Search

Introduction:  In 2021, the area of CRAS (Centro de Reabilitação de Animais Selvagens - Wild Animal Rehabilitation Center), located in a state park in Campo Grande City, Mato Grosso do Sul, Brazil, was suffering from a tick infestation affecting wild animals that inhabit the area and humans that visited its trails. Following a formal technical-scientific cooperation agreement between IMASUL, an institute for the environment in Mato Grosso do Sul state, and SIGO Homeopatia, a formulated homeopathic complex ( Formula Parques Urbanos , FPU) was designed and prepared specifically to treat the animals. This environmental intervention used specially designed slow-release water biodegradable devices. Tracking the FPU signal in water was necessary to monitor and manage the intervention., Aims:  Our aims were (1) to evaluate, among six previously standardized solvatochromic dyes, which would serve as a marker for the homeopathic complex under study; and (2) to evaluate whether the chosen solvatochromic dye could map the propagation of the homeopathic complex activity throughout the stream system from water samples harvested at different locations over time., Method:  Water samples were harvested from each point at different times, filtered, frozen, and sent to the laboratory, where they were prepared at 1cH potency for analysis using 30% ethanol as the vehicle. Solvatochromic dyes were used to analyze the samples since they alter their absorbance when in contact with homeopathic potencies. Of the six dyes tested, Coumarin 7 was found to be the most suitable for tracking the FPU complex. A static and average unidirectional magnetic field of 2,400 Gauss (240 mT), generated by a neodymium magnet, was applied to the samples immediately before reading., Results:  There were significant differences in the delta absorbance of dyes when adding treated/potentized water samples, making it possible to map the propagation of the FPU signal throughout the park over time. The signals were identifiable at the same point 1 minute and 32 days after the insertion of the device into the water. These signals were also identifiable after 75 minutes and 8 days at a point far from the insertion place., Conclusion:  Coumarin 7 was the best marker for the homeopathic complex (FPU) used to treat the wild animals living in the park. The microplates/enzyme-linked immunosorbent assay reader method and the application of a magnetic field to samples were shown to be effective in tracing homeopathic signals by changes in dye absorbance ( p ≤ 0.02) in a real-life situation, with large volumes of water, involving many environmental variables, and over large distances., Competing Interests: The samples were made available for analysis by SIGO Homeopatia company in partnership with the governmental organization IMASUL (Instituto do Meio Ambiente do Mato Grosso do Sul). No other resource was given to the researchers., (Faculty of Homeopathy. This article is published by Thieme.)

Background: Whilst it is assumed that water plays a crucial role in homeopathy, it is unclear what specific functions water has in the generation, propagation and maintenance of homeopathic potencies. Solvatochromic dyes have demonstrated themselves to be reliable indicators of the presence of potencies and so are well placed to begin to address these questions., Objective: The aim of the current study was to ascertain whether pure water could be potentised. If pure water can be potentised, this would indicate that water plays a crucial role in the generation of homeopathic potencies. If it cannot be potentised, then this result points to water being an inert carrier and medium for the potentisation of substances placed in it., Methods: A specially designed potentiser capable of consistently and reliably producing homeopathic potencies to rigorous standards (Pinkus Potentiser) was used to run up potencies of "Water 10M" and Arsenicum 10M in two interchangeable stainless-steel vials. "Water 10M" and Arsenicum 10M, along with unsuccussed control water, were then tested against solutions of the solvatochromic dye phenol blue by following the absorbance of the dye at 658 nm. RESULTS ": Water 10M" gave results no different statistically from unsuccussed water ( p  > 0.05) with no change over the time course of the experiments, whereas Arsenicum 10M gave statistically different results ( p  < 0.001) with increasing divergence from both "Water 10M" and unsuccussed water over time. Similar results were obtained when the vials in which Arsenicum 10M and "Water 10M" were produced were exchanged, so that the vial in which Arsenicum 10M had been produced was then used to produce "Water 10M" and vice versa, indicating the results could not be explained by any anomalies in the production vials themselves., Conclusion: Results using the solvatochromic dye phenol blue to detect and measure homeopathic potencies indicate that bulk water itself does not potentise and that unsuccussed water and water that has been succussed are no different. This indicates that what is potentised during cycles of dilution and succussion are the substances placed in water and not the water itself. In turn, this indicates that water is simply a carrier for the homeopathic signal. Evidence from the practice of homeopathy supports this conclusion., Competing Interests: None declared. Tony Pinkus holds the position of Superintendent Pharmacist & Technical Director of Ainsworth Homeopathic Pharmacy. He also owns Ainsworth Pharmacy and designed the Pinkus Potentiser. However, all experimental work and assays were executed independently of Ainsworths pharmacy., (Faculty of Homeopathy. This article is published by Thieme.)

Introduction:  Cyanobacteria are microorganisms found in many parts of the world and several genera, such as Raphidiopsis raciborskii , are producers of cyanotoxins. Homeopathic potencies have been found to modulate toxicity in different biological models, and the present study endeavors to discover whether this might also be the case with cyanobacteria., Objectives:  Our objective was to investigate the possible effects of homeopathic potencies on the resilience of Artemia franciscana (brine shrimp) embryos to saxitoxin (STX; cyanotoxin) and on controlling the growth of R. raciborskii in vitro ., Method:   A. franciscana cysts were cultivated in seawater in 96-well plates to evaluate the hatching rate and vitality, plus the gene expression of heat shock proteins (HSPs), after being challenged with R. raciborskii extract containing 2.5 µg/L of STX and treated with different homeopathic potencies. Untreated wells were used as controls ("base-line"). Potencies were chosen from a screening process based on seven selected homeopathic preparations according to the similitude of STX symptoms ( Sulphur , Zincum metallicum , Nitric acidum , Plumbum metallicum , Mercurius solubilis , Phosphoric acidum , Isotherapic from R. raciborskii extract; all at 6cH, 30cH and 200cH). Cultures of R. raciborskii maintained in an artificial seawater medium were equally treated with screened homeopathic potencies selected from the same list but specifically for their growth control as a function of time., Results:  A 15% lower rate of hatching of A. franciscana cysts was observed after treatment with Nitric acidum 6cH in comparison with baseline ( p  = 0.05). A complete toxicity reversal was seen after treatment with Isotherapic 200cH, with a 23-fold increase of Hsp 26 gene expression ( p  = 0.023) and a 24-fold increase of p26 gene expression ( p ≤ 0.001) in relation to baseline. Nitric acidum 200cH and Mercurius solubilis 30cH limited the exponential growth of cyanobacteria up to 95% and 85% respectively ( p ≤ 0.003) in relation to baseline. Succussed water presented only a transitory 50% inhibition effect., Conclusion:  Isotherapic 200cH improved A. franciscana bioresilience to STX; Nitric acidum 200cH and Mercurius solubilis 30cH showed the optimal performance on limiting R. raciborskii growth. The results point to the potential of homeopathic potencies to mitigate environmental problems related to water quality., Competing Interests: None declared., (Faculty of Homeopathy. This article is published by Thieme.)

Introduction:  The control of cyanobacterial toxicity and growth by homeopathic potencies was described in Part 1 of this two-part report. Here, a parallel approach characterized the physico-chemical features of the potencies used and the liquid media treated with them, correlating these results with their respective biological effects., Objectives:  Our objective was to establish if physico-chemical parameters can track homeopathic potencies in seawater or artificial seawater medium (ASM)-1 and to discover whether these parameters correlate with previously described biological effects., Method:   Artemia franciscana (brine shrimp) cysts were cultivated in seawater challenged with Raphidiopsis raciborskii extract and treated with different homeopathic potencies chosen from a screening process. Cultures of R. raciborskii maintained in ASM-1 were also treated with previously screened homeopathic potencies, and their growth was monitored as a function of time. The physico-chemical properties of the treated media (seawater or ASM-1) were evaluated by their interaction with solvatochromic dyes and changes in pH, conductivity and temperature., Results:  Coumarin 7 was found to be a marker for Nitric acidum 6cH and Isotherapic ( R. raciborskii extract) 200cH in seawater (analysis of variance [ANOVA], p  = 0.0015). Nile red was found to be a marker for Nitric acidum 200cH and Mercurius solubilis 30cH in ASM-1 (ANOVA, p ≤ 0.001). An increase in pH of ASM-1 and endothermic effects were observed after these treatments (two-way ANOVA, p  = 0.0001). Seawater and ASM-1 to which potencies had been added were also subjected to a constant unidirectional 2,400 Gauss static magnetic field and found to have enhanced effects on the solvatochromic dyes tested., Conclusion:  Homeopathic potencies were specifically traceable in aqueous media using solvatochromic dyes, especially when the samples were subjected to a magnetic field. Results from monitoring other physical parameters, such as pH and temperature, were less specific in relation to potency tracking. However, potency-induced endothermic effects might provide valuable thermodynamic data relating to the nature of potencies., Competing Interests: None declared., (Faculty of Homeopathy. This article is published by Thieme.)

Glyphosate is an herbicide widely used in agriculture but can present chronic toxicity in low concentrations. Artemia salina is a common bio-indicator of ecotoxicity; it was used herein as a model to evaluate the effect of highly diluted-succussed glyphosate (potentized glyphosate) in glyphosate-based herbicide (GBH) exposed living systems. Artemia salina cysts were kept in artificial seawater with 0.02% glyphosate (corresponding to 10% lethal concentration or LC10) under constant oxygenation, luminosity, and controlled temperature, to promote hatching in 48 h. Cysts were treated with 1% ( v / v ) potentized glyphosate in different dilution levels (Gly 6 cH, 30 cH, 200 cH) prepared the day before according to homeopathic techniques, using GBH from the same batch. Controls were unchallenged cysts, and cysts treated with succussed water or potentized vehicle. After 48 h, the number of born nauplii per 100 µL, nauplii vitality, and morphology were evaluated. The remaining seawater was used for physicochemical analyses using solvatochromic dyes. In a second set of experiments, Gly 6 cH treated cysts were observed under different degrees of salinity (50 to 100% seawater) and GBH concentrations (zero to LC 50); hatching and nauplii activity were recorded and analyzed using the ImageJ 1.52, plug-in Trackmate. The treatments were performed blind, and the codes were revealed after statistical analysis. Gly 6 cH increased nauplii vitality ( p = 0.01) and improved the healthy/defective nauplii ratio ( p = 0.005) but delayed hatching ( p = 0.02). Overall, these results suggest Gly 6cH treatment promotes the emergence of the more GBH-resistant phenotype in the nauplii population. Also, Gly 6cH delays hatching, another useful survival mechanism in the presence of stress. Hatching arrest was most marked in 80% seawater when exposed to glyphosate at LC10. Water samples treated with Gly 6 cH showed specific interactions with solvatochromic dyes, mainly Coumarin 7, such that it appears to be a potential physicochemical marker for Gly 6 cH. In short, Gly 6 cH treatment appears to protect the Artemia salina population exposed to GBH at low concentrations.

Background: Highly diluted and succussed solutions (homeopathic potencies) have been shown to interact with a wide range of solvatochromic dyes based on changes in their UV-visible spectra. Studies so far have involved free dyes in solution, but there is a pressing need to find ways to investigate the potency-dye interaction using isolated dye molecules to ask more searching physico-chemical questions regarding the fundamental nature of potencies., Aims and Methods: The aims of the present study have been to look for ways to covalently immobilize solvatochromic dyes onto transparent cellulose films and hence be in a position to investigate dye-potency interactions without the complication of dye-dye interactions, including dye aggregation, which can occur with free dyes in solution., Results: To date, a total of nine different dyes have been immobilized on cellulose films using epoxide activation of hydroxyl groups on the cellulose surface. Using this method, studies have begun looking at the time course of potency action on one of these immobilized dyes, Brooker's merocyanine. Results show that the interaction of Arsenicum 10M with this dye consists of three phases-an initial growth phase, a sustained plateau of interaction, and a final decline phase lasting several days., Conclusion: A method has been developed that successfully immobilizes solvatochromic dyes onto transparent cellulose film. These films can then be used in a spectrophotometer to study at a much more detailed level how potencies interact with dyes compared with using free dyes in solution. Results indicate that the information gained in this way provides new insights regarding the fundamental nature of potencies. Specifically, studies using immobilized Brooker's merocyanine with Arsenicum 10M reveal that the lifetime of the potency is much longer than expected and that its action consists of three distinct phases, suggesting a resonant interaction with the dye. How resonant interaction might help to explain the clinical action of potencies is discussed., Competing Interests: None declared., (Faculty of Homeopathy. This article is published by Thieme.)

Introduction: Finding solutions to mitigate the impact of pollution on living systems is a matter of great interest. Homeopathic preparations of toxic substances have been described in the literature as attenuation factors for intoxication. Herein, an experimental study using Artemia salina and mercury chloride was developed as a model to identify aspects related to bioresilience., Aims: The aim of the study was to describe the effects of homeopathic Mercurius corrosivus (MC) on Artemia salina cysts hatching and on mercury bioavailability., Methods: Artemia salina cysts were exposed to 5.0 µg/mL of mercury chloride during the hatching phase. MC potencies (6cH, 30cH, and 200cH) were prepared in sterile purified water and poured into artificial sea water. Different controls were used (non-challenged cysts and challenged cysts treated with water, succussed water, and Ethilicum 1cH). Four series of nine experiments were performed to evaluate the percentage of cyst hatching. Soluble total mercury (THg) levels and precipitated mercury content were also evaluated. Solvatochromic dyes were used to check for eventual physicochemical markers of MC biological activity., Results: Significant delay ( p  < 0.0001) in cyst hatching was observed only after treatment with MC 30cH, compared with controls. This result was associated with an increase of THg concentration in water ( p  = 0.0018) and of chlorine/oxygen ratio ( p  < 0.0001) in suspended micraggregates, suggesting changes in mercury bioavailability. A specific interaction of MC 30cH with the solvatochromic dye ET33 ( p  = 0.0017) was found., Conclusion: Changes in hatching rate and possible changes in Hg bioavailability are postulated as protective effects of MC 30cH on Artemia salina , by improving its natural bioresilience processes., Competing Interests: None declared., (Faculty of Homeopathy. This article is published by Thieme.)

Background: Highly diluted and succussed solutions interact with solvatochromic dyes, indicating that changes in solvent and solute polarity could be related to their mechanism of action. It is not known, however, how the activity associated with succussed high dilutions is transferred to untreated water and what the limits of this process are., Aims: The aims of the present study were to ascertain whether a succussed high dilution of phosphorus (1.5 × 1 -59 M; Phos 30cH) seeded into a natural water source that fed a fjord and two connected lakes could propagate itself through the lake system (total volume 2200 m 3 ) and, moreover, whether the process could be tracked using solvatochromic dyes., Methods: Samples of water were collected before and after seeding, at different times and places throughout the lake system. Controls comprised water taken from an untreated and adjacent, but independent, lake (1385 m 3 )., Results: Water samples taken up to 72 hours after the source treatment produced significant increases ( p  ≤ 0.03) in the absorbance of the solvatochromic dye methylene violet (MV), while samples from the control lake produced no changes., Conclusions: The study indicates that activity associated with Phos 30c can propagate itself through large volumes of water, causing changes throughout a whole connected lake system, and that these changes can be tracked using the solvatochromic dye MV. This in turn means the use of homeopathic medicines in large volumes of drinking water, in farming and ecological contexts, now has the potential to be assessed with physico-chemical monitoring., Competing Interests: The authors declare no conflict of interest. The sponsors had no participation in the design, organization, and execution of the study nor in the interpretation of data or in writing of this manuscript or the submission decision. The scope of this study was outside of their commercial interests., (The Faculty of Homeopathy.)

Background: The mechanism by which highly diluted and agitated solutions have their effect is still unknown, but the development in recent years of new methods identifying changes in water and solute dipole moments is providing insights into potential modes of action., Objective: The objective of the current study was to compare the biological effects of Antimonium crudum (AC) previously obtained by our group and already described in the literature with now measurable physico-chemical effects on solvatochromic dyes., Methods: Different dilutions of AC and succussed water have been characterized with respect to their effect on the visible spectra of the solvatochromic dyes methylene violet (MV), a pyridinium phenolate (ET33), and a dimethylamino naphthalenone (BDN) compared with in-vitro action against Leishmania amazonensis -infected macrophages., Results: Dye responses varied according to the dye used and the level of AC dilution and results were found to corroborate previously published in-vivo and in-vitro effects of AC. In addition, a very significant enhancement in the absorbance increase of MV was seen using the supernatant from AC 200cH-treated cells (15%; p  < 0.0001) over that seen with AC 200cH itself (4%; p  = 0.034), suggesting the amplification of ultra-high dilution effects by biological systems. Furthermore, supernatants from AC-treated cells increased the range of dilutions of AC that were capable of producing effects on the spectra of MV. The effect of AC dilutions on dye ET33 was eliminated by a weak electric current passed through potency solutions., Conclusion: The data confirm a correspondence between the biological effects of dilutions of AC in-vitro and physico-chemical effects on solvatochromic dyes as measured by changes in their visible spectra. Results also indicate high dilutions of AC are sensitive to exposure to electric currents and biological systems., Competing Interests: None declared., (The Faculty of Homeopathy.)

Background: Homeopathic potencies have been shown to interact with a range of solvatochromic dyes to produce spectroscopic changes in the visible region of the electromagnetic spectrum. Furthermore, the nature of the changes observed under different experimental conditions is beginning to limit the number of possible hypotheses that can be put forward regarding the fundamental identity of potencies., Aims and Methods: The present study uses β-cyclodextrins to encapsulate solvatochromic dyes of widely varying structures. The purpose of this approach is to de-couple the primary dye-potency interaction from any subsequent aggregation effects., Results: Despite large differences in molecular structure between dyes, results show that potencies affect all dyes according to the same fundamental principles. Specifically, positively and negatively solvatochromic dyes collectively respond in opposite and complementary ways to potencies in accordance with the differential stabilisation of their excited and ground electronic states. Under the conditions of encapsulation, positively solvatochromic dyes display a bathochromic shift of, on average, 0.4 nm with a 2% absorbance change, and negatively solvatochromic dyes display a hypsochromic shift of, on average, 0.2 nm with a 1% absorbance change. This behaviour is only ever seen in two situations-where solvent becomes more polar or where an electric field is applied to solutions of dyes., Conclusions: The conditions used in this and previous studies to investigate the interaction of potencies with solvatochromic dyes preclude increased polarity of solvent as being responsible for the observed effects and that an explanation in which potencies carry an electric field (or electric field-like) component is by far the more likely. From the magnitude of the spectral changes induced in the dye Brooker's merocyanine by Arsenicum 10M, an estimate of the strength of the postulated electric field of 1.16 × 10 7  V/m can be made, which is comparable with the potential difference across cell membranes., Competing Interests: None declared., (The Faculty of Homeopathy.)

Background: The use of solvatochromic dyes to investigate homeopathic potencies holds out the promise of understanding the nature of serially succussed and diluted solutions at a fundamental physicochemical level. Recent studies have shown that a range of different dyes interact with potencies and, moreover, the nature of the interaction is beginning to allow certain specific characteristics of potencies to be delineated., Aims and Methods: The study reported in this article takes previous investigations further and aims to understand more about the nature of the interaction between potencies and solvatochromic dyes. To this end, the UV-visible spectra of a wide range of potential detectors of potencies have been examined using methodologies previously described., Results: Results presented demonstrate that solvatochromic dyes are a sub-group of a larger class of compounds capable of demonstrating interactions with potencies. In particular, amino acids containing an aromatic bridge also show marked optical changes in the presence of potencies. Several specific features of molecular detectors can now be shown to be necessary for significant interactions with homeopathic potencies. These include systems with a large dipole moment, electron delocalisation, polarizability and molecular rigidity., Conclusions: Analysis of the optical changes occurring on interaction with potencies suggests that in all cases potencies increase the polarity of molecular detectors to a degree that correlates with the size of the compound's permanent or ground dipole moment. These results can be explained by inferring that potencies themselves have polarity. Possible candidates for the identity of potencies, based on these and previously reported results, are discussed., Competing Interests: No source of funding had any influence on the design, analysis, interpretation or outcome of the research contained within this manuscript, nor on the writing of the manuscript., (The Faculty of Homeopathy.)

The third international conference on "Cutting Edge Research in Homeopathy" organised by the Homeopathy Research Institute (HRI) was held on the inspiring and historic island of Malta from 9th to 11th of June, 2017. One hundred and two abstracts underwent peer review by the HRI Scientific Advisory Committee and external experts to produce the programme of 36 oral presentations and 37 posters, presented by researchers from 19 countries. The 2.5-day programme covered a diverse range of topics, including quantitative and qualitative clinical research, basic research, veterinary research, and provings. These intensive plenary and parallel sessions were interspersed with multiple opportunities for delegates to discuss and exchange ideas, in particular through interactive panel discussions and a pre-conference workshop. The continuing commitment of the homeopathy research community to generate high-quality studies in this rapidly evolving field was clear. In this conference report, we present highlights from this memorable event., Competing Interests: None., (The Faculty of Homeopathy.)

Introduction: Previous studies have demonstrated the potential of solvatochromic dyes for investigating the physical chemistry of homeopathic potencies., Results: Following examination of this class of dyes in organic solvents, results obtained using the positively solvatochromic dye Bis-dimethylaminofuchsone (BDF) in aqueous solution are now reported. Spectral changes observed with this dye in the presence of potencies are both substantial and reproducible. Studies across a wide range of pH values reveal an unusual pH dependence for the dye's interaction with homeopathic potencies. Results indicate potency enhances dye protonation at pH values below c. 7.0, whilst protecting the dye from attack by hydroxyl ions above c. pH 7.5., Conclusion: A possible explanation for these observations is offered in terms of a potency - induced electron density shift in BDF. The interaction of homeopathic potencies with solvatochromic dyes, particularly BDF, points towards a possible physico-chemical model for the nature of potencies, how they may be interacting with this class of dyes, and moreover how their biological effects may be mediated., (Copyright © 2017 The Author. Published by Elsevier Ltd.. All rights reserved.)

Effective management of infectious disease relies upon understanding mechanisms of pathogen transmission. In particular, while models of disease dynamics usually assume transmission through direct contact, transmission through environmental contamination can cause different dynamics. We used Global Positioning System (GPS) collars and proximity-sensing contact-collars to explore opportunities for transmission of Mycobacterium bovis [causal agent of bovine tuberculosis] between cattle and badgers (Meles meles). Cattle pasture was badgers' most preferred habitat. Nevertheless, although collared cattle spent 2914 collar-nights in the home ranges of contact-collared badgers, and 5380 collar-nights in the home ranges of GPS-collared badgers, we detected no direct contacts between the two species. Simultaneous GPS-tracking revealed that badgers preferred land > 50 m from cattle. Very infrequent direct contact indicates that badger-to-cattle and cattle-to-badger M. bovis transmission may typically occur through contamination of the two species' shared environment. This information should help to inform tuberculosis control by guiding both modelling and farm management., (© 2016 John Wiley & Sons Ltd/CNRS.)

A systematic approach to the design of simple, chemical systems for investigating the nature of homeopathic medicines has led to the development of an experimental protocol in which solvatochromic dyes are used as molecular probes of serially diluted and agitated solutions. Electronic spectroscopy has been used to follow changes in the absorbance of this class of dyes across the visible spectrum in the presence of homeopathic potencies. Evidence is presented using six different solvatochromic dyes in three different solvent systems. In all cases homeopathic potencies produce consistent and reproducible changes in the spectra of the dyes. Results suggest that potencies influence the supramolecular chemistry of solvatochromic dyes, enhancing either dye aggregation or disaggregation, depending upon dye structure. Comparable dyes lacking the intramolecular charge transfer feature of solvatochromic dyes are unaffected by homeopathic potencies, suggesting potencies require the oscillating dipole of solvatochromic dyes for effective interaction. The implications of the results presented, both for an eventual understanding of the nature of homeopathic medicines and their mode of action, together with future directions for research in this area, are discussed., (Copyright © 2015 The Author. Published by Elsevier Ltd.. All rights reserved.)

Background: The successful execution of lateral osteotomies in rhinoplasty is an important step that can influence the functional and aesthetic outcome of the procedure., Objective: This paper describes an alternative method for achieving mobilisation of the nasal bones by careful application of Walsham forceps during primary rhinoplasty.

Habitat conversion for agriculture is a major driver of biodiversity loss, but our understanding of the demographic processes involved remains poor. We typically investigate the impacts of agriculture in isolation even though populations are likely to experience multiple, concurrent changes in the environment (e.g. land and climate change). Drivers of environmental change may interact to affect demography, but the mechanisms have yet to be explored fully in wild populations.Here, we investigate the mechanisms linking agricultural land use with breeding success using long-term data for the formerly Critically Endangered Mauritius kestrel Falco punctatus, a tropical forest specialist that also occupies agricultural habitats. We specifically focused on the relationship between breeding success, agriculture and the timing of breeding because the latter is sensitive to changes in climatic conditions (spring rainfall) and enables us to explore the interactive effects of different (land and climate) drivers of environmental change.Breeding success, measured as egg survival to fledging, declines seasonally in this population, but we found that the rate of this decline became increasingly rapid as the area of agriculture around a nest site increased. If the relationship between breeding success and agriculture was used in isolation to estimate the demographic impact of agriculture, it would significantly under-estimate breeding success in dry (early) springs and over-estimate breeding success in wet (late) springs.Analysis of prey delivered to nests suggests that the relationship between breeding success and agriculture might be due, in part, to spatial variation in the availability of native, arboreal geckos. Synthesis and applications . Agriculture modifies the seasonal decline in breeding success in this population. As springs are becoming wetter in our study area and since the kestrels breed later in wetter springs, the impact of agriculture on breeding success will become worse over time. Our results suggest that forest restoration designed to reduce the detrimental impacts of agriculture on breeding may also help reduce the detrimental effects of breeding late due to wetter springs. Our results therefore highlight the importance of considering the interactive effects of environmental change when managing wild populations.

Recent work suggests that the environment experienced in early life can alter life histories in wild populations, but our understanding of the processes involved remains limited. Since anthropogenic environmental change is currently having a major impact on wild populations, this raises the possibility that life histories may be influenced by human activities that alter environmental conditions in early life. Whether this is the case and the processes involved remain unexplored in wild populations. Using 23 years of longitudinal data on the Mauritius kestrel (Falco punctatus), a tropical forest specialist, we found that females born in territories affected by anthropogenic habitat change shifted investment in reproduction to earlier in life at the expense of late life performance. They also had lower survival rates as young adults. This shift in life history strategy appears to be adaptive, because fitness was comparable to that of other females experiencing less anthropogenic modification in their natal environment. Our results suggest that human activities can leave a legacy on wild birds through natal environmental effects. Whether these legacies have a detrimental effect on populations will depend on life history responses and the extent to which these reduce individual fitness., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Methods which aid and enhance the teaching of surgical procedures to trainees are beneficial to both trainer and trainee. In this article, we suggest a simple way of performing suction diathermy which allows the trainer to provide a template for the trainee to reproduce. Related articles have suggested the use of additional equipment, such as an endoscope; however, the method we describe requires no additional technical elements. Thus, it represents a sound and efficient teaching tool.

The amino acid sequence deduced from the L1 beta-lactamase gene of Xanthomonas maltophilia shows a significant variation from that of the CphA and Blm metallo-beta-lactamases of Aeromonas hydrophila and Bacillus cereus, respectively. Whilst the N-terminus of the L1 protein shows some similarity, particularly at the histidine residues previously suggested as a zinc-binding motif, the C-terminus of the protein demonstrates very little similarity. Such differences amongst this group of enzymes would argue for at least three subclasses within the Group 3 beta-lactamases. However, in order to predict their phylogenetic ancestry more sequence data are required from other possible metallo-beta-lactamases.

Several beta-lactamases, enzymes that play an important part in antibiotic resistance, have been purified by affinity chromatography on boronic acid gels. The procedure is rapid, appears to be selective for beta-lactamases, and allows a one-step purification of large amounts of enzyme from crude cell extracts. We have found the method useful for any beta-lactamase that is inhibited by boronic acids. Two kinds of boronic acid column have been prepared, the more hydrophobic one being reserved for those beta-lactamases that bind boronic acids relatively weakly. beta-Lactamase I from Bacillus cereus, P99 beta-lactamase and K 1 beta-lactamase from Gram-negative bacteria are among the better-known beta-lactamases that have been purified by this method. The procedure has also been used to purify a novel beta-lactamase from Pseudomonas maltophilia in high yield; the enzyme has an exceptionally broad substrate profile and hydrolyses monocyclic beta-lactams such as azthreonam and desthiobenzylpenicillin.

Cryoenzymology techniques were used to facilitate trapping an acyl-enzyme intermediate in beta-lactamase I catalysis. The enzyme (from Bacillus cereus) was investigated in aqueous methanol cryosolvents over the 25 to -75 degrees C range, and was stable and functional in 70% (v/v) methanol at and below 0 degree C. The value of kcat. decreased linearly with increasing methanol concentration, suggesting that water is a reactant in the rate-determining step. In view of this, the lack of incorporation of methanol into the product means that the water molecule involved in the deacylation is shielded from bulk solvent in the enzyme-substrate complex. From the lack of adverse effects of methanol on the catalytic and structural properties of the enzyme we conclude that 70% methanol is a satisfactory cryosolvent system for beta-lactamase I. The acyl-enzyme intermediate from the reaction with 6-beta-(furylacryloyl)amidopenicillanic acid was accumulated in steady-state experiments at -40 degrees C and the reaction was quenched by lowering the pH to 2. H.p.l.c. experiments showed covalent attachment of the penicillin to the enzyme. Digestion by pepsin and trypsin yielded a single labelled peptide fragment; analysis of this peptide was consistent with Ser-70 as the site of attachment.

In summary, Table XVI shows the inhibition profiles of representative beta-lactamases from each major class of Richmond and Sykes. Either resistance (R) or sensitivity (S) is given as a general guide to the type of compounds likely to inhibit each class. Thus the (qualitative) statements regarding the effectiveness of clavulanic acid can be taken to represent those for the penam sulfones and similarly for MM4550 and the other olivanic acids, carpetimycins, PS series, and asparenomycins. This can also be said of cloxacillin and the other aromatic carboxamido penicillins. Compounds are also included which are specifically or particularly inhibitory to certain beta-lactamases.

An inactivator of serine beta-lactamases, 6 beta-iodopenicillanate, can be utilized as a probe in the classification of beta-lactamases. It is a substrate for class-B Zn2+-containing beta-lactamase II. Although it inactivates enzymes from both classes A and C, it is much more efficient for the former group, with which it sometimes interacts following a branched pathway. On the basis of these observations, predictions are made concerning the class to which several enzymes belong.

A detailed understanding of the mechanism of enzyme action requires a correspondingly detailed knowledge of the structures of the intermediates and transition states on the catalytic pathway, as well as the kinetics and thermodynamics of their interconversion. Cryoenzymology, the use of subzero temperatures and fluid cryosolvents, has the potential to supply this type of information. In this article recent investigations illustrating the advantages of cryoenzymology are reviewed. The major advantage lies in the ability to accumulate and stabilize productive enzyme-substrate intermediates for sufficient lengths of time to allow the collection of high resolution structural data, e.g. by X-ray diffraction. In addition, intermediates which are not detectable under normal conditions, due to too short lifetimes and/or low concentrations, may be detected at low temperatures, in some cases through changes in the rate-limiting step. The method seems of rather general applicability, judging by the number of different types of enzymes, including oligomeric and membrane-associated ones, which have successfully been studied using cryoenzymology. In some cases, at least, good agreement between observations at subzero temperatures and normal conditions has been found, demonstrating the relevance of the technique. Potential limitations of the techniques, as well as questions regarding the effects on the protein structure, are also discussed.

The cryoenzymology of several different beta-lactamases has been investigated. Particular attention has been paid to the experimental pitfalls of the technique. These include such factors as false bursts at the start of the reaction, instability of the enzymes during turnover, and Km values so high that little of the enzyme is present as a complex. Many of the difficulties in cryoenzymology stem from the use of organic cryosolvents. A novel "salt" cryosolvent has been tested: ammonium acetate solutions can be used down to about -60 degrees C. The enzymes examined are readily soluble, and stable, in this solvent. Nevertheless, out of 17 beta-lactamase beta-lactam systems, only 4 proved suitable for detailed investigation. In two of these, the hydrolysis of nitrocefin or 7-(thienyl-2-acetamido)-3-[[2-[[4- (dimethylamino)phenyl]azo]pyridinio]-methyl]cephem-4-carboxylic acid (PADAC), by beta-lactamase I from Bacillus cereus, substrate was converted into product at a slow enough rate (at -60 or -55 degrees C, respectively) for it to be possible to do successive scans during the course of the reaction. The spectra were those of substrate and product, and no intermediate was detected. The results argue against the accumulation of intermediate acyl-enzyme. The hydrolysis of PADAC by the P99 beta-lactamase from Enterobacter cloacae again showed spectra characteristic of substrate and product, and there was, moreover, a break in the Arrhenius plot; it is possible that a conformational change is (at least partially) rate-determining. The hydrolysis of dinitrophenylpenicillin by the P99 beta-lactamase did show features suggesting the accumulation of acyl-enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

A pair of strains of Pseudomonas aeruginosa (3-Pre: cefsulodin-sensitive, inducible beta-lactamase; and 3-Post: cefsulodin-resistant, elevated beta-lactamase, derived from 3-Pre by subculture in the presence of cefsulodin) were taken as representative of the class of bacteria resistant to third-generation cephalosporins due to elevated synthesis of the normally inducible, chromosomally encoded beta-lactamase. These two strains were used to differentiate between 'trapping' and 'hydrolytic' mechanisms of cefsulodin resistance by (a) measuring the outer-membrane permeabilities to cefsulodin, (b) measuring the kinetics of cefsulodin hydrolysis and the stoichiometry of cefsulodin trapping by the periplasmic beta-lactamase, and (c) comparing the predictions of the trapping and hydrolysis hypotheses with the minimum inhibitory concentrations (MIC) of cefsulodin. The MIC of cefsulodin for strains 3-Pre and 3-Post were 2.35 microM (1.25 micrograms ml-1) and 37.6 microM (20.0 micrograms ml-1) respectively. The permeability parameter for cefsulodin of the outer membrane of the resistant strain was 0.0034 cm3 min-1 mg dry mass-1, so the flux of cefsulodin across its outer membrane at the MIC was calculated to be 0.120 nmol min-1 mg dry mass-1. Hydrolysis of cefsulodin by the beta-lactamase in the periplasm occurred at a rate of 0.118 nmol min-1 mg dry mass-1 which can thus account for resistance by matching the above rate of inflow. Trapping by the beta-lactamase, even with a 1:1 stoichiometry, would require the enzyme to be synthesized at 5.0 micrograms protein min-1 mg dry mass-1 or about 40% of the dry mass/generation. We conclude that hydrolysis, but not trapping, adequately explains the resistance to cefsulodin in P. aeruginosa 3-Post. A similar calculation for latamoxef resistance, using data taken from the literature, led to the same conclusion.