Genetic and biochemical analyses of Pfh1 DNA helicase function in fission yeast

The Schizosaccharomyces pombe pfh1⁺ gene (PIF1 homolog) encodes an essential enzyme that has both DNA helicase and ATPase activities and is implicated in lagging strand DNA processing. Mutations in the pfh1⁺ gene suppress a temperature-sensitive allele of cdc24⁺, which encodes a protein that functions with Schizosaccharomyces pombe Dna2 in Okazaki fragment processing. In this study, we describe the enzymatic properties of the Pfh1 helicase and the genetic interactions between pfh1 and cdc24, dna2, cdc27 or pol 3, all of which are involved in the Okazaki fragment metabolism. We show that a full-length Pfh1 fusion protein is active as a monomer. The helicase activity of Pfh1 displaced only short (<30 bp) duplex DNA regions efficiently in a highly distributive manner and was markedly stimulated by the presence of a replication-fork-like structure in the substrate. The temperature-sensitive phenotype of a dna2-C2 or a cdc24-M38 mutant was suppressed by pfh1-R20 (a cold-sensitive mutant allele of pfh1) and overexpression of wild-type pfh1⁺ abolished the ability of the pfh1 mutant alleles to suppress dna2-C2 and cdc24-M38. Purified Pfh1-R20 mutant protein displayed significantly reduced ATPase and helicase activities. These results indicate that the simultaneous loss-of-function mutations of pfh1⁺ and dna2⁺ (or cdc24⁺) are essential to restore the growth defect. Our genetic data indicate that the Pfh1 DNA helicase acts in concert with Cdc24 and Dna2 to process single-stranded DNA flaps generated in vivo by pol δ-mediated lagging strand displacement DNA synthesis.     

Relationships between biological soil crusts,bacterial diversity and abundance,and ecosystem functioning: Insights from a semi‐arid Mediterranean environment

Questions: To what degree do biological soil crusts (BSCs), which are regulators of the soil surface boundary, influence associated microbial communities? Are these associations important to ecosystem functioning in a Mediterranean semi‐arid environment? Location: Gypsum outcrops near Belmonte del Tajo, Central Spain. Methods: We sampled a total of 45 (50 cm × 50 cm) plots, where we estimated the cover of every lichen and BSC‐forming lichen species. We also collected soil samples to estimate bacterial species richness and abundance, and to assess different surrogates of ecosystem functioning. We used path analysis to evaluate the relationships between the richness/abundance of above‐ and below‐ground species and ecosystem functioning. Results: We found that the greatest direct effect upon the ecosystem function matrix was that of the biological soil crust (BSC) richness matrix. A few bacterial species were sensitive to the lichen community, with a disproportionate effect of Collema crispum and Toninia sedifolia compared to their low abundance and frequency. The lichens Fulgensia subbracteata and Toninia spp. also had negative effects on bacteria, while Diploschistes diacapsis consistently affected sensitive bacteria, sometimes positively. Despite these results, very few of the BSC effects on ecosystem function could be ascribed to changes within the bacterial community. Conclusion: Our results suggest the primary importance of the richness of BSC‐forming lichens as drivers of small‐scale changes in ecosystem functioning. This study provides valuable insights on semi‐arid ecosystems where plant cover is spatially discontinuous and ecosystem function in plant interspaces is regulated largely by BSCs.     

Retracted: Tusk shells in trouble? Physiology and behavior in response to changing temperature in a scaphopod (Mollusca: Scaphopoda: Dentaliida)

Scaphopods (tusk shells) are infaunal marine predators that occur at locally high densities in coastal and deep‐sea mud habitats, and as consumers of foraminifera they are important in carbon cycling. We investigated oxygen metabolism and burying behavior of the scaphopod Rhabdus rectius and its responses to altered temperatures. These are the first measurements of oxygen uptake rates for any member of this taxonomic class. In response to elevated temperatures, oxygen uptake rates increased, but the ability of animals to bury themselves in sediment was compromised. Female scaphopods were significantly larger than males and, when corrected for body mass, oxygen uptake rates were consistently higher for female individuals than for males. This is consistent with previous anecdotal observations of females in other scaphopod species being larger and potentially more active. In conditions of declining oxygen availability, individuals of Rhabdus rectius showed strong oxyregulatory ability by maintaining the same oxygen uptake rate displayed in normoxic conditions. The ability to maintain normal metabolic functioning even in conditions of oxygen limitation would benefit a species living in a benthic environment that may be prone to temporary or transient anoxic events. Yet the decrease in normal escape response in moderately elevated temperatures indicates these animals may be at risk from rising sea temperatures.     

Antibody fingerprints in lyme disease deciphered with high density peptide arrays

Lyme disease is the most common tick‐borne infectious disease in Europe and North America. Previous studies discovered the immunogenic role of a surface‐exposed lipoprotein (VlsE) of Borreliella burgdorferi. We employed high density peptide arrays to investigate the antibody response to the VlsE protein in VlsE‐positive patients by mapping the protein as overlapping peptides and subsequent in‐depth epitope substitution analyses. These investigations led to the identification of antibody fingerprints represented by a number of key residues that are indispensable for the binding of the respective antibody. This approach allows us to compare the antibody specificities of different patients to the resolution of single amino acids. Our study revealed that the sera of VlsE‐positive patients recognize different epitopes on the protein. Remarkably, in those cases where the same epitope is targeted, the antibody fingerprint is almost identical. Furthermore, we could correlate two fingerprints with human autoantigens and an Epstein‐Barr virus epitope; yet, the link to autoimmune disorders seems unlikely and must be investigated in further studies. The other three fingerprints are much more specific for B. burgdorferi. Since antibody fingerprints of longer sequences have proven to be highly disease specific, our findings suggest that the fingerprints could function as diagnostic markers that can reduce false positive test results.     

Signs of stabilisation and stable coexistence

Many empirical studies motivated by an interest in stable coexistence have quantified negative density dependence, negative frequency dependence, or negative plant–soil feedback, but the links between these empirical results and ecological theory are not straightforward. Here, we relate these analyses to theoretical conditions for stabilisation and stable coexistence in classical competition models. By stabilisation, we mean an excess of intraspecific competition relative to interspecific competition that inherently slows or even prevents competitive exclusion. We show that most, though not all, tests demonstrating negative density dependence, negative frequency dependence, and negative plant–soil feedback constitute sufficient conditions for stabilisation of two‐species interactions if applied to data for per capita population growth rates of pairs of species, but none are necessary or sufficient conditions for stable coexistence of two species. Potential inferences are even more limited when communities involve more than two species, and when performance is measured at a single life stage or vital rate. We then discuss two approaches that enable stronger tests for stable coexistence‐invasibility experiments and model parameterisation. The model parameterisation approach can be applied to typical density‐dependence, frequency‐dependence, and plant–soil feedback data sets, and generally enables better links with mechanisms and greater insights, as demonstrated by recent studies.     

Characterization of highly pathogenic H5N1 avian influenza A viruses isolated from South Korea

An unprecedented outbreak of H5N1 highly pathogenic avian influenza (HPAI) has been reported for poultry in eight different Asian countries, including South Korea, since December 2003. A phylogenetic analysis of the eight viral genes showed that the H5N1 poultry isolates from South Korea were of avian origin and contained the hemagglutinin and neuraminidase genes of the A/goose/Guangdong/1/96 (Gs/Gd) lineage. The current H5N1 strains in Asia, including the Korean isolates, share a gene constellation similar to that of the Penfold Park, Hong Kong, isolates from late 2002 and contain some molecular markers that seem to have been fixed in the Gs/Gd lineage virus since 2001. However, despite genetic similarities among recent H5N1 isolates, the topology of the phylogenetic tree clearly differentiates the Korean isolates from the Vietnamese and Thai isolates which have been reported to infect humans. A representative Korean isolate was inoculated into mice, with no mortality and no virus being isolated from the brain, although high titers of virus were observed in the lungs. The same isolate, however, caused systemic infections in chickens and quail and killed all of the birds within 2 and 4 days of intranasal inoculation, respectively. This isolate also replicated in multiple organs and tissues of ducks and caused some mortality. However, lower virus titers were observed in all corresponding tissues of ducks than in chicken and quail tissues, and the histological lesions were restricted to the respiratory tract. This study characterizes the molecular and biological properties of the H5N1 HPAI viruses from South Korea and emphasizes the need for comparative analyses of the H5N1 isolates from different countries to help elucidate the risk of a human pandemic from the strains of H5N1 HPAI currently circulating in Asia.