Distribution change in South African frogs

Range change is a common species response to global change. Comparing historical species distribution data with recent biological surveys has the potential to quantify changes to species geographic ranges. However, the broad-scale sampling strategies typically employed to acquire primary species distribution data are prone to errors of omission. The aim of this study was to evaluate the South African Frog Atlas Project (SAFAP) as a means for detecting changes in amphibian species distributions and to relate observed range changes to extrinsic environmental factors and intrinsic species characteristics. The SAFAP provided historical (1905 – 1995) and recent (1996 – 2003) species distributions of the amphibians of South Africa. Geographic sampling bias in the dataset was assessed by relating collection density and species richness to hypothesised sources of bias. Several methods for managing differing sampling intensity were tested on hypothetical ranges. The best methods were applied to the South African species to investigate range dynamics. Changes to the size of species ranges and shifts in mean range centre were assessed. An Ecological Niche Factor Analysis provided comparative measures of climate and habitat niche breadth for each species. SAFAP sampling was concentrated around cities, roads and protected areas, resulting in relatively overestimated species richness and range sizes near to these features. Large parts of the arid northwestern regions were under-sampled. An increase in sampling intensity over time resulted in the false detection of range expansions. The most reliable method to correct for increased sampling was a mathematical correction factor, according to which, 60.2% of South African frog species have undergone range contractions. Upslope shifts of 47.6 m were found for South African species and species of the Bushveld region shifted towards an area of Savanna Biome resilience. While several of the observed changes to species ranges were consistent with global change predictions, southern hemisphere amphibians may show a differing response to global change to that which is commonly predicted. Small range size, habitat specialisation and climate specialisation were significant predictors of range contractions for all species. Contracting habitat specialists were concentrated within two areas of endemism that also had high levels of land transformation. The use of methods that correct for sampling variation has allowed the SAFAP to be valuable in investigating species range change

Discovery of a Natural Microsporidian Pathogen with a Broad Tissue Tropism in Caenorhabditis elegans

International audienc

Discovery of a Natural Microsporidian Pathogen with a Broad Tissue Tropism in Caenorhabditis elegans.

Microbial pathogens often establish infection within particular niches of their host for replication. Determining how infection occurs preferentially in specific host tissues is a key aspect of understanding host-microbe interactions. Here, we describe the discovery of a natural microsporidian parasite of the nematode Caenorhabditis elegans that displays a unique tissue tropism compared to previously described parasites of this host. We characterize the life cycle of this new species, Nematocida displodere, including pathogen entry, intracellular replication, and exit. N. displodere can invade multiple host tissues, including the epidermis, muscle, neurons, and intestine of C. elegans. Despite robust invasion of the intestine very little replication occurs there, with the majority of replication occurring in the muscle and epidermis. This feature distinguishes N. displodere from two closely related microsporidian pathogens, N. parisii and N. sp. 1, which exclusively invade and replicate in the intestine. Comparison of the N. displodere genome with N. parisii and N. sp. 1 reveals that N. displodere is the earliest diverging species of the Nematocida genus. Over 10% of the proteins encoded by the N. displodere genome belong to a single species-specific family of RING-domain containing proteins of unknown function that may be mediating interactions with the host. Altogether, this system provides a powerful whole-animal model to investigate factors responsible for pathogen growth in different tissue niches

More than just a (red) list : Over a decade of using South Africa's threatened ecosystems in policy and practice

One of the stated applications of the IUCN Red List of Ecosystems (RLE) is to influence government policy and decision-making. We share 15 years' experience in integrating an independently developed indicator of ecosystem threat status into government policies and practice. South Africa's ecosystem threat status indicator was conceptualised in the early 2000s and progressed from a project-based indicator to listing of threatened ecosystems in terms of national legislation in 2011. We show the range of applications of the indicator, from its use as a headline indicator in the National Biodiversity Assessment to its role as a direct trigger for Environmental Impact Assessment. The strong link between threatened ecosystems and systematic conservation planning in South Africa also enabled ecosystem threat status to inform multi-sectoral development planning and decision-making. We show how bridging products, data availability, persistent mainstreaming and stakeholder engagement have encouraged the use of the indicator in government policy. The advantages and disadvantages of legislative listing are shared. Sound scientific foundations, combined with pragmatism, have provided a policy-relevant tool for focussing management on threatened ecosystems. We make active recommendations that will facilitate the policy uptake of the IUCN RLE in other countries.</p

<i>N</i>. <i>displodere</i> likely accesses non-intestinal tissues from the intestinal lumen.

<p>(a) <i>C</i>. <i>elegans</i> intestinal GFP expression strain ERT413 at 1 dpi stained with <i>N</i>. <i>displodere</i> rRNA FISH. Sporoplasms are seen inside and outside of the GFP-labeled intestine, in close proximity to the intestine, but never anterior to the posterior bulb (<i>PB</i>). Scale bar is 10 μm. (b) Exterior polar tubes associated with a spore were measured for <i>N</i>. <i>displodere</i> and <i>N</i>. <i>parisii</i> small spores. Each data point represents a measured polar tube, with the line and error bars showing the mean and SD of n = 40 for <i>N</i>. <i>displodere</i> and n = 41 for <i>N</i>. <i>parisii</i>. Note polar tubes of <i>N</i>. <i>displodere</i> and <i>N</i>. <i>parisii</i> were measured with separate techniques on separate occasions. The image (<i>right</i>) shows an <i>N</i>. <i>displodere</i> spore stained by Calcofluor white (CFW) with the associated polar tube stained by Concanavalin A-rhodamine (ConA). Scale bar is 10 μm. (c) The widths of GFP-labeled intestine from L3 larvae and young adults were measured in the anterior and posterior regions of the animal and halved to estimate the distance from the lumen to the basal lateral side of the intestine. The mean and SD from n = 14 L3 animals and n = 8 young adults are indicated (***p = 0.0002, two-tailed Mann-Whitney test). (d) The tissue distribution of invasion events of <i>N</i>. <i>displodere</i> infection (sporoplasms) was analyzed after 30 minutes of infection in L3 larvae versus young adults, and was calculated in the tissue-specific strains expressing GFP in the intestine (<i>left</i>) and muscle (<i>right</i>). Invasion events were calculated as the percent of FISH-stained sporoplams occurring outside of the GFP-expressing intestine (<i>left)</i> or inside the GFP-expressing muscle (<i>right</i>) compared to the total number of events throughout the animals. Data are represented as mean with SD of four replicates across two experiments, with a total of 25 animals counted for each replicate (*p = 0.0286, two-tailed Mann-Whitney test).</p

<i>N</i>. <i>displodere</i> spores exit through a bursting route.

<p>(a) Time course comparing the total number of internal spores compared to shed spores in <i>N</i>. <i>displodere</i>-infected (<i>left</i>) and <i>N</i>. <i>parisii</i>-infected (<i>right</i>) animals at 15°C. Note that only intact (non-burst) animals were picked for this assay. Internal spores indicate the average number of internal spores per animal, while external spores indicate the average number of spores shed by twenty animals into the media in four hours. Data points indicate the mean with SD of n = 6 replicates of 20 animals across 3 experiments for internal <i>N</i>. <i>displodere</i> spores and n = 4 replicates of 20 animals across 2 experiments for <i>N</i>. <i>displodere</i> shed spores and all <i>N</i>. <i>parisii</i> data. (b) Time course depicting the percent of animals with a burst vulva phenotype of uninfected, <i>N</i>. <i>displodere</i>-infected, and <i>N</i>. <i>parisii</i>-infected animals at 15°C. Data points depict the mean and SD from n = 4 independent experiments for uninfected and <i>N</i>. <i>displodere</i> and n = 3 experiments for <i>N</i>. <i>parisii</i> where each experiment consisted of triplicate samples containing at least 150 animals per replicate. (c) Image from a plate of wild-type <i>C</i>. <i>elegans</i> infected with <i>N</i>. <i>displodere</i> at 10 dpi. Indicated are adult animals with a burst vulva (<i>BV</i>) and internal organs spilling out. Image taken from a Nikon SMZ800 dissecting scope with an iPhone 5S. (d) Analysis of spores shed by late stage <i>N</i>. <i>displodere</i>-infected animals split into two groups, intact animals versus animals with a burst phenotype. Each data point indicates the number of spores shed by twenty animals for four hours of a single replicate, with the line and error bars showing the mean and SD of n = 4 replicates across two independent experiments (*p = 0.0211, two-tailed Mann-Whitney test; ns = not significant, p = 0.298).</p

Analysis and comparison of <i>N</i>. <i>displodere</i>, <i>N</i>. <i>parisii</i>, <i>and N</i>. sp. 1 genomes.

<p>(a) Phylogenomic tree of <i>N</i>. <i>displodere</i> and 18 other microsporidia genomes, with <i>Rozella allomycis</i> as an outgroup. Bootstrap support is indicated next to each node. Scale bar indicates changes per site. The tree was created with FigTree 1.4.2 (<a href="http://tree.bio.ed.ac.uk/software/figtree/" target="_blank">http://tree.bio.ed.ac.uk/software/figtree/</a>). (b) Histogram of intergenic region lengths of the three <i>Nematocida</i> species. (c) Comparison of protein content among the three <i>Nematocida</i> species. Proteins were classified into 7 categories: proteins shared with all <i>Nematocida</i> and at least 1 other non-microsporidian eukaryotic species (<i>eukaryotic</i>), proteins shared between all <i>Nematocida</i> and at least 1 other microsporidian species (<i>microsporidia</i>), proteins shared only between all the three <i>Nematocida</i> species (Nematocida), proteins shared by <i>N</i>. <i>displodere</i> and <i>N</i>. <i>parisii</i>, proteins shared by <i>N</i>. <i>displodere</i> and <i>N</i>. sp. 1, proteins shared by <i>N</i>. <i>parisii</i> and <i>N</i>. sp. 1, and proteins not in any other species (<i>unique</i>). (d) Protein schematic of a generalized member of each of the large gene families in the <i>Nematocida</i> species, which contain signal peptides (<i>SP</i>). The average size of the gene family and the number of proteins in each species are indicated at the right.</p

<i>N</i>. <i>displodere</i> infects multiple tissues but shows preferential proliferation in non-intestinal tissues.

<p>(a) The anterior region of a <i>C</i>. <i>elegans</i> animal co-infected with <i>N</i>. <i>displodere</i> (green) and <i>N</i>. <i>parisii</i> (red), visualized by FISH using species-specific rRNA probes and DAPI (blue). This image was captured by confocal microscopy with a single z-plane represented in the main inset, and orthogonal views of the x- and y-planes on the top and right insets, respectively, which show a cross-sectional view of the captured z-stacks within those planes. Scale bar is 50 μm. (b) <i>C</i>. <i>elegans</i> tissue-specific GFP-expression strains in the epidermis (<i>top</i>), body wall muscle (<i>middle</i>), and neurons (<i>bottom</i>), were infected with <i>N</i>. <i>displodere</i> and imaged at 3 dpi by FISH and DAPI. The neuron infected was in the ventral nerve cord (<i>bottom</i>). (c) Tissue-specific GFP strains were infected and imaged at 5 dpi with FISH and DY96 to stain clusters of spores (<i>Sp</i>). GFP-positive tissues that are difficult to see due to heavy infection are outlined with dashed white lines. The neuron infected was in the pre-anal ganglia (<i>bottom</i>). Scale bars are 20 μm. (d) The mid-body of the <i>C</i>. <i>elegans</i> intestinal GFP-expression strain infected with <i>N</i>. <i>displodere</i> at 1 dpi (<i>top</i>) and 3 dpi (<i>bottom</i>). Infection events are labeled as either inside (<i>in</i>) or outside (<i>out</i>) of the GFP-labeled intestine. Scale bar is 10 μm. (e) The tissue distribution of proliferating <i>N</i>. <i>displodere</i> infection was analyzed at 3 dpi, and was calculated individually in each <i>C</i>. <i>elegans</i> tissue-expression strain as the percent of FISH-stained meront clusters occurring in the GFP-positive tissues compared to the total number of events throughout the animals. Data are represented as the mean with SD of four replicates across two experiments, with a total of 50 animals counted for each replicate. (f) A comparison of the percent of animals infected in the specified GFP-positive tissue at three time points at which the three main stages of <i>N</i>. <i>displodere</i> infection occur, with sporoplasms analyzed at 1 dpi, meronts at 3 dpi, and new spores at 6 dpi. Each time point was calculated individually in each <i>C</i>. <i>elegans</i> tissue-expression strain as the percent of 50 animals that show a given symptom in the GFP-positive tissues. Data are represented as the mean with SD of four replicates across two experiments (ns = not significant, comparing intestine to muscle (p = 0.55) or intestine to epidermis (p = 0.11) at 1 dpi; *p = 0.03 comparing intestine to muscle and comparing intestine to epidermis at 6 dpi, two-tailed Mann-Whitney test).</p

A new microsporidian species that infects <i>C</i>. <i>elegans</i>.

<p>(a) Infected head region of a live <i>C</i>. <i>elegans</i> animal from strain JU2807 (derived from the wild-isolated P₀ animal, see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005724#ppat.1005724.s001" target="_blank">S1 Fig</a>) showing a large group of structures that appear to be meronts (<i>Me</i>) adjacent to the pharynx (<i>Ph</i>) and intestine (<i>In</i>). (b) Infected head region with the area adjacent to the pharynx filled with spores (<i>Sp</i>). (c) The mid-posterior to tail region of N2 <i>C</i>. <i>elegans</i> infected with <i>N</i>. <i>displodere</i> from 1 dpi to 7 dpi at 15°C visualized by FISH to stain parasite rRNA (red), DAPI to stain nuclei (blue), and DY96 to stain the chitin of parasite spore walls (green). Animals were at the L2 larval stage at 1 dpi, L3 stage at 2 dpi, L4 stage at 3 dpi, and adult stage at 4–7 dpi. Sporoplasms (<i>Sppl</i>), meronts (<i>Me</i>), sporonts (<i>Spnt</i>), and spores (<i>Sp</i>) are indicated. Scale bars are 10 μm. (d) Quantification of symptoms of <i>N</i>. <i>displodere</i> infection over time at 15°C with N2 animals infected as starved L1 larvae at T₀. Sporoplasms are mononucleated structures, meronts are multinucleated structures, and sporoblasts are rounded, mononucleated structures stained by FISH (see c above). Spores are oblong DY96-stained structures in infected animals. Fifty animals were quantified for each replicate at each time point, and data points indicate the mean and standard deviation (SD) from four replicates across two experiments. Each symptom was fit to a Boltzmann sigmoidal curve (R square > 0.99 for each curve), and the time to 50% of the animals exhibiting symptoms (T₅₀) is shown.</p

<i>N</i>. <i>displodere</i> induces an intestinal response, and host feeding is required for infection.

<p>(a) Normalized GFP induction after <i>N</i>. <i>displodere</i> or <i>N</i>. <i>parisii</i> infection of an intestinal infection reporter strain (ERT54 <i>F26F2</i>.<i>1p</i>::<i>GFP</i>, <i>left</i>), and an epidermal infection/cuticle damage reporter (AU189 <i>nlp-29p</i>::<i>GFP</i>, <i>col-12p</i>::<i>dsRed</i>, <i>right</i>), as measured by a COPAS Biosort. Experimental replicates were normalized by animal body size for ERT54 or by red fluorescence (<i>col12p</i>::<i>dsred</i>) for AU189. For ERT54, data are represented as mean values with SD from n = 882 animals from six replicates across two independent experiments (****p<0.0001, two-tailed Mann-Whitney test). For AU189, due to a batch effect, only data are shown from three replicates in one experiment, with mean values shown with SD from n = 900 animals (****p<0.0001, ns = not significant, two-tailed Mann-Whitney test). Data from the other AU189 replicates are shown in the supplement (<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005724#ppat.1005724.s004" target="_blank">S4 Fig</a>). (b) Comparison of <i>N</i>. <i>displodere</i> infection of <i>daf-2(ts)</i> animals at the L3 stage (maintained at 15°C) or <i>daf-2(ts)</i> animals induced to form dauer larvae (maintained at 25°C). As controls, N2 animals were maintained at 15°C and infected with <i>N</i>. <i>displodere</i> as L3 animals at 25°C, and <i>N</i>. <i>parisii</i> spores were used to infect <i>daf-2(ts)</i> dauer larvae. (c) Comparison of the number of invasion events (counted as sporoplasms) occurring in N2 and <i>eat-2</i> animals at 1 dpi. Events were counted as either intestinal (co-localizing with intestinal gut) or non-intestinal. Data are represented as mean values with SD from n = 75 animals from three independent experiments (****p<0.0001, two-tailed Mann-Whitney test). (d) Comparison of the number of invasion events (counted as sporoplasms) occurring in <i>dyn-1(ts)</i> and N2 animals at 30°C for 30 minutes for <i>N</i>. <i>displodere</i> (<i>left</i>) and <i>N</i>. <i>parisii</i> infection (<i>right</i>). Infection events were distinguished as either intestinal or non-intestinal as above. <i>dyn-1(ts)</i> animals are paralyzed and cease to feed at the non-permissive temperature (30°C). Data are represented as mean values with SD from n = 80 <i>dyn-1(ts)</i> animals and n = 50 N2 animals across two independent experiments (****p<0.0001, two-tailed Mann-Whitney test).</p