Animals maintain complex associations with a diverse microbiota living in their guts. Our understanding of the ecology of these associations is extremely limited in reptiles. Here, we report an in-depth study into the microbial ecology of gut communities in three syntopic and viviparous lizard species (two omnivores: Liolaemus parvus and Liolaemus ruibali and an herbivore: Phymaturus williamsi). Using 16S rRNA gene sequencing to inventory various bacterial communities, we elucidate four major findings: (i) closely related lizard species harbour distinct gut bacterial microbiota that remain distinguishable in captivity; a considerable portion of gut bacterial diversity (39.1%) in nature overlap with that found on plant material, (ii) captivity changes bacterial community composition, although host-specific communities are retained, (iii) faecal samples are largely representative of the hindgut bacterial community and thus represent acceptable sources for nondestructive sampling, and (iv) lizards born in captivity and separated from their mothers within 24 h shared 34.3% of their gut bacterial diversity with their mothers, suggestive of maternal or environmental transmission. Each of these findings represents the first time such a topic has been investigated in lizard hosts. Taken together, our findings provide a foundation for comparative analyses of the faecal and gastrointestinal microbiota of reptile hosts.

While herbivory is a common feeding strategy in a number of vertebrate classes, less than 4% of squamate reptiles feed primarily on plant material. It has been hypothesized that physiological or microbial limitations may constrain the evolution of herbivory in lizards. Herbivorous lizards exhibit adaptations in digestive morphology and function that allow them to better assimilate plant material. However, it is unknown whether these traits are fixed or perhaps phenotypically flexible as a result of diet. Here, we maintained a naturally omnivorous lizard, Liolaemus ruibali, on a mixed diet of 50% insects and 50% plant material, or a plant-rich diet of 90% plant material. We compared parameters of digestive performance, gut morphology and function, and gut microbial community structure between the two groups. We found that lizards fed the plant-rich diet maintained nitrogen balance and exhibited low minimum nitrogen requirements. Additionally, lizards fed the plantrich diet exhibited significantly longer small intestines and larger hindguts, demonstrating that gut morphology is phenotypically flexible. Lizards fed the plant-rich diet harbored small intestinal communities that were more diverse and enriched in Melainabacteria and Oscillospira compared with mixed diet-fed lizards. Additionally, the relative abundance of sulfate-reducing bacteria in the small intestine significantly correlated with whole-animal fiber digestibility. Thus, we suggest that physiological and microbial limitations do not sensu stricto constrain the evolution of herbivory in lizards. Rather, ecological context and fitness consequences may be more important in driving the evolution of this feeding strategy.

Vertebrate diets and digestive physiologies vary tremendously. Although the contribution of ecological and behavioral features to such diversity is well documented, the roles and identities of individual intestinal enzymes shaping digestive traits remain largely unexplored. Here, we show that the sucrase-isomaltase (SI)/maltase-glucoamylase (MGAM) dual enzyme system long assumed to be the conserved disaccharide and starch digestion framework in all vertebrates is absent in many lineages. Our analyses indicate that independent duplications of an ancestral SI gave rise to the mammalian-specific MGAM, as well as to other duplicates in fish and birds. Strikingly, the duplicated avian enzyme exhibits similar activities to MGAM, revealing an unexpected case of functional convergence. Our results highlight digestive enzyme variation as a key uncharacterized component of dietary diversity in vertebrates.

A simple method for the identification of brush-border membrane α-glucosidases is described. The proteins were first solubilized and separated in a gel under native, non-denaturing, conditions. The gel was then incubated in substrate solutions (maltose or sucrose), and the product (glucose) exposed in situ by the oxidation of o-dianisidine, which yields a brown-orange color. Nano-liquid chromatography coupled to mass spectrometry analyses of proteins (nano LC-MS/MS) present in the colored bands excised from the gels, was used to confirm the presence of the enzymes. The stain is inexpensive and the procedure permits testing several substrates in the same gel. Once enzymes are identified, their abundance, relative to that of other proteins in the brush border, can be semi-quantified using nano LC-MS/MS.

Terbuthylazine technical and its 80% wettable powder were analyzed in a CIPAC collaborative study. The content and the identity were determined by gas-liquid chromatography on a column of 3% Carbowax K20M on Gas-Chrom Q, using di-n-pentylphthalate as the internal standard. Results obtained from 27 government, university, and industrial collaborators showed within-laboratory repeatability of 1.3% for the technical and 1.0% for the wettable powder. Reproducibility was 2.1% for both samples. The method has been adopted official first action

The gas-liquid chromatographic determination of tetradifon technical and formulations was collaboratively studied in duplicate with 12 laboratories. Six samples were dissolved in dichloroethane with n-hexacosane as the internal standard, chromatographed on a column of 3% SE-52, and detected by flame ionization. The average coefficients of variation were 1.2% for the 2 technical samples, 1.6% for the 2 wettable powders, and 1.5% for the 2 emulsifiable concentrates. The method has been adopted official first action.

We describe developmental changes in maltasic activity and its mRNA until adulthood, and in response to an increase in dietary starch. We studied house sparrows (), which undergo a natural switch from insects to a starch-containing seed diet during development, and zebra finches (), which have a relatively fixed starchy seed diet during development. In zebra finches, in which maltasic activity increased with age but not with dietary starch, α-glycosidase (AG) mRNA was not affected by either age or dietary starch level. In house sparrow nestlings, in which maltasic activity increased with age and with added starch, AG mRNA was higher when birds were fed a diet with added starch but did not increase with age. These results are consistent with the idea that the apparent programmed developmental increase in maltasic activity is not mainly under transcriptional control of AG mRNA, whereas induction of maltasic activity by increased dietary starch is.

Although dietary flexibility in digestive enzyme activity (i.e. reaction rate) is widespread in vertebrates, mechanisms are poorly understood. When laboratory rats are switched to a higher protein diet, the activities of apical intestinal peptidases increase within 15 h, in some cases by rapid increase in enzyme transcription followed by rapid translation and translocation to the intestine's apical, brush-border membrane (BBM). Focusing on aminopeptidase-N (APN), we studied intestinal digestive enzyme flexibility in birds, relying on activity and mRNA data from the same animals. Our model was nestling house sparrows (Passer domesticus), already known to modulate intestinal peptidase activity when switching between lower and higher protein diets. Twenty-four hours after a switch from an adequate, lower protein diet to a higher protein diet, APN activity was increased in both whole intestinal tissue homogenates and in isolated BBM, but not at 12 h post-diet switch. Twenty-four hours after a reverse switch back to the lower protein diet, APN activity was decreased, but not at 12 h post-diet switch. Changes in APN activity in both diet switch experiments were associated with parallel changes in APN mRNA. Although transcriptional changes seem to be an important mechanism underlying dietary modulation of intestinal peptidase in both nestling house sparrows and laboratory rodents, the time course for modulation in nestlings seemed slower (taking approximately twice as long) compared with laboratory rodents. It may be ecologically advantageous if nestlings biochemically restructure their gut in response to a sustained increase in insects and protein intake rather than one or a few lucky insect meals.