Matan Shelomi

Where Are We Now? Bergmann’s Rule Sensu Lato in Insects

Bergmann’s rule states that individuals of a species/clade at higher altitudes or latitudes will be larger than those at lower ones. A systemic review of the known literature on inter- and intraspecific variation in insect size along latitudinal or altitudinal clines was done to see how often such clines appeared and whether they reflected classwide, species-specific, or experimentally biased tendencies. Nearly even numbers of studies showed Bergmann clines and converse-Bergmann clines, where insects get smaller as latitude/altitude increases. In fact, the majority of studies suggested no clines at all. Small ranges may have obscured certain clines, while giant ranges may have introduced artifacts. Researchers examining interspecific patterns found clines less frequently than those examining intraspecific patterns because of variation among species within the clades, which renders interspecific studies unhelpful. Bergmann’s rule does not apply to hexapods with nearly the same consistency as it does to endothermic vertebrates. The validity of Bergmann’s rule for any group and range of insects is highly idiosyncratic and partially depends on the study design. I conclude that studies of Bergmann’s rule should focus within species and look at widespread but contiguous populations to account for all sources of variation while minimizing error.

Analysis of the gut microbiota of walking sticks (Phasmatodea)

BackgroundLittle is known about the Phasmatodea gut microbial community, including whether phasmids have symbiotic bacteria aiding in their digestion. While symbionts are near ubiquitous in herbivorous insects, the Phasmatodea’s distinctively thin body shape precludes the gut enlargements needed for microbial fermentation. High-throughput sequencing was used to characterize the entire microbiota of the fat bodies, salivary glands, and anterior and posterior midguts of two species of walking stick.ResultsMost bacterial sequences belonged to a strain of Spiroplasma (Tenericutes) found primarily in the posterior midgut of the parthenogenetic species Ramulus artemis (Phasmatidae). Beyond this, no significant differences were found between the R. artemis midgut sections or between that species and Peruphasma schultei (Pseudophasmatidae). Histological analysis further indicated a lack of bacteriocytes.ConclusionsPhasmids are unlikely to depend on bacteria for digestion, suggesting they produce enzymes endogenously that most other herbivorous insects obtain from symbionts. This conclusion matches predictions based on phasmid anatomy. The role of Spiroplasma in insects warrants further study.

Endogenous cellulase enzymes in the stick insect (Phasmatodea) gut.

High cellulase (endo-beta-1,4-glucanase) activity was detected in the anterior midgut of the walking stick (Phasmatodea) Eurycantha calcarata. The enzyme was isolated and analyzed via mass spectrometry. RT-PCR revealed two endoglucanase genes, EcEG1 and EcEG2. Mascot analysis of the purified enzyme confirms it to be the product of gene EcEG1. Homologous cDNAs were also isolated from a distantly related species, Entoria okinawaensis, suggesting a general distribution of cellulase genes in phasmids. Phasmid cellulases showed high homology to endogenously-produced glycoside hydrolase family 9 (GH9) endoglucanases from insects, especially to those of termites, cockroaches, and crickets. The purified E. calcarata enzyme showed clear antigency against an anti-serum for termite GH9 cellulase, which, together with the sequence homology, further suggests an endogenous origin of the enzyme. This discovery suggests a possible nutritive value for cellulose in the leaf-feeding phasmids, unlike in herbivorous Lepidoptera.

Differential expression of endogenous plant cell wall degrading enzyme genes in the stick insect (Phasmatodea) midgut

BackgroundStick and leaf insects (Phasmatodea) are an exclusively leaf-feeding order of insects with no record of omnivory, unlike other “herbivorous” Polyneoptera. They represent an ideal system for investigating the adaptations necessary for obligate folivory, including plant cell wall degrading enzymes (PCWDEs). However, their physiology and internal anatomy is poorly understood, with limited genomic resources available.ResultsWe de novo assembled transcriptomes for the anterior and posterior midguts of six diverse Phasmatodea species, with RNA-Seq on one exemplar species, Peruphasma schultei. The latter’s assembly yielded >100,000 transcripts, with over 4000 transcripts uniquely or more highly expressed in specific midgut sections. Two to three dozen PCWDE encoding gene families, including cellulases and pectinases, were differentially expressed in the anterior midgut. These genes were also found in genomic DNA from phasmid brain tissue, suggesting endogenous production. Sequence alignments revealed catalytic sites on most PCWDE transcripts. While most phasmid PCWDE genes showed homology with those of other insects, the pectinases were homologous to bacterial genes.ConclusionsWe identified a large and diverse PCWDE repertoire endogenous to the phasmids. If these expressed genes are translated into active enzymes, then phasmids can theoretically break plant cell walls into their monomer components independently of microbial symbionts. The differential gene expression between the two midgut sections provides the first molecular hints as to their function in living phasmids. Our work expands the resources available for industrial applications of animal-derived PCWDEs, and facilitates evolutionary analysis of lower Polyneopteran digestive enzymes, including the pectinases whose origin in Phasmatodea may have been a horizontal transfer event from bacteria.

Horizontal Gene Transfer of Pectinases from Bacteria Preceded the Diversification of Stick and Leaf Insects

Genes acquired by horizontal transfer are increasingly being found in animal genomes. Understanding their origin and evolution requires knowledge about the phylogenetic relationships from both source and recipient organisms. We used RNASeq data and respective assembled transcript libraries to trace the evolutionary history of polygalacturonase (pectinase) genes in stick insects (Phasmatodea). By mapping the distribution of pectinase genes on a Polyneoptera phylogeny, we identified the transfer of pectinase genes from known phasmatodean gut microbes into the genome of an early euphasmatodean ancestor that took place between 60 and 100 million years ago. This transfer preceded the rapid diversification of the suborder, enabling symbiont-free pectinase production that would increase the insects’ digestive efficiency and reduce dependence on microbes. Bacteria-to-insect gene transfer was thought to be uncommon, however the increasing availability of large-scale genomic data may change this prevailing notion.

Ancestral gene duplication enabled the evolution of multifunctional cellulases in stick insects (Phasmatodea).

The Phasmatodea (stick insects) have multiple, endogenous, highly expressed copies of glycoside hydrolase family 9 (GH9) genes. The purpose for retaining so many was unknown. We cloned and expressed the enzymes in transfected insect cell lines, and tested the individual proteins against different plant cell wall component poly- and oligosaccharides. Nearly all isolated enzymes were active against carboxymethylcellulose, however most could also degrade glucomannan, and some also either xylan or xyloglucan. The latter two enzyme groups were each monophyletic, suggesting the evolution of these novel substrate specificities in an early ancestor of the order. Such enzymes are highly unusual for Metazoa, for which no xyloglucanases had been reported. Phasmatodea gut extracts could degrade multiple plant cell wall components fully into sugar monomers, suggesting that enzymatic breakdown of plant cell walls by the entire Phasmatodea digestome may contribute to the Phasmatodea nutritional budget. The duplication and neofunctionalization of GH9s in the ancestral Phasmatodea may have enabled them to specialize as folivores and diverge from their omnivorous ancestors. The structural changes enabling these unprecedented activities in the cellulases require further study.

Review of the Gross Anatomy and Microbiology of the Phasmatodea Digestive Tract

Abstract The sparse descriptions of the stick insect (Phasmatodea) digestive system as reported/provided in the literature are highly contradictory. This paper describes the digestive systems of several families of Phasmatodea (Timematidae, Heteropterygidae, Diapheromeridae, Pseudophasmatidae, and Phasmatidae) plus the gut microbiome of these and one other (Phylliidae) to both verify past findings and provide a general description of the Phasmatodea alimentary canal. The constrictions imposed by this anatomy on phasmid gut microbiology, its connections to recently released Phasmatodea transcriptomes, and how it differs from the anatomy of related orders in the Polyneoptera are discussed. All Phasmatodea have ridged proventriculi lined or covered with small spines. Anterior projections of the midgut, sometimes described as gastric caeca, are only found in Euphasmatodea and often obscure the proventriculus. We define the cardia as the complex of foregut and midgut tissue where the type II peritrophic matrix is produced. Appendices of the midgut are an autapomorphy for Phasmatodea, but Timema have fewer and larger appendices relative to body size. We suggest caeca-like projections and the loss of large, proventricular teeth are apomorphies of Euphasmatodea. We identify a possible facultative symbiosis in Eucalyptus-feeding species that requires further study.

Vital staining of the stick insect digestive system identifies appendices of the midgut as novel system of excretion.

The stick insects or phasmids (Phamsatodea) have a series of pyriform ampulles with long, thin filaments on the posterior end of their midgut referred to as the “appendices of the midgut.” Found only in phasmids, their function had never been determined until now. Their similarity to the Malpighian tubules, which are ubiquitous insect organs of excretion, suggested a similar function. To differentiate between the appendices and the Malpighian tubules and compare functional differences between the two tissue types, vital staining (the injection of histological stains into living organisms) was done in conjunction with light and scanning electron microscopy in multiple phasmid species. The results showed that the appendices originated in the basal phasmids (Timematidae) and grew more numerous in derived species. The appendices stain selectively, notably failing to pick up the indicators of the two known systems of invertebrate excretory function, indigo carmine and ammonium carmine. Appendices sequester stains in the ampule portion before eliminating the compounds into the midgut. We conclude by confirming that the appendices do have an excretory function, but one unlike any other known in invertebrates. Their function is likely cation excretion, playing a role in calcium regulation and/or organic alkaloid sequestration. The appendices must thus be considered distinct organs from the Malpighian tubules. J. Morphol. 275:623–633, 2014.

Review of Black Soldier Fly (Hermetia illucens) as Animal Feed and Human Food

Food futurists accept that sustainability-minded humanity will increasingly incorporate insects as alternative protein. The most studied and easily reared species are not necessarily the most sustainable, acceptable, or delicious. Here, we review the literature on the black soldier fly, Hermetia illucens, which is capable of efficiently converting a wide variety of organic materials, from food waste to manure, into insect biomass. They can be grown and harvested without dedicated facilities and are not pestiferous. Their larvae are 42% crude protein and 29% fat, although they are higher in saturated fats than most insects. They do not concentrate pesticides or mycotoxins. They are already grown and recommended for use as animal feed, but with regional legal restrictions on how this is done. For commercial use in human foods, larvae could potentially be milled and converted into a textured protein with a strong flavor. Their biggest advantage over other insects is their ability to convert waste into food, generating value and closing nutrient loops as they reduce pollution and costs. This general advantage is also their greatest disadvantage, for the social stigmas and legal prohibitions against eating organisms that eat waste are added to extant taboos facing insect consumption.

Evidence of Photo Manipulation in a Delusional Parasitosis Paper

Abstract: In 2004, an article in the Journal of the New York Entomological Society claimed that individuals with delusory parasitosis actually suffer from collembola infestations. The article has been critiqued for poor methodology and results that contradict all knowledge about collembolans. Less easily accounted for has been a figure in the article purporting to show a collembolan in a skin scraping. The image appears to have been altered using photo manipulation software to an unacceptable degree, and this paper demonstrates that to be the case. The altered figure represents creation of nonexistent data, a form of scientific misconduct. Whether the deception is deliberate or a product of an otherwise well-meaning author ignorant of the limits of acceptable image manipulation is unknown, but the result is peer-reviewed support for a conclusion that complicates patient treatment. In the current era of computers, even regional entomology journals must have detailed standards for what kind of images and image manipulations are acceptable for publication.