Designing probiotic therapies with broad-spectrum activity against a wildlife pathogen

Host-associated microbes form an important component of immunity that protect against infection by pathogens. Treating wild individuals with these protective microbes, known as probiotics, can reduce rates of infection and disease in both wild and captive settings. However, the utility of probiotics for tackling wildlife disease requires that they offer consistent protection across the broad genomic variation of the pathogen that hosts can encounter in natural settings. Here we develop multi-isolate probiotic consortia with the aim of effecting broad-spectrum inhibition of growth of the lethal amphibian pathogen Batrachochytrium dendrobatidis (Bd) when tested against nine Bd isolates from two distinct lineages. Though we achieved strong growth inhibition between 70 and 100% for seven Bd isolates, two isolates appeared consistently resistant to inhibition, irrespective of probiotic strategy employed. We found no evidence that genomic relatedness of the chytrid predicted similarity of inhibition scores, nor that increasing the genetic diversity of the bacterial consortia could offer stronger inhibition of pathogen growth, even for the two resistant isolates. Our findings have important consequences for the application of probiotics to mitigate wildlife diseases in the face of extensive pathogen genomic variation

Amphibian skin defences show variation in ability to inhibit growth of Batrachochytrium dendrobatidis isolates from the Global Panzootic Lineage

The fungal pathogen Batrachochytrium dendrobatidis has caused declines and extinctions in hundreds of amphibian species across the world. Virulence varies among and within lineages; the Global Panzootic Lineage (GPL) is the most pathogenic, although there is also variation in lethality between GPL isolates. Amphibians have a number of defences against pathogens, and skin products including the microbiota and host peptides have been shown to have considerable influence over disease progression. Here we show the collective skin products (the mucosome) of two amphibian species show significant variation in their ability to inhibit different globally- distributed isolates of GPL. This may in part explain the variation in disease susceptibility of hosts to different strains of Batrachochytrium dendrobatidis. More work is required to identify particular traits associated with mucosomes that confer broad- spectrum inhibition across GPL in order to facilitate the development of prophylaxis and/or treatments for chytridiomycosis in situ

The effect of different UV and calcium provisioning on health and fitness traits of red-eyed tree frogs (Agalychnis callidryas)

In response to global amphibian declines and extinctions, the IUCN has recommended the establishment of ex situ conservation breeding programmes. However, there are a limited number of studies that scientifically assess amphibian husbandry practices, even at a basic level of nutrition and lighting. One component of captive husbandry that is increasingly discussed is the provision of ultraviolet radiation (UVR), which is required for the synthesis of vitamin D3 and subsequent assimilation of calcium and phosphorous from the diet. Here we used two methods of UV provision (“background UV” and “background UV with UV boost”) and two calcium gut-loading diets (5% and 10%) to assess the effects on a range of fitness measures in the red-eyed tree frog (Agalychnis callidryas). We found no effects of either UV treatment or calcium diet on growth, body condition or cutaneous bacterial communities of frogs, although subsequent to the UV boost, frogs had a significantly greater fungal load in comparison to frogs that were not UV-boosted. There were negligible differences in the breeding success of females according to UV exposure. Provision of the UV boost was not demonstrated to provide any real advantages for A. callidryas in terms of growth or breeding success. In addition, there were no benefits of a 10% calcium diet over a 5% calcium diet (in conjunction with regular dusting). Further studies that investigate the UV requirements of other amphibian species and ecotypes are required, particularly in conjunction with naturalistic cricket gut-loading diets

Mixed-cropping between field pea varieties alters root bacterial and fungal communities

Modern agricultural practices have vastly increased crop production but negatively affected soil health. As such, there is a call to develop sustainable, ecologically-viable approaches to food production. Mixed-cropping of plant varieties can increase yields, although impacts on plant-associated microbial communities are unclear, despite their critical role in plant health and broader ecosystem function. We investigated how mixed-cropping between two field pea (Pisum sativum L.) varieties (Winfreda and Ambassador) influenced root-associated microbial communities and yield. The two varieties supported significantly different fungal and bacterial communities when grown as mono-crops. Mixed-cropping caused changes in microbial communities but with differences between varieties. Root bacterial communities of Winfreda remained stable in response to mixed-cropping, whereas those of Ambassador became more similar to Winfreda. Conversely, root fungal communities of Ambassador remained stable under mixed-cropping, and those of Winfreda shifted towards the composition of Ambassador. Microbial co-occurrence networks of both varieties were stronger and larger under mixed-cropping, which may improve stability and resilience in agricultural soils. Both varieties produced slightly higher yields under mixed-cropping, although overall Ambassador plants produced higher yields than Winfreda plants. Our results suggest that variety diversification may increase yield and promote microbial interactions

Genetic diversity of honeybee colonies predicts gut bacterial diversity of individual colony members.

The gut microbiota of social bees is relatively simple and dominated by a set of core taxa found consistently in individuals around the world. Yet, variation remains, and can affect host health. We characterised individual- and regional-scale variation in honeybee (Apis mellifera) gut microbiota from 64 colonies in North-West England by sequencing the V4 region of the 16S rRNA gene, and asked whether microbiota were influenced by host genotype and landscape composition. We also characterised the genotypes of individual bees and the land cover surrounding each colony. The literature-defined core taxa dominated across the region despite the varied environments. However, there was variation in the relative abundance of core taxa, and colony membership explained much of this variation. Individuals from more genetically diverse colonies had more diverse microbiotas, but individual genetic diversity did not influence gut microbial diversity. There were weak trends for colonies in more similar landscapes to have more similar microbiota, and for bees from more urban landscapes to have less diverse microbiota. To our knowledge, this is the first report for any species that the gut bacterial communities of individuals are influenced by the genotypes of others in the population. This article is protected by copyright. All rights reserved

Making amphibian conservation more effective

Amphibians face an extinction crisis. Hundreds of species may be lost as conservation scientists and practitioners struggle to identify remedies to poorly understood declines spanning several decades. Due to various life history characteristics and a range of drivers, amphibians continue to be especially hard- hit, more so than any other vertebrate group. In this special issue of Conservation Evidence, studies that report the effectiveness of amphibian conservation interventions are presented to add to the rapidly growing body of literature on this topic. We here summarise the current understanding of global amphibian declines to highlight the importance of applying evidence-based strategies to amphibian conservation

Nitrogen addition alters composition, diversity, and functioning of microbial communities in mangrove soils : an incubation experiment

Mangrove ecosystems are important for carbon storage due to their high productivity and low decomposition rates. Waterways have experienced increased nutrient loads as a result of anthropogenic activities and it is unclear how this may affect carbon and nutrient cycles in downstream mangroves that receive these nutrient-rich waters. Using a laboratory-based incubation experiment, this study aimed to assess the effects of nutrient addition on the diversity and structure of mangrove soil bacterial communities, as well as biomass and activity of the soil microbial community, under different oxygen conditions. Bacterial community diversity and composition was characterised using 16S rRNA gene sequencing and microbial activity was examined through the measurement of microbial respiration and the activities of enzymes associated with organic matter decomposition. Nitrogen addition caused clear shifts in bacterial community composition, with decreases in bacterial diversity and the abundance of sulfate-reducing bacteria. Microbial biomass also decreased with nitrogen addition under reduced oxygen incubations. Changes in bacterial community structure were accompanied by changes in the activity of some enzymes involved in carbon, nitrogen and phosphorus cycling. Under reduced oxygen conditions, nitrogen addition resulted in a significant increase in the microbial metabolic quotient but no accompanying change in microbial respiration, which was explained by a decrease in microbial biomass. The findings of this study indicate that nitrogen loading has potential implications for microbial communities and carbon and nutrient cycling in mangrove environments that warrant further investigation under field conditions

Manipulation of the calcium content of insectivore diets through supplementary dusting

Insects fed to captive insectivores are deficient in calcium with inverse calcium to phosphorous ratios (Ca:P), and supplementation is required to avoid nutritional metabolic bone disease (NMBD). One method of improving the nutritional value of feeder insects is by “dusting” with powdered supplements, although it is often suggested that these are rapidly shed from prey insects. Here we analysed the calcium content of hatchling, second, fourth and adult instars of black field crickets and silent crickets at increasing time intervals after dusting, as well as comparing three commercially available brands of supplement in fourth instar black field crickets. Our data show these brands do not differ from one another in terms of calcium delivery, despite differences in calcium content. We also show that dusting can be used to increase Ca:P ratios above 1:1 in crickets up to 5.5 hours after dusting, with the exception of adult black field crickets, and thus dusting is a useful method of calcium supplementation

Tagging frogs with passive integrated transponders causes disruption of the cutaneous bacterial community and proliferation of opportunistic fungi

Symbiotic bacterial communities play a key role in protecting amphibians from infectious diseases including chytridiomycosis,caused by the pathogenic fungus Batrachochytrium dendrobatidis. Events that lead to the disruption of the bacterial community may have implications for the susceptibility of amphibians to such diseases. Amphibians are often marked both in the wild and in captivity for a variety of reasons, and although existing literature indicates that marking techniques have few negative effects, the response of cutaneous microbial communities has not yet been investigated. Here we determine the effects of passive integrated transponder (PIT) tagging on culturable cutaneous microbial communities of captive Morelet’s tree frogs (Agalychnis moreletii) and assess the isolated bacterial strains for anti-B. dendrobatidis activity in vitro. We find that PIT tagging causes a major disruption to the bacterial community associated with the skin of frogs (∼12-fold increase in abundance), as well as a concurrent proliferation in resident fungi (up to ∼200-fold increase). Handling also caused a disruption the bacterial community, although to a lesser extent than PIT tagging. However, the effects of both tagging and handling were temporary, and after 2 weeks, the bacterial communities were similar to their original compositions. We also identify two bacterial strains that inhibit B. dendrobatidis, one of which increased in abundance on PIT-tagged frogs at 1 day postmarking, while the other was unaffected. These results show that PIT tagging has previously unobserved consequences for cutaneous microbial communities of frogs and may be particularly relevant for studies that intend to use PIT tagging to identify individuals involved in trials to develop probiotic treatments

Evidence for the genetic similarity rule at an expanding mangrove range limit

Premise Host-plant genetic variation can shape associated communities of organisms. These community-genetic effects include (1) genetically similar hosts harboring similar associated communities (i.e., the genetic similarity rule) and (2) host-plant heterozygosity increasing associated community diversity. Community-genetic effects are predicted to be less prominent in plant systems with limited genetic variation, such as those at distributional range limits. Yet, empirical evidence from such systems is limited. Methods We sampled a natural population of a mangrove foundation species (Avicennia germinans) at an expanding range limit in Florida, USA. We measured genetic variation within and among 40 host trees with 24 nuclear microsatellite loci and characterized their foliar endophytic fungal communities with internal transcribed spacer (ITS1) gene amplicon sequencing. We evaluated relationships among host-tree genetic variation, host-tree spatial location, and the associated fungal communities. Results Genetic diversity was low across all host trees (mean: 2.6 alleles per locus) and associated fungal communities were relatively homogeneous (five sequence variants represented 78% of all reads). We found (1) genetically similar host trees harbored similar fungal communities, with no detectable effect of interhost geographic distance. (2) Host-tree heterozygosity had no detectable effect, while host-tree absolute spatial location affected community alpha diversity. Conclusions This research supports the genetic similarity rule within a range limit population and helps broaden the current scope of community genetics theory by demonstrating that community-genetic effects can occur even at expanding distributional limits where host-plant genetic variation may be limited. Our findings also provide the first documentation of community-genetic effects in a natural mangrove system