DNase I and Proteinase K eliminate DNA from injured or dead bacteria but not from living bacteria in microbial reference systems and natural drinking water biofilms for subsequent molecular biology analyses

Molecular techniques, such as polymerase chain reaction (PCR) and quantitative PCR (qPCR), are very sensitive, but may detect total DNA present in a sample, including extracellular DNA (eDNA) and DNA coming from live and dead cells. DNase I is an endonuclease that non-specifically cleaves single- and double-stranded DNA. This enzyme was tested in this study to analyze its capacity of digesting DNA coming from dead cells with damaged cell membranes, leaving DNA from living cells with intact cell membranes available for DNA-based methods. For this purpose, an optimized DNase I/Proteinase K (DNase/PK) protocol was developed. Intact Staphylococcus aureus cells, heat-killed Pseudomonas aeruginosa cells, free genomic DNA of Salmonella enterica, and a mixture of these targets were treated according to the developed DNase/PK protocol. In parallel, these samples were treated with propidium monoazide (PMA) as an already described assay for live-dead discrimination. Quantitative PCR and PCR-DGGE of the eubacterial 16S rDNA fragment were used to test the ability of the DNase/PK and PMA treatments to distinguish DNA coming from cells with intact cell membranes in the presence of DNA from dead cells and free genomic DNA. The methods were applied to three months old autochthonous drinking water biofilms from a pilot facility built at a German waterworks. Shifts in the DNA patterns observed after DGGE analysis demonstrated the applicability of DNase/PK as well as of the PMA treatment for natural biofilm investigation. However, the DNase/PK treatment demonstrated some practical advantages in comparison with the PMA treatment for live/dead discrimination of bacterial targets in drinking water systems.     

Strategic Management of an Invasive Ant‐scale Mutualism Enables Recovery of a Threatened Tropical Tree Species

Mutualisms between invasive ants and honeydew‐producing insects can have widespread negative effects on natural ecosystems. This is becoming an increasingly serious problem worldwide, causing certain ecosystems to change radically. Management of these abundant and influential mutualistic species is essential if the host ecosystem is to recover to its former non‐invaded status. This negative effect is particularly prevalent on some tropical islands, including Cousine Island, Seychelles. On this island, the invasive ant Pheidole megacephala has caused serious indirect damage to the threatened native Pisonia grandis trees via a mutualism with an invasive scale insect, Pulvinaria urbicola. We aimed to suppress the ant, thereby decoupling the mutualism and enabling recovery of the Pisonia trees. We treated all areas where ant pressure was high with a selective formicidal bait, which was deployed in custom‐made bait stations designed to avoid risk of treatment to endemic fauna. In the treated area, ant foraging activity was reduced by 93 percent and was followed by a 100 percent reduction in scale insect density. Abundance of endemic herbivorous insects and herbivorous activity increased significantly, however, after the decline in mutualistic species densities. Despite the native herbivore increase, there was considerable overall improvement in Pisonia shoot condition and an observed increase in foliage density. Our results demonstrate the benefit of strategic management of highly mutualistic alien species to the native Pisonia trees. It also supports the idea that area‐wide suppression is a feasible alternative to eradication for achieving positive conservation management at the level of the forest ecosystem.     

A plant‐traits approach to assessing the success of alien weed species in irrigated Mediterranean orchards

In addition to their impact on natural habitats, invasive alien plants can have a significant negative effect on agricultural systems and cause economic losses. Flood‐irrigated orchards in the Mediterranean Basin are vulnerable to the invasion of alien weeds, primarily because of the traditional management practices used in the orchards, which are characterized by high soil moisture during the dry summer period, nutrient availability and high levels of disturbance. This study sought to determine whether their biological traits can explain the success of alien weed species. To answer this question, 408 floristic relevés were conducted in 136 flood‐irrigated orchards on the Plains of Lleida (Catalonia, NE of Spain). Richness and cover of native and alien weeds were compared. Furthermore, a set of biological traits were compared between successful and non‐successful weeds for the whole data and separately between native and alien weeds using logistic regression and classification trees. In flood‐irrigated orchards, alien species covered most of their area, even though the richness of alien species was lower than that of the native species. The most important species were C4 species with seeds dispersed by water, and on the other hand, rosulate and caespitose‐reptant hemicryptophytes with long flowering period. Most of these traits fitted with those of the invasive alien weeds, which were mostly C4 species with seeds dispersed by water. Perennial life form characterized successful native weeds. In this study, we discuss how the traditional management of flood irrigation in fruit‐tree orchards favours invasive alien weeds that have specific traits, acting as a reservoir for the spread of alien weeds into other crops and surrounding riparian habitats. We also propose changing management practices in order to avoid the selection of alien weeds and to promote native species.     

Microbial analysis of cucumbers (Cucumis sativus) produced with tap or treated waste water

There is increasing evidence for a significant role of fruits and vegetables in infectious diseases in humans. Their consumption is increasing and environmental factors such as water availability are impacting their production. In this study, adding fertiliser to tap water (TW) increased the microbial load above that found in treated waste water (TWW); coliforms were also introduced. Low numbers of Bacillus spp. were recovered from inside some healthy cucumber fruits. No visible differences were observed between cucumber plants irrigated with TWW or TW or cucumbers with and without endophytic Bacillus spp. This is noteworthy when considering the use of TWW for crop irrigation.     

Food constraints explain the restricted distribution of wintering Lesser White‐fronted Geese Anser erythropus in China

More than 90% of the Lesser White‐fronted Geese Anser erythropus in the Eastern Palearctic flyway population winter at East Dongting Lake, China. To explain this restricted distribution and to understand better the winter feeding ecology and habitat requirements of this poorly known species, we assessed their food availability, diet and energy budgets at this site through two winters. Lesser White‐fronted Geese maintained a positive energy budget when feeding on above‐ground green production of Eleocharis and Alopecurus in recessional grasslands in autumn and spring to accumulate fat stores. Such food was severely depleted by late November and showed no growth in mid‐winter. Geese fed on more extensive old‐growth Carex sedge meadows in mid‐winter where they were in energy deficit and depleted endogenous fat stores. Geese failed to accumulate autumn fat stores in one year when high water levels prevented the Geese from using recessional grassland feeding areas. Fat stores remained lower throughout that winter and Geese left for breeding areas later in spring than in the previous year, perhaps reflecting the need to gain threshold fat stores for migration. Sedge meadows are widespread at other Yangtze River floodplain wetlands, but recessional grasslands are rare and perhaps restricted to parts of East Dongting Lake, which would explain the highly localized distribution of Lesser White‐fronted Geese in China and their heavy use of these habitats at this site. Sympathetic management of water tables is essential to maintain the recessional grasslands in the best condition for Geese. Regular depletion of fat stores whilst grazing sedge meadows in mid‐winter also underlines the need to protect the species from unnecessary anthropogenic disturbances that enhance energy expenditure. The specialized diet of the Lesser White‐fronted Goose may explain its highly restricted winter distribution and global rarity.     

Conservation implications of past and present nesting habitat selection of the endangered Osprey Pandion haliaetus population of the Canary Islands

We studied nesting habitat selection of the endangered non‐migratory Osprey Pandion haliaetus population of the Canary Islands and evaluated the effect of human expansion in recent decades. Compared with randomly selected potential nest‐sites, Osprey nests were more frequently found on taller, southwest‐facing cliffs, characterized by lower human pressure and closer to Yellow‐legged Gull Larus michahellis colonies and Barbary Falcon Falco pelegrinoides breeding sites. Furthermore, changes in some breeding habitat features have been detected in recent decades. According to our predictive models, large areas of suitable habitat are available but unoccupied in the Canaries, and human activities are probably limiting the settlement and dispersion of new pairs.     

A comparison of pedigree‐ and DNA‐based measures for identifying inbreeding depression in the critically endangered Attwater's Prairie‐chicken

The primary goal of captive breeding programmes for endangered species is to prevent extinction, a component of which includes the preservation of genetic diversity and avoidance of inbreeding. This is typically accomplished by minimizing mean kinship in the population, thereby maintaining equal representation of the genetic founders used to initiate the captive population. If errors in the pedigree do exist, such an approach becomes less effective for minimizing inbreeding depression. In this study, both pedigree‐ and DNA‐based methods were used to assess whether inbreeding depression existed in the captive population of the critically endangered Attwater's Prairie‐chicken (Tympanuchus cupido attwateri), a subspecies of prairie grouse that has experienced a significant decline in abundance and concurrent reduction in neutral genetic diversity. When examining the captive population for signs of inbreeding, variation in pedigree‐based inbreeding coefficients (f_pedigree) was less than that obtained from DNA‐based methods (f_DNA). Mortality of chicks and adults in captivity were also positively correlated with parental relatedness (r_DNA) and f_DNA, respectively, while no correlation was observed with pedigree‐based measures when controlling for additional variables such as age, breeding facility, gender and captive/release status. Further, individual homozygosity by loci (HL) and parental r_DNA values were positively correlated with adult mortality in captivity and the occurrence of a lethal congenital defect in chicks, respectively, suggesting that inbreeding may be a contributing factor increasing the frequency of this condition among Attwater's Prairie‐chickens. This study highlights the importance of using DNA‐based methods to better inform management decisions when pedigrees are incomplete or errors may exist due to uncertainty in pairings.     

Using multilevel models to identify drivers of landscape‐genetic structure among management areas

Landscape genetics offers a powerful approach to understanding species' dispersal patterns. However, a central obstacle is to account for ecological processes operating at multiple spatial scales, while keeping research outcomes applicable to conservation management. We address this challenge by applying a novel multilevel regression approach to model landscape drivers of genetic structure at both the resolution of individuals and at a spatial resolution relevant to management (i.e. local government management areas: LGAs) for the koala (Phascolartos cinereus) in Australia. Our approach allows for the simultaneous incorporation of drivers of landscape‐genetic relationships operating at multiple spatial resolutions. Using microsatellite data for 1106 koalas, we show that, at the individual resolution, foliage projective cover (FPC) facilitates high gene flow (i.e. low resistance) until it falls below approximately 30%. Out of six additional land‐cover variables, only highways and freeways further explained genetic distance after accounting for the effect of FPC. At the LGA resolution, there was significant variation in isolation‐by‐resistance (IBR) relationships in terms of their slopes and intercepts. This was predominantly explained by the average resistance distance among LGAs, with a weaker effect of historical forest cover. Rates of recent landscape change did not further explain variation in IBR relationships among LGAs. By using a novel multilevel model, we disentangle the effect of landscape resistance on gene flow at the fine resolution (i.e. among individuals) from effects occurring at coarser resolutions (i.e. among LGAs). This has important implications for our ability to identify appropriate scale‐dependent management actions.     

The impact of strain diversity and mixed infections on the evolution of resistance to Bacillus thuringiensis

Pesticide mixtures can reduce the rate at which insects evolve pesticide resistance. However, with live biopesticides such as the naturally abundant pathogen Bacillus thuringiensis (Bt), a range of additional biological considerations might affect the evolution of resistance. These can include ecological interactions in mixed infections, the different rates of transmission post-application and the impact of the native biodiversity on the frequency of mixed infections. Using multi-generation selection experiments, we tested how applications of single and mixed strains of Bt from diverse sources (natural isolates and biopesticides) affected the evolution of resistance in the diamondback moth, Plutella xylostella, to a focal strain. There was no significant difference in the rate of evolution of resistance between single and mixed-strain applications although the latter did result in lower insect populations. The relative survivorship of Bt-resistant genotypes was higher in the mixed-strain treatment, in part owing to elevated mortality of susceptible larvae in mixtures. Resistance evolved more quickly with treatments that contained natural isolates, and biological differences in transmission rate may have contributed to this. Our data indicate that the use of mixtures can have unexpected consequences on the fitness of resistant and susceptible insects.     

Ion transport through dimethyl sulfoxide (DMSO) induced transient water pores in cell membranes

Abstract

It is well known that dimethyl sulphoxide (DMSO) increases membrane permeability, which makes it widely used as a vehicle to facilitate drug delivery across biological membranes. However, the mechanism of how DMSO increases membrane permeability has not been well understood. Recently, molecular dynamics simulations have demonstrated that DMSO can induce water pores in biological membranes, but no direct experimental evidence is so far available to prove the simulation result. Using FluxOR Tl⁺ influx assay and intracellular Ca²⁺ imaging technique, we studied the effect of DMSO on Tl⁺ and Ca²⁺ permeation across cell membranes. Upon application of DMSO on CHO-K1 cell line, Tl⁺ influx was transiently increased in a dose-dependent manner. The increase in Tl⁺ permeability induced by DMSO was not changed in the presence of blockers for K⁺ channel and Na⁺-K⁺ ATPase, suggesting that Tl⁺ permeates through transient water pores induced by DMSO to enter into the cell. In addition, Ca²⁺ permeability was significantly increased upon application of DMSO, indicating that the transient water pores induced by DMSO were non-selective pores. Furthermore, similar results could be obtained from RAW264.7 macrophage cell line. Therefore, this study provided experimental evidence to support the prediction that DMSO can induce transient water pores in cell membranes, which in turn facilitates the transport of active substances across membranes.