A novel method for estimating killing ability and digestion of<i>Paracoccidioides brasiliensis</i>by phagocytic cells<i>in vitro</i>

Goihman-Yahr, Mauricio; Rothenberg, A.; Rosquete, R.; Avila-Millán, Enrique; Albornoz, María B. de; Gómez, María Helena; Martín, Blanca San; Ocanto, Ana; Pereira, José Renato; Molina, Tulio

doi:10.1080/00362178585380371

Cited by 14 publications

(7 citation statements)

References 6 publications

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“…A later experiment showed that the yeast form of P. brasiliensis produced higher levels of iron chelants in media containing low concentrations of iron (11). The production of germ tubes in a slide culture is also a dependable procedure for determining the viability of P. brasiliensis yeast form cells (2,127,249).…”

Section: Microbiologymentioning

confidence: 99%

Paracoccidioidomycosis: an update

1993

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This review summarizes knowledge on various aspects of paracoccidioidomycosis. Mycelial propagules, chlamydospores, and arthroconidia exhibit thermal dimorphism; arthroconidia are infectious in animals and, by electron microscopy, appear well provided for survival. The mycelial-to-yeast-phase transformation requires a strict control of glucan synthesis probably mediated by membrane enzymes. Hormonal influences on the transformation of the fungus (mycelium or conidium to yeast phase) have been demonstrated. Estrogen-binding proteins have been detected in the fungal cytosol, and during the transformation novel proteins are produced as a result of estradiol incorporation. Clinical forms have been better defined on the basis of better experimental models. Emphasis has been placed on the lungs as the portal of entry and on the existence of silent pulmonary infections. A specific Paracoccidioides brasiliensis antigen, the 43-kDa glycoprotein (Gp43), has been identified, characterized, and cloned. This has led to improved reproducibility and specificity of serologic tests. The depression of cell-mediated immune responses has been associated with severe disease in humans and in the experimental host. T-cell subsets in patients' tissues were characterized by means of monoclonal antibodies, and a reduced CD4/CD8 ratio was demonstrated. This has been related to alterations in lymphokine and tumor necrosis factor production, production of antigen-antibody complexes, etc. Amphotericin B has provided effective therapy. Azole derivatives have also improved prognosis and facilitated therapy. Itraconazole is presently the drug of choice, yet incapacitating sequelae (mainly pulmonary fibrosis) still constitute major problems.

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Section: Microbiologymentioning

confidence: 99%

Paracoccidioidomycosis: an update

1993

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“…We also studied the role of complement in mediating PMN CL in response to FI. PMN cells seem to play an important role in host defense against P. brasiliensis [ 18,24,34]. In addition to P. brasiliensis, several other infectious agents are able to stimulate the PMN oxidative metabolism leading to oxygen toxic metabolite release and consequently to CL production [9,19,20,27,40].…”

Section: Discussionmentioning

confidence: 99%

“…A similar inflammatory response was also observed when animals were injected with isolated components of the fungus cell wall such as lipids and polysaccharides [36,37], suggesting that these substances may play an important role Since some peptides derived from complement activation are known to be important chemoattractants for PMN, the present study was designed to investigate the involvement of the classical and alternative complement pathways in the in vitro PMN chemotaxis induced by the F1 fraction of P. brasiliensis. Moreover, since the fungicidal activity of PMN in this infection is still controversial [17,18,24,34], we also looked for the participation of the complement system and opsonins of other origins in the stimulation of rat PMN oxidative metabolism by the F1 fraction.…”

Section: (Accepted 27 August 1992)mentioning

confidence: 99%

The role of the complement system in the neutrophil functions stimulatedin vitroby an alkali-insoluble cell wall fraction ofParacoccidioides brasiliensis

Crott¹,

Valim

Silva

et al. 1993

Med Mycol

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We investigated the capacity of an alkali-insoluble cell wall polysaccharide fraction (FI) ofParacoeeidioides brasiliensis to induce rat polymorphonuclear neutrophil (PMN) migratory and chemiluminescence (CL) responses. Normal rat serum pre-incubated with F1 induced a chemotactic neutrophil response which was fully abolished by heat-inactivation. The participation of the alternative complement pathway was more effective than that of the classical pathway since depletion of factor B by heating at 50°C reduced PMN migration, whereas blockade of the classical pathway with EGTA left the migratory response practically unchanged. Opsonized serum F 1 induced a significant release of oxygen radicals from PMN as measured by CL. The complement system was also found to be involved in this activity since serum inactivation at 56°C altered the CL response. In addition to complementderived fragments, other serum opsonins, probably cross-reacting antibodies, were required for optimal interaction between PM N and opsonized particles. These results contribute to the understanding of the role of fungal components and of the complement system in the inflammatory response observed in paracoccidioidomycosis.Paracoccidioidomycosis or South American blastomycosis is a deep mycosis caused by the dimorphic fungus Paracoccidioides brasiliensis and is the most widespread systemic mycosis in South America [23]. The role of phagocytic cells in the host defense against this fungus is still not well understood. The importance of polymorphonuclear neutrophils (PMN) has been suggested by the finding of a massive infiltration of these cells at the inflammatory sites during the initial stage of the disease in man [12] as well as in experimentally infected animals [3,37]. The presence of PMN has also been detected in the suppurative region of the granulomatous lesions characteristic of the chronic phase of this mycosis [4,11]. A similar inflammatory response was also observed when animals were injected with isolated components of the fungus cell wall such as lipids and polysaccharides [36,37], suggesting that these substances may play an important role in the virulence and pathogenicity of P. brasiliensis. One of these components, the alkali-insoluble polysaccharide fraction (F1),was able to induce accumulation of PMN during the early phase of the inflammatory response in experimental models [3,11]. Furthermore, the F 1 fraction was shown to be able to activate the classical and alternative complement pathways of rats both in vivo and in vitro (L. S. P. Crott, R. T. P. Sobreira, C. L. Silva & J. E. Barbosa, unpublished results).

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“…An ultrasonic processorsonicator (Heat Systems-Ultrasonic, Farmingdale, NY, USA) model W-385 was used. Sonication was carried out at a low intensity (amplitude #1 of the apparatus' output control; energy: 40% output0/0.4 )/ 475 W available 0/190 W delivered) with continuous energy for exactly 20 s according to the method described by Goihman-Yahr et al [5] and used in several other assays involving experiments with P. brasiliensis [1,5 Á/ 14]. All preparations were then washed and the fungal cells were prepared for biological and morphological observations as described below.…”

Section: Sonicationmentioning

confidence: 99%

“…It is well established that when maintained in axenic cultures P. brasiliensis grows forming large clumps due to the fact that multiple budding takes place and the new-formed cells remain attached to the mother cells [5], thus rendering it difficult to obtain free cells. This fact makes it difficult to carry out basic quantitative studies on the interactions of the fungus with host cells.…”

Section: Introductionmentioning

confidence: 99%

Viability of yeast form cells ofParacoccidioides brasiliensisafter sonication

et al. 2004

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To perform in-vitro studies with Paracoccidioides brasiliensis yeast cells it is necessary to avoid the presence of clumps of cells while maintaining their integrity. Because of the multiple budding type of growth, the bud cells are always attached to the mother cell and the yeast cells keep growing, resulting in the formation of large clumps. In order to obtain free cells, the cultures are usually sonicated. The present study shows that sonication induces lesions in a significant number of cells, as evaluated by labelling of the cells with acridine orange and Janus green vital dyes. In some cases labelling was initially observed in only one cell of the clump; however, the other cells also became labelled after a few minutes. These observations were confirmed by scanning and transmission electron microscopy of treated cells. Colony forming units (c.f.u.) on BHI plates also confirmed the decrease in cell viability following sonication.

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A novel method for estimating killing ability and digestion ofParacoccidioides brasiliensisby phagocytic cellsin vitro

Cited by 14 publications

References 6 publications

Paracoccidioidomycosis: an update

Paracoccidioidomycosis: an update

The role of the complement system in the neutrophil functions stimulatedin vitroby an alkali-insoluble cell wall fraction ofParacoccidioides brasiliensis

Viability of yeast form cells ofParacoccidioides brasiliensisafter sonication

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