House flies are well recognized as filth-associated organisms and public nuisances. House flies create sanitation issues when they bridge the gap between microbe-rich breeding environments and animal/human habitations. Numerous scientific surveys have demonstrated that house flies harbor bacterial pathogens that pose a threat to humans and animals. More extensive and informative surveys incorporating next-generation sequencing technologies have shown that house fly carriage of pathogens and harmful genetic elements, such as antimicrobial resistance genes, is more widespread and dangerous than previously thought. Further, there is a strong body of research confirming that flies not only harbor but also transmit viable, and presumably infectious, bacterial pathogens. Some pathogens replicate and persist in the fly, permitting prolonged shedding and dissemination. Finally, although the drivers still have yet to be firmly determined, the potential range of dissemination of flies and their associated pathogens can be extensive. Despite this evidence, the house flies’ role as reservoirs, disseminators, and true, yet facultative, vectors for pathogens have been greatly underestimated and underappreciated. In this review, we present key studies that bolster the house fly’s role both an important player in microbial ecology and population biology and as transmitters of microbial threats to animal and human health.

House flies are important nuisance pests in a variety of confined livestock operations. More importantly, house flies are known mechanical vectors of numerous animal and human pathogens. Bovine respiratory disease (BRD) is an economically important, complex illness of cattle associated with several bacteria and viruses. The role of flies in the ecology and transmission of bacterial pathogens associated with BRD is not understood. Using culture-dependent and culture-independent methods, we examined the prevalence of the BRD bacterial complex Mannheimia haemolytica, Pasteurella multocida and Histophilus somni in house flies collected in a commercial feedlot from a pen with cattle exhibiting apparent BRD symptoms. Using both methods, M. haemolytica was detected in 11.7% of house flies, followed by P. multocida (5.0%) and H. somni (3.3%). The presence of BRD bacterial pathogens in house flies suggests that this insect plays a role in the ecology of BRD pathogens and could pose a risk as a potential reservoir and/or a vector of BRD pathogens among individual cattle and in their environment.

Background: Ticks are obligate hematophagous arthropods transmitting a wide range of pathogens to humans and animals. They also harbor a non-pathogenic microbiota, primarily in the ovaries and the midgut. In the previous study on Ixodes ricinus, we used a culture-independent approach and showed a diverse but quantitatively poor midgut bacterial microbiome. Our analysis also revealed the absence of a core microbiome, suggesting an environmental origin of the tick midgut microbiota. Methods: A bacterial analysis of the midgut of adult females collected by flagging from two localities in the Czech Republic was performed. Using the culture-independent approach, we tested the hypothesis that the midgut microbiome is of the environmental origin. We also cultured indigenous bacteria from the tick midgut and used these to feed ticks artificially in an attempt to manipulate the midgut microbiome. Results: The midgut showed a very low prevalence and abundance of culturable bacteria, with only 37% of ticks positive for bacteria. The culture-independent approach revealed the presence of Borrelia sp., Spiroplasma sp., Rickettsia sp., Midichloria sp. and various mainly environmental Gram-positive bacterial taxa. The comparison of ticks from two regions revealed that the habitat influenced the midgut bacterial diversity. In addition, the midgut of ticks capillary fed with the indigenous Micrococcus luteus (Gram-positive) and Pantoea sp. (Gram-negative) could not be colonized due to rapid and effective clearance of both bacterial taxa. Conclusions: The midgut microbiome of I. ricinus is diverse but low in abundance, with the exception of tick-borne pathogens and symbionts. The environment impacts the diversity of the tick midgut microbiome. Ingested extracellular environmental bacteria are rapidly eliminated and are not able to colonize the gut. We hypothesize that bacterial elimination triggered in the midgut of unfed adult females is critical to maintain low microbial levels during blood-feeding. Graphical

Adult house flies frequent microbe-rich sites such as urban dumpsters and animal facilities, and encounter and ingest bacteria during feeding and reproductive activities. Due to unique nutritional and reproductive needs, male and female flies demonstrate different interactions with microbe-rich substrates and therefore dissemination potential. We investigated culturable aerobic bacteria and coliform abundance in male and female flies (n = 107) collected from urban (restaurant dumpsters) and agricultural (dairy farm) sites. Whole-fly homogenate was aerobically cultured and enumerated on nonselective (tryptic soy agar; culturable bacteria) and selective (violet-red bile agar, VRBA; coliforms) media. Unique morphotypes from VRBA cultures of agricultural flies were identified and tested for susceptibility to 14 antimicrobials. Female flies harbored more bacteria than males and there was a sex by site interaction with sex effects on bacterial abundance at the urban site. Coliform abundance did not differ by sex, site or sex within site. Both male and female flies carried antimicrobial-resistant (AMR) bacteria: 36/38 isolates (95%) were resistant to ≥1 antimicrobial, 33/38 were multidrug-resistant (≥2), and 24/38 isolates were resistant to ≥4 antimicrobials. Our results emphasize the role of house flies in harboring bacteria including AMR strains that pose a risk to human and animal health.

Strain CS-1^T, a novel facultative anaerobic bacterium, was isolated from the larval gastrointestinal tract of the biting midge, Culicoides sonorensis, a vector of the epizootic haemorrhagic disease virus and the bluetongue virus. Cells were Gram-stain-positive, non-motile, non-spore-forming, pleomorphic rods. Optimal growth occurred at pH 7.5 and 37 °C. The G+C content of the genomic DNA was 38.3 mol%, estimated by using HPLC. The dominant cellular fatty acids were C_14 : 0 (45.9 %) and C_16 : 0 (26.6 %). The polar lipid profile comprised glycolipids, diphosphatidylglycerol, phospholipids and phosphoglycolipids. Respiratory quinones were not detected. Strain CS-1^T had very low 16S rRNA gene similarity to members of the phylum Firmicutes : Macrococcus canis KM45013^T (85 % similarity) and Turicibacter sanguinis MOL361^T (88 % similarity). Phylogenetic analysis based on 16S rRNA, rpoB, gyrB genes, and conserved protein sequences of the whole genome revealed that strain CS-1^T was related to members of the classes Bacilli and Erysipelotrichia within the phylum Firmicutes . Furthermore, average nucleotide identity and digital DNA–DNA hybridization analyses of the whole genome revealed very low sequence similarity to species of Bacilli and Erysipelotrichaceae ( Macrococcus canis KM45013^T and Turicibacter sp. H121). These results indicate that strain CS-1^T belongs to the phylum Firmicutes and represents a new species of a novel genus, family, order and class. Based on the phenotypic, chemotaxonomic, phylogenetic and genomic characteristics, we propose the novel taxon Culicoidibacter larvae gen. nov., sp. nov. with the type strain CS-1^T (=CCUG 71726^T=DSM 106607^T) within the hereby new proposed novel family Culicoidibacteraceae fam. nov., new order Culicoidibacaterales ord. nov. and new class Culicoidibacteria classis nov. in the phylum Firmicutes .

House flies (Musca domestica Linnaeus) are vectors of human and animal pathogens at livestock operations. Microbial communities in flies are acquired from, and correlate with, their local environment. However, variation among microbial communities carried by flies from farms in different geographical areas is not well understood. We characterized bacterial communities of female house flies collected from beef and dairy farms in Oklahoma, Kansas, and Nebraska using 16S rDNA amplicon sequencing and PCR. Bacterial community composition in house flies was affected by farm type and location. While the shared number of taxa between flies from beef or dairy farms was low, those taxa accounted >97% of the total bacterial community abundance. Bacterial species richness was 4% greater in flies collected from beef than in those collected from dairy farms and varied by farm type within states. Several potential pathogenic taxa were highly prevalent, comprising a core bacterial community in house flies from cattle farms. Prevalence of the pathogens Moraxella bovis and Moraxella bovoculi was greater in flies from beef farms relative to those collected on dairy cattle farms. House flies also carried bacteria with multiple tetracycline and florfenicol resistance genes. This study suggests that the house flies are significant reservoirs and disseminators of microbial threats to human and cattle health.