Tobacco Hornworm (Manduca sexta) caterpillars as a novel host model for the study of fungal virulence and drug efficacy

Abstract

29Pathogenic yeast species can cause life-threatening infections in humans. The two leading 30 yeast pathogens, Candida albicans and Cryptococcus neoformans, cause systemic infections 31 in >1.4 million patients world-wide with mortality rates approaching 75%. It is thus 32 imperative to study fungal virulence mechanisms, stress response pathways, and the efficacy 33 of antifungal drugs. This is commonly done using mammalian models. To address ethical and 34 practical concerns, invertebrate models, such as wax moth larvae, nematodes, or flies, have 35 been introduced over the last two decades. To address short-comings in existing invertebrate 36 host models, we developed fifth instar caterpillars of the Tobacco Hornworm moth Manduca 37 sexta as a novel host model for the study of fungal virulence and drug efficacy. These 38 caterpillars can be raised at standardised conditions, maintained at 37˚C, can be injected with 39 defined amounts of yeast cells, and are susceptible to the most threatening yeast pathogens, 40 including C. albicans, C. neoformans, C. auris, and C. glabrata. Infected caterpillars can be 41 rescued by treatment with commonly deployed antifungal drugs and importantly, fungal 42 burden can be assessed daily throughout the course of infection in a single caterpillar's faeces 43 and hemolymph. Notably, these animals are large enough so that weight provides a reliable 44 and reproducible measure of fungal virulence. This model combines a suite of parameters that 45 recommend it for the study of fungal virulence. 46 47 50 tuberculosis 1 . The leading yeast pathogens, Candida albicans and Cryptococcus neoformans, 51 together account for >1,400,000 life-threatening infections world-wide with mortality rates 52 approaching 75% 1 . Candidemia, most commonly caused by C. albicans, is the fourth most 53 common cause of nosocomial blood stream infections, only surpassed by infections with 54Staphylococci and Enterococcus spp. Disturbingly, candidemia incidence rates are on the 55 rise. Within less than ten years, incidence rates increased by 36% 2 . Although cryptococcosis 56 incidence rates are on the decline in North America, this AIDS-defining illness is responsible 57 for 15% of all AIDS-related deaths world-wide 2,3 . This already dire situation is further 58 confounded by the emergence of drug resistant yeast species. Patients at risk of developing 59 invasive candidemia are often prophylactically treated with fluconazole, while the 60 echinocandins are considered a first line defence strategy 4 . Yet, C. glabrata, the most 61 3 common non-albicans Candida species associated with nosocomial blood stream infections 5 , 62 is intrinsically less susceptible to azole drugs and acquires resistance to echinocandins 63 rapidly 6 . The rapid global spread of multi-drug resistant C. auris has further exacerbated the 64 threat posed by fungi. C. auris was first reported in 2009 in Japan 7 . In 2015, C. auris arrived 65 in Europe causing an outbreak involving 72 patients in a cardio-thoracic hospital i...

De novogenome assembly of the Tobacco Hornworm moth (Manduca sexta)

“…As a large, easily grown insect, the tobacco hornworm ( M. sexta, NCBI:txid7130 ) is a key model for studying biochemical mechanisms in insect biochemistry, physiology, neurobiology, development, and immunity [1 , 2 , 3 , 4] . In particular, its innate immune pathways, many of which share mammalian homologs, have been characterized in detail and used as a model system for studying fungal virulence and drug efficacy [5,6] . M. sexta metamorphosis has also proved an excellent system for studying development, particularly the complex interactions between hormones and environmental regulators, e.g.…”