Heterokaryosis is the initial step of the parasexual cycle, a process that provides genetic variability in filamentous fungi through the production of heterozygous diploid nuclei. To characterize the parasexual cycle in Colletotrichum lindemuthianum, we evaluated the presence of heterokaryosis, vegetative compatibility reactions, and diploid formation among isolates of Race 65 collected from different Brazilian states. Vegetative compatibility groups were identified among the isolates according to their ability to form heterokaryons. Two heterozygous diploids were selected from compatible heterokaryons, which were characterized by the segregation of the parental auxotrophic markers and by RAPD profiles.

Metformin is a hypoglycemiant drug prescribed for the treatment and control of the type 2 diabetes mellitus. Recently, the potential efficacy of this antidiabetic drug as an anticancer agent has been demonstrated in various mammalian cancer cells. This report evaluates the mutagenic as well as the recombinogenic potentials of the metformin drug in therapeutically relevant plasma concentrations (12.5 µM, 25.0 µM or 50.0 µM). Since the loss of heterozygosity is a process associated with carcinogenesis, the recombinogenic potential of such a drug was evaluated by the homozygotization assay using a heterozygous diploid strain of Aspergillus nidulans. The homozigotization indices (HI) for the genetic markers from the metformin-treated diploids were not statistically different from the negative control (non-treated diploids). For the first time, this indicated a lack of recombinogenic activity of the antidiabetic drug. The mutagenic potential of the metformin drug was evaluated by the chromosome aberrations and the micronuclei tests in human lymphocytes cultures. The metformin drug did not show any significant increase either in the numerical or in the structural chromosome aberrations and did not affect significantly the mitotic index when compared to the negative control. In the in vitro micronucleus test, the drug did not increase the number of micronuclei or nuclear buds when compared with the negative control. The data in this study suggest that the metformin drug is not a secondary cancer inducer, since it has neither showed recombinogenic nor mutagenic activities when used in pharmacological concentrations.

The genetic variation among nine soybean-originating isolates of Colletotrichum truncatum from different Brazilian states was studied. Nitrate non-utilizing (nit) mutants were obtained with potassium chlorate and used to characterize vegetative compatibility reactions, heterokaryosis and RAPD profile. Based on pairings of nit mutants from the different isolates, five vegetative complementation groups (VCG) were identified, and barriers to the formation of heterokaryons were observed among isolates derived from the same geographic area. No complementation was observed among any of the nit mutants recovered from the isolate A, which was designed heterokaryon-self-incompatible. Based on RAPD analysis, a polymorphism was detected among the wild isolate C and their nit1 and NitM mutants. RAPD amplification, with five different primers, also showed polymorphic profiles among Brazilian C. truncatum isolates. Dendrogram analysis resulted in a similarity degree ranging between 0.331 and 0.882 among isolates and identified three RAPD groups. Despite the lack of a correlation between the RAPD analysis and the vegetative compatibility grouping, results demonstrated the potential of VCG analysis to differentiate C. truncatum isolates genotypically similar when compared by RAPD.

Aspergillus flavus, a haploid organism found worldwide in a variety of crops, including maize, cottonseed, almond, pistachio, and peanut, causes substantial and recurrent worldwide economic liabilities. This filamentous fungus produces aflatoxins (AFLs) B1 and B2, which are among the most carcinogenic compounds from nature, acutely hepatotoxic and immunosuppressive. Recent efforts to reduce AFL contamination in crops have focused on the use of nonaflatoxigenic A. flavus strains as biological control agents. Such agents are applied to soil to competitively exclude native AFL strains from crops and thereby reduce AFL contamination. Because the possibility of genetic recombination in A. flavus could influence the stability of biocontrol strains with the production of novel AFL phenotypes, this article assesses the diversity of vegetative compatibility reactions in isolates of A. flavus to identify heterokaryon self-incompatible (HSI) strains among nonaflatoxigenic isolates, which would be used as biological controls of AFL contamination in crops. Nitrate nonutilizing (nit) mutants were recovered from 25 A. flavus isolates, and based on vegetative complementation between nit mutants and on the microscopic examination of the number of hyphal fusions, five nonaflatoxigenic (6, 7, 9 to 11) and two nontoxigenic (8 and 12) isolates of A. flavus were phenotypically characterized as HSI. Because the number of hyphal fusions is reduced in HSI strains, impairing both heterokaryon formation and the genetic exchanges with aflatoxigenic strains, the HSI isolates characterized here, especially isolates 8 and 12, are potential agents for reducing AFL contamination in crops.

This report evaluates the potential of the antidepressant drug citalopram to induce homozygotization of genes previously present in a heterozygous condition, by homologous recombination. In order to address this question, a heterozygous diploid strain of the filamentous fungus Aspergillus nidulans and the homozygotization assay were utilized. Non-cytotoxic concentrations of citalopram (50, 75 and 100 μmol/L) showed a strong recombinogenic effect in A. nidulans, inducing homozygosis of the diploid strain's nutritional markers. The genetic markers exhibited homozygotization index (HI) rates higher than 2.0 and significantly different from HI control ones. Since citalopram has been previously characterized as a DNA synthesis inhibitor, the recombinogenic potential of this antidepressant in A. nidulans may be associated with the recombinational repair of citalopram-induced DNA strand breaks.

Mitotic recombination is a process involved in carcinogenesis which can lead to genetic loss through the loss of heterozygosity. The recombinogenic potentials of two anticancer drugs topoisomerase I inhibitors, camptothecin (CPT) and irinotecan (CPT-11), were evaluated in the present study. The homozygotization assay, which assess the induction of mitotic recombination and gene homozygosis, as well as the heterozygous A757//UT448 diploid strain of Aspergillus nidulans were employed. The three non-cytotoxic concentrations of CPT (3.5 ng mL -1 , 10.5 ng mL -1 and 17.4 ng mL -1 ) were found to induce both mitotic recombination and gene homozygosis. CPT treatment produced three diploids homozygous, for nutritional and conidia color genes, and Homozygotization Indices (HI) significantly different from negative control. On the other hand, only the highest CPT-11 concentration tested (18 µg mL -1 ), corresponding to the maximal single chemotherapeutic dose, produced HI values higher than 2.0 and significantly different from negative control HI values. The recombinogenic effects of both topoisomerase I blockers were associated with the recombinational repair of DNA strand breaks induced by CPT and CPT-11. The anticancer drugs CPT and CPT-11 may be characterized as secondary malignancies promoters in cancer patients after chemotherapy treatment.

Heterokaryosis is an important mechanism which provides genetic variability increase in filamentous fungi. In order to assess the diversity of vegetative compatibility reactions existing among Colletotrichum acutatum isolates derived from different hosts, complementary nit mutants of each isolate were obtained and paired in all possible combinations. Vegetative compatibility groups (VCG) were identified among the isolates according to their ability to form viable heterokaryons. Seven VCGs were identified among the isolates, one of which contained isolates from different hosts. VCGs 2 and 6 contained two and three members, respectively; VCG-3 contained four members, and four VCGs (1, 4, 5, and 7) contained a single one. This study shows, for the first time, the isolation and the parasexual segregation of a heterozygous diploid sector derived from the heterokaryon formed with nit mutants from VCG-6. Diploid, named DE-3, showed nit+ phenotype and growth rate similar to the parental wild isolate. When inoculated in the presence of the haploidizing agent benomyl, the diploid strain produced parasexual haploid segregants exhibiting the nit phenotypes of the crossed mutants. Since viable heterokaryons and diploid may be formed among vegetative compatible isolates of C. acutatum, this study suggests that the parasexual cycle may be an alternative source of genetic variability in C. acutatum isolates.

ABSTRACT. Imidocarb dipropionate (IMD) is a chemotherapeutic agent prescribed for the treatment and control of babesiosis; it is known to be a nucleic acid synthesis inhibitor. Although it is an effective babesicide, there are reports of persistent IMD residues retained at high levels in edible tissues of cattle, swine and sheep, raising concerns about potential effects on humans. Since the carcinogenic potential of a chemical compound can be assessed through its effect on the homologous recombination, we investigated whether IMD is recombinogenic in Aspergillus nidulans diploid cells and whether it is capable of inducing homozygosis in genes that were previously heterozygous. This analysis was done with a homozygotization assay applied to a heterozygous diploid strain of A. nidulans. IMD used at non-toxic concentrations (2.5 to 10.0 μM) was recombinogenic, demonstrated by homozygotization indices higher than 2.0 for diploid markers. A diploid homozygous for genetic markers from chromosomes I and II was also produced. Since DNA replication blockers that induce DNA strand breaks have been classified as potent inducers of homologous recombination, Genotoxicity of imidocarb in Aspergillus nidulansthe recombinogenic potential of IMD may be due to induction of recombinational repair.