Staphylococcus aureus in the house fly: temporospatial fate of bacteria and expression of the antimicrobial peptide defensin

Authors: Nayduch, Dana; CHO, HANNA - Georgia Southern University; JOYNER, CHESTER - Emory University

Submitted to: Journal of Medical Entomology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/1/2012
Publication Date: 1/1/2013
Citation: Nayduch, D., Cho, H., Joyner, C. 2013. Staphylococcus aureus in the house fly: temporospatial fate of bacteria and expression of the antimicrobial peptide defensin. Journal of Medical Entomology. 50(1): 171-178.

Interpretive Summary: House flies feed and breed in feces, garbage and other microbe-rich habitats. Because of these activities, flies often harbor and disseminate bacteria, some pathogenic to humans and livestock. Previous surveys of house flies have detected the pathogen Staphylococcus aureus, a bacteria species that causes skin and wound infections, toxic shock syndrome and food poisoning. The potential of flies to disseminate, transport or vector this bacterium is not known. In this paper, we used a transformed strain of S. aureus that expressed a green-flourescent protein (GFP) tag so that we could track its association with house flies. Flies were fed known amounts of bacteria (ranging from about 150,000 to 350,000 colonies) and then processed as follows. In the first experiment, flies were dissected to remove the alimentary canal and view bacteria within regions of the gut by microscopy at 2, 4 and 6 h post ingestion. In the second experiment, flies were fed bacteria and then homogenized for culture analysis (at 2, 4, and 6 h), or flies were housed individually so we could collect and culture their excrement (at 2, 3, and 4 h). Finally, we fed flies bacteria and looked at the expression of an antimicrobial gene defensin, part of the house fly immune system involved in destroying bacteria, in respect to location in the gut or systemically in the body at 2, 4, and 6 h. At 2 h post-ingestion (PI), live bacteria were present in the crop, an anterior storage sack in the fly gut from which vomitus is expelled, and the midgut, which is analogous to the intestine. As soon as 4 h PI, GFP-S. aureus were visualized in feces in the hindgut and rectum of flies, but many were destroyed and non-viable. Bacteria persisted up to 6 h PI but significantly decreased over time. Excretion of viable GFP-S. aureus peaked at 2 h PI and, although significantly less, continued up to 4 h PI. defensin was detected at high levels locally in the digestive tract (proximal to bacteria) and systemically in fat body (which sits in the fly body cavity) at 2, 4 and 6 h PI. To our knowledge, this is the first study to report a local and systemic response to a bacterium with lysine-type peptidoglycan, such as S. aureus, in flies exposed to bacteria by feeding. While flies harbored S. aureus for up to 6 h PI, our data indicate that the highest probability of vectoring biologically-relevant amounts of bacteria occurred between 0-2 h PI. The combined effects of excretion, digestion and antimicrobial responses likely contribute to loss of ingested bacteria. Nonetheless, we demonstrated that house flies are relevant vectors for S. aureus up to 2 h PI and environmental reservoirs up to 6 h PI. These findings indicate that in both human and livestock environments where flies have opportunities to ingest this pathogen, they would be able to disseminate bacteria effectively, and therefore serve as source of cross contamination between otherwise clean and filthy areas.

Technical Abstract: House flies disseminate numerous species of bacteria acquired during feeding and breeding activities in microbe-rich habitats. Previous house fly surveys have detected the pathogen Staphylococcus aureus, which causes cutaneous and septic infections in mammals and enterotoxic food poisoning. We assessed the fate of GFP-expressing S. aureus (GFP-S. aureus) in the house fly alimentary canal with microscopy and by culture of whole flies and excreta. Furthermore, the concurrent expression of the antimicrobial peptide gene defensin (def) was measured in the crop, proventriculus, midgut and fat body. As soon as 4 h post-ingestion (PI), GFP-S. aureus were visualized as cocci or diplococci in the hindgut and rectum of flies fed ˜105 CFU. Bacteria persisted up to 6 h PI but significantly decreased. Excretion of viable GFP-S. aureus peaked at 2 h PI and, although significantly less, continued up to 4 h PI. def was highly upregulated locally in the alimentary canal and systemically in fat body at 2, 4 and 6 h PI making this study the first to report, to our knowledge, an epithelial and systemic response to a bacterium with lysine-type peptidoglycan in flies exposed via feeding. While flies harbored S. aureus for up to 6 h PI, the highest probability of vectoring biologically-relevant amounts of bacteria occurred 0-2 h PI. The combined effects of excretion, digestion and antimicrobial effectors likely contribute to loss of ingested bacteria. Nonetheless, house flies are relevant vectors for S. aureus up to 2 h PI and environmental reservoirs up to 6 h PI.