Axillary bud outgrowth in herbaceous shoots: how do strigolactones fit into the picture?

Strigolactones have recently been identified as the long sought-after signal required to inhibit shoot branching (Gomez-Roldan et al. 2008; Umehara et al. 2008; reviewed in Dun et al. 2009). Here we briefly describe the evidence for strigolactone inhibition of shoot branching and, more extensively, the broader context of this action. We address the central question of why strigolactone mutants exhibit a varied branching phenotype across a wide range of experimental conditions. Where knowledge is available, we highlight the role of other hormones in dictating these phenotypes and describe those instances where our knowledge of known plant hormones and their interactions falls considerably short of explaining the phenotypes. This review will focus on bud outgrowth in herbaceous species because knowledge on the role of strigolactones in shoot branching to date barely extends beyond this group of plants.     

A model of direction selectivity in the starburst amacrine cell network

Displaced starburst amacrine cells (SACs) are retinal interneurons that exhibit GABA _A receptor-mediated and Cl ^? cotransporter-mediated, directionally selective (DS) light responses in the rabbit retina. They depolarize to stimuli that move centrifugally through the receptive field surround and hyperpolarize to stimuli that move centripetally through the surround (Gavrikov et al, PNAS 100(26):16047–16052, 2003, PNAS 103(49):18793–18798, 2006). They also play a key role in the activity of DS ganglion cells (DS GC; Amthor et al, Vis Neurosci 19:495–509 2002; Euler et al, Nature 418:845–852, 2002; Fried et al, Nature 420:411– 414, 2002; Gavrikov et al, PNAS 100(26):16047–16052, 2003, PNAS 103(49):18793–18798, 2006; Lee and Zhou, Neuron 51:787–799 2006; Yoshida et al, Neuron 30:771–780, 2001). In this paper we present a model of strong DS behavior of SACs which relies on the GABA-mediated communication within a tightly interconnected network of these cells and on the glutamate signal that the SACs receive from bipolar cells (a presynaptic cell that receives input from cones). We describe how a moving light stimulus can produce a large, sustained depolarization of the SAC dendritic tips that point in the direction that the stimulus moves (i.e., centrifugal motion), but produce a minimal depolarization of the dendritic tips that point in the opposite direction (i.e., centripetal motion). This DS behavior, which is quantified based on the relative size and duration of the depolarizations evoked by stimulus motion at dendritic tips pointing in opposite directions, is robust to changes of many different parameter values and consistent with experimental data. In addition, the DS behavior is strengthened under the assumptions that the Cl^? cotransporters Na^?+?-K^?+?-Cl^?? and K^?+?-Cl^?? are located in different regions of the SAC dendritic tree (Gavrikov et al, PNAS 103(49):18793–18798, 2006) and that GABA evokes a long-lasting response (Gavrikov et al, PNAS 100(26):16047–16052, 2003, PNAS 103(49):18793–18798, 2006; Lee and Zhou, Neuron 51:787–799, 2006). A possible mechanism is discussed based on the generation of waves of local glutamate and GABA secretion, and their postsynaptic interplay as the waves travel between cell compartments.     

Heme-bound nitroxyl, hydroxylamine, and ammonia ligands as intermediates in the reaction cycle of cytochrome c nitrite reductase: a theoretical study

In this article, we consider, in detail, the second half-cycle of the six-electron nitrite reduction mechanism catalyzed by cytochrome c nitrite reductase. In total, three electrons and four protons must be provided to reach the final product, ammonia, starting from the HNO intermediate. According to our results, the first event in this half-cycle is the reduction of the HNO intermediate, which is accomplished by two PCET reactions. Two isomeric radical intermediates, HNOH^? and H₂NO^?, are formed. Both intermediates are readily transformed into hydroxylamine, most likely through intramolecular proton transfer from either Arg₁₁₄ or His₂₇₇. An extra proton must enter the active site of the enzyme to initiate heterolytic cleavage of the N–O bond. As a result of N–O bond cleavage, the H₂N⁺ intermediate is formed. The latter readily picks up an electron, forming H₂N^+?, which in turn reacts with Tyr₂₁₈. Interestingly, evidence for Tyr₂₁₈ activity was provided by the mutational studies of Lukat (Biochemistry 47:2080, 2008), but this has never been observed in the initial stages of the overall reduction process. According to our results, an intramolecular reaction with Tyr₂₁₈ in the final step of the nitrite reduction process leads directly to the final product, ammonia. Dissociation of the final product proceeds concomitantly with a change in spin state, which was also observed in the resonance Raman investigations of Martins et al. (J Phys Chem B 114:5563, 2010).     

Missense mutations in the APOL1 gene are highly associated with end stage kidney disease risk previously attributed to the MYH9 gene

MYH9 has been proposed as a major genetic risk locus for a spectrum of nondiabetic end stage kidney disease (ESKD). We use recently released sequences from the 1000 Genomes Project to identify two western African-specific missense mutations (S342G and I384M) in the neighboring APOL1 gene, and demonstrate that these are more strongly associated with ESKD than previously reported MYH9 variants. The APOL1 gene product, apolipoprotein L-1, has been studied for its roles in trypanosomal lysis, autophagic cell death, lipid metabolism, as well as vascular and other biological activities. We also show that the distribution of these newly identified APOL1 risk variants in African populations is consistent with the pattern of African ancestry ESKD risk previously attributed to MYH9. Mapping by admixture linkage disequilibrium (MALD) localized an interval on chromosome 22, in a region that includes the MYH9 gene, which was shown to contain African ancestry risk variants associated with certain forms of ESKD (Kao et al. 2008; Kopp et al. 2008). MYH9 encodes nonmuscle myosin heavy chain IIa, a major cytoskeletal nanomotor protein expressed in many cell types, including podocyte cells of the renal glomerulus. Moreover, 39 different coding region mutations in MYH9 have been identified in patients with a group of rare syndromes, collectively termed the Giant Platelet Syndromes, with clear autosomal dominant inheritance, and various clinical manifestations, sometimes also including glomerular pathology and chronic kidney disease (Kopp 2010; Sekine et al. 2010). Accordingly, MYH9 was further explored in these studies as the leading candidate gene responsible for the MALD signal. Dense mapping of MYH9 identified individual single nucleotide polymorphisms (SNPs) and sets of such SNPs grouped as haplotypes that were found to be highly associated with a large and important group of ESKD risk phenotypes, which as a consequence were designated as MYH9-associated nephropathies (Bostrom and Freedman 2010). These included HIV-associated nephropathy (HIVAN), primary nonmonogenic forms of focal segmental glomerulosclerosis, and hypertension affiliated chronic kidney disease not attributed to other etiologies (Bostrom and Freedman 2010). The MYH9 SNP and haplotype associations observed with these forms of ESKD yielded the largest odds ratios (OR) reported to date for the association of common variants with common disease risk (Winkler et al. 2010). Two specific MYH9 variants (rs5750250 of S-haplotype and rs11912763 of F-haplotype) were designated as most strongly predictive on the basis of Receiver Operating Characteristic analysis (Nelson et al. 2010). These MYH9 association studies were then also extended to earlier stage and related kidney disease phenotypes and to population groups with varying degrees of recent African ancestry admixture (Behar et al. 2010; Freedman et al. 2009a, b; Nelson et al. 2010), and led to the expectation of finding a functional African ancestry causative variant within MYH9. However, despite intensive efforts including re-sequencing of the MYH9 gene no suggested functional mutation has been identified (Nelson et al. 2010; Winkler et al. 2010). This led us to re-examine the interval surrounding MYH9 and to the detection of novel missense mutations with predicted functional effects in the neighboring APOL1 gene, which are significantly more associated with ESKD than all previously reported SNPs in MYH9.     

An analysis of a ‘community-driven’ reconstruction of the human metabolic network

Following a strategy similar to that used in baker’s yeast (Herrgård et al. Nat Biotechnol 26:1155–1160, 2008). A consensus yeast metabolic network obtained from a community approach to systems biology (Herrgård et al. 2008; Dobson et al. BMC Syst Biol 4:145, 2010). Further developments towards a genome-scale metabolic model of yeast (Dobson et al. 2010; Heavner et al. BMC Syst Biol 6:55, 2012). Yeast 5—an expanded reconstruction of the Saccharomyces cerevisiae metabolic network (Heavner et al. 2012) and in Salmonella typhimurium (Thiele et al. BMC Syst Biol 5:8, 2011). A community effort towards a knowledge-base and mathematical model of the human pathogen Salmonella typhimurium LT2 (Thiele et al. 2011), a recent paper (Thiele et al. Nat Biotechnol 31:419–425, 2013). A community-driven global reconstruction of human metabolism (Thiele et al. 2013) described a much improved ‘community consensus’ reconstruction of the human metabolic network, called Recon 2, and the authors (that include the present ones) have made it freely available via a database at http://humanmetabolism.org/ and in SBML format at Biomodels (http://identifiers.org/biomodels.db/MODEL1109130000). This short analysis summarises the main findings, and suggests some approaches that will be able to exploit the availability of this model to advantage.     

Plant biotechnology for food security and bioeconomy

This year is a special year for plant biotechnology. It was 30 years ago, on January 18 1983, one of the most important dates in the history of plant biotechnology, that three independent groups described Agrobacterium tumefaciens—mediated genetic transformation at the Miami Winter Symposium, leading to the production of normal, fertile transgenic plants (Bevan et al. in Nature 304:184–187, 1983; Fraley et al. in Proc Natl Acad Sci USA 80:4803–4807, 1983; Herrera-Estrella et al. in EMBO J 2:987–995, 1983; Vasil in Plant Cell Rep 27:1432–1440, 2008). Since then, plant biotechnology has rapidly advanced into a useful and valuable tool and has made a significant impact on crop production, development of a biotech industry and the bio-based economy worldwide.     

Reply to: Comment on ‘neutron imaging reveals internal plant water dynamics’

Background

Our recent publication (Warren et al., Plant Soil 366:683–693, 2013) described how pulses of deuterium oxide (D₂O) or H₂O combined with neutron radiography can be used to indicate root water uptake and hydraulic redistribution in maize. This technique depends on the large inherent differences in neutron cross-section between D and H atoms resulting in strong image contrast.

Scope and Conclusions

However, as illustrated by Carminati and Zarebanadkouki (2013) there can be a change in total water content without a change in contrast simply by a change in the relative proportions of D₂O and H₂O. We agree with their premise and detailed calculations (Zarebanadkouki at al. 2012, 2013), and present further evidence that mixing of D₂O and H₂O did not confound evidence of hydraulic redistribution in our study.     

Mitochondrial complex I inhibitors, acetogenins, induce HepG2 cell death through the induction of the complete apoptotic mitochondrial pathway

The development of new anti-neoplastic drugs is a key issue for cancer chemotherapy due to the reality that, most likely, certain cancer cells are resistant to current chemotherapy. The past two decades have witnessed tremendous advances in our understanding of the pathogenesis of cancer. These advances have allowed identification new targets as oncogenes, tumor supressor genes and the possible implication of the mitochondria (Fulda et al. Nat Rev Drug Discov 9:447–464, 2010). Annonaceous Acetogenins (ACGs) have been described as the most potent inhibitors of the respiratory chain because of their interaction with mitochondrial Complex I (Degli Esposti and Ghelli Biochim Biophys Acta 1187:116–120, 1994; Zafra-Polo et al. Phytochemistry 42:253–271, 1996; Miyoshi et al. Biochim Biophys Acta 1365:443–452, 1998; Tormo et al. Arch Biochem Biophys 369:119–126, 1999; Motoyama et al. Bioorg Med Chem Lett 12:2089–2092, 2002). To explore a possible application of natural products from Annonaceous plants to cancer treatment, we have selected four bis-tetrahydrofuranic ACGs, three from Annona cherimolia (cherimolin-1, motrilin and laherradurin) and one from Rollinia mucosa (rollinianstatin-1) in order to fully describe their mechanisms responsible within the cell (Fig. 1). In this study, using a hepato-carcinoma cell line (HepG2) as a model, we showed that the bis-THF ACGs caused cell death through the induction of the apoptotic mitochondrial pathway. Their potency and behavior were compared with the classical mitochondrial respiratory chain Complex I inhibitor rotenone in every apoptotic pathway step.

Two ants (Insecta: Hymenoptera: Formicidae: Formicinae) from the Late Pliocene of Willershausen, Germany, with a nomenclatural note on the genus Camponotites

Two species of the genus Camponotites (Formicidae, Formicinae) are described from the Late Pliocene deposits of Willershausen, Lower Saxony, northern Germany: C. silvestris Steinbach, 1967, and C. steinbachi n. sp. The generic name Camponotites has been established for fossil (Tertiary) ants independently by Steinbach (Bericht der Naturhistorischen Gesellschaft zu Hannover 111:95–102, 1967) and by Dlussky (Trudy paleontologi?eskogo instituta, akademiâ nauk SSSR, 1981), each for materials of different stratigraphical and geographical origin. Though poorly described, Camponotites Steinbach, 1967, and the single included (type) species C. silvestris Steinbach, 1967 (a monotypic species from the Late Pliocene of Willershausen), were based upon indication in the sense of the ICZN. Therefore, both the generic and specific names are valid and available. Camponotites Dlussky, 1981 (and its type species C. macropterus Dlussky, 1981) were certainly introduced correctly and are therefore available, too; but due to its homonymy the generic name is not valid. The revision shows that in this rare case both generic names are not only homonyms but also synonyms.     

ORIUM: Optimized RDC-based Iterative and Unified Model-free analysis

Residual dipolar couplings (RDCs) are NMR parameters that provide both structural and dynamic information concerning inter-nuclear vectors, such as N–H^N and Cα–Hα bonds within the protein backbone. Two approaches for extracting this information from RDCs are the model free analysis (MFA) (Meiler et al. in J Am Chem Soc 123:6098–6107, 2001; Peti et al. in J Am Chem Soc 124:5822–5833, 2002) and the direct interpretation of dipolar couplings (DIDCs) (Tolman in J Am Chem Soc 124:12020–12030, 2002). Both methods have been incorporated into iterative schemes, namely the self-consistent RDC based MFA (SCRM) (Lakomek et al. in J Biomol NMR 41:139–155, 2008) and iterative DIDC (Yao et al. in J Phys Chem B 112:6045–6056, 2008), with the goal of removing the influence of structural noise in the MFA and DIDC formulations. Here, we report a new iterative procedure entitled Optimized RDC-based Iterative and Unified Model-free analysis (ORIUM). ORIUM unifies theoretical concepts developed in the MFA, SCRM, and DIDC methods to construct a computationally less demanding approach to determine these structural and dynamic parameters. In all schemes, dynamic averaging reduces the actual magnitude of the alignment tensors complicating the determination of the absolute values for the generalized order parameters. To readdress this scaling issue that has been previously investigated (Lakomek et al. in J Biomol NMR 41:139–155, 2008; Salmon et al. in Angew Chem Int Edit 48:4154–4157, 2009), a new method is presented using only RDC data to establish a lower bound on protein motion, bypassing the requirement of Lipari–Szabo order parameters. ORIUM and the new scaling procedure are applied to the proteins ubiquitin and the third immunoglobulin domain of protein G (GB3). Our results indicate good agreement with the SCRM and iterative DIDC approaches and signify the general applicability of ORIUM and the proposed scaling for the extraction of inter-nuclear vector structural and dynamic content.     

Mitochondrial iron metabolism in plants: frataxin comes into play

Friedreich ataxia (FRDA), an autosomal recessive neurological dysfunction that severely impairs motor coordination and reduction of life expectancy in humans, is caused by a deficiency in frataxin, a nuclear-encoded mitochondrial protein. Recently, a frataxin ortholog has been identified in Arabidopsis thaliana, named AtFH, with a transit peptide for localization in mitochondria and 65% sequence identity with human frataxin (Busi et al. FEBS Lett 576:141–144, 2004). Complementation of S. cerevisiae mutant strain Δyfh1 deficient in frataxin with AtFH, proved that the plant isoform is a functional protein, able to restore normal respiration and growth rates in the mutant yeast (Busi et al. FEBS Lett 576:141–144, 2004). AtFH is localized in mitochondria as its animal counterparts (Busi et al. Plant J 48:873–882, 2006); it is expressed mainly in flowers and developing embryos and it is an essential protein, since the knocking out of AtFH gene causes arrest of embryo development at the globular stage (Vazzola et al. FEBS Lett 581:667–672, 2007). A T-DNA insertional A.thaliana mutant showing a greater than 50% reduction of AtFH protein content, named atfh-1, has impaired activity of two mitochondrial enzymes possessing [Fe-S] clusters: aconitase and succinate dehydrogenase (Busi et al. Plant J 48:873–882, 2006). The results obtained in the last ten years on animal systems can contribute, without any doubt, to the elucidation of the role of frataxin in plant mitochondria; however, mitochondria of photosynthetically active cells, differently from animal ones, are not the major source of Reactive Oxygen Species (ROS) which could suggest possible differences in function between plant and animal frataxin.     

Constructions and headedness in derivation and compounding

The ??traditional?? distinction of compounds into endocentric (Eng. doorknob) and exocentric (pickpocket) is based on the presence or absence of a head constituent (Bloomfield, Language, Holt, New York, 1933); since the early eighties, the syntactic notion of ??head?? has been extended also to derivation, claiming that English derivational suffixes, as e.g. -ness, are heads, either in an absolute sense or in a categorial sense (see Williams Linguist Inq 12:245?C274, 1981; Lieber On the organization of the lexicon, Indiana university Linguistics Club, Bloomington, 1981; Lieber, in Yearbook of morphology 1989, Foris, Dordrecht, 1989; among others). In this paper, we shall first review some key issues in the morphological notion of head, illustrating well-known problematic cases, and then we shall discuss the Construction Morphology approach to headedness in derivation and compounding (Booij in The Oxford handbook of compounding, Oxford University Press, Oxford, 2009; Booij, in Cross-disciplinary issues in compounding, John Benjamins, Amsterdam, 2010a, 2010 Booij, Construction morphology, Oxford University Press, Oxford, 2010b). The stipulation of a hierarchical lexicon with subschemas expressing intermediate generalizations is a powerful theoretical device in accounting for a phenomenon as headedness variation, as we shall show with a Vietnamese case study; also, inconsistencies in word-class assignment in derivation will be dealt with in a constructionist perspective. Moreover, we shall discuss the consequences of a constructionist approach to the distinction between compounding and derivation.     

Typical Cyclical Behavioural Patterns: The Case of Routines,Rituals and Celebrations

The dynamics inherent to the life activity of all living systems presents itself in the form of regular patterns viewed by the observer as taking place in an extended timeline. Routines, rituals and celebrations, each in their own way, are defined by the typical cyclical behavioural patterns exhibited by individuals embedded in specific semiospheres. The particular nature of these semiospheres will determine the distinct patterns of behaviour to be adopted in different life contexts so that existential functions are fulfilled. The restricted and protected family circle normally provides the initial learning environment where the definition of the individual’s Umwelt (von Uexküll 1909, 1934; Ferreira, Biosemiotics 3(1):107–130, 2010, 2011) his/her meaningful world, starts to take shape. This learning process comprehends the progressive identification of distinct physical entities, the development of basic patterns of physical and social behaviour- the incorporation of basic routines and the assignment of meaning to particular events.     

Leucothoid and Maerid Amphipods (Crustacea) from deep regions of the North Atlantic

From amphipod collections from the deep Norwegian Sea, three species are here redescribed: Maera loveni (Bruzelius 1859) (Maeridae), Leucothoe aff. spinicarpa (Abildgaard, 1789) and L. uschakovi Gurjanova, 1951 (Leucothoidae). A key to all Atlantic and Mediterranean Leucothoe species (males only) is provided.     

A Mathematical and Numerical Investigation of the Hemodynamical Origins of Oscillations in Microvascular Networks

Evidence is presented to show that self-sustained oscillations of purely hemodynamical origin are possible in some arcade-type microvascular networks supplied with steady boundary conditions, but that in others the oscillations disappear with sufficient reduction of the time step Δt, showing them to be numerical artefacts. In an attempt to elucidate the mechanisms involved in the onset of fluctuations, we proceed to perform a linear stability analysis for the convective model of Kiani et al. (Microvasc. Res. 45:219–232, 1993; Am. J. Physiol. 266(35):H1822–H1828, 1994), and show that this leads via a system of delay differential equations to a nonlinear eigenvalue problem. This result generalises the characteristic equation obtained by Carr et al. (Ann. Biomed. Eng. 33:764–771, 2005) and Geddes et al. (SIAM J. Appl. Dyn. Syst. 6(4):694–727, 2007) who solved a special case in a two node network. An implicit numerical method is proposed for the computation of blood flows in networks using the convective model. In a moderate size subnetwork of one of the networks chosen by Kiani et al. (Am. J. Physiol. 266(35):H1822–H1828, 1994), the topology, vessel lengths, and diameters of which were based on microvascular networks in the rat mesentery, we compare results generated using the original explicit numerical method of Kiani et al. (Am. J. Physiol. 266(35):H1822–H1828, 1994) with those from our implicit scheme. From the linear stability theory, a critical value D _RBC,crit of a red blood cell diameter parameter D _RBC in the plasma skimming model of Fenton et al. (Pflügers Arch. 403:396–401, 1985b) is identified for the onset of oscillations about steady state and both the explicit and implicit methods are used to calculate the inflow hematocrit solutions in all vessels of the subnetwork at the critical parameter value, subject to perturbed initial conditions. The results of the implicit method are demonstrated to be in excellent and superior agreement with the predictions of the linear analysis in this case. For values of D _RBC slightly larger than D _RBC,crit the bifurcating periodic solutions calculated using either the explicit or implicit schemes are characteristic of those of a supercritical Hopf bifurcation and the graphs of D _RBC vs. oscillation amplitude would seem to converge as Δt→0.     

Integrating participatory approaches into social life cycle assessment: the SLCA participatory approach

Purpose

This article discusses the choice of stakeholder categories and the integration of stakeholders into participatory processes to define impact categories and select indicators.

Methods

We undertook a literature review concerning the roles and the importance of stakeholders in participatory processes, and the use of such processes in environmental and social LCAs (Biswas et al. Int J Life Cycle Assess 3(4):184-190, 1998; Sonnemann et al. Int J Life Cycle Assess 6(6):325-333, 2001; Baldo Int J Life Cycle Assess 7(5):269-275, 2002; James et al. Int J Life Cycle Assess 7(3):151-157, 2002; Bras-Kapwijk Int J Life Cycle Assess 8(5):266-272, 2003; Mettier et al. Int J Life Cycle Assess 11(6):468-476, 2006). As part of the French National Research Agency Piscenlit project, we adapted the Principle, Criteria, Indicator (PCI) method (Rey-Valette et al. 2008), which is an assessment method of sustainable development, as a way to integrate the participatory approach into Social Life Cycle Assessment (SLCA) methodology, mainly at the impact definition stage.

Results and discussion

Different views of participation were found in the literature; there is no consensual normative approach for the implication of stakeholders in LCA development. Some attempts have been made to integrate stakeholders into environmental LCAs but these attempts have not been generalized. However, they strongly emphasize the interrelationship between research on the growing integration of stakeholders and on the choice of stakeholders. We then propose criteria from stakeholder theory (Freeman 1984; Mitchell et al. Acad Manage Rev 22(4):853-886, 1997; Geibler et al. Bus Strat Environ 15:334-346, 2006) in order to identify relevant stakeholders for SLCA participatory approach. The adaptation of the PCI method to Principles, Impacts, and Indicators (PII) enables stakeholders to express themselves and hence leads to definitions of relevant social indicators that they can appropriate. The paper presents results regarding the selection of stakeholders but no specific results regarding the choice of impact categories and indicators.

Conclusions and recommendations

Integrating a participatory approach into SLCAs is of interest at several levels. It enables various factors to be taken into account: plurality of stakeholder interests, local knowledge, and impact categories that make sense for stakeholders in different contexts. It also promotes dialogue and simplifies the search for indicators. However, it requires a multidisciplinary approach and the integration of new knowledge and skills for the SLCA practitioners.     

Simple signaling games of sexual selection (Grafen’s revisited)

We investigate several versions of a simple game of sexual selection, to explore the role of secondary sexual characters (the “handicap paradox”) with the tools of signaling theory. Our models admit closed form solutions. They are very much inspired by Grafen’s (J Theor Biol 144:517–546, 1990a; J Theor Biol 144:473–516, 1990b) seminal companion papers. By merging and simplifying his two approaches, we identify a not so minor artifact in the seminal study. We propose an alternative model to start with Grafen’s sexual selection theory, with several similarities with Getty (Anim Behav 56:127–130, 1998).     

Detection of Legionella spp. and Legionella pneumophila in water samples of Spain by specific real-time PCR

Legionella pneumophila is the primary cause of the legionellosis diseases (90 %) (Yu et al. in J Infect Dis 186:127–128, 2002; Doleans et al. in J Clin Microbiol 42:458–460, 2004; Den Boer et al. in Clin Microbiol Infect 14:459–466, 2008). In this study, methodologies based on molecular biology were developed in order to provide a quick diagnosis of the bacterial presence in water samples of Spain. Multiplex real-time polymerase chain reaction assays were realized to target the 16S rRNA and macrophage infectivity potentiator (mip) genes of, respectively, Legionella spp. and L. pneumophila including in the design of an internal control. The results obtained by the culture and the gene amplification methods agreed in 94.44 % for the 16S rRNA gene, and a concordance of 66.67 % of the cases was obtained for the mip gene.