Geologically ancient DNA: fact or artefact?

Studies continue to report ancient DNA sequences and viable microbial cells that are many millions of years old. In this paper we evaluate some of the most extravagant claims of geologically ancient DNA. We conclude that although exciting, the reports suffer from inadequate experimental setup and insufficient authentication of results. Consequently, it remains doubtful whether amplifiable DNA sequences and viable bacteria can survive over geological timescales. To enhance the credibility of future studies and assist in discarding false-positive results, we propose a rigorous set of authentication criteria for work with geologically ancient DNA.     

Cyclic electron transport in C3 plants: fact or artefact?

The phenomenon of cyclic electron transport was first characterized in higher plant chloroplasts 50 years ago, yet there is still a debate about whether or not this is a physiological process. The recent isolation of mutants that appear to lack cyclic electron transport, as well as new data providing functional evidence for its occurrence, support the notion that this pathway plays an important role in plant responses to stress, providing a pH gradient across the thylakoid membrane to trigger non-photochemical quenching of chlorophyll fluorescence. At present, little is known about the regulation of cyclic electron transport, but it is possible that this is activated in response to a low redox potential in the chloroplast stroma.     

The carbon-quality temperature hypothesis: Fact or artefact?

Climate warming can reduce global soil carbon stocks by enhancing microbial decomposition. However, the magnitude of this loss remains uncertain because the temperature sensitivity of the decomposition of the major fraction of soil carbon, namely resistant carbon, is not fully known. It is now believed that the resistance of soil carbon mostly depends on microbial accessibility of soil carbon with physical protection being the primary control of the decomposition of protected carbon, which is insensitive to temperature changes. However, it is still unclear whether the temperature sensitivity of the decomposition of unprotected carbon, for example, carbon that is not protected by the soil mineral matrix, may depend on the chemical recalcitrance of carbon compounds. In particular, the carbon-quality temperature (CQT) hypothesis asserts that recalcitrant low-quality carbon is more temperature-sensitive to decomposition than labile high-quality carbon. If the hypothesis is correct, climate warming could amplify the loss of unprotected, but chemically recalcitrant, carbon and the resultant CO₂ release from soils to the atmosphere. Previous research has supported this hypothesis based on reported negative relationships between temperature sensitivity and carbon quality, defined as the decomposition rate at a reference temperature. Here we show that negative relationships can arise simply from the arbitrary choice of reference temperature, inherently invalidating those tests. To avoid this artefact, we defined the carbon quality of different compounds as their uncatalysed reaction rates in the absence of enzymes. Taking the uncatalysed rate as the carbon quality index, we found that the CQT hypothesis is not supported for enzyme-catalysed reactions, which showed no relationship between carbon quality and temperature sensitivity. The lack of correlation in enzyme-catalysed reactions implies similar temperature sensitivity for microbial decomposition of soil carbon, regardless of its quality, thereby allaying concerns of acceleration of warming-induced decomposition of recalcitrant carbon.     

Tendon conditioning: artefact or property?

Isolated tendons subjected to cyclic tensile loads higher than those experienced in the tendons' recent history exhibit 'conditioning', i.e. gradually increasing elongations upon loading and gradually increasing residual elongations after unloading in the first few loading-unloading cycles. The present study examines whether this behaviour is a measurement artefact or an actual time-dependent property. The gastrocnemius tendons of six men who refrained from rigorous physical activities prior to the experiment were loaded cyclically by 10 repeated isometric plantarflexion contractions at 80% of the moment generated during plantarflexion maximal voluntary contraction (MVC). In each contraction, the elongation of the gastrocnemius tendon at 80% of MVC and the residual tendon elongation after relaxation were obtained from the analysis of sonographs recorded during the test. The tendon elongation during activation and the residual tendon elongation after relaxation increased by ca. 5 mm from the first contraction to the tenth contraction, with no changes obtained after the fifth contraction. The behaviour of the tendon in the first five contractions indicates the presence of conditioning. It is therefore concluded that conditioning is a relevant property and not an artefact associated with in vitro testing. This has implications for joint kinematics and muscle excursion.     

The cytochemical localization of adenylate cyclase: fact or artifact?

In a study of the location of adenylate cyclase activity in rat pancreas with the method of Reik et al. (Science 168:382, 1970), as modified by Howell and Whitfield (J Histochem Cytochem 20:873, 1972) it was found that (a) unspecific staining occurs in rat pancreatic tissue fragments incubated in the Reik-Howell medium in the absence of substrate; (b) addition of adenylyl-imidodiphosphate (AMP-PNP) as substrate, either alone or together with stimulants of rat pancreas adenylate cyclase (secretin. NaF), does not result in increased precipitation; (c) cytochemical incubation of isolated rat pancreatic acinar cells and of rat liver and kidney fragments does not lead to substrate-specific precipitation. In subsequent chemical studies we have found that cyclic adenosine monophosphate (AMP) formation from [alpha32P]AMP-PNP in the presence of rat pancreatic particulate matter is very low in the Reik-Howell medium without lead ions, but is stimulated by addition of lead nitrate (4 mM). Whereas heat-treatment of the particulate matter abolishes all cyclic AMP formation in the absence of lead ions, it actually increases cyclic AMP production in the presence of 4 mM lead nitrate. This indicates that the cyclic AMP formation in the complet Reik-Howell medium occurs by a nonenzymatic mechanism. In addition, this medium shows a tendency to become turbid, particularly when calcium ions are added to the medium, suggesting a possible explanation for the apparently specific cytochemical detection observed by other authors. A revised cytochemical medium, with barium replacing lead and with a pH of 8.9 (optimal for adenylate cyclase with AMP-PNP substrate), leaves rat pancreatic adenylate cyclase activity intact and hormone sensitive, while it is still able to precipitate imidodiphosphate. However, cytochemical incubation of isolated rat pancreatic acinar cells in this revised medium in the presence of AMP-PNP and secretin does not yield an electron-dense precipitate, showing that the enzyme activity is to low to produce sufficient imidodiphosphate. These findings throw further doubt on the validity of the cytochemical detection of adenylate cyclase, reported by other investigators, notwithstanding the alleged positive results.     

The cytokine-prostaglandin cascade in fever production: fact or fancy?

(1) The classical view of the genesis of infectious fevers is that they develop in sequential steps, starting with the production by peripheral mononuclear phagocytes activated by the infectious noxa (i.e., the invading pathogens and/or their products, e.g., bacterial endotoxic lipopolysaccharides (LPS)) of pyrogenic cytokines, principally tumor necrosis factor-, interleukin(IL)-1β and IL-6, the release of these cytokines into the bloodstream, their transport to targets accessible from blood (e.g., cerebral microvessels, the organum vasculosum laminae terminalis) in close proximity to the ventromedial preoptic area (VMPO, the presumptive brain site of the febrigenic controller), and the consequent generation of stimulatory signals directed to the VMPO. An alternative view is that the message of the pyrogenic cytokines elaborated in the periphery is conveyed to the VMPO via a neural rather than a humoral pathway. In both views, cyclooxygenase (COX)-2-dependent prostaglandin (PG) E₂ is considered to be the proximal fever mediator induced in the VMPO by these cytokines, modulating the activity of thermosensitive neurons contained in this region and effecting the development of fever. (2) However, peripheral cytokines are not consistently detectable in febrile illnesses, and it was recently reported that neither circulating LPS nor cytokine levels are increased at the onset of robust fevers induced in rats by subcutaneous injections of replicating Escherichia coli. (3) And it was reported long ago that the intracerebroventricular (icv) injection of PGE₂ did not evoke fever in newborn lambs and goats although these animals responded normally to the intravenous injection of LPS, and that the icv-injection of synthetic PGE₂ antagonists that prevented the febrile response of rabbits to icv-injected PGE₂ did not inhibit that to a simultaneously injected endogenous pyrogen. (4) Other, more recent data indicate that the pyrogenic chemokine macrophage inflammatory protein-1β produces fever independently of PGE₂. (5) We found recently that the febrile response of adult guinea pigs to intravenously injected LPS is initiated significantly before the appearance of cytokines in the blood and, moreover, is evoked and sustainable in the absence of preoptic PGE₂. (6) What is amiss? Are the contradictory data fallacious or should the conventional wisdom be revisited? This issue is considered in this article and an explanation suggested for these disparate findings.     

Tumor counterattack: fact or fiction?

Cancer development relies on a variety of mechanisms that facilitate tumor growth despite the presence of a functioning immune system. Understanding these mechanisms may foster novel therapeutic approaches for oncology and organ transplantation. By expression of the apoptosis-inducing protein CD95L (FasL, APO-1L, CD178), tumors may eliminate tumor-infiltrating lymphocytes and suppress anti-tumor immune responses, a phenomenon called tumor counterattack. On the one hand, preliminary evidence of tumor counterattack in human tumors exists, and CD95L expression can prevent T-cell responses in vitro. On the other hand, CD95L-expressing tumors are rapidly rejected and induce inflammation in mice. Here, we summarize and discuss the consequences of CD95L expression of tumor cells and its contribution to immune escape.This article is a symposium paper from the conference Tumor Escape and Its Determinants, held in Salzburg, Austria, on 10–13 October 2004.     

Superoxide reductase: fact or fiction?

It was recently proposed that anaerobic microorganisms contain a new pathway for detoxification of reactive oxygen species. This is centered around a novel mononuclear iron-containing enzyme, superoxide reductase (SOR), which catalyzes the reduction, rather than the dismutation, of superoxide to hydrogen peroxide. A surprisingly large amount of relevant data has accumulated in the two years or so since the proposal was made. Herein we address the questions: to what extent has the pathway been validated, and what fundamental issues have yet to be answered in considering the response of anaerobes to reactive oxygen species? The evidence for superoxide reduction by SOR is now overwhelming and comes from a variety of anaerobic and microaerophilic species. Moreover, the available spectroscopic and structural information provide a convincing case that the catalytic Fe site of SOR is structurally and electronically tuned to mediate superoxide reduction rather than oxidation. Kinetic analyses also support the original proposal of NAD(P)H, via rubredoxin and NAD(P)H:rubredoxin oxidoreductase, as the source of reductant. What is still to be determined is the fate of the peroxide generated by the SOR reaction. In particular, the role of otherwise well-characterized proteins like rubrerythrin, NADH peroxidase, and rubredoxin:oxygen oxidoreductase in "anaerobic" oxygen metabolism has yet to be established.     

The hemolymph proteome of the honeybee: Gel‐based or gel‐free?

The honeybee has an invaluable economic impact and is a model for studying immunity, development and social behavior. The recent sequencing and annotation of the honeybee genome facilitates the study of its hemolymph, which reflects the physiological condition and mediates immune responses. We aimed at making a proteomic reference map of honeybee hemolymph and compared gel‐free and gel‐based techniques. One hundered and four 2‐DE spots corresponding to 62 different proteins were identified. Eight identical 2‐DLC experiments resulted in the identification of 32 unique proteins. One repeat was clearly not representative for the potential of the given 2‐DLC setup. Only 27% of the identified hemolymph proteins were found by both techniques. In addition, we found proteins of three different viruses which creates possibilities for biomarker design. Future hemolymph studies will benefit from this work.     

Structure of small tropical island freshwater fish and crustacean communities: A niche‐or dispersal‐based process?

Determining the relative importance of niche‐ and dispersal‐based processes in the structuring of animal communities is central in ecology. Freshwater fish and crustacean communities of small tropical islands can bring new insights for understanding these processes as all their species present a pelagic larval stage which gives them important dispersal capacities. Consequently, we hypothesized that dispersal‐based process may be preponderant for structuring these communities from the regional to the local, that is, survey site, scales. Gradient forest analyses allowed us to model the effect of 12 environmental variables on freshwater taxa abundances in two southwestern Indian Ocean islands: Mayotte (26 taxa) and Reunion (21). A total of 153 surveys in Mayotte and 266 in Reunion were used for building the models. Despite the strong heterogeneity of environmental conditions between the two islands, the main factors structuring freshwater fish and crustacean communities in both islands were the elevation and the slope of the sites. The observed structure appeared more pronounced for predatory species than for primary consumers and omnivores. As predators generally have limited locomotor capacities, it is concluded that dispersal‐based process structures these communities not only at large geographical scales but also at the intra‐watershed scale, by limiting the inland penetration (or dispersion) of species in relation to their locomotor capacities. However, more knowledge concerning ecological traits and taxonomic status of many species is needed to confirm this assumption. Abstract in French is available with online material     

Propagule pressure as a driver of establishment success in deliberately introduced exotic species: fact or artefact?

A central paradigm in invasion biology is that more releases of higher numbers of individuals increase the likelihood that an exotic population successfully establishes and persists. Recently, however, it has been suggested that, in cases where the data are sourced from historical records of purposefully released species, the direction of causality is reversed, and that initial success leads to higher numbers being released. Here, we explore the implications of this alternative hypothesis, and derive six a priori predictions from it. We test these predictions using data on Acclimatization Society introductions of passerine bird species to New Zealand, which have previously been used to support both hypotheses for the direction of causality. All our predictions are falsified. This study reaffirms that the conventional paradigm in invasion biology is indeed the correct one for New Zealand passerine bird introductions, for which numbers released determine establishment success. Our predictions are not restricted to this fauna, however, and we keenly anticipate their application to other suitable datasets.     

Haemophilus influenzae biofilms: hypothesis or fact?

Many publications state that nontypeable Haemophilus influenzae (NTHi) produces biofilms. Here, we review many of the publications that have led to acceptance by some that NTHi expresses a biofilm-specific phenotype as a distinct part of its life cycle. Biofilm formation was originally invoked to explain the failure to culture NTHi from middle-ear effusions, recalcitrance to antibiotics and its pathogenic behaviour. We argue that the current evidence for NTHi biofilm formation in vitro and in vivo is inconclusive. We consider that NTHi biofilm is hypothesis not fact, and although it might yet prove to be correct, there has been little or no consideration of alternative interpretations for the in vitro and in vivo observations. Uncritical acceptance of a distinctive NTHi biofilm phenotype has the potential to mislead and could confuse and compromise research efforts aimed at improving management and prevention of NTHi diseases of the human respiratory tract.     

Allergy to penicillin: fable or fact?

OBJECTIVE--To assess whether, on the basis of one blood test, penicillin allergy might be excluded sufficiently for general practitioners to give oral penicillin to patients claiming a history of penicillin allergy. DESIGN--Prospective study of patients referred by general practitioners. SETTING--Outpatient allergy clinic in a district general hospital. PATIENTS--175 referred patients who gave a history of immediate type reaction to penicillin, of whom 144 attended as requested and 132 completed the investigations. MAIN OUTCOME MEASURES--History and examination, serum radioallergosorbent test to phenoxymethylpenicillin and benzylpenicillin, and oral challenge with penicillin. RESULTS--Of 132 patients, four were confirmed to have penicillin allergy by the radioallergosorbent test and 128 had an oral penicillin challenge without ill effect. CONCLUSIONS--Most patients who gave a history of penicillin allergy are not so allergic, and their actual allergic state should be substantiated whenever feasible. For patients reporting minor or vague reactions negative findings with a radioallergosorbent test to phenoxymethylpenicillin and benzylpenicillin provide sufficient evidence to give oral penicillin safely.     

Radiation induced isostaining: fact or fiction?

Two different experiments were set up to induce differential staining in M-1¹ cells or to induce isostaining in M-2 cells after -irradiation of human lymphocytes. Neither of these two unusual staining patterns previously described by Luchnik and Porjadkova (1977) were observed. Possible explanations for this disagreement are discussed.Abbreviations SCE Sister chromatid exchange - IS Isostained regions - S-1, S-2, S-3 and S-4 First, second, third and fourth DNA synthetic periods after addition of 5-bromodeoxyuridine - M-1, M-2, M-3, M-4 and M-5 First, second, third, fourth and fifth mitosis after addition of 5-bromodeoxyuridine - BrdU 5-bromodeoxyuridine - EMEM Eagle's Minimum Essential Medium     

Mangrove growth rings: fact or fiction?

The analysis of tree rings in the tropics is less straightforward than in temperate areas with a demarcated unfavourable winter season. But especially in mangroves, the highly dynamic intertidal environment and the overriding ecological drivers therein have been a reason for questioning the existence of growth rings. This study aimed at casting light on growth rings in mangroves. In six mangrove species growing in Gazi Bay, Kenya (Sonneratia alba, Heritiera littoralis, Ceriops tagal, Bruguiera gymnorrhiza, Xylocarpus granatum and Lumnitzera racemosa), the occurrence of growth rings was examined. Growth rate of each species was determined based on a 1-year period using the cambial marking technique. The effect of climate was furthermore considered by comparing the results with a number of wood samples originating from contrasting climatic regions. We can conclude that for growth rings to appear in mangroves more than one condition has to be fulfilled, making general statements impossible and explaining the prevalent uncertainty. Climatic conditions that result in a range of soil water salinity experienced over the year are a prerequisite for the formation of growth rings. For species with an anatomy characterized by indistinct ring boundaries, this should be combined with a growth rate of at least 0.3 mm/year. The use of growth rings for age or growth rate determinations should thus be evaluated on a case by case basis.     

Reproductive clonality in protozoan pathogens—truth or artefact?

The debate around the frequency and importance of genetic exchange in parasitic protozoa is now several decades old. Recently, fresh assertions have been made that predominant clonal evolution explains the population structures of several key protozoan pathogens. Here, we present an alternative perspective. On the assumption that much apparent clonality may be an artefact of inadequate sampling and study design, we review current research to define why sex might be so difficult to detect in protozoan parasite populations. In doing so, we contrast laboratory models of genetic exchange in parasitic protozoa with natural patterns of genetic diversity and consider the fitness advantage of sex at different evolutionary scales. We discuss approaches to improve the accuracy of efforts to characterize genetic exchange in the field. We also examine the implications of the first population genomic studies for the debate around sex and clonality in parasitic protozoa and discuss caveats for the future.