Genetic diversity studies based on morpho-pathological and molecular variability of the Sclerotinia sclerotiorum population infecting potato (Solanum tuberosum L.).

White mould or stem rot, caused by Sclerotinia sclerotiorum (Lib.) de Bary, is a devastating fungal disease found in major potato cultivation areas worldwide. The aim of this study was to characterize genetic diversity in the S. sclerotiorum population from the main potato producing regions of India by means of morphological (mycelial growth, colony colour, number and distribution pattern of sclerotia) and molecular characteristics, as well as to evaluate the virulence of S. sclerotiorum isolates in potato for the first time. Among the S. sclerotiorum population analyzed, high phenotypic and genotypic diversity were observed. Using all the morphological characteristics, a dendrogram was constructed based on Gower's similarity coefficient that distributed all the isolates into three clusters at the 0.62 similarity coefficient. Carpogenic germination of apothecia revealed that larger sclerotia produced a greater number of apothecia while smaller sclerotia produced fewer apothecia. Pathogenicity test results revealed that out of 25 isolates, seven were highly aggressive, 14 were moderate and four had low aggressiveness, whilst isolates from Punjab were more pathogenic than those of Uttar Pradesh. Phylogenetic analysis of universal rice primer polymorphism showed high genetic variability within the isolates that grouped all the isolates in three evolutionary lineages in the resulting dendrogram and showed partial relationship with geographical locations of the isolates. Further, the findings suggest the occurrence of higher heterogeneity and genetic diversity among the S. sclerotiorum isolates that indicates the existence of both clonal and sexual reproduction in the pathogen population of potato producing areas in India.

Genome sequence and analysis of the tuber crop potato.

Potato (Solanum tuberosum L.) is the world's most important non-grain food crop and is central to global food security. It is clonally propagated, highly heterozygous, autotetraploid, and suffers acute inbreeding depression. Here we use a homozygous doubled-monoploid potato clone to sequence and assemble 86% of the 844-megabase genome. We predict 39,031 protein-coding genes and present evidence for at least two genome duplication events indicative of a palaeopolyploid origin. As the first genome sequence of an asterid, the potato genome reveals 2,642 genes specific to this large angiosperm clade. We also sequenced a heterozygous diploid clone and show that gene presence/absence variants and other potentially deleterious mutations occur frequently and are a likely cause of inbreeding depression. Gene family expansion, tissue-specific expression and recruitment of genes to new pathways contributed to the evolution of tuber development. The potato genome sequence provides a platform for genetic improvement of this vital crop.

Development and application of reverse transcription-loop mediated isothermal amplification assay for sensitive detection of groundnut bud necrosis virus infecting potato.

Reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detection of groundnut bud necrosis virus (GBNV) causing potato stem necrosis disease. The isothermal temperatures, reaction periods and concentrations of reaction mixture were optimized where, the assay worked well at 65 °C for 50 min, 6 U of WarmStart Bst 2.0 DNA polymerase, 1.4 mM dNTPs and 2.0 mM MgSO4. The optimized assay proved to be specific to GBNV with no cross reactivity to other viruses infecting potato in India. The specificity of RT-LAMP assay was found to be 100 fold more sensitive than that of RT-PCR. The developed assay was applied for the detection of GBNV from 80 potato leaf samples where 24 samples were found infected which was confirmed by RT-PCR. It was concluded that the RT-LAMP assay developed for detection of GBNV was specific, sensitive and suitable for its use in virus indexing under potato seed production programme.

Influence of drying of biosludge on organochlorine compounds from pulp and paper industry.

Pulp and paper industry is one of the major sources of man-made generation of organochlorine compounds. During biological treatment of wastewater, part of organochlorine compounds is discharged with treated effluent and part is retained on biomass and disposed of as waste activated sludge. Due to presence of these compounds, the disposal of biosludge from pulp and paper industry has become an issue. The estimation of adsorbable organic halogen (AOX) compounds after drying and grinding resulted in 49% lower concentration of AOX due to stripping of purgeable compounds. These purgeable compounds are not released at 60 degrees C in aqueous medium during estimation of purgeable organic halogen (POX) compounds. Dispersion of sludge by sonication overcomes the loss of POX compounds and results in higher concentration ofAOX compounds. The drying of biosludge samples at 45, 100 degrees C and in presence of sun light resulted in 20.1, 49.0 and 29.6% removal of purgeable AOX compounds, respectively. The lab scale sorption study using dichloromethane (as volatile organochlorine compound) reveal that biosludge from pulp and paper industry is a good adsorbent of volatile organochlorine compounds and results in poor release of these compounds during estimation of POX compounds.

Spraying of dsRNA molecules derived from Phytophthora infestans, along with nanoclay carriers as a proof of concept for developing novel protection strategy for potato late blight.

BACKGROUND: Phytophthora infestans is a late blight-causing oomycetes pathogen. It rapidly evolves and adapts to the host background and new fungicide molecules within a few years of their release, most likely because of the predominance of transposable elements in its genome. Frequent applications of fungicides cause environmental concerns. Here, we developed target-specific RNA interference (RNAi)-based molecules, along with nanoclay carriers, that when sprayed on plants are capable of effectively reducing late blight infection. RESULTS: Targeted the genes unique to sporulation, early satge infection and the metabolism pathway stages based on in an our own microarray data. We used nanoclay as a carrier for sorbitol dehydrogenase, heat shock protein 90, translation elongation factor 1-α, phospholipase-D like 3 and glycosylphosphatidylinositol-anchored acidic serine-threonine-rich HAM34-like protein double-stranded (ds)RNAs, which were assessed by culture bioassay, detached leaf assay and spray methods, and revealed a reduction in growth, sporulation and symptom expression. Plants sprayed with multigene targeted dsRNA-nanoclay showed enhanced disease resistance (4% disease severity) and less sporulation (<1 × 103 ) compared with plants sprayed with dsRNA alone. CONCLUSION: The use of nanoclay with multigene targeted dsRNA was assumed to be involved in effective delivery, protection and boosting the action of RNAi as a spray-induced gene silencing approach (SIGS). A significant reduction in growth, sporulation, disease severity and decreased gene expression authenticates the effects of SIGS on late blight progression. This study demonstrated as a proof of concept the dsRNA-nanoclay SIGS approach, which could be used as an alternative to chemical fungicides and transgenic approaches to develop an environmentally friendly novel plant protection strategy for late blight. © 2022 Society of Chemical Industry.

Degradation of pentachlorophenol by Kocuria sp. CL2 isolated from secondary sludge of pulp and paper mill.

A pentachlorophenol (PCP) degrading bacterium was isolated and characterized from sludge of pulp and paper mill. This isolate used PCP as its sole source of carbon and energy and was capable of degrading this compound, as indicated by stoichiometric release of chloride and biomass formation. Based on morphology, biochemical tests, and 16S rRNA gene sequence analysis this strain was identified as Kocuria sp. CL2. High Performance Liquid Chromatography (HPLC) analysis revealed that this strain was able to degrade PCP up to a concentration of 600 mg/l. This is first time we are reporting the degradation of PCP by the Kocuria species. This isolate was also able to remove 58.64% of PCP from the sludge within two weeks. This study showed that the removal efficiency of PCP by CL2 was found to be very effective and can be used in degradation of PCP containing pulp paper mill waste in the environment.

Effect of ozonation on activated sludge from pulp and paper industry.

Aerobic biological treatment with activated sludge is the predominant process all over the world for treatment of pulp and paper industry wastewater. 50-70% of the biodegradable organic material is oxidized to CO2 and the rest is converted to bacterial biomass, typically termed as excess sludge or waste activated sludge (WAS). Handling and disposal of WAS in general and in particular from the pulp and paper industry face different processing difficulties, regulatory stringency due to organochlorine contamination and reluctance of people for reuse. With an objective of reducing the net disposable biomass, ozonation of WAS from a pulp and paper mill and from a laboratory scale batch activated sludge process operated with the wastewater and bacterial seed of the same pulp and paper mill have been carried out. With the mill sludge having predominant filamentous organisms 18% MLSS was reduced at an ozone dosage of 55 mg O3/g dry MLSS solid (DS) resulting in 2.5 times COD increase. With the laboratory sludge which is well structured and flocculating, only 6% MLSS was reduced at an ozone dosage of 55 mg O3/g DS. Ozonation mineralizes 26% and 20% AOX compounds embedded in the secondary sludge in the mill and laboratory sludge respectively at an ozone dosage of 55 mg O3/g DS. During ozonation, absorbed/adsorbed lignin on biomass was released which resulted in increased colour concentration. Ozonation can be a potential oxidative pretreatment process for reducing the WAS and paving the way for cost effective overall treatment of WAS.

Development of a visual detection method for Potato virus S by reverse transcription loop-mediated isothermal amplification.

A reverse transcription-loop mediated isothermal amplification (RT-LAMP) assay was developed to detect the Potato virus S (PVS) in potato. Two sets of six novel primers that recognize the coat protein gene sequence of the PVS were designed and RT-LAMP assay was optimized for the parameters such as different concentrations of primers, MgSO4, betaine, dNTPs, Bst DNA polymerase, temperature and duration. The RT-LAMP was carried out under isothermal conditions without the thermal cycler using PVS infected leaf and tuber samples, LAMP specific primers with amplification at 65 °C for 60 min, and 80 °C for 5 min. The results were assessed by gel electrophoresis and visual observation of colour change using SYBR Green I dye. The detection limit of the developed RT-LAMP assay was determined and compared with a conventional reverse transcription-polymerase chain reaction (RT-PCR). RT-LAMP was found 100 times more sensitive than RT-PCR. The optimized RT-LAMP assay is robust, reliable, sensitive and convenient for the detection of the PVS in infected potato tubers including asymptomatic plants. No cross-reactions were observed with healthy plants and other potato viruses. The assay is economical and can be employed in large scale testing of potato plants against PVS under healthy seed potato production programme.

Multiple displacement amplification as a pre-polymerase chain reaction (pre-PCR) to detect ultra low population of Ralstonia solanacearum (Smith 1896) Yabuchi et al. (1996).

AIMS: To develop a reliable and sensitive protocol for detection of Ralstonia solanacearum using MDA-PCR (Multiple displacement amplification-PCR amplification). METHODS AND RESULTS: MDA-PCR technique was performed on pure cell lysates as well as soil samples. Pure cell lysate as well as that of soil DNA was used as template in MDA reaction. MDA of template DNA was carried out in the presence of sample buffer, reaction buffer and enzyme mix (Phi 29 DNA polymerase and random hexamers). The MDA amplified DNA was used for PCR amplification using R. solanacearum -specific PCR primers. MDA-PCR could detect as low as 1 colony forming unit (CFU ml(-1)) of bacteria within 8 h including DNA isolation. CONCLUSION: MDA followed by standard PCR facilitated the detection of pathogen from very low count samples. The method is of great importance in managing the brown rot disease of potato. SIGNIFICANCE AND IMPACT OF STUDY: The ultrasensitive detection technique developed in the present study is sensitive and speedy enough to be included into integrated wilt disease control programmes.

Assessment of genetic fidelity of micropropagated apple rootstock plants, EMLA 111, using RAPD markers.

EMLA 111(East Malling Long Ashton), a clonal rootstock of apple (Malus pumila Mill.) was micropropagated using axillary bud and shoot apices. The present work was carried out to assess the genetic fidelity of micropropagated plants using random amplified polymorphic DNA (RAPD). Ten arbitrary decamer primers have been used to amplify genomic DNA from in vitro raised field material and mother plant. A total of 57 amplified products were obtained, out of which 53 were monomorphic across the mother tree and its tissue culture raised progenies. Of the ten primers used, 8 showed RAPD profiles which were identical to the mother plant. Similarity matrix based on Jaccard's coefficient revealed that pairwise value between the mother plant and its tissue cultured plants ranged from 0.93 to 1.00 and among tissue cultured plants, it was 0.92 to 1.00, thus indicating a high degree of genetic fidelity.

Complete mitogenome mapping of potato late blight pathogen, Phytophthora infestans A2 mating type.

Complete mitochondrial genome of Phytophthora infestans, A2 mating type (MT) with a size of ≅37,767 bp was sequenced. A total of 53 protein-coding genes are predicted on both strands, including 25 tRNA, 2 rRNA, and 18 respiratory proteins. Gene order of A2MT was consistent with that established in A1, despite high level of polymorphism in both coding and non-coding regions. The mtDNA of A2MT was found to have 99.5% and 99.4% homology with Ia and Ib, whereas 94.7% and 94.3% with IIa and IIb, respectively. Study of repeats revealed a dinucleotide (AT)9 specific to A1 and homology of cox1 gene sequence revealed the relationship among 50 Phytophthora species.

Genetic diversity and differentiation of Indian isolates of Phytophthora infestans as revealed by RAPD analysis.

Sixty-seven isolates of Phytophthora infestans collected from Himalayan hill regions and subtropical planes of India were characterized by RAPD markers to assess diversity and differentiation based on location of origin. Ten random decamer primers generated 161 polymorphic fragments. Association of P. infestans isolates on the dendrogram and PCO plot revealed two clear grouping based on geographical location of origin-hill isolates and plane isolates. Quantification of diversity by Shannon index of diversity analysis demonstrated that most of the diversity was present with a particular population (hill or plane) of P. infestans isolates, with 85% variation being within and 15% being between hill and plane isolates. Subtropical plane isolates of P. infestans exhibited higher variability compared to hill isolates and they were more dispersed on the PCO plot. No clear differentiation of isolates based on mating type was reflected on the dendrogram and PCO plot.

Role of oxidative stress in nickel chloride-induced cell injury in rat renal cortical slices.

Nickel chloride (NiCl2) induced lactate dehydrogenase (LDH) release and lipid peroxidation (LPO) in rat renal cortical slices in vitro in a concentration- (0-2 mM) and time- (0-4 hr) dependent manner, with initial significant LDH release occurring as early as 1 hr, whereas significant increase in LPO started 3 hr after exposure, suggesting that LPO results from renal cell injury. Both NiCl2-induced LDH release and LPO were prevented significantly by glutathione and dithiothreitol, suggesting that NiCl2-induced renal cell injury is dependent on thiols. However, such injury is not dependent solely on thiols, because (a) these thiols failed to inhibit completely the uptake of Ni2+ by the renal cortex, and (b) diethylmaleate pretreatment failed to increase NiCl2-induced cell injury further. Superoxide dismutase partially reduced the NiCl2-induced LDH release without affecting LPO and glutathione, whereas catalase did not affect such LDH release and LPO. Dimethylthiourea and DMSO completely prevented NiCl2-induced LPO, but only partially reduced LDH release. Deferoxamine prevented NiCl2-induced renal cell injury without affecting LPO and without significantly reducing Ni2+ uptake by the renal cortex, suggesting that nickel chelation is not important in such prevention of injury. NiCl2-induced inhibition of para-aminohippurate uptake was prevented significantly by thiols, deferoxamine, and dimethylthiourea. NiCl2-induced loss of cellular glutathione content was prevented significantly by thiols and deferoxamine, but not by superoxide dismutase and dimethylthiourea. These results suggest that LPO was not related to NiCl2-induced lethal renal cell injury, whereas such injury may be caused by the induction of the Fenton reaction, generating hydroxyl radicals.

Effects of various pretreatments on the acute nephrotoxic potential of styrene in Fischer-344 rats.

The effects of various inducers and inhibitors of hepatic microsomal mixed-function oxidase (MFO) system and diethylmaleate treatment on styrene-induced acute nephrotoxicity in male Fischer-344 rats were studied. Groups of rats were pretreated with either 3-methylcholanthrene (15 mg/kg, i.p., 3 days), or phenobarbital (80 mg/kg, i.p., 3 days), or SKF525-A (50 mg/kg, i.p., 1 h), or piperonyl butoxide (300 mg/kg, i.p., 2 h), or diethylmaleate (400 mg/kg, i.p., 90 min) prior to an i.p. administration of styrene (0, 0.6 and 0.9 g/kg) in corn oil. The uptake of p-aminohippurate (PAH) by renal cortical slices, the morphology of renal cortices, as well as urinary excretion of N-acetyl-beta-D-glucosaminidase (NAG) and gamma-glutamyl transpeptidase (gamma-GT) of control and pretreated rats were examined 24 h after styrene. The inducers and inhibitors of MFO system failed to modify further the acute nephrotoxicity of styrene. On the other hand, diethylmaleate pretreatment not only reduced further the uptake of PAH, but also produced further significant increase in the urinary excretion of NAG and gamma-GT observed at the higher dose of styrene. Similarly, ultrastructural studies showed a moderate increase in the severity of kidney damage induced at the higher dose of styrene due to pretreatment with diethylmaleate. These data suggest that tissue glutathione concentrations and hence, corresponding conjugating activity might be important determinants of styrene nephrotoxicity. The results further indicate that a metabolic activation system not involving certain cytochrome P-450 might be responsible in styrene-induced nephrotoxicity.

Studies on the subchronic nephrotoxic potential of styrene in Sprague-Dawley rats.

The nephrotoxic potential low non-toxic dose of styrene was studied in male Sprague-Dawley rats. Groups of rats received i.p. injections of styrene in corn oil at doses 0, 2.9, and 5.8 mmol/kg once daily, 5 days/week for 6 consecutive weeks. After collection of urine for 0-24 and 24-48 h following the end of the treatment, the rats were sacrificed. A significant increase in the excreted urinary volume was noticed at 5.8 mmol styrene during 0-24 and 24-48 h, relative to control, whereas urinary concentrations of gamma-glutamyl transpeptidase and glucose were significantly elevated during the 24-48-h period. Urinary activity of N-acetyl-beta-D-glucosaminidase was increased at the higher dose of styrene during 0-24 and 24-48 h. The capacity of renal cortical slices to accumulate p-aminohippurate was significantly reduced 48 h after the exposure to any dose of styrene. Electron microscopic examination of renal cortex 48 h after the exposure to a higher dose revealed the presence of enlarged mitochondria having more electron dense matrix. The data suggest that subchronic exposure to a very low non-toxic dose of styrene may have the potential to elicit nephrotoxicity preferentially in the proximal tubular region of the rat kidney.

Effects of different nickel compounds on the transport of para-aminohippurate ion by rat renal cortical slices.

In vitro rat renal cortical slice uptake of para-aminohippurate ion (PAH) expressed as tissue slice-to-medium ratio was used as a model to predict nephrotoxicity of different nickel compounds. Pretreatment with nickel compounds for a period of up to 4 h was followed by an incubation with 74 microM PAH for 2 h. The PAH uptake by renal cortical slices was reduced by nickel compounds in both concentration- and time-dependent manner. Furthermore, the extent of such reduction of PAH uptake was found to depend on the chemical form (speciation) of nickel. Thus, slice-to-medium ratios were 41, 49, 48, and 66% of control values after a 4-h pretreatment with 0.378 mM nickel carbonate hydroxide, 1.5 mM nickel subsulfide, 4 mM nickel sulfate and 2.98 mM nickel oxide, respectively. Such an inhibition of PAH transport was not due to cytotoxicity. The results suggest that the nephrotoxic potency decreases in the following order: nickel carbonate hydroxide>nickel subsulfide>nickel sulfate>nickel oxide. Treatment of renal cortical slices with high concentrations of either mannitol, a hydroxyl radical scavenger, or glutathione significantly reduced the inhibition of PAH uptake by soluble form of nickel subsulfide, with concomitant reduction of nickel ion uptake by cortical slices. But no such oxidative stress seems to be involved in nickel carbonate hydroxide-induced inhibition of PAH uptake.

Altered regulation of dopaminergic activity and impairment in motor function in rats after subchronic exposure to styrene.

Animal and human studies suggest a dopamine-mediated effect of styrene neurotoxicity. However, the results reported to date are incomplete and not consistent. As such, the mechanism of its neurotoxicity is still unclear. The present study has, therefore, reexamined the central dopaminergic system in relation to some neurobehavioral effects in rats following subchronic exposure to styrene. Groups of adult male Sprague-Dawley rats received 0, 0.25, or 0.5 g styrene per kg b.wt. by gavage for 13 consecutive weeks. Twenty-four hours after cessation of such treatment with the higher dose (0.5 g/kg), the contents of dopamine (DA) and its metabolites were significantly reduced in the corpus striatum, hypothalamus, and lateral olfactory tract regions. In vitro styrene showed a significant increase in DA release from rat striatal synaptosomes similar to that of tyramine. Significant loss of motor function was observed on days 56, 70, and 84 during the styrene treatment with the higher dose, and lasted over a month after such treatment. However, the treated animals recovered their motor function within 45-60 days after cessation of such treatment, along with the recovery of normal levels of dopamine and its metabolites. Furthermore, styrene-induced initial impairments in measures of dopaminergic activity cannot be attributed to altered regulation of tyrosine hydroxylase activity. Specific [3H]-spiroperidol binding was also unaltered 7 or 15 days after subchronic treatment with styrene. These data imply that despite the dopaminergic neuronal loss due to styrene, dopaminergic transmission was not reduced to a level that would result in an overall development of dopamine receptor supersensitivity in the striatum. Collectively, these studies indicate that the subchronic neurotoxic action of styrene may be primarily presynaptic in nature and may involve impaired regulation of DA content and stimulation of DA release.

Influence of caffeine on allyl alcohol-induced hepatotoxicity in rats. I. In vivo study.

Cotreatment of rats with a low hepatotoxic dose (30.7 mg/kg, i.p.) of allyl alcohol (AA) and a higher, but nontoxic, dose (150 mg/kg, oral) of caffeine (CF) potentiated the hepatotoxicity of AA. This was verified by significantly higher levels of plasma alanine aminotransferase (ALT) activity and histopathologically greater severity of lesions in the periportal hepatocytes than those due to AA alone. Treatment of rats with 4-methylpyrazole (4-MP) (0.5 mmol/kg, i.p.) (an inhibitor liver alcohol dehydrogenase) for 30 minutes, followed by similar cotreatment with AA and CF, completely prevented the elevation of plasma levels of ALT and histological damage induced by cotreatment with CF and AA 24 hours following their administration. Severe liver damage induced by cotreatment with CF and AA was further, markedly enhanced by phenobarbital pretreatment (80 mg/kg, i.p., 3 days). Thus, extensive necrosis of periportal hepatocytes was noted, as well as edema and accumulation of inflammatory cells in the necrotic foci caused by such pretreatment. The depression of hepatic nonprotein sulfhydryls resulting from CF plus AA was much more severe than that caused by AA or CF alone and appeared as early as 30 minutes after administration. However, much less marked depletion of protein thiols was observed following similar treatments. Significant increase in lipid peroxidation (as measured by melondialdehyde [MDA] formation) was also observed in rat liver but only 24 hours after administration. The production ofMDA in the rat liver was significantly higher after administration of AA plus CF than after administration of AA alone. Pretreatment of rats with phenobarbital further significantly enhanced the formation of 2,4-dinitrophenylhydrazine (DNP)-reactive metabolite(s) (measured as DNP-acrolein adduct equivalents) in rat liver induced by AA (30.7 mg/kg) plus CF (150 mg/kg) within 1 hour following such treatment. Cotreatment with AA and a higher dose of CF resulted in significantly higher excretion of urinary thioethers or mercapturic acids than in rats treated with AA alone. Thus, these data suggest that an increased bioactivation pathway of acrolein involving a P450 mixed-function oxidase system caused by CF may be involved in such potentiating effects of CF on AA-induced hepatotoxicity in rats.

Caffeine potentiation of allyl alcohol-induced hepatotoxicity. II. In vitro study.

We examined the effects of caffeine (C) on allyl alcohol (AA)- and acrolein (A)-induced hepatotoxicity on freshly-isolated, rat hepatocytes obtained from livers of adult, male, Sprague-Dawley rats. Isolated rat hepatocytes in suspension were incubated in each test with one of the following: 0, 1.0, or 2.5 mM of AA alone; or with 0, 2.5, or 5 mM of C alone; or a combination of AA and C at the same range of concentrations as used alone, for 15, 30, 60, 90, and 120 minutes at 37 degrees C. A dose- and time-dependent potentiation of cytotoxicity as measured by cellular viability (using trypan blue exclusion) were observed. The AA (2.5 mM)-induced lactate dehydrogenase (LDH) leakage observed after 60 minutes incubation was completely prevented when pretreated for 15 minutes with 4-methylpyrazole (MP) (0.5 mM). Such pretreatment, even with a double dose of 4-MP, only partially, and not significantly, prevented LDH leakage when the hepatocytes were incubated with a mixture of 2.5 mM AA and 5 mM C. The depletion of hepatocyte nonprotein sulfhydryl (NPSH) content caused by AA was further enhanced in the presence of C, as early as 15 minutes after their exposure. The AA-induced increase in lipid peroxidation was also potentiated by C; however, potentiation started later, and only after sufficient depletion of NPSH (mostly glutathione) occurred resulting from the presence of AA plus C. A significant loss of protein sulfhydryls in rat hepatocytes could be noted following a 60-minute incubation period with either AA (1 mM) or AA (1 mM) plus C (5 mM). Similarly, C produced a dose-and time-dependent potentiation of A-induced liver cytotoxicity, which was preceded by severe loss of NPSH content within 15 minutes of exposure, whereas the potentiation of lipid peroxidation (LPO) resulting from A plus C was found to be a relatively late event, as with AA plus C. Furthermore, combined treatment with AA and C produced a significantly higher cytotoxicity (as measured by cellular viability) than that due to the combined treatment with A plus C based on equimolar concentration. These results suggest that two increased bioactivation pathways of AA involving the P-450 mixed-function oxidase system resulting from C may be involved in the potentiation of AA hepatotoxicity.

Modulation of monoamine oxidase activity in different brain regions and platelets following exposure of rats to methylmercury.

Monoamine oxidase (MAO; EC 1.4.3.4) is known to have an important role in the regulation of biogenic amines in the brain and peripheral tissues. It is also known that circulating platelets represent an excellent model for an easy assessment of the effect of MAO-B inhibitors in extracerebral tissue. The present study was carried out to determine the effects of methylmercury (MeHg) on the activity of MAO in synaptosomes of different brain regions of male Sprague-Dawley rats as well as in rat blood platelets both in vitro and in vivo. MeHg pretreatment inhibited the activity of MAO in the synaptosomes of the cortex, hypothalamus, hippocampus, striatum, cerebellum, and brain stem in a concentration-dependent (0-10 microM) manner. The threshold concentration of MeHg for such inhibition in different brain synaptosomes was found to be the same (i.e., 1 microM) except for in the rat striatum it was 2.5 microM, and the IC50 value for MeHg was found to be around 2.1 microM. Significant inhibition of the MAO activity was also observed in synaptosomes of the cortex, cerebellum, hypothalamus, and hippocampus as well as in platelets of rats 24 h after treatment by gavage with a total cumulative dose of 35 mg/kg (5 mg/kg/day for 7 days). The decrease of such activity was found to be at maximum in different brain synaptosomes and platelets 24 h following treatment with a cumulative total dose of 75 mg/kg (7.5 mg/kg/day for 10 days); the treated animals showed signs of ataxia under these conditions. The data have further shown that methylmercury is capable of inhibiting the MAO activity in different brain synaptosomes to different degrees but without showing any specificity towards any specific brain region. The present in vivo results suggest that the platelet MAO activity may be used as a potential biomarker of early neurotoxicity due to repeated exposure to MeHg in rats.