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We retrieved 34 medieval ovicaprine remains, from three archaeological sites of central Italy dating to about 1000 years old, and analyzed them using mitochondrial DNA. We compared the reconstructed haplogroups with modern sheep samples from Europe and the Middle East and sequences from the literature. In modern sheep, haplogroup HA is present in countries with access to the Mediterranean and close to the domestication center, whereas it is very rare or absent in the rest of Europe. The haplogroup HB was predominant in ancient samples (90%), whereas haplogroup HA was found at 10%. Ancient haplogroups match the present distribution in modern sheep in Italy, indicating that the current proportion of HA/HB was already established in the Middle Ages and is not the result of subsequent events such as selective breeding practices., (© 2015 Stichting International Foundation for Animal Genetics.)

A 721-bp fragment from 15,541 to 16,261 bp (NC_001941.1) of the mtDNA control region from different Egyptian and Italian sheep breeds was amplified. The PCR products were purified and sequenced. From the amplified fragment of 721-bp, a region of 423 bp after excluding a central region rich in tandem repeats was analyzed. Within all tested breeds, the haplotype diversity and average number of pairwise differences were 0.97571 and 7.01484, respectively. The genetic distances (D) and the average number of pairwise differences (Dxy) between breeds were estimated. The lowest distance was observed between Laticauda and Italian Muflon followed by distance between Sarda and Italian Muflon while the highest distance was observed between Barki and Sarda followed by distance between Barki and Laticauda. Phylogenetic analysis showed the presence of three haplogroups - HapA, HapB and HapC - in the examined samples with the absence of other two haplogroups HapD and HapE. All Italian samples cluster with B haplogroup and also in the Egyptian breeds the most dominant haplogroup was B (62 out of 67 analyzed samples). In Egyptian Barki breed one individual clusters with A haplogroup and another individual with C haplogroup. In Ossimi breed two individuals cluster with C haplogroup and in Rahmani there is one sample belonging to A haplogroup. The matrix of pairwise differences among breeds was used to perform a Principal Component Analysis (PCA). This analysis showed that the Italian breeds are clearly separated from the Egyptian breeds; moreover the Egyptian Barki breed is separated from Ossimi and Rahmani.

Analysis of DNA from archeological remains is a valuable tool to interpret the history of ancient animal populations. So far most studies of ancient DNA target mitochondrial DNA (mtDNA), which reveals maternal lineages, but only partially the relationships of current breeds and ancient populations. In this study we explore the feasibility of nuclear DNA analysis. DNA was extracted from 1000-years old cattle bone collected from Ferento, an archeological site in central Italy. Amplification of 15 microsatellite FAO-recommended markers with PCR products yielded genotypes for four markers. Expected heterozygosity was comparable with values of modern breeds, but observed heterozygosity was underestimated due to allelic loss. Genetic distances suggested a position intermediate between (1) Anatolian, Balkan, Sicilian and South-Italian cattle and (2) the Iberian, North-European and Central-European cattle, but also a clear relationship with two central-Italian breeds, Chianina and Romagnola. This suggests that these breeds are derived from medieval cattle living in the same area. Our results illustrate the potential of ancient DNA for reconstructing the history of local cattle husbandry.

In this study we used a medium density panel of SNP markers to perform population genetic analysis in five Italian cattle breeds. The BovineSNP50 BeadChip was used to genotype a total of 2,935 bulls of Piedmontese, Marchigiana, Italian Holstein, Italian Brown and Italian Pezzata Rossa breeds. To determine a genome-wide pattern of positive selection we mapped the F st values against genome location. The highest F st peaks were obtained on BTA6 and BTA13 where some candidate genes are located. We identified selection signatures peculiar of each breed which suggest selection for genes involved in milk or meat traits. The genetic structure was investigated by using a multidimensional scaling of the genetic distance matrix and a Bayesian approach implemented in the STRUCTURE software. The genotyping data showed a clear partitioning of the cattle genetic diversity into distinct breeds if a number of clusters equal to the number of populations were given. Assuming a lower number of clusters beef breeds group together. Both methods showed all five breeds separated in well defined clusters and the Bayesian approach assigned individuals to the breed of origin. The work is of interest not only because it enriches the knowledge on the process of evolution but also because the results generated could have implications for selective breeding programs.

Italy represented a crucial zone for migration and formation of sheep breeds. However, few data on Italian breeds have been published so far. We analysed seven Italian sheep breeds using mitochondrial DNA (mtDNA) sequencing to gain information on their genetic diversity and history. A 721 bp mtDNA control region fragment was amplified and sequenced in a total of 138 samples belonging to seven breeds and to Italian mohuflon (Ovis orientalis musimon) to investigate genetic diversity and phylogenetic evolution. We retrieved 68 variable sites and 79 haplotypes. The sheep breeds in our study are quite diverse, and phylogenetic analyses resulted in 3.6% of the samples belonging to A, 2.2% to D and 94.2% to B mtDNA haplogroups. Principal Component Analysis (PCA) showed a separation of breeds on both dimensions. The results of this study provide data on Italian breeds, presently scarcely investigated, and contribute to the knowledge of Italian sheep breeds and will be useful to the understanding of population genetics and breed evolution.

Background: Bovine Spastic Paresis (BSP) is a neuromuscular disorder which affects both male and female cattle. BSP is characterized by spastic contraction and overextension of the gastrocnemious muscle of one or both limbs and is associated with a scarce increase in body weight. This disease seems to be caused by an autosomal and recessive gene, with incomplete penetration, although no genes clearly involved with its onset have been so far identified. We employed cDNA microarrays to identify metabolic pathways affected by BSP in Romagnola cattle breed. Investigation of those pathways at the genome level can help to understand this disease., Results: Microarray analysis of control and affected individuals resulted in 268 differentially expressed genes. These genes were subjected to KEGG pathway functional clustering analysis, revealing that they are predominantly involved in Cell Communication, Signalling Molecules and Interaction and Signal Transduction, Diseases and Nervous System classes. Significantly enriched KEGG pathway's classes for the differentially expressed genes were calculated; interestingly, all those significantly under-expressed in the affected samples are included in Neurodegenerative Diseases. To identify genome locations possibly harbouring gene(s) involved in the disease, the chromosome distribution of the differentially expressed genes was also investigated., Conclusions: The cDNA microarray we used in this study contains a brain library and, even if carrying an incomplete transcriptome representation, it has proven to be a valuable tool allowing us to add useful and new information to a poorly studied disease. By using this tool, we examined nearly 15000 transcripts and analysed gene pathways affected by the disease. Particularly, our data suggest also a defective glycinergic synaptic transmission in the development of the disease and an alteration of calcium signalling proteins. We provide data to acquire knowledge of a genetic disease for which literature still presents poor results and that could be further and specifically analysed in the next future. Moreover this study, performed in livestock, may also harbour molecular information useful for understanding human diseases.

In this work, the genetic variation of milk FA was investigated in three different bovine breeds, the Jersey, the Piedmontese and the Valdostana, and at different lactation stages. All animals were genotyped for 21 Single Nucleotide Polymorphisms located within nine candidate genes involved in lipid synthesis: diacylglycerol acyltransferase 1 and 2 (DGAT1, 2); stearoyl-CoA desaturase (SCD); growth hormone receptor (GHR); fatty acid synthase (FASN); acyl-CoA dehydrogenase (ACAD); fatty acid binding protein (FABP4); lipoprotein lipase (LPL); and leptin gene (LEP). The highest milk-fat Jersey breed also showed the highest content of saturated FA. Throughout lactation, the breeds showed a similar variation in the FA, with a decrease in the short-chain, this was accompanied by a general increase in the long chain FA at the end of lactation. The increase in long chain saturated FA was particularly evident in the case of the Jersey. The effect of SCD gene on the C14 desaturation index was confirmed; the DGAT1 gene was polymorphic only in the Jersey breed, but its effect was confirmed only on milk fat content; three further potential candidate genes were identified: first, the FABP4 gene, which was found to influence medium and long chain FA in all the breeds, but not the desaturation indices; second, the FASN gene, which was found to influence the amount of PUFA in the Piedmontese and the Valdostana, and third, the LPL gene, which was found to affect fat content in the Piedmontese.

Calving in cattle is affected by calf morphology and by dam characteristics. It is described by two different traits: maternal calving ease, which is the ability to generate dams with good physiological predisposition to calving, and direct calving ease, which is the ability to generate calves that are easily born. The aim of this study was to identify regions of cattle genome harboring genes possibly affecting direct calving ease in the Piedmontese cattle breed. A population of 323 bulls scored for direct calving ease (EBV) was analyzed by a medium-density SNP marker panel (54,001 SNPs) to perform a genome-wide scan. The strongest signal was detected on chromosome 6 between 37.8 and 38.7 Mb where 13 SNPs associated to direct calving ease were found. Three genes are located in this region: LAP3, encoding for a leucine aminopeptidase involved in the oxytocin hydrolysis; NCAPG, encoding for a non-SMC condensin I complex, which has been associated in cattle with fetal growth and carcass size; and LCORL, which has been associated to height in humans and cattle. To further confirm the results of the genome-wide scan we genotyped additional SNPs within these genes and analyzed their association with direct calving ease. The results of this additional analysis fully confirmed the findings of the GWAS and particularly indicated LAP3 as the most probable gene involved. Linkage Disequilibrium (LD) analysis showed high correlation between SNPs located within LAP3 and LCORL indicating a possible selection signature due either to increased fitness or breeders' selection for the trait.

Background: X-linked anhidrotic ectodermal dysplasia is a disorder characterized by abnormal development of tissues and organs of ectodermal origin caused by mutations in the EDA gene. The bovine EDA gene encodes the ectodysplasin A, a membrane protein expressed in keratinocytes, hair follicles and sweat glands, which is involved in the interactions between cell and cell and/or cell and matrix. Four mutations causing ectodermal dysplasia in cattle have been described so far., Results: We identified a new single nucleotide polymorphism (SNP) at the 9th base of exon 8 in the EDA gene in two calves of Holstein Friesian cattle breed affected by ectodermal dysplasia. This SNP is located in the exonic splicing enhancer (ESEs) recognized by SRp40 protein. As a consequence, the spliceosome machinery is no longer able to recognize the sequence as exonic and causes exon skipping. The mutation determines the deletion of the entire exon (131 bp) in the RNA processing, causing a severe alteration of the protein structure and thus the disease., Conclusion: We identified a mutation, never described before, that changes the regulation of alternative splicing in the EDA gene and causes ectodermal dysplasia in cattle. The analysis of the SNP allows the identification of carriers that can transmit the disease to the offspring. This mutation can thus be exploited for a rational and efficient selection of unequivocally healthy cows for breeding.

We employed mtDNA and nuclear SNPs to investigate the genetic diversity of sheep breeds of three countries of the Mediterranean basin: Albania, Greece, and Italy. In total, 154 unique mtDNA haplotypes were detected by means of D-loop sequence analysis. The major nucleotide diversity was observed in Albania. We identified haplogroups, A, B, and C in Albanian and Greek samples, while Italian individuals clustered in groups A and B. In general, the data show a pattern reflecting old migrations that occurred in postneolithic and historical times. PCA analysis on SNP data differentiated breeds with good correspondence to geographical locations. This could reflect geographical isolation, selection operated by local sheep farmers, and different flock management and breed admixture that occurred in the last centuries.

Italian Maremmana, Turkish Grey and Hungarian Grey breeds belong to the same Podolic group of cattle, have a similar conformation and recently experienced a similar demographic reduction. The aim of this study was to assess the relationship among the analysed Podolic breeds and to verify whether their genetic state reflects their history. To do so, approximately 100 single nucleotide polymorphisms (SNPs) were genotyped on individuals belonging to these breeds and compared to genotypes of individuals of two Italian beef breeds, Marchigiana and Piemontese, which underwent different selection and migration histories. Population genetic parameters such as allelic frequencies and heterozygosity values were assessed, genetic distances calculated and assignment test performed to evaluate the possibility of recent admixture between the populations. The data show that the physical similarity among the Podolic breeds examined, and particularly between Hungarian Grey and Maremmana cattle that experienced admixture in the recent past, is mainly morphological. The assignment of individuals from genotype data was achieved using Bayesian inference, confirming that the set of chosen SNPs is able to distinguish among the breeds and that the breeds are genetically distinct. Individuals of Turkish Grey breed were clearly assigned to their breed of origin for all clustering alternatives, showing that this breed can be differentiated from the others on the basis of the allelic frequencies. Remarkably, in the Turkish Grey there were differences observed between the population of Enez district, where in situ conservation studies are practised, and that of Bandirma district of Balikesir, where ex situ conservation studies are practised out of the original raising area. In conclusion, this study demonstrates that molecular data could be used to reveal an unbiased view of past events and provide the basis for a rational exploitation of livestock, suggesting appropriate cross-breeding plans based on genetic distance or breeding strategies that include the population structure., (© 2010 Blackwell Verlag GmbH.)

The aim of the present study was to investigate the genetic control of the fatty acid (FA) composition in milk from 3 breeds of sheep: Altamurana, Gentile di Puglia, and Sarda. Single nucleotide polymorphisms within genes, encoding enzymes putatively involved in the synthesis and metabolism of milk fat, were selected for analysis, and the allele substitution effects were determined for 16 genes, which were polymorphic in the 3 sheep breeds, upon the milk fat composition. Four genes (alpha-1-antichymotrypsin-2; diacylglycerol O-acyltransferase homolog-2; propionyl Coenzyme A carboxylase, beta polypeptide; and insulin-like growth factor-I) play a role in the desaturation of stearic FA into polyunsaturated fatty acids. Furthermore, 2 genes (growth hormone receptor and zona pellucida glycoprotein-2) affect the variability of the total fat content in addition to the butyric and stearic FA profile, and the fatty acid synthetase gene has an influence on the medium-chain FA. Milk FA profiles play an important role in dairy sheep farming because they have a large effect on cheese characteristics and also because sheep milk may be marketed as a source of nutraceuticals because it contains higher levels of conjugated linoleic acid than milk from other ruminants. The current study evaluated the global effects of a large number of single nucleotide polymorphisms and haplotypes on traits that are not commonly investigated in sheep but that are potentially very useful for improving milk quality., (Copyright (c) 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

The genetic variation and relationships among six Turkish water buffalo populations, typical of different regions, were assessed using a set of 26 heterologous (bovine) microsatellite markers. Between seven and 17 different alleles were identified per microsatellite in a total of 254 alleles. The average number of alleles across all loci in all the analysed populations was found to be 12.57. The expected mean heterozygosity (H(e)) per population ranged between 0.5 and 0.58. Significant departures from Hardy-Weinberg equilibrium were observed for 44 locus-population combinations. Population differentiation was analysed by estimation of the F(st) index (values ranging from 0.053 to 0.123) among populations. A principal component analysis of variation revealed the Merzifon population to show the highest differentiation compared with the others. In addition, some individuals of the Danamandira population appeared clearly separated, while the Afyon, Coskun, Pazar and Thural populations represented a single cluster. The assignment of individuals to their source populations, performed using the Bayesian clustering approach implemented in the structure 2.2 software, supports a high differentiation of Merzifon and Danamandira populations. The results of this study are useful for the development of conservation strategies for the Turkish buffalo.

The Holstein Friesian and Chianina cattle breeds are representative of extreme selection for milk and meat traits, respectively, with significant changes in metabolism resulting from human selection over the past centuries. In the present study, we wanted to assess whether selection for different purposes has had a measurable effect on liver metabolism through a comparison of the protein and gene expression profiles of the two breeds. We applied 2-DE in order to identify proteins which were differentially expressed in the livers of the two breeds and relate them to different liver functions. We expected to find that only a small number of proteins would be differentially expressed, due to the relatively short phylogenetic distance between these cattle breeds. Nonetheless, thirty nine differentially-expressed proteins were characterized between Chianina and Holstein Friesian, out of a total of 560+/-57 spots that matched. Microarray analyses evidenced the differential expression of 167 genes (148 for the Holstein Friesian and 19 for the Chianina). Despite being closely related at the genetic level, the disparity of the proteomic and transcriptomic profiles of these two breeds allows us to perform pathway analysis thus to pinpoint proteins whose expression might render the latter capable of greater milk production, or proteins involved in altered thermoregulatory ability or hormone production. On the other hand, we found proteins and gene transcripts in Chianina, not expressed in Holstein, which, upon interaction pathway analysis, were mainly involved in anabolic pathways. In brief, our integrated study provides molecular evidences to support the physiological differences between Holstein and Chianina cattle breeds.

Background: SNP data of goats of three Mediterranean countries were used for population studies and reconstruction of geographical patterning. 496 individuals belonging to Italian, Albanian and Greek breeds were genotyped to assess the basic population parameters., Results: A total of 26 SNPs were used, for a total of 12,896 genotypes assayed. Statistical analysis revealed that breeds are not so similar in terms of genetic variability, as reported in studies performed using different markers. The Mantel test showed a strongly significant correlation between genetic and geographic distance. Also, PCA analysis revealed that breeds are grouped according to geographical origin, with the exception of the Greek Skopelos breed., Conclusion: Our data point out that the use of SNP markers to analyze a wider breed sample could help in understanding the recent evolutionary history of domestic goats. We found correlation between genetic diversity and geographic distance. Also PCA analysis shows that the breeds are well differentiated, with good correspondence to geographical locations, thus confirming the correlation between geographical and genetic distances. This suggests that migration history of the species played a pivotal role in the present-day structure of the breeds and a scenario in which coastal routes were easier for migrating in comparison with inland routes. A westward coastal route to Italy through Greece could have led to gene flow along the Northern Mediterranean.

Microarrays produce a measurement of gene expression based on the relative measures of dye intensities that correspond to the amount of target RNA. This technology is fast developing and its application is expanding from Homo sapiens to a wide number of species, where enough information on sequences and annotations exist. Anyway, the number of species for which a dedicated platform exists is not high. The use of heterologous array hybridization, screening for gene expression in one species using an array developed for another one, is still quite frequent, even though cross-species microarray hybridization has raised many arguments. Some methods which are high throughput and do not rely on knowledge of the DNA/RNA sequence exist, namely serial analysis of gene expression (SAGE), Massively Parallel Signature Sequencing (MPSS) and deep sequencing of full transcriptome. Although very powerful, particularly the latter, they are still quite costly and cumbersome methods. In some species where genome sequences are largely unknown, several anonymous sequences are deposited in gene banks as a result of Expressed Sequence Tags (ESTs) sequencing projects. The ESTs databases represent a valuable knowledge that can be exploited with some bioinformatic effort to build species-specific microarrays. We present here a method of high-density in situ synthesized microarrays starting from available EST sequences in, Ovis aries. Our data indicate that the method is very efficient and can be easily extended to other species of which genetic sequences are present in public databases, but neglected so far with advanced devices like microarrays. As a perspective, the approach can be applied also to species of which no sequences are available to date, thanks to high-throughput deep sequencing methods.

Background: In this study we compare outlier loci detected using a FST based method with those identified by a recently described method based on spatial analysis (SAM). We tested a panel of single nucleotide polymorphisms (SNPs) previously genotyped in individuals of goat breeds of southern areas of the Mediterranean basin (Italy, Greece and Albania). We evaluate how the SAM method performs with SNPs, which are increasingly employed due to their high number, low cost and easy of scoring., Results: The combined use of the two outlier detection approaches, never tested before using SNP polymorphisms, resulted in the identification of the same three loci involved in milk and meat quality data by using the two methods, while the FST based method identified 3 more loci as under selection sweep in the breeds examined., Conclusion: Data appear congruent by using the two methods for FST values exceeding the 99% confidence limits. The methods of FST and SAM can independently detect signatures of selection and therefore can reduce the probability of finding false positives if employed together. The outlier loci identified in this study could indicate adaptive variation in the analysed species, characterized by a large range of climatic conditions in the rearing areas and by a history of intense trade, that implies plasticity in adapting to new environments.

Milk and dairy products are an essential food and an economic resource in many countries. Milk component synthesis and secretion by the mammary gland involve expression of a large number of genes whose nutritional regulation remains poorly defined. The purpose of this study was to gain an understanding of the genomic influence on milk quality and synthesis by comparing two sheep breeds with different milking attitude (Sarda and Gentile di Puglia) using sheep-specific microarray technology. From sheep ESTs deposited at NCBI, we have generated the first annotated microarray developed for sheep with a coverage of most of the genome.

The effectiveness of single nucleotide polymorphisms (SNPs) for the assignment of cattle to their source breeds was investigated by analysing a panel of 90 SNPs assayed on 24 European breeds. Breed assignment was performed by comparing the Bayesian and frequentist methods implemented in the STRUCTURE 2.2 and GENECLASS 2 software programs. The use of SNPs for the reallocation of known individuals to their breeds of origin and the assignment of unknown individuals was tested. In the reallocation tests, the methods implemented in STRUCTURE 2.2 performed better than those in GENECLASS 2, with 96% vs. 85% correct assignments respectively. In contrast, the methods implemented in GENECLASS 2 showed a greater correct assignment rate in allocating animals treated as unknowns to a reference dataset (62% vs. 51% and 80% vs. 65% in field tests 1 and 2 respectively). These results demonstrate that SNPs are suitable for the assignment of individuals to reference breeds. The results also indicate that STRUCTURE 2.2 and GENECLASS 2 can be complementary tools to assess breed integrity and assignment. Our findings also stress the importance of a high-quality reference dataset in allocation studies.

Identification of proteins involved in milk production is important to understand the biology of lactation. Many studies have advanced the understanding of mammary function and milk secretion, but the critical molecular mechanisms implicated in milk fat secretion is still incomplete. Milk fat globules are secreted from the apical surface of the mammary cells, surrounded by a thin membrane bilayer, the milk fat globule membrane (MFGM), formed by proteins which have been suggested to be cholesterolemia-lowering factors, inhibitors of cancer cell growth, vitamin binders, bactericidal, suppressors of multiple sclerosis. Using a proteomic approach, we compared MFGM from milk samples of individuals belonging to two different cattle breeds, Chianina and Holstein, representative of selection for milk and meat traits, respectively. We were able to isolate some of the major MFGM proteins in the examined samples and to identify differences between the protein fractions of the two breeds. We detected differences in the amount of proteins linked to mammary gland development and lipid droplets formation, as well as host defence mechanisms. We have shown that proteomics is a suitable, unbiased method for the study of milk fractions proteins and a powerful tool in nutritional genomics.

The use of SNPs in combination with Bayesian statistics for the geographic traceability of cattle was evaluated using a dataset comprising 24 breeds from Italy, France, Spain, Denmark, the Netherlands, Switzerland and UK genotyped with 90 polymorphic markers. The percentage of correct assignment of the individuals to their Country of origin was 90%, with an average assignment probability of 93% and an average specificity of 92%. The higher value was observed for UK breeds (97% of correct assignment) while Swiss animals were the most difficult to allocate (77% of correct assignment). Tracing of Protected Geographic Indication (PGI) products, the approach correctly assigned 100% of Guaranteed Pure Highland Beef; 97% of "Vitellone dell'Appennino Centrale" breeds; 84% of Ternera de Navarra, and 80% of Boeuf de Chalosse. Methods to verify Products of Designated Origin (PDO) and Protected Geographic Indication (PGI) products will help to protect regional foods and promote the economic growth of marginal rural areas by encouraging the production of high quality niche market foods.

We identified 37 single-nucleotide polymorphisms (SNPs) in sheep and screened 16 individuals from 8 different sheep breeds selected throughout Europe. Population genetic measures based on the genotyping of about 30 sheep from the same 8 breeds are reported. To date, there are no sheep SNPs documented in the National Center for Biotechnology Information dbSNP database. Therefore, the markers presented here contribute significantly to those currently available.

The discovery of SNPs was performed using animals from eight European sheep breeds. Eleven SNPs were further characterized using about 1,700 sheep belonging to 57 breeds. A method for the identification of loci that were likely subject to selection was applied; three of the 11 SNPs lying outside the 95% confidence region of the conditional joint distribution of F(ST) and mean heterozygosity were identified as outliers.

Single nucleotide polymorphisms (SNPs) may be used in biodiversity studies and commercial tasks like traceability, paternity testing and selection for suitable genotypes. Twenty-seven SNPs were characterized and genotyped on 250 individuals belonging to eight Italian goat breeds. Multilocus genotype data were used to infer population structure and assign individuals to populations. To estimate the number of groups (K) to test in population structure analysis we used likelihood values and variance of the bootstrap samples, deriving optimal K from a drop in the likelihood and a rise in the variance plots against K.

Chromosome 2 (chr 2) deletions are recurrent abnormalities in acute myeloid leukemia (AML) induced by ionizing radiation in the mouse. The localization of deletion sites has proven extremely useful in providing information on the molecular mechanisms of leukemogenesis. The models available for the study of AML are mostly represented by inbred mouse strains, in which the molecular resolution of breakpoints is problematic. In this study, we have examined five leukemic cell lines exhibiting hemizygous chr 2 loss, derived from CBA, C3H, or (C57BLxCBA/H) F1 mice in which AML had been induced by a whole-body dose of radiation. By application of a somatic cell hybridization technique, we have generated interspecific cell hybrids retaining the deleted murine chr 2 homologue. This strategy permitted a very detailed genetic analysis allowing the utilization of any genetic marker on chr 2 without a requirement for polymorphism. Somatic cell hybrid clones were subjected to a high-density polymerase chain reaction-based microsatellite screening using 62-106 informative markers for each cell line. Detailed maps accurately defining chr 2 breakpoints were obtained. The identification of critical breakpoint markers allowed the construction of partial yeast artificial chromosome contigs across chr 2 breakpoints. These maps represent an essential resource for cloning of the breakpoint regions.

For the assessment of radiation risk at low doses, it is presumed that the shape of the low-dose-response curve in humans for cancer induction is linear. Epidemiological data alone are unlikely to ever have the statistical power needed to confirm this assumption. Another approach is to use oncogenic transformation in vitro as a surrogate for carcinogenesis in vivo. In mid-1990, six European laboratories initiated such an approach using C3H 10T1/2 mouse cells. Rigid standardisation procedures were established followed by collaborative measurements of transformation down to absorbed doses of 0.25 Gy of x-radiation resulting in a total of 759 transformed foci. The results clearly support a linear dose-response relationship for cell transformation in vitro with no evidence for a threshold dose or for an enhanced, supralinear response at doses approximately 200-300 mGy. For radiological protection this represents a large dose, and the limitations of this approach are apparent. Only by understanding the fundamental mechanisms involved in radiation carcinogenesis will further knowledge concerning the effects of low doses become available. These results will, however, help validate new biologically based models of radiation cancer risk thus providing increased confidence in the estimation of cancer risk at low doses.

Workplace description is an operation indispensable for the identification of professional risks and, in fulfillment of that objective, can best be accomplished by listing the constituent tasks associated with each job. This article presents a software in Visual-Basic to be run on a portable micro-computer, which facilitates this process according to these aims. The occupational physician and safety engineer can make good use of the exposure database resulting from the application of this program. Job histories are built automatically, while workers with similar exposures are grouped together and job-exposure matrices are elaborated. This information, computerized, sorted and re-assembled, can then be used effectively for studies in the field of occupational epidemiology.

An experimental study of male and female CBA/Cne mice was set up at Casaccia primarily to investigate the influence of sex on long-term survival and tumor induction after exposure to high- and low-LET radiation. Mice were whole-body-irradiated at 3 months of age with fission-neutron doses of 0.1, 0.2, 0.4, 0.8, 1.2 and 1.8 Gy at the RSV-TAPIRO reactor (mean neutron energy 0.4 MeV, in terms of kerma, y(D) = 51.5 keV/micron), or with 250 kVp X-ray doses of 1, 3, 5 and 7 Gy. Control and irradiated animals were then followed for their entire life span. As a general finding, male CBA/Cne mice appear more susceptible to tumorigenesis than females. In particular, the incidences of induced acute myeloid leukemia and malignant lymphomas are significant only in male mice. Benign and malignant solid tumors of many types are observed in mice of both sexes, the most frequent being in the lung, liver and ovary. However, evidence for a radiation response is limited to the case of Harderian gland neoplasms. In addition, a comparison of the observed frequency of all irradiated compared to unirradiated animals bearing solid tumors shows that the total tumor occurrence is not altered markedly by radiation exposure. A decrease in survival time is observed for both sexes and radiation types and correlates well with increasing dose. Moreover, both sex and radiation quality appear to influence the life shortening. A similar dose dependence of survival time is found when tumor-free animals alone are considered, suggesting a non-specific component of life-shortening.

Cell-age sensitivity to both cell killing and neoplastic transformation induced by radiation was investigated using synchronized populations of C3H10T1/2 cells. Mitotic-cell suspensions, collected using a mitotic shake-off procedure, were irradiated with 4Gy 250 kVp X-rays or 0.5 Gy fission neutrons from the RSV-TAPIRO reactor at CR-Casaccia. For study of cell killing the mitotic-cell suspensions were either irradiated immediately after collection, or plated for subsequent irradiation, which was performed every hour, covering an interval of 17 h. The response pattern observed was similar after X-rays and neutron irradiation, but the magnitude of the variation through the cell cycle was smaller in the case of neutrons (1.3- compared with 5-fold). For study of neoplastic transformation induction the irradiation was performed immediately after collection, i.e. in M phase, or at later times corresponding to mid-G1, G1/S and G2 phases. The sensitivity of the G2/M phase was examined by irradiating the cells with 4Gy X-rays while still attached to the flask bottom, and dislodging them after 25 min. SimilarLy to cell survival, the transformation frequency showed a small variation after neutron irradiation (1.4- compared with 3.1-fold) for the phases examined.

An experimental study of the biological effectiveness of multifractionated low doses of high-LET radiation was carried out using BC3F1 male mice. They were treated with whole-body irradiation with five equal daily fractions of fission neutrons to yield cumulative doses of 0.025, 0.05, 0.10, 0.17, 0.25, 0.36, 0.535 and 0.71 Gy at the RSV-TAPIRO reactor (mean neutron energy 0.4 MeV, in terms of kerma, y D = 51.5 keV/microns, dose rate 0.004 Gy/min) and were followed for their entire life span. The statistical method described by Peto et al. (IARC Monograph, Suppl. 2, 1980) to establish the existence of a carcinogenic effect in long-term animal experiments was applied to the data sets. This analysis was done for myeloid leukemia and for the presence of selected solid tumors. Myeloid leukemia was absent in the control group and was rarely found in irradiated animals. However, a positive significant trend was found in the dose ranges 0-0.17 Gy and higher. Epithelial tumors were induced at doses from 0.17 Gy on. Tumor occurrence was evaluated further as final incidences with age adjustment for the differences in mortality rates. Survival and incidence data for selected classes of tumors after 0.17, 0.36 and 0.71 Gy were compared with those from a previous experiment at corresponding doses given acutely (dose rate between 0.05 and 0.25 Gy/min). This indicated no marked overall influence of the time regimen of neutron irradiation on survival and tumor induction.

As most occupational and environmental exposures to ionizing radiation are at low dose rates or in small dose fractions, risk estimation requires that the effects of the temporal distribution of dose are taken into account. Previous in vitro studies of oncogenic transformation, as well as in vivo studies of carcinogenesis induced by high-LET radiation, yielded controversial results concerning the presence of an inverse dose-rate effect. The present study tested the influence of one scheme of dose fractionation of monoenergetic neutrons on neoplastic transformation of C3H 10T1/2 cells. Neutrons of 0.5, 1.0 and 6.0 MeV were used. Cells were exposed to doses of 0.25 and 0.5 Gy, given acutely or in five fractions at 2-h intervals. The acute and fractionated irradiations with each energy were done on the same day. No significant difference between the two irradiation modes was found for both cell inactivation and neoplastic transformation at all energies. These results are in agreement with our data for fractionated fission-spectrum neutrons from the RSV-TAPIRO reactor.