List of Drugs/vaccines screened or developed against Infectious diseases in
Drosophila melanogaster is a widely used, dynamic model organism to study various pathogenic diseases observed ubiquitously in the human population. Drosophila, at present, is extensively used to conduct preclinical studies besides its counterpart rodents. The epidemic and pandemic diseases are discussed in this review to demonstrate Drosophila melanogaster as a key model. Epidemic and pandemic diseases are still claiming more than 5 million lives every year, and these diseases were well studied in flies. Currently there is no cure for the disease like HIV; the bacterial and fungal infections usually seen in HIV/AIDS patients could be demonstrated elaborately in Drosophila melanogaster. Diseases like myocardial infractions and cancer causing viral infection are long term effects of ART (anti-retroviral therapy) that could be experimented in flies. Stable Drosophila S2 cell line, Transgenic flies, transfusion of bacteria and fungi could be implemented to study several infectious diseases and for vaccine development. The latest trends in understanding pathogenic diseases and its potential biochemical markers in flies are discussed in this review to utilize the fruit flies as a functional tool and to explore further it in drug development. The advantages and disadvantages of the fly as a model of infection are discussed along with the epidemiology and the cellular pathophysiology
- Drosophila melanogaster
- viral diseases
- epidemic and pandemic diseases
1.1 Epidemic disease
The term epidemic is derived from Greek word “epi” meaning “upon” and “demos” meaning “people”. It refers to a communicable disease which spreads rapidly in a given population within a very short period of time. Any infectious disease existing in a region does not make it epidemic unless it causes faster mortality. A death rate of around 1.6 folds higher than usual death rate (baseline) caused by a disease in a population within a fixed period could be considered as an epidemic disease. A disease lower than this fold increase, observed in a population could be designated as an outbreak of a disease .
Diseases like tuberculosis, hepatitis, yellow fever, chikungunya, ebola virus disease, marburg virus disease, Crimean-Congo haemorrhagic fever, rift valley fever, typhoid fever, Shigellosis, plague, lassa fever, West Nile fever, zika virus disease, meningitis, MERS-CoV, plague, monkeypox, nodding syndrome, nipah virus infection are considered as epidemic diseases as per World Health Organization . Epidemic diseases like plague, small pox and cholera caused unsurpassed deaths in human population till the end of eighteenth century .
1.2 Pandemic disease
The term pandemic is derived from Greek word “pan” meaning “all” and “demos” meaning “people”. It refers to an epidemic disease which spreads among large population possibly across geographic locations or continents within a short time span .
Influenza, along with viral pneumonia, HIV and cholera are considered as pandemic disease and caused millions to die beside high rate of hospitalization and life threatening conditions across the globe . The viral diseases like Influenza, cholera and HIV caused maximum deaths in the twenty-first century .
Vaccines are available for most of the epidemic and pandemic diseases . Vaccination is the most effective prevention technique to suppress the infection in healthy population . However, poor and conflicted regions of Asia and Africa are deprived of these vaccines . World Health Organization plays a major role in epidemic preparedness in these regions and provides extended healthcare facilities during an epidemic outbreak .
1.4 Global requirement
Considering, the disease outbreak and its transmission is high in a poor population of developing region . First line and second line antibiotics are the most effective medicines for infected subjects as the vaccines are ineffective after the infection had taken place. First line therapy includes antibiotics are the most commonly prescribed medicines to alleviate the infection process, often not responsive on several types of multi drug resistant infection . Hence it is important to select a cost effective model to screen the first line antibiotics or antivirals.
1.5 Fruit fly as a model organism for drug screening
Today, we need to discover more efficacious antibiotics to fight the infectious diseases.
2. Epidemiology of infectious diseases
2.1 Lower respiratory tract infection epidemiology
As per World Health Organization lower respiratory tract infections are caused mainly by influenza, pneumococcal pneumonia and viral pneumonia are responsible for 3 million deaths . The WHO reported in 2018, 3–5 million cases of Influenza with 290,000–650,000 death cases annually . As per the Global Disease burden (GDB) study report of 2015 there were around 1.5 million deaths in all age groups caused due to pneumococcal pneumonia . SARS (Corona virus) causes viral pneumonia; it is epidemic to more than 30 countries with 8000 reported cases and 774 deaths during the year 2002–2003. MERS is a viral pneumonia causing infections in 688 persons and 282 deaths reported by WHO in 20 countries during 2012 .
Influenza originates from
2.3 Pneumonia and viral pneumonia
Pneumonia is caused due to several communicable infections usually known as community acquired pneumonia (CAP), often seen in hospitalized patients. Pneumonia can be caused by bacteria like
Tuberculosis is caused by
Currently HIV is the most fatal disease observed in human population across the globe. It caused maximum number of deaths around the world in the last 3 decades. As per the latest WHO report of 2019, HIV/AIDS have claimed more than 35 million deaths till date. Currently 36.9 million (31.1–43.9 million) peoples are living with HIV as of 2017 . Although the rate of infection has decreased in the recent years, still HIV remains a global burden on world economy.
2.6 Diarrhoeal disease
The term “cholera” was derived from Sanskrit meaning “stomach disturbance” . Since, early 1800 century cholera outbreak turned out to be pandemic and caused millions to die, altogether six different pandemics took place the seventh started in the year 1961 and is still ongoing [23, 24]. In 2019 WHO report suggests 1.3 million to 4.0 million cases of cholera with an estimated 21,000–143,000 deaths worldwide .
Viral hepatitis is one of the most life threatening disease, it causes death to 1.4 million peoples across the globe reported in 2018 . Globally around 260 million peoples are infected with HBV and 71 million with HCV infections are reported causing 90% of deaths among viral hepatitis patients . The HBV and HCV has the highest prevalence rate in the global population at present, hepatitis viruses like HAV, HAD and HEV are endemic in many countries . Currently there is no vaccine available for HCV till date.
The term Typhoid was coined from the Greek word “typhus” which means “Smoky” was used to relate the delirium symptom often associated with typhoid fever . Typhoid fever is caused by gram-negative bacteria known as
Malaria fever is a severe parasitic disease caused by
2.10 Viral meningitis, viral encephalitis and hemorrhagic fever viruses
Viruses like herpes simplex virus HSV, HIV, mumps virus, measles virus and west Nile virus causes meningitis which causes frequent outbreaks in some regions . Japanese encephalitis virus along with genus
3. Drosophila model to study highly infectious diseases
There are at present several bacterial, fungal and viral models of infection which were successfully demonstrated to infect flies and used it to understand drug efficacy. Drosophila model of infectious disease could be very low cost model to study drug efficacy in-vivo; it could help to save lives by saving time during an epidemic outbreak. Understanding the disease pathogenesis in humans and drawing out a similar model in
In the recent past several research works has been conducted to understand the immune system of
4. Host-pathogen interaction
The bacterial and fungal infection leads to the activation of dToll, Imd, Eiger (TNF family homolog) and insulin like receptors (FOXO) in
The fungal pathogen was found to be recognized by GNBP3 along with PGRPSA and GNBP1 it activates the drosophila toll receptors . The Drosophila toll-5 (Tehao) and toll-9 plays major role during fungal infection by inducing Drosomycin gene .
During the preliminary stage of viral infection Drosophila toll receptor homolog of human TLR, Imd (TNF-alpha), Domeless (Jak–STAT), and RNAi plays a major role against viral infection these are components of innate immune system . Similar to humans the viral glycoproteins are recognized by toll receptors like toll-4, while toll-7 dependent autophagy observed during viral infection in flies [42, 46]. Jak–STAT and Imd together mediates effective immunity against viral attack in flies . The domeless-hop-stat2 pathway stimulated by upd1/2/3 activates Jak–STAT regulated genes responsible for controlling viral load; it is homologous to mammalian Jak–STAT pathway . The Drosophila P53 and dP38 mediates apoptosis in flies upon stress response generated due to DNA damage, P53 mediated apoptotic genes are regulated by Jak–STAT-MAPK . The dP38 stimulation in flies triggers Unpraired gene (upd protein) a mammalian IL-6 homolog further activates Jak–STAT-
5. Markers of infectious diseases
In the recent decades extensive research has been conducted to understand the regulation of immune system in
6. Behavioral and physiological characterization of infected flies
6.1 Negative geotaxis assay
Negative geotaxis assay serves the purpose to manifest ongoing pathogenesis inside the live model. It was demonstrated previously that infected flies display significantly lesser motility than healthy flies when exposed under bright light. It could be considered as an important parameter to explain drug efficacy while screening anti-microbial drugs in flies .
6.2 Circadian rhythm
Circadian rhythm in flies was studied, the genes timeless or period controls the circadian rhythm of activity-sleep cycle during day-night respectively. It has been observed that infected flies exhibit interrupted circadian control of locomotion thus flies with this deficit shows restlessness at the same time gets lesser sleep than normal flies. The behavioral changes could also be studied in infected flies beside the control/uninfected flies .
7. Factors contributing to suitable infection model
It was previously reported that in order to replicate the outcome of future studies it is important to optimize the lethal dosage selection and the route of inoculum . It is suggested that the selection of microbial strain and gender of flies are two important factors which could potentially impact the findings of future research.
7.1 Route of inoculum
There are two prime techniques for inducing infection in flies, primarily by feeding the flies with the microbes secondly by pricking micro needles dipped in bacterial liquid (inoculum) into fly’s abdomen or thorax [62, 63]. Flies could be pricked in the abdomen with micro-needle dipped in the microbial solution, known amount can be useful in pharmacodynamic as well as pharmacokinetic studies .
7.2 Flies gender selection
Selecting gender should be considered strictly, few studies do not prefer to report the reason behind choosing the gender male/female type. In a study with
8. In-vivo models for epidemic and pandemic diseases
The existing models using live bacterial infusion, feeding fungal strains and transgenic flies expressing viral proteins. Under immuno-suppressed condition would serve multiple purposes like studying host pathogen interaction and conducting preclinical trials [62, 66].
8.1 HIV models
Since human viruses do not usually invade insects the use of
The incapacity of Drosophila S2 cells is only associated with the expression of HIV-1 envelope proteins. It is possible to express gycosylated and cleaved Gp120 in S2 cells but fusion with CD4+ receptors of T-helper cells could not be achieved in the model expression system . In another study the expression of Gp120 in drosophila was carried out in S2 cell line, the antigen Gp-120 did not exhibited T-helper cell mediated humoral immune system activation and IgG antibody generation, when introduced in mice . Due to this usual challenge in a different study they expressed HIV-1 virus like proteins in Drosophila S2 cells .
The nef transgenic flies exhibited JNK mediated apoptosis further nef inhibits NF-kB necessary for Relish gene activation similarly decreased immune response is common in AIDS patients . In a study transfected viral protein Vpu was shown to cause immune suppression in fat body of flies via toll dependent pathway, in wings the Vpu expression caused apoptosis and hindered wing development, in mammals Vpu is known for causing T-cell lymphocyte death in infected patients [74, 75]. Active microbial invasion in nef flies should be further confirmed before targeting with potent anti-nef drug candidate. The
Cryptococcosis, Candidiasis and Aspergillosis are common types of fungal infections observed as clinical challenge in HIV-positive patients . Under immunosuppressed condition the invasion of fungi in flies causes fatality. In Drosophila fungal infection could be difficult to achieve as the innate immune system mediates anti-fungal peptide production by haemocyte causing decrease of fungal load and increases fly survival rate . Hence Toll mutant flies were generated and used to induce fungal infection.
Fluconazole and voriconazole showed anti-fungal activity against
In order to study HPV and EBV there are two model systems to study the effect in flies. In the study with HPV co-expression of viral oncoprotein E6 and human UBE3A did not resulted in tumorigenesis requires Ras or Notch pathway in flies, E6-UBE3A requires insulin receptors for cancer to develop . Upon introducing the BZLF1 gene of EBV led to interaction with shaven gene in flies a homolog of pax gene family of humans responsible for B-cell development . Expression of BRLF1 and BZLF1 genes using
8.2 Influenza infection models
Influenza virus like most other viruses fails to infect the
8.3 Pneumococcal pneumonia models
There are several pneumococcal pneumonia infection model studied in drosophila using
8.4 Tuberculosis models
There are at present two bacterial models for studying mycobacterium infection in flies, induced by
8.5 Cholera models
9. Importance of in-vitro model infection in Drosophila
The Drosophila S2 cells were first discovered by I. Schneider in 1972 . S2 cells are derived from primary cell culture of late phase embryo of
10. In-vitro model of epidemic and pandemic infectious disease
Using drosophila S2 cell model a study showed that intercellular
11. Viral meningitis, encephalitis and hemorrhagic fever S2 cell line model
Herpes simplex virus was studied in Drosophila S2 cells where transfection of two viral proteins PILRα and gB responsible for binding to mammalian cells were expressed found to be poorly glycosylated . The RNAi pathway was indulged by host cells to inhibit the Dengue virus (Flavivirus family) infection, by knocking down Argonaute (Ago1/2) and Dicer (Dcr1/2) showed sustained viral infection, currently clinical trial is underway NCT00936429 [115, 116]. Japanese encephalitis virus envelope glycoprotein E transfected in Drosophila S2 cells resulted in stable protein expression, this glycoprotein exposure in mice led antibody production against it . Infection of Sindbis virus in live flies led activation of Notch, Jak–STAT and ImD pathway to intervene viral invasion . Notch pathway mediated assimilation of ankyrin, plap, syx13, unc-13, csp, rab1 and rab8 during Sindbis virus infection in S2 cells . The human antibody MR191 specific against Marburg virus was fused with recombinant RAVV GP ectodomain produced in S2 cell line . The Zika virus structural envelope (E) protein were efficiently produced and secreted from transfected Drosophila S2 cell line model . Flies produces RNAi against west Nile virus infection as a result of innate immune response similarly it was seen in S2 cell line, S2 cell lines were used for WNV infection, currently vaccine development NCT01477580 and NCT00707642 is underway [116, 120]. In a study mice were injected with glycoprotein GP of Ebola virus expressed in Drosophila S2 cell line found to produce antibodies against the infused antigen  (Table 1).
|Epidemic/Pandemic Disease||Microbes||Vaccine/Drugs screened or derived out of fly model (in-vitro/in-vivo)||References|
|HIV/AIDS||Human Immuno virus||Leptomycin B (In-Vitro) Unapproved under clinical trials|
Zidovudine, lamivudine, stavudine, didanosine, Abacavir
|Tuberculosis||Rifampicin, Tigecycline + Linezolid||[100, 101]|
|SARS||SARS Corona virus||—|
|MERS||MERS corona virus||—|
|Hepatitis||Hepatitis A and B||HBsAg expressed in S2 cell line|||
|Small pox||Variola virus||—|
|Malaria||VAR2CSA/PfRH5 viral protein expressed in S2 cell line||[112, 113]|
|Zika Fever||Zika virus||Structural envelope (E) protein expressed in S2 cells|||
|Dengue Fever||Dengue Virus||DEN1-80E expressed in S2 cells||[115, 116]|
|Encephalitis||Japanese encephalitis virus||JEV E protein expression in S2 cell line|||
|Haemorrhagic fever||Ebola virus||glycoprotein GP expressed in S2 cell line|||
|Haemorrhagic fever||Marburg Virus||MR191 expressed in S2 cell line|||
|Yellow fever||Yellow fever virus||—|
|West Nile Fever||West Nile Virus||WN-80E expressed in S2 cell line||[116, 120]|
12. Disadvantages of Drosophila model for drug screening
13. Future perspective
Irrespective of multiple disadvantages flies could be used for studying drug efficacy. Multi-drug resistance tuberculosis infection could be studied in flies. The ART medication impairs human heart by causing prolonged QT, prolonged arrhythmic condition leads to myocardial infraction,
Currently the existing models of infection in drosophila are capable of causing infection using viruses, bacteria (gram negative and gram positive) and fungi. These models are of great use since the efficacy of a drug capable of modifying diseased condition could be studied in detail in live
I would like to thank Professor Sarat Chandra Yenisetti, Nagaland University, India for the effort and advices given for this article. I would like to thank Professor David S. Schneider of Stanford University, USA for clearing doubts regarding tuberculosis and typhoid infection in flies.
Conflict of interest
The author has no conflict of interest.
I would like to thank the IntechOpen Journal for giving 100% waiver to publish this review article. I would like to thank all the researchers for providing their complete articles which are unavailable online.