Number of succussion strokes affects effectiveness of ultra-high-diluted arsenic on in vitro wheat germination and polycrystalline structures obtained by droplet evaporation method.

OBJECTIVES: The aim of this study is to investigate whether the number of succussion strokes applied after each dilution step when preparing the homeopathic treatments influences the effectiveness of ultra-high-diluted (UHD) arsenic trioxide at the 45th decimal dilution/dynamization (As2O3 45x). DESIGN: Wheat seeds, previously stressed with ponderal As2O3, were treated with: As2O3 45x, H2O 45x (dynamized control), or pure water (negative control). The succussion was done manually, and various succussion durations (numbers of strokes) were tested for each treatment. Treatment effectiveness was tested blind using the in vitro germination test and the droplet evaporation method (DEM). Data were processed by the Poisson test (germination test) and by two-way analysis of variance (DEM). MAIN OUTCOME MEASURES: We evaluated both the in vitro germination rate, by counting the non-germinated seeds, and the complexity of polycrystalline structures (PCS) (local connected fractal dimension (LCFD)) obtained by evaporating leakage droplets from stressed seeds that had been watered with the different treatments. RESULTS: We observed a highly significant increase in germination rate when the number of strokes (NS) was ≥32 for both As2O3 45x and H2O 45x, and a significant increase in the LCFD of PCS for As2O3 45x when the NS was ≥32 and for H2O 45x when it was 70. CONCLUSIONS: Both experimental approaches showed increased effectiveness for treatments prepared with a higher number of succussion strokes. These results indicate that succussion may have an important influence on treatment effectiveness, and so highlight the need for further research.

The efficacy of ultramolecular aqueous dilutions on a wheat germination model as a function of heat and aging-time.

This study evaluates the effects of temperature and aging on the efficacy of As(2)O(3) at the 45th decimal potency in a wheat germination model, compared against a control and potentized H(2)O 45×. Each treatment-temperature combination was tested on seeds (Triticum aestivum L.) of Pandas variety, using six Petri dishes (33 seeds/dish) per trial, performing eight trials. Seeds were pre-treated by poisoning with 0.1% As(2)O(3) solution to reduce germination, to allow a better evaluation of homeopathic treatment effects. The outcome variable was the number of non-germinated seeds after 96 h. Temperature effect was investigated by heating each treatment in a water bath for 30 min (at 20, 40 or 70°C), or for 5 min (at 100°C), and that of aging by dividing experimental data, collected over a period of nearly five months, into two groups: early and late experiments. Results seem to show that the efficacy of As(2)O(3) 45× is unaltered at 20 and 40°C, increases at 70°C and decreases at 100°C. As regards aging, a notable difference was found between early trials, with no significant efficacy, and late trials, where As(2)O(3) 45× exhibits a repeated significant effect versus control, except at 100°C. A reduction in variability was observed for As(2)O(3) 45× at 20°C versus control, confirming the findings of previous work. The main conclusion suggested by this experiment is that the efficacy of As(2)O(3) 45× on wheat germination may be influenced by heating degree and seems to have an increasing trend as a function of aging.

Self-organized crystallization patterns from evaporating droplets of common wheat grain leakages as a potential tool for quality analysis.

We studied the evaporation-induced pattern formation in droplets of common wheat kernel leakages prepared out of ancient and modern wheat cultivars as a possible tool for wheat quality analysis. The experiments showed that the substances which passed into the water during the soaking of the kernels created crystalline structures with different degrees of complexity while the droplets were evaporating. The forms ranged from spots and simple structures with single ramifications, through dendrites, up to highly organized hexagonal shapes and fractal-like structures. The patterns were observed and photographed using dark field microscopy in small magnifications. The evaluation of the patterns was performed both visually and by means of the fractal dimension analysis. From the results, it can be inferred that the wheat cultivars differed in their pattern-forming capacities. Two of the analyzed wheat cultivars showed poor pattern formation, whereas another two created well-formed and complex patterns. Additionally, the wheat cultivars were analyzed for their vigor by means of the germination test and measurement of the electrical conductivity of the grain leakages. The results showed that the more vigorous cultivars also created more complex patterns, whereas the weaker cultivars created predominantly poor forms. This observation suggests a correlation between the wheat seed quality and droplet evaporation patterns.

A review of three simple plant models and corresponding statistical tools for basic research in homeopathy.

In this paper, we review three simple plant models (wheat seed germination, wheat seedling growth, and infected tobacco plants) that we set up during a series of experiments carried out from 1991 to 2009 in order to study the effects of homeopathic treatments. We will also describe the set of statistical tools applied in the different models. The homeopathic treatment used in our experiments was arsenic trioxide (As2O3) diluted in a decimal scale and dynamized. Since the most significant results were achieved with the 45th decimal potency, both for As2O3 (As 45x) and water (W 45x), we here report a brief summary of these results. The statistical analysis was performed by using parametric and nonparametric tests, and Poisson distribution had an essential role when dealing with germination experiments. Finally, we will describe some results related to the changes in variability, which seems to be one of the targets of homeopathic treatment effect.

A biostatistical insight into the As(2)O(3) high dilution effects on the rate and variability of wheat seedling growth.

BACKGROUND: Most criticism of homeopathy concerns the lack of scientific bases and theoretical models. Fundamental research could make important contributions to our understanding of the mechanisms of action of homeopathic treatments. Plant-based bioassays are suitable for basic research -- lacking the placebo effect and ensuring large data samples for structured statistical analyses. OBJECTIVE: The aim of this study was to reproduce a previous experiment on the effects of arsenic trioxide (As(2)O(3)) high dilutions on wheat seedling growth in order to verify whether the same significant results could be obtained working in a different place and with a different experimental team. A further goal was to investigate high dilution effects on variability. MATERIAL AND METHODS: A structured experiment was performed blind over 9 weeks, using wheat seeds previously stressed with a sublethal dose of As(2)O(3). The seeds were then treated with either potentized As(2)O(3) (5x, 15x, 25x, 35x, 45x), potentized water (equivalent potencies) or diluted As(2)O(3) (10(-5), 10(-15), 10(-25), 10(-35), 10(-45)). The working variable was the stem length, measured after 4, 5, 6 and 7 days. RESULTS: Some potencies (As(2)O(3) 45x and H(2)O 45x) induced a relevant increase in seedling growth and/or a variability decrease. Diluted As(2)O(3) did not induce any significant results. CONCLUSIONS: Confirmation of a significant stimulating effect on seedling growth and a significant decrease of variability was obtained with ultra-high dilutions at the 45x potency. The model of wheat germination and growth has been confirmed to be a good tool for basic research in homeopathy.

Nonthermal microwave radiations affect the hypersensitive response of tobacco to tobacco mosaic virus.

OBJECTIVES: The aim of the present study was to evaluate the effects of nonthermal extremely high-frequency microwave radiations in a plant-based bioassay, represented by tobacco plants reacting to tobacco mosaic virus with a hypersensitive response leading to the appearance of necrotic lesions at the infection sites. DESIGN: This study was performed blind and different experimental protocols on tobacco plants inoculated with tobacco mosaic virus were used. BIO-OBJECTS: Tobacco plants (Nicotiana tabacum L. cultivar Samsun) carrying the resistance gene N against tobacco mosaic virus. INTERVENTIONS: Tobacco plants or leaf disks were either directly or indirectly (water-mediated) irradiated using a medical device, designed for microwave resonance therapy. It produces nonthermal weak-intensity extremely high-frequency radiations, either modulated at extremely low frequency or in continuous flux of waves, coupled with a nonthermal red/near-infrared radiation. OUTCOME MEASUREMENTS: The working variable was the number of hypersensitive lesions per leaf disk. RESULTS: Both direct and indirect nonthermal microwave radiations led to significant effects on the hypersensitive response of tobacco plants: modulated radiations generally induced a resistance increase, whereas a continuous flux of waves induced a resistance decrease with direct treatments only. CONCLUSIONS: Nonthermal microwave radiations are effective on the hypersensitive response of tobacco to tobacco mosaic virus and their low-frequency modulation seems to be more bioactive than the continuous-flux of waves, particularly in the indirect water-mediated treatments.

Transcriptome Profiling of Wheat Seedlings following Treatment with Ultrahigh Diluted Arsenic Trioxide.

Plant systems are useful research tools to address basic questions in homeopathy as they make it possible to overcome some of the drawbacks encountered in clinical trials (placebo effect, ethical issues, duration of the experiment, and high costs). The objective of the present study was to test the hypothesis whether 7-day-old wheat seedlings, grown from seeds either poisoned with a sublethal dose of As2O3 or unpoisoned, showed different significant gene expression profiles after the application of ultrahigh diluted As2O3 (beyond Avogadro's limit) compared to water (control). The results provided evidence for a strong gene modulating effect of ultrahigh diluted As2O3 in seedlings grown from poisoned seeds: a massive reduction of gene expression levels to values comparable to those of the control group was observed for several functional classes of genes. A plausible hypothesis is that ultrahigh diluted As2O3 treatment induced a reequilibration of those genes that were upregulated during the oxidative stress by bringing the expression levels closer to the basal levels normally occurring in the control plants.

Droplet evaporation method as a new potential approach for highlighting the effectiveness of ultra high dilutions.

OBJECTIVE: This study sought to verify whether the droplet evaporation method (DEM) can be applied to assess the effectiveness of ultra-high dilutions (UHDs). We studied the shape characteristics of the polycrystalline structures formed during droplet evaporation of wheat seed leakages. METHODS: The experimental protocol tested both unstressed seeds and seeds stressed with arsenic trioxide 5mM, treated with either ultra-high dilutions of the same stressor substance, or with water as a control. The experimental groups were analyzed by DEM and in vitro growth tests. DEM patterns were evaluated for their local connected fractal dimension (measure of complexity) and fluctuating asymmetry (measure of symmetry exactness). RESULTS: Treatment with arsenic at UHD of both stressed and non-stressed seeds increased the local connected fractal dimension levels and bilateral symmetry exactness values in the polycrystalline structures, as compared to the water treatment. The results of in vitro growth tests revealed a stimulating effect of arsenic at UHD vs. control, and a correlation between the changes in growth rate and the crystallographic values of the polycrystalline structures was observed. CONCLUSIONS: The results indicate that polycrystalline structures are sensitive to UHDs, and so for the first time provide grounds for the use of DEM as a new tool for testing UHD effectiveness. DEM could find application as a treatment pre-selection tool, or to monitor sample conditions during treatment. Moreover, when applied to biological liquids (such as saliva, blood, blood serum, etc.), DEM might provide information about UHD effectiveness on human and animal health.

The role of variability in evaluating ultra high dilution effects: considerations based on plant model experiments.

A series of experiments, performed on plant models with ultra high dilutions (UHD) of arsenic trioxide at 45th decimal potency has been reviewed with a particular focus on variability. The working variables considered are: the number of germinated seeds out of a fixed set of 33, the stem length of wheat seedlings and the number of necrotic lesions in tobacco leaf disks inoculated with tobacco mosaic virus (TMV). A thorough comparison between treatment and control group has been proposed, considering the two main sources of variability in each series of experiments: variability within and between experiments. In treated groups, a systematic decrease in variability between-experiments, as well as a general decrease, with very few exceptions, in variability within experiments has been observed with respect to control. Variability is traditionally considered as control parameter of model systems. Our hypothesis, based on experimental evidences, proposes a new role of variability as a target of UHD action. This hypothesis may help interpret unanswered questions that keep rising in basic and clinical research in homeopathy.