A quantitative PCR assay for antiviral activity screening of medicinal plants against Herpes simplex 1.

Herpes simplex virus one is one of the most prevalent pathogens worldwide. Strains resistant to current treatment have been reported, so it is necessary to search for new antiviral molecules. The most common method to quantify antiviral activity from natural products is the plaque reduction assay, a technically demanding method. In order to provide a simple alternative to this method, we have established a procedure for viral quantification by qPCR, and coupled with a cytotoxicity evaluation system using resazurin. In this way, it is possible to obtain both the estimation of cytotoxicity and the antiviral activity simultaneously, allowing rapid screening of plant extracts. Ten out of twenty-eight Paraguayan medicinal plant extracts evaluated using this method showed antiviral activity, and the EC50, CC50, and SI values were calculated for each extract. Our experience supports the employment of the described method for a rapid identification of plant extracts with antiviral activity.

MRI and EEG findings in surgically treated patients with partial seizures due to neuronal migration disorders, their relations to each other and to surgery outcome.

OBJECTIVES: To evaluate magnetic resonance imaging (MRI) and electro-encephalogram (EEG) findings of patients with therapy-resistant partial seizures due to neuronal migration disorders (NMD), and compare them with each other and with surgery outcome. MATERIAL AND METHODS: The MRI, interictal and ictal EEG findings, and their relations to each other in 41 patients with NMD were compared with class IA surgery outcome. RESULTS: The patients showed an MRI lesion in decreasing frequency from the frontal to the occipital areas. A predominantly extratemporal location of the interictal EEG foci (73.3%) and ictal patterns (82.4%) was therefore apparent, also showing a diminishing frequency from the anterior to the posterior areas. Comparing the EEG foci with the MRI lesions, the same location of the interictal foci was found in 68.4% and of the ictal foci in 52.7%, including several cases with a more widespread EEG focus or MRI lesion. The same location of interictal as well as ictal foci was evident in 85.7%. The most favourable surgery outcome (class IA) was, on average, apparent in patients with an MRI lesion (28.6% vs 25%), a main interictal EEG focus (50% vs 18%) and an ictal seizure pattern (37.5% vs 16.7%) located in the temporal area; likewise, if the interictal focus (30.8% vs 18.2%) and the ictal pattern (31.8% vs 12.5%) showed the same location as the MRI lesion, as well as when the interictal and ictal EEG changes had an equal location (30% vs 20%). CONCLUSION: MRI and EEG recordings show relatively close relations to each other and might be important methods to predict surgery outcome in NMD patients.

Plasmodium falciparum: cloned and expressed CIDR domains of PfEMP1 bind to chondroitin sulfate A.

Adherence of erythrocytes infected with mature asexual Plasmodium falciparum parasites (iRBC) to microvascular endothelial cells contributes to the pathology of P. falciparum malaria. It has been shown that the variant P. falciparum erythrocyte membrane protein 1 (PfEMP1) confers adhesion to a wide range of cell surface receptors. Previously, the cysteine-rich interdomain region (CIDR) of PfEMP1 has been identified as binding site to CD36. We provide evidence that the same region can also mediate binding to chondroitin sulfate A (CSA). CIDR domains of two different parasite strains were expressed in Escherichia coli as a 6xHis-tagged protein. Purified recombinant protein bound to Chinese hamster ovary (CHO) cells which naturally express chondroitin sulfate A. Treatment of wild-type CHO cells with chondroitinase ABC reduced binding up to 94.4%. Competitive binding using soluble CSA inhibited binding to CHO cells by up to 100% at 2 mg/ml and by 62.4% at 0.5 mg/ml, whereas 1 mg/ml heparan sulfate had only a little effect (18.1%). In contrast, a recombinant 6xHis-tagged DBL1 domain showed no binding to wild-type CHO cells. Such an approach of analyzing various domains of PfEMP1 as recombinant proteins may elucidate their functions and may lead to novel anti-adherence therapeutics, especially for maternal malaria infections.

Benign epileptic discharges in patients with lesional partial epilepsies.

Case reports of four patients with therapy-resistant lesional partial epilepsies and additional foci of benign epileptic discharges of childhood, in addition to the usual electroencephalogram (EEG) changes, are presented. A family history of epileptic or febrile seizures in childhood was reported in all four patients. A distant relative of one patient, not manifesting seizures, demonstrated rolandic spikes on EEG. An abnormal pregnancy (polyhydramnion, premature pains, induced labor because of an abnormal CTG , placenta insufficiency) was reported in one patient, risk factors during birth (birth 14 days after term, placenta insufficiency) were reported in one, and bacterial meningitis at 4 weeks of age was reported in one. All patients manifested a retarded, partly severe, unfavorable infantile psychomotor development. An early seizure onset was observed in all patients (in three patients during the first year of life and in one patient during the second year). A hemifacial seizure symptomatology was seen, in addition to other symptoms, in two patients, possibly indicating the seizure pattern indicative of benign partial seizures; seizures occurred exclusively in sleep in one patient. The benign focus was never located in the lesional area. It was recorded over the same hemisphere in two patients and over the other hemisphere in the other two.

The genetics of localization-related symptomatic epilepsy: risk of a family history with seizures in patients who have undergone surgery.

A family history of epileptic seizures including febrile convulsions was found in 15 of 103 patients (15%) with localization related epilepsy with partial seizures with and without secondary generalization, who were operated on because of drug resistance. This rate was significantly higher than that of the cumulative incidence in the general population (4%). The localization of the brain damage did not play a role (temporal lobe resection left: 15%, right: 17%, extra-temporal lesion excision: 20%, hemispherectomy: 11%). Various family members were involved. Some patients had more than one relative with seizures. Thus, 21 relatives suffered from seizures. Eleven of them had generalized tonic-clonic seizures (one grand mal on awakening), 7 had febrile convulsions (4 complicated), and in 1 patient the grand mal seizures on awakening were preceded by absences; 1 had generalized tonic-clonic and complex partial seizures; 1 after complicated febrile seizures likewise had complex partial seizures; another mentally retarded patient suffered from generalized tonic-clonic, axial tonic and myoclonic-astatic seizures. The seizure type of 3 remote relatives was not known. The first seizure occurred in 16 family members during childhood, in 3 in adolescence and in only 1 in adulthood (1 unknown). Eight showed mental retardation of slight degree in most. It is interesting that only one-third of the patients with a family history with seizures were seizure-free after the operation; 5 still had seizures, mostly reduced in frequency, 3 had seizures and isolated auras and 2 had only isolated auras. On comparing the findings in patients with and without a family history with seizures, those with family members with epileptic seizures showed a lower rate of an intellectual deficit (7 vs 47%) and brain tumours (13 vs 44%). Our earlier findings with a different group of patients are thus confirmed: that genetics play a role in symptomatic epilepsies.

Contribution to the genetics of symptomatic generalized tonic-clonic seizures: waking and sleep EEGs in siblings.

The presence of epileptiform activity (EA) in the EEG of patients' relatives points to the significance of genetics in the etiology of epilepsies. Waking and sleep EEGs were recorded in 83 siblings of 54 patients suffering from symptomatic generalized tonic-clonic seizures. EA was recorded in at least one sibling of 27 (50%) of the 54 patients. When the 83 siblings are taken as a basis, EA was found in 34 (41%) of them. Generalized spike-wave discharges were seen in 32 cases; 2 siblings showed benign sharp wave foci in the right parietal area. EA was seen only in sleep in 44.1%. The highest rates of EA were seen in the age range up to 15 years. EA was found in 15 of 50 male siblings (30%), but in 18 of 33 female siblings (54.5%). Therefore genetics also play an import role in the etiology of symptomatic generalized tonic-clonic seizures.

Sexual compatibility among Trypanosoma brucei isolates from an epidemic area in southeastern Uganda.

For the first time, two clones of Trypanosoma brucei from the same epidemic area in southeastern Uganda were successfully crossed. The cotransmission experiments were as close to natural conditions as possible in that two uncloned isolates from wild-caught Glossina fuscipes fuscipes were directly cotransmitted in an initial recombination experiment. From the first uncloned progeny population, which revealed a majority of recombinants, two clones with different parental phenotypes [characterized by isocitrate dehydrogenase (ICD) analysis] were isolated. These clones were cotransmitted in a second recombination experiment. Nine clones could be isolated from two different progeny populations of the second experiment, and all showed a recombinant phenotype. These nine clones belonged to three different karyotypes with respect to the large chromosomes (1-3 Mb), which were different from those of either parental karyotype or the sum of both parental karyotypes. The results indicate that genetic recombination might well occur between trypanosome populations transmitted within the same epidemic area.

[Genetic aspects of symptomatic epilepsy based on waking and sleep EEG recordings in siblings]. / Zur Genetik der symptomatischen Epilepsien aufgrund von Wach- und Schlaf-EEG-Ableitungen bei Geschwistern.

Waking and sleep EEG-recordings were carried out in siblings of patients with various idiopathic and symptomatic seizure types. Rates of epileptic activity (e.a.) were found in the symptomatic ones varying between 24.1% (Complex partial seizures) and 46.7% (Symptomatic absences). 1/4 to 1/2 of the e.a. was recorded exclusively in sleep, so that sleep recordings are also necessary for such investigations. 2.5-4/sec. spike wave-complexes were predominantly seen; benign foci and photosensitivity were recorded in a smaller number of siblings. More e.a. was observed in idiopathic (72%) than in symptomatic absences (46.7%). On the other hand the same rates (42:41%) as well as almost the same EEG-patterns were found in idiopathic and symptomatic generalized tonic-clonic seizures. When counting the single epileptic discharges more e.a. was seen in siblings of patients with the idiopathic type than the symptomatic one (one discharge every 53 sec.:229.3 sec.). Most e.a. was found in the age group 6-14 years in siblings of all seizure types; therefore, this age dependent penetrance does not depend on the seizure type, but on the recorded spike wave-complexes, benign foci and photosensitivity which occur most frequently in this age range. A multi-factorial mode of inheritance is assumed.

On the genetics of complex partial seizures: waking and sleep EEGs in siblings.

Waking and sleep EEGs were recorded in 29 siblings of 19 patients with complex partial seizures. At least 1 sibling with epileptic activity (EA) was found for 36.8% of the patients. Taking the 29 siblings as a basis, in 7 EA was recorded. Most EA was seen during sleep in stage C (29%). More EA was recorded in female siblings (28%: 18%) and in siblings of female patients (56%: 20%). All EA was seen in the age range 5-14 years. Siblings with occipital theta-delta activity with a generalization tendency showed more EA (59%) than those without this pattern (8%). Of the siblings of patients with generalized EA 50% showed EA, but only 25% of those of patients with localized EEG patterns.

Contribution to the genetics of rolandic epilepsy: waking and sleep EEGs in siblings.

Waking and sleep EEGs were recorded in 69 siblings of 43 patients with rolandic spikes. 36 suffered from rolandic epilepsy, 7 from other diseases or symptoms (headaches, migraine, learning problems). At least one sibling with epileptic activity was found in 51.16% of the patients. Taking the 69 siblings as a basis, in 26 (37.68%) epileptic activity was recorded. Benign spike foci were recorded in only 4 siblings, generalized spike-wave complexes were seen in 22. Most epileptic activity was recorded in the age group of 5-12 years (54.3%). Nearly one-half (17.4%) was recorded exclusively in sleep, predominantly in sleep stage C (88%). Siblings of patients with (40%) and without seizures (37.5%) showed approximately the same rate, likewise siblings with (40%) and without seizures (34.8%). An autosomal-dominant mode of inheritance is assumed, but a multifactorial mode is also discussed.

A contribution to the genetics of febrile seizures: waking and sleep EEG in siblings.

Waking and sleep EEGs were recorded in 67 siblings of 52 patients with febrile seizures (FS). Epileptic activity was noted in at least 1 sibling for 28 of the 52 patients (53.8%). Epileptic discharges were noted in 33 (49.2%) of the 67 siblings. Thirty-two siblings had 3-4 Hz spike wave complexes, and 1 sibling had independent centrotemporal spike foci (rolandic foci). Epileptic activity was noted exclusively in the waking state in only 3%, in waking and sleep in 31.3%, and only in sleep in 14.9%. The greatest number of epileptic discharges occurred in waking during hyperventatilation (32.8%) and during stage C sleep (38.8%). In 5 photosensitive siblings, additional epileptic discharges were noted in sleep in 1 and in waking and sleep in 4. At least 1 sibling in 5 (55.6%) of 9 patients with complicated febrile seizures, in 23 (53.5%) of 43 patients with simple FS, and in 4 of 9 patients (44.4%) with later onset epileptic seizures had seizure discharges. At least 1 sibling in 24 of 43 patients (55.8%) with exclusively FS in 13 of 30 (43.3%) male patients and in 15 of 22 (68.2%) female patients had seizure discharges. Siblings aged 6-10 years had (66.7%) the highest rates of activation. Epileptic discharges were noted in 83.3% of siblings with seizures, but in only 45.9% of siblings without seizures. Seizure activity was recorded in 68.2% of siblings who had occipital 3-4-Hz theta-delta-activity but in only 13.0% of siblings without this pattern.(ABSTRACT TRUNCATED AT 250 WORDS)

Some genetic aspects of idiopathic and symptomatic absence seizures: waking and sleep EEGs in siblings.

Epileptic activity was recorded in the waking and sleep EEG of 62.5% of 80 siblings of 38 patients with absence seizures. Epileptic discharges were noted in waking only in 8.7%, in waking as well as sleep in 28.8%, and in sleep only in 25%. Generalized, partly irregular, and slow spike-wave complexes were found, twice with lateral emphasis. Spike-wave complexes were recorded in 72% of 50 siblings of patients with idiopathic absence and in 46.7% of 30 siblings of patients with symptomatic absence. One epileptic discharge was observed every 108.6 s on the average, without striking differences between siblings of patients with idiopathic (99.7 s) and symptomatic absence (119.3 s). Without any differences between siblings of children with idiopathic and symptomatic absence, the most epileptic discharges were activated in sleep stages C and D, followed by stages A and B. The highest activation rate was observed in the 7-14-year-old group (73.5%) and to a somewhat lesser degree in the group between 15 and 20 years of age (66.7%); fewer epileptic discharges were recorded in younger (25%) and older patients (28.6%). The higher activation rates in the male sex were significant only in siblings of patients with idiopathic absence. Although only five patients (13.2%) were photosensitive, a photosensitivity was found in 24% of siblings of children with idiopathic absence and in 20% of siblings of patients with symptomatic absence. Three siblings of patients with idiopathic absence also had absence seizures; in one of them a febrile seizure occurred at an earlier age. All of them showed generalized spike wave discharges in waking as well as sleep. Occipital theta delta activity with generalization was observed more frequently in siblings of patients with idiopathic absence (82.2%) than in those of patients with symptomatic absence (63.6%). Our waking and sleep EEG recordings prove that concerning etiology-genetic factors play a striking role in idiopathic absence, but are also of considerable significance in the symptomatic types.

Some genetic aspects of rolandic epilepsy: waking and sleep EEGs in siblings.

Epileptic activity was recorded electroencephalographically in at least one sibling in 22 (51.16%) of 43 patients with rolandic epilepsy and/or centrotemporal spikes. In 26 of 69 (37.68%) siblings, epileptic discharges were observed. These were recorded only in waking in 1 subject (1.5%), in waking and sleep in 13 (18.8%), and in sleep only in 12 (17.4%). The greatest number of epileptic discharges was noted in waking during hyperventilation (52.4%) and in sleep stage C (88%). Foci were recorded in only 4 (5.8%) of the 26 cases with epileptic discharges, and generalized spike-wave complexes were recorded in 22 (31.9%). In one sibling, the sharp-wave focus was located in the right centrotemporal area, in a second in the left occipital, and in a third in the left frontal region with spreading to the centrotemporal; in the fourth, two independent foci were observed in the left and right centrotemporal area. One epileptic discharge was observed every 74.9 s in waking and every 150.9 s in sleep. Epileptic activity was greatest in the group between 5 and 12 years of age (54.3%). The same activation rates were noted in siblings of patients with (47.2%) and without seizures (42.9%), and no differences were noted in siblings with (40%) or without (37.5%) seizures. Family history and sex of the siblings of patient did not play a role in the rate of activation. An autosomal dominant inheritance is assumed, but further investigations are necessary.

The development of independent foci in epileptic patients.

The electroencephalograms of 309 unselected epileptic patients were reexamined to ascertain the incidence of independent secondary discharges. In 41 patients (13%), a simple one-sided focus, without evidence of independent secondary discharges, was found (group 1); in 33 patients (11%), an epileptic focus with certain evidence of independent secondary discharges was found (group 2). These two groups were compared with respect to multiple variables (eg, seizure type, time course of the epileptic illness, site of foci, medication, neurological status), and significant differences between the two groups were obtained (eg, with respect to frequency of seizures, duration of seizures, number of anticonvulsive drugs being taken, and in other respects). Our findings are compared with earlier work, and it is concluded that they do not support the conventional models of secondary epileptogenesis.

Sleep EEG with or without sleep deprivation? Does sleep deprivation activate more epileptic activity in patients suffering from different types of epilepsy?

A sleep EEG of 190 patients without sleep deprivation was recorded, followed by a sleep EEG after 24 h of sleep deprivation on the next day. The patients suffered from various types of epilepsy, in their routine EEGs no epileptic discharges were seen. Both sleep EEGs were recorded under the same antiepileptic drugs. A waking EEG was recorded immediately before each sleep EEG. The activation rates of epileptic activity in 52.6% (without sleep deprivation) and 53.2% (with sleep deprivation) of the patients showed no significant differences. Also on classifying the epileptic discharges no real difference was found between the 2 methods (generalized: 29.5 vs. 29.5%, generalized with lateral emphasis: 11.1 vs. 9.5%, focal: 12.1 vs. 14.2%). Only in the waking EEG, recorded immediately before the sleep EEG after sleep deprivation, a few more patients showed epileptic discharges (33.6 vs. 27.4%). Without there being any significant differences between the 2 methods there were some different results in comparing the EEG with the clinical findings: significantly more epileptic activity was shown in patients who had their first seizure before the age of 20 (55.6 and 55.6% vs. 26.3 and 31.6%), amongst females (59.8 and 61.9% vs. 45.2 and 44.1%), in awakening grand mal (= primary generalized tonic-clonic seizures, 76.5 and 70%) and in absences (69 and 72.4%). The higher activation rates in young subjects, in patients with a family history of seizures, with pathological neurological findings, mental retardation and delayed psychomotoric development in early childhood, were not statistically significant.(ABSTRACT TRUNCATED AT 250 WORDS)

The activating effect of sleep EEGs in epileptic patients with epileptic activity in their waking EEGs.

A sleep EEG was recorded in 44 patients whose waking EEG showed epileptic activity. The time in seconds in which one epileptic discharge occurred was determined. Considering the waking and sleep EEGs together one epileptic discharge occurred every 7.8 seconds. Focal epileptic activity was recorded nearly twice as frequently (1 paroxysm every 4.6 sec.), as generalized activity (1 paroxysm every 9.8 sec.) or generalized activity with lateral emphasis (1 paroxysm every 8.8 sec.). Considering the sleep and waking EEGs separately it was found that on average more epileptic activity occurred in sleep than in the waking EEG (6.4 vs. 10.8 sec.). There was no difference between generalized (8.9 vs. 13.5 sec.), generalized activity with lateral emphasis (7.7 vs. 12.9 sec.) and focal activity (3.9 vs. 7.6 sec.). Comparing, however, the frequency of epileptic activity in each individual patient during wakefulness and sleep it was seen that during sleep more activity was found in only 56.8% of the patients. If a difference of 3 seconds is considered to be due to chance, then more epileptic activity was found in 34.1% of the patients during sleep and in 24.5% during wakefulness. Thus, the sleep EEG by no means always activates epileptic activity.