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Detection and Isolation of Exotic Newcastle Disease Virus from Field-Collected Flies

  • U.S. National Poultry Research Center


Flies were collected by sweep net from the vicinity of two small groups of "backyard" poultry (10-20 chickens per group) that had been identified as infected with exotic Newcastle disease virus (family Paramyxoviridae, genus avulavirus, ENDV) in Los Angeles County, CA, during the 2002-2003 END outbreak. Collected flies were subdivided into pools and homogenized in brain-heart infusion broth with antibiotics. The separated supernatant was tested for the presence of ENDV by inoculation into embryonated chicken eggs. Exotic Newcastle disease virus was isolated from pools of Phaenicia cuprina (Wiedemann), Fannia canicularis (L.), and Musca domestica L., and it was identified by hemagglutination inhibition with Newcastle disease virus antiserum. Viral concentration in positive pools was low (<1 egg infectious dose50 per fly). Isolated virus demonstrated identical monoclonal antibody binding profiles as well as 99% sequence homology in the 635-bp fusion gene sequence compared with ENDV recovered from infected commercial egg layer poultry during the 2002 outbreak.
Detection and Isolation of Exotic Newcastle Disease Virus
from Field-Collected Flies
J. Med. Entomol. 44(5): 840Ð844 (2007)
ABSTRACT Flies were collected by sweep net from the vicinity of two small groups of backyard
poultry (10Ð20 chickens per group) that had been identiÞed as infected with exotic Newcastle disease
virus (family Paramyxoviridae, genus avulavirus, ENDV) in Los Angeles County, CA, during the
2002Ð2003 END outbreak. Collected ßies were subdivided into pools and homogenized in brain-heart
infusion broth with antibiotics. The separated supernatant was tested for the presence of ENDV by
inoculation into embryonated chicken eggs. Exotic Newcastle disease virus was isolated from pools
of Phaenicia cuprina (Wiedemann), Fannia canicularis (L.), and Musca domestica L., and it was
identiÞed by hemagglutination inhibition with Newcastle disease virus antiserum. Viral concentration
in positive pools was low (1 egg infectious dose
per ßy). Isolated virus demonstrated identical
monoclonal antibody binding proÞles as well as 99% sequence homology in the 635-bp fusion gene
sequence compared with ENDV recovered from infected commercial egg layer poultry during the
2002 outbreak.
KEY WORDS ßies, Newcastle disease virus, poultry, chickens, mechanical vector
Exotic Newcastle disease (END) is a contagious and
fatal viral disease affecting the respiratory, nervous,
and digestive systems, of poultry and other birds. END
is so virulent that many birds may die without ever
showing clinical signs of illness. There is a near 100%
mortality in unvaccinated poultry, and even poultry
vaccinated against the endemic low-virulence New-
castle disease virus strains are not adequately pro-
tected against END virus (family Paramyxoviridae,
genus avulavirus, ENDV) (USDAÐAPHIS 2006).
Newcastle disease control in poultry has been on-
going in the United States since the Þrst infections
were identiÞed in the early 1940s. Introductions of the
more virulent strains of Newcastle disease virus, the
cause of END, have occurred, and in most cases, the vi-
rus has been quickly eradicated (Utterback and
Schwartz 1973). During two END outbreaks, virus was
not quickly eradicated. The Þrst occurred during
1971Ð1973, and this outbreak resulted in the quaran-
tine of eight California counties, the destruction of
11.9 million birds, and eradication costs of $56 million
(USDAÐAPHIS 1978). The second occurred during
2002Ð2003, and it resulted in the quarantine of 18,345
premises, the destruction of 3.2 million birds, and
eradication efforts cost of $170 million (Breitmeyer et
al. 2003).
ENDV is primarily spread by direct contact be-
tween infected and healthy birds. However, it also can
be transmitted indirectly via contaminated equipment
and persons (Utterback and Schwartz 1973, Burridge
et al. 1975). During the 1971Ð1973 California END
outbreak, Rogoff et al. (1975) isolated virus from pools
of Fannia spp. collected at commercial poultry oper-
ations with ENDV infected birds, implicating this spe-
cies as a possible vector of ENDV.
Many insects, especially ßies, are commonly asso-
ciated with poultry operations, and they are known to
disperse into surrounding areas (Lysyk and Axtell
1986, Axtell 1999, Sawabe et al. 2006). Wet manure is
an excellent breeding environment for several ßy spe-
cies, and it also likely to support the survival of ENDV
(Kinde et al. 2004). Flies are capable of harboring a
diverse range of animal and human pathogens (Cali-
beo-Hayes et al. 2003, Graczyk et al. 2005, Sawabe et
al. 2006), and they may be involved in transmission of
these pathogens to otherwise healthy hosts (Shane et
al. 1985, Calibeo-Hayes et al. 2003, Ahmad et al. 2007).
However, the extent to which insects may be involved
in the dispersal and transmission of ENDV to unin-
fected birds remains unknown.
In this study, ßy species collected at two private
homes with ENDV-infected poultry in Los Angeles
County, CA, were examined for the presence of
ENDV. Isolated ENDV was characterized by mono-
clonal antibody binding proÞle and sequence analysis
of the fusion gene.
Department of Entomology, University of California, Riverside,
CA 92521.
Veterinary Medicine Extension, University of California, Davis,
CA 95616.
University of California Cooperative Extension, Riverside, CA
Corresponding author, e-mail:
Materials and Methods
Flies were captured by sweep net from the imme-
diate vicinity of two private homes in Los Angeles Co.,
southern California. Both locations, one location in
the city of Compton (home A), and the other location
in Inglewood (home B), contained chicken ßocks
(20 birds) that had been identiÞed by the California
Department of Food and Agriculture (CDFA) as hav-
ing one or more ENDV-infected birds within the pre-
ceding 5 d. Flies were collected before poultry were
euthanized and removed from the private homes by
the END Task Force (a joint United States Depart-
ment of Agriculture and CDFA organization) respon-
sible for efforts to eradicate END in California. Fly
collectors followed all biosecurity precautions recom-
mended by the CDFA while sampling at these homes.
Multiple 5-min sweep collections were made at each
home, taking care not to touch poultry cages or the
ground with the net while sweeping. Sweep net col-
lections continued at each home until ßies were no
longer readily captured. Captured insects were killed
on dry ice, placed into labeled vials by using sterilized
forceps, and held on dry ice until they could be re-
turned to the laboratory to be identiÞed and counted.
Sweep nets and forceps were sterilized between each
5-min collection by submergence in 10% Lysol (Rec-
kitt Benckiser Inc., Wayne, NJ) after which nets were
allowed to dry before reuse.
Flies were kept frozen on dry ice during identiÞ-
cation in a biological safety cabinet, and then they
were pooled into groups of Þve or fewer ßies of the
same species from the same home. Fly pools were
stored at 80C until shipment on dry ice to the USDA
Southeast Poultry Research Laboratory (SEPRL), in
Athens, GA, where they were again placed at 80C
until tested for the presence of ENDV. Pools were
homogenized in 1.5 ml of brain-heart infusion (BHI)
broth with antibiotics (200
g of gentamicin/ml, 2000
U of penicillin/ml, and 4
g of amphotericin B/ml,
Sigma-Aldrich, St. Louis, MO) by using a tissue grinder
with sterile plastic pestles in microfuge tubes and
centrifuged at 16,000 g for 10 min. Virus isolation
was performed by inoculating 100
l of the superna-
tant into the allantois of each of three 9- or 10-d-old
embryonated chicken eggs (ECE). Eggs were incu-
bated at 37C in a standard humidiÞed incubator. The
embryonated chicken eggs were obtained from the
SEPRL speciÞc-pathogen-free White Leghorn ßock.
Eggs were candled to determine embryo death each
24 h through 7 d postinoculation. Embryos that died
within the Þrst 24 h were discarded. Embryos that died
between 24 h and7daswell as all survivors at 7 d were
chilled at 4C. Amnio-allantoic ßuid (AAF) harvested
from chilled eggs was tested for hemagglutination
(HA) activity to detect ENDV. Virus presence in HA-
positive samples was conÞrmed by hemagglutination-
inhibition (HI) with Newcastle disease virus (family
Paramyxoviridae, genus Avulavirus, NDV)-speciÞc
antiserum (King 1996a). Amnio-allantoic ßuids from
HA negative dead embryos and embryos alive at 7 d
postinoculation were subjected to a second serial pas-
sage by inoculation of 100
l of the AAF into each of
three additional embryonated chicken eggs. Eggs
were candled, and killed embryos were handled as
before. If by day 7 postinoculation there was no HA
activity in the AAF of the second passage dead or
surviving embryos, the specimen was regarded as neg-
ative for ENDV.
A monoclonal antibody proÞle was determined for
each isolated ENDV using nine monoclonal antibodies
(mAbs) with different Newcastle disease virus HI
speciÞcities by using previously described methods
(King 1996a, Kommers et al. 2001). The mAbs in-
cluded 15C4, AVS, B79, 161/167, P11C9, P3A11,
10D11, P15D7, and P10B8 (Kommers et al. 2003).
RNA was extracted from AAF by using TRIzol LS
according to manufacturerÕs instructions (Invitrogen,
Carlsbad, CA); 750
l of TRIzol LS reagent was added
to 250
l of allantoic ßuid. The ßuid was vortexed and
incubated at room temperature for 7 min. RNA was
separated into the aqueous phase with the addition of
l of chloroform, followed by precipitation with
isopropanol. After one wash with 70% ethanol, RNA
was dried and resuspended in RNase-free water. The
5 end of the ENDV fusion gene was ampliÞed by
polymerase chain reaction (PCR) and sequenced us-
ing primers targeting a 635-bp fragment that included
the fusion protein cleavage site (forward primer, 5-
reverse primer, 5-TCA TTA ACA AAY TGC TGC
ATC TTC CCW AC-3). These primers amplify the
NDV genomic region between positions 4317 and
5084. Standard 50
l reverse transcription (RT)-PCR
reactions were carried out using a kit (SuperScript III
One Step RT-PCR, Invitrogen) with annealing tem-
perature at 56C. PCR-ampliÞed samples were sepa-
rated on a 1% agarose gel, and the bands were excised
and eluted using the QuickClean 5M gel extraction kit
(GenScript Corp., Piscataway, NJ). Once the PCR
products were cleaned, samples were quantiÞed using
a standard spectrophotomer and sequenced. All dou-
ble-stranded nucleotide-sequencing reactions were
performed with ßuorescent dideoxynucleotide termi-
nators in an automated sequencer (ABI 3700 auto-
mated sequencer, Applied Biosystems, Foster City,
CA). Nucleotide sequence editing and analysis were
conducted with the LaserGene sequence analysis soft-
ware package (LaserGene, version 5.07, DNAStar,
Inc., Madison, WI). The virus was sequenced and
compared by Blast analysis (Altschul et al. 1990) to an
END viral sequence [chicken/U.S.(AZ)/236498/03]
obtained from commercial poultry infected with
ENDV during the 2002 outbreak. It also was compared
with other available GenBank ENDV sequences.
Genomic sequences from viruses recovered from ßies
at each collection site were deposited in GenBank
(accession nos. EF424375 and EF424376).
Overall, 87 ßies composing nine species were col-
lected from the area surrounding the two private
homes housing ENDV-infected birds (Table 1). Most
September 2007 C
of the collected ßies were Phaenicia cuprina (Wiede-
mann), Musca domestica L., or Fannia canicularis (L.).
There were 24 ßy pools representing groups of 5 ßies
of the same species from the same home. Five pools of
ßies contained ENDV; three pools of P. cuprina (two
pools from home A and the other pools from home B)
and one each of F. canicularis and M. domestica (both
from home B). The amount of virus recovered from
each of the ßy pools was very low (1 egg infectious
dose [EID]
per ßy) and required a second serial
passage in ECE for detection.
The mAb binding proÞle (Table 2) of the virus
isolated from each of the ßy pools was identical to an
ENDV strain obtained from infected commercial egg-
layer poultry during the 2002Ð2003 END outbreak
[chicken/U.S.(CA)/12430/02] and different than the
proÞle of the heterologous NDV B1 strain included as
a control. The NDV B1 strain is commonly used as a
live virus vaccine in commercial poultry. The partial
sequence of the fusion gene of the virus isolated from
the ßies showed 99% homology (634/635 bp) to a
2002Ð2003 ENDV isolate [chicken/U.S.(AZ)/236498/
03] and 87% homology (552/635 bp) to a 1971Ð1973
ENDV isolate [chicken/U.S.(CA)/1083(Fontana)/
72], both obtained from commercial egg-layer poultry
during ENDV outbreaks. At the amino acid level, the
partial sequence of the ßy isolate was 100% identical
to the 2002Ð2003 outbreak strain and 92% identical to
the 1971Ð1973 outbreak strain. Viruses recovered from
ßies at both collection sites were 100% identical to
each other.
Although all ßy species were collected in small
numbers, ENDV was isolated from the three most
abundant species collected during this study. This is
the Þrst report of ENDV isolated from Þeld-collected
P. cuprina and M. domestica; ENDV had previously
been isolated from F. canicularis and Fannia femoralis
(Stein) from a commercial poultry operation during
the 1971 END outbreak (Rogoff et al. 1975). Given the
small numbers of ßies collected, the prevalence of
ENDV-infected ßies was high with 30% of the pools
containing the three species mentioned above having
one or more ßies per pool infected with virus. The
virus prevalence in Fannia spp. collected from com-
mercial poultry operations during the 1971 END out-
break was far lower with only two of 78 pools of Fannia
spp. (3,926 total ßies) collected before or within2dof
poultry removal containing ENDV, and no virus was
recovered from other common ßy species (Rogoff et
al. 1975). It is unclear to what extent Rogoff et al.
(1975) may have collected ßies from poultry facilities
where poultry or even manure had already been re-
moved from the site, perhaps resulting in the much
lower ENDV infection prevalence in ßies relative to
this study. The higher ENDV prevalence in this study
also may be due to differences between commercial
poultry and backyard poultry in animal housing, ma-
nure handling, or vaccination status of the poultry
against Newcastle disease virus. Backyard poultry are
often housed together on the ground or in wooden
boxes, giving them greater access to an infected bird
and its manure relative to commercial egg-layer poul-
try that are separated into cages suspended above the
ground where the manure is allowed to accumulate.
Additionally, backyard poultry are usually not vacci-
nated against Newcastle disease virus and infected
Table 1. Detection of ENDV from flies collected at two homes
(A and B) in Los Angeles County with ENDV-infected poultry during
2002–2003 ENDV outbreak
Fly species
No. ßies
Positive pools
Calliphora spp. 1 1
Fannia canicularis 10 10 2 2 1 (n 5)
Fannia femoralis 11
Musca domestica 8152 3 1(n 5)
Muscina stabulans 2111
Phaenicia sericata 21
Phaenicia cuprina 14 21 3 5 2 (n 10) 1 (n 5)
Phormia regina 11
Stomoxys calcitrans 11
Table 2. Hemagglutination-inhibition test results of ENDV isolated from field-collected flies against NDV-specific mAbs
Viral isolate
15C4 AVS B79 161/617 P11C9 P3A11 10D11 P15D7 P10B8
P. cuprina (A1) ⫹⫺
P. cuprina (A2) ⫹⫺
P. cuprina (B1) ⫹⫺
F. canicularis (B1) ⫹⫺
M. domestica (B1) ⫹⫺
CK/CA/12430/02 (outbreak strain) ⫹⫺
CK/US/B1/48 (vaccine strain) ⫹⫹
Antibody-inhibited HA (); no HA inhibition (). 15C4 inhibits all NDV but pigeon PMV-1. AVS inhibits lentogenic strains like the B1
and LaSota ND vaccine virus. B79 inhibits most NDV including most pigeon PMV-1. 161/617 inhibits only pigeon PMV-1. P11C9, P3A11, P15D7,
and P10B8 inhibit various NDV strains. 10D11 inhibits neurotropic NDV mesogens and velogens such as virulent Texas GB. CK/CA/12430/02
is an ENDV strain recovered from commercial poultry during the 2002Ð2003 outbreak in California. CK/US/B1/48 is a strain widely used as
a Newcastle disease vaccine in the United States. The antibody inhibition proÞle of the viruses tested indicates the ENDV isolates from three
ßy species at premises A and B are antigenically similar as well as being similar to the CK/CA/12430/02 ENDV isolate from chickens but are
different than the vaccine strain B1.
birds would be expected to shed a greater concentra-
tion of ENDV in their manure relative to vaccinated
commercial poultry (Kapczynski and King 2005).
The mAb proÞle and sequence analysis of the virus
recovered from each of the ßy pools indicated that
these viruses were the same ENDV strain circulating
in commercial and backyard poultry during this same
period. Viruses recovered during 2002Ð2003 were ge-
notypically different from viruses recovered during
the earlier 1971Ð1973 ENDV outbreak (genotypes V
and VI, respectively) (Czegledi et al. 2006) with virus
isolates from the 2002Ð2003 ENDV outbreak being
most similar to ENDV isolated from chickens in Mex-
ico during 1998Ð2000 (Pedersen et al. 2004).
Flies are thus contacting infectious ENDV in the
environment and are capable of harboring at least low
levels of this virus. The ENDV infective dose for a
chicken is reported to range from 10
(King 1996b)
to 10
(Alexander et al. 2006), far greater than
the 1 EID
per ßy found in our Þeld-collected ßy
pools. However, given the small number of ßies col-
lected, it would be premature to assume ßies are not
capable of carrying substantially higher viral loads. It
is entirely possible that these ßies had contacted a
source of ENDV several days before our sampling and
the low viral load represented residual virus still as-
sociated with the ßies at the time of their capture.
The mechanism by which the ßies might be acquir-
ing virus has not been determined. It has been shown
that ENDV-infected poultry shed a considerable
amount of virus in their feces (Kapczynski and King
2005) and that virus can persist in the manure for up
to 16 d (Kinde et al. 2004). Wet manure provides an
excellent developmental environment for several ßy
species as well as a substrate on which adult ßies feed.
Manure also provides an excellent media for main-
taining and transmitting infectious agents to ßies as the
viscosity of the manure enhances manure attachment
to ßy tarsi and cuticular bristles (Tan et al. 1997). In
addition, ßies feeding on bacteria in the manure would
be expected to consume some amount of the infec-
tious agent. Further studies are needed to determine
whether ENDV is associated with these ßy species
simply as an external contaminant or if the virus is
protected or even ampliÞed within the gut or other
tissues of the ßy.
Shane et al. (1985) demonstrated the potential for
house ßies to transmit pathogens by placing ßies into
cages with Campylobacter jejuni-infected chickens
and subsequently moving the ßies to cages with patho-
gen-free chickens, resulting in C. jejuni infection in the
previously uninfected chickens. Similarly, turkey
poults housed in isolation units became infected with
turkey corona virus (family Coronaviridae, genus
Coronavirus, TCV) when house ßies orally inoculated
with TCV were released into the isolation units for
24 h (Calibeo-Hayes et al. 2003), and cattle held in
conÞned pens became infected with Escherichia coli
O157:H7 after house ßies orally inoculated with the
bacteria were released into the pens for 48 h (Ahmad
et al. 2007). However, the actual mechanism of bac-
terial transmission from the ßies to the chickens or
cattle in the studies mentioned above remains un-
Poultry are known to consume adult ßies in labo-
ratory studies (Calibeo-Hayes et al. 2003) as well as in
the Þeld (A.C.G., unpublished observation). Addi-
tionally, ßies may regurgitate and defecate virus while
feeding or resting on the surface of foods (Greenberg
1973). It is possible that chickens and other birds can
consume virus with recently infected ßies or contam-
inated ßy feces and regurgitated matter (Sawabe et al.
2006). The consumption of many adult ßies in a short
period might result in infection even if the viral load
of each individual ßy is lower than the required in-
fectious dose.
Fly larvae developing in infected manure also might
serve as a source of infection when consumed by
poultry having access to the manure (typical of back-
yard poultry and some commercial poultry), although
this has not been evaluated. Additional studies are
required to evaluate the vector competence of ßies
and other poultry associated arthropods.
Dispersal of infectious virus by ßies moving from
premises with infected poultry to nearby premises
with uninfected poultry is an important biosecurity
concern for END and other avian diseases. Greenberg
(1973) reported that house ßies were capable of ßights
ranging from 2.3 to 11.8 km within 24 h. Although ßy
involvement in virus movement between commercial
poultry facilities is probably minor relative to other
means of virus movement (e.g., on personnel, shared
equipment, or manure hauling vehicles), ßy involve-
ment in virus movement among poultry houses at a
single commercial facility or among backyard poultry
may be signiÞcant given the short distances ßies would
need to move between infected and uninfected poul-
try. Backyard poultry are geographically clumped in
southern California with small numbers of poultry at
many adjacent homes and only a few meters often
separating poultry at one home from the next. During
the 2002Ð2003 END outbreak, homes with ENDV-
infected poultry were often very close to each other,
forming a cluster that was distinct within the some-
what larger geographic clumping of backyard poultry
(C.J.C., unpublished data). In addition to ßy dispersal,
movement of birds or personnel between these clus-
tered homes also may have occurred.
Although this study has not shown ßies to be com-
petent vectors of ENDV, a conservative approach
would indicate that biosecurity measures should in-
clude ßy control on and near premises with END-
infected poultry as also suggested by Bram et al.
(1974). Fly control measures would be especially im-
portant for a quarantined poultry facility (to include
backyard ßocks) before removal of birds and their
manure by a state and/or federal task force as part of
the disease eradication effort. Removal of birds and
manure would be expected to encourage ßy dispersal
into the surrounding area. It is recommended that
future END task force organizations consider the use
of insecticides providing rapid knockdown of adult
ßies immediately before initiating eradication efforts
at commercial and backyard poultry facilities and that
September 2007 C
control measures continue throughout the period of
manure removal and site disinfection.
We thank Diane Zhang for ßy colony maintenance and
Timothy Olivier and Dawn Williams-Coplin for help in con-
ducting the experiments. This study was funded by USDAÐ
CSREES grant 2003-34439-13366 (Exotic Pests and Dis-
eases) managed by the UCANR Statewide IPM Program and
awarded to A.C.G. and C.J.C.; and USDAÐARS CRIS project
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Received 21 March 2007; accepted 3 June 2007.
... M. domestica has also been reported to mechanically transmit several types of viral pathogens to livestock including: avian influenza virus (AIV), both high and low pathogenic strains [46,[212][213][214][215][216], turkey coronavirus (TCV) [217], Newcastle disease virus (NDV) [218][219][220][221][222], reticuloendotheliosis virus (REV) [223], porcine reproductive and respiratory syndrome virus (PRRSV) [224][225][226][227][228], porcine circovirus genotype 2 (PCV2b) [229], porcine epidemic diarrhea virus (PEDV) [230], African swine fever virus (ASF) [231,232], Aujeszky's virus (PRV-1) [233], senecavirus A (SVA) [234], Rift Valley fever virus (RVFV) [235], Aleutian mink disease virus (AMDV) [236,237], and lumpy skin disease (LSDV) [238,239].Viruses detected in dipteran edible species are listed in Table 4. ...
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Edible insects are expected to become an important nutrient source for animals and humans in the Western world in the near future. Only a few studies on viruses in edible insects with potential for industrial rearing have been published and concern only some edible insect species. Viral pathogens that can infect insects could be non-pathogenic, or pathogenic to the insects themselves, or to humans and animals. The objective of this systematic review is to provide an overview of the viruses detected in edible insects currently considered for use in food and/or feed in the European Union or appropriate for mass rearing, and to collect information on clinical symptoms in insects and on the vector role of insects themselves. Many different virus species have been detected in edible insect species showing promise for mass production systems. These viruses could be a risk for mass insect rearing systems causing acute high mortality, a drastic decline in growth in juvenile stages and in the reproductive performance of adults. Furthermore, some viruses could pose a risk to human and animal health where insects are used for food and feed.
... Because F. canicularis can develop in animal feces and adult flies will also contact feces to oviposit or feed, they are potential vectors for several important animal and human pathogens. Viruses include virulent Newcastle disease virus (Rogoff et al. 1975, Chakrabarti et al. 2007) and Aleutian mink disease virus (Prieto et al. 2018 Burnett et al. 1957, Steve 1960. ...
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The little house fly, Fannia canicularis (L.) (Diptera: Fanniidae), is a significant pest associated with livestock and animal systems worldwide. This species commonly develops in poultry production systems. The males of this species are a nuisance to people because they form mating swarms in enclosed spaces. The pest status of F. canicularis has not lessened since it was identified as a critical arthropod pest of veterinary importance over 50 yr ago. During this period, there has been little research progress to control this pest, especially when compared with other filth fly species. This article reviews the biology, distribution, pest status (including nuisance and pathogen transmission risk), monitoring, and control techniques, and identifies knowledge gaps for F. canicularis.
... The disease is endemic worldwide and causes highly economic losses due to high mortalities and reduced production [1][2][3][4][5]. The main route of NDV transmission is airborne route and also it can spread through direct contact with infected birds, contaminated poultry products, people with contaminated clothes or shoes, equipment, vaccines [6], contaminated water [7] and insects in a poultry house [8]. ...
... (Lignieres 1900), Shigella spp. ( Castellani and Chalmers 1919), Vibrio cholera (Pacini 1854), enterohemorrhagic Escherichia coli O157:H7 (Migula 1895, Castellani and Chalmers 1919), Newcastle virus, and porcine respiratory virus (Olsen 1998, Kobayashi et al. 1999, Graczyk et al. 2001, Schurrer et al. 2004, Chakrabarti et al. 2007, Pitkin et al. 2009, and this list is steadily growing. One management method used to control adult house flies is through the application of granular fly baits containing an insecticidal active ingredient (a.i.) formulated into a food matrix, which encourages fly feeding and consumption of the insecticide (Mayeux 1954, Keller 1955. ...
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The mortality rate of a field population of house fly (Musca domestica L.) was determined for a granular fly bait containing the active ingredient indoxacarb, which was compared to two commercially available granular fly baits containing either dinotefuran or cyantraniliprole. Indoxacarb was applied at three different application rates 0.498, 0.986, and 1.972 g/m2 (low, medium, and high). Time to 50% mortality was fastest for dinotefuran (5.7 h) and slowest for the low application rate of indoxacarb (10.3 h). Time to 90% mortality was fastest for the high application rate of indoxacarb (27.7 h) and slowest for dinotefuran (51.0 h) and cyantraniliprole (45.9 h). Among the three indoxacarb application rates, the high rate reached both 50 and 90% fly mortality significantly faster than the low rate. The medium rate did not significantly differ from either the high or low application rates. Dinotefuran bait produced greater fly mortality than all other treatments at 30-min post-exposure, with mortality for remaining baits exceeding controls by 3- to 6-h post-exposure. All insecticidal baits produced similar fly mortality by 6-h post-exposure and >94% fly mortality by 96-h post-exposure, indicating that each may be effective in a fly management program. Flies consumed a similar amount of the indoxacarb (regardless of application rate) and dinotefuran baits, but consumed less of the cyantraniliprole bait, suggesting a feeding irritancy or toxicity effect manifested during consumption. Nevertheless, flies consumed enough cyantraniliprole bait to cause mortality similar to other baits by 6-h post-exposure.
... This may be attributed to inadequate protection against vaccine strains used in the poultry industry and poor biosecurity (Chukwudi et al., 2012). Likewise, it is also possible that NDV strain circulating in the environment is a different pathotype or strain, or exotic birds may be responsible for spreading the infection (Chakrabarti et al., 2007). Vaccine strains are of genotype II, which has different genetic properties than genotype VII and VIII that are causing sporadic outbreaks (Forrester et al., 2013). ...
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This study aimed to characterize the lentogenic NDV isolate from native chickens in Surabaya, Indonesia. Thirty seven samples of cloacal swabs from infected native chickens were collected, and pathotypes were characterized using Mean Death Time (MDT) analysis; six isolates were found to be lentogenic. Lentogenic NDV isolates were then analyzed via RT-PCR, using primers specific for cleavage site of fusion protein (F). PCR product was sequenced and analyzed using Epitope Prediction Tools (IEDB) Analysis Resource to determine the epitopes. The results showed some shifts in nucleotides, but no change was observed in amino acids. Five samples were found with similar sequence in the cleavage site, except NDV isolated from sample Ck/sby27, which had a different amino acid sequence, “RRQKRFI.” Epitope characterization specific for T-cell NDV was found in Ck/sby27 at position 42-142, with highest real score in epitopes CLDYLQLVY, SIDGRPLAA, and TAEQITAAA. These findings reinforce the assumption that the original NDV lentogenic strain is of wild origin and is still circulating in the environment. Therefore, NDV isolate Ck/ sby27 can be used in developing vaccines in the future. © University of the Philippines at Los Banos. All rights reserved.
... Rogoff et al. (1977) clearly showed that virulent velogenic NDV can be transmitted to young chickens by Fanniacanicularis, either from a highly infective source or directly from infected birds. Exotic NDV was isolated from houseflies collected from backyard flocks during an outbreak in the U.S.A. ( Chakrabarti et al., 2007). Also, laboratory-reared flies that were experimentally exposed to NDV La Sota strain, the virus was detected in the dissected gastrointestinal tract of flies for up to 72 h post-exposure ( Barin et al., 2010) But, these infected flies could infect sensitive chickens under laboratory conditions the housefly did not carry sufficient quantities of NDV (Roakin strain) to cause disease in chickens ( Watson et al., 2007). ...
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Flies are the most important insect pest associated with poultry, where the accumulated organic waste and favorable environmental conditions often promote rapid development of large populations. This study aims to evaluate flies as a vector for avian viral pathogens. A total of 90 flies were collected from campus of School of Veterinary Medicine, Shiraz University, Iran. Reverse transcription-polymerase chain reaction test using specific published primers for Newcastle Disease virus, Avian Influenza virus, Infectious Bronchitis virus and Infectious Bursal Disease virus was carried out to detect the viruses. The M1 gene of avian influenza and F gene of Newcastle disease virus genes were detected from 18 and 32 separated samples respectively by reverse transcription-polymerase chain reaction. Infectious Bronchitis and Infectious Bursal Disease viruses were not detected from all houseflies. These results suggest that it is possible that flies could become a mechanical transmitter of avian influenza and Newcastle disease viruses.
... For example, Newcastle disease virus (NDV) RNA was detected in, and even virions were isolated from, flies collected in the vicinity of infected chickens. Similarly, H5N1 RNA was found in flies collected in the surroundings of a poultry farm with infected birds 22,23 . ...
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Monitoring wildlife infectious agents requires acquiring samples suitable for analyses, which is often logistically demanding. A possible alternative to invasive or non-invasive sampling of wild-living vertebrates is the use of vertebrate material contained in invertebrates feeding on them, their feces, or their remains. Carrion flies have been shown to contain vertebrate DNA; here we investigate whether they might also be suitable for wildlife pathogen detection. We collected 498 flies in Taï National Park, Côte d'Ivoire, a tropical rainforest and examined them for adenoviruses (family Adenoviridae), whose DNA is frequently shed in feces of local mammals. Adenoviral DNA was detected in 6/142 mammal-positive flies. Phylogenetic analyses revealed that five of these sequences were closely related to sequences obtained from local non-human primates, while the sixth sequence was closely related to a murine adenovirus. Next-generation sequencing-based DNA-profiling of the meals of the respective flies identified putative hosts that were a good fit to those suggested by adenoviral sequence affinities. We conclude that, while characterizing the genetic diversity of wildlife infectious agents through fly-based monitoring may not be cost-efficient, this method could probably be used to detect the genetic material of wildlife infectious agents causing wildlife mass mortality in pristine areas.
The Paramyxoviridae family has several genera that include important human and veterinary pathogens such as Rubulavirus, Respiroviruses, Henipavirus, and the Avulavirus genus that contains Newcastle disease virus (NDV) and other avian paramyxoviruses (APMV). This chapter focuses on infections of poultry with NDV. It offers detailed coverage of the history, etiology, pathobiology, epidemiology, diagnosis, and intervention strategies of Newcastle disease, APMV, and avian Metapneumovirus Infections. AMPV infections continue to emerge as a disease threat with four defined subtypes, A–D, being recognized and producing clinical disease in both turkeys and chickens. For effective disease management, it is important to be able to identify birds that are infected with NDV and distinguish vaccine viruses from virulent viruses. Regardless of whether ND control is applied at the international, national, or farm level, the objective is either to prevent susceptible birds from becoming infected or to reduce the number of susceptible birds by vaccination.
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The presence of various species of filth flies is a widespread problem where livestock, including poultry, are maintained and where manure accumulates. The house fly, Musca domestica L.; the stable fly, Stomoxys calcitrans (L.); and the little house fly, Fannia canicularis (L.) (each Diptera: Muscidae), the target pests in our study, can mechanically spread diseases, and S. calcitrans can bite cattle, causing losses in meat and milk production. Chemical control is widely used to suppress filth flies, but resistance to conventional insecticides has become problematic. Hence, an alternative approach, insect growth regulators (IGRs), has been adopted by many livestock producers. We assessed the ability of the IGR cyromazine in granular and granular-based aqueous formulations to suppress the three muscid species from developing in poultry, cattle, and swine manure collected from commercial livestock production facilities. Each of the two formulations provided either strong or complete control of the pests for the 4-wk duration of the study, excluding the granular formulation that provides control of only F. canicularis developing in poultry manure for 2 wk. The two cyromazine-based IGR formulations appear to be effective tools that, if rotated appropriately with other insecticides, can be incorporated into integrated pest management strategies for filth fly suppression.
In recent years, avian influenza virus (AIV) and Newcastle disease virus (NDV) have caused large-scale outbreaks in many countries, including Egypt. The culling and vaccination strategies have failed to control both viruses in Egypt. In this study, we investigated the outbreaks of nervous manifestations and deaths in pigeons between 2013 and 2015. The H5N1 subtype of the highly pathogenic avian influenza (HPAI) virus and pigeon paramyxovirus-1 (PPMV-1), an antigenic variant of NDV, were found to be the cause; AIV and PPMV-1 were isolated from 61.3% (19/31) and 67.8% (21/31) of tested pigeons, respectively. Co-infection with both viruses was detected in 51.6% of pigeons (16/31). The AIV sequences showed PQGEKRRKKR/GLF motif at the HA gene cleavage site, which is typical of highly pathogenic H5N1 subtype. The phylogenetic tree showed that the HPAI belonged to clade The NDV sequences carried one of the three motifs, ¹¹²GKQGRL¹¹⁷, ¹¹²KRQKRF¹¹⁷ or ¹¹²RRQKRF¹¹⁷, at the fusion protein cleavage site and were classified as genotypes I, VI and II in NDV­-class II, respectively. This indicated that different genotypes of NDV can circulate simultaneously among pigeons. Further analysis revealed the clustering of some sequences in subgenotypes Ia and VIb.2. To the best of our knowledge, these subgenotypes have not been previously reported from pigeons in Egypt. Our results should serve as a base for future studies on both viruses in Egypt.
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A new approach to rapid sequence comparison, basic local alignment search tool (BLAST), directly approximates alignments that optimize a measure of local similarity, the maximal segment pair (MSP) score. Recent mathematical results on the stochastic properties of MSP scores allow an analysis of the performance of this method as well as the statistical significance of alignments it generates. The basic algorithm is simple and robust; it can be implemented in a number of ways and applied in a variety of contexts including straightforward DNA and protein sequence database searches, motif searches, gene identification searches, and in the analysis of multiple regions of similarity in long DNA sequences. In addition to its flexibility and tractability to mathematical analysis, BLAST is an order of magnitude faster than existing sequence comparison tools of comparable sensitivity.
Exotic Newcastle disease (also known as velogenic viscerotropic Newcastle disease, Asiatic Newcastle disease, and Doyle's form of Newcastle disease) is a virus disease affecting domestic poultry and other avian species. It is that form or expression of Newcastle disease which is characterized by the causative virus having high lethality and an affinity for visceral organs, especially the digestive tract (Walker et al. 1973). Observed almost invariably in victims are hemorrhagic lesions and necrotic areas on the mucosal surface of the various subdivisions of the intestinal tract, especially the cecal tonsils, proventriculus, and Peyer's patches. Edema of the tissue along the trachea and near the thoracic inlet is also observed frequently. Susceptible chickens, artificially infected, die in 5-7 days with virtually 100 percent mortality.
Modern commercial poultry production under large companies is expanding worldwide with similar methods and housing, and the accompanying arthropod and rodent pest problems. The pests increase the cost of production and are factors in the spread of avian diseases. The biology, behavior and control of ectoparasites and premise pests are described in relation to the different housing and production practices for broiler breeders, turkey breeders, growout (broilers and turkeys), caged-layers, and pullets. Ectoparasites include Ornithonyssus fowl mites, Dermanyssus chicken mites, lice, bedbugs, fleas, and argasid fowl ticks. Premise pests include Alphitobius darkling beetles, Dermestes hide beetles, the house fly and several related filth fly species, calliphorid blow flies, moths, cockroaches, and rodents. Populations of these pests are largely determined by the housing, waste, and flock management practices. An integrated pest management (IPM) approach, tailored to the different production systems, is required for satisfactory poultry pest control. Biosecurity, preventing the introduction of pests and diseases into a facility, is critical. Poultry IPM, based on pest identification, pest population monitoring, and methods of cultural, biological, and chemical control, is elucidated. The structure of the sophisticated, highly integrated poultry industry provides a situation conducive to refinement and wider implementation of IPM.
Field collections of insects in the vicinity of poultry flocks infected with the virus of exotic viscerotropic, velogenic Newcastle disease (VVND) were made in Riverside and San Bernardino Counties, southern California. Virus was isolated from 3 pools of Fannia canicularis (L.) and 1 of F. femoralis (Stein). Another pool of F. canicularis contained a mesogenic ND virus, and 1 pool of Musca domestica L. larvae had VVND virus. Newcastle disease virus was not isolated from 117 pools of M. domestica adults nor from 19 pools of Muscina stabulans (Fallén) tested. None of the other insect species collected showed evidence of the presence of Newcastle disease virus.
Data collected during the velogenic viscerotropic Newcastle disease (VVND) epidemic that occurred in southern California from 1971 to 1973 were analyzed to determine the methods of spread of the disease. Spread between chicken flocks was extensive and due mainly to the movement of live birds and mechanical transport of virus by man, especially by vaccination and poultry service crews. Spread to exotic birds was from contact with infected imported stock. Spread to other species was most probably through contact with infected chickens. Infection persisted in commercial chicken flocks because of intensive vaccination programs, heavy traffic and contact between layer operations, and the maintenance of multi-age flocks. These foci of infection probably led to spread of the disease to areas from which VVND had been eradicated several months before. There was no evidence of significant wind-borne spread of virus between flocks.
House fly, Musca domestica L., movement between breeding sites (dairies and poultry houses) and into surrounding habitats (buildings, fields, and pastures) was mea- sured by releasing marked flies in poultry houses and dairies at two farms. Relative density of wild and marked flies was greater in the dairy and poultry houses than in other habitats at similar distances from the release areas. Calculated proportions of wild flies in each habitat were greatest in the dairies and poultry houses; averages of 25 and 36% of the wild popu- lations were estimated to be in the other habitats. After 5 days, an average of 60 and 53% of marked flies released in the poultry houses remained there, 13% moved to the dairies at both farms, and 27 and 34% moved from the poultry houses to the nonbreeding habitats. An average of 56 and 73% of the marked flies released in the dairies remained there after 5 days, while 8 and 10% moved into the poultry houses, and 34 and 19% moved from the dairies into the nonbreeding habitats at both farms.
Houseflies (Musca domestica) were infected with Campylobacter jejuni after being confined for 5 days in a Horsfall isolator containing 25-day-old chickens known to be fecal excretors of the organism. Contaminated flies, when subsequently transferred to a second unit, transmitted C. jejuni to specific-pathogen-free chickens. Allowing a sample of 32 houseflies to ingest C. jejuni in a liquid suspension resulted in recovery rates of 20% from the feet and ventral surface of the body and 70% from the viscera. These experiments demonstrated the potential role of flies in the dissemination of avian campylobacteriosis.
Velogenic viscerotropic Newcastle disease (VVND) was first diagnosed in southern California near Fontana in San Bernardino County in late November, 1971. The virus appeared to have escaped from a nearby exotic bird importer. Once established in poultry, the virus spread rapidly to other areas, primarily by movement of poultry, service people, and equipment among the poultry ranches. Initial quarantine and eradication measures proved inadequate to curb the spread. By March 14, 1972, when the national emergency was declared, more than 150 flocks were known to be infected, including most of the major poultry areas of southern California. A massive effort was mounted by a state federal task force to contain VVND in southern California and to eradicate the disease before it became established as an enzootic disease. Eight southern California counties were quarantined. The eradication effort was organized to diagnose and depopulate all known infected flocks, vaccinate the entire avian population rapidly, and to restrict movement of birds and poultry products. This program contained VVND within the quarantine area and, with some adjusments in surveillance methods, has allowed the eradication of the disease from most of the 8 counties.