Comparative study on the deamidation of three recombinant human insulins using capillary electrophoresis.

The applicability of capillary zone electrophoresis (CZE) for the separation of different recombinant human insulins and their deamidated isoforms was studied. The high resolving power of CZE is demonstrated by its ability to separate insulin isoforms differing only by 0.984 Da (different-fold deamidated forms) and even components having the exacts same mass but slightly different shapes (same-fold deamidated forms). From among the several insulins available, humulin, glargine and glulisine were selected for our study because their sequences and chemical parameters are quite similar, however, the small differences present in their amino acid sequences influence the deamidation processes. Using a background electrolyte with basic pH was favourable not only for the separation of the different types of insulin but also for the separation of deamidated protein forms even in a bare fused silica capillary. The LOD values ranged between 0.6 - 0.93 mg/L and 2.17 - 4.37 mg/L for UV and ESI-MS detection, respectively. At -20 - -80 °C, the deamidation is minimal, but at temperatures above +5 °C deamidation is accelerated. At +5 °C only 1-fold deamidation forms could be observed for each insulin. Acidified samples incubated for 1-month at room temperature showed varying levels of deamidation: 1-fold, 1-2-fold and 1-2-3-fold forms for glargine, glulisine and humulin, respectively.

Vitamin D and iron levels correlate weakly with hepcidin levels in postoperative patients with digestive neoplasms undergoing open abdominal surgery.

OBJECTIVE: Patients with digestive neoplasms have both iron deficiency and chronic inflammatory anemia with hepcidin upregulation, which may be aggravated in the postoperative period. Vitamin D impacts hepcidin levels. We aimed to investigate the correlations between vitamin D and iron status vs. hepcidin levels in patients with digestive tumors undergoing open abdominal surgery. PATIENTS AND METHODS: This prospective observational study was performed during 2016-2018 in a University teaching hospital. After obtaining the Ethical Approval and the patients' informed consent, 30 adult patients with digestive tumors were included. Hepcidin, vitamin D and iron levels were measured in the first 24 hours after surgery. RESULTS: We observed a negligible/weak correlation between serum iron and hepcidin levels in the first 24 hours after surgery, with a correlation coefficient of 0.24 and a weak/low correlation between hepcidin and vitamin D levels, with a correlation coefficient of 0.37. CONCLUSIONS: The correlations between vitamin D and hepcidin levels, as well as between hepcidin and serum iron levels, are weak. Interindividual variability in iron-hepcidin-vitamin D regulation might be wide and other regulatory mechanisms might also play important roles in inflammatory anemia modulation in the perioperative period.

Managing menopausal symptoms after cancer.

The joint burden of cancer and menopause impacts millions of women globally. This review provides an approach to management of menopausal symptoms after cancer in all settings. This includes an overview of current evidence for both hormonal and non-hormonal treatments for vasomotor symptoms and vaginal dryness after cancer. Systemic menopausal hormone therapy provides symptom control and may be used after most cancers but should be avoided after estrogen receptor-positive breast cancer and after some other estrogen-dependent cancers. Non-hormonal therapies have been minimally studied in women after a cancer diagnosis and, where they have been studied, it is usually in women with breast cancer. Non-hormonal methods to manage vasomotor symptoms include cognitive behavioral therapy, hypnosis, selective serotonin reuptake inhibitors, serotonin noradrenaline reuptake inhibitors, clonidine, and gabapentin. Vaginal estrogen may be useful to address vaginal dryness. However, safety data in breast cancer patients are still lacking and there is currently no consensus. Lubricants may also help with pain with sexual activity. Management of menopausal symptoms after cancer may be challenging and should include information about induced menopause and possible symptoms as well as available treatments. Management then requires a holistic and multidisciplinary approach with individualized care.

Regulatory T Cells in the Context of New-Onset Diabetes After Renal Transplant: A Single-Center Experience.

BACKGROUND: New-onset diabetes mellitus after transplant (NODAT) is a well-known complication of renal transplant that severely affects graft and patient survival. It is necessary to explore further risk factors and reveal the underlying pathomechanism. METHODS: Renal transplants performed between January 2010 and June 2018 were involved. Exclusion criteria were the recipient age younger than 18 years, follow-up period less than 6 months, and patients with diabetes at the time of transplant. Only primary kidney transplants were involved in our study, which totaled 223 cases. Besides donor and recipient demographic data, the type of immunosuppression, the average fasting glucose level, and T-subset profiles were compared. RESULTS: Of 223 cases there were 33 patients (14.8%) with NODAT (17 female; mean age, 54.2 [SD, 10.3] years; mean body mass index [calculated as weight in kilograms divided by height in meters squared], 27.8 [SD, 5.1]; mean follow-up, 43.3 [SD, 25.5] months). The control group consisted of 190 patients. The average fasting blood glucose level was higher in the NODAT group vs the control group (P < .001). The average fasting blood glucose level above diabetic threshold (≥7 mmol/L) was in association with a 6-fold higher risk of NODAT (odds ratio, 5.86; 95% CI, 2.46-13.97; P < .001). Absolute value of CD4+CD25brightCD127dim regulatory T cells was lower in the NODAT group at the first month after transplant (P = .048) Immunosuppressive protocol and survival data did not differ. CONCLUSIONS: Intensive management of the carbohydrate excursions during the early post-transplant period may decrease the incidence of NODAT. Further investigations will be required to decide whether the reduced CD4+CD25brightCD127dim/regulatory T-cell count contributes the development of NODAT.

Surgical Treatment of Hyperparathyroidism After Kidney Transplant.

Chronic renal impairment is often associated with complex bone disorders. Improvement of secondary hyperparathyroidism (HPT) is expected after kidney transplant (KT) if the glomerular filtration rate is normalized. PATIENTS AND METHODS: There were 888 KTs performed between 1996 and 2017 at our department. A total of 558 general patients have been operated on for HPT during the same period. The 2 populations had a common part: out of the 558, a total of 69 (12.4%) were in end-stage renal failure when operated on because of secondary HPT. That also means that 7.8% of all KTs were associated with HPT. Retrospective, single-center analysis was performed using the patients' medical records. The aim of our study was to analyze the results of parathyroidectomies after KT. RESULTS: Parathyroid surgery was performed on 19 patients (2.14%) because of HPT after KT. The applied surgical technique was total parathyroidectomy with autotransplant in 6 cases, subtotal parathyroidectomy in 3 cases, and selective parathyroidectomy in 10 cases. In all cases, histology revealed benign disease. Complications were observed in 10 cases (52%); there were 6 cases of postoperative hypocalcaemia (31.58%), 1 case of transient laryngeal recurrent nerve paresis (5.26%), and 6 cases of recurrent HPT (31.58%). SUMMARY: The first step of HPT management is calcimimetic drug treatment. It is essential to prevent possible complications with regular laboratory monitoring. If the proper conservative therapy is refractory or severe in complications, surgery should be chosen. If the patient is already waiting for a KT, it is worth performing the parathyroid surgery before KT. Close collaboration with endocrinologists and nephrologists is needed to achieve successful therapy.

Patient Attitudes Toward Transplantation as Preferred Treatment Modality in Different Stages of Renal Disease.

OBJECTIVE: Kidney transplantation is generally considered to be the best treatment for end-stage renal disease. Not every patient can be operated, but many of suitable patients refuse this possibility. We aimed to explore the attitudes of patients with chronic kidney disease towards renal replacement therapies to recognize motives, thoughts, and feelings concerning accepting or refusing the treatment. We studied the attitudes towards the illness and the treatment, the appearance of depression, and the disease burden during different stages of the disease. METHODS: For this study we implemented a questionnaire that we developed (which has been described in an earlier publication of this journal) with 99 pre-dialysis patients, 99 dialysis patients, and 87 transplantation patients. We completed the attitude questionnaire designed by our team to include disease burden and depression questionnaires. RESULTS: We used discriminant analysis to describe different stages of the disease. There was a significant difference in the following factors between the three patient groups: accepting the new kidney, lack of confidence in transplantation therapy, fear of surgery, accepting self-responsibility in recovery, dependency on the transplanted kidney, confidence in recovery, subjective burden of dialysis, and denial of personal responsibility in maintaining the transplanted kidney. Significant differences were also detected in these three groups regarding the level of depression and disease burden: we measured the highest value among the dialysis patients, and the lowest value among the pre-dialysis patients. Comparing patients accepting and refusing transplantation, we found a correlation between the refusal of transplantation and the attitudes towards the illness and treatment. CONCLUSIONS: Most patients remain unmotivated to change treatment modality and refuse transplantation. Misbelief about transplantation shows a correlation with the motive of refusal. Dissemination of information may facilitate a change in the situation.

Autosomal Dominant Polycystic Kidney Disease Transplant Recipients After Kidney Transplantation: A Single-center Experience.

Kidney transplantation is indicated for end-stage renal disease. Autosomal dominant polycystic kidney disease (ADPKD) causes structural degeneration of the kidney and eventually becomes end-stage renal disease. ADPKD patients usually have several renal and nonrenal complications. We analyzed our kidney transplantation activities between 1991 and 2010 regarding ADPKD. We followed up with patients to December 31, 2016. Data were collected as patient and graft survival rates, the prevalence of polycystic manifestation of the gastrointestinal tract and other organs, and the attendance of urinary tract infection. Among the 734 kidney transplantations, 10.9% (n = 80) had an ADPKD. Four patients (5%) had diverticulum perforation. The prevalence of post-transplantation urinary tract infection was higher in ADPKD patients (55.9%) compared to non-ADPKD patients (44.1%). The 1-, 3-, and 5-year overall survival rates in ADPKD recipients versus non-ADPKD patients are 77.5%, 70.0%, and 67.5% versus 86.4%, 83.0%, and 80.1%, respectively. Patients with ADPKD were transplanted at an elder age compared to others (median: 47.5 years vs. 39.9 years). Female patients had longer graft survival times than males. ADPKD implies multiple cystic degeneration of the kidneys; however, it can cause structural degeneration in other organs. It is typical for ADPKD patients to have an acute abdominal-like syndrome. Immunosuppressive drugs can hide the clinical picture, which makes early diagnosis difficult.

Ophthalmic Findings in Patients After Renal Transplantation.

The aim of the study was to perform complete ophthalmic examinations in patients after renal transplantation to determine ocular alterations and condition of the eyes. Moreover, ophthalmic findings were correlated with certain clinical characteristics related to transplantation such as post-operative renal functions and immunosuppressive regimen. The study was performed on 84 eyes of 42 patients who had received a renal transplant at least 6 months earlier. All patients underwent a complete ophthalmologic examination. In addition, in 33 (78.6%) patients peripapillary retinal nerve fiber layer (RNFL) thickness was determined using optical coherence tomography (Stratus OCT, Zeiss, Germany), which is a unique finding among renal transplantation patients. Recipients received immunosuppressive therapy consisting of tacrolimus, mycophenolate mofetil, and corticosteroid. Of 42 patients, 19 (45%) were women and 23 (55%) were men. The age of the patients ranged from 26 to 74 years, the mean age was 53.4 years. At least one ocular abnormality could be detected in 37 patients (88%), including impaired visual acuity (n = 31, 74%), keratoconjunctivitis sicca (n = 6, 14.3%), pinguecula (n = 3, 7.1%), arcus lipoides (n = 1, 2.4%), cataracts (n = 24, 57.1%), glaucoma (n = 2, 5%), retinal drusen (n = 6, 14.3%), and hypertensive or atherosclerotic retinopathy (n = 22, 52.4%). Twenty-five patients (75.8%) have reduced RNFL thickness. Cataract formation was positively correlated with age and usage of methylprednisolone. Moreover, RNFL thickness loss was correlated with transplantation duration and postoperative infections. Our study suggests that ocular disorders are frequent among renal transplantation patients. Besides immunosuppression and postoperative infection, aging is a high-risk factor in such cases.

Antiviral therapy and prevention against hantavirus infections.

Hantaviruses are emerging zoonoses hosted by small mammals. In humans, they cause two diseases. Hemorrhagic fever with renal syndrome is mainly caused by Dobrava-Belgrade virus, Puumala virus, Seoul virus and Hantaan virus in Asia and Europe. On the other hand, the most important causes of hantavirus cardiopulmonary syndrome are Sin Nombre virus and Andes virus in Americas. Ribavirin yet remains the only licensed drug against the hantavirus infections, but its sufficient antiviral activity remains an issue under discussion. There are still no available vaccines against hantaviruses except of some inactivated virus vaccines licensed only in East-Asian countries. Some of the vaccines are under development in pre-clinical stages. The review discuses about specific compounds with approved antiviral activity against hantaviruses. Other approaches such as development of vaccines, are compiled as well.

Next generation tau models in Alzheimer's disease research - virus based gene delivery systems.

Alzheimer's disease (AD) the most common form of dementia is characterized by cognitive decline and progressive loss of neurons in the central nervous system. Despite huge scientific progress, there are only few animal models that recapitulate at least majority of the AD pathology and related symptomatology. Therefore, alternative methods to develop animal models for neurodegenerative diseases are constantly explored. Recently, recombinant adeno-associated viruses (AAVs) are widely used viral vectors in development of novel models for neurodegenerative diseases. AAV vectors expressing full length, mutant or truncated forms of tau demonstrate early and robust pathology characterized by AT8 positivity, NFT formation, motor and cognitive deficits. Furthermore, AAVs have been used in expression of tau in amyloid rodent models thus developing both lesions of amyloid and tau therefore recapitulating AD like features. Major advantage of AAV as a delivery system is the site specific expression of tau, mostly in hippocampus and cortex, and thus elimination of unwanted ectopic transgene expression. These novel models may help in better understanding of AD etiopathogenesis and provide a platform for development and testing of disease modifying drugs in preclinical efficacy studies.

Ghrelin-Induced Enhancement of Vasopressin and Oxytocin Secretion in Rat Neurohypophyseal Cell Cultures.

The effects of ghrelin on vasopressin and oxytocin secretion were studied in 13-14-day cell cultures of isolated rat neurohypophyseal tissue. The vasopressin and oxytocin contents of the supernatant were determined by radioimmunoassay after a 1- or 2-h incubation. Significantly increased levels of vasopressin and oxytocin production were detected in the cell culture media following ghrelin administration, depending on the ghrelin doses. The oxytocin level proved to be more elevated than that of vasopressin. The increase of vasopressin and oxytocin secretion could be totally blocked by previous administration of the ghrelin receptor antagonist ([D-Lys3]-growth hormone-releasing peptide-6). Application of the ghrelin receptor antagonist after ghrelin administration proved ineffective. The results indicate that vasopressin and oxytocin release is influenced directly by the ghrelin system, and the effects of ghrelin on vasopressin and oxytocin secretion from the neurohypophyseal tissue in rats can occur at the level of the posterior pituitary. Our observations lend support to the view that neurohypophysis contains ghrelin receptors.

Bacterial Infections After Kidney Transplantation: A Single-Center Experience.

BACKGROUND: Bacterial infections significantly affect graft loss and mortality after kidney transplantation (KT). We reviewed the frequencies, risk factors, and sources of bacterial infections after KT and their impact on graft and patient survivals. METHODS: The data of 154 kidney recipients who underwent transplantation from 2010 to 2015 were explored. Donor, recipient, and surgical parameters were collected, and source, type, and frequency of infectious complications, number of infective episodes, multidrug-resistant (MDR) bacteria, and the bacterial spectrum were established. RESULTS: The most common infection was urinary tract infection, which is in line with the literature. Out of the 154 recipients, 72.1% (n = 111) had at least 1 occasion of a bacterial infection episode with clinical symptoms. It occurred 0-43 months (mean, 19.5 mo) after transplantation. Ninety-three KT recipients (67.9%) developed 274 episodes of infection in the postoperative 1st year (1.8 episodes/patient/y), and 42 patients had admission to the hospital ward (5.2 d/patient/y). MDR was detected in 19.8% of the infections. CONCLUSIONS: A bacterial infection had no significant impact on survival by itself. However, in case of sepsis graft and patient survivals were lower compared with normal control subjects.

Passenger mutations and aberrant gene expression in congenic tissue plasminogen activator-deficient mouse strains.

UNLABELLED: Essentials C57BL/6J-tissue plasminogen activator (tPA)-deficient mice are widely used to study tPA function. Congenic C57BL/6J-tPA-deficient mice harbor large 129-derived chromosomal segments. The 129-derived chromosomal segments contain gene mutations that may confound data interpretation. Passenger mutation-free isogenic tPA-deficient mice were generated for study of tPA function. SUMMARY: Background The ability to generate defined null mutations in mice revolutionized the analysis of gene function in mammals. However, gene-deficient mice generated by using 129-derived embryonic stem cells may carry large segments of 129 DNA, even when extensively backcrossed to reference strains, such as C57BL/6J, and this may confound interpretation of experiments performed in these mice. Tissue plasminogen activator (tPA), encoded by the PLAT gene, is a fibrinolytic serine protease that is widely expressed in the brain. A number of neurological abnormalities have been reported in tPA-deficient mice. Objectives To study genetic contamination of tPA-deficient mice. Materials and methods Whole genome expression array analysis, RNAseq expression profiling, low- and high-density single nucleotide polymorphism (SNP) analysis, bioinformatics and genome editing were used to analyze gene expression in tPA-deficient mouse brains. Results and conclusions Genes differentially expressed in the brain of Plat(-/-) mice from two independent colonies highly backcrossed onto the C57BL/6J strain clustered near Plat on chromosome 8. SNP analysis attributed this anomaly to about 20 Mbp of DNA flanking Plat being of 129 origin in both strains. Bioinformatic analysis of these 129-derived chromosomal segments identified a significant number of mutations in genes co-segregating with the targeted Plat allele, including several potential null mutations. Using zinc finger nuclease technology, we generated novel 'passenger mutation'-free isogenic C57BL/6J-Plat(-/-) and FVB/NJ-Plat(-/-) mouse strains by introducing an 11 bp deletion into the exon encoding the signal peptide. These novel mouse strains will be a useful community resource for further exploration of tPA function in physiological and pathological processes.

Cardiovascular Screening and Management Among Kidney Transplant Candidates in Hungary.

INTRODUCTION: Cardiovascular disease is a major cause of morbidity and mortality in end-stage renal disease patients on dialysis and the most common cause of death in the immediate post-transplantation period. The aim of our study was to describe a novel approach of cardiovascular screening and management of dialysis patients evaluated for the transplant waiting list. METHODS: Twenty-eight patients with end-stage renal disease put on the waiting list between July 2013 and July 2014 were subjected to a prespecified cardiovascular screening protocol utilizing noninvasive and/or invasive tests. Patients were subsequently divided into 3 strata in terms of their estimated cardiovascular risk. Each of these groups were then prescribed interventions aiming to improve their cardiovascular condition. RESULTS: According to our prespecified protocol of cardiovascular screening studies, 15 (54%) patients were identified as low, 5 (18%) as intermediate, and 8 (28%) as high risk. Four (14%) patients were current smokers. In the low-risk group, we initiated a patient education program involving counseling on regular exercise such as swimming or cycling to improve their functional capacity. In the high-risk group revascularization was done in 5 cases (63%), including 3 percutaneous transluminal coronary angioplasties (PTCA) with stents for single-vessel disease, and coronary artery bypass graft surgeries (CABG) for triple-vessel disease in 2 cases. In the medium-risk group medical management was opted for, including introduction of beta-blockers, inhibitors, statins, and ezetimibe, as well as efforts to optimize anemia management, indices of bone-mineral disease, and fluid status. CONCLUSION: In our regional transplant program, we introduced a comprehensive multidisciplinary approach to treat potential transplant candidates according to cardiovascular risk stratification based on a prespecified screening protocol. Further studies are needed to correlate this novel strategy with post-transplantation outcomes.

TOXICITY OF CHLORPYRIPHOS CONTAINING FORMULATION AND HEAVY ELEMENTS (Cd, Pb) TO CHICKEN EMBRYOS.

The phytotoxic effect of cadmium sulphate, lead acetate and chlorpyriphos containing insecticide Pyrinex 48 EC were investigated on chicken embryos on late developmental stage (day 19 of incubation). The eggs were injected by 0.1 ml amount of lead acetate (0.1%), of cadmium sulphate (0.01%), of Pyrinex 48 EC (chlorpyriphos, 480 g/l; 1%) and as a control 0.1 ml of bird-physiologic saline solution (0.75%). The treatments were performed on day 0 of incubation, and the embryos were examined on day 19. Number of embryonic death, developmental abnormalities and body weight of embryos were recorded on day 19. The body weight of embryos reduced significantly, the rate of embryo mortality increased, but teratogenic effect was not realised by the single treatment of lead acetate and cadmium sulphate. By the single administration of PYRINEX 48 EC the average body weight of embryos resulted in a significant decrease and the rate of embryo mortality and developmental anomalies were increased and it was found to be embryo toxic and teratogenic in the embryos. The increased embryo toxic effect can occur by the additive effect of the heavy elements (cadmium sulphate and lead acetate) and the PYRINEX 48 EC.

Non-hematopoietic PAR-2 is essential for matriptase-driven pre-malignant progression and potentiation of ras-mediated squamous cell carcinogenesis.

The membrane-anchored serine protease, matriptase, is consistently dysregulated in a range of human carcinomas, and high matriptase activity correlates with poor prognosis. Furthermore, matriptase is unique among tumor-associated proteases in that epithelial stem cell expression of the protease suffices to induce malignant transformation. Here, we use genetic epistasis analysis to identify proteinase-activated receptor (PAR)-2-dependent inflammatory signaling as an essential component of matriptase-mediated oncogenesis. In cell-based assays, matriptase was a potent activator of PAR-2, and PAR-2 activation by matriptase caused robust induction of nuclear factor (NF)κB through Gαi. Importantly, genetic elimination of PAR-2 from mice completely prevented matriptase-induced pre-malignant progression, including inflammatory cytokine production, inflammatory cell recruitment, epidermal hyperplasia and dermal fibrosis. Selective ablation of PAR-2 from bone marrow-derived cells did not prevent matriptase-driven pre-malignant progression, indicating that matriptase activates keratinocyte stem cell PAR-2 to elicit its pro-inflammatory and pro-tumorigenic effects. When combined with previous studies, our data suggest that dual induction of PAR-2-NFκB inflammatory signaling and PI3K-Akt-mTor survival/proliferative signaling underlies the transforming potential of matriptase and may contribute to pro-tumorigenic signaling in human epithelial carcinogenesis.

Potent and specific inhibition of the biological activity of the type-II transmembrane serine protease matriptase by the cyclic microprotein MCoTI-II.

Matriptase is a type-II transmembrane serine protease involved in epithelial homeostasis in both health and disease, and is implicated in the development and progression of a variety of cancers. Matriptase mediates its biological effects both via as yet undefined substrates and pathways, and also by proteolytic cleavage of a variety of well-defined protein substrates, several of which it shares with the closely-related protease hepsin. Development of targeted therapeutic strategies will require discrimination between these proteases. Here we have investigated cyclic microproteins of the squash Momordica cochinchinensis trypsin-inhibitor family (generated by total chemical synthesis) and found MCoTI-II to be a high-affinity (Ki 9 nM) and highly selective (> 1,000-fold) inhibitor of matriptase. MCoTI-II efficiently inhibited the proteolytic activation of pro-hepatocyte growth factor (HGF) by matriptase but not by hepsin, in both purified and cell-based systems, and inhibited HGF-dependent cell scattering. MCoTI-II also selectively inhibited the invasion of matriptase-expressing prostate cancer cells. Using a model of epithelial cell tight junction assembly, we also found that MCoTI-II could effectively inhibit the re-establishment of tight junctions and epithelial barrier function in MDCK-I cells after disruption, consistent with the role of matriptase in regulating epithelial integrity. Surprisingly, MCoTI-II was unable to inhibit matriptase-dependent proteolytic activation of prostasin, a GPI-anchored serine protease also implicated in epithelial homeostasis. These observations suggest that the unusually high selectivity afforded by MCoTI-II and its biological effectiveness might represent a useful starting point for the development of therapeutic inhibitors, and further highlight the role of matriptase in epithelial maintenance.

TOXIC INTERACTION OF CHLORPYRIPHOS AND COPPER SULPHATE ON CHICKEN EMBRYO.

The single and simultaneous phytotoxic effect of copper sulphate and Pyrinex 48 EC insecticide were investigated on chicken embryos. The eggs were injected by 0.1 ml of copper sulphate solution (0.05%) and/or by 0.1 ml of Pyrinex 48 EC (chlorpyriphos, 480 g/l; 1%). The treatments were performed on day 0 of incubation, and the embryos were examined on day 3 and 19 of it. Germinal disc was prepared to study the early stage of development. Number of embryonic death, developmental abnormalities and body weight of embryos were recorded on day 19. Liver samples were taken for histology and the skeleton was stained by Dawson method. The embryo mortality was not influenced by single treatment of copper sulphate, however, Pyrinex 48 EC and the combination of the test items significantly increased it on day 3. Same tendency was observed in the case of developmental aberration. Single administration of both test items and their combination significantly increased the embryo mortality on day 19. Frequency of abnormalities was not influenced by copper sulphate but single and simultaneous application of insecticide increased it significantly. Pyrinex 48 EC alone and in combination with copper sulphate significantly reduced the body weight, however, the copper sulphate alone did not influence it. Developmental abnormalities were observed sporadically in all cases. There were no findings of drug-induced hepatopathy, however, the ratio of the mitotic cells were markedly reduced. Based on the results, addition and synergistic toxic interaction may be between the copper sulphate and Pyrinex 48 EC that can highly reduce the viability of the embryos.

WEED SURVEYING OF PHACELIA (PHACELIA TANACETIFOLIA L.) AND EVALUATING THE EFFICIENCY OF THE WEED CONTROL.

The experiment was set up in an area of 9 ha that was split into 4 plots: in plot 1 the row spacing was 12 cm and the seeding rate was 10 kg; in plot 2 the row spacing was 24 cm and the seeding rate was 10 kg; in plot 3 the row spacing was 24 cm and the seeding rate was 8 kg; in plot 4 the row spacing was 12 cm and the seeding rate was 8 kg. After the weed surveying, the total weed coverage was established as follows: in plot 1 the total weed coverage was 11.34%, in plot 2 it was 12.3%, in plot 3 it was 18%, and in plot 4 the total weed coverage was 15%. Based on the weed survey, on the test area the following dicotyledon weeds belonging to the T4 Raunkiaer plant life-form category occupied the highest percentage: heal-all, black-bindweed, goosefoot. The proportion of the perennial dicotyledons: field bindweed (G3), tuberous pea (G1), white campion (H3) was negligible. In all four cases the weed control was executed using the same herbicide in the same doses and with regard to the weed species it showed the same level of efficiency. The smaller row spacing and higher seeding rate has a beneficial effect on the weed suppressing capacity of the crop, the crop's weed suppressing capacity is better and the development of the weeds becomes worse.