Host-Parasite Relationships in Brucellosis: II. Destruction of Macrophage Cultures by Brucella of Different Virulence

Freeman, Bob A.; Kross, Damon J.; Circo, Richard

doi:10.1093/infdis/108.3.333

Cited by 26 publications

(35 citation statements)

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“…In addition to recent studies suggesting that apoptotic cell death may be blocked by O antigen but continues unabated in cells infected with rough mutants (17), the cell death induced by the rough mutants described here appeared to be necrotic based on early LDH release, annexin V and PI staining, morphological appearance of infected cells, lack of nuclear condensation, and glycine protection. In agreement with the work of Freeman et al (19)(20)(21), the CPE observed was restricted to live bacteria, as it was eliminated by both heat treatment and chloramphenicol pretreatment of the B. abortus (data not shown). Furthermore, these data are in accordance with the work by Freeman and coworkers (19)(20)(21), and more recently by others (24,40,49), describing the ability of rough organisms to replicate within macrophages and/or monocytes.…”

Section: Discussionsupporting

confidence: 92%

“…In agreement with the work of Freeman et al (19)(20)(21), the CPE observed was restricted to live bacteria, as it was eliminated by both heat treatment and chloramphenicol pretreatment of the B. abortus (data not shown). Furthermore, these data are in accordance with the work by Freeman and coworkers (19)(20)(21), and more recently by others (24,40,49), describing the ability of rough organisms to replicate within macrophages and/or monocytes.…”

Section: Discussionsupporting

confidence: 92%

“…Although Brucella invasion, intracellular trafficking, and growth in professional phagocytes and epithelial cells have been widely investigated (2, 12-14, 25, 39), only antiapoptotic effects of Brucella infection on phagocytes have been reported (17,22,27). The cytopathic effects (CPE) of Brucella infection have been overlooked (19)(20)(21).…”

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confidence: 99%

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Brucella abortusRough Mutants Are Cytopathic for Macrophages in Culture

Pei¹,

Ficht²

2004

Infect Immun

148

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Rough mutants of Brucella spp. are attenuated for survival in animal models. However, conflicting in vitro evidence has been obtained concerning the intracellular survival of rough mutants. Transposon-derived rough mutants isolated in our laboratory were previously shown to exhibit small but significant reductions in intracellular survival in a 12-h in vitro assay. Several recent publications report that rough mutants exhibited increased macrophage uptake relative to their smooth parental strains, and a reduction in numbers at the end of the assay has been interpreted as intracellular killing. In an effort to explore the role of O antigen in the interaction between Brucella abortus and macrophages, we have monitored the uptake of rough mutants and survival in vitro by using the murine macrophage cell line J774.A1. The results confirm a 10-to 20-foldincreased uptake of rough mutants over that of smooth organisms under standard conditions. Recovery of the rough mutants persisted up to 8 h postinfection, but at the point when intracellular replication of the smooth organisms was observed, the number of rough organisms recovered declined. Fluorescence microscopy revealed the intracellular multiplication of both smooth and rough organisms, and assays performed in the absence of antibiotic confirmed the replication of the rough organisms. Examination by phase-contrast microscopy revealed the lytic death of macrophages infected with the rough mutants, which was confirmed by the release of lactate dehydrogenase (LDH) from the cell cytoplasm. Thus, the decline in the number of rough organisms was the result of the lysis of macrophages and not from intracellular killing. The cytopathic effect is characterized as necrotic rather than apoptotic cell death based on early LDH release, annexin V and propidium iodide staining, morphological changes of infected cells and nuclei, and glycine protection. The cytopathic effect was observed with macrophages at multiplicities of infection (MOIs) of as low as 20 and was not observed with epithelial cells at MOIs of as high as 2000. These findings suggest a role for O antigen during the early stages of host-agent interaction that is essential in establishing an intracellular niche that maintains and supports persistent intracellular infection resulting in disease.Brucella spp. are facultative intracellular bacteria that cause brucellosis in a variety of animals and undulant fever in humans (3). Brucellosis is a worldwide zoonosis characterized by persistence of the organism in the reticuloendothelial system in secondary hosts and in the reproductive system in primary hosts. The ability of these organisms to survive in professional and nonprofessional phagocytic cells is the basis for disease (18). The gene products and mechanisms that are essential for the intracellular lifestyle are currently under investigation. Among these, the O antigen has been classified as a major virulence determinant of Brucella and is essential for survival of the classical species in the host. Although it is...

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Section: Discussionsupporting

confidence: 92%

Section: Discussionsupporting

confidence: 92%

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Brucella abortusRough Mutants Are Cytopathic for Macrophages in Culture

Pei¹,

Ficht²

2004

Infect Immun

148

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“…Longer times were not feasible because in these experiments the monocytes infected with brucellas, strain SA-R, underwent degeneration much more rapidly than parallel cultures infected with the same inocula of strain SA-S. These observations remain unexplained, but recall similar findings reported by Freeman, Kross & Circo (1961).…”

Section: Brucella Abortus Strain Sa-rsupporting

confidence: 57%

Electron Microscope Observations of Brucella abortus Grown within Monocytes in vitro

Karlsbad

Kessel

Petris

et al. 1964

Journal of General Microbiology

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SUMMARYCells of Brucella abortus, strain S A -S , grown within normal guinea-pig monocytes maintained in vitro, were examined by electron microscopy. Intracellular multiplication of brucellae was followed for 48 hr. During the entire period of observation the bacteria were present in the cytoplasm of the monocytes, localized within vacuoles limited by a 'unit membrane'.No obvious ultrastructural differences (particularly in surface structures) were detected between monocyte-grown bacteria and sister cells cultivated on a lifeless medium, which might account for the known differences between the two (e.g. differences in resistance to serum bactericidal factors and in intracellular growth patterns).In spite of the large numbers of intracellular parasites present the host cells did not show evident structural modifications.Some observations made on intracellular B. abortus, strain SA-R, showed that after up to 24 hr within guinea-pig monocytes most of the bacteria had retained their morphological integrity and were indistinguishable from R organisms grown on agar,

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“…The virulence of Brucella relies on its ability to survive and replicate in the vacuolar compartments of macrophages (18). In contrast, many rough derivatives of B. suis, B. abortus, and B. melitensis, which are deficient in the O antigen (or O side chain) associated with lipopolysaccharide (LPS), cannot survive inside macrophages and hence are attenuated (11,33).While smooth virulent Brucella strains inhibit programmed macrophage cell death (17, 18), many rough Brucella strains are cytotoxic to macrophages and induce macrophage cell death (7,11,14,30,32). Freeman et al first reported that rough Brucella induced cytopathic cell death (14).…”

mentioning

confidence: 99%

Proinflammatory Caspase-2-Mediated Macrophage Cell Death Induced by a Rough Attenuated Brucella suis Strain

Chen

Ding

et al. 2011

Infect Immun

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Brucella spp. are intracellular bacteria that cause an infectious disease called brucellosis in humans and many domestic and wildlife animals. B. suis primarily infects pigs and is pathogenic to humans. The macrophage-Brucella interaction is critical for the establishment of a chronic Brucella infection. Our studies showed that smooth virulent B. suis strain 1330 (S1330) prevented programmed cell death of infected macrophages and rough attenuated B. suis strain VTRS1 (a vaccine candidate) induced strong macrophage cell death. To further investigate the mechanism of VTRS1-induced macrophage cell death, microarrays were used to analyze temporal transcriptional responses of murine macrophage-like J774.A1 cells infected with S1330 or VTRS1. In total 17,685 probe sets were significantly regulated based on the effects of strain, time and their interactions. A miniTUBA dynamic Bayesian network analysis predicted that VTRS1-induced macrophage cell death was mediated by a proinflammatory gene (the tumor necrosis factor alpha [TNF-␣] gene), an NF-B pathway gene (the IB-␣ gene), the caspase-2 gene, and several other genes. VTRS1 induced significantly higher levels of transcription of 40 proinflammatory genes than S1330. A Mann-Whitney U test confirmed the proinflammatory response in VTRS1-infected macrophages. Increased production of TNF-␣ and interleukin 1␤ (IL-1␤) were also detected in the supernatants in VTRS1-infected macrophage cell culture. Hyperphosphorylation of IB-␣ was observed in macrophages infected with VTRS1 but not S1330. The important roles of TNF-␣ and IB-␣ in VTRS1-induced macrophage cell death were further confirmed by individual inhibition studies. VTRS1-induced macrophage cell death was significantly inhibited by a caspase-2 inhibitor but not a caspase-1 inhibitor. The role of caspase-2 in regulating the programmed cell death of VTRS1-infected macrophages was confirmed in another study using caspase-2-knockout mice. In summary, VTRS1 induces a proinflammatory, caspase-2-and NF-B-mediated macrophage cell death. This unique cell death differs from apoptosis, which is not proinflammatory. It is also different from classical pyroptosis, which is caspase-1 mediated.Brucella spp. are facultative, intracellular, Gram-negative bacteria that cause a zoonotic disease called brucellosis in swine, cattle, other animals, and humans. Human brucellosis remains the most common zoonotic disease worldwide, with more than 500,000 new cases reported annually (28). B. suis is the main causative agent of brucellosis in swine and humans (28). Brucella enters and replicates efficiently in a variety of cells, including macrophages, epithelial cells, dendritic cells, and trophoblasts (34). Interaction between Brucella and macrophages is critical for the establishment of a chronic infection. The virulence of Brucella relies on its ability to survive and replicate in the vacuolar compartments of macrophages (18). In contrast, many rough derivatives of B. suis, B. abortus, and B. melitensis, which are deficient in the O antigen (o...

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Host-Parasite Relationships in Brucellosis: II. Destruction of Macrophage Cultures by Brucella of Different Virulence

Cited by 26 publications

References 0 publications

Brucella abortusRough Mutants Are Cytopathic for Macrophages in Culture

Brucella abortusRough Mutants Are Cytopathic for Macrophages in Culture

Electron Microscope Observations of Brucella abortus Grown within Monocytes in vitro

Proinflammatory Caspase-2-Mediated Macrophage Cell Death Induced by a Rough Attenuated Brucella suis Strain

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