The Attenuated Nine Mile Phase II Clone 4/RSA439 Strain of Coxiella burnetii is Highly Virulent for Severe Combined Immunodeficient (SCID) Mice

Islam, Aminul; Lockhart, Michelle G.; Stenos, John; Graves, Stephen

doi:10.4269/ajtmh.12-0653

Cited by 13 publications

(11 citation statements)

References 23 publications

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“…SCID mice have been used to study virulence of intracellular bacterial pathogens including phase I C. burnetii (Andoh et al, 2007 ). It was also recently determined that a quantitative lethal dose 50 (LD 50 ) could be determined with NMII in SCID mice using longer experimental periods (Islam et al, 2013 ) The most physiologically relevant route of challenge would be aerosol, however, the disease progression after IT challenge with NMII in SCID mice was comparably slow and therefore not well-suited to large-scale screening. Furthermore, the increase in splenomegaly and GEs in organs was moderate, indicating a fundamental attenuation in dissemination.…”

Section: Discussionmentioning

confidence: 99%

The SCID Mouse Model for Identifying Virulence Determinants in Coxiella burnetii

Schaik

Case

Martínez

et al. 2017

Front. Cell. Infect. Microbiol.

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Coxiella burnetii is an intracellular, zoonotic pathogen that is the causative agent of Q fever. Infection most frequently occurs after inhalation of contaminated aerosols, which can lead to acute, self-limiting febrile illness or more serve chronic infections such as hepatitis or endocarditis. Macrophages are the principal target cells during infection where C. burnetii resides and replicates within a unique phagolysosome-like compartment, the Coxiella-containing vacuole (CCV). The first virulence determinant described as necessary for infection was full-length lipopolysaccarride (LPS); spontaneous rough mutants (phase II) arise after passage in immuno-incompetent hosts. Phase II C. burnetii are attenuated in immuno-competent animals, but are fully capable of infecting a variety of host cells in vitro. A clonal strain of the Nine Mile isolate (RSA439, clone 4), has a 26 KDa chromosomal deletion that includes LPS biosynthetic genes and is uniquely approved for use in BL2/ABL2 conditions. With the advances of axenic media and genetic tools for C. burnetii research, the characterization of novel virulence determinants is ongoing and almost exclusively performed using this attenuated clone. A major problem with predicting essential virulence loci with RSA439 is that, although some cell-autonomous phenotypes can be assessed in tissue culture, no animal model for assessing pathogenesis has been defined. Here we describe the use of SCID mice for predicting virulence factors of C. burnetii, in either independent or competitive infections. We propose that this model allows for the identification of mutations that are competent for intracellular replication in vitro, but attenuated for growth in vivo and predict essential innate immune responses modulated by the pathogen during infection as a central pathogenic strategy.

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Section: Discussionmentioning

confidence: 99%

The SCID Mouse Model for Identifying Virulence Determinants in Coxiella burnetii

Schaik

Case

Martínez

et al. 2017

Front. Cell. Infect. Microbiol.

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“…There are several animal models of virulent phase I C. burnetii infection available, including guinea pig, mouse and primate models; however, SCID mice are the only lethal animal model of C. burnetii NMII (Islam et al, 2013). C. burnetii NMII strains are used commonly by research laboratories at containment level 2 to characterize host-pathogen interactions in vitro and genetic manipulation has been achieved recently which allows specific virulence determinants to be evaluated (Voth and Heinzen, 2007;Beare et al, 2012).…”

Section: Discussionmentioning

confidence: 99%

Galleria mellonella as an alternative model of Coxiella burnetii infection

et al. 2014

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Coxiella burnetii is a Gram-negative intracellular bacterium and is the causative agent of the zoonotic disease Q fever. Several rodent and non-human primate models of virulent phase I C. burnetii [Nine Mile (NM)I] have been developed, and have been used to determine the efficacy of antibiotics and vaccine candidates. However, there are several advantages to using insect models to study hostmicrobe interactions, such as reduced animal use, lowered cost and ease of manipulation in high containment. In addition, many laboratories use the avirulent phase II C. burnetii clone (NMII) to study cellular interactions and identify novel virulence determinants using genetic manipulation. We report that larvae of the greater wax moth, Galleria mellonella, were susceptible to infection with both C. burnetii NMI and NMII. Following subcutaneous infection, we report that intracellular bacteria were present within haemocytes and that larval death occurred in a dose-dependent manner. Additionally, we have used the model to characterize the role of the type 4 secretion system in C. burnetii NMII and to determine antibiotic efficacy in a non-mammalian model of disease.

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“…Microscope slides were coated with phase 1 (Henzerling strain from the Australian Q vax vaccine) and phase 2 (Nine Mile) C. burnetii antigens grown in Vero cells as previously described (40). Fluorescein-labeled anti-goat IgG and anti-sheep IgM (KPL, USA) were used to detect IgG and IgM antibody-antigen complexes, respectively, as described previously (41).…”

Section: Methodsmentioning

confidence: 99%

Bayesian Validation of the Indirect Immunofluorescence Assay and Its Superiority to the Enzyme-Linked Immunosorbent Assay and the Complement Fixation Test for Detecting Antibodies against Coxiella burnetii in Goat Serum

Muleme

Stenos²,

Vincent³

et al. 2016

Clin Vaccine Immunol

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dAlthough many studies have reported the indirect immunofluorescence assay (IFA) to be more sensitive in detection of antibodies to Coxiella burnetii than the complement fixation test (CFT), the diagnostic sensitivity (DSe) and diagnostic specificity (DSp) of the assay have not been previously established for use in ruminants. This study aimed to validate the IFA by describing the optimization, selection of cutoff titers, repeatability, and reliability as well as the DSe and DSp of the assay. Bayesian latent class analysis was used to estimate diagnostic specifications in comparison with the CFT and the enzyme-linked immunosorbent assay (ELISA). The optimal cutoff dilution for screening for IgG and IgM antibodies in goat serum using the IFA was estimated to be 1:160. The IFA had good repeatability (>96.9% for IgG, >78. Coxiella burnetii causes Q fever in humans as well as abortions, stillbirths, and infertility in ruminants (1-5). The organism replicates in the placenta of infected ruminants, reaching a level of up to 10 9 bacteria per gram of placenta tissue (4-6). C. burnetii organisms are shed in an extremely high concentration in birth fluids, placental tissues, and membranes of aborted fetuses as well as in milk, urine, and feces of infected ruminant animals around the parturition period (4, 5). The high concentration of C. burnetii organisms shed in tissues, fluids, and excreta of infected ruminants is the primary source of human infections (7).Caprine and ovine infections have been reported to result in severe placentitis and consequently in shedding of higher numbers of C. burnetii organisms than infections of cattle (8). Studies have also revealed that goats and sheep shed higher quantities of C. burnetii in feces, vaginal mucus, and birth tissues than other livestock (9). Thus, the risk of human transmission is higher when infections occur in herds of small ruminants than when they occur with other livestock. Unsurprisingly, the majority of reported large outbreaks of Q fever have been associated with infected sheep and goat flocks, including a major outbreak of more than 4,000 human Q fever cases in the Netherlands that was linked to sheep and goat farms with over 50 animals (9)(10)(11)(12)(13)(14). Infection with C. burnetii can be asymptomatic in many animals and may be detected in ruminants only when infection causes abortions and reproductive abnormalities in pregnant animals (1). Delay in diagnosis in livestock slows the implementation of appropriate control strategies, thus increasing the risk of human infection.Coxiellosis in animals can be diagnosed through microscopic examination of stained tissues, culture, detection of C. burnetii DNA using PCR, and detection of antibodies to C. burnetii in blood and milk (15,16). The microscopic diagnosis of coxiellosis is mainly undertaken on placental tissues using Stamp-Macchiavello coloration or Giemsa stain. The organism can be cultured in cells, embryonated hen eggs, or cell-free media (15,17). However, microscopy and culture are expensive and requi...

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The Attenuated Nine Mile Phase II Clone 4/RSA439 Strain of Coxiella burnetii is Highly Virulent for Severe Combined Immunodeficient (SCID) Mice

Cited by 13 publications

References 23 publications

The SCID Mouse Model for Identifying Virulence Determinants in Coxiella burnetii

The SCID Mouse Model for Identifying Virulence Determinants in Coxiella burnetii

Galleria mellonella as an alternative model of Coxiella burnetii infection

Bayesian Validation of the Indirect Immunofluorescence Assay and Its Superiority to the Enzyme-Linked Immunosorbent Assay and the Complement Fixation Test for Detecting Antibodies against Coxiella burnetii in Goat Serum

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