Clastogenicity and aneuploidy in newborn and adult mice exposed to 50 Hz magnetic fields.

PURPOSE: To detect possible clastogenic and aneugenic properties of a 50 Hz, 650 muT magnetic field. MATERIALS AND METHODS: The micronucleus test with CREST (Calcinosis, Raynaud's phenomenon, Esophageal dismotility, Sclerodactility, Telangectasia) antibody staining was performed on liver and peripheral blood sampled from newborn mice exposed to an ELF (Extremely Low Frequency) magnetic field during the whole intra-uterine life (21 days), and on bone marrow and peripheral blood sampled from adult mice exposed to the same magnetic field for the same period. RESULTS: Data obtained in newborn mice show a significant increase in micronuclei frequencies. In absolute terms, most of the induced micronuclei were CREST-negative (i.e., formed by a chromosome fragment). However, in relative terms, ELF exposure caused a two-fold increase in CREST-negative micronuclei and a four-fold increase in CREST-positive micronuclei (i.e., formed by a whole chromosome). No significant effect was recorded on exposed adults. CONCLUSIONS: These findings suggest the need for investigation of aneugenic properties of ELF magnetic fields in order to establish a possible relationship to carcinogenesis.

Environmental impact of the Chernobyl accident: mutagenesis in bank voles from Sweden.

An investigation was carried out in Sweden aimed at studying the possible genetic effects of the Chernobyl fallout on wild small mammals. The bank voles (Clethrionomys glareolus Schreb.) were obtained from three differently contaminated areas in Sweden and, for control, in an area with negligible contamination by fallout. Radionuclide determinations to assess the content of 137Cs and mutagenicity tests (bone marrow micronucleus test and sperm abnormality assay) were performed. The results obtained showed a positive correlation between the increase of micronucleated polychromatic erythrocytes (MPCE/1000 PCE) and both 137Cs content in muscle and in soil contamination. The estimated doses absorbed by the animals were far lower than those required for the same effect in laboratory experiments. An explanation of this discrepancy between dose and measured biological effect is not available, yet similar results have been repeatedly reported after the Chernobyl accident and should be a matter for further discussion. An increased frequency of micronucleated cells might occur at minimal dose gradients, and the micronucleus test appears to be a valid tool to show such effects.

Genetic damage in urban mice exposed to traffic pollution.

Wild rodents (Mus domesticus) were collected in three areas in Rome exposed to different traffic flows to ascertain a possible correlation between genetic damage and heavy metal concentration. The concentration of lead, cadmium and zinc were determined in liver, kidney and bones and two mutagenicity tests (micronucleus test and sperm abnormality assay) were employed. The results obtained showed that the contents of lead and cadmium were higher in animals collected in areas with high traffic flows than in those from control areas. A statistically significant increase of the frequency of micronucleated erythrocytes and of abnormal sperm cells was also obtained in animals collected in sites with high traffic flows. The investigation confirmed the suitability of using wild rodents as bioindicators of environmental pollution and as key-organisms in programs of pollution monitoring and environmental conservation.

137Cs determination and mutagenicity tests in wild Mus musculus domesticus before and after the Chernobyl accident.

Wild rodents were collected alive in a suburban area to the north of Rome before, during, 6 months and 1 year after the Chernobyl accident. Radionuclide determinations to assess their content of (137)Cs and mutagenicity tests (bone-marrow micronucleus test and sperm abnormalities assay) were performed on the captured animals. The results obtained for the species Mus musculus domesticus (which was the rodent captured in highest numbers), compared with the results obtained before the fallout, showed an increase of both micronucleated polychromatic erythrocytes (MPCEs/1000 PCEs) and (137)Cs content during and after the fallout.

Genotoxic damage in free-living Algerian mouse (Mus spretus) after the Coto Doñana ecological disaster.

The Doñana National Park (Spain), one of the most important wildlife sites in the West of Europe, was affected (25 April 1998) by the spill of acidic waste rich in toxic metals (mainly zinc, lead, copper, etc.), arsenic and aromatic amines from the Aznalcollar mine accident. Micronuclei test with May Grunwald-Giemsa and with CREST-antikinetocore staining using DAPI as counter-staining were performed on peripheral blood erythrocytes from Algerian mice to evaluate genotoxic damage. Animals were collected in four locations each differently affected by the disaster. Higher frequencies of micronuclei and CREST-positive micronuclei were observed in the sites, which were reached by toxic sludge and contaminated water in comparison with those located within the park. The results obtained applying the two methods indicate that DAPI staining is more sensitive in detecting micronuclei. Genotoxic biomonitoring should be further carried out in the area to control the mutagenetic level in natural populations.

Specific synaptic systems in reaggregated spherules from dissociated chick cerebellum cultivated in vitro.

Studies were performed on spherules of approximately 100-300 micrometer in diameter obtained from in vitro cultures of reaggregates of embryonic fragments of cerebellum from 10-12 day-incubated chick embryos, dissociated with trypsin and cultivated in a rotating shaker for a maximum of 21 days. The differentiated neurons within these spherules included a few Purkinje cells, many granule cells and type II Golgi cells, as well as many glial cells. Zones rich in synaptic knobs and other simple synaptic structures as well as complex synaptic systems with numerous active points of contact, were visible in various parts of the spherule. Typical "glomeruli" consisting of a varicosity of "rosette" joined to the dendritic claws of the granule cells, and "en marron" systems with perikarya of type II Golgi cells were easily recognised. The complete absence of extracerebellar afferents confirms that both the granule and Golgi cells are capable of making synaptic connections with afferents different from those normally formed by extracerebellar mossy or climbing fibres. The experimental findings confirm that the recipient neurons determine the specific synaptic pattern regardless of the nature of the afferents, and furthermore demonstrate that the clinging activity of the recipient neuron determines the synaptogenic behaviour of nervous pathways.

The Comet assay for the detection of DNA damage in Mus spretus from Doñana National Park.

Doñana Park (Spain), a protected area in Europe, was affected by an environmental disaster in April 1998 that caused the spreading of acidic water and mud full of toxic metals from the Aznalcollar pyrite mine. In order to assess the contamination in the area and to monitor the possible biological effects of the toxic spill, a series of coordinated studies was performed utilizing several animal species living in that area. We performed genotoxicity monitoring using the Comet assay on peripheral blood leukocytes of the Algerian mouse (Mus spretus), a nonprotected rodent suitable as a bioindicator. The mice were sampled in different areas 6 months after the ecological disaster and again 1 year later. Our results showed that in 1998 all the areas examined were contaminated, as determined by an increase in the Comet assay parameters in the analyzed animals, whereas a significant decrease in the values of these parameters was observed in the 1999 samples, which were collected in a riverside area subject to tide flows. Thus, the Comet assay has proven to be an interesting and sensitive tool in studies of environmental genotoxicity.

[Antibodies against Hantaan virus and Leptospira in subjects at risk in Rome]. / Anticorpi contro hantavirus e leptospire in soggetti a rischio in Roma.

A survey on the prevalence of Hantaan and leptospiral antibodies on mammalogists and rodent control personnel was performed. None of the 66 trappers studied (using IFI ) had detectable Hantaan antibody, while only 2 out of 20 mammalogists presented antibody at low titer (1:32). For leptospiral antibody the microagglutination test (MAT) using live leptospires as antigen was performed. 14 out of 66 trappers, or 21.2 per cent, had antibodies, at titer of 1:50 or more, to various leptospiral serovars: L.icterohaemorrhagiae in 12 cases, L.hardjo in 1 case, L.bratislava in 1 case. On the contrary, none of the mammalogists showed positivity for any of 16 serovars used. The environmental risk factors could justify the high prevalence of leptospiral antibodies in the field workers (trappers), while continuous laboratory contacts with rodents explain the presence of Hantaan virus antibodies in mammalogists .

CREST staining of micronuclei from free-living rodents to detect environmental contamination in situ.

In this work immunofluorescent antikinetochore (CREST) staining was used to analyse bone marrow micronuclei (MN) from free-living animals belonging to four different rodent species. Yellow-necked mice (Apodemus flavicollis) and bank voles (Clethrionomys glareolus) were trapped in the Czech Republic, Algerian mice (Mus spretus) in Spain and house mice (Mus musculus domesticus) in Italy. Animals were collected in areas displaying low or high environmental pollution in order to investigate the sensitivity of CREST analysis on bone marrow MN as a biomarker of environmental stress in situ. Differences in total MN frequencies between animals collected in control or contaminated areas were statistically significant for two species, whereas the differences in CREST+ MN were statistically significant for three species. Interestingly, the percentages of CREST+ MN in animals collected in the control areas were very low (3. 2-8.7%), suggesting that activities inducing alterations in the distribution of chromosomes are very rare in natural conditions. The increased frequencies of CREST+ MN observed in areas with high environmental impact indicate that activities producing loss of chromosomes at mitosis may be characteristic of anthropogenic environments such as industrial settlements around petrochemical factories. Our data suggest that the analysis of CREST+ MN may represent a sensitive end-point for the detection of environmental contamination by genotoxic xenobiotics, offering the advantage of providing information on the mechanism of action of environmental contaminants.