Potential neoplastic evolution of Vero cells: in vivo and in vitro characterization.

Vero cell lines are extensively employed in viral vaccine manufacturing. Similarly to all established cells, mutations can occur during Vero cells in vitro amplification which can result in adverse features compromising their biological safety. To evaluate the potential neoplastic evolution of these cells, in vitro transformation test, gene expression analysis and karyotyping were compared among low- (127 and 139 passages) and high-passage (passage 194) cell lines, as well as transformed colonies (TCs). In vivo tumorigenicity was also tested to confirm preliminary in vitro data obtained for low passage lines and TCs. Moreover, Vero cells cultivated in foetal bovine serum-free medium and derived from TCs were analysed to investigate the influence of cultivation methods on tumorigenic evolution. Low-passage Vero developed TCs in soft agar, without showing any tumorigenic evolution when inoculated in the animal model. Karyotyping showed a hypo-diploid modal chromosome number and rearrangements with no difference among Vero cell line passages and TCs. These abnormalities were reported also in serum-free cultivated Vero. Gene expression revealed that high-passage Vero cells had several under-expressed and a few over-expressed genes compared to low-passage ones. Gene ontology revealed no significant enrichment of pathways related to oncogenic risk. These findings suggest that in vitro high passage, and not culture conditions, induces Vero transformation correlated to karyotype and gene expression alterations. These data, together with previous investigations reporting tumour induction in high-passage Vero cells, suggest the use of low-passage Vero cells or cell lines other than Vero to increase the safety of vaccine manufacturing.

Genome-wide association analysis in Italian Simmental cows for lactation curve traits using a low-density (7K) SNP panel.

High-throughput cow genotyping has opened new perspectives for genome-wide association studies (GWAS). Directly recorded phenotypes and several records per animal could be used. In this study, a GWAS on lactation curve traits of 337 Italian Simmental cows genotyped with the Illumina (San Diego, CA) low-density BeadChip (7K) was carried out. Scores of the first 2 principal components extracted from test-day records (7 for each lactation) for milk yield, fat and protein percentages, and somatic cell score were used as phenotypes. The first component described the average level of the lactation curve, whereas the second summarized its shape. Data were analyzed with a mixed linear model that included fixed effects of herd, calving month, calving year, parity, SNP genotype, and random effects of animal and permanent environment. All statistically significant markers (Bonferroni corrected) were associated with the average level component (2 for milk yield, 9 for fat percentage, 6 for protein percentages, and 1 for somatic cell score). No markers were found to be associated with the lactation curve shape. Gene discovery was performed using windows of variable size, according to the linkage disequilibrium level of the specific genomic region. Several suggestive candidate genes were identified, some of which already reported to be associated with dairy traits, such as DGAT1. Others were involved in lipid metabolism, in protein synthesis, in the immune response, in cellular processes, and in early development. The large number of genes flagged in the present study suggests interesting perspectives for the use of low-density genotyped females for GWAS, also for novel phenotypes that are not currently considered as breeding goals.

MUGBAS: a species free gene-based programme suite for post-GWAS analysis.

UNLABELLED: Genome Wide Association Studies between molecular markers and phenotypes are now routinely run in model and non-model species. However, tools to estimate the probability of association of functional units (e.g. genes) containing multiple markers are not developed for species other than humans. Here we introduce MUGBAS (MUlti species Gene-Based Association Suite), software that estimates the P-value of a gene using information on annotation, single marker GWA results and genotype. The software is species and annotation independent, fast, highly parallelized and ready for high-density marker studies. AVAILABILITY AND IMPLEMENTATION: https://bitbucket.org/capemaster/mugbas

Detecting population structure and recent demographic history in endangered livestock breeds: the case of the Italian autochthonous donkeys.

Since its domestication, about 5000 years ago, the donkey (Equus asinus) has been extensively used as a work or draft animal in agricultural activities and for the transportation of people and goods. In the last century, technology improvement and growing mechanization strongly affected agriculture and the management and use of this livestock species in the industrialized countries. Nowadays, the use of donkeys for work or transport has almost disappeared, together with the need for mules or hinny breeding. During the last five decades, Italian autochthonous donkey populations suffered from a severe reduction in population size, which led to the extinction of several breeds. At present, eight breeds remain, all classified by FAO as critically endangered or endangered: Asinara, Pantesco, Grigio Siciliano, Romagnolo, Amiatino, Sardo Grigio, Martina Franca, and Ragusano. To evaluate the extant genetic variability of Italian donkeys, we typed 16 microsatellite loci in 258 individuals from these breeds. The results highlighted moderate levels of inbreeding ( F (IS) = 0.127) and a significant partition of genetic variation into breeds, as suggested by fixation index ( F (ST) = 0.109) and analysis of molecular variance (10.86% of total variation assigned to the between-breeds level) analyses. This was confirmed by a Bayesian clustering procedure that also highlighted a further partitioning at lower hierarchical levels corresponding to the farms of origin. This evidence suggests that an effective management strategy for Italian donkey populations should focus on breeds as conservation units. However, this requires a synergic management strategy at the farm level to maintain diversity and avoid inbreeding.

[The comparative isoenzyme characteristics of Old World Leishmania strains from different geographical regions (Africa, the Near East, Central Asia)]. / Sravnitel'naia izofermentnaia kharakteristika shtammov leishmanii Starogo Sveta iz razlichnykh geograficheskikh raionov (Afrika, Blizhnii Vostok, Sredniaia Aziia).

Thirty-four strains of seven Old World Leishmania species were analyzed for 12 enzymes. The findings indicate isoenzymatic profile geographic changeability of some Leishmania species: Tested L. major, L. tropica, and L. infantum strains from Africa, Middle East, and Central Asia were found to be represented by individual zymodemes differing between themselves by 3-5 enzymes. Many identical zymodemes could induce in some cases skin, in other visceral symptoms in the carriers. Great isoenzymatic affinity between L. tropica and L. donovani sensu lato strains was detected. The nature of intraspecies Leishmania zymodeme variability being virtually not studied at present, the authors emphasize the usefulness of clone studies.

[The serological examination of the population for leishmaniasis and the detection of Leishmania in rodents in the Republic of Guinea]. / Serologicheskoe obsledovanie naseleniia na leishmanioz i obnaruzhenie leishmanii u gryzunov v Gvineiskoi Respublike.

A serological study on leishmaniasis in human population of Guinea revealed the percentage of seropositive persons varying in different parts of the country from 0.84 to 4.76 (according to C-ELISA) and from 1.0 to 5.1 (according to IFAT). The majority of sera positively reacting in C-ELISA with the antigen of Leishmania major in dilutions of 1:800 and higher were received from Kundara District (northwestern part of the country), and with the antigen of L. donovani sensu lato from Sigiri and Kankan districts (northeastern part of the country). All the districts were situated in the savanna zone. The amastigotes of Leishmania were found in the liver and spleen of Tatera gambiana caught near Kindia. These data suggest the circulation of Leishmania over the territory of Guinea.