Ultrastructural localization of carbohydrate determinants of lipopolysaccharide of coxiella burnetii (Phase I) by monoclonal antibodies

1988 ◽  
Vol 46 ◽  
pp. 374-375
Author(s):  
T. F. Mccaul ◽  
J. C. Williams
Keyword(s):  
Monoclonal Antibodies ◽  
Phase I ◽  
Coxiella Burnetii ◽  
Antigenic Variation ◽  
Q Fever ◽  
Picric Acid ◽  
Gradient Centrifugation ◽  
Ultrastructural Localization ◽  
Large Cell ◽  
Developmental Cycle

Coxiella burnetii, the etiological agent of Q fever, is an obligate phagolysosomal bacterium and an unorthodox spore former. Morphologically distinct cell types, produced by a developmental cycle, are defined as large cell (LCV) and small cell variants (SCV). Recent study has shown phase I lipopolysaccharide (LPS-I) is differentially expressed among the cell variants. This antigenic variation was investigated further by use of a series of monoclonal antibodies (MAbs), directed against LPS-I, and post-embedding immunolabelling techniques. Since these MAbs recognize carbohydrate determinants of LPS-I, the specificity of the labelling was tested by carbohydrate reducing and oxidizing reagents prior to labelling.C. burnetii (9MIC7 strain), which was purified from infected yolk sac material of hen eggs by Renografin gradient centrifugation, was fixed for 3 h in 1.5% glutaraldehyde and 0.2% picric acid in 66mM Na-cacodylate buffer, pH = 6.8; pre-embedded in 2% Difco Nobel agar; and then rinsed once (15 min) in the same buffer.

scholarly journals Proteome and Antigen Profiling of Coxiella burnetii Developmental Forms

2006 ◽  
Vol 75 (1) ◽  
pp. 290-298 ◽  
Author(s):  
Sherry A. Coleman ◽  
Elizabeth R. Fischer ◽  
Diane C. Cockrell ◽  
Daniel E. Voth ◽  
Dale Howe ◽  
...  
Keyword(s):  
Coxiella Burnetii ◽  
Q Fever ◽  
Proteome Analysis ◽  
Large Cell ◽  
Human Infection ◽  
Developmental Cycle ◽  
Functional Roles ◽  
Immunogenic Proteins ◽  
Acute Q Fever ◽  
Insight Into

ABSTRACT A biphasic developmental cycle whereby highly resistant small-cell variants (SCVs) are generated from large-cell variants (LCVs) is considered fundamental to the virulence of Coxiella burnetii, the causative agent of human Q fever. In this study a proteome analysis of C. burnetii developmental forms was conducted to provide insight into their unique biological and immunological properties. Silver-stained gels of SCV and LCV lysates separated by two-dimensional (2-D) gel electrophoresis resolved over 675 proteins in both developmental forms. Forty-eight proteins were greater than twofold more abundant in LCVs than in SCVs, with six proteins greater than twofold more abundant in SCVs than in LCVs. Four and 15 upregulated proteins of SCVs and LCVs, respectively, were identified by mass spectrometry, and their predicted functional roles are consistent with a metabolically active LCV and a structurally resistant SCV. One-dimensional and 2-D immunoblots of cell form lysates probed with sera from infected/vaccinated guinea pigs and convalescent-phase serum from human patients who had recovered from acute Q fever, respectively, revealed both unique SCV/LCV antigens and common SCV/LCV antigens that were often differentially synthesized. Antigens recognized during human infection were identified by mass spectroscopy and included both previously described immunodominant proteins of C. burnetii and novel immunogenic proteins that may be important in the pathophysiology of clinical Q fever and/or the induction of protective immunity.

scholarly journals Contributions of lipopolysaccharide and the type IVB secretion system to Coxiella burnetii vaccine efficacy and reactogenicity

npj Vaccines ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Carrie M. Long ◽  
Paul A. Beare ◽  
Diane C. Cockrell ◽  
Jonathan Fintzi ◽  
Mahelat Tesfamariam ◽  
...  
Keyword(s):  
Phase I ◽  
Coxiella Burnetii ◽  
Vaccine Efficacy ◽  
Q Fever ◽  
Secretion System ◽  
Cell Vaccine ◽  
Post Vaccination ◽  
Whole Cell Vaccine ◽  
Type Ivb Secretion ◽  
Type Ivb Secretion System

AbstractCoxiella burnetii is the bacterial causative agent of the zoonosis Q fever. The current human Q fever vaccine, Q-VAX®, is a fixed, whole cell vaccine (WCV) licensed solely for use in Australia. C. burnetii WCV administration is associated with a dermal hypersensitivity reaction in people with pre-existing immunity to C. burnetii, limiting wider use. Consequently, a less reactogenic vaccine is needed. Here, we investigated contributions of the C. burnetii Dot/Icm type IVB secretion system (T4BSS) and lipopolysaccharide (LPS) in protection and reactogenicity of fixed WCVs. A 32.5 kb region containing 23 dot/icm genes was deleted in the virulent Nine Mile phase I (NMI) strain and the resulting mutant was evaluated in guinea pig models of C. burnetii infection, vaccination-challenge, and post-vaccination hypersensitivity. The NMI ∆dot/icm strain was avirulent, protective as a WCV against a robust C. burnetii challenge, and displayed potentially altered reactogenicity compared to NMI. Nine Mile phase II (NMII) strains of C. burnetii that produce rough LPS, were similarly tested. NMI was significantly more protective than NMII as a WCV; however, both vaccines exhibited similar reactogenicity. Collectively, our results indicate that, like phase I LPS, the T4BSS is required for full virulence by C. burnetii. Conversely, unlike phase I LPS, the T4BSS is not required for vaccine-induced protection. LPS length does not appear to contribute to reactogenicity while the T4BSS may contribute to this response. NMI ∆dot/icm represents an avirulent phase I strain with full vaccine efficacy, illustrating the potential of genetically modified C. burnetii as improved WCVs.

scholarly journals Animals withCoxiella burnetiiInfection Demonstrate a Western Immunoblot Profile of Chronic Q Fever in Humans

1996 ◽  
Vol 7 (1) ◽  
pp. 45-48
Author(s):  
TJ Marrie ◽  
Linda Yates
Keyword(s):  
Immune Response ◽  
Phase I ◽  
Phase Ii ◽  
Coxiella Burnetii ◽  
Q Fever ◽  
Western Immunoblotting ◽  
Western Immunoblot ◽  
Chronic Q Fever

Western immunoblotting was used to compare the immune response toCoxiella burnetiiphase I and phase II antigens of humans with acute and chronic Q fever with that of infected cats, rabbits, cows and raccoons. The cats, rabbits, cows and raccoons had an immunoblot profile similar to that of the human with chronic Q fever.

scholarly journals Characterization of a Stress-Induced Alternate Sigma Factor, RpoS, of Coxiella burnetii and Its Expression during the Development Cycle

2001 ◽  
Vol 69 (8) ◽  
pp. 4874-4883 ◽  
Author(s):  
Rekha Seshadri ◽  
James E. Samuel
Keyword(s):  
Stationary Phase ◽  
Coxiella Burnetii ◽  
Sigma Factor ◽  
Stationary Phases ◽  
Large Cell ◽  
Polyclonal Antiserum ◽  
Developmental Cycle ◽  
Growth Stages ◽  
Extracellular Environment ◽  
Sigma Factor Rpos

ABSTRACT Coxiella burnetii is an obligate intracellular bacterium that resides in an acidified phagolysosome and has a remarkable ability to persist in the extracellular environment.C. burnetii has evolved a developmental cycle that includes at least two morphologic forms, designated large cell variants (LCV) and small cell variants (SCV). Based on differential protein expression, distinct ultrastructures, and different metabolic activities, we speculated that LCV and SCV are similar to typical logarithmic- and stationary-phase growth stages. We hypothesized that the alternate sigma factor, RpoS, a global regulator of genes expressed under stationary-phase, starvation, and stress conditions in many bacteria, regulates differential expression in life cycle variants of C. burnetii. To test this hypothesis, we cloned and characterized the major sigma factor, encoded by an rpoD homologue, and the stress response sigma factor, encoded by an rpoS homologue. TherpoS gene was cloned by complementation of anEscherichia coli rpoS null mutant containing an RpoS-dependent lacZ fusion (osmY::lacZ). Expression ofC. burnetii rpoS was regulated by growth phase inE. coli (induced upon entry into stationary phase). A glutathione S-transferase–RpoS fusion protein was used to develop polyclonal antiserum against C. burnetii RpoS. Western blot analysis detected abundant RpoS in LCV but not in SCV. These results suggest that LCV and SCV are not comparable to logarithmic and stationary phases of growth and may represent a novel adaptation for survival in both the phagolysosome and the extracellular environment.

scholarly journals Truckin' pneumonia—an outbreak of Q fever in a truck repair plant probably due to aerosols from clothing contaminated by contact with newborn kittens

1989 ◽  
Vol 102 (1) ◽  
pp. 119-127 ◽  
Author(s):  
Thomas J. Marrie ◽  
Donald Langille ◽  
Vasilia Papukna ◽  
Linda Yates
Keyword(s):  
Phase I ◽  
Phase Ii ◽  
Coxiella Burnetii ◽  
Attack Rate ◽  
Q Fever ◽  
The Other ◽  
High Antibody ◽  
First Case ◽  
The Family ◽  
Antibody Titres

SUMMARYWe describe an outbreak of Q fever affecting 16 of 32 employees at a truck repair plant. None of the cases were exposed to cattle, sheep or goats. the traditional reservoirs of Q fever. The cases did not work, live on, or visit farms or attend livestock auctions. One of the employees had a cat which gave birth to kittens 2 weeks prior to the first case of Q fever in the plant. The cat owner fed the kittens every day before coming to work as the cat would not let the kittens suckle. Serum from the cat had high antibody titres to phase I and phase IICoxiella burnetiiantigens. The attack rate among the employees where the cat owner worked. 13 of 19 (68%), was higher than that of employees elsewhere, 3 of 13 (28%) [P <0·01]. The cat owner's wife and son also developed Q fever. None of the family members of the other employees with Q fever was so affected.We conclude that this outbreak of Q fever probably resulted from exposure to the contaminated clothing of the cat owner.

scholarly journals Phase Variation Analysis of Coxiella burnetii during Serial Passage in Cell Culture by Use of Monoclonal Antibodies

2002 ◽  
Vol 70 (8) ◽  
pp. 4747-4749 ◽  
Author(s):  
Akitoyo Hotta ◽  
Midori Kawamura ◽  
Ho To ◽  
Masako Andoh ◽  
Tsuyoshi Yamaguchi ◽  
...  
Keyword(s):  
Cell Culture ◽  
Monoclonal Antibodies ◽  
Phase I ◽  
Coxiella Burnetii ◽  
Phase Variation ◽  
Serial Passage ◽  
Variation Analysis ◽  
Group 3 ◽  
Group 2 ◽  
Group 1

ABSTRACT Antigenic changes in Coxiella burnetii Nine Mile strain phase I during serial passages in cell culture were analyzed with three groups of monoclonal antibodies (MAbs) against lipopolysaccharide. The MAbs of group 1 did not react with organisms that were passaged over five times, and the MAbs of group 2 did not react with organisms that were passaged over eight times. The MAbs of group 3 reacted with organisms passaged up to 15 times but did not react with phase II cells. These results suggest that C. burnetii could be differentiated into four phase states during phase variation.

scholarly journals Early Diagnosis and Treatment of Patients with Symptomatic Acute Q Fever Do Not Prohibit IgG Antibody Responses to Coxiella burnetii

2012 ◽  
Vol 19 (10) ◽  
pp. 1661-1666 ◽  
Author(s):  
C. C. H. Wielders ◽  
L. M. Kampschreur ◽  
P. M. Schneeberger ◽  
M. M. Jager ◽  
A. I. M. Hoepelman ◽  
...  
Keyword(s):  
Early Diagnosis ◽  
Phase I ◽  
Phase Ii ◽  
Coxiella Burnetii ◽  
Q Fever ◽  
Antibody Responses ◽  
Igg Antibody ◽  
Antibody Titers ◽  
Acute Q Fever

ABSTRACTLittle is known about the effect of timing of antibiotic treatment on development of IgG antibodies following acute Q fever. We studied IgG antibody responses in symptomatic patients diagnosed either before or during development of the serologic response toCoxiella burnetii. Between 15 and 31 May 2009, 186 patients presented with acute Q fever, of which 181 were included in this retrospective study: 91 early-diagnosed (ED) acute Q fever patients, defined as negative IgM phase II enzyme-linked immunosorbent assay (ELISA) and positive PCR, and 90 late-diagnosed (LD) acute Q fever patients, defined as positive/dubious IgM phase II ELISA and positive immunofluorescence assay (IFA). Follow-up serology at 3, 6, and 12 months was performed using IFA (IgG phase I and II). High IgG antibody titers were defined as IgG phase II titers of ≥1:1,024 together with IgG phase I titers of ≥1:256. At 12 months, 28.6% of ED patients and 19.5% of LD patients had high IgG antibody titers (P= 0.17). No statistically significant differences were found in frequencies of IgG phase I and IgG phase II antibody titers at all follow-up appointments for adequately and inadequately treated patients overall, as well as for ED and LD patients analyzed separately. Additionally, no significant difference was found in frequencies of high antibody titers and between early (treatment started within 7 days after seeking medical attention) and late timing of treatment. This study indicates that early diagnosis and antibiotic treatment of acute Q fever do not prohibit development of the IgG antibody response.

scholarly journals Seroprevalence of Q fever in small ruminants in the northeast Anatolian region in Turkey

2021 ◽  
Vol 77 (05) ◽  
pp. 6522-2021
Author(s):  
PERIHAN SERIFOĞLU BAGATIR ◽  
BIRAY OKUMUS ◽  
EDIZ KAAN OZGEN ◽  
MUSTAFA ULUCAN ◽  
BERNA YANMAZ ◽  
...  
Keyword(s):  
Phase I ◽  
Coxiella Burnetii ◽  
Indirect Elisa ◽  
Q Fever ◽  
Small Ruminants ◽  
Animal Husbandry ◽  
Fold Increase ◽  
Igg Antibodies ◽  
Serum Samples ◽  
Goat Population

The aim of our study was to determine the seroepidemiological profile of Q fever in small ruminants in Turkey and to examine its prevalence changes over the years. The study included 573 serum samples taken in 2013 and 472 samples taken in 2017 from animals in mixed herds of sheep and goats from 84 farms in Northeast Anatolia. Phase I and phase II IgG antibodies against Coxiella burnetii in serum samples were investigated by IDEXX ELISA (Q fever Ab Test IDEXX Laboratories, USA) indirect ELISA kits. Seroprevalence of Coxiella burnetii IgG in Artvin, Gümüşhane and Iğdır provinces was 5.6% in sheep, 1.8% in goats and 4.5% in total in 2013. In contrast, it was 24.4% in sheep, 1.1% in goats and 20.1% in total in 2017. According to the total seroprevalence rates calculated by including both sheep and goat population, it was seen that the province with the highest seroprevalence change in these animals was Iğdır with a 7.3-fold increase. Herd-level seroprevalence was 29.4% in 2013 and 57.6% in 2017. According to these results, the C. burnetii IgG seroprevalence nearly doubled after four years. This increase has been evaluated as a major risk for animal and human health as well as for the livestock economy in Northeastern Anatolia, where animal husbandry is intense.

scholarly journals Culture under normoxic conditions and enhanced virulence of phase II Coxiella burnetii transformed with a RSF1010-based shuttle vector

2019 ◽  
Author(s):  
Shengdong Luo ◽  
Zemin He ◽  
Zhihui Sun ◽  
Yonghui Yu ◽  
Yongqiang Jiang ◽  
...  
Keyword(s):  
Phase I ◽  
Phase Ii ◽  
Coxiella Burnetii ◽  
Q Fever ◽  
Shuttle Vector ◽  
Axenic Culture ◽  
Hypoxic Condition ◽  
Kanamycin Resistance ◽  
Temperature Sensitive ◽  
Egfp Gene

AbstractCoxiella burnetii is a Gram-negative, facultative intracellular microorganism that can cause acute or chronic Q fever in human. It was recognized as an obligate intracellular organism until the revolutionary design of an axenic cystine culture medium (ACCM). Present axenic culture of C. burnetii strictly requires a hypoxic condition (<10% oxygen). Here we investigated the normoxic growth of C. burnetii strains in ACCM-2 with or without tryptophan supplementation. Three C. burnetii strains - Henzerling phase I, Nine Mile phase II and a Nine Mile phase II transformant, were included. The transformant contains a pMMGK plasmid that is composed of a RSF1010 ori, a repABC operon, an eGFP gene and a kanamycin resistance cassette. We found that, under normoxia if staring from an appropriate concentration of fresh age inocula, Nine Mile phase II can grow significantly in ACCM-2 with tryptophan, while the transformant can grow robustly in ACCM-2 with or without tryptophan. In contrast, long-term frozen stocks of phase II and its transformant, and Henzerling phase I of different ages had no growth capability under normoxia under any circumstances. Furthermore, frozen stocks of the transformant consistently caused large splenomegaly in SCID mice, while wild type Nine Mile phase II induced a lesser extent of splenomegaly. Taken together, our data show that normoxic cultivation of phase II C. burnetii can be achieved under certain conditions. Our data suggests that tryptophan and an unknown temperature sensitive signal are involved in the expression of genes for normoxic growth regulated by quorum sensing in C. burnetii.

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