Efficient isolation and propagation of human immunodeficiency virus on recombinant colony-stimulating factor 1-treated monocytes.

Monocytes were maintained in tissue culture for greater than 3 mo in media supplemented with rCSF-1. These cultures provided susceptible target cells for isolation and propagation of virus from PBMC of HIV-infected patients. HIV isolated into monocytes readily infected other rCSF-1-treated monocytes but only inefficiently infected PHA-stimulated lymphoblasts. Similarly, laboratory HIV strains passaged in T cell lines or virus isolated from patients' leukocytes into PHA-stimulated lymphoblasts inefficiently infected rCSF-1-treated monocytes. Persistent, low-level virion production was detected in macrophage culture fluids by reverse transcriptase activity or HIV antigen capture through 6-7 wk. Marked changes in cell morphology with cell death, syncytia, and giant cell formation were observed in monocyte cultures 2 wk after infection, but at 4-6 wk, all cells appeared morphologically normal. However, the frequency of infected cells in these cultures at 6 wk was 60-90% as quantified by in situ hybridization with HIV RNA probes or by immunofluorescence with AIDS patients' sera. Ultrastructural analysis by EM also showed a high frequency of infected cells; virtually all HIV budded into and accumulated within cytoplasmic vacuoles and virus particles were only infrequently associated with the plasma membrane. Retention of virus within macrophages and the macrophage tropism of HIV variants may explain mechanisms of both virus persistence and dissemination during disease.

[Fungal keratitis: A 5-year monocentric retrospective study on Reunion Island]. / Kératites fongiques : étude rétrospective monocentrique sur 5ans à l'île de la Réunion.

OBJECTIVES: Fungal keratitis is rare in France, but could be a severe sight-threatening condition. Here, we aimed to describe the epidemiology of fungal keratitis in Réunion Island. METHODS: In a retrospective study, we analyzed 13 culture-proven keratitis episodes, occurred between January 2013 and July 2017 in the ophthalmology ward of a University Hospital, Saint-Pierre. Twelve isolates were genotyped and antifungal susceptibility testing was performed. RESULTS: Corneal abrasion caused by vegetable matter was the main predisposing factor. Stromal infiltration was observed in 12 patients. Six patients did not response to medical treatment, requiring surgical care, including two enucleations surgery. Fusarium solani (n = 6) and Fusarium dimerum (n = 4) were the main fungal species involved in fungal keratitis. Clinical failures were more prevalent with F. solani infections. The lowest minimal inhibitory concentrations for Fusarium sp. were observed with voriconazole and amphotericin B. CONCLUSION: In Reunion Island, the epidemiology of fungal keratitis is characterized by the predominance of Fusarium species, potentially involved in visual loss. This pattern is consistent with the epidemiology usually observed in tropical areas.

Effects of ibuterol, a beta-2 adrenergic prodrug, on intraocular pressure.

Ibuterol, a prodrug of terbutaline, was approximately 100 times more potent than terbutaline in producing ocular hypotension in the treated eye of normal and sympathectomized (SX) rabbits. In normal rabbits, ibuterol produced no change in IOP of the contralateral eye whereas in unilaterally SX rabbits a rise in IOP occurred in the SX (contralateral) eye when the normal eye was treated with ibuterol. Ibuterol also suppressed ocular hypertension induced by water loading and delayed the IOP recovery rate although its onset of action was delayed. Aqueous flow was increased significantly at 1 hr after ibuterol in fluorophotometric studies in normal rabbits. Pretreatment with forskolin antagonized rather than enhanced ibuterol-induced ocular hypotension. Pretreatment with diclofenac failed to suppress the development of tachyphylaxis to the ocular hypotensive effect of ibuterol. Although ibuterol is an effective ocular hypotensive agent in rabbits, the effects of this agent on aqueous flow are complex and tachyphylaxis to the ocular hypotensive effect develops fairly rapidly.

Neuroendocrine carcinoma of the skin (Merkel cell carcinoma) in a black.

Neuroendocrine carcinoma of the skin is an uncommon, small-cell neoplasm most commonly found on white, sun-exposed skin. Diagnosis by clinical and histologic means may be difficult. Immunohistochemical and ultrastructural analysis are often required. Because of the aggressive nature of neuroendocrine carcinoma of the skin, prompt diagnosis and treatment are essential. We present the rare occurrence of a neuroendocrine carcinoma of the skin on sun protected skin in a black. Clinical, histologic, immunohistochemical, and ultrastructural features are reviewed, and therapeutic options are discussed.

Enzyme-linked immunosorbent assay for Pseudomonas aeruginosa exotoxin A.

An enzyme-linked immunosorbent assay (ELISA) is described for Pseudomonas aeruginosa exotoxin A. A double antibody sandwich method was used, employing polyvinyl microtiter plates as the solid phase, a primary coat of monospecific rabbit antitoxin serum, an outer layer composed of a horseradish peroxidase-sheep antitoxin immunoglobulin G conjugate, and an ortho-phenylene-diamine substrate. Absorbance (optical density) of hydrolyzed end product was read spectrophotometrically at 492 nm. ELISA detected as little as 30 pg (0.3 ng/ml) of purified toxin, and absorbance was linear over a 20-fold or greater concentration range. Toxin was demonstrated in culture filtrates from 42 of 48 (88%) consecutive clinical P. aeruginosa isolates compared with 37 of 48 (77%) positive by hemagglutination inhibition. Results of the two assays correlated closely (r = 0.82, P less than 0.001). Specificity was confirmed by neutralizability of ELISA activity with monospecific antitoxin. ELISA was thus a sensitive, specific, and quantifiable technique for the assay of P. aeruginosa exotoxin A in both purified and crude culture materials.

Cytoplasmic assembly and accumulation of human immunodeficiency virus types 1 and 2 in recombinant human colony-stimulating factor-1-treated human monocytes: an ultrastructural study.

Recombinant human colony-stimulating factor-1-treated human peripheral blood-derived monocytes-macrophages are efficient host cells for recovery of the human immunodeficiency virus (HIV) from blood leukocytes of patients with acquired immunodeficiency syndrome. These cells can be maintained as viable monolayers for intervals exceeding 3 months. Infection with HIV resulted in virus-induced cytopathic effects, accompanied by relatively high levels of released progeny virus, followed by a prolonged low-level release of virus from morphologically normal cells. In both acutely and chronically infected monocytes, viral particles were seen budding into and accumulating within cytoplasmic vacuoles. The number of intravacuolar virions far exceeded those associated with the plasma membrane, especially in the chronic phase, and were concentrated in the perinuclear Golgi zone. In many instances, the vacuoles were identified as Golgi elements. Fusion of virus-laden vacuoles with primary lysosomes were rare. The pattern of cytoplasmic assembly of virus was observed with both HIV types 1 and 2 and in brain macrophages of an individual with acquired immunodeficiency syndrome encephalopathy. Immunoglobulin-coated gold beads added to acutely infected cultures were segregated from the vacuoles containing virus; relatively few beads and viral particles colocalized. The assembly of HIV virions within vacuoles of macrophages is in contrast to the exclusive surface assembly of HIV by T lymphocytes. Intracytoplasmic virus hidden from immune surveillance in monocytes-macrophages may explain, in part, the persistence of HIV in the infected human host.

Survey of H2-antagonist usage in acute upper gastrointestinal hemorrhage.

H2-antagonists are frequently used in the management of upper gastrointestinal (UGI) hemorrhage despite their lack of proven efficacy. In order to determine the pattern of H2-antagonist usage for this indication, we retrospectively reviewed the charts of 137 patients admitted with acute UGI bleeding over a 1-year period at two teaching hospitals in West Texas. An H2-antagonist was ordered in 89% of patients (77%) intravenous, 12% oral). It was administered within 2 h of admission in 25% of these patients, within 4 h in 54%, and within 8 h in 78%. An H2-antagonist was ordered among the initial six orders in 49% and among the initial 10 orders in 77% of patients. Considering orders for specific therapies, an H2-antagonist was in the initial three orders in 60% of patients and among the initial six orders in 97%. Of the patients who were prescribed an H2-antagonist and who also had upper endoscopy, the drug was ordered prior to endoscopy in 86%. This review of H2-antagonist usage in the management of acute UGI bleeding has identified a prescribing pattern of writing for these drugs early in the sequence of order writing, with the drugs being given early in the course of hospitalization.

Protection of macaques with a simian immunodeficiency virus envelope peptide vaccine based on conserved human immunodeficiency virus type 1 sequences.

This report describes the vaccination of rhesus macaques with peptides selected from regions of the simian immunodeficiency virus (SIV) envelope that are hydrophilic, immunoreactive, and highly homologous with corresponding conserved envelope sequences of the human immunodeficiency virus (HIV). The peptides, produced as beta-galactosidase fusion proteins, induced virus-neutralizing and peptide-specific antibodies. After challenge with virulent virus, controls became virus positive and developed gradually rising antibody titers to SIV over 63 weeks. Immunized macaques developed a postchallenge anamnestic response to SIVenv antigens within 3-6 weeks followed by a gradual, fluctuating decline in SIV antibody titers and partial or total suppression of detectable SIV. Virus suppression correlated with prechallenge neutralizing antibody titers. Although the average CD4+ cell count in the blood of immunized macaques remained constant, the control macaques exhibited a progressive decrease developing about week 55 after challenge. The conserved nature of the HIV and SIV peptides and the similar humoral immunoreactivity in the respective hosts suggest that homologous HIV peptides may be important components of a successful immunization strategy.