Effects of ivermectin on development of Calliphora vicina, Robineau-Desvoidy 1830 (Diptera, Calliphoridae).

Ivermectin is one of the most widely used drugs for parasite control. Previous studies have shown a reduction in the abundance and diversity of "non-target" coprophilous organisms due to the presence of ivermectin (IVM) in bovine faecal matter (FM). Due to its breadth of behavioural habits, Calliphora vicina is a suitable dipteran species to evaluate the effects of IVM in FM. The aim of this work was to evaluate the effect of five concentrations of IVM in FM (3000, 300, 100, 30, and 3 ng/g) on the development of C. vicina. The following endpoints were evaluated: survival (between the first larval stage and emergence of new adults), larval development times to pupation and pupation times to adult, and adult emergence (% sex) and LC50. Sampling was performed from larval hatching at 60 and 120 min and at 3, 4, 5, and 12 h, and every 24 h specimens were weighed until pupae were observed. Data were analysed by ANOVA using a non-parametric Kruskal-Wallis test and as a function of elapsed development time and accumulated degree hours (ADH). Mortality at 3000 and 300 ng/g was 100% and 97%, respectively. There were statistically significant delays in adult emergence time (p = 0.0216) and in the ADH (p = 0.0431) between the control group (C) and 100 ng/g. The LC50 was determined at 5.6 ng/g. These results demonstrate the lethal and sub-lethal effects of IVM on C. vicina, while highlighting the usefulness of this species as a bioindicator for ecotoxicological studies.

Transcription Factors AhR and ARNT Regulate the Expression of CYP6SX1 and CYP3828A1 Involved in Insecticide Detoxification in Bradysia odoriphaga.

Aryl hydrocarbon receptor (AhR) and aryl hydrocarbon receptor nuclear translocator (ARNT) mediate the responses of adaptive metabolism to various xenobiotics. Here, we found that BoAhR and BoARNT are highly expressed in the midgut of Bradysia odoriphaga larvae. The expression of BoAhR and BoARNT was significantly increased after exposure to imidacloprid and phoxim. The knockdown of BoAhR and BoARNT significantly decreased the expression of CYP6SX1 and CYP3828A1 as well as P450 enzyme activity and caused a significant increase in the sensitivity of larvae to imidacloprid and phoxim. Exposure to ß-naphthoflavone (BNF) significantly increased the expression of BoAhR, BoARNT, CYP6SX1, and CYP3828A1 as well as P450 activity and decreased larval sensitivity to imidacloprid and phoxim. Furthermore, CYP6SX1 and CYP3828A1 were significantly induced by imidacloprid and phoxim, and the silencing of these two genes significantly reduced larval tolerance to imidacloprid and phoxim. Taken together, the BoAhR/BoARNT pathway plays key roles in larval tolerance to imidacloprid and phoxim by regulating the expression of CYP6SX1 and CYP3828A1.

Hemolymph metabolism of black soldier fly (Diptera: Stratiomyidae), response to different supplemental fungi.

The black soldier fly, Hermetia illucens L. (Diptera: Stratiomyidae), is commonly used for organic waste recycling and animal feed production. However, the often inadequate nutrients in organic waste necessitate nutritional enhancement of black soldier fly larvae, e.g., by fungal supplementation of its diet. We investigated the amino acid composition of two fungi, Candida tropicalis (Castell.) Berkhout (Saccharomycetales: Saccharomycetaceae) and Pichia kudriavzevii Boidin, Pignal & Besson (Saccharomycetales: Pichiaceae), from the black soldier fly gut, and commercial baker's yeast, Saccharomyces cerevisiae Meyen ex E.C. Hansen (Saccharomycetales: Saccharomycetaceae), and their effects on larval growth and hemolymph metabolites in fifth-instar black soldier fly larvae. Liquid chromatography-mass spectrometry was used to study the effect of fungal metabolites on black soldier fly larval metabolism. Amino acid analysis revealed significant variation among the fungi. Fungal supplementation led to increased larval body mass and differential metabolite accumulation. The three fungal species caused distinct metabolic changes, with each over-accumulating and down-accumulating various metabolites. We identified significant alteration of histidine metabolism, aminoacyl-tRNA biosynthesis, and glycerophospholipid metabolism in BSF larvae treated with C. tropicalis. Treatment with P. kudriavzevii affected histidine metabolism and citrate cycle metabolites, while both P. kudriavzevii and S. cerevisiae treatments impacted tyrosine metabolism. Treatment with S. cerevisiae resulted in down-accumulation of metabolites related to glycine, serine, and threonine metabolism. This study suggests that adding fungi to the larval diet significantly affects black soldier fly larval metabolomics. Further research is needed to understand how individual amino acids and their metabolites contributed by fungi affect black soldier fly larval physiology, growth, and development, to elucidate the interaction between fungal nutrients and black soldier fly physiology.

Effects of imidacloprid on the survival and biomarker responses of Eristalis tenax larvae (Diptera: Syrphidae): a comparative study between indoor and outdoor exposures.

Imidacloprid is a widely used pesticide in agriculture. It is being found in aquatic ecosystems in agricultural regions. This study aimed to evaluate its effects on the survival rates, acetylcholinesterase (AChE) and catalase (CAT) responses of larval Eristalis tenax hoverflies. The larvae were exposed for 3, 7 and 14 days to increasing concentrations of imidacloprid (0, 0.1, 0.5 and 2 mg L-1) both indoors at a constant temperature of 20 °C and outdoors under varying environmental conditions. The results revealed that indoors and outdoors, the mortality of E. tenax significantly increased with increasing imidacloprid concentration and duration of exposure. Median lethal concentrations (LC50) varied from 0.03 to 0.17 mg L-1 depending on the duration and conditions of exposure. Indoors, AChE activity decreased in all the treatments for all three exposure durations, whereas outdoors the decrease was observed after the short (3-day) and long (14-day) exposure durations. AChE inhibition ranged from 6% to 62% (indoors) and 12% to 62% (outdoors). Variations in CAT activity were observed for both experimental setups, with a decrease outdoors in larvae exposed to 0.5 mg L-1 for 7 days and a gradual dose-dependent increase indoors for exposure lasting 3 and 7 days. This study sheds light on the potential ecological implications of imidacloprid contamination which may cause the decline of aquatic insect populations and pollination rates, leading to disruptions of the food chain and the overall decline of aquatic and terrestrial ecosystem health.

Valorization of Hermetia illucens breeding rejects by chitins and chitosans production. Influence of processes and life cycle on their physicochemical characteristics.

Breeding of the black soldier fly is carried out to produce proteins. It is accompanied by releases during the life cycle of this insect. This work is a study of the valorization of these rejects through the production of chitins and chitosans with controlled characteristics. An extraction process is developed with an order of treatments and reaction conditions that provide chitins with high contents. These contents increase as the stages of the life cycle progress and drop for the adult. However, the exuviae chitins present organic impurities which will be eliminated at the N-deacetylation reaction for pupe and after a purification treatment for chitosan from larval stages. All these chitins have an α structure although certain physicochemical characteristics of the larval exuviae chitins are close to those presented by γ chitin. The observed shifts are linked to the effect of impurities rather than to a difference in structure. N-deacetylation of chitins makes possible the valorization of all rejects by the production of pure chitosans with high yields which retain a porous structure for the exuviae and fibrous for the adult which allow complementary applications. These chitosans are highly to completely deacetylated and their molar masses can vary depending on the process and life stage.

Case of Wound Myiasis in a Squamous Cell Carcinoma Lesion of the Scalp.

Myiasis refers to the infestation of living humans and vertebrate animals by dipterous larvae. Many organs can be infested by fly larvae, but cutaneous and wound myiases are the most frequently encountered clinical forms. Persistent ulcer or non-healing wound is one of the symptoms of squamous cell carcinoma which is the second most common skin cancer in the world. Here we report a case of an elderly man with a severe wound myiasis in a squamous cell carcinoma lesion of the scalp. The maggots were confirmed to be Lucilia sericata which are widespread flies in Japan. Human myiasis is rarely reported in Japan, but patients with necrotic, hemorrhaging, or pus-filled wounds are susceptible to infestation. It is necessary for doctors and nurses to ensure that their patients change their dressings daily and keep their wounds clean.

An optimized method for Nasonia germ-free rearing.

A germ-free rearing system is a crucial method for host-microbiota interactions using Nasonia as a model system. The previous rearing media in 2012 introduced toxic factors like bleach and antibiotics, required significant effort and volume of media preparation, and the rearing protocols in 2012 and 2016 often resulted in embryos, larvae, and enclosing pupae drowning, underfed, or desiccating. In this work, we optimize the germ-free rearing media that excludes the toxic factors and provide a substrate for the developing animals to have constant access to media without the risk of drowning or desiccation. The new process resulted in an increase in full maturation of larvae to adults from 33 to 65%, with no effect on the rate of growth or final adult size. This significantly improves the applicability of germ-free rearing of Nasonia and potentially other parasitoids.

Characterization and Phylogenetic Analysis of the Complete Mitochondrial Genomes of Two Tiny Necrophagous Phorid Flies, Metopina sagittata and Puliciphora borinquenensis (Diptera: Phoridae).

The mitochondrial genome is frequently used for species identification and phylogenetic studies. In this study, we first sequenced and annotated the complete mitochondrial genomes of two phorid species that are forensically important in buried or enclosed environments: Metopina sagittata (Liu) and Puliciphora borinquenensis (Wheeler). The complete mitochondrial genome sequences of M. sagittata and P. borinquenensis were 15,640 bp with an A+T content of 75.97% and 15,429 bp with an A+T content of 75.38%, respectively. Their circular genomes both contained 13 protein-coding genes (PCGs), 22 transfer RNA genes, 2 ribosomal RNA genes, and 1 control region located between rrnS and trnI which was 808 bp for M. sagittata and 746 bp for P. borinquenensis. All the PCGs of both species started with ATN codons except for cox1 which used TTG codon. In addition to the common stop codon TAA and TAG, the incomplete stop codon T was used in two PCGs (cox1 and nad4) of M. sagittata and five PCGs (cox1, cox2, cox3, nad5, and nad4) of P. borinquenensis. There were 3 and 10 mismatched base pairs in the tRNA secondary structures from M. sagittata and P. borinquenensis, respectively. Both maximum likelihood and Bayesian inference analyses indicated that Platypezidae and Phoridae are sister taxa. M. sagittata is closely related to P. borinquenensis within the subfamily Metopininae. This work enhances the databases of Phoridae genomes and contributes to the further study of species identification and phylogenetics of this family.

Staphylococcus aureus in Substrates for Black Soldier Fly Larvae (Hermetia illucens) and Its Dynamics during Rearing.

Black soldier fly larvae (BSFL; Hermetia illucens) are promising insects for the conversion of organic waste streams into valuable biomolecules. Such waste streams can contain foodborne pathogens. To assess this risk factor, this study evaluated the presence of Staphylococcus aureus in waste streams as a substrate ingredient for BSFL production as well as in the rearing process. First, the general microbiological quality and the occurrence of S. aureus were investigated for different waste streams. Staphylococcus aureus was abundantly present. Control of pH and water activity should avoid pathogens, which cannot grow in single-substrate ingredients, redeveloping when mixing streams for optimal substrate conditions for BSFL production. Next, it was investigated whether S. aureus present in the substrate was ingested and/or eradicated by BSFL. In inoculation trials, with S. aureus added to chicken feed as the substrate at 3 or 7 log CFU/g, the larvae showed a reducing effect on S. aureus. After 6 days, S. aureus counts were below the detection limit (2.0 log CFU/g) in all larvae samples and decreased in the substrate to <2.0 and <3.1 log CFU/g for inoculation levels of 3 and 7 log CFU/g, respectively. While this is promising, it is still recommended to monitor and control this pathogen in BSFL rearing. Intriguingly, screening of antimicrobial activity of dominant microorganisms associated with BSFL showed a clear activity of Trichosporon isolates against S. aureus. Future research should explore whether Trichosporon, which is frequently observed in BSFL, plays a role in controlling specific microorganisms, such as S. aureus. IMPORTANCE Given the increasing need for (more sustainable) methods to upcycle organic waste streams, the interest to rear insects, like black soldier fly larvae (BSFL), on such streams is increasing. This study reveals that S. aureus is abundantly present in such waste streams, which might be a point of attention for insect producers. At the same time, it reveals that when S. aureus was inoculated in chicken feed as the substrate, it was not detected in the larvae and was reduced in the substrate after 6 days. Future inoculation trials should investigate whether this reduction is substrate dependent or not. Toward the future, the role of the BSFL microbiota in controlling intestinal bacterial community homeostasis should be explored, because one of the dominant microorganisms associated with BSFL, Trichosporon spp., showed clear activity against S. aureus.

Assessing the effects of a novel biostimulant to enhance leafminer resistance and plant growth on common bean.

The leafminer Liriomyza trifolii is one of the major insects that affect Phaseolus vulgaris production worldwide. Novel and safe biobased stimulator compound (BSTC) with micronutrient-amino acid chelated compounds was developed from natural compounds and was used for foliar spray of P. vulgaris. Treated plants showed significantly increased in quality and productivity as well as significant reduction in leafminer infestation by close the tunnel end resulting in larvae suffocation and death. BSTC contains chemical composition that has important function in inducing immunity and resistance against insects, enhance plant growth and production. Also, HPLC showed that the assembled BSTC is rich in nucleobases than yeast extract (> 56 fold). Aminochelation zinc enhanced the rate of absorption of nutrient compounds and could participate in safe biofortification strategy. The expression of plant defense related genes under BSTC treatment revealed strong correlations between the transcription rates of defense related genes. Based on binding energies and interacting residues of six vital insect proteins, the best-docked complexes was obtained with disodium 5'-inosinate, delphinidin 3-glucoside and hyperoside. Obtained findings indicate that the foliar application of BSTC can enhance plant growth and productivity, uptake of important elements, expression of defense related genes and inhibit insect essential genes.

Comparative Lipid Profile Analysis of Hermetia illucens Larvae Fed Food Waste at Different Days of Age Using an LC-MS-Based Lipidomics Approach.

A lipidomics approach based on liquid chromatography-tandem mass spectrometry (LC-MS) was applied to analyze the molecular-level mechanism of lipid deposition in Hermetia illucens (H. illucens) larvae fed food waste (FW) at different days of age. The H. illucens larvae reared on FW substrates generally became larger, heavier, and fatter at 5-15 d of age. A large amount of glycerolipids (GL) were deposited, while glycerophospholipids (GP), sphingolipids, and derivatized lipids became relatively less abundant during the growth stage of the larvae. Forty-three subclasses of 3,205 lipid molecules were identified in larvae, and 139 lipids (79 upregulated and 60 downregulated during larval growth and development) were identified as potential biomarkers (variable importance in projection > 1; P < 0.05). The differential lipids were mainly enriched in 19 metabolic pathways, of which 9 metabolic pathways related to lipids, including GL and GP metabolisms. The results demonstrate that the lipid composition and mechanisms changed during the growth and development stage of H. illucens larvae. To the best of our knowledge, this is the first work exploring the molecular-level mechanism of lipid deposition during the growth and development stage of H. illucens larvae. The findings provide novel information for determining and utilizing the nutritional value of H. illucens larvae.

Comprehensive thermal performance curves for yellow dung fly life history traits and the temperature-size-rule.

Ambient temperature strongly determines the behaviour, physiology, and life history of all organisms. The technical assessment of organismal thermal niches in form of now so-called thermal performance curves (TPC) thus has a long tradition in biological research. Nevertheless, several traits do not display the idealized, intuitive dome-shaped TPC, and in practice assessments often do not cover the entire realistic or natural temperature range of an organism. We here illustrate this by presenting comprehensive sex-specific TPCs for the major (juvenile) life history traits of yellow dung flies (Scathophaga stercoraria; Diptera: Scathophagidae). This concerns estimation of prominent biogeographic rules, such as the temperature-size-rule (TSR), the common phenomenon in ectothermic organisms that body size decreases as temperature increases. S. stercoraria shows an untypical asymptotic TPC of continuous body size increase with decreasing temperature without a peak (optimum), thus following the TSR throughout their entire thermal range (unlike several other insects presented here). Egg-to-adult mortality (our best fitness estimator) also shows no intermediate maximum. Both may relate to this fly entering pupal winter diapause below 12 °C. While development time presents a negative exponential relationship with temperature, development rate and growth rate typify the classic TPC form for this fly. The hitherto largely unexplored close relative S. suilla with an even more arctic distribution showed very similar responses, demonstrating large overlap among two ecologically similar, coexisting dung fly species, thus implying limited utility of even complete TPCs for predicting species distribution and coexistence.

Ovitrap Preference in the Black Soldier Fly, Hermetia illucens (L.) (Diptera: Stratiomyidae).

<b>Background and Objective:</b> The adult female of the Black Soldier Fly (BSF) usually performs oviposition near decaying organic matter in cracks found in dry areas. In mass-rearing facilities or on a laboratory scale, females usually lay eggs on the provided ovitrap which is made of various types of materials. This study was aimed to observe the female preferential behaviour for ovitrap types during the oviposition period. <b>Materials and Methods:</b> The study was conducted in a semi-outdoor screen house with direct sunlight. The ovitrap materials used were: Dry leaves, wood, cardboard and plastic (infraboard). The parameters calculated in this study were ovitrap preference, oviposition duration, the number and weight of the eggs, fertility of eggs, development time and mortality of offspring. <b>Results:</b> The BSF females preferred the wood ovitrap to other ovitrap types. This was indicated by the high number of females visits, the number of egg-laying females and the total number of eggs on the wood ovitrap. Eggs found in the plastic ovitrap had the highest mortality compared to eggs found in the other ovitraps. <b>Conclusion:</b> The difference of ovitrap material affected female visitation preferences, the oviposition frequency and the duration of egg-laying of female BSF but did not affect offspring development time in any of the stages (egg-adults).

Mechanism of Action of Endophytic Fungi Hypocrea lixii and Beauveria bassiana in Phaseolus vulgaris as Biopesticides against Pea Leafminer and Fall Armyworm.

Endophytic fungal isolates Hypocrea lixii F3ST1 and Beauveria bassiana G1LU3 were evaluated for their potential to endophytically colonize and induce active compounds in Phaseolus vulgaris, as a defense mechanism against pea leafminer (Liriomyza huidobrensis) and fall armyworm (Spodoptera frugiperda). Endophytic colonization was achieved through seed inoculation with the volatile emissions from P. vulgaris plants being analyzed using GC-MS. The crude extracts of P. vulgaris obtained using methanol and dichloromethane were assayed against leafminer and fall armyworm larvae using leaf dipping and topical application, respectively. The two isolates successfully colonized the entire host plant (roots, stems, and leaves) with significant variation (p < 0.001) between fungal isolates and the controls. The results showed qualitative differences in the volatile profiles between the control plants, endophytically colonized and insect-damaged plants attributed to fungal inoculation and leafminer damage. The crude methanol extracts significantly reduced the percentage pupation of 2nd instar leafminer larvae (p < 0.001) and adult-flies emergence (p < 0.05). The survival of the 1st instar fall armyworm larvae was also significantly reduced (p < 0.001) compared to the controls. This study demonstrated the high potential of endophytic fungi H. lixii and B. bassiana in inducing mainly specific defense compounds in the common bean P. vulgaris that can be used against pea leafminer and fall armyworm.

Fluctuating temperatures extend longevity in pupae and adult stages of the sepsid Themira biloba.

Fluctuating Thermal Regimes (FTR), where organisms are held at low temperatures with a brief, daily warm pulse, have been shown to increase longevity in adult insects and improve pupa survival while reducing sublethal effects. We used FTR to extend the longevity and thus generation time of the fly species Themira biloba (Diptera: Sepsidae). T. biloba can be maintained in continuous culture and requires an insecticide-free dung substrate for larval growth and development. Our objective was to decrease labor and consumable materials required to maintain insect species in critical scientific collections using FTR. We extended pupation time from 4 days up to 8 weeks with no increase in mortality, and mean adult longevity was increased from 12 days to 50 days. FTR is a valuable tool for reducing the investment required to maintain rare and exotic insects.

Non-lethal effects of entomopathogenic nematode infection.

Entomopathogenic nematodes are typically considered lethal parasites of insect hosts. Indeed they are employed as such for biological control of insect pests. The effects of exposure to entomopathogenic nematodes are not strictly limited to mortality, however. Here we explore non-lethal effects of exposure to entomopathogenic nematodes by introducing the relatively non-susceptible pupal stage of Delia antiqua to thirteen different strains. We specifically chose to inoculate the pupal stage because it tends to be more resistant to infection, yet resides in the soil where it could come into contact with EPN biological control agents. We find that there is no significant mortality at the pupal stage, but that there are a host of strain-dependent non-lethal effects during and after the transition to adulthood including altered developmental times and changes in risk of death compared to controls. We also find that exposure to specific strains can reduce risk of mortality. These results emphasize the strain-dependent nature of entomopathogenic nematode infection and highlight the positive and negative ramifications for non-lethal effects for biological control of insect pests. Our work emphasizes the need for strain-specific screening of biological control agents before wide-spread adoption.

Local intestinal microbiota response and systemic effects of feeding black soldier fly larvae to replace soybean meal in growing pigs.

Black soldier fly (Hermetia illucens; BSF) larvae as dietary protein source have the ability to deliver nutrients and could possess functional properties that positively support animal productivity and health. More knowledge, however, is needed to assess the impact of feeding a BSF based diet on gut and animal health. Sixteen post-weaned male pigs were randomly assigned to two groups and fed for three weeks with iso-caloric and iso-proteinaceous experimental diets prepared with either soybean meal (SBM) as reference protein source or with BSF as single source of dietary protein. At the end of the trial, the pigs were sacrificed to collect relevant digesta, gut tissue and blood samples to study changes induced by the dietary treatments using ~ omics based analyses. Inclusion of BSF in the diet supports the development of the intestinal microbiome that could positively influence intestinal health. By amine metabolite analysis, we identified two metabolites i.e. sarcosine and methionine sulfoxide, in plasma that serve as markers for the ingestion of insect based ingredients. BSF seems to possess functional properties indicated by the appearance of alpha-aminobutyric acid and taurine in blood plasma of pigs that are known to induce health beneficial effects.

Evidence for competition and cannibalism in wormlions.

Trap-building predators, such as web-building spiders and pit-building antlions, construct traps to capture their prey. These predators compete over sites that either enable the construction of suitable traps, are prey rich, or simply satisfy their abiotic requirements. We examined the effect of intraspecific competition over suitable space in pit-building wormlions. As expected, the ability of wormlions to select their favorable microhabitats-shaded or deep sand over lit or shallow sand-decreased with increasing density. Favorable microhabitats were populated more frequently by large than by small individuals and the density of individuals in the favorable microhabitat decreased with their increase in body mass. The advantage of large individuals in populating favorable microhabitats is nevertheless not absolute: both size categories constructed smaller pits when competing over a limited space compared to those constructed in isolation. The outcome of competition also depends on the type of habitat: deep sand is more important for large wormlions than small ones, while shade is similarly important for both size classes. Finally, in contrast to previous reports, cannibalism is shown here to be possible in wormlions. Its prevalence however is much lower compared to that documented in other trap-building predators. Our findings show that the advantage of large individuals over small ones should not be taken for granted, as it can depend on the environmental context. We present suggestions for the relative lack of competitive advantage of large wormlion individuals compared to other trap-building predators, which may stem from the absence of obvious weaponry, such as sharp mandibles.

Evaluation of Development Datasets for Hermetia illucens (L.) (Diptera: Stratiomyidae) for Estimating the Time of Placement of Human and Swine Remains in Texas, USA.

A basic tenet of forensic entomology is development data of an insect can be used to predict the time of colonization (TOC) by insect specimens collected from remains, and this prediction is related to the time of death and/or time of placement (TOP). However, few datasets have been evaluated to determine their accuracy or precision. The black soldier fly, Hermetia illucens (L.) (Diptera: Stratiomyidae) is recognized as an insect of forensic importance. This study examined the accuracy and precision of several development datasets for the black soldier fly by estimating the TOP of five sets of human and three sets of swine remains in San Marcos and College Station, TX, respectively. Data generated from this study indicate only one of these datasets consistently (time-to-prepupae 52%; time-to-eclosion 75%) produced TOP estimations that occurred within a day of the actual TOP of the remains. It is unknown if the precolonization interval (PreCI) of this species is long, but it has been observed that the species can colonize within 6 d after death. This assumption remains untested by validation studies. Accounting for this PreCI improved accuracy for the time-to-prepupae group, but reduced accuracy in the time-to-eclosion group. The findings presented here highlight a need for detailed, forensic-based development data for the black soldier fly that can reliably and accurately be used in casework. Finally, this study outlines the need for a basic understanding of the timing of resource utilization (i.e., duration of the PreCI) for forensically relevant taxa so that reasonable corrections may be made to TOC as related to minimum postmortem interval (mPMI) estimates.

Effects of Gasterophilus pecorum infestation on the intestinal microbiota of the rewilded Przewalski's horses in China.

Horse botflies have been a threat to the Przewalski's horses in the Kalamaili Nature Reserve in Xinjiang of China since their reintroduction to the original range. As larvae of these parasites could infest the intestine of a horse for months, they could interact with and alter the structure and composition of its intestinal microbiota, affecting adversely its health. Nonetheless, there are no such studies on the rewilded Przewalski's horses yet. For the first time, this study characterizes the composition of the intestinal microbiota of 7 rewilded Przewalski's horses infected severely by Gasterophilus pecorum following and prior to their anthelmintic treatment. Bioinformatics analyses of the sequence data obtained by amplicon high throughput sequencing of bacterial 16S rRNA genes showed that G. pecorum infestation significantly increased the richness of the intestinal microbial community but not its diversity. Firmicutes and Bacteroidetes were found the dominant phyla as in other animals, and the parasitic infestation decreased the F/B ratio largely by over 50%. Large reduction in relative abundances of the two genera Streptococcus and Lactobacillus observed with G. pecorum infestation suggested possible changes in colic and digestion related conditions of the infected horses. Variations on the relative abundance of the genus groups known to be pathogenic or symbiotic showed that adverse impact of the G. pecorum infestation could be associated with reduction of the symbiotic genera Lactobacillus and Bifidobacterium that are probiotics and able to promote immunity against parasitic infection.