Assessment of the mutagenic, recombinogenic, and carcinogenic potential of amphotericin B in somatic cells of Drosophila melanogaster.

Amphotericin B (AmB) is an antifungal antibiotic extracted from Streptomyces nodosus. Its fungicidal activity depends primarily on its binding to the sterol group that is present in fungal membranes. In view of the toxicity of this drug, the purpose of this study was to evaluate its mutagenic, carcinogenic, and recombinogenic activity, based on the wing somatic mutation and recombination test (SMART) and the epithelial tumor detection test (wts) applied to Drosophila melanogaster. Larvae were chronically treated with different concentrations of AmB (0.01, 0.02, and 0.04 mg/mL). The results revealed that AmB is a promutagen exhibiting increase in the number of spots on individuals from high bioactivation (HB) cross with a high level of cytochrome P450. The results also indicate that the main genotoxic event induced by AmB is recombinogenicity. Homologous recombination can act as a determinant at different stages of carcinogenesis. For verification of carcinogenic potential of this compound, larvae from the wts/mwh and wts/ORR, flr3 were treated with the same three AmB concentrations used in the SMART assay. The results did not provide evidence that AmB has carcinogenic potential in wts/mwh individuals. However, individuals from wts/ORR, flr3 developed tumors at the highest concentration tested.

Protective effect of aspirin against mitomycin C-induced carcinogenicity, assessed by the test for detection of epithelial tumor clones (warts) in Drosophila melanogaster.

The present study assessed the protective effect of aspirin against carcinogenicity induced by mitomycin C (MMC) by the test for detection of warts/epithelial tumor clones in Drosophila melanogaster. Larvae were treated with different concentrations of aspirin alone (10, 20 or 40 mg/mL) or aspirin in association with MMC. MMC and ultrapure water were employed as the positive and negative control, respectively. Antioxidant activity was determined using the DPPH method. For performing cytotoxicity assay on HeLa cells, the aspirin concentrations used ranged from 200 mmol/L to 3,125 mmol/L. For assessment of apoptosis and necrosis, cells were incubated for 24 h with complete medium in the absence (control group) or presence of aspirin (12.5 mmol/L and 25 mmol/L). The results obtained in the assessment of the possible carcinogenic effects of aspirin at the three concentrations tested indicate no statistically significant increase in tumor frequency compared to the negative control. The anticarcinogenic activity assessment, where the larvae of D. melanogaster were previously induced to tumor formation by MMC and later treated with aspirin, showed a statistically significant reduction in the number of tumors compared to the positive control. Antioxidant activity across the three aspirin concentrations (10, 20 or 40 mg/mL) ranged from 20.81% to 26.5%. It was observed that aspirin reduced growth viability of HeLa cells in a concentration-dependent manner in comparison with the control. These results indicate that aspirin did not induce tumors in Drosophila and reduced MMC-induced carcinogenicity. The antioxidant activity and apoptosis induction appear to be the main mechanisms involved in reducing the frequency of tumors.

Assessment of genotoxic, mutagenic, and recombinogenic potential of water resources in the Paranaíba River basin of Brazil: A case study.

Exposure to certain pollutants induces a series of alterations in deoxyribonucleic acid (DNA) that may result in genotoxic/mutagenic effects in exposed individuals. The present study aimed to monitor genotoxic, mutagenic, and recombinogenic potential and consequently water quality in two streams in the Paranaíba River basin in the state of Minas Gerais, Brazil, using two bioindicator fish (Rhamdia quelen and Geophagus brasiliensis). The micronucleus (MN) test and somatic recombination and mutation test (SMART) were employed to assess DNA damage. The water quality index (WQI) at the reference site control (S1) due to its proximity to the river source was compared to Córrego do Óleo (S2) with respect to chemical parameter levels of biochemical oxygen demand (BOD), dissolved-oxygen rates (DO), and total solid and fecal coliform counts. These chemical parameters were above the permitted limits at Córrego do Óleo (S2). At a third site, Córrego Liso (S3), a poor WQI was detected, attributed to the influence of domestic and industrial activities where BOD, DO, total solid, fecal coliform, total phosphorus, and turbidity rates exceeded premissible limits. The MN frequencies and the numbers of MN per cell (CMN) at sites S2 and S3 were significantly higher than those at S1 in both species. It is of interest that the increased frequency of MN was similar to the positive control cyclophosphamide only at S3, suggesting that the effects of water contaminants were most severe at this site. At sites assessed (S2 and S3), there was a significant rise in somatic mutation and recombination in the wings of Drosophila melanogaster, indicating the presence of trace elements, mainly lead (Pb) and cadmium (Cd), in the effluents in the Paranaíba River basin sites.

Micronucleus as biomarker of genotoxicity in birds from Brazilian Cerrado.

Birds are considered efficient bioindicators, by their behavioral characteristics, diversified diet, and use of several vegetation layers, including in contaminated environments. The accumulation of contaminants can harm the reproductive process and survival of species, in addition to causing severe metabolic disorders. Air pollution can also affect the birds' health. Micronucleus analysis, a technique able to evaluate the organisms' sensitivity to contaminant agents, has been regarded as a practical tool for evaluating and monitoring the clastogenic and aneugenic effects caused by pollutants. The purpose of this study was to evaluate the presence of micronuclei in bird species that use forest environments and their surroundings; and to verify if the frequency and amount of micronuclei varies between species, areas and populations. Birds transiting between the Brazilian Cerrado forest and open formations were analyzed, coming from four forest fragments of Triângulo Mineiro, two close to urban areas and two more distant. Birds were captured with mist-nets for collecting blood extensions, which were used for counting micronuclei. In total, 103 individuals of 21 species were captured, and the micronucleus rate for every 5000 erythrocytes analyzed was 1.30. Only six populations had sampling sufficiency. There was no difference between the number of individuals with and without micronuclei (χ²=3.18, df=1, p=0.08). In areas closer to the urban perimeter, the micronuclei averages in birds were greater compared to the most isolated areas (H=27.534, df=3, p<0,001). In São José, the individuals of Myiothlypis flaveola presented a number of micronuclei significantly greater than the Galheiro and Água Fria (H=9.601, df=2, p=0.008). M. flaveola clearly reflected the area quality. The micronuclei analysis in birds was effective for evaluating the area quality as well as the intensity with which the birds respond to impacts caused by the surrounding matrix.

Recombinogenic activity of Pantoprazole(®) in somatic cells of Drosophila melanogaster.

Pantoprazole(®) is one of the leading proton pump inhibitors (PPIs) used in the treatment of a variety of diseases related to the upper gastrointestinal tract. However, studies have shown an increased risk of developing gastric cancer, intestinal metaplasia and hyperplasia of endocrine cells with prolonged use. In the present study, the somatic mutation and recombination test (SMART) was employed to determine the mutagenic effects of Pantoprazole on Drosophila melanogaster. Repeated treatments with Pantoprazole were performed on 72-hour larvae of the standard (ST) and high bioactivation (HB) crosses at concentrations of 2.5, 5.0, and 10.0 µM. In addition, doxorubicin (DXR) was administered at 0.4 mM, as a positive control. When administered to ST descendants, total number of spots were statistically significant at 2.5 and 5.0 µM concentrations. For HB descendants, a significant increase in the total number of spots was observed among the marked transheterozygous (MH) flies. Through analysis of balancer heterozygous (BH) descendants, recombinogenic effects were observed at all concentrations in descendants of the HB cross. In view of these experimental conditions and results, it was concluded that Pantoprazole is associated with recombinogenic effects in Drosophila melanogaster.

In situ assessment of the paraguay river water, in Brazilian Pantanal, by means of micronucleus assay with fish and chemical analysis.

The aim of this research was to assess water quality in a stretch of the Paraguay River within the Brazilian Pantanal by means of a micronucleus assay with fish, and by water and sediment physicochemical analysis. Significant increases (p >0.05) in the frequency of micronuclei (MN) and micronucleated cells (MNC) occurred in erythrocytes of Pimelodus maculatus and Leporinus friderici at two river sites in the town of Caceres relative to an upstream reference site. The results demonstrate that the Paraguay River water near Caceres has been receiving genotoxic effluents, which may be associated with the presence of chromium, sulfides, oil and grease, and/or other chemicals.

Molecular characterization of adenoviruses from children presenting with acute respiratory disease in Uberlândia, Minas Gerais, Brazil, and detection of an isolate genetically related to feline adenovirus.

Human adenoviruses (HAdV) are a major cause of acute respiratory diseases (ARD), gastroenteritis, conjunctivitis and urinary infections. Between November 2000-April 2007, a total of 468 nasopharyngeal aspirate samples were collected from children with ARD at the Clinics Hospital of Uberlândia. These samples were tested by immunofluorescence assay (IFA) and 3% (14/468) tested positive for the presence of HAdV. By performing polymerase chain reaction (PCR) to detect HAdV DNA in samples that tested negative or inconclusive for all viruses identifiable by IFA (respiratory syncytial virus, parainfluenza viruses 1, 2 and 3, influenza viruses A and B and HAdV), as well as negative for rhinoviruses by reverse transcription-PCR, additional 19 cases were detected, for a total of 33 (7.1%) HAdV-positive samples. Nucleotide sequences of 13 HAdV samples were analyzed, revealing that they belonged to species B, C and E. Further analyses showed that species C (HAdV-2) was the most prevalent among the sequenced samples. To our knowledge, this is the first report describing the presence of HAdV-4 in Brazil. We also detected an isolate that was 100% identical to a part of the feline adenovirus hexon gene sequence.

Assessment of the carcinogenic potential of high intense-sweeteners through the test for detection of epithelial tumor clones (warts) in Drosophila melanogaster.

High intensity-sweeteners (HIS) are natural or synthetic substances, sweeter than sugar, providing sweetness without calories. Sweeteners are mainly used as an aid in losing weight, preventing obesity and controlling blood sugar levels for diabetics. The objective of this study was to evaluate the carcinogenic potential of the sweeteners aspartame, sucralose, sodium saccharin and steviol glycoside, using the test for detection of epithelial tumor clones in Drosophila melanogaster. Larvae of 72 ± 4h, obtained from wts/TM3 female mated with mwh/mwh males, were treated for approximately 48h with different concentrations of aspartame (0.85, 1.7, 3.4, 6.8 or 13.6 mM ); sucralose (0.5, 1.25, 2.5, 5.0 or 10 mM); sodium saccharin (25; 50; 100; 200 or 400 mM) and steviol glycoside (2.5; 5.0; 10; 20 or 40 mM). Water (Reverse Osmosis) and doxorubicin (DXR 0.4 mM) were used as negative and positive controls, respectively. No statistically significant differences were observed (p > 0.05) in tumor frequencies in individuals treated with all concentrations of these sweeteners when compared to negative control. It was therefore concluded that, in these experimental conditions, aspartame, sucralose, sodium saccharin and steviol glycoside have no carcinogenic effect in D. melanogaster.

Modulatory effects of metformin on mutagenicity and epithelial tumor incidence in doxorubicin-treated Drosophila melanogaster.

Metformin (MET) is an anti-diabetic drug used to prevent hepatic glucose release and increase tissue insulin sensitivity. Diabetic cancer patients are on additional therapy with anticancer drugs. Doxorubicin (DXR) is a cancer chemotherapeutic agent that interferes with the topoisomerase II enzyme and generates free radicals. MET (2.5, 5, 10, 25 or 50 mM) alone was examined for mutagenicity, recombinogenicity and carcinogenicity, and combined with DXR (0.4 mM) for antimutagenicity, antirecombinogenicity and anticarcinogenicity, using the Somatic Mutation and Recombination Test and the Test for Detecting Epithelial Tumor Clones in Drosophila melanogaster. MET alone did not induce mutation or recombination. Modulating effects of MET on DXR-induced DNA damage were observed at the highest concentrations. In the evaluation of carcinogenesis, MET alone did not induce tumors. When combined with DXR, MET also reduced the DXR-induced tumors at the highest concentrations. Therefore, in the present experimental conditions, MET alone did not present mutagenic/recombinogenic/carcinogenic effects, but it was able to modulate the effect of DXR in the induction of DNA damage and of tumors in D. melanogaster. It is believed that this modulating effect is mainly related to the antioxidant, anti-inflammatory and apoptotic effects of this drug, although such effects have not been directly evaluated.

Protective effects of a mixture of antioxidant vitamins and minerals on the genotoxicity of doxorubicin in somatic cells of Drosophila melanogaster.

Antioxidant vitamins are able to deactivate highly bioactive molecules, such as free radicals, that are generated during cellular biochemical processes. Doxorubicin (DXR) is a cancer chemotherapeutic agent that generates free radicals as a byproduct. In the present study, the Drosophila melanogaster somatic wing spot test was used to evaluate the effects of a mixture of vitamins (Vitamins C, E, and beta-carotene) and minerals (copper, selenium, and zinc), commercially known as Vitergan Zinc Plus, on the genotoxicity of DXR in standard and high-bioactivation crosses of flies. 12.5, 25, and 50 mg/ml of the vitamin/mineral mixture by itself was nongenotoxic in the trans-heterozygous descendants of both crosses, while the mixture produced a significant reduction in the genotoxicity produced by 0.125 mg/ml DXR in the trans-heterozygous descendants of both crosses. The protective effect was observed when the larvae received either pre- or cotreatments of the multivitamin/mineral (MV) mixture. The results indicate that, under these experimental conditions, the MV mixture is not genotoxic; however, it protects against the genotoxic effects of the chemotherapeutic free-radical generator DXR.

Evaluation of mutagenic, recombinogenic and carcinogenic potential of (+)-usnic acid in somatic cells of Drosophila melanogaster.

The main of this study was to evaluate the mutagenic and carcinogenic potential of (+) - usnic acid (UA), using Somatic Mutation and Recombination Test (SMART) and the test for detecting epithelial tumor clones (wts) in Drosophila melanogaster. Larvae from 72 ± 4 h from Drosophila were fed with UA (5.0, 10.0 or 20.0 mM); urethane (10.0 mM) (positive control); and solvent (Milli-Q water, 1% Tween-80 and 3% ethanol) (negative control). ST cross produced increase in total mutant spots in the individuals treated with 5.0, 10.0 or 20.0 mM of UA. HB cross produced spot frequencies in the concentration of 5.0 mM that were higher than the frequency for the same concentration in the ST cross. In the highest concentrations the result was negative, which means that the difference observed can be attributed, in part, to the high levels of P450, suggesting that increasing the metabolic capacity maximized the toxic effect of these doses. In the evaluation of carcinogenesis using the wts test, the results obtained for the same concentrations of UA show a positive result for the presence of tumors when compared to the negative control. We conclude that UA has recombinogenic, mutagenic and carcinogenic effects on somatic cells in D. melanogaster.

Genotoxicity testing of Plantago major extracts in somatic cells of Drosophila melanogaster.

Plantago major is used in many parts of the world for the treatment of diseases and to promote the healing of wounds. In the present study, the somatic mutation and recombination test (SMART) in Drosophila melanogaster was used to evaluate the genotoxic activity of an aqueous extract of P. major. The following Drosophila crosses were made: standard (ST) cross, in which virgin flare females (flr3/TM3, Bds) were mated with mwh/mwh males, and high-bioactivation (HB) cross, in which virgin ORR females (ORR/ORR; flr3/TM3, Bds) were mated with mwh/mwh males. Each cross produced two types of descendents, marker-transheterozygous (MH) (mwh +/+ flr3) and balancer-heterozygous (BH) (mwh +/+ TM3, Bds) flies. Three-day-old larvae of both types of descendents were treated with undiluted and diluted (1:1 and 1:2 in water) aqueous extracts of P. major. The extracts were genotoxic in both crosses, producing similar induced frequencies in ST and HB flies. Comparison of the frequencies of wing spots in the BH and MH descendents indicated that recombination was a major response. The results indicate that, under these experimental conditions, aqueous extracts of P. major are genotoxic (recombinagenic).

Genotoxicity of lapachol evaluated by wing spot test of Drosophila melanogaster.

This study investigated the genotoxicity of Lapachol (LAP) evaluated by wing spot test of Drosophila melanogaster in the descendants from standard (ST) and high bioactivation (HB) crosses. This assay detects the loss of heterozygosity of marker genes expressed phenotypically on the fly's wings. Drosophila has extensive genetic homology to mammals, which makes it a suitable model organism for genotoxic investigations. Three-day-old larvae from ST crosses (females flr(3)/TM3, Bd(s) x males mwh/mwh), with basal levels of the cytochrome P450 and larvae of high metabolic bioactivity capacity (HB cross) (females ORR; flr(3)/TM3, Bd(s) x males mwh/mwh), were used. The results showed that LAP is a promutagen, exhibiting genotoxic activity in larvae from the HB cross. In other words, an increase in the frequency of spots is exclusive of individuals with a high level of the cytochrome P450. The results also indicate that recombinogenicity is the main genotoxic event induced by LAP.

Genotoxicity of water from the Paraguay River near Cáceres-MT, Brazil in the Drosophila wing-spot test.

The genotoxic activity of surface water samples from four sites along the Paraguay River, near Cáceres, Mato Grosso State, Brazil, was investigated using the Drosophila melanogaster Somatic Mutation and Recombination test (SMART). Effluents from sanitary sewers and agroindustrial effluents (residual effluents from slaughterhouses, leather tanneries, and dairies) flow into the Paraguay River, and directly or indirectly contaminate water from sampling sites 1-3. Site 4 was an upriver reference site that received no domestic or agroindustrial discharges. Water was collected at 4 time periods: September 2003 and August 2004 (periods of low water or drought); and April 2004 and March 2005 (periods of high water or flood). Chromium concentrations above statutory limits were detected at sites 1-3 (August 2004), and sites 1, 2 and 4 (March 2005). Sulfur compounds were also detected at sites 1-3. The SMART performed using standard (ST) cross flies detected genotoxic responses in only two samples, the August 2004 site 1 sample and the March 2005 site 2 sample. Many more samples were positive using high bioactivation (HB) cross flies: site 1 (all collection periods), site 2 (September 2003 and April 2004), and site 3 (September 2003 and August 2004). Mutant frequency comparisons between marker-heterozygous and balancer-heterozygous flies from the HB cross indicated that the positive genotoxic responses for the site 2 (April 2004) and site 3 (September 2003) samples were due mainly to mitotic recombination. Our findings indicate that the section of the Paraguay River within the urban perimeter of Cáceres is contaminated with genotoxic agents.

Genotoxicity of lapachol evaluated by wing spot test of Drosophila melanogaster

This study investigated the genotoxicity of Lapachol (LAP) evaluated by wing spot test of Drosophila melanogaster in the descendants from standard (ST) and high bioactivation (HB) crosses. This assay detects the loss of heterozygosity of marker genes expressed phenotypically on the fly's wings. Drosophila has extensive genetic homology to mammals, which makes it a suitable model organism for genotoxic investigations. Three-day-old larvae from ST crosses (females flr³/TM3, Bd s x males mwh/mwh), with basal levels of the cytochrome P450 and larvae of high metabolic bioactivity capacity (HB cross) (females ORR; flr³/TM3, Bd s x males mwh/mwh), were used. The results showed that LAP is a promutagen, exhibiting genotoxic activity in larvae from the HB cross. In other words, an increase in the frequency of spots is exclusive of individuals with a high level of the cytochrome P450. The results also indicate that recombinogenicity is the main genotoxic event induced by LAP.

Molecular characterization of adenoviruses from children presenting with acute respiratory disease in Uberlândia, Minas Gerais, Brazil, and detection of an isolate genetically related to feline adenovirus

Human adenoviruses (HAdV) are a major cause of acute respiratory diseases (ARD), gastroenteritis, conjunctivitis and urinary infections. Between November 2000-April 2007, a total of 468 nasopharyngeal aspirate samples were collected from children with ARD at the Clinics Hospital of Uberlândia. These samples were tested by immunofluorescence assay (IFA) and 3 percent (14/468) tested positive for the presence of HAdV. By performing polymerase chain reaction (PCR) to detect HAdV DNA in samples that tested negative or inconclusive for all viruses identifiable by IFA (respiratory syncytial virus, parainfluenza viruses 1, 2 and 3, influenza viruses A and B and HAdV), as well as negative for rhinoviruses by reverse transcription-PCR, additional 19 cases were detected, for a total of 33 (7.1 percent) HAdV-positive samples. Nucleotide sequences of 13 HAdV samples were analyzed, revealing that they belonged to species B, C and E. Further analyses showed that species C (HAdV-2) was the most prevalent among the sequenced samples. To our knowledge, this is the first report describing the presence of HAdV-4 in Brazil. We also detected an isolate that was 100 percent identical to a part of the feline adenovirus hexon gene sequence.

Dieta e câncer: vitaminas antioxidantes / Diet and cancer: antioxidant vitamins

A dieta é importante tanto para minimizar, quanto para aumentar o risco de desenvolvimento do câncer. Vida sedentária, meio ambiente, vírus, fumo, álcool em excesso, são fatores que contribuem e são, também, pontos estratégicos que devem ser trabalhados na prevenção do câncer. O consumo de bebidas alcoólicas, em particular nas pessoas fumantes, aumenta o risco de câncer nos tratos respiratório e digestivo. Muitos agentes que protegem contra o câncer foram identificados em laboratório. Podemos incluir as vitaminas A e seus análogos, vitamina C e vitamina E, e os compostos sem valor nutricional tais como, isotiocianatos e ditioltionas. Ditioltionas, por exemplo, são agentes com potencial de quimioprevenção, encontrados no brócolis, couve-flor e couve. Nos últimos anos, o consumo de vitaminas, minerais e ervas como suplemento alimentar tem aumentado acentuadamente. As suplementações com vitaminas e minerais são geralmente mais utilizadas do que ervas medicinais. A suplementação alimentar com vitaminas e minerais, na prevenção do câncer, ainda é uma questão polêmica e sua eficácia não está suficientemente comprovada.

Diet is important to minimize but also to increase the risk of cancer. Sedentary life, environment, viruses, smoking, alcohol in excess, are all elements that contribute and are also strategic points that must be developed in cancer prevention. The consume of liquor, especially in smokers, increase the risk of cancer in breathin and digestive tracts. Lots of agents that protect against cancer have been identified in labs. We could incluse vitamin A and its analogues, vitamin C and vitamin E, and the compounds without nutrition values, such as isothiocynates and dithiolthione. Dithiolthiones, for example, are agents with a chemioprevention potential, found in broccolis, cauliflower and cabbage. In later years, the consume of vitamins, minerals and herbs as feed supplements have increased dramatically. The feed supplements with vitamins and minerals, in the prevention of cancer, is still a polemic matter and its efficacy is not sufficiently proved.

Ausência de efeitos genotóxicos do antibiótico amicacina em células de medula óssea de ratos Wistar, in vivo / Absence of genotoxic effects of the antibiotic amikacin on Wistar rat bone marrow cells, in vivo

O sulfato de amicacina (AK) é um antibiótico aminoglicosídico semi-seintético, indicado no tratamento de infecçöes graves produzidas por cepas sensíveis de bactérias Gram-negativas, e algumas Gram-positivas. O potencial genotóxico do Ak foi avaliado por meio de análise de células de medula óssea de ratos Wistar, tratados in vivo. A droga foi injetada intraperitonealmente (i.p.), em dose única, nas concentraçöes de 1,5; 3,0 e 6,0 mg/ 100g de peso do animal. Para cada concentraçäo foram tratados nove animais. O experimento foi realizado de modo a permitir correlacionar a dose-efeito genotóxico, com diferentes tempos de tratamento (6, 12 ou 24 h). Foram analisados três animais, de cada concentraçäo, para cada tempo de tratamento. Foi observado aumento estatisticamente significativo na frequência de falhas acromáticas nas células dos animais tratados com 1,5 e 3,0 mg/100 g de peso do animal, sacrificados após 6 h de tratamento. A frequência de quebras cromossômicas e os índices mitóticos foram estatisticamente näo significativos, quando comparados com o controle negativo, para todas as concentraçöes de Ak , para todos os tempos de tratamento. Os resultados obtidos permitem-nos concluir que o Ak, nestas condiçöes experimentais, näo possui efeito genotóxico

Ausência de efeitos mutagênicos e recombinogênicos do ketoconazole em células somáticas de Drosophila Melanogaster / Absence of mutagenic and recombinogenic effects of ketoconazole in somatic cells of Drosophila Melanogaster

Ketoconazole (Kc) é um derivado imidazol, utilizado em medicina clínica como antifúngico oral de amplo espectro. Para realizar o estudo de seus possíveis efeitos mutagênicos e recombinogênicos, foi empregado o teste da mancha da asa, denominado Somatic Mutation And Recombination Test (SMART), que detecta mutaçöes e recombinaçöes somáticas em células de asas de Drosophila melanogaster. Foram realizados os seguintes cruzamentos: [1] cruzamento padräo (ST - "standard cross"), no qual fêmeas flr3 / In (3LR)TM3, ri pp sep l(3)89Aa bx34e e BdS foram cruzadas com machos mwh (GRAF et al., 1989); [2] cruzamento aprimorado de alta capacidade de bioativaçäo (IHB - "improved high bioactivation cross") no qual fêmeas ORR; flr3 / In(3LR)TM3, ri pp sep l(3)89Aa bx34e e BdS foram cruzadas com machos mwh (GRAF & VAN SCHAIK, 1992). Este último, é caracterizado por uma alta sensibilidade a promutágenos e a procarcinógenos. As larvas resultantes desses dois cruzamentos foram tratadas por aproximadamente 48 horas com diferentes concentraçöes de Kc (1,5 a 12,5 mg / 5 ml de solvente). As asas dos adultos emergentes foram analisadas quanto à ocorrência de diferentes tipos de manchas mutantes. Näo foi verificado aumento, estatisticamente significante, na frequência de manchas mutantes em nenhum dos cruzamentos realizados, o que permite concluir que o Kc, nas concentraçöes testadas, é um agente näo mutagênico e näo recombinogênico em somáticas de Drosophila melanogaster