevalence of Listeria spp. and L. monocytogenes in individual thawed fish was found to be 14.3% and 8.8%, respectively. These numbers tend to overestimate and underestimate the real situation because not all fish in pooled samples were necessarily contaminated and in some of the Listeria positive pooled samples all individual samples turned out to be Listeria free. The prevalence of L. monocytogenes varied greatly between different fish farms from zero to 100% in pooled samples and from zero to 75% according to individually studied fish samples. Some indications of the influence of weather conditions and seasonal variations that strongly affected the Listeria contamination of fish were also noticed. The location of Listeria spp. and L. monocytogenes in different parts of the fish differed with statistical significance in rainbow trout. Up to 95.6% of the L. monocytogenes and 84.5% of Listeria spp. positive samples were gill samples. Only 4.4% (2/45) of the L. monocytogenes positive samples were obtained from skin or viscera. Closer study at one fish farm revealed that there was only one L. monocytogenes ribotype present in the contaminated fish, although water and surfaces were heavily contaminated with six other L. monocytogenes ribotypes.

Effects of in utero and lactational TCDD exposure on bone development in differentially sensitive rat lines.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a notorious model compound of highly toxic environmental pollutants, polychlorinated dibenzo-p-dioxins (PCDDs). Their toxic effects are mediated via cytosolic aryl hydrocarbon receptor (AHR). We studied the effects of several dose levels of TCDD on developing rat bone after maternal exposure at different times of gestation and lactation in three differentially sensitive rat lines. Rat lines A, B, and C differ in their sensitivity to TCDD due to mutated AHR (Ahr(hw)) in line A and another TCDD-resistance allele (B(hw)) in line B. Line C rats have no resistance alleles. Offspring were analyzed for bone mineral density and geometry by peripheral quantitative computed tomography (pQCT) and for bone biomechanics by three-point bending at mid-diaphysis of tibia and femur and by axial loading at femoral neck. TCDD treatment resulted in bone defects, mainly in offspring of the most sensitive line C at a maternal dose of 1 microg/kg. They included decreased bone length, cross-sectional area of cortex, and bone mineral density. Mechanical testing revealed significantly reduced bending breaking force and stiffness of tibia, femur, and femoral neck. The effects were exposure time-dependent, and earlier exposure caused more severe defects. Gestational exposure alone was not sufficient, but lactational exposure was required to cause the bone defects. Most of the defects were recovered at the age of 1 year. The results indicate that dioxins affect developing bone by interfering with bone growth and mechanical strength and that the effects are mainly reversible. The dioxin-resistance alleles, Ahr(hw) and B(hw) increase the resistance to these defects.

Pasteurization of rainbow trout roe: Listeria monocytogenes and sensory analyses.

D- and z-values for a mixture of four Listeria monocytogenes strains originating from the roe of different fish species were determined in rainbow trout (Oncorhynchus mykiss) roe. The D60- and D63-values obtained were 1.60 and 0.44 min, respectively, and the z-value was 5.36 degrees C accordingly. In pilot-scale experiments, rainbow trout roe (100 g) was vacuum packaged into glass jars and pasteurized both at 62 and 65 degrees C for 10 min. These treatments were enough to destroy 10(8) CFU/ g of L. monocytogenes cells, which was the highest possible Listeria cell count to grow in roe. On the basis of the determined z-value and calculation of pasteurization values, these experimental pasteurizations were found to theoretically destroy at least 45 log units of L. monocytogenes cells in rainbow trout roe. In addition, these pasteurization treatments did not significantly affect the sensory quality of the roe. The sensory quality of pasteurized vacuum-packaged rainbow trout roe stored at 3 degrees C was evaluated as good after 6 months of storage and not statistically different from the control that was frozen from the same roe lot as the pasteurized roe samples. Pasteurization of rainbow trout roe was proven to be an appropriate method for ensuring product safety with regard to L. monocytogenes and to stabilizing the sensory and microbial quality of roe. However, the safety risk caused by spore-forming bacteria still exists in pasteurized roe. Therefore, it has to be stored below 3 degrees C.