A Lack of Specificity for Ethylene-induced Mitochondrial Changes

A critical evaluation was made of the hypothesis that the primary mode of action of ethylene in inducing physiological responses is by changing the permeability of cell organelles. The parameter investigated was the evaluation of the influence of ethylene and other gases on mitochondrial oxidation and swelling. Spectrometric evidence demonstrated that mitochondria prepared with good respiratory control can be induced to swell more rapidly with ethylene and other aliphatic gases (ethane, propene, propane, I-butene) in test solutions of 0.125 m KCl. The fact that saturated as well as unsaturated hydrocarbon gases elicited similar changes provides evidence that ethylene does not directly alter membrane permeability as its mechanism of action.     

Species Specificity of Brain Cyclic AMP Receptor Proteins

The cyclic AMP binding proteins present in mouse, rat, bovine, and sheep brains were compared. Extracts were isotopically labeled with 8-azido-cyclic [32P]AMP, a photoaffinity analog specific for cyclic AMP binding sites, and then subjected to two-dimensional gel electrophoresis. The resulting autoradiographic patterns were generally similar, but showed consistent species variations. Proteins identified by their size and phosphorylatability as regulatory subunits of Type II protein kinase isozymes were present in all species, but with slight variations in pI. A series of charge variants identified as regulatory subunits of Type I kinase isozymes on the basis of their size was also ubiquitously present, as were several smaller proteins postulated to be proteolytic fragments derived from the regulatory subunits. The major species difference was a series of labeled proteins found only in rodent brains and not in the brains of any ruminant, or in other rodent tissues. These proteins had a molecular weight of 54,000 and a pI range of 5.89--6.26, and could not be endogenously phosphorylated. The identities of these proteins and their relationship to the protein kinase regulatory subunits are unknown.     

Lack of Specificity of Commercially Available Antisera: Better Specifications Needed

The ideal antiserum for immunohistochemical (IHC) applications contains monospecific high-affinity antibodies with little nonspecific adherence to sections. Many commercially available antibodies are “affinity” purified, but it is unknown if they meet “hard” specificity criteria, such as absence of staining in tissues genetically deficient for the antigen or a staining pattern that is identical to that of an antibody raised against a different epitope on the same protein. Reviewers, therefore, often require additional characterization. Although the affinity-purified antibodies used in our study on the distribution of muscarinic receptors produced selective staining patterns on sections, few passed the preabsorption test, and none produced bands of the anticipated size on Western blots. More importantly, none showed a difference in staining pattern on sections or Western blots between wild-type and knockout mice. Because these antibodies were used in most studies published thus far, our findings cast doubts on the validity of the extant body of morphological knowledge of the whole family of muscarinic receptors. We formulate requirements that antibody-specification data sheets should meet and propose that journals for which IHC is a core technique facilitate consumer rating of antibodies. “Certified” antibodies could avoid fruitless and costly validation assays and should become the standard of commercial suppliers. (J Histochem Cytochem 56:1099–1111, 2008)     

Cell Surface Saccharides in Three Phytomonas Species Differing in Host Specificity

ABSTRACT. Cell surface carbohydrates of three phytoflagellates, Phytomonas francai. Phytomonas serpens and Phytomonas sp. from different hosts including cassava, coreid insect Phthia picta and the milkweed plant Euphorbia hyssopifolia, respectively, were analysed by agglutination assays employing a battery of highly purified lectins with affinity for receptor molecules containing N-acetylglucosamine (d-GlcNAc), N-acetylgalactosamine (D-GalNAc), galactose, mannose-like (D-Man-like) residues and fucose, and by binding assay using radiolabeled [¹²⁵I]-wheat germ agglutinin (WGA) and fluorescent WGA lectin, as well as glycosidases of known sugar specificity, Escherichia coli K with mannose-affinity fimbrial lectin was also used as an agglutination probe. In general, the presence of D-GlcNAc. D-GalNAc and D-Man-like residues was detected in the phytomonads' plasma membrane. These sugar moieties were confirmed in whole cell hydrolysates as assessed by gas-liquid chromatography (GLC) which in addition, also showed the presence of galactose and xylose. However, marked differences in cell surface carbohydrate structures were observed. Wheat germ agglutinin, which binds to sialic acid and/or d-GlcNAc-containing residues, shows selective agglutinin activities for P. francai and Phytomonas sp., while Bandeiraea simplicifolia II agglutinin (which recognizes d-GlcNAc units) specifically bound to Phytomonas sp. Helix pomatia agglutinin which binds to D-GalNAc-containing residues reacted preferentially with Phytomonas sp. and P. serpens. Con A, which recognizes D-Man-like receptors, agglutinates all the phytomonads; however, the higher interaction was observed with Phytomonas sp. P. francai was selectively agglutinated in the presence of E. coli fimbrial lectin. Fluorescence WGA binding was significantly decreased by N-acetylglucosaminidase activities and the cell agglutination was not altered by neuraminidase treatment, suggesting the presence of an exposed D-GlcNAc moiety on the P. francai and Phytomonas sp. surfaces. Binding studies with [¹²⁵I]-WGA essentially confirmed the fluorescence WGA binding and agglutination assays.     

The Site Specificity of Two Species of Gregarina in Tenebrio molitor Larvae

The site specificity of the apicomplexans Gregarina cuneata and Gregarina sleini , in larval Tenebrio molitor was investigated. Gregarina cuneata was found to inhabit the anteriormost region of the larval midgut, while G. steini was restricted to the posterior portion of the intestine. The site specificity of the pair was conserved in single and concurrent infections. Interspecific interactions do not seem to be presently responsible for the resource partitioning by the 2 gregarine species. Key words. Gregarina, site specificity, Tenebrio molitor.     

Plant-Associated Symbiotic Burkholderia Species Lack Hallmark Strategies Required in Mammalian Pathogenesis

Burkholderia is a diverse and dynamic genus, containing pathogenic species as well as species that form complex interactions with plants. Pathogenic strains, such as B. pseudomallei and B. mallei, can cause serious disease in mammals, while other Burkholderia strains are opportunistic pathogens, infecting humans or animals with a compromised immune system. Although some of the opportunistic Burkholderia pathogens are known to promote plant growth and even fix nitrogen, the risk of infection to infants, the elderly, and people who are immunocompromised has not only resulted in a restriction on their use, but has also limited the application of non-pathogenic, symbiotic species, several of which nodulate legume roots or have positive effects on plant growth. However, recent phylogenetic analyses have demonstrated that Burkholderia species separate into distinct lineages, suggesting the possibility for safe use of certain symbiotic species in agricultural contexts. A number of environmental strains that promote plant growth or degrade xenobiotics are also included in the symbiotic lineage. Many of these species have the potential to enhance agriculture in areas where fertilizers are not readily available and may serve in the future as inocula for crops growing in soils impacted by climate change. Here we address the pathogenic potential of several of the symbiotic Burkholderia strains using bioinformatics and functional tests. A series of infection experiments using Caenorhabditis elegans and HeLa cells, as well as genomic characterization of pathogenic loci, show that the risk of opportunistic infection by symbiotic strains such as B. tuberum is extremely low.     

Affinity and Specificity of Ciprofloxacin-Bovine Serum Albumin Interactions: Spectroscopic Approach

Fluorescence spectroscopy in combination with UV–Vis absorption spectroscopy were employed to investigate the binding of an antibacterial drug Ciprofloxacin (CPFX) to bovine serum albumin (BSA) under the physiological conditions. In the discussion of the quenching mechanism, it was proved that the fluorescence quenching of BSA by CPFX is a result of the formation of CPFX-BSA complex. Binding parameters were determined using the modified Stern-Volmer equation and Scatchard equation to provide a measure of the binding affinity between CPFX and BSA. The results of thermodynamic parameters ΔG, ΔH, ΔS, at different temperatures indicate that the electrostatic interactions play a major role for CPFX-BSA association. Site marker competitive experiments indicated that the binding of CPFX to BSA primarily took place in site I. Furthermore, the effect of metal ions to CPFX-BSA system was studied, and the distance r between donor (BSA) and acceptor (CPFX) was obtained according to fluorescence resonance energy transfer (FRET). The conformation of BSA upon CPFX binding was evaluated by measuring synchronous fluorescence properties of the CPFX-BSA complex.     

Humoral Recognition Factors in the Arthropoda. The Specificity of Chelicerata Serum Lectins

Arthropods have the capacity of recognizing self from non-selfin various defense phenomena including hemolymph clotting, phagocytosis,encapsulation, melanization and clearance of the foreign matterwhich must be initiated by a first step of molecular recognition."Natural" and experimentally induced humoral factors have beendescribed which act as hemagglutinins, bacterial agglutinins,precipitins, bactericidal factors, bacteriolysins, hemolysins,opsonins, clotting factors, and lysozymes. Their exact rolein recognition functions has not been fully explored and theirfunction remains unclear. Among these factors, the carbohydrate-bindingmolecules (lectins) are the best characterized in their specificity,physicochemical properties and molecular structure. Arthropodlectins are multimeric, high molecular weight protein (glycoprotein)molecules with a certain degree of heterogeneity in their specificityand structure. In particular, serum lectins from chelicerates(horseshoe crabs, scorpions and spiders) share a common property:the specificity for sialic acids. Arachnids and merostomes divergedin the earliest Cambrian. Since they occupy markedly differenthabitats, the sialic acid specific lectins most probably area relict rather than an adaptative character. In addition tothis common feature of specificity, lectins from cheliceratesand other arthropods represent heterogeneous populations whichcan bind a wide variety of carbohydrates, many of them presenton bacteria as D-galactose, 2-keto-3-deoxyoctonate, glucuronicacid, N-acetylmuramic acid, and colominic acid. Multiplicityin specificity suggests that serum lectins might contributeas a carbohydrate-based recognition system for the non-self.The requirement for avoiding self recognition would be thatcarbohydrate structures potentially recognized by the systemwould be absent, masked or out of reach of this humoral factoror cell associated recognition factors.     

Lack of Specificity in Plant Extracts and Chemicals as Inducers of Systemic Resistance in Cucumber Plants to Anthracnose

Canoia, lettuce, green bean, dwarf pea, tobacco, tomato, poplar, corn, wheat, and cucumber leaves were extracted by boiling with 95% ethanol. The ethanol was removed from the ethanol-water extract, and the water-soluble fraction was centrifuged and filtered to remove compounds with a molecular weight > 1 000 D. The undersides of the first true leaf of a cucumber plant (leaf 1) and the leaf above (leaf 2) were sprayed with each plant extract (induction), and 7 days later leaf 3 was inoculated with a conidial suspension of Colletotrichym lagenarium (challenge). Extracts from all plants tested induced systemic resistance in leaf 3 to C. lagenarium . Commercially available compounds were screened to determine structural similarities among the compounds which can induce systemic resistance. All compounds which induced resistance in leaf 3 caused some damage to leaves 1 and 2. but not all compounds which damaged leaves 1 and 2 induced resistance in leaf 3. Galacturonic acid, glucuronic acid, salicyhc acid, m-hydroxybenzoic acid, p-hydroxybenzoic acid, y-resorcylic acid, protocatecbuic acid, gallic acid, hemimelletic acid, trimelletic acid, trimesic acid, phloroglucinol, and glycine induced systemic resistance, whereas catechol, resorcinol, hydroquinone, pyrogaLol, hydroxyhydroquinone, benzoic acid, phthalic acid, α-resorcylic acid, β-resorcylic acid, gentisic acid. 2, 3-dihydroxybenzoic acid, benzaldehyde, 2,3-dihydroxybenzaldehyde, β-resorcaldehyde, protocatechuic aldehyde, benzyl alcohol, salicyl alcohol, myo-inositol, phytic acid, pinitol, and quebrachitol did not. Structural similarities were not evident among either compounds which induced systemic resistance or those that did not.     

Carotenoids in Rhodoplanes Species: Variation of Compositions and Substrate Specificity of Predicted Carotenogenesis Enzymes

Phototrophic bacteria necessarily contain carotenoids for photosynthesis, and accumulate unusual carotenoids in some cases. The carotenoids in all established species of Rhodoplanes (Rpl.), a representative of phototrophic genera, were identified using spectroscopic methods. The major carotenoid was spirilloxanthin in Rpl. roseus and Rpl. serenus, and rhodopin in "Rpl. cryptolactis". Rpl. elegans contained rhodopin, anhydrorhodovibrin, and spirilloxanthin. Rpl. pokkaliisoli contained not only rhodopin but also 1,1'-dihydroxylycopene and 3,4,3',4'-tetrahydrospirilloxanthin. These variations in carotenoid composition suggested that Rpl. roseus and Rpl. serenus had normal substrate specificity of the carotenogenesis enzymes of CrtC (acyclic carotene 1,2-hydratase), CrtD (acyclic carotenoid 3,4-desaturase), and CrtF (acyclic 1-hydroxycarotenoid methyltransferase). On the other hand, CrtC of Rpl. elegans, CrtD of "Rpl. cryptolactis", and CrtC, CrtD, and CrtF of Rpl. pokkaliisoli might have different characteristics from the usual activity of these normal enzymes in the normal spirilloxanthin pathway. These results suggest that the variation of carotenoids among the species of Rhodoplanes results from modified substrate specificity of the carotenogenesis enzymes involved.     

Specificity of Lipoxygenase Pathways Supports Species Delineation in the Marine Diatom Genus Pseudo-nitzschia

Oxylipins are low-molecular weight secondary metabolites derived from the incorporation of oxygen into the carbon chains of polyunsaturated fatty acids (PUFAs). Oxylipins are produced in many prokaryotic and eukaryotic lineages where they are involved in a broad spectrum of actions spanning from stress and defense responses, regulation of growth and development, signaling, and innate immunity. We explored the diversity in oxylipin patterns in the marine planktonic diatom Pseudo-nitzschia. This genus includes several species only distinguishable with the aid of molecular markers. Oxylipin profiles of cultured strains were obtained by reverse phase column on a liquid chromatograph equipped with UV photodiode detector and q-ToF mass spectrometer. Lipoxygenase compounds were mapped on phylogenies of the genus Pseudo-nitzschia inferred from the nuclear encoded hyper-variable region of the LSU rDNA and the plastid encoded rbcL. Results showed that the genus Pseudo-nitzschia exhibits a rich and varied lipoxygenase metabolism of eicosapentaenoic acid (EPA), with a high level of specificity for oxylipin markers that generally corroborated the genotypic delineation, even among genetically closely related cryptic species. These results suggest that oxylipin profiles constitute additional identification tools for Pseudo-nitzschia species providing a functional support to species delineation obtained with molecular markers and morphological traits. The exploration of the diversity, patterns and plasticity of oxylipin production across diatom species and genera will also provide insights on the ecological functions of these secondary metabolites and on the selective pressures driving their diversification.     

Lack of Differences in Serum Binding of Calcium in Cystic Fibrosis,Carriers and Controls

Attempts have been made to detect carriers for cystic fibrosis (CF) by some means other than genetic inference from affected children to their otherwise normal parents¹. The ability to detect CF carriers would result in substantial advances in genetic counselling. Moreover, an assay by which heterozygotes are identifiable must involve a parameter close to the elusive primary gene abnormality. Accordingly the paper by Fitz-patrick et al.² in which serum binding of tracer amounts of calcium (determined by equilibrium dialysis) of CF > heterozygotes > controls was a potentially significant report. We were, however, sceptical about data in which healthy carrier individuals demonstrated so great a difference from normal (an increase in serum binding of ⁴⁵Ca in heterozygotes over controls of 65 ± 11%) for a clearly nonspecific characteristic. As we have substantial serum and salivary electrophoretic data³ in which no consistent protein differences were apparent in carriers and because we wondered about the biological explanation of the findings of Fitzpatrick et al.², we decided to repeat their serum binding of calcium experiments. We repeated the reported procedures as closely as possible. We also assayed calcium binding with the additional buffer and salt concentration conditions described below. Sera from different individuals [N (controls = 30; N (heterozygotes) = 31; N (CF) = 33] were used in 4 experiments.     

Robust Detection of Rare Species Using Environmental DNA: The Importance of Primer Specificity

Environmental DNA (eDNA) is being rapidly adopted as a tool to detect rare animals. Quantitative PCR (qPCR) using probe-based chemistries may represent a particularly powerful tool because of the method’s sensitivity, specificity, and potential to quantify target DNA. However, there has been little work understanding the performance of these assays in the presence of closely related, sympatric taxa. If related species cause any cross-amplification or interference, false positives and negatives may be generated. These errors can be disastrous if false positives lead to overestimate the abundance of an endangered species or if false negatives prevent detection of an invasive species. In this study we test factors that influence the specificity and sensitivity of TaqMan MGB assays using co-occurring, closely related brook trout (Salvelinus fontinalis) and bull trout (S. confluentus) as a case study. We found qPCR to be substantially more sensitive than traditional PCR, with a high probability of detection at concentrations as low as 0.5 target copies/µl. We also found that number and placement of base pair mismatches between the Taqman MGB assay and non-target templates was important to target specificity, and that specificity was most influenced by base pair mismatches in the primers, rather than in the probe. We found that insufficient specificity can result in both false positive and false negative results, particularly in the presence of abundant related species. Our results highlight the utility of qPCR as a highly sensitive eDNA tool, and underscore the importance of careful assay design.     

The Extracellular A-loop of Dual Oxidases Affects the Specificity of Reactive Oxygen Species Release

NADPH oxidase (Nox) family proteins produce superoxide (O₂^⨪) directly by transferring an electron to molecular oxygen. Dual oxidases (Duoxes) also produce an O₂^⨪ intermediate, although the final species secreted by mature Duoxes is H₂O₂, suggesting that intramolecular O₂^⨪ dismutation or other mechanisms contribute to H₂O₂ release. We explored the structural determinants affecting reactive oxygen species formation by Duox enzymes. Duox2 showed O₂^⨪ leakage when mismatched with Duox activator 1 (DuoxA1). Duox2 released O₂^⨪ even in correctly matched combinations, including Duox2 + DuoxA2 and Duox2 + N-terminally tagged DuoxA2 regardless of the type or number of tags. Conversely, Duox1 did not release O₂^⨪ in any combination. Chimeric Duox2 possessing the A-loop of Duox1 showed no O₂^⨪ leakage; chimeric Duox1 possessing the A-loop of Duox2 released O₂^⨪. Moreover, Duox2 proteins possessing the A-loops of Nox1 or Nox5 co-expressed with DuoxA2 showed enhanced O₂^⨪ release, and Duox1 proteins possessing the A-loops of Nox1 or Nox5 co-expressed with DuoxA1 acquired O₂^⨪ leakage. Although we identified Duox1 A-loop residues (His¹⁰⁷¹, His¹⁰⁷², and Gly¹⁰⁷⁴) important for reducing O₂^⨪ release, mutations of these residues to those of Duox2 failed to convert Duox1 to an O₂^⨪-releasing enzyme. Using immunoprecipitation and endoglycosidase H sensitivity assays, we found that the A-loop of Duoxes binds to DuoxA N termini, creating more stable, mature Duox-DuoxA complexes. In conclusion, the A-loops of both Duoxes support H₂O₂ production through interaction with corresponding activators, but complex formation between the Duox1 A-loop and DuoxA1 results in tighter control of H₂O₂ release by the enzyme complex.     

Species Richness and Host Specificity among Caterpillar Ensembles on Shrubs in the Andes of Southern Ecuador

Caterpillar ensembles were sampled on 16 species of shrubs from the family Asteraceae and the genus Piper (Piperaceae) in open and forest habitats in the Andean montane forest zone of southern Ecuador between August 2007 and May 2009. Trophic affiliations of caterpillars to the host plants were confirmed in feeding trials. Overall, species richness of herbivorous caterpillars was high (191 species across all plants), but varied strongly between ensembles associated with different plant species (2?C96 lepidopteran species per shrub species). Ensembles on Piper species were characterized by low effective species numbers and high dominance of one or two species of the Geometridae genus Eois Hübner. Low species number and high dominance were also found on latex-bearing Erato polymnioides, whereas ensembles on two other Asteraceae species were far more diverse and less strongly shaped by a few dominant species. The observed diversity patterns fit well to the concept that anti-herbivore defenses of plants are the major factors regulating associated insect ensembles. Local abundance and geographic range of host plants appear to have less influence. Lepidopteran species feeding on Asteraceae were found to be more generalistic than those feeding on Piper species. We conclude that caterpillar ensembles on most, but not all, studied plant species are defined by a small number of dominant species, which usually are narrow host specialists. This pattern was more distinct on Piper shrubs in forest understory, whereas Asteraceae in disturbed habitats had more open caterpillar ensembles.     

Zonadhesin Is Essential for Species Specificity of Sperm Adhesion to the Egg Zona Pellucida

Interaction of rapidly evolving molecules imparts species specificity to sperm-egg recognition in marine invertebrates, but it is unclear whether comparable interactions occur during fertilization in any vertebrate species. In mammals, the sperm acrosomal protein zonadhesin is a rapidly evolving molecule with species-specific binding activity for the egg zona pellucida (ZP). Here we show using null mice produced by targeted disruption of Zan that zonadhesin confers species specificity to sperm-ZP adhesion. Sperm capacitation selectively exposed a partial von Willebrand D domain of mouse zonadhesin on the surface of living, motile cells. Antibodies to the exposed domain inhibited adhesion of wild-type spermatozoa to the mouse ZP but did not inhibit adhesion of spermatozoa lacking zonadhesin. Zan^−/− males were fertile, and their spermatozoa readily fertilized mouse eggs in vitro. Remarkably, however, loss of zonadhesin increased adhesion of mouse spermatozoa to pig, cow, and rabbit ZP but not mouse ZP. We conclude that zonadhesin mediates species-specific ZP adhesion, and Zan^−/− males are fertile because their spermatozoa retain adhesion capability that is not species-specific. Mammalian sperm-ZP adhesion is therefore molecularly robust, and species-specific egg recognition by a protein in the sperm acrosome is conserved between invertebrates and vertebrates, even though the adhesion molecules themselves are unrelated.     

Concentration and Composition of Serum and Plasma Glycosaminoglycans in Domestic Animal Species

Acid glycosaminoglycans (GAGs) were isolated from serum and/or plasma of some domestic animals and the composition of the isolated GAG mixtures were studied. Mean values of total GAG concentration, in terms of hexuronic acid, ranged from a maximum of about 12 mg/l in serum of calves, trained horses and sheep to about 9 mg/l in serum of cows and donkeys and in plasma of trained horses to about 6.5 mg/l in sedentary horse serum and rabbit plasma to a minimum of 4 mg/l in dog serum and sedentary horse plasma. Statistically significant differences in total GAG concentrations (P < 0.0005) were found in horses between plasma and serum and also between sedentary and trained subjects. Chondroitin sulphate was the main component in serum and plasma GAG mixtures, accounting for 81–84% of total GAGs in the examined animals, except in cow serum (72%), trained horse plasma (75.5%) and sheep serum (87%). Keratan sulphate-like structures, measured as galactose, ranged from 12% in sheep serum to 17% in cattle serum. Fucose was associated with galactose in GAG fractions, which supports the hypothesis that articular cartilage is among the sites of origin of circulating GAGs.     

Affinity and Specificity of Levamlodipine-Human Serum Albumin Interactions: Insights into Its Carrier Function

The affinity and specificity of drugs with human serum albumin (HSA) are crucial factors influencing the bioactivity of drugs. To gain insight into the carrier function of HSA, the binding of levamlodipine with HSA has been investigated as a model system by a combined experimental and theoretical/computational approach. The fluorescence properties of HSA and the binding parameters of levamlodipine indicate that the binding is characterized by one binding site with static quenching mechanism, which is related to the energy transfer. As indicated by the thermodynamic analysis, hydrophobic interaction is the predominant force in levamlodipine-HSA complex, which is in agreement with the computational results. And the hydrogen bonds can be confirmed by computational approach between levamlodipine and HSA. Compared to predicted binding energies and binding energy spectra at seven sites on HSA, levamlodipine binding HSA at site I has a high affinity regime and the highest specificity characterized by the largest intrinsic specificity ratio (ISR). The binding characteristics at site I guarantee that drugs can be carried and released from HSA to carry out their specific bioactivity. Our concept and quantification of specificity is general and can be applied to other drug-target binding as well as molecular recognition of peptide-protein, protein-protein, and protein-DNA interactions.     

Pollinator Specificity and Convergence in Fly-pollinated Pleurothallis(Orchidaceae) Species: A Multiple Population Approach

We studied the floral biology of 12 populations of five rupicolousPleurothallis(Orchidaceae) species occurring in campo rupestre vegetationat nine localities in Brazil. All of these species are pollinatedby flies belonging to the families Chloropidae and Phoridae.In the five Pleurothallis species studied, all conspecific populationsattracted the same pollinator species. All pollinators werefemales; they laid eggs in flowers of the two nectarless species,but never in the flowers of nectar-presenting species. The twopairs ofPleurothallis species with similar flower morphologiesand odours attracted the same pollinators: P. johannensis -P. fabiobarrosii, pollinated by Tricimba sp. (Chloropidae) andP. teres - P. ochreata pollinated by Megaselia spp. (Phoridae).There was no overlap in the distribution of thePleurothallisspecies that shared pollinators. Despite similarities in floralmorphology and odour, genetic data show that these species pairsare not each other's closest relatives. We hypothesize thatthese similarities are due to convergence in allopatric speciesthat evolved similar pollination mechanisms. Conversely, thereare reasons to believe that adaptation to different pollinationmechanisms occurred in the closely related species P. johannensisand P. teres.Copyright 2001 Annals of Botany Company Pleurothallis, Orchidaceae, floral biology, fly-pollination, Chloropidae, Phoridae, pollinator specificity, campo rupestre