A comparison of virus isolation, indirect immunofluorescence and nested multiplex polymerase chain reaction for the diagnosis of primary and recurrent herpes simplex type 1 and type 2 infections

134 swabs in viral transport medium were received from 126 patients with suspected clinical HSV-1 and HSV-2 infections. They were tested by (i) nested multiplex polymerase chain reaction NMPCR (strongly positive specimens had visible bands on both rounds of PCR) without prior extraction, (ii) culture in primary rhesus monkey kidney, E6-Vero, RD and HEp-2 cells and (iii) antigen detection by immunofluorescence (IF). Antigen detection employed four novel pools (A-D) of monoclonal antibodies (Mab): A was HSV-1 specific, B was HSV-2 specific while C and D were generic. In comparison to NMPCR the sensitivity and specificity of (i) culture was 59% (22/37) and 100% (134/134), (ii) IF by Pool A was 59% (16/27) and 100% (117/117), (iii) IF by Pool B was 40% (4/10) and 100% (130/130) and (iv) IF by Pools C and D were 60% (18/30) and 100% (96/96). Specimens positive by culture were more likely to be strongly positive by NMPCR (chi2 P = 0.004). Typing by each method concurred on all occasions. NMPCR was cost effective, easier to perform and was the most sensitive method for HSV detection. It should become the method of choice for HSV diagnosis.     

Fluorescent amplified fragment length polymorphism genotyping of Campylobacter jejuni and Campylobacter coli strains and its relationship with host specificity, serotyping, and phage typing

Fluorescent amplified fragment length polymorphism (FAFLP) analysis was applied to 276 Campylobacter jejuni strains and 87 Campylobacter coli strains isolated from humans, pigs, cattle, poultry, and retail meats to investigate whether certain FAFLP genotypes of C. jejuni and C. coli are associated with a particular host and to determine the degree of association between FAFLP-defined genotypes and heat-stable serotypes and/or phage types. Within C. coli, the poultry strains clustered separately from those of porcine origin. In contrast, no evidence of host specificity was detected among C. jejuni strains. While C. coli strains show host specificity by FAFLP genotyping, C. jejuni strains that are genotypically similar appear to colonize a range of hosts, rather than being host adapted. Some serotypes and/or phage types (C. jejuni serotype HS18, phage type PT6, and serophage type HS19/PT2 and C. coli HS66, PT2, and HS56/PT2) were the most homogeneous by FAFLP genotyping, while others were more heterogeneous (C. jejuni HS5 and PT39, and C. coli HS24 and PT44) and therefore poor indicators of genetic relatedness between strains. The lack of host specificity in C. jejuni suggests that tracing the source of infection during epidemiological investigations will continue to be difficult. The lack of congruence between some serotypes and/or phage types and FAFLP genotype underlines the need for phenotypic testing to be supplemented by genotyping. This study also demonstrates how, in general, FAFLP generates "anonymous" genetic markers for strain characterization and epidemiological investigation of Campylobacter in the food chain.     

Correlation between macrophage activation and bactericidal function and Mycobacterium leprae antigen presentation in macrophages of leprosy patients and normal individuals.

The killing of Mycobacterium leprae by resting and gamma interferon (IFN-gamma)-activated macrophages in normal subjects and leprosy patients was assessed. Resting macrophages from normal individuals demonstrated the ability to kill M. leprae. For macrophages from tuberculoid patients, killing of M. leprae was only achieved in the presence of IFN-gamma, suggesting that initial T-cell activation occurs prior to the killing of M. leprae. In contrast, though activation with IFN-gamma rendered the lepromatous macrophages microbicidal, it failed to induce lymphocyte proliferation, suggesting a defect at either the antigen-presenting cell or the lymphocyte level or both. The concept that T-cell anergy is primarily due to lack of lymphokine generation was ruled out by our results, since responsiveness was restored in only a small proportion of lepromatous patients after exogenous lymphokine addition. In conclusion, this study demonstrated that killing and antigen presentation are two independent events. It appears that the ability of the macrophages per se to kill M. leprae may be of greater importance than lymphocyte-mediated activation for protection against M. leprae infection.     

Lymphoblastic crisis of chronic myelogenous leukemia. Hand mirror variant

We present, to our knowledge, the first extensively studied case of lymphoid L2 blast crisis of chronic myelogenous leukemia with a hand mirror cell (HMC) variant. Special stains revealed the leukemic cells to be terminal deoxynucleotidyl transferase positive by immunofluorescence and cytochemically positive for alpha-naphthyl acetate esterase and acid phosphatase (diffuse granular). Immunophenotyping identified the major leukemic cell population as B-cells that expressed CD10+, CD19+, and HLA-DR+. It was not possible to separate the HMC and the non-HMC leukemic population by gating various cell populations, dual staining, cytochemistry, or by terminal deoxynucleotidyl transferase. Gene rearrangements were observed in both Ig heavy-chain alleles and one T-cell antigen receptor gamma-subunit allele. The rearrangements occupied all of the cells, indicating that the HMC and non-HMC were of a common clonal origin. The patient had a mosaic karyotype, with 90% of the cells having t(9;22), t(8;14), and t(9;15) translocations, an additional chromosome 8, and deleted chromosomes 9 and 15. Antibodies to simian sarcoma-associated virus and baboon endogenous virus were isolated in the patient's peripheral blood plasma.     

Ante mortem diagnosis of human rabies using saliva samples: comparison of real time and conventional RT-PCR techniques.

BACKGROUND: Rabies is an enzootic and fatal disease and is still a major problem in developing countries. Ante mortem diagnosis in human cases is necessary for medical management of the patient and to ensure appropriate post-exposure treatment of contacts. Both conventional RT-PCR and Real time PCR (TaqMan) have been described for the detection of rabies virus RNA from saliva and tissue respectively, however to date, there have been no studies comparing conventional and real time PCR assays for detection of rabies virus nucleic acid using saliva samples for ante mortem diagnosis. OBJECTIVES: In this study, we evaluated the utility of conventional RT-PCR and SYBR Green I Real time PCR in the ante mortem diagnosis of rabies using saliva samples. STUDY DESIGN: Saliva samples collected from twenty-four patients presenting with typical clinical manifestations of rabies were tested in the two assays. RESULTS: Amongst the 24 samples tested, 21 samples (87.5%) were positive by either of the two molecular methods. Of these 21, rabies virus RNA was detected in 6/21 in the conventional RT-PCR assay while SYBR Green I Real time PCR could detect RNA in 18/21 samples. CONCLUSION: Real time PCR assay was more sensitive than conventional RT-PCR assay (sensitivity 75% versus 37%, p=0.0189). This study highlights the utility of molecular diagnostic tests in establishing ante mortem diagnosis of rabies using saliva samples within a few hours.     

Validity of self-report of recent opiate use in treatment setting

Self-report validity of recent drug use among heroin abusers depends on many factors including the population being studied and the setting in which the study is carried out. This study was conducted by the treating physicians to assess the self-report validity of recent heroin use by heroin dependent patients in the outdoor setting using 'thin layer chromatography' (TLC) and two highly sensitive methods of urinalysis viz. 'gas liquid chromatography' (GLC) and 'high performance liquid chromatography' (HPLC). Out of seventy-six heroin dependent patients who entered the study, 64 provided urine sample on the same day. Patients' self-report about recent opiate use was found to have a moderate agreement with urinalysis report. However, it is important to validate it with urinalysis during the treatment process because a substantial proportion of patients fails to report recent opiate use. It is recommended that all drug dependence treatment centres should be equipped with a sensitive urinalysis facility. Otherwise, the outcome of the treatment process should be considered with caution.     

Fungal endophthalmitis in a tertiary care cancer center: a review of 23 cases

Few data exist on the etiology, presentation, prognosis, and management of fungal endophthalmitis (FE) in cancer patients. FE cases were identified by reviewing the ophthalmology reports and microbiology cultures of patients at The University of Texas M. D. Anderson Cancer Center. We retrospectively reviewed the medical records and obtained information related to malignancy, fungal infection and its management, visual outcome, and mortality. We compared FE caused by Candida spp. (CE) to FE caused by molds (ME). Of the 102 cancer patients with a fungal infection for whom an ophthalmology consult was requested, 23 met the criteria for definite (N = 6) or probable (N = 17) FE (8 with CE, 15 with ME). All of the patients with ME had hematologic malignancies, whereas half of the patients with CE had solid tumor (P = .008). Only patients with CE had a history of surgery within 30 days of FE diagnosis (38%, P = .03). Fungal pneumonia [17 (74%)] and disseminated infection [14, (61%)] were common. The most common presenting symptoms were decreased vision [16 (70%)] and ocular pain [14 (61%)]. All treated patients received systemic antifungals (combination therapy in 72% of the cases). Seven patients (30%) underwent vitrectomy. Only one patient received intraocular injection of amphotericin B along with systemic antifungals. Four-week mortality was high [13 (57%)], especially in ME (73%, P = .04). Among the eight surviving patients where visual acuity could be assessed, visual outcome improved or remained stable in five (63%). FE in cancer patients occurs in the setting of severe, frequently disseminated opportunistic mycoses, is caused predominantly by hyalohyphomycetes, and is a marker for high 4-week mortality.     

Influence of nanoporous alumina membranes on long-term osteoblast response

A major goal of bone tissue engineering is to design better scaffold configuration and materials to better control osteoblast behavior. Nanoporous architecture has been shown to significantly affect cellular response. In this work, nanoporous alumina membranes were fabricated by a two-step anodization method to investigate bone cell response. Osteoblasts were seeded on nanoporous alumina membranes to investigate both short-term adhesion and proliferation and long-term functionality and matrix production. Cell adhesion and proliferation were characterized using a standard MTT assay and cell counting. The total protein content was measured after cell lysis using the BCA assay. Matrix production was characterized in terms of surface concentrations of calcium and phosphorous, components of bone matrix, using X-ray photoelectron spectroscopy (XPS). The results from nanoporous alumina membranes were compared with those of amorphous alumina, aluminum, commercially available ANOPORE and glass. Results indicate improved osteoblast adhesion and proliferation and increased matrix production after 4 weeks of study.     

Effect of temperature, pH, and ions on sweet taste

The purpose of this experiment was to determine the effects of temperature (50 degrees C and 6 degrees C), pH (pH 3.0, 4.0, 5.0, 6. 0, and 7.0) and the addition of monovalent and divalent cations (5 mM Na(+), 5 mM K(+), and 5 mM Ca(2)+ ) on the sweetness intensity ratings of sweeteners ranging widely in chemical structure. A trained panel provided intensity evaluations for prototypical tastes (sweet, bitter, sour, and salty) as well as aromatic and mouth-feel attributes. The following sweeteners were included in this experiment: three sugars (fructose, glucose, sucrose), three terpenoid glycosides (monoammonium glycyrrhizinate, rebaudioside-A, stevioside), two polyhydric alcohols (mannitol, sorbitol), two dipeptide derivatives (alitame, aspartame), two N-sulfonylamides (acesulfame-K, sodium saccharin), one sulfamate (sodium cyclamate), one protein (thaumatin), one dihydrochalcone (neohesperidin dihydrochalcone), and one chlorodeoxysugar (sucralose). Two to five levels of each sweetener reflecting a range of sweetness intensities were tested, using formulae developed by DuBois et al. The main finding from this three-part study was that temperature, pH, and ions had little effect on perceived sweetness intensity. Even when significant differences were found in the temperature study, the effects were very small.