Low local but high beta diversity of tropical forest dung beetles in Madagascar

Aims We have compared local (alpha) and regional (beta) species diversities of dung beetles in wet forests in the main tropical regions including Madagascar. Madagascar is exceptional in lacking native large herbivorous mammals which produce the key resource for dung beetles elsewhere. Location Central and South America, mainland Africa, Madagascar and Southeast Asia. Methods Trapping data on dung beetles and data on mammalian faunas were obtained from published and unpublished studies. We used our original data for Madagascar. Results Species richness of dung beetles and that of large‐bodied (> 15 mm length) species in particular were highly significantly explained by the regional number of large‐bodied (> 10 kg) mammals (R² from 50 to 80%). For a given pairwise spatial distance between two communities, beta diversity was significantly higher in Madagascar than elsewhere, explaining the very high total species richness in Madagascar in spite of low local diversity. Main conclusion The presence and numbers of large herbivorous mammals greatly influence the species richness of dung beetles in tropical wet forests. The lack of native large herbivores rather than a limited species pool explains the low local diversity in Madagascar. Exceptionally high beta diversity in Madagascar suggests a pattern of old radiation involving extensive allopatric speciation.     

Orthogonal protein purification--expanding the repertoire of GST fusion systems

We have previously developed a labeling scheme that can be used to site-specifically link human glutathione transferases (hGSTs) from the alpha class to chemical entities such as fluorophores and aldehydes. The reagents are in-house synthesized derivatives of glutathione (GS-derivatives). We have focused on a lysine mutant of hGST A1:A216K. In this study, we wanted to utilize these findings and improve on protein purification schemes that are using GSTs as fusion partners. We have used random mutagenesis to scramble the hydrophobic binding site of A216K through mutations at position M208 and isolated a library of 11 A216K/M208X mutants. All mutants were easily expressed and purified and retained all or parts of the catalytic properties of the parent GST. The mutants were stable over several days at room temperature. The A216K/M208X mutants could be site-specifically labeled using our designed fluorescent reagents. Furthermore, reaction with an aldehyde-containing reagent termed GS-Al results in site-specific introduction of an orthogonal handle for subsequent conjugation with aldehyde-reactive probes. Labeling with coumarin results in a fluorescent protein-conjugate that can bind glutathione (GSH) derivatives for subsequent affinity purification. The K(d) for S-hexyl-GSH of coumarin-labeled A216K was measured to be 2.5 microM. The candidate proteins A216K and A216K/M208F could be purified in high yield in a one-step procedure through affinity chromatography (Glutathione Sepharose 4B). The proteins can readily be perceived as improved GST fusion partners.     

Use of CFSE staining of borreliae in studies on the interaction between borreliae and human neutrophils

Background  

Species of the tick-transmitted spirochete group Borrelia burgdorferi sensu lato (B. burgdorferi) cause Lyme borreliosis. Acute borrelial infection of the skin has unusual characteristics with only a mild local inflammatory response suggesting that the interaction between borreliae and the cells of the first-line defence might differ from that of other bacteria. It has been reported that human neutrophils phagocytose motile borreliae through an unconventional mechanism (tube phagocytosis) which is not observed with non-motile borreliae. Therefore, it would be of great interest to visualise the bacteria by a method not affecting motility and viability of borreliae to be able to study their interaction with the cells of the innate immunity. Carboxyfluorescein diacetate, succinimidyl ester (CFSE) labelling has been previously used for studying the adhesion of labelled bacteria to host cells and the uptake of labelled substrates by various cells using flow cytometry.     

Evaluation of a pumping system for sampling zooplankton

This study describes the sampling efficiency of a plankton pumpdesigned to operate from a moving research vessel. The samplingefficiency of the pump was evaluated by comparing the catchper unit volume of several planktonic taxa sampled simultaneouslywith the pump and with a modified Gulf V plankton sampler ora tube sampler. The estimates of plankton density obtained withthe pump sampler corresponded well with those obtained withthe modified plankton net and tube sampler. We conclude thatthe use of a plankton pump in zooplankton sampling gives resultscomparable to those found with traditional sampling, exceptthat the results were biased for the sampling of rotifers andcopepod eggs.     

The susceptibility of conjugative resistance transfer in Gram-negative bacteria to physicochemical and biochemical agents

Abstract Over thirty years of studies have established that conjugative transfer of plasmid-encoded resistance to drugs and heavy metals can take place at high frequency between various organisms under laboratory conditions. The detected transfer frequencies in soil, in aquatic environments, and in the urogenital and respiratory tracts of healthy animals and man have generally been low. However, the conversion of bacteria from susceptible to resistant to antibiotics has been observed often during antimicrobial therapy. This has formed a challenge for the antibacterial treatment of pathogenic bacteria and called for the evaluation of the extent of conjugative transfer in various environments. Several biochemical and physicochemical factors inhibit conjugation, show preferential toxicity against plasmid-bearing cells, or stimulate plasmid curing. These factors include various agents such as detergents, anesthetics, mutagens and antibiotics which affect membrane potential, membrane permeability, protein synthesis and the processing of DNA. The application of the data on these agents, summarized in this review, might be helpful in preventing drug multi-resistance from spreading. Also these data might be valuable in studies which use conjugation as a tool or which treat the molecular mechanisms involved in conjugation.     

The susceptibility of conjugative resistance transfer in gram-negative bacteria to physicochemical and biochemical agents

Over thirty years of studies have established that conjugative transfer of plasmid-encoded resistance to drugs and heavy metals can take place at high frequency between various organisms under laboratory conditions. The detected transfer frequencies in soil, in aquatic environments, and in the urogenital and respiratory tracts of healthy animals and man have generally been low. However, the conversion of bacteria from susceptible to resistant to antibiotics has been observed often during antimicrobial therapy. This has formed a challenge for the antibacterial treatment of pathogenic bacteria and called for the evaluation of the extent of conjugative transfer in various environments. Several biochemical and physicochemical factors inhibit conjugation, show preferential toxicity against plasmid-bearing cells, or stimulate plasmid curing. These factors include various agents such as detergents, anesthetics, mutagens and antibiotics which affect membrane potential, membrane permeability, protein synthesis and the processing of DNA. The application of the data on these agents, summarized in this review, might be helpful in preventing drug multi-resistance from spreading. Also these data might be valuable in studies which use conjugation as a tool or which treat the molecular mechanisms involved in conjugation.     

Lack of correlation between calcium mobilization and respiratory burst activation induced by chemotactic factors in rabbit polymorphonuclear leukocytes

Low concentrations of FMLP, partially purified rabbit C5a, leukotriene B4 and platelet activating factor induced a rapid rise of intracellular free Ca2+ in rabbit polymorphonuclear leukocytes. However, the four factors differed markedly in their ability to activate the respiratory burst. The peptides FMLP and C5a induced a single, strong chemiluminescence response whereas the lipids leukotriene B4 and platelet activating factor induced a markedly less intense response with a two-peak profile. Respiratory burst activation by the peptides was dependent on extracellular Ca2+ whereas the lipids required both Mg2+ and Ca2+. The results indicate that mobilization of intracellular Ca2+ is insufficient by itself to induce respiratory burst activation and that the intracellular pathways leading to activation differ depending on the nature of the stimulus.     

Susceptibility of gram-negative bacteria to the synergistic bactericidal action of serum and polymyxin B nonapeptide

Polymyxin B nonapeptide was able to sensitize Escherichia coli strains and strains of Salmonella typhimurium, Klebsiella spp., Enterobacter cloacae, Pseudomonas aeruginosa, and Haemophilus influenzae to the bactericidal action of fresh normal human serum. The degree of sensitization varied significantly within the strains. Strains of Proteus mirabilis, Neisseria gonorrhoeae, and N. meningitidis remained resistant.