A novel LTBP2 gene variant in a Turkish family with juvenile-onset open-angle glaucoma.

BACKGROUND: Juvenile-onset open-angle glaucoma (JOAG) is a rare form of primary open-angle glaucoma (POAG) with an early age of onset before 40 years. Latent transforming growth factor-beta binding protein 2 (LTBP-2) is an extracellular matrix protein with a multi-domain structure and homology to fibrillins. LTBP2 gene variants have been associated with JOAG in a small number of patients. Herein, we report a novel missense variant in the LTBP2 gene in a Turkish family with JOAG. MATERIALS AND METHODS: Blood samples were obtained from three siblings (a 20-year-old woman with JOAG, 26-year-old man with JOAG, and 15-year-old girl with posterior embryotoxon) for genetic analysis. Their father had moderate-severe POAG and the 24-year-old brother had JOAG. The mother and 32-year-old sister were healthy. Although the parents reported no consanguinity, they come from the same village. RESULTS: Clinical exome sequencing analysis of the two siblings with JOAG revealed a novel c.607C>T p.(R203C) (rs777450651) homozygous LTBP2 variant, while the variant was heterozygous in their 15-year-old sister. There were no mutations in the MYOC, CYP1B1, or FBN1 genes. CONCLUSION: We documented a novel missense mutation in the LTBP2 gene leading to a severe form of JOAG with refractory IOP and progressive optic nerve damage, which seems to show autosomal recessive inheritance.

KRAS Mutation Reduces Thymoquinone Anticancer Effects on Viability of Cells and Apoptosis.

BACKGROUND: Cancer is a life-threatening condition with an economic burden on societies. Phytotherapy is rapidly taking place in cancer research to increase the success of treatment and quality of life. Thymoquinone (TQ) is the main active phenolic compound obtained from the essential oil of the Nigella sativa (black cumin) plant seed. For a long time, black cumin has been used traditionally for the remedy of different diseases because of its various biological effects. It has been shown that most of these effects of black cumin seeds are due to TQ. TQ became a popular research topic for phytotherapy studies for its potential therapeutic applications, and more research is going on to fully understand its mechanisms of action, safety, and efficacy in humans. KRAS is a gene that regulates cell division and growth. Monoallelic variants in KRAS result in uncontrollable cell division, leading to cancer development. Studies have shown that cancer cells with KRAS mutations are often resistant to certain types of chemotherapy and targeted therapies. OBJECTIVE: This study aimed to compare the effect of TQ on cancer cells with and without KRAS mutation to better understand the reason why TQ may have different anticancer effects in the different types of cancer cells. METHODS: TQ was investigated for its cytotoxic and apoptotic effects in laryngeal cancer cells (HEp-2) without KRAS mutation and compared to mutant KRAS-transfected larynx cancer cells and KRAS mutation-carrying lung cancer cells (A549). RESULTS: We showed that TQ has more cytotoxic and apoptotic effects on laryngeal cancer cells without KRAS mutation than in cells with mutation. CONCLUSION: KRAS mutations decrease the effect of TQ on cell viability and apoptosis, and further studies are needed to fully understand the relationship between KRAS mutations and thymoquinone effectiveness in cancer treatment.

Contribution of genotypes in Prothrombin and Factor V Leiden to COVID-19 and disease severity in patients at high risk for hereditary thrombophilia.

Thrombotic and microangiopathic effects have been reported in COVID-19 patients. This study examined the contribution of the hereditary thrombophilia factors Prothrombin (FII) and Factor V Leiden (FVL) genotypes to the severity of COVID-19 disease and the development of thrombosis. This study investigated FII and FVL alleles in a cohort of 9508 patients (2606 male and 6902 female) with thrombophilia. It was observed that 930 of these patients had been infected by SARS-CoV-2 causing COVID-19. The demographic characteristics of the patients and their COVID-19 medical history were recorded. Detailed clinical manifestations were analyzed in a group of cases (n = 4092). This subgroup was age and gender-matched. FII and FVL frequency data of healthy populations without thrombophilia risk were obtained from Bursa Uludag University Medical Genetic Department's Exome Databank. The ratio of males (31.08%; 27.01%) and the mean age (36.85 ± 15.20; 33.89 ± 14.14) were higher among COVID-19 patients compared to non-COVID-19 patients. The prevalence of FVL and computerized tomography (CT) positivity in COVID-19 patients was statistically significant in the thrombotic subgroup (p < 0.05). FVL prevalence, CT positivity rate, history of thrombosis, and pulmonary thromboembolism complication were found to be higher in deceased COVID-19 patients (p < 0.05). Disease severity was mainly affected by FVL and not related to genotypes at the Prothrombin mutations. Overall, disease severity and development of thrombosis in COVID-19 are mainly affected by the variation within the FVL gene. Possible FVL mutation should be investigated in COVID-19 patients and appropriate treatment should be started earlier in FVL-positive patients.

Clinical and molecular evaluation of MEFV gene variants in the Turkish population: a study by the National Genetics Consortium.

Familial Mediterranean fever (FMF) is a monogenic autoinflammatory disorder with recurrent fever, abdominal pain, serositis, articular manifestations, erysipelas-like erythema, and renal complications as its main features. Caused by the mutations in the MEditerranean FeVer (MEFV) gene, it mainly affects people of Mediterranean descent with a higher incidence in the Turkish, Jewish, Arabic, and Armenian populations. As our understanding of FMF improves, it becomes clearer that we are facing with a more complex picture of FMF with respect to its pathogenesis, penetrance, variant type (gain-of-function vs. loss-of-function), and inheritance. In this study, MEFV gene analysis results and clinical findings of 27,504 patients from 35 universities and institutions in Turkey and Northern Cyprus are combined in an effort to provide a better insight into the genotype-phenotype correlation and how a specific variant contributes to certain clinical findings in FMF patients. Our results may help better understand this complex disease and how the genotype may sometimes contribute to phenotype. Unlike many studies in the literature, our study investigated a broader symptomatic spectrum and the relationship between the genotype and phenotype data. In this sense, we aimed to guide all clinicians and academicians who work in this field to better establish a comprehensive data set for the patients. One of the biggest messages of our study is that lack of uniformity in some clinical and demographic data of participants may become an obstacle in approaching FMF patients and understanding this complex disease.

Determination of cytokine profile and associated genes of the signaling pathway in HNSCC.

Head and neck squamose cell carcinoma (HNSCC) is an aggressive group of tumors that are generally heterogeneous. Despite treatment advances, disease-free survival has not significantly improved. Therefore, it is of great importance to understand the molecular etiology of HNSCC and genetic alterations in the signal pathways in order to develop new therapeutic approaches. In this study, firstly we used a cytokine array to analyze the secretomes of HNSCC patients and healthy controls. In the next step, the results from the cytokine sequence were validated by qRT-PCR and western blot, including genes in the associated signaling pathway. In array analysis, the levels of EGF, IGF-1, IGFBP-1, and PDGFBB were significantly higher in patients than in the controls. The results of qRT-PCR analyses showed that expression levels of PDGFRB gene were significantly up-regulated (p = 0.006) and PTEN (p > 0.001) were significantly down-regulated in tumors compared with normal tissues. When groups (early vs. advanced) were compared, higher expression of IGFBP-1 was observed in the larynx (p = 0.045) and larynx + oral cavity tumors (p = 0.010) in an advanced stage. In western blot analysis, pEGFR, pIGF-IR, pIR-ß, pPDGFRB, and pAKT levels were upregulated, and pPTEN was downregulated in tumors. Based on our observations, determining the interactions of EGFR, PDGFRB, IGF-1R and PTEN or the activation of each might represent a promising new and innovative treatment approach in HNSCC patients. It seems clear that, in most cancers, effective targeted therapy may be involved the blockade of each one or multiple targets.

NF-Ä¸ß upregulates ADAMTS5 expression by direct binding after TNF-α treatment in OUMS-27 chondrosarcoma cell line.

Inflammation caused-aggrecan degradation is a critical event in the pathogenesis of osteoarthritis (OA). The aggrecanases like a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5) are assumed to be key players in the aggrecan destruction. To develop the comprehensive therapy method for OA, it is essential to elucidate the activation mechanism of ADAMTS5 gene after stimulation of inflammatory cytokines like tumor necrosis factor-α (TNF-α). The cell lines of human chondrosarcoma (OUMS-27) and embryonic kidney (HEK293T) were incubated with tumor necrosis factor-α (TNF-α) for certain time periods, and the expression level of ADAMTS5 was measured in both mRNA and protein levels. Tissue-specific ADAMTS5 activation was founded to be induced after TNF-α treatment. Then, the constructs for the promoter region of ADAMTS5 were prepared and luciferase assay was conducted to understand the involvement mechanism of nuclear factor-kappa beta (NF-ĸß) in ADAMTS5 activation. It was demonstrated that NF-Ä¸ß induces the ADAMTS5 expression level by directly binding the promoter region of ADAMTS5. Although the TNF-α blocker is used for OA treatment, the development of a more comprehensive treatment strategy is an urgent need. Our experimental data contributes in terms of selecting NF-Ä¸ß as a target molecule. Up to date, NF-Ä¸ß has been proven to involve in the ADAMTS5 up-regulation after several pro-inflammatory cytokines stimulation. In conclusion, our findings make important contributions to the knowledge about the roles of NF-Ä¸ß in ADAMTS5 activation under inflammatory conditions. So, NF-Ä¸ß could be considered to be a potential target for OA treatment.

Investigation of Human Papillomavirus (HPV) and Epstein-Barr Virus (EBV) in antrochoanal polyps.

OBJECTIVES/HYPOTHESES: This study aimed to investigate the presence of HPV (HPV types 11 and 16) and EBV in antrochoanal polyps and to contribute to the current literature in this regard. STUDY DESIGN: A case-control study. METHODS: A total of 100 patients (including 43 patients undergoing surgery for antrochoanal polyp, 27 patients undergoing surgery for nasal polyp, and 30 patients undergoing surgery for hypertrophic inferior turbinate) were included in this study. DNA was isolated from formalin-fixed, paraffin-embedded samples with the aid of the Bioneer's AccuPrep Genomic DNA Extraction Kit. In the obtained genomic DNAs, while the detection of HPV DNA was performed using the nested-PCR method, the detection of HPV types 11/16 and EBV DNA was performed using the RT-PCR method. RESULTS: The mean age of the patients with antrochoanal polyp was 26.7 ±â€¯15.4 years (range 7-70). There were 20 (46.5%) women and 23 (53.5%) men in the antrochoanal polyp group. HPV DNA was positively detected using the nested-PCR method in 14 (32.6%) of the patients with antrochoanal polyp and in 3 (11.1%) of the patients with nasal polyp. HPV DNA was not detected in the hypertrophic inferior turbinate group (control group). There was a statistically significant difference between all groups in terms of HPV DNA positivity. In the antrochoanal polyp group, 2 patients had HPV 11 positivity and 12 patients had HPV 16 positivity. In the nasal polyp group, 1 patient had HPV 11 positivity and 2 patients had HPV 16 positivity. EBV DNA was positively detected in 16 (37.2%) of the patients with antrochoanal polyp, in 11 (40.7%) of the patients with nasal polyp and in 8 (26.7%) of the patients with hypertrophic inferior turbinate, respectively. There was no statistically significant difference between the groups in terms of EBV DNA positivity. CONCLUSIONS: This study demonstrates that there is a need for further studies investigating the presence of viruses in antrochoanal polyps.

Glutathione S-transferase M1 and T1 genotypes and myocardial infarction.

Myocardial infarction (MI), which is the most important manifestation of coronary artery disease, is the leading cause of morbidity and mortality in the world. Glutathione S transferases (GSTs) are enzymes responsible for the metabolism of numerous xenobiotics and are known to be polymorphic in humans. We investigated the association between the GSTM1 and GSTT1 gene polymorphisms and MI. The study consists of 296 healthy controls and 324 consecutive patients who had undergone coronary angiography for suspicion of coronary artery disease and with a past history of myocardial infarction. DNA was extracted from whole blood of patient and control. GSTM1 and GSTT1 gene polymorphisms were examined using multiplex PCR. We found that the null GSTM1 was associated with protective effect on MI, although this increase was not significant for GSTM1 (p < 0.054). However, GSTT1 genotype was associated with an increase in the risk of developing MI. In addition to after adjusting other all coronary risk factors, the interactive effect of GSTT1 null genotype remained statistically significant (p < 0.001) for MI disease but GSTM1 null genotype was not statistically significant. Patients, who smoke having the null genotypes of GSTM1, were at a higher risk for developing MI (p < 0.001, OR = 0.41, 95 % CI = 0.240-0.207). There was an effect of interaction of GSTM1 null genotype and smoking on MI development between patient and control groups (p < 0.001). Our results showed that individuals with the null genotypes for GSTM1 had protective effect, while GSTT1 was at a higher risk for MI disease. In addition, there was additional effects of smoking when smoking and non-smoking groups were compared.

HLA-B27 subtypes in Turkish patients with ankylosing spondylitis and healthy controls.

The aim of this study was to determine human leukocyte antigen (HLA)-B27 subtypes frequency in ankylosing spondylitis (AS) and related spondyloartropathy (SpA) patients. Therefore, we investigated the differences in HLA-B27 subtypes between HLA-B27-positive patients and controls. Sixty six patients were included in this study (51 AS and 15 SpA). Thirty-five individuals were diagnosed with leukemia or chronic renal failure, and their donors without any rheumatological problem (no SpA history) were selected as the control group. HLA-B27 subtyping was performed by PCR-SSP (polymerase chain reaction with sequence-specific primer) method in serologically HLA-B27-positive 46 AS patients, 9 SpA patients and control group. When the frequency of HLA-B27 was 4.5% in Turkish population, this frequency was 90.2% in AS patients. Four different HLA-B27 subtypes found in AS patients were B 2705 (65.2%), B 2702 (26.1%), B 2704 (6.5%) and B 2707 (2.2%). In SpA patients, B 2705 and B 2702 found in equal frequency. Five B27 alleles were identified in our control group: B 2705 (54.3%), B 2702 (31.4) %, B 2703 (2.9%), B 2704 (2.9%) and B 2702/B 2705 (8.5%). Both in the patient group and in the control group, we also observed B 2705 as most frequent allele, and B 2702 was second common allele. Our results show that the frequency of HLA-B27 subtypes is not significantly different between patients and controls (P > 0.10).

Association of the polymorphisms of vitamin D receptor and aggrecan genes with degenerative disc disease.

The aim of this study was to investigate the association between the polymorphisms of the vitamin D receptor (VDR) and aggrecan genes and degenerative disc disease in young Turkish patients. Aggrecan and VDR proteins are the main components of bone and cartilage. In our study, the polymorphisms of the VDR and aggrecan genes were investigated in a total of 300 individuals regarding disc degeneration and herniation. An association was found in the patients having VDR gene TT, Tt, FF, and Ff genotypes with the protrusion type of disc herniation, whereas the patients having tt and ff genotypes were associated with extrusion/sequestration types of the disease. Also, an association was observed between TT and FF genotypes of the VDR gene and mild forms of disc degeneration; and tt, ff, and Ff genotypes and severe forms of the disease. There was also an association between shorter, normal, and longer alleles of the aggrecan gene and a protrusion type of disc herniation. An association was found between short alleles and multilevel and severe disc degeneration, as well as normal and long alleles and mild disc degeneration. This study revealed that the polymorphisms of the VDR and aggrecan genes are associated with disc degeneration and herniation.

Platelet glycoprotein Ibalpha gene polymorphism and massive or submassive pulmonary embolism.

OBJECTIVE: The -5T/C polymorphism in the Kozak sequence of glycoprotein Ibalpha, a component of the platelet glycoprotein Ib-IX-V receptor complex, is associated with an increase in this receptor density on the surface of the platelet. This study was designed to investigate the effect of platelet glycoprotein Ibalpha Kozak polymorphism on the clinical presentation of the patients with acute pulmonary embolism. METHODS: Forty-two patients with pulmonary embolism were genotyped for Kozak polymorphism of the glycoprotein Ibalpha by polymerase chain reaction/restriction fragment length polymorphism. RESULTS: Carriers of the -5T/C polymorphism of glycoprotein Ibalpha were significantly over-represented in the patient group with clinically massive or submassive pulmonary embolism (odds ratio 5.5, 95% confidence interval 1.4 to 22.2, P = 0.023). Also the association between this polymorphism and massive or submassive pulmonary embolism still existed even after being adjusted for conventional risk factors. CONCLUSION: The -5T/C polymorphism in the Kozak sequence of glycoprotein Ibalpha may present as a risk factor for clinical manifestation of pulmonary embolism in which clot burden plays an important role.

Evaluation of HLA-A, -B, -Cw, and -DRB1 alleles frequency in Turkish patients with nasal polyposis.

OBJECTIVE: To evaluate whether there is a relationship between HLA-A, -B, -Cw, and -DRB1 alleles and developing nasal polyposis (NP). STUDY DESIGN: Data from 66 patients with NP were compared with data from 100 healthy randomly selected controls. Asthma, ASA (acetylsalicylic acid) triad, polyp score, and previous sinonasal surgery were also recorded. SUBJECTS AND METHODS: Genotyping of the HLA-A, -B, -Cw, and -DRB1 alleles were performed with polymerase chain reaction (PCR) with the sequence-specific primer (SSP) method. Data were analyzed by using a Pearson chi(2) test. RESULTS: The HLA-B*07 and -Cw*12 alleles were found to be significantly higher in the NP patients compared with the control group, whereas the HLA-B*57 and HLA-Cw*04 alleles were significantly lower (P < 0.05). The HLA-A*24, HLA-Cw*12, and HLA-DRB1*04 alleles were determined to be significantly higher in the NP patients with asthma and ASA triad (P < 0.05). CONCLUSIONS: Our results show that some of the HLA alleles seem to be associated with the genetic susceptibility to develop NP in the Turkish population.

46,XX maleness and 46,XX 21-hydroxylase deficiency in dizygotic twins: association or coincidence?

46,XX maleness is a rare abnormality of gonadal differentiation. We present dizygotic twins, one having ambiguous genitalia due to 2-hydroxylase deficiency, and the other having normal male genitalia with 46,XX maleness. One of the twins was referred to the endocrinology unit at 2 days old because of ambiguous genitalia. The other twin with bilateral undescended testes located in the inguinal canal was diagnosed with 46,XX maleness. The karyotype was 46,XX. Molecular analysis revealed the presence of SRY in the latter twin without Müllerian structures. Association of congenital adrenal hyperplasia (46,XX 21-hydroxylase deficiency) and 46,XX maleness in twins has not been previously reported.

Relation of glutathione S-transferase genotypes (GSTM1 and GSTT1) to laryngeal squamous cell carcinoma risk.

The purpose of the present study was to investigate GSTM1 and GSTT1 genotypes by using multiplex polymerase chain reaction (PCR) in patients with laryngeal squamous cell carcinoma (LSCC). The genotypes of 110 patients with LSCC and of 197 healthy subjects as the control group were determined by PCR analysis for GSTM1 and GSTT1 genes. Results showed that frequencies of GSTM1-null, GSTT1-null, and both GSTs-null genotypes were 51.8, 30, and 16.4%, respectively, in the patients with LSCC and 37.6, 15.7, and 5.6% in the control group. There was a significant difference between the genotype distributions of all GSTs in patients and in control groups (P < 0.05). The results support the hypothesis that null genotypes of GSTM1 and GSTT1 can reduce detoxification capacity of GSTs as members of the xenobiotic enzyme system. GSTM1-null, GSTT1-null, and both GSTs-null genotypes were more common in the patients with LSCC than in the control group. Patients with both GSTs-null genotypes had the highest risk for supraglottic LSCC in the early period, even if they were light-to-medium smokers. Investigation and determination of the genetic basis of LSCC may contribute to detection of risk groups and to prevent LSCC in the population.

Lack of association between the glutathione-s-transferase genes (GSTT1 and GSTM1) and nasal polyposis.

OBJECTIVES: To evaluate the glutation-S-transferase (GST) polymorphisms (GSTM1 and GSTT1) in nasal polyposis (NP). METHODS: The study population consisted of 102 unrelated healthy individuals and 98 patients with NP (67 without asthma, 31 with asthma). Genotyping of the polymorphism in the GSTM1 and GSTT1 genes was performed using the multiplex polymerase chain reaction (PCR)-based method. RESULTS: GSTM1 and GSTT1 null-genotypes were found in 46.1% and 23.5% of the controls, and in 43.9% and 33.7% of the NP patients, respectively. These differences were not significant (for GSTM1 null odds ratio (OR) = 0.92; 95% confidence interval (CI) = 0.52-1.6 and for GSTT1, OR = 1.65; 95% CI = 0.89-3.07). Although no significant difference for combined GSTM1 and GSTT1 null genotypes between control (8.8%) and NP patients (17.3%) was found, there was a 2.16-fold increased proportion in the NP with the combined GSTM1-null and GSTT1-null genotype (OR = 2.16; 95% CI = 0.91-5.13). CONCLUSION: These results suggest that there is lack of association between GSTM1 and GSTT1 polymorphisms and NP. The GSTM1 or GSTT1 polymorphisms had also no relevant developing effect on NP patients without or with asthma.

Analysis of chromosomes 9 and 11 aneuploidy frequency in pleural effusion of patients with and without malignancy: interphase FISH technique.

Fluids of body cavities result in a series of pathophysiological events associated with non-malignant and malignant conditions that lead to the formation of exudative effusion. Diagnosis of effusion from the patients is frequently troublesome for the cytologist because of the differentiation and biological behavior of different cells type in effusion. In the present study, chromosomal aneuploidy status in effusion cells derived from 32 patients including 14 patients with non-malignant and 18 patients with malignant diseases [including malign mesothelioma (n = 6), adeno carcinoma (n = 10), small cell carcinoma (n = 2)] was analyzed by using fluorescence in situ hybridization (FISH) with centromere specific probes for chromosomes 9 and 11. There was significant difference in the incidence of chromosomal 9 and 11 aneuploidies when compared with controls (P = 0.000). However, aneuploidies of chromosomes 9 and 11 in effusion cells from patients with malignant disease had significantly higher than in effusion cells from patients with non-malignant (P = 0.000), suggesting that chromosomes 9 and 11 are frequently involved in the status of disease. The present study indicates that there is a association between chromosomes aneuploidies and pleural effusion cell status. Chromosome aneuploidies in non-malignant group may be an indicator of premalignancy.

Molecular cytogenetic detection of meiotic segregation patterns in sperm nuclei of carriers of 46,XY,t(15;17)(q21; q25).

Structural chromosomal abnormalities in gonadal tissue represent an important category of parentally transmittable unbalanced chromosomal abnormalities to the offspring. A child with multiple anomalies was sent for cytogenetic analysis, and his karyotype was 46,XY,der(17)t(15;17)(q21; q25). This abnormality was transferred from his grandfather to his father and to the proband. In this family, 5 persons (1 female and 4 male) are the carriers of this abnormality. In this study, fluorescence in situ hybridization (FISH) on sperm nuclei of 4 male carriers was studied to determine the distribution of segregation patterns of the balanced translocation 15q;17q. The segregation results showed that the segregation products in the third carrier (the grandfather) were different, but they were not statistically significant. The segregation patterns in the other carriers were similar. Overall, 50.3% of the sperm nuclei (mean value for 4 carriers) analyzed were the result of alternate segregation; 36.9%, of adjacent I segregation; 9.0%, of adjacent II segregation; and 2.4%, of 3:1 segregation; the remaining 1.3% could be diploid sperm nuclei or of 4:0 segregation. Multicolor FISH analysis appears to be a rapid and reliable method for the direct analysis of segregation patterns in sperm nuclei of carriers of balanced reciprocal translocation, and it also provides interesting information for determining the possible risks for the offspring.

Evaluation of segregation patterns of 21;21 Robertsonian translocation along with sex chromosomes and interchromosomal effects in sperm nuclei of carrier by FISH technique.

Meiotic segregation patterns of carriers of Robertsonian translocations (RT) are important for assessing the risk of unbalanced forms. We investigated the ratio of sperm with t(21;21) to sperm with nullisomy for chromosome 21; the segregation of the t(21;21) along with sex chromosomes, and also interchromosomal effects on chromosome 10 by using three color fluorescence in situ hybridization (FISH) with telomere specific (Tel 21q) and centromere-specific alpha satellite probes for chromosomes X, Y, and 10. The percentage of cosegregation of t(21;21) with sex chromosomes (49.50%) and without sex chromosomes (46.98%) was not significant. There are no significant differences between the percentages of cosegregation of t(21;21) with chromosome X (23.36%) and with chromosome Y (26.16%). No evidence of an interchromosomal effect on chromosome 10 was detected, the percentage of chromosome 10 aneuploidy being similar to that in controls. In addition, the frequency of diploid sperm nuclei was not significantly higher in the carrier (0.32%) than in the controls (0.44%) (P > 0.05). The sex ratio was similar within the carrier and the controls and between the carrier and the control. Three color-FISH analysis, using different probe combinations, seems a rapid and accurate tool for direct analysis of meiotic segregation product.