The study investigated the transmission of Salmonella enterica, Cronobacter sakazakii, Escherichia coli O157:H7, and Listeria monocytogenes from adult house flies to their eggs and first filial (F1) generation adults. The researchers fed adult house flies food contaminated with low, medium, and high levels of each pathogen. They found that all pathogens were present in samples of pooled house fly eggs. Transmission was highest when adults consumed medium bacterial loads. Cronobacter sakazakii was most likely to be transmitted to eggs. S. enterica and C. sakazakii were transmitted to F1 adults and more likely to be found on their surfaces than in their guts
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
All manuscripts are subject to rapid peer review. Those of high quality (not previously published and not under consideration for publication in another journal) will be published without delay.
Depopulation options as welfare indicator for layer systemsHarm Kiezebrink
Egg production systems have become subject to heightened levels of scrutiny. Multiple factors such as disease, skeletal and foot health, pest and parasite load, behavior, stress, affective states, nutrition, and genetics influence the level of welfare hens experience. Although the need to evaluate the influence of these factors on welfare is recognized, research is still in the early stages.
In this paper conventional cages are compared to furnished cages, non-cage systems, and outdoor systems. Specific attributes of each system are shown to affect welfare, and systems that have similar attributes are affected similarly.
Environments such as conventional cages, which limit movement, can lead to osteoporosis, but environments that have increased complexity, such as non-cage systems, expose hens to an increased incidence of bone fractures.
Less is understood about the stress that each system imposes on the hen, but it appears that each system has its unique challenges. Selective breeding for desired traits such as improved bone strength and decreased feather pecking and cannibalism may help to improve welfare.
It appears that no single housing system is ideal from a hen welfare perspective. Although environmental complexity increases behavioral opportunities, it also introduces difficulties in terms of disease and pest control.
One specific circumstance has not been taken into consideration in this paper: how to depopulate the hens in case of an outbreak situation. Emergency control is not an economic parameter to choose a specific production system, but comparing a production system with or without cages, it is clear that it is much easier to depopulate chickens in a system without cages. Without a proper technique to cull the animals in a animal welfare friendly way and to transport the carcasses out of the house mechanically, the chickens are killed and transported manually.
This is not only increasing the risks for humans to get infected, it also influences the risks that animals suffer unnecessary during depopulation. Handling animals during outbreak situations is mostly done by inexperienced responders who have little to no knowledge about animal welfare. Veterinary authorities in charge of the response activities have issues like effectiveness and efficiency to consider.
How to depopulate the chickens in an outbreak situation is an important welfare indicator and the producer of these systems need to be kept responsible for the technical solution.
Harm Kiezebrink
Research Fellow Queensland University /
CEO AVT Europe AB
AVT Applied Veterinary Technologies Europe AB
Address details: c/o INTRED, Södra Hamnen 2,
45142 Uddevalla, Sweden
Phone: +44 7452 272 358
E-mail: harm.kie@gmail.com
Presented by Kristina Roesel and Delia Grace at “Microsporidia in the Animal to Human Food Chain: An International Symposium to Address Chronic Epizootic Disease”, Vancouver, Canada, 9-13 August 2015.
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
All manuscripts are subject to rapid peer review. Those of high quality (not previously published and not under consideration for publication in another journal) will be published without delay.
Depopulation options as welfare indicator for layer systemsHarm Kiezebrink
Egg production systems have become subject to heightened levels of scrutiny. Multiple factors such as disease, skeletal and foot health, pest and parasite load, behavior, stress, affective states, nutrition, and genetics influence the level of welfare hens experience. Although the need to evaluate the influence of these factors on welfare is recognized, research is still in the early stages.
In this paper conventional cages are compared to furnished cages, non-cage systems, and outdoor systems. Specific attributes of each system are shown to affect welfare, and systems that have similar attributes are affected similarly.
Environments such as conventional cages, which limit movement, can lead to osteoporosis, but environments that have increased complexity, such as non-cage systems, expose hens to an increased incidence of bone fractures.
Less is understood about the stress that each system imposes on the hen, but it appears that each system has its unique challenges. Selective breeding for desired traits such as improved bone strength and decreased feather pecking and cannibalism may help to improve welfare.
It appears that no single housing system is ideal from a hen welfare perspective. Although environmental complexity increases behavioral opportunities, it also introduces difficulties in terms of disease and pest control.
One specific circumstance has not been taken into consideration in this paper: how to depopulate the hens in case of an outbreak situation. Emergency control is not an economic parameter to choose a specific production system, but comparing a production system with or without cages, it is clear that it is much easier to depopulate chickens in a system without cages. Without a proper technique to cull the animals in a animal welfare friendly way and to transport the carcasses out of the house mechanically, the chickens are killed and transported manually.
This is not only increasing the risks for humans to get infected, it also influences the risks that animals suffer unnecessary during depopulation. Handling animals during outbreak situations is mostly done by inexperienced responders who have little to no knowledge about animal welfare. Veterinary authorities in charge of the response activities have issues like effectiveness and efficiency to consider.
How to depopulate the chickens in an outbreak situation is an important welfare indicator and the producer of these systems need to be kept responsible for the technical solution.
Harm Kiezebrink
Research Fellow Queensland University /
CEO AVT Europe AB
AVT Applied Veterinary Technologies Europe AB
Address details: c/o INTRED, Södra Hamnen 2,
45142 Uddevalla, Sweden
Phone: +44 7452 272 358
E-mail: harm.kie@gmail.com
Presented by Kristina Roesel and Delia Grace at “Microsporidia in the Animal to Human Food Chain: An International Symposium to Address Chronic Epizootic Disease”, Vancouver, Canada, 9-13 August 2015.
Ensuring successful introduction of Wolbachia in natural populations of Aedes...FGV Brazil
The control of the spread of dengue fever by introduction of the intracellular parasitic bacterium Wolbachia in populations of the vector Aedes aegypti, is presently one of the most promising tools for eliminating dengue, in the absence of an efficient vaccine. The success of this operation requires locally careful planning to determine the adequate number of individuals carrying the wolbachia parasite that need to be introduced into the natural population. The introduced mosquitoes are expected to eventually replace the Wolbachia-free population and guarantee permanent protection against the transmission of dengue to human. In this study, we propose and analyze a model describing the fundamental aspects of the competition between mosquitoes carrying Wolbachia and mosquitoes free of the parasite. We then use feedback control techniques to devise an introduction protocol which is proved to guarantee that the population converges to a stable equilibrium where the totality of mosquitoes carry Wolbachia.
Date: 2015-03-19
Authors:
Bliman, Pierre-Alexandre
Soledad Aronna, Maria
Coelho, Flávio Codeço
Silva, Moacyr da
Relations between pathogens, hosts and environmentEFSA EU
Presentation of the EFSA's second scientific conference, held on 14-16 October 2015 in Milan, Italy.
DRIVERS FOR EMERGING ISSUES IN ANIMAL AND PLANT HEALTH
Discovering novel pathways of cross-species pathogen transmissionEFSA EU
Presentation of the EFSA's second scientific conference, held on 14-16 October 2015 in Milan, Italy.
DRIVERS FOR EMERGING ISSUES IN ANIMAL AND PLANT HEALTH
MERS has to be tackled more practically ,its nothing to scare unless you find and suspicious case around you.The contributing factors are ,weather ,closed homes ,shisha culture ,and the anatomy of Arabs nostrils play a aggressive role in spread of this new disease.The virus mutated recently in a more more cases in humans appeared in hospitals in Jeddah, which may indicate increased virus transmission from man to man due to mutation in the genome leading to virus adaptation. This event may be associated with loss of some virulence elements in the virus.”to survive, viruses adapt or evolve, changing its surface proteins enough to trick the host cell into allowing it to attach.
Abstract
Study was conducted to record prevalence of gastrointestinal parasites of cat. A total of 100 fecal samples from cat (50 from
male and 50 from female) were collected and examined for the presence of GIT parasites. Samples were collected and
transported for the laboratory diagnosis. Animal data such as age, sex, and breed were recorded. Results of the present study
revealed that 24% cats found positive for the gastrointestinal parasites. The percentage of infection was found higher in female
cats (28%) than males (20%). The prevalence in adults and kitten was recorded as 21.42% and 30% respectively. Dipylidium
caninum was found more prevalent with the infection rate of 9% followed by Toxocara cati, Aeluroslonglun obstrusus, Taenia
taeniaeformis and Paragonimus kellikotti with the infection rate of 5, 5, 3 and 2% respectively.
Key words: Cat, Felus catus, GIT parasites, Prevalence
Human-bat interactions and diseases: transmission risks in GhanaNaomi Marks
Presentation by Professor Yaa Ntiamoa-Baidu of the University of Ghana at the One Health for the Real World: zoonoses, ecosystems and wellbeing symposium, London 17-18 March 2016
Prevalence and resistance of bacterial strains isolated from chicken beddings...IOSRJAVS
The main interest of researchers is focused on the microbiology of the industrial poultry beddings. In this study the microbiology and the microbial resistance of strains isolated from composite samples of poultry (gallus gallus domesticus) beddings originating from rural households has been investigated. In the area of Arta (Epirus, Greece) samples were collected from 300 rural households. These samples were classified regarding the following 4 criteria: (a) the size of the chicken flock, (b) the presence of different poultry species in the same household, (c) the presence of small ruminants in the same household and (d) differences in feeding practices. Results reveal that the microbiology of the beddings was mostly affected by the presence of small ruminants in the same household and the administration of concentrated feeds. Microbial resistance followed the same distribution pattern. The most resistant strains were isolated from samples originating from households breeding both poultry and small ruminants. Feeding with concentrated feeds was a determinant factor and probably the link between resistance and prevalence.
Ensuring successful introduction of Wolbachia in natural populations of Aedes...FGV Brazil
The control of the spread of dengue fever by introduction of the intracellular parasitic bacterium Wolbachia in populations of the vector Aedes aegypti, is presently one of the most promising tools for eliminating dengue, in the absence of an efficient vaccine. The success of this operation requires locally careful planning to determine the adequate number of individuals carrying the wolbachia parasite that need to be introduced into the natural population. The introduced mosquitoes are expected to eventually replace the Wolbachia-free population and guarantee permanent protection against the transmission of dengue to human. In this study, we propose and analyze a model describing the fundamental aspects of the competition between mosquitoes carrying Wolbachia and mosquitoes free of the parasite. We then use feedback control techniques to devise an introduction protocol which is proved to guarantee that the population converges to a stable equilibrium where the totality of mosquitoes carry Wolbachia.
Date: 2015-03-19
Authors:
Bliman, Pierre-Alexandre
Soledad Aronna, Maria
Coelho, Flávio Codeço
Silva, Moacyr da
Relations between pathogens, hosts and environmentEFSA EU
Presentation of the EFSA's second scientific conference, held on 14-16 October 2015 in Milan, Italy.
DRIVERS FOR EMERGING ISSUES IN ANIMAL AND PLANT HEALTH
Discovering novel pathways of cross-species pathogen transmissionEFSA EU
Presentation of the EFSA's second scientific conference, held on 14-16 October 2015 in Milan, Italy.
DRIVERS FOR EMERGING ISSUES IN ANIMAL AND PLANT HEALTH
MERS has to be tackled more practically ,its nothing to scare unless you find and suspicious case around you.The contributing factors are ,weather ,closed homes ,shisha culture ,and the anatomy of Arabs nostrils play a aggressive role in spread of this new disease.The virus mutated recently in a more more cases in humans appeared in hospitals in Jeddah, which may indicate increased virus transmission from man to man due to mutation in the genome leading to virus adaptation. This event may be associated with loss of some virulence elements in the virus.”to survive, viruses adapt or evolve, changing its surface proteins enough to trick the host cell into allowing it to attach.
Abstract
Study was conducted to record prevalence of gastrointestinal parasites of cat. A total of 100 fecal samples from cat (50 from
male and 50 from female) were collected and examined for the presence of GIT parasites. Samples were collected and
transported for the laboratory diagnosis. Animal data such as age, sex, and breed were recorded. Results of the present study
revealed that 24% cats found positive for the gastrointestinal parasites. The percentage of infection was found higher in female
cats (28%) than males (20%). The prevalence in adults and kitten was recorded as 21.42% and 30% respectively. Dipylidium
caninum was found more prevalent with the infection rate of 9% followed by Toxocara cati, Aeluroslonglun obstrusus, Taenia
taeniaeformis and Paragonimus kellikotti with the infection rate of 5, 5, 3 and 2% respectively.
Key words: Cat, Felus catus, GIT parasites, Prevalence
Human-bat interactions and diseases: transmission risks in GhanaNaomi Marks
Presentation by Professor Yaa Ntiamoa-Baidu of the University of Ghana at the One Health for the Real World: zoonoses, ecosystems and wellbeing symposium, London 17-18 March 2016
Prevalence and resistance of bacterial strains isolated from chicken beddings...IOSRJAVS
The main interest of researchers is focused on the microbiology of the industrial poultry beddings. In this study the microbiology and the microbial resistance of strains isolated from composite samples of poultry (gallus gallus domesticus) beddings originating from rural households has been investigated. In the area of Arta (Epirus, Greece) samples were collected from 300 rural households. These samples were classified regarding the following 4 criteria: (a) the size of the chicken flock, (b) the presence of different poultry species in the same household, (c) the presence of small ruminants in the same household and (d) differences in feeding practices. Results reveal that the microbiology of the beddings was mostly affected by the presence of small ruminants in the same household and the administration of concentrated feeds. Microbial resistance followed the same distribution pattern. The most resistant strains were isolated from samples originating from households breeding both poultry and small ruminants. Feeding with concentrated feeds was a determinant factor and probably the link between resistance and prevalence.
Active listening is a work in progress because listening never ends. It takes ongoing practice to learn to be active listens, but the effort is worthwhile.
Las mentiras de Donald Trump de acuerdo con Economistas de USASusana Gallardo
Un documento firmado por miles de economistas en E.E.U.U. hacen un análisis de su discurso dado en campaña; y dan cuenta de las mentiras que realizó a diestra y siniestra con tal de ganar votos.
EconomistLetter2016.
Human Noroviruses (HuNoVs) are important enteric pathogens, which affect the stomach and intestines, leading to
gastroenteritis or more commonly called the "stomach flu" or “winter vomiting bug". HuNoVs are mainly transmitted by the fecal-oral
route, either by directly infected person-to-person contact or directly via contaminated foods, water and surface areas. The virus is highly
contagious as 10-100 virus particles are sufficient to cause diseases. HuNoVs can spread easily and cause prolonged outbreaks. This is
due to their environmental persistence, high infectivity, being resistance to disinfection and difficulty in preventing transmission.
HuNoVs are the most common causative agent leading to acute gastroenteritis among infectious diseases worldwide and poses a serious
public health problem, especially among children being the most susceptible. In developing countries, the highest cost of medical care
after respiratory infections is listed for acute gastroenteritis. In this study, Norovirus outbreaks, precautions, its identification and
struggles were informed and some suggestions were made about this case.
FLI Seminar on different response strategies: Stamping out or NeutralizationHarm Kiezebrink
During this spring, American poultry producers are losing birds by the millions, due to the High Pathogenic Avian Influenza outbreaks on factory farms. USDA APHIS applied the stamping out strategy in an attempt to prevent the flu from spreading.
With stamping out as the highest priority of the response strategy, large numbers of responders are involved. With in average almost 1 million caged layers per farm in Iowa, there is hardly any room for a proper bio security training for these responders. And existing culling techniques had insufficient capacity, the authorities had to decide to apply drastic techniques like macerating live birds in order to take away the source of virus reproduction.
This strategy didn't work; on the contrary. Instead of slowing down the spreading of the virus, the outbreaks continue to reoccur and have caused death and destruction in 15 USA states, killing almost 50 million birds on mote than 220infected commercial poultry farms, all within a very small time frame.
The question is whether the priority of the response strategy should be on neutralizing the transmission routes instead of on stamping out infections after they occur. All indicators currently point out into the direction that the industry should prioritize on environmental drivers: the connection between outbreaks and wild ducks; wind-mediated transmission; pre-contact probability; on-farm bio security; transmission via rodents etc.
Once the contribution of each transmission route has been determined, a revolutionary new response strategy can be developed based on the principle of neutralizing transmission routes. Neutralizing risks means that fully new techniques need to be developed, based on culling the animals without human – to – animal contact; integrating detergent application into the culling operations; combining culling & disposal into one activity.
This new response strategy will be the main subject of the FLI Animal Welfare and Disease Control Seminar, organized at September 23, 2015 in Celle, Germany
Dynamic Aspects of Schistosoma Haematobium Infection as Experimental Model.pdfAlim A-H Yacoub Lovers
Abdul-Hussein H Awad, Alim A-H Yacoub, Sabeeh H Al-Mayah. Dynamic Aspects of Schistosoma Haematobium Infection as Experimental Model. Medical Journal of Basra University 1995;13(1&2):21-30
Frequency and Risk-Factors Analysis of Escherichia coli O157:H7 in Bali-CattleUniversitasGadjahMada
Cattle are known as the main reservoir of zoonotic agents verocytotoxin producing Escherichia coli. These bacteria are usually isolated from calves with diarrhea and / or mucus and blood. Tolerance of these agents to the environmental conditions will strengthen of their transmission among livestock. A total of 238 cattle fecal samples from four sub-districts in Badung, Bali were used in this study. Epidemiological data observed include cattle age, sex, cattle rearing system, the source of drinking water, weather, altitude, and type of cage floor, the cleanliness of cage floor, the slope of cage floor, and the level of cattle cleanliness. The study was initiated by culturing of samples onto eosin methylene blue agar, then Gram stained, and tested for indole, methyl-red, voges proskauer, and citrate, Potential E.coli isolates were then cultured onto sorbitol MacConkey agar, and further tested using O157 latex agglutination test and H7 antisera. Molecular identification was performed by analysis of the 16S rRNA gene, and epidemiological data was analyzed using
STATA 12.0 software. The results showed, the prevalence of E. coli O157:H7 in cattle at Badung regency was 6.30% (15/238) covering four sub districts i.e. Petang, Abiansemal, Mengwi, and Kuta which their prevalence was 8.62%(5/58), 10%(6/60), 3.33%(2/60), and 3.33(2/60)%, respectively. The analysis of 16S rRNA gene confirmed of isolates as an E. coli O157:H7 strain with 99% similarities. Furthermore, the risk factors analysis showed that the slope of the cage floor has a highly significant effect (P<0.05) to the distribution of infection. Consequently, implementing this factor must be concerned in order to decrease of infection.
Current Understanding of the Transmission, Diagnosis, and Treatment of H. pyl...AI Publications
H. pylori infection is a prevalent bacterial infection that affects the gastric mucosa of humans, with a prevalence ranging from 30% to 90%, depending on the region. The infection is a significant cause of gastritis, peptic ulcer disease, and gastric cancer. In this comprehensive review, we discuss the current understanding of the transmission, diagnosis, and treatment of H. pylori infection. We describe the risk factors and epidemiology of the infection, along with its pathogenesis, which involves multiple virulence factors that contribute to the colonization and survival of the bacteria in the acidic stomach environment. Diagnostic tests for H. pylori infection include invasive and non-invasive methods, with the choice of test depending on several factors. Treatment of H. pylori infection is aimed at eradicating the bacteria and preventing complications. Antibiotic-based triple or quadruple therapy, in combination with acid-suppressing agents, is the standard treatment, but antibiotic resistance is an emerging problem that needs to be addressed. This comprehensive review provides a useful resource for clinicians, researchers, and public health officials involved in managing H. pylori infection and its associated complications.
potassium, chloride, bicarbonate, blood urea nitrogen (BUN), magnesium, creatinine, glucose, and sometimes calcium. Tests that focus on cholesterol levels can determine LDL and HDL cholesterol levels, as well as triglyceride levels.[6]
PREVALENCE AND DEGREE OF INFECTION OF TOXOCARIASIS IN DAIRY CALVES (HOLSTEIN ...IAEME Publication
Background: Worm infection is one of the most common diseases affecting
livestock, one of those diseases is Toxocara vitulorum. Infection can cause diarrhea,
reduced productivity, intestinal and bile obstruction, to death in livestock. However,
this disease is often ignored by farmers. Observing from an economic perspective, this
disease results in very high losses for farmers.
Objective: This study aimed to determine the effect of dairy calves age on the
prevalence of toxocariasis and determine the effect of different degrees of toxocariasis
infection on dairy calves (Holstein Friesian).
Methods: This study used a descriptive laboratory method. The samples studied
were 120 stool or feces samples of calves aged 0-6 months. The number of samples in
each age group of 0-2 months, 2-4 months and 4-6 months were 40 samples. The
independent variable of this study was the age of dairy calves. The dependent variables
were the prevalence and degree of toxocariasis infection. Whereas, the control
variables were feed and cattle nation.
Results: From 120 samples studied, 5 feces samples were positively infected with
T. vitulorum. From 5 samples that were positively infected, 4 of them came from the
age group of 0-2 months and the other 1 from the age group of 2-4 months. Whereas,
in the 4-6 month age group all negative samples were from T. vitulorum infection. The
mean value of infection rates in the 0-2 month age group was 4.219, in the 2-4 month
age group was 1.066 and in the 4-6 month age group was 0.707.
Conclusion: The prevalence of toxocariasis and the degree of toxocariasis infection
based on the most influential age differences were at the age of 0-2 months.
Sub-clinical necrotic enteritis: its aetiology and predisposing factors in co...Chamari Palliyeguru
How does the multi-factorial disease sub-clinical necrotic enteritis induced in poultry fed without antibiotic growth promoters?
Clostridium perfringes a commensal bacterium on the large intestines, induces enteritis and necroses in upper intestines.
Many dietary and management stress factors affect the multiplication of bacteria in the upper intestines. Thus, causes a severe damage in the absorptive mucosae causing a significant loss in the growth performances.
1ScIeNtIFIc REPORTS | (2018) 8:1250 | DOI:10.1038/s41598-018-19638-x
www.nature.com/scientificreports
Histology, immunohistochemistry,
and in situ hybridization reveal
overlooked Ebola virus target
tissues in the Ebola virus disease
guinea pig model
Timothy K. Cooper1, Louis Huzella 1, Joshua C. Johnson 1, Oscar Rojas1, Sri Yellayi1,3,
Mei G. Sun2, Sina Bavari2, Amanda Bonilla1, Randy Hart1, Peter B. Jahrling 1, Jens H. Kuhn 1
& Xiankun Zeng 2
Survivors of Ebola virus infection may become subclinically infected, but whether animal models
recapitulate this complication is unclear. Using histology in combination with immunohistochemistry
and in situ hybridization in a retrospective review of a guinea pig confirmation-of-virulence study, we
demonstrate for the first time Ebola virus infection in hepatic oval cells, the endocardium and stroma of
the atrioventricular valves and chordae tendinae, satellite cells of peripheral ganglia, neurofibroblasts
and Schwann cells of peripheral nerves and ganglia, smooth muscle cells of the uterine myometrium
and vaginal wall, acini of the parotid salivary glands, thyroid follicular cells, adrenal medullary cells,
pancreatic islet cells, endometrial glandular and surface epithelium, and the epithelium of the vagina,
penis and, prepuce. These findings indicate that standard animal models for Ebola virus disease are not
as well-described as previously thought and may serve as a stepping stone for future identification of
potential sites of virus persistence.
Ebola virus disease (EVD) is a severe and frequently lethal affliction of humans caused by infection with any
of three members of the mononegavirus family Filoviridae: Bundibugyo virus (BDBV), Ebola virus (EBOV),
and Sudan virus (SUDV). A fourth virus, Taï Forest virus (TAFV), has thus far caused only a single reported
human infection, which was nonlethal1. EVD is an exotic disease with case numbers rarely surpassing the lower
hundreds1; however, from 2013–2016, EBOV caused an EVD outbreak in Western Africa encompassing 28,616
infections and 11,310 deaths in Guinea, Liberia, and Sierra Leone2. Long term sequelae in individual survivors of
acute EVD and the similar Marburg virus disease (MVD) and filovirus persistence followed by disease relapse or
sexual transmission had been reported before this outbreak3–8. However, observations during and following the
Western African EVD outbreak suggest that sequelae and filovirus persistence may be common events9. Reported
sequelae include arthralgia, cardiac valvulopathy, parotid gland inflammation, peripheral paresthesia or dyses-
thesia, and gastrointestinal motility disorders10–14. Semen may contain detectable EBOV RNA for more than 500
days following recovery, and EBOV RNA has been detected in breast milk of a subclinically infected mother15,16.
Replicating EBOV has been isolated from the cerebrospinal fluid of an EVD survivor suffering a disease relapse
and from the aqueous hu.
2. Background
The biology and ecology of synanthropic insects like flies
make them efficient carriers of disease-causing microorgan-
isms. Their breeding habits, mode of feeding and indiscrim-
inate traveling between decomposed waste and human
settings highly contribute to the dissemination of pathogens
in the environment. Approximately 350 fly species in 29
families are potentially associated with the transmission of
diseases of public health importance [1]. However, fewer
numbers of fly species have been associated with the trans-
mission of foodborne pathogens [1, 2]. Although there are
scarce reports of filth flies being the causative agent of
foodborne outbreaks, several studies have demonstrated a
steady decrease in the incidence of foodborne diarrhea after
suppressing fly populations [3–5], indirectly implicating
filth flies as the source of the foodborne pathogen.
The presence of flies in food and food facilities has always
been a concern of the U.S. Food and Drug Administration
(FDA). The FDA’s regulatory action criteria for filth in-
cludes a five-attribute profile that needs to be fulfilled be-
fore including a particular fly species as reasonably likely to
act as a contributing factor of the spread of foodborne
pathogens. These five attributes are synanthropy, endophily,
communicative behavior, attraction to filth and human
food, and the isolation of pathogens from wild populations
[1, 2]. Other fly species fulfilling at least four of those attri-
butes are considered opportunistic pests and their presence
in food and/or food-related environments is an indication
of insanitation [2].
Foodborne pathogens transmitted by synanthropic filth
flies are found not only externally on the fly surface
(which includes body, head, legs, and wings), but also in-
ternally, mainly in the alimentary canal (which runs the
length of the body, from pharynx to anus) [6]. In fact, we
previously reported that foodborne pathogens were up to
three times more likely to be found in the alimentary canal
than on the body surface of wild flies caught in and
around urban restaurant dumpsters [7]. Consequently,
flies can contaminate food or food-contact surfaces mech-
anically or through regurgitation or defecation. The poten-
tial spread of foodborne pathogens increases when there is
a focus of infection for a particular bacterium [8]. Our pre-
vious study showed a statistically significant association be-
tween the presence of Salmonella enterica, Listeria
monocytogenes and Cronobacter spp. (former Enterobacter
sakazakii) on the surface and in the guts of wild flies and
the sites where those flies were collected [7]; thus, empha-
sizing that bacteria inhabiting the alimentary canal of flies
are acquired from the surrounding environment. Filth flies
also travel quickly and may move several miles [9]; there-
fore, they can rapidly intensify the risk of foodborne dis-
eases by transporting pathogens from places where the
pathogens pose no hazard to places where they do, such as
food preparation areas [1].
The transmission process of a particular pathogen in
populations of synanthropic filth flies determines the spread
and persistence of that pathogen. Thus, information about
the transmission dynamics of a particular pathogen within
a fly population is essential to appropriately avoid the
spread of foodborne diseases. It is important to note that
understanding the epidemiology of an illness caused by a
pathogen transmitted by flies, requires a deeper knowledge
of the ecology, physiology, immunology, and genetics of the
pathogen as well as the morphology, physiology, and behav-
ior of the fly. Nevertheless, it is even more important to
understand how pathogen and fly interact in a particular
environment [10, 11].
Filth flies can be transient or definitive hosts of patho-
gens and, like vertebrates, they may be immune or sus-
ceptible to infection. Although flies can internally harbor
foodborne bacteria, it is not well known if these patho-
gens are beneficial or harmful to them. However, flies
have shown remarkable resilience to these pathogens.
For instance, several species of Cronobacter have been
isolated from the alimentary canal of several flies col-
lected in the wild [7, 12–16] and have also shown to
support the development of stable fly larvae in the ab-
sence of other microbes, by colonizing the alimentary
canal of newly emerged flies [16]. Flies have also shown
efficient and rapid responses to ingested Escherichia coli
O157:H7 since excretion of this pathogen was observed
6 to 24 h after being ingested [17, 18].
There are plenty of studies reporting the mechanical
transmission of foodborne pathogens by filth flies (some ex-
amples include [7, 19–22]) and there are other studies
reporting the fate and the temporospatial distribution of
ingested foodborne pathogens by flies [17, 18, 23–26].
However, there is little scientific information about the
transmission dynamics of foodborne bacteria to the fly’s
progeny after parental flies have ingested those pathogens.
The objective of this study was to estimate the probability
of transmission of four foodborne bacteria (S. enterica, C.
sakazakii, E. coli O157:H7, and L. monocytogenes ) to the
progeny of the common house fly, Musca domestica (Lin-
neaus) (Diptera: Muscidae), after parental house flies were
fed with food contaminated with low, medium, and high
levels of each bacterium. The presence of each pathogen
was evaluated on pooled house fly eggs laid by parental fe-
males and on the surface and in the alimentary canal of
newly emerged first filial (F1) generation adults.
Results and discussion
All parental house flies used in our experiments were ob-
served feeding from contaminated food and the presence of
each pathogen was confirmed from all alimentary canals
dissected from randomly selected parental females. Al-
though the focus of this study was not to evaluate the dy-
namics of the parental population of adult house flies,
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 2 of 12
3. anecdotal evidence suggests that the feeding and mating
behaviors were not influenced by the ingestion of bacteria,
and although not measured, we did not observe apparent
increases in the mortality of parental flies or reductions in
their oviposition rate, when compared to control groups.
We observed clusters of house fly eggs on the oviposition
substrate approximately 10-16 h after they were placed on
the mesh of all jars.
The combined molecular and culture approach that
we used to detect and isolate the foodborne pathogens
from samples of pooled house fly eggs, and the surfaces
and alimentary canals of single adult house flies was
straightforward when evaluating for the presence of S.
enterica, E. coli O157:H7, and L. monocytogenes. We eas-
ily obtained pure colonies of these three pathogens from
the enrichment media of all PCR-positive samples. Like-
wise, we easily obtained pure C. sakazakii colonies from
the enrichment media of all PCR-positives from pooled
house fly eggs and alimentary canals of parental flies.
However, the isolation of this pathogen from PCR-
positive samples from F1 adults was more challenging
and required several subculturing steps on selective
media. Consequently, we could only obtain pure col-
onies of C. sakazakii from eight out of 15 PCR-positive
samples from F1 adults.
We have previously reported that C. sakazakii colonies
could not be recovered from some PCR-positive samples
while using this combined approach, likely due to the PCR
being positive when other closely related bacterial genera,
such as Citrobacter freundii, are present in the samples
[7, 27]. Additionally, besides C. sakazakii a number of other
Enterobacteriaceae are α-glucosidase positive, therefore the
co-isolation of those organisms from samples with highly
complex microbiota (such as the fly’s alimentary canal)
could lower the efficiency of recovery of C. sakazakii from
the chromogenic media used [28]. As a result, only those
samples from which pure C. sakazakii colonies were iso-
lated, were considered positive for the presence of the
pathogen and included for statistical analysis. No pathogens
were observed on chromogenic media from any of the
PCR-negative samples that were randomly selected.
Pure colonies of S. enterica, L. monocytogenes, and E.
coli O157:H7 isolated from PCR-positive samples were
confirmed to be identical to the strains ingested by
parental house flies by showing indistinguishable PFGE
profiles (see Additional file 1). Likewise, matching nu-
cleotide sequences were obtained from pure colonies
of C. sakazakii when performing nucleotide compari-
son of the amplified fragment (463 bp) of the cgcA C.
sakazakii gene.
Probability of bacterial transmission to house fly eggs
Our study reports the probability of the presence of the
target pathogen in a sample containing pooled house fly
eggs laid by several females fed from contaminated food.
This study does not attempt to report the transmission
rate of individual eggs laid by one or several female flies.
The stepwise selection model of the logistic regression
analysis indicated that the predicted probability of the
presence of bacteria in samples of pooled house fly eggs
was associated with the type of foodborne pathogen and
the level of bacterial contamination of the food given to
parental house flies. However, there was not a significant
interaction between these two variables; thus, the inter-
action was removed from the full model described in Eq.
1. The model fit statistics and the AUC value of 0.89 (ex-
cellent discrimination) shows that our data fit the model
relatively well. Results from the analysis of the maximum
likelihood estimates of the parameters included in the
logistic regression model and the model fit statistics for
house fly eggs are included in Additional file 2(A).
For all bacteria evaluated, there was a higher chance of
the presence of the pathogen in samples with house fly
eggs after parental flies received food containing
medium levels of bacteria (Table 1A). In fact, when par-
ental house flies received food containing medium bac-
terial loads, the pathogens were two and six times more
likely to be present in the samples than when parental
flies fed from food contaminated with high and low bac-
terial levels, respectively. Therefore, there was not a
positive correlation between the levels of contaminated
food given to parental flies and the presence of the path-
ogens in samples with pooled house fly eggs.
The transmission potential of ingested bacteria to the
house fly progeny is a very complex process. Flies harbor
many microorganisms (including human pathogens) in
their alimentary canals and they require the ingestion of
live bacteria for their development. However, feeding
from contaminated food does not imply that flies will
become infected themselves or that ingested pathogens
will survive, proliferate, and/or invade the reproductive
system to be transovarially transmitted to house fly eggs
and to subsequent life stages or generations. House flies
can fight ingested opportunistic invaders by using phys-
ical barriers (i.e. the type II peritrophic matrix of the
midgut epithelium), physiological defenses (i.e. digestive
processes: pH, and digestive enzymes such as lysozyme),
and innate immune response (i.e. the secretion of anti-
microbial peptides, AMPs, by the fat body) [23, 29].
House flies also carry symbiotic bacteria from one
source to another and from one generation to the next
[30]. Other studies have suggested that the presence of
inherited symbiotic bacteria in insects increases the in-
sect’s resistance to pathogens; thus, inherited symbionts
may have important effects on the ecology and evolu-
tionary dynamics of host-pathogen interactions [31–33].
For instance, symbiotic Klebsiella oxytoca has been asso-
ciated with house fly eggs. This bacterium is deposited
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 3 of 12
4. on the surface of the eggs, inducing female oviposition.
However, when K. oxytoca is above the threshold abun-
dance levels, it causes oviposition inhibition [34]. None-
theless, the threshold levels of many other ingested
bacteria that will trigger a particular defense mechanism
(s) or particular behaviors in the house fly are not yet
well known. Nayduch and Joyner [35] detected lysozyme
protein in adult house flies that ingested 1.2×105
CFU/μl
of Staphylococcus aureus, and in their life history stages
(eggs, larval instars, and F1 adults), providing evidence
that the digestive and defensive dual role of lysozymes
was activated by the ingestion of high levels of these
bacteria. These facts could help to explain the lower
rates of contamination found in samples containing
pooled house fly eggs laid by females that ingested high
levels of contaminated food. However, more research is
needed to determine the role of specific foodborne bac-
teria in house flies and the threshold levels that will trig-
ger defense mechanisms or behaviors in these insects.
The highest rates of contamination of house fly eggs
were observed when parental flies fed from food con-
taminated with C. sakazakii (Fig. 1). Percentages of con-
tamination of 87, 98, and 96 % were observed after
parental house flies fed from food containing low,
medium, and high levels of C. sakazakii, respectively.
This was followed by the ingestion of food contaminated
with L. monocytogenes and E. coli O157:H7. The con-
tamination rate of house fly eggs with S. enterica was
lower than other pathogens: 30, 72, and 58 %, after par-
ental house flies received food containing low, medium,
and high levels of this pathogen, respectively (Fig. 1). Re-
gardless of the level of contamination of the food given
to parental house flies, C. sakazakii was 16, 6, and 3
times more likely to contaminate house fly eggs than S.
enterica, E. coli O157:H7, and L. monocytogenes, respect-
ively. Similarly, L. monocytogenes was 5 and 2 times
more probable to contaminate house fly eggs than S.
enterica and E. coli O157:H7, respectively (Table 1A).
Although the groups of collected house fly eggs were
surface-disinfected and we obtained no bacterial growth
from aliquots of water from the last rinse of the surface-
disinfection process, this only demonstrates that no
more bacteria could be dislodged from the surface of the
eggs (also known as chorion). To verify that bacterial
cells were not adsorbed onto the surface of house fly
Table 1 Odds ratios estimates of the presence of foodborne
pathogens
Foodborne
pathogen
Bacterial levels
in food
Fly’s body part Odds ratio
(95 % CL)
A) House fly eggs
Medium vs. high 1.9 (0.5, 6.8)
Medium vs. low 6.0 (1.7, 20.4)
High vs. low 3.2 (1.1, 9.6)
C. sakazakii vs. S. enterica 15.5 (2.9, 82.6)
C. sakazakii vs. E. coli O157:H7 5.7 (1.0, 31.3)
C. sakazakii vs. L. monocytogenes 3.0 (0.5, 16.4)
L. monocytogenes vs. S. enterica 5.2 (1.5, 18.7)
L. monocytogenes vs. E. coli O157:H7 1.9 (0.5, 7.2)
E. coli O157:H7 vs. S. enterica 2.7 (0.8, 8.6)
B) F1 female adults
S. enterica Medium vs. high 2.4 (1.7, 3.4)
Surface vs.
alimentary canal
2.4 (1.7, 3.4)
C. sakazakii High vs. medium 2.2 (1.3, 3.5)
Surface vs.
alimentary canal
2.4 (1.5, 3.8)
(A) house fly eggs and (B) first filial (F1) generation adults
Fig. 1 Probability of bacterial transmission to house fly eggs. Numbers in parenthesis represent lower and upper 95 % confidence limits (CL)
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 4 of 12
5. eggs, we randomly selected several surface-disinfected
eggs and used them either to take scanning electron mi-
croscopy (SEM) images or to plate them on the surface
of chromogenic media specific for each pathogen. Even
though SEM images of individual eggs did not reveal the
attachment of bacterial cells to the chorion (see
Additional file 3), we observed the presence of typical
bacterial colonies surrounding some of the surface-
disinfected house fly eggs that were individually plated.
Thus, indicating that some bacterial cells remained at-
tached to the chorion of surface-disinfected eggs.
Ingested bacteria could be adsorbed onto the surface
of house fly eggs during or after oviposition because in
female house flies the vaginal opening is in close prox-
imity to the anal opening [36], which may facilitate con-
tamination of the egg’s surface with waste products of
the fly’s digestive tract. Bacterial cells could remain at-
tached to the chorion due to the adhesive fluid that
covers the eggs when they are laid. This fluid is secreted
by the accessory glands of the female reproductive sys-
tem and causes the eggs to adhere to each other and to
the material where they were laid [37]. Additionally, the
chorionic sculpture of house fly eggs has minute hex-
agonal markings, distinct curved rib-like thickenings
(the hatching line), and some elevations and depressions
[36, 37] that could hinder the dislodgement of bacterial
cells during the surface-disinfection process.
We did not perform histological studies or transmis-
sion electron microscopy (TEM) to demonstrate the
presence and/or possible development of the target bac-
teria in the internal tissues of the eggs. Thus, in this
study we cannot confirm that the presence of pathogens
in samples containing pooled house fly eggs was due to
the transfer of bacteria at early stages of oogenesis and
embryogenesis, as required during true transovarial
transmission. Instead, the presence of pathogens in sam-
ples with pooled house fly eggs was probably due to the
adsorption of bacterial cells onto the surface of the eggs
during or after oviposition. Bacteria adsorbed on the sur-
face of the eggs can proliferate in the larval rearing sub-
strate and re-contaminate the hatching larvae, creating new
focus of infection from where the newly hatched larvae can
re-acquire the pathogen. In fact, random samples from lar-
val rearing substrates taken the same day that pupae were
removed from the rearing chambers evidenced the pres-
ence of the target pathogens (data not shown). Bacteria as-
sociated with house fly eggs have been found to
supplement the rearing substrate of the developing larvae
[38]. However, in this study we did not evaluate the pres-
ence of pathogens in any of the F1 larval stages. Future
studies in our lab will assess the temporospatial fate of
green fluorescent protein (GFP)-expressing S. enterica
and/or C. sakazakii from individual eggs laid by female
house flies fed with contaminated food. We will also
evaluate the presence of the pathogen on the surface and
internal tissues of the developing stages of the house fly
(three larval instars, puparia, and newly-emerged adults)
to have a better understanding of the trans-stadial trans-
mission of those pathogens during metamorphosis.
Probability of bacterial transmission to house fly F1 adults
House fly F1 adults were observed in all treatments, indi-
cating the successful completion of the house fly’s life
cycle. No pathogens were detected from the surface or
the alimentary canal of any of the adult specimens that
were sampled from the control groups. Even though L.
monocytogenes and E. coli O157:H7 were present in sam-
ples of pooled house fly eggs (Fig. 1), they were not de-
tected from either the surface or the alimentary canal of
any of the house fly F1 adults that were sampled. There-
fore, no statistics were computed for these two patho-
gens when included in the model, because all
observations had the same response.
We previously reported that L. monocytogenes was found
in 3 % of wild filth flies [7] and later confirmed that isolated
strains belonged to serotype 4b (unpublished data), respon-
sible for most major outbreaks of human listeriosis [39].
However, studies providing evidence of L. monocytogenes
being vectored by synanthropic filth flies are scarce. The in-
nate immune response elicited by L. monocytogenes infec-
tions has shown that this bacterium is rapidly detected by
the insect, inducing autophagy and inhibiting its intracellu-
lar growth to enhance insect survival [40, 41]. Additionally,
L. monocytogenes are not restricted to localized tissues or
specialized cells within the insect [42] and their release
from the alimentary canal during metamorphosis may in-
duce both localized and humoral insect immune responses
[43], decreasing the overall bacterial population [44]. Thus,
the absence of L. monocytogenes from house fly F1
adults was probably due to the flies’ innate immune re-
sponse towards this foodborne pathogen. However, the
ubiquitous abundance of L. monocytogenes in the en-
vironment, their ability to survive for long periods of
time in acidic soils containing high endogenous micro-
biota [45], and their capability to attach to environ-
mental surfaces and form biofilms [46] gives them the
ability to create new focus of infection that can be used
by filth flies to widely spread this pathogen.
Escherichia coli O157:H7 was also absent from house
fly F1 generation adults. While some studies have dem-
onstrated that house flies that ingested high E. coli
O157:H7 concentrations (109
CFU/ml), retained this
pathogen inside the alimentary canal for up to three days
[18, 47], some others have reported that immune mo-
lecular effectors such as AMPs and lysozymes prevent
the proliferation of this pathogen in the fly’s alimentary
canal [17]. Thus, the question that E. coli O157:H7 is
pathogenic to house flies needs to be further
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 5 of 12
6. investigated. Although E. coli O157:H7 was present in
samples containing pooled house fly eggs, this pathogen
did not persist throughout metamorphosis. This finding
was opposite to other studies that have reported the in-
gestion of non-pathogenic E. coli by house fly larva and
their persistence throughout pupae and newly emerged
adults [48, 49]. However, in this study we did not quantify
the amount of E. coli O157:H7 present in the larval rearing
substrate; hence, the levels of this pathogen that were likely
to be ingested by house fly larvae were unknown and prob-
ably low enough to avoid their persistence through the
house fly life cycle. Nevertheless, the association of synan-
thropic filth flies with E. coli O157:H7 is broadly supported
[21, 50–54], strongly suggesting that house flies can indis-
criminately disseminate this foodborne pathogen.
Salmonella enterica and C. sakazakii were the only
pathogens present on F1 generation adults and only
when parental house flies were given food contaminated
with medium and high bacterial loads. The analysis of the
maximum likelihood estimates of the parameters of this lo-
gistic regression model and the model fit statistics are in-
cluded in Additional file 2(B). As shown by the model fit
statistics and AUC values of 0.87 and 0.82 (excellent dis-
crimination) for S. enterica and C. sakazakii, respectively,
our data fit the model in Eq. 2 relatively well. The estimated
probability of transmitting these pathogens to any single fe-
male adult fly from the F1 generation was associated with
the bacterial concentration given to parental flies and the
body part of the fly.
The presence of S. enterica and C. sakazakii was 2.4
times more likely on the body surface than in the ali-
mentary canal of newly emerged F1 adults (Table 1B).
This is in agreement with early studies performed by
Radvan [55] who determined that some bacteria includ-
ing Bacillus anthracis, B. subtilis, Shigella sonnei, and
non-pathogenic E. coli were mainly located on the sur-
face of recently emerged flies. This is probably due to
the release of the intestinal content of the larvae into the
pupal cavity, one of the changes that take place while
the larvae re-organizes into an adult house fly [43, 56].
The probability of finding S. enterica on a single F1
adult house fly was greater than the probability of find-
ing C. sakazakii (Fig. 2a, b). When parental flies received
food with medium levels of S. enterica the probability of
60 (52, 67)
38 (32, 45)38 (31, 45)
20 (16, 26)
0
10
20
30
40
50
60
70
Surface
15 (11, 22)
28 (22, 35)
7 (4, 11)
14 (10, 19)
0
10
20
30
40
50
60
70
Alimentary canal
Medium High
b
Fig. 2 Probability of bacterial transmission to house fly first filial (F1) generation adults. a Salmonella enterica and (b) Cronobacter sakazakii.
Numbers in parenthesis represent lower and upper 95 % confidence limits (CL)
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 6 of 12
7. finding this pathogen on the surface and in the alimen-
tary canal of a single F1 adult fly was 60 and 38 %, re-
spectively. However, this probability decreased 38 and
20 % for the body surface and the alimentary canal, re-
spectively, when parental flies fed from food with high S.
enterica levels (Fig. 2a). Overall, it was 2.4 times more
likely to find S. enterica on F1 adults after parental flies
fed from food containing medium bacterial loads
(Table 1B). Even though the presence of S. enterica in
samples containing pooled house fly eggs was lower than
other bacteria evaluated, this pathogen has developed
strategies to deal with environmental changes brought
on by the whole microbial community of a specific niche
[57]. This could allow Salmonella to colonize the larval
rearing substrate, be re-acquired by the developing lar-
vae and persist through the adult stage.
Contrary to our findings with S. enterica, it was 2.2
times more likely to find C. sakazakii in a single F1 adult
house fly after parental flies fed from food contaminated
with high levels of this pathogen (Table 1B). The prob-
abilities of finding C. sakazakii on the fly’s body surface
and in the alimentary canal were 28 and 14 %, respect-
ively, after parental flies fed from highly contaminated
food (Fig. 2b). This probability decreased 15 and 7 % for
the body surface and the alimentary canal, respectively,
when parental flies fed from food contaminated with
medium C. sakazakii levels. Thus, our results emphasize
that pathogen concentration is an important parameter
to determine the transmission of bacteria to the house
fly progeny. Other authors have also stressed the signifi-
cance of bacterial concentrations in the transmission of
microorganisms since low bacterial inocula are insuffi-
cient to colonize the insect and the ingestion of exces-
sive bacteria may be either pathogenic [48, 58, 59] or
alter population dynamics or behavior [60].
Cronobacter sakazakii and S. enterica have probably
evolved several mechanisms to evade the fly’s immune
system. Bacteria that are associated with food can access
the fly’s digestive tract and if they tolerate digestive pro-
cesses and evade the immune system, they are able to
access an environment that allows them to disseminate
via regurgitation or defecation [32, 61]. Some ingested
pathogenic bacteria can also produce a chronic infection
in the host that makes it difficult to distinguish between
a pathogenic or beneficial insect-microbe association
[32, 62]. If C. sakazakii and S. enterica provide some
benefit to synanthropic filth flies needs to be studied fur-
ther. Examples of beneficial facultative symbionts by sev-
eral arthropods include Serratia symbiotica in the pea
aphid, Acyrthosiphon pisum (Hemiptera: Aphididae),
which confers resistance against natural enemies such as
parasitic wasps [63–65], and Hamiltonella defensa in
whiteflies Bemisia tabaci (Hemiptera: Aleyrodidae) that
increases the development and fitness of the host [66].
Consequently, the transmission mechanisms of both C.
sakazakii and S. enterica need to be studied through
more than one generation of flies to elucidate the type
of associations these bacteria can potentially establish
with these insects. Additionally, the interactions of these
foodborne pathogens with other microorganisms present
in flies need to be further explored. Understanding the
type of associations that synanthropic flies establish with
foodborne pathogens will help to elucidate transmission
mechanisms as well as possible ways to mitigate the
spread of foodborne pathogens.
It is important to mention that there is zero tolerance
for the presence of S. enterica, C. sakazakii, E. coli
O157:H7, or L. monocytogenes in foods. The mere pres-
ence of any of these four foodborne pathogens deems
the food to be adulterated. Because the concentration of
these pathogens in foods is usually not quantified, it is
difficult to associate the three levels of contaminated
food given to flies, to contamination concentrations of
these pathogens in foods. Interestingly, this study dem-
onstrated that adult house flies feeding from food con-
taminated with levels of bacteria as low as 100 cells/ml
are able to transfer ingested pathogens to their progeny.
Even though food can become contaminated at any
point during production, the presence of pests, such as
flies, increases the potential risk of pathogen transmis-
sion. Synanthropic filth flies that feed from any level of
contaminated food are able to disseminate pathogens in-
discriminately, not only mechanically or through regur-
gitation and defecation but also to their progeny, greatly
increasing their vector potential.
To better protect public health, it is important to high-
light the need for effective preventative measures that
minimize the hazard posed by pests that may come in con-
tact with food or food-contact surfaces and utensils. The
implementation of pest control programs is one of the
frequently and highly recommended measures to avoid
the indirect transmission of foodborne pathogens by
synanthropic insects like flies. The effectiveness of the
program should be constantly monitored and filthy
breading sites should be eliminated. By targeting con-
trol measures towards synanthropic filth flies, the po-
tential transmission of foodborne pathogens can be
interrupted, contributing to the prevention of future
foodborne illness outbreaks.
Conclusion
In this study, we demonstrated that adult house flies that
fed from food contaminated with low, medium, and high
levels of S. enterica, C. sakazakii, E. coli O157:H7 or L.
monocytogenes transmit these pathogens to their eggs.
Salmonella enterica and C. sakazakii were further trans-
mitted to F1 generation house fly adults, and they were
more commonly found on the surface than in the
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 7 of 12
8. alimentary canal of newly emerged house flies. Results
from this research emphasize the public health signifi-
cance and the regulatory importance of the presence of
flies in food and food facilities.
Methods
House fly source
House fly (M. domestica) puparia were obtained from
Spider Pharm, Inc. (Yarnell, AZ) and placed in plastic
cages inside a Percival growth chamber at 30 °C and
16:8 h light:dark (L:D) photoperiod until eclosion.
Emerged house flies were fed with a dry mixture of 1:1
granulated sugar and powdered milk. Cotton balls
soaked in autoclaved water were also provided as a water
source. Adult house flies (2-4 days old) were immobi-
lized by placing the plastic cages at -30 °C for 5-7 min.
Groups of approximately 40 adults (mixed sex) were
transferred to autoclaved wide-mouth quart Mason glass
jars. A disinfected 6-inch square piece of fiberglass win-
dow screen (New York Wire, Hanover, PA) was placed
on top of each jar and secured with a rubber band. All
glass jars were kept in the Percival growth chamber
under the same conditions described above.
Preparation of contaminated food
Four bacterial foodborne pathogens (S. enterica, C.
sakazakii, E. coli O157:H7, and L. monocytogenes) were
used in our study. Information about bacterial strains,
serotypes, and their origin is specified in Additional
file 4. Bacterial strains were reconstituted from 30 %
glycerol stock cultures, plated on Trypticase Soy
Agar (TSA; Oxoid, Cambridge, UK), and incubated
at 37 °C overnight. Stock suspensions of each bacter-
ium were prepared by scraping bacterial cells from
overnight cultures and adding them to buffered pep-
tone water (BPW; Difco, Becton, Dickinson and
Company, Sparks, MD). The optical density of the
stock suspension was measured at 600 nm (OD600)
using a GENESYS™ 20 Spectrophotometer (Thermo
Fisher Scientific, Rochester, NY), and the bacterial
concentration was calculated assuming that 0.1
OD600 = 108
bacterial cells/ml [67, 68]. A known vol-
ume of the stock bacterial suspension was added to
a known volume of liquid fly food (18 g of dried
powdered milk, 4 g of sugar, 2 g of protein powder,
and 200 ml sterile distilled water) to obtain final
bacterial concentrations of 108
, 104
, and 102
CFU/ml
of each foodborne pathogen.
Adult house fly feeding bioassay
For each pathogen, approximately ten ml of fly food with
the corresponding level of bacteria was added to three
autoclaved cotton balls that were previously placed in
the base of a sterile 60 mm diameter Petri dish. Fly food
with no bacteria was used to feed the control groups. Fly
food was given to parental house flies by inverting the
Petri dish onto the mesh screen on top of each glass jar
(see Additional file 5(A)), replacing with the correspond-
ing fresh food after 18-20 h. Jars were kept in the Perci-
val growth chamber under the same conditions
described before and adult house flies were allowed to
mate and feed ad libitum for a total of 30-32 h. Al-
though the level of bacterial contamination of the fly
food provided to parental house flies was known, the
amount of bacteria actually ingested by adult house flies
was not quantified. Thus, fly food containing final bac-
terial concentrations of 108
, 104
, and 102
CFU/ml will be
referred hereinafter as high, medium, and low, respect-
ively. After completing the feeding time, the Petri dish
and cotton balls were removed and the mesh screen was
thoroughly cleaned and disinfected with 70 % ethanol
before adding the oviposition substrate.
Collection of house fly eggs
To create an oviposition substrate, several pieces of
dehydrated beef liver (approximately 1 cubic inch and
hydrated overnight) were placed on top of the mesh
screen of each glass jar and covered with the lid of a
sterile Petri dish to prevent dehydration (see Additional
file 5(B)). Once fly eggs were visible on the surface of
the liver, the glass jars were removed from the Percival
growth chamber and clusters of approximately 100 eggs
(laid by several females) were carefully removed using
autoclaved forceps. To remove microbiota from the outer
surface of the eggs, each cluster of eggs was transferred to a
two ml tube with 70 % ethanol for 1 min, then submersed
in 0.05 % bleach for 1 min, and finally rinsed three times
with autoclaved distilled water (see Additional file 5(C)).
One-hundred μl aliquots of water from the last rinse were
plated on chromogenic media specific for the target food-
borne pathogen (see Additional file 4). Surface-disinfected
house fly eggs were divided in two groups approximately
equal in number (~40-50). To assess the presence of patho-
gens, the first group of pooled eggs was added to one ml of
enrichment media specific for each bacterial pathogen and
incubated accordingly (see Additional file 4). The second
group of eggs was added to a larval rearing substrate and
allowed to hatch and complete their life cycle to evaluate
the presence of foodborne pathogens in adult house flies of
the F1 generation.
Validation that parental house fly adults ingested
bacteria
After eggs were collected, glass jars containing parental
house flies were placed at -20 °C for 5-7 min until flies
were immobilized. Immobilized flies were then trans-
ferred to a disposable Petri dish containing 70 % alcohol
for 2 min. Using a dissecting scope, three adult female
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 8 of 12
9. house flies were randomly sub-sampled per each glass
jar (n = 48 per each foodborne bacterium) and individu-
ally transferred to an autoclaved two ml tube to be
surface-disinfected and their alimentary canals dissected
as described by Pava-Ripoll, et al. [27]. The alimentary
canals of maternal house flies were individually evalu-
ated for the presence of the target bacteria as described
in sections below.
House fly F1 offspring rearing procedure
The larval rearing substrate was prepared by pre-mixing
dry ingredients (1 cup of autoclaved alfalfa pellets, 1 cup
of autoclaved wheat bran, 1 cup of autoclaved bone
meal, 1 cup of autoclaved poultry litter, 1/3 cup of dried
milk powder, and 1 teaspoon of Brewer’s yeast) and add-
ing 4 ½ cups of autoclaved tap water. Half cup of the
prepared larval rearing substrate was added to individual
plastic containers and then the group of surface-
disinfected house fly eggs was added to the substrate
using a disposable plastic pipette. The container was
then nested in a larger plastic container that was ap-
proximately 1/8th
filled with autoclaved sand to give fly
larvae a dry place to pupate (see Additional file 5(D)).
The rearing chambers were covered with an autoclaved
paper towel, secured with a rubber band and placed in a
Percival growth chamber at 32-35 °C and 16:8 h L:D
photoperiod until pupation (approximately 4-5 days; see
Additional file 5(E)). Using a disposable 1000 μl sterile
pipette tip the larval substrate was gently mixed every
day to inhibit mold growth. House fly pupae from each
rearing chamber were carefully separated from the sand
using sterile forceps and transferred to an extra-deep
sterile, disposable Petri dish (Fisherbrand, Thermo
Fisher Scientific, Rochester, NY) to allow F1 adults to
emerge avoiding cross-contamination with the larval
rearing substrate (see Additional file 5(F)). Petri dishes
containing pupae were kept in the Percival growth
chamber under same conditions until emergence of F1
generation adults (approximately 2-3 additional days;
see Additional file 5(G)).
Collection of female F1 generation house fly adults
Recently emerged (0-1 days old) F1 adults were immobi-
lized by placing extra-deep Petri dishes at -20 °C for 5-7
min. Under a dissecting scope, three females were ran-
domly sub-sampled per each Petri dish (n = 48 per each
foodborne bacterium) and individually transferred to auto-
claved two ml tubes containing one ml of enrichment
media specific for the target pathogen (see Additional
file 4) to collect microbiota from the surface of the
newly emerged house fly. Each house fly was then
removed from the enrichment media, surface-
disinfected and their alimentary canals aseptically
dissected as described by Pava-Ripoll, et al. [27]. Tubes
with enrichment media containing microbiota from the
surface (s) and the alimentary canal (ac) of each F1
adult house fly were incubated at times and tempera-
tures recommended for each bacterial pathogen (see
Additional file 4).
Detection and isolation of the target bacteria
Enriched samples were assessed for the presence/ab-
sence of the target bacteria using a combined molecular
and culture approach. The molecular approach was per-
formed using a commercial PCR cycler/detector system
(BAX® System Q7, DuPont Qualicon, Wilmington, DE)
and assay kits specific for each bacteria (see Additional
file 4) following manufacturer’s instructions and as de-
scribed by Pava-Ripoll, et al. [27]. Each assay kit con-
tains PCR-ready tablets with an intercalating dye that
emits a fluorescence signal when binding to the target
double-stranded DNA. The signal is detected by the
PCR system and interpreted by the software as positive
or negative. The culture approach was performed by
plating ten μl of the enrichment media of PCR-positive
samples on chromogenic media specific for each bacter-
ium (see Additional file 4) until pure colonies were ob-
tained. The culture approach was performed to confirm
that isolated pathogens were the same strains given to par-
ental house flies. Isolated S. enterica, L. monocytogenes,
and E. coli O157:H7 were confirmed through pulsed-field
gel electrophoresis (PFGE), following the protocols de-
scribed by PulseNet and only using primary enzyme re-
striction [69, 70]. Isolated C. sakazakii was confirmed
by polymerase chain reaction (PCR) amplification of
the diguanylate cyclase (cgcA) gene using primers
Cmstu-825 F and Csak-1317R as described by Carter,
et al. [71]. Amplicons of expected size (463 bp) of the
singleton PCR reaction were purified and sequenced by
Retrogen, Inc. (San Diego, CA) and sequence files were
imported into Sequencher 5.0 (GeneCodes, Ann Arbor,
MI) to be processed and assembled. Contigs were
exported and aligned using the CLUSTALX software
(Lasergene, Madison, WI) and aligned sequences were
used to generate a variance table report (Sequencher 5.0)
where nucleotide bases of each sequence were compared
to the reference C. sakazakii sequence. Four to five ran-
domly selected PCR-negative samples were also plated on
specific chromogenic media to confirm the absence of the
target pathogen.
Experimental design
This experiment was set up as a completely randomized
design and was performed at four different times with a
one-month interlude. One foodborne pathogen (S.
enterica, C. sakazakii, E. coli O157:H7, or L. monocyto-
genes) and fly food with three levels of bacterial contam-
ination (high, medium, and low) plus a control,
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 9 of 12
10. consisting of fly food with no bacteria, were evaluated
each time. Each treatment combination (foodborne
pathogen by levels of contaminated food) was replicated
four times. Thus, 16 glass jars containing parental gen-
eration of adult flies were prepared each time the ex-
periment was run. The presence/absence of the target
bacterium was assessed as follows: a) from the ali-
mentary canals of three parental females that were
randomly sub-sampled per replicate (n = 48 per each
foodborne pathogen); b) from pooled house fly eggs
laid by several parental females (n = 16 per each food-
borne pathogen); and c) from body surfaces and ali-
mentary canals of three F1 female house flies that were
randomly sub-sampled per replicate (n surface = 48 and
n alimentary canal = 48 per each foodborne pathogen).
Statistical analysis
We used the SAS logistic regression procedure (PROC
LOGIT; SAS Institute Inc., 2005) to predict the prob-
ability of bacterial contamination to house fly eggs and
to the surface and the alimentary canal of F1 female
adults. The presence/absence of foodborne pathogens
was the categorical dichotomous response variable and
its relationship with the predictor variables was ana-
lyzed using the two full logistic probability models
described in Eq. 1 (for house fly eggs) and Eq. 2 (for F1
generation house fly adults).
Logit Pð Þeggs ¼ β0 þ β1 Ã foodborne pathogen
þ β2 Ã bacterial levels of contaminated food
þ β3 Ã foodborne pathogen Ã
bacterial levels of contaminated food
ð1Þ
Logit Pð ÞF1 ¼ β0 þ β1
à bacterial levels of contaminated food
þ β2 Ã house fly’s body part
ð2Þ
Where logit (P) = ln [P/1-P], ln is the natural log, P is
the probability of the presence of bacteria, β0 is the P
intercept, βi are regression coefficients. The predictor
variables for the probabilistic model of house fly eggs
(Eq. 1) were the type foodborne pathogen, the level of
bacterial contamination of the food given to parental
house flies and their interaction. The predictor variables
for the probabilistic model of F1 generation of house fly
adults were the level of bacterial contamination of the
food given to parental house flies and the fly’s body part
(surface and alimentary canal) and the model was ana-
lyzed by each foodborne pathogen. The stepwise selec-
tion method with analysis of maximum likelihood
estimates based on a Wald Chi-square p value <0.05 was
used to determine the best probability model. The re-
ceiver operating characteristics (ROC) curve was used as a
measurement of the goodness-of-fit of the model. The
ROC curve quantifies the power of the predicted values
using the area under the ROC curve (AUC). AUC values
>0.7 are considered acceptable discrimination, >0.8 are
considered excellent discrimination and >0.9 are consid-
ered outstanding discrimination [72].
Additional files
Additional file 1: Pulsed-field gel electrophoresis (PFGE) profiles.
The PFGE fingerprinting shows an indistinguishable pattern between the
bacterial strains used to feed parental flies and the bacterial colonies
isolated from the alimentary canal of parental flies (Pac), house fly eggs
(e), and the surface (s) and alimentary canal (ac) of adult flies from the
first filial (F1) generation. Profiles were obtained from (A) Salmonella
enterica serotype Schwarzengrund (strain SAL3542; PFGE PulseNet pattern
JM6X01.0289); (B) enterohemorrhagic Escherichia coli O157:H7 (strain
ESC0786; PFGE PulseNet pattern EXHX01.0125); and (C) Listeria
monocytogenes serotype 4b (strain LIS0150; PFGE PulseNet combined
pattern GX6A16.0059_GX6A12.1652).
Additional file 2: Analysis of Maximum Likelihood Estimates (MLE)
of the logistic regression model. (A) house fly eggs and (B) house fly
first filial (F1) generation of adults.
Additional file 3: Scanning Electron Microscopy (SEM) of
surface-disinfected house fly (Musca domestica) eggs (A) house fly
egg; (B) the hatching line, with distinct curved rib-like thickenings;
and (C) adhesive fluid on the egg surface.
Additional file 4: Information about foodborne bacteria, culture
media, incubation conditions, and PCR-based kits used in this
study.
Additional file 5: Experimental setup. (A) feeding of the parental
population of house flies, (B) oviposition substrate, (C) collected house fly
eggs, (D) surface-disinfected eggs placed in the larval rearing substrate,
(E) house fly larval rearing container, (F) transfer of house fly pupae to
plates, (G) emergence of first filial (F1) generation of house fly adults.
Competing interests
The authors certify that there is no competing interest with any financial
organization regarding the materials discussed in this manuscript. The use of
specified instrumentation is not an endorsement by the U.S. Food and Drug
Administration.
Authors’ contributions
MPR and REGP conceived and designed the experiments. MPR, REGP, and
AKM carried out laboratory work including feeding bioassays, rearing
procedures, and detected and isolated bacteria from individual flies. MPR
performed PCR analysis of C. sakazakii. BDT performed Scanning Electron
Microscopy (SEM). CEK performed pulsed-field gel electrophoresis (PFGE)
profiles of S. enterica, L. monocytogenes, and E. coli O157:H7. BDT and GCZ
contributed by giving their point of view to the discussion of the results.
MPR performed statistical analysis and wrote the manuscript. All authors read
and approved the final manuscript.
Financial disclosure
The authors received no specific funding for this work.
Author details
1
U.S. Food and Drug Administration, Center for Food Safety and Applied
Nutrition, Office of Food Safety, 5100 Paint Branch Pkwy, College Park, MD
20740, USA. 2
U.S. Food and Drug Administration, Center for Food Safety and
Applied Nutrition, Office of Applied Research and Safety Assessment, 8301
Muirkirk Rd, Laurel, MD 20708, USA. 3
U.S. Food and Drug Administration,
Center for Food Safety and Applied Nutrition, Office of Regulatory Science,
5100 Paint Branch Pkwy, College Park, MD 20740, USA.
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 10 of 12
11. Received: 27 February 2015 Accepted: 6 July 2015
References
1. Olsen AR. Regulatory action criteria for filth and other extraneous materials
III. Review of flies and foodborne enteric disease. Regul Toxicol Pharmacol.
1998;28(3):199–211.
2. Olsen AR, Gecan JS, Ziobro GC, Bryce JR. Regulatory action criteria for filth
and other extraneous materials V. Strategy for evaluating hazardous and
nonhazardous filth. Regul Toxicol Pharmacol. 2001;33(3):363–92.
3. Emerson PM, Lindsay SW, Walraven GEL, Faal H, Bogh C, Lowe K, et al. Effect
of fly control on trachoma and diarrhoea. Lancet. 1999;353(9162):1401–3.
4. Esrey SA, Potash JB, Roberts L, Shiff C. Effects of improved water-supply and
sanitation on ascariasis, diarrhea, dracunculiasis, hookworm infection,
schistosomiasis, and trachoma. Bull WHO. 1991;69(5):609–21.
5. Cohen D, Green M, Block C, Slepon R, Ambar R, Wasserman SS, et al.
Reduction of transmission of shigellosis by control of house flies (Musca
domestica). Lancet. 1991;337(8748):993–7.
6. Steinhaus EA. Insect microbiology. vol. November. New York: Hafner
Publishing Co Ltd.; 1967.
7. Pava-Ripoll M, Pearson REG, Miller AK, Ziobro GC. Prevalence and relative
risk of Cronobacter spp., Salmonella spp., and Listeria monocytogenes
associated with the body surfaces and guts of individual filth flies. Appl
Environ Microbiol. 2012;78(22):7891–902.
8. Greenberg B, Bornstein AA. Fly dispersion from rural Mexican
slaughterhouse. Am J Trop Med Hyg. 1964;13(6):881.
9. Savage EP. Disease Vector. In: Purdom PW, editor. Environ Health. London,
U.K: Academic Pres; 1971. p. 48.
10. Gorham JR. Food, filth, and disease: A review. In: Foodborne disease
handbook: Diseases caused by hazardous substances. Edited by Hui YH,
Gorham JR, Murrell KD, Cliver DO, vol. 3: New York, USA: Marcel Dekker, Inc.;
1994: 627–634.
11. Reisen WK. Landscape epidemiology of vector-borne diseases. Annu Rev
Entomol. 2010;55:461–83.
12. Butler JF, Garcia-Maruniak A, Meek F, Maruniak JE. Wild Florida house flies
(Musca domestica) as carriers of pathogenic bacteria. Fla Entomol. 2010;93(2):218–23.
13. Gupta AK, Nayduch D, Verma P, Shah B, Ghate HV, Patole MS, et al.
Phylogenetic characterization of bacteria in the gut of house flies (Musca
domestica L.). FEMS Microbiol Ecol. 2012;79(3):581–93.
14. Hamilton JV, Lehane MJ, Braig HR. Isolation of Enterobacter sakazakii from
midgut of Stomoxys calcitrans. Emerging Infect Dis. 2003;9(10):1355–6.
15. Mramba F, Broce A, Zurek L. Isolation of Enterobacter sakazakii from stable flies,
Stomoxys calcitrans L. (Diptera:Muscidae). J Food Prot. 2006;69(3):671–3.
16. Mramba F, Broce AB, Zurek L. Vector competence of stable flies, Stomoxys
calcitrans L. (Diptera:Muscidae), for Enterobacter sakazakii. J Vector Ecol.
2007;32(1):134–9.
17. Fleming A. Spatial and temporal immune response in house flies in
response to ingestion of Bacillus cereus and Eschericha coli O57:H7.
Electronic Theses & Dissertations: Georgia Southern University; 2012.
18. Kobayashi M, Sasaki T, Saito N, Tamura K, Suzuki K, Watanabe H, et al.
Houseflies: Not simple mechanical vectors of enterohemorrhagic Escherichia
coli O157:H7. Am J Trop Med Hyg. 1999;61(4):625–9.
19. Bidawid SP, Edeson JFB, Ibrahim J, Matossian RM. Role of non-biting flies
in transmission of enteric pathogens (Salmonella species and Shigella
species) in Beirut, Lebanon. Ann Trop Med Parasitol. 1978;72(2):117–21.
20. Mian LS, Maag H, Tacal JV. Isolation of Salmonella from muscoid flies at
commercial animal establishments in San Bernardino county, California. J
Vector Ecol. 2002;27(1):82–5.
21. Moriya K, Fujibayashi T, Yoshihara T, Matsuda A, Sumi N, Umezaki N, et al.
Verotoxin-producing Escherichia coli O157:H7 carried by the housefly in
Japan. Med Vet Entomol. 1999;13(2):214–6.
22. Olsen AR, Hammack TS. Isolation of Salmonella spp. from the housefly,
Musca domestica L., and the dump fly, Hydrotaea aenescens (Wiedemann)
(Diptera:Muscidae), at caged-layer houses. J Food Prot. 2000;63(7):958–60.
23. Fleming A, Kumar HV, Joyner C, Reynolds A, Nayduch D. Temporospatial fate of
bacteria and immune effector expression in house flies fed GFP-Escherichia coli
O157:H7. Med Vet Entomol. 2014;28(4):364–71.
24. Olafson PU, Lohmeyer KH, Edrington TS, Loneragan GH. Survival
and Fate of Salmonella enterica serovar Montevideo in Adult Horn Flies
(Diptera: Muscidae). J Med Entomol. 2014;51(5):993–1001.
25. Joyner C, Mills MK, Nayduch D. Pseudomonas aeruginosa in Musca domestica L.:
Temporospatial examination of bacteria population dynamics and house fly
antimicrobial responses. Plos One. 2013, 8(11):e79224.
26. Nayduch D, Cho H, Joyner C. Staphylococcus aureus in the House Fly:
temporospatial fate of bacteria and expression of the antimicrobial peptide
defensin. J Med Entomol. 2013;50(1):171–8.
27. Pava-Ripoll M, Pearson REG, Miller AK, Ziobro GC. Detection of foodborne
bacterial pathogens from individual filth flies. J Vis Exp. 2015;96:e52372.
28. Iversen C, Forsythe SJ. Comparison of media for the isolation of Enterobacter
sakazakii. Appl Environ Microbiol. 2007;73(1):48–52.
29. Brandt SM, Dionne MS, Khush RS, Pham LN, Vigdal TJ, Schneider DS.
Secreted bacterial effectors and host-produced eiger/TNF drive death in a
Salmonella-infected fruit fly. PLoS Biol. 2004;2(12):2067–75.
30. Kellner R. The role of microorganisms for eggs and progeny. In:
Chemoecology of Insect Eggs and Egg Deposition. Edited by Hilker M,
Meiners T: Oxford, UK: Blackwell Publishing; 2002: 149–164.
31. Brownlie JC, Johnson KN. Symbiont-mediated protection in insect hosts.
Trends Microbiol. 2009;17(8):348–54.
32. Eleftherianos I, Atri J, Accetta J, Castillo JC. Endosymbiotic bacteria in insects:
Guardians of the immune system? Front Physiol. 2013, 4(46):1–10.
33. Hurst GDD, Darby AC. The inherited microbiota of arthropods, and their
importance in understanding resistance and immunity. In: Insect Infection and
Immunity. Evolution, Ecology, and Mechanisms. Edited by Rolff J, Reynolds SE:
Oxford, UK: Oxford University Press 2009: 119-135.
34. Lam K, Babor D, Duthie B, Babor EM, Moore M, Gries G. Proliferating
bacterial symbionts on house fly eggs affect oviposition behaviour of adult
flies. Anim Behav. 2007;74:81–92.
35. Nayduch D, Joyner C. Expression of lysozyme in the life history of the house
Fly (Musca domestica L.). J Med Entomol. 2013;50(4):847–52.
36. Leopold RA, Meola S, Degrugillier ME. The egg fertilization site within the
house fly, Musca domestica (L.) (Diptera: Muscidae). Int J Insect Morphol
Embryol. 1978;7(2):111–20.
37. Hewitt CG. The housefly, its structure, habits, development, relation to
disease and control: Cambridge, UK: Cambridge University Press; 1914.
38. Lam K, Geisreiter C, Gries G. Ovipositing female house flies provision
offspring larvae with bacterial food. Entomol Exp Appl. 2009;133(3):292–5.
39. Pan YW, Breidt F, Kathariou S. Competition of Listeria monocytogenes
Serotype 1/2a and 4b strains in mixed-culture biofilms. Appl Environ
Microbiol. 2009;75(18):5846–52.
40. Tindwa H, Patnaik BB, Kim DH, Mun S, Jo YH, Lee BL, et al. Cloning,
characterization and effect of TmPGRP-LE gene silencing on survival
of Tenebrio molitor against Listeria monocytogenes infection. Int J Mol
Sci. 2013;14(11):22462–82.
41. Yano T, Mita S, Ohmori H, Oshima Y, Fujimoto Y, Ueda R, et al.
Autophagic control of Listeria through intracellular innate immune recognition in
Drosophila. Nat Immunol. 2008;9(8):908–16.
42. Kikuchi Y. Endosymbiotic bacteria in insects: their diversity and
culturability. Microbes Environ. 2009;24(3):195–204.
43. Regan JC, Brandão AS, Leitão AB, Mantas Dias AR, Sucena E, Jacinto
A, et al. Steroid hormone signaling is essential to regulate innate
immune cells and fight bacterial infection in Drosophila. PLoS Path.
2013;9(10):e1003720. doi:10.03710.1001371/journal.ppat.1003720.
44. Greenberg B. Persistence of bacteria in the developmental stages of
the housefly. 3. Quantitative distribution in prepupae and pupae. Am
J Trop Med Hyg. 1959;8(6):613–7.
45. Locatelli A, Spor A, Jolivet C, Piveteau P, Hartmann A. Biotic and abiotic soil
properties influence survival of Listeria monocytogenes in soil. Plos One.
2013, 8(10):e75969.
46. Lemon KP, Higgins DE, Kolter R. Flagellar motility is critical for Listeria
monocytogenes biofilm formation. J Bacteriol. 2007;189(12):4418–24.
47. Sasaki T, Kobayashi M, Agui N. Epidemiological potential of excretion
and regurgitation by Musca domestica (Diptera:Muscidae) in the
dissemination of Escherichia coli O157:H7 to food. J Med Entomol.
2000;37(6):945–9.
48. Rochon K, Lysyk TJ, Selinger LB. Retention of Escherichia coli by house
fly and stable fly (Diptera:Muscidae) during pupal metamorphosis and
eclosion. J Med Entomol. 2005;42(3):397–403.
49. Schuster GL, Donaldson JR, Buntyn JO, Duoss HA, Callaway TR,
Carroll JA, et al. Use of bioluminescent Escherichia coli to determine
retention during the life cycle of the Hhusefly, Musca domestica
(Diptera: Muscidae, L). Foodborne Pathog Dis. 2013;10(5):442–7.
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 11 of 12
12. 50. Ahmad A, Nagaraja TG, Zurek L. Transmission of Escherichia coli O157:H7
to cattle by house flies. Prev Vet Med. 2007;80(1):74–81.
51. Alam MJ, Zurek L. Association of Escherichia coli O157:H7 with houseflies on
a cattle farm. Appl Environ Microbiol. 2004;70(12):7578–80.
52. Iwasa M, Makino S, Asakura H, Kobori H, Morimoto Y. Detection
of Escherichia coli O157:H7 from Musca domestica (Diptera: Muscidae)
at a cattle farm in Japan. J Med Entomol. 1999;36(1):108–12.
53. Rahn K, Renwick SA, Johnson RP, Wilson JB, Clarke RC, Alves D, et al.
Persistence of Escherichia coli O157:H7 in dairy cattle and the dairy farm
environment. Epidemiol Infect. 1997;119(2):251–9.
54. Talley JL, Wayadande AC, Wasala LP, Gerry AC, Fletcher J, DeSilva U,
et al. Association of Escherichia coli O157:H7 with filth flies (Muscidae
and Calliphoridae) captured in leafy greens fields and experimental
transmission of E. coli O157:H7 to spinach leaves by house flies
(Diptera: Muscidae). J Food Prot. 2009;72(7):1547–52.
55. Radvan R. Persistence of bacteria during development in flies. III.
Localization of the bacteria and transmission after emergence of the fly.
Folia Microbiol. 1960;5:149–56.
56. Radvan R. Persistence of bacteria during development in flies. I. Basic
possibilities of survival. Folia Microbiol. 1960;5:50–6.
57. Anjam Khan CM. The dynamic interactions between Salmonella and the
microbiota, within the challenging niche of the gastrointestinal tract. Int
Sch Res Notices. 2014;2014(846049):1–23.
58. Hosokawa T, Kikuchi Y, Fukatsu T. How many symbionts are provided by
mothers, acquired by offspring, and needed for successful vertical
transmission in an obligate insect-bacterium mutualism? Mol Ecol.
2007;16(24):5316–25.
59. Min KT, Benzer S. Wolbachia, normally a symbiont of Drosophila, can be
virulent, causing degeneration and early death. P Natl Acad Sci USA.
1997;94(20):10792–6.
60. Goodacre SL, Martin OY. Modification of insect and Arachnid behaviours
by vertically transmitted endosymbionts: Infections as drivers of
behavioural change and evolutionary novelty. Insects. 2012;3(1):246–61.
61. Douglas AE, Francois CLMJ, Minto LB. Facultative ‘secondary’ bacterial
symbionts and the nutrition of the pea aphid, Acyrthosiphon pisum. Physiol
Entomol. 2006;31(3):262–9.
62. Mateos M, Castrezana SJ, Nankivell BJ, Estes AM, Markow TA, Moran NA.
Heritable endosymbionts of Drosophila. Genetics. 2006;174(1):363–76.
63. Oliver KM, Degnan PH, Burke GR, Moran NA. Facultative symbionts in aphids
and the horizontal transfer of ecologically important traits. Annu Rev
Entomol. 2010;55:247–66.
64. Oliver KM, Moran NA, Hunter MS. Costs and benefits of a superinfection
of facultative symbionts in aphids. P Roy Soc B-Biol Sci.
2006;273(1591):1273–80.
65. Oliver KM, Russell JA, Moran NA, Hunter MS. Facultative bacterial symbionts
in aphids confer resistance to parasitic wasps. P Natl Acad Sci USA.
2003;100(4):1803–7.
66. Su Q, Oliver KM, Pan H, Jiao X, Liu B, Xie W, et al. Facultative symbiont
Hamiltonella confers benefits to Bemisia tabaci (Hemiptera: Aleyrodidae), an
invasive agricultural pest worldwide. Environ Entomol. 2013;42(6):1265–71.
67. Li WR, Xie XB, Shi QS, Zeng HY, Ou-Yang YS, Chen YB. Antibacterial activity
and mechanism of silver nanoparticles on Escherichia coli. Appl Microbiol
Biotechnol. 2010;85(4):1115–22.
68. Sondi I, Salopek-Sondi B. Silver nanoparticles as antimicrobial agent: a case
study on E. coli as a model for Gram-negative bacteria. J Colloid Interface
Sci. 2004;275(1):177–82.
69. Halpin JL, Garrett NM, Ribot EM, Graves LM, Cooper KL. Re-evaluation,
optimization, and multilaboratory validation of the PulseNet-standardized
pulsed-field gel electrophoresis protocol for Listeria monocytogenes. Foodborne
Pathog Dis. 2010;7(3):293–8.
70. Ribot EM, Fair MA, Gautom R, Cameron DN, Hunter SB, Swaminathan B, et
al. Standardization of pulsed-field gel electrophoresis protocols for the
subtyping of Escherichia coli O157: H7 Salmonella, and Shigella for
PulseNet. Foodborne Pathog Dis. 2006;3(1):59–67.
71. Carter L, Lindsey LA, Grim CJ, Sathyamoorthy V, Jarvis KG, Gopinath G, et al.
Multiplex PCR assay targeting a diguanylate cyclase-encoding gene, cgcA, to
differentiate species within the genus Cronobacter. Appl Environ Microbiol.
2013;79(2):734–7.
72. Hosmer DW, Lemeshow S. Applied logistic regression. 2nd ed. New York:
John Wiley and Sons; 2000.
Submit your next manuscript to BioMed Central
and take full advantage of:
• Convenient online submission
• Thorough peer review
• No space constraints or color figure charges
• Immediate publication on acceptance
• Inclusion in PubMed, CAS, Scopus and Google Scholar
• Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit
Pava-Ripoll et al. BMC Microbiology (2015) 15:150 Page 12 of 12