Investigating antimicrobial resistance in the gut bacteria of insects feeding on plants

It has been previously described that antibiotic-resistant bacteria can be found in the guts of insects feeding on a variety of plants and not exposed to significant levels of antibiotics. Such naturally-occurring resistance has implications for clinically-relevant antibiotic resistance, which is a worldwide problem, and for using plants as a source of potential novel antibiotics. We investigated this phenomenon further. Firstly, we searched for antibiotic resistance in different insects’ guts and explored its origin, using two lepidopteran hosts feeding on artificial food containing either ciprofloxacin or oxytetracycline. We discovered that these antibiotics have a diverse impact on the insect gut microbiome, beyond simply inducing antibiotic resistance. Secondly, we used the insect gut bacteria to identify plant extracts with antibacterial activity, and purified their active fractions. We found that vindoline, from leaf extract, and serpentine, from root extract, are the most abundant metabolites in active fractions of Madagascar periwinkle extracts. Finally, we developed one of the insect species we used, Galleria mellonella, into a laboratory model for antibiotic efficacy testing, toxicity testing and as a model for human baby gut. In summary, in this project we explored different aspects of the antibiotic resistance that can be found in the insect gut and used it to guide us towards plant metabolites with antibacterial properties

Amylin and beta amyloid proteins interact to form amorphous heterocomplexes with enhanced toxicity in neuronal cells

Human pancreatic islet amyloid polypeptide (hIAPP) and beta amyloid (Aβ) can accumulate in Type 2 diabetes (T2D) and Alzheimer’s disease (AD) brains and evidence suggests that interaction between the two amyloidogenic proteins can lead to the formation of heterocomplex aggregates. However, the structure and consequences of the formation of these complexes remains to be determined. The main objective of this study was to characterise the different types and morphology of Aβ-hIAPP heterocomplexes and determine if formation of such complexes exacerbate neurotoxicity. We demonstrate that hIAPP promotes Aβ oligomerization and formation of small oligomer and large aggregate heterocomplexes. Co-oligomerized Aβ42-hIAPP mixtures displayed distinct amorphous structures and a 3-fold increase in neuronal cell death as compared to Aβ and hIAPP alone. However, in contrast to hIAPP, non-amyloidogenic rat amylin (rIAPP) reduced oligomer Aβ-mediated neuronal cell death. rIAPP exhibited reductions in Aβ induced neuronal cell death that was independent of its ability to interact with Aβ and form heterocomplexes; suggesting mediation by other pathways. Our findings reveal distinct effects of IAPP peptides in modulating Aβ aggregation and toxicity and provide new insight into the potential pathogenic effects of Aβ-IAPP hetero-oligomerization and development of IAPP based therapies for AD and T2D

Level of oxytocin prior to rugby and handball matches: An exploratory study among groups of Polish players

The aim of the present exploratory study was to assess the changes in urinary oxytocin (OT) concentration during the period between five days before, and on the day of match, among rugby and handball players. Nine male rugby players with a mean age of 27.62 years (SD = 4.21) and 18 male handball players with a mean age of 17.03 years (SD = 0.57) participated. Urinary oxytocin level was measured by ELISA immunoassay as a ratio to the concentration of creatinine [mg/ml] measured through colorimetric detection. The relative level of OT to creatinine (OT/CRE) significantly differed between the type of player (rugby or handball) but not between times of measurements. Significant differences were only between OT/CRE level in a day of match in rugby players and in 5 days before match in handball players (p<0.05). There was no change in oxytocin levels during the time periods between five days before and on the day of a match, in either of the two kinds of players. The change in oxytocin might be traceable during the match but not before a match and this perhaps depends on a more subtle context of competition, but not on the assumption of competition. Further studies are needed based on more homogenous group with higher number of matches

Galleria mellonella (greater wax moth) larvae as a model for antibiotic susceptibility testing and acute toxicity trials

Abstract Background Infectivity trials and toxicity testing in rodents are important prerequisites to the use of compounds in man. However, trials in rats and mice are expensive and there are ethical considerations. Galleria mellonella (greater wax moth) larvae are a potential alternative. We have assessed the use of these insects in infectivity trials and toxicity testing. Findings Using four bacterial species (two Gram-negative and two Gram-positive) we have assessed the efficacy of four antibiotics against infections in Galleria and compared the antibiotic susceptibility with that in humans. In general, we find a good correlation. Similarly, we have assessed 11 compounds (initially tested blind) for their toxicity in Galleria and compared this with toxicity trials in mice and rats. Again we found a good correlation between toxicity in Galleria and that in rodents. Conclusion We have found, in our hands, that G. mellonella larvae can be used in infectivity trials and toxicity testing, and that these assays represent an inexpensive and readily executable alternative to testing in rodents

Oxytetracycline reduces the diversity of tetracycline-resistance genes in the Galleria mellonella gut microbiome

Abstract Background Clinically-relevant multidrug resistance is sometimes present in bacteria not exposed to human-made antibiotics, in environments without extreme selective pressures, such as the insect gut. The use of antibiotics on naïve microbiomes often leads to decreased microbe diversity and increased antibiotic resistance. Results Here we investigate the impact of antibiotics on the insect gut microbiome by identifying tetracycline-resistance genes in the gut bacteria of greater wax moth (Galleria mellonella) larvae, feeding on artificial food containing oxytetracycline. We determined that G. mellonella can be raised on artificial food for over five generations and that the insects tolerate low doses of antibiotics in their diets, but doses of oxytetracycline higher than sub-inhibitory lead to early larval mortality. In our experiments, greater wax moth larvae had a sparse microbiome, which is consistent with previous findings. Additionally, we determined that the microbiome of G. mellonella larvae not exposed to antibiotics carries a number of tetracycline-resistance genes and some of that diversity is lost upon exposure to strong selective pressure. Conclusions We show that G. mellonella larvae can be raised on artificial food, including antibiotics, for several generations and that the microbiome can be sampled. We show that, in the absence of antibiotics, the insect gut microbiome can maintain a diverse pool of tetracycline-resistance genes. Selective pressure, from exposure to the antibiotic oxytetracycline, leads to microbiome changes and alteration in the tetracycline-resistance gene pool

Antibiotic-resistant bacteria in the guts of insects feeding on plants: prospects for discovering plant-derived antibiotics

Abstract Background Although plants produce many secondary metabolites, currently none of these are commercial antibiotics. Insects feeding on specific plants can harbour bacterial strains resistant to known antibiotics suggesting that compounds in the plant have stimulated resistance development. We sought to determine whether the occurrence of antibiotic-resistant bacteria in insect guts was a widespread phenomenon, and whether this could be used as a part of a strategy to identify antibacterial compounds from plants. Results Six insect/plant pairs were selected and the insect gut bacteria were identified and assessed for antibiotic susceptibilities compared with type strains from culture collections. We found that the gut strains could be more or less susceptible to antibiotics than the type strains, or show no differences. Evidence of antibacterial activity was found in the plant extracts from five of the six plants, and, in one case Catharanthus roseus (Madagascar Periwinkle), compounds with antibacterial activity were identified. Conclusion Bacterial strains isolated from insect guts show a range of susceptibilities to antibiotics suggesting a complex interplay between species in the insect gut microbiome. Extracts from selected plants can show antibacterial activity but it is not easy to isolate and identify the active components. We found that vindoline, present in Madagascar Periwinkle extracts, possessed moderate antibacterial activity. We suggest that plant-derived antibiotics are a realistic possibility given the advances in genomic and metabolomic methodologies

Additional file 1: Figure S1. of Antibiotic-resistant bacteria in the guts of insects feeding on plants: prospects for discovering plant-derived antibiotics

Rarefaction curves of metagenomic samples from two D. gigantea guts. (DOCX 240 kb

Megacystis-Microcolon-Intestinal Hypoperistalsis Syndrome (MMIHS): Series of 4 Cases Caused by Mutation of ACTG2 (Actin Gamma 2, Smooth Muscle) Gene

MMIHS, also known as Berdon’s syndrome, is a rare disease that belongs to primary causes of CIPOS (chronic intestinal pseudoobstruction syndrome). Clinical characteristics of MMIHS are differential, but we come across the following classic symptoms: disorders of intestinal peristalsis, microcolon, and megacystis. In this article, we present a series of 4 patients with Berdon’s syndrome, in whom we managed to identify the genetic causes of MMIHS. All infants showed clinical features of bowel obstruction and dysfunction of the urinary system after birth. Two of them also manifested disorders from other systems. The prognosis for these patients is poor, but a constant betterment of management in MMIHS, in which the leading role plays TPN (total parental nutrition), causes improvement of patients’ survival

Additional file 2: Figure S2. of Antibiotic-resistant bacteria in the guts of insects feeding on plants: prospects for discovering plant-derived antibiotics

The composition of D. gigantea gut community. (DOCX 179 kb