Article

Surveying the microbiome of ants: Comparing 454 pyrosequencing with traditional methods to uncover bacterial diversity.

Field Museum of Natural History, Department of Zoology, 1400 S. Lake Shore Dr., Chicago, IL 60605, USA.
Applied and Environmental Microbiology (Impact Factor: 3.95). 11/2012; DOI: 10.1128/AEM.03107-12
Source: PubMed

ABSTRACT We are only beginning to understand the depth and breadth of microbial associations across the eukaryotic tree of life. Reliably assessing bacterial diversity is a key challenge and next-generation sequencing approaches are facilitating this endeavor. Here, we used 16S rRNA amplicon pyrosequencing to survey microbial diversity in ants. We compared 454 libraries with those from Sanger sequenced clone libraries as well as cultivation of live bacteria. Pyrosequencing yielded 95,656 bacterial 16S rRNA reads from 19 samples derived from four colonies of one ant species. The most dominant bacterial orders in the microbiome of the turtle ant Cephalotes varians were Rhizobiales, Burkholderiales, Opitutales, Xanthomonadales, and Campylobacterales as revealed through both 454 sequencing and cloning. Even after stringent quality filtering, pyrosequencing recovered 445 microbe OTUs not detected with traditional techniques. In comparing bacterial communities associated with specific tissues, we found that gut tissues had significantly higher diversity than non-gut tissues and many of the OTUs identified from these groups clustered within ant specific lineages indicating a deep coevolutionary history of Cephalotes ants and their associated microbes. These lineages likely function as nutritional symbionts. One of four ant colonies investigated was infected with a Spiroplasma sp. (order Entomoplasmatales), a potential ant pathogen. Our work shows that the microbiome associated with Cephalotes varians is dominated by a few dozen bacterial lineages, and that 454 sequencing is a cost efficient tool to screen ant symbiont diversity.

0 Followers
 · 
110 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The recent development of methods applying next-generation sequencing to microbial community characterization has led to the proliferation of these studies in a wide variety of sample types. Yet, variation in the physical properties of environmental samples demands that optimal DNA extraction techniques be explored for each new environment. The microbiota associated with many species of insects offer an extraction challenge as they are frequently surrounded by an armored exoskeleton, inhibiting disruption of the tissues within. In this study, we examine the efficacy of several commonly used protocols for extracting bacterial DNA from ants. While bacterial community composition recovered using Illumina 16S rRNA amplicon sequencing was not detectably biased by any method, the quantity of bacterial DNA varied drastically, reducing the number of samples that could be amplified and sequenced. These results indicate that the concentration necessary for dependable sequencing is around 10,000 copies of target DNA per microliter. Exoskeletal pulverization and tissue digestion increased the reliability of extractions, suggesting that these steps should be included in any study of insect-associated microorganisms that relies on obtaining microbial DNA from intact body segments. Although laboratory and analysis techniques should be standardized across diverse sample types as much as possible, minimal modifications such as these will increase the number of environments in which bacterial communities can be successfully studied.
    09/2014; DOI:10.1002/mbo3.216
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Insect guts are often colonized by multi-species microbial communities that play integral roles in nutrition, digestion, and defense. Community composition can differ across host species with increasing dietary and genetic divergence, yet gut microbiota can also vary between conspecific hosts and across an individual's lifespan. Through exploration of such intraspecific variation and its correlates, molecular profiling of microbial communities can generate and test hypotheses on the causes and consequences of symbioses. In this study, we used 454 pyrosequencing and TRFLP to achieve these goals in an herbivorous ant, Cephalotes varians, exploring variation in bacterial communities across colonies, populations, and workers reared on different diets. C. varians bacterial communities were dominated by 16 core species present in over two-thirds of the sampled colonies. Core species comprised multiple genotypes, or strains, and hailed from ant-specific clades containing relatives from other Cephalotes species. Yet three were detected in environmental samples, suggesting the potential for environmental acquisition. In spite of their prevalence and long-standing relationships with Cephalotes ants, the relative abundance and genotypic composition of core species varied across colonies. Diet-induced plasticity is a likely cause, but only pollen-based diets had consistent effects, altering the abundance of two types of bacteria. Additional factors such as host age, genetics, chance, or natural selection must therefore shape natural variation. Future studies on these possibilities and on bacterial contributions to the use of pollen, a widespread food source across Cephalotes, will be important steps in developing C. varians as a model for studying widespread social insect-bacteria symbioses. This article is protected by copyright. All rights reserved.
    Molecular Ecology 11/2013; DOI:10.1111/mec.12607
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Enterococcus faecalis is a Gram-positive commensal member of the gut microbiota of a wide range of organisms. With the advent of antibiotic therapy, it has emerged as a multidrug resistant, hospital-acquired pathogen. Highly virulent strains of E. faecalis express a pore-forming exotoxin, called cytolysin, which lyses both bacterial and eukaryotic cells in response to quorum signals. Originally described in the 1930s, the cytolysin is a member of a large class of lanthionine-containing bacteriocins produced by Gram-positive bacteria. While the cytolysin shares some core features with other lantibiotics, it possesses unique characteristics as well. The current understanding of cytolysin biosynthesis, structure/function relationships, and contribution to the biology of E. faecalis are reviewed, and opportunities for using emerging technologies to advance this understanding are discussed.
    Toxins 05/2013; 5(5):895-911. DOI:10.3390/toxins5050895