RESUMO
Background Homozygous familial hypercholesterolemia (HoFH) is a rare, treatment-resistant disorder characterized by early-onset atherosclerotic and aortic valvular cardiovascular disease if left untreated. Contemporary information on HoFH in the United States is lacking, and the extent of underdiagnosis and undertreatment is uncertain. Methods and Results Data were analyzed from 67 children and adults with clinically diagnosed HoFH from the CASCADE (Cascade Screening for Awareness and Detection) FH Registry. Genetic diagnosis was confirmed in 43 patients. We used the clinical characteristics of genetically confirmed patients with HoFH to query the Family Heart Database, a US anonymized payer health database, to estimate the number of patients with similar lipid profiles in a "real-world" setting. Untreated low-density lipoprotein cholesterol levels were lower in adults than children (533 versus 776 mg/dL; P=0.001). At enrollment, atherosclerotic cardiovascular disease and supravalvular and aortic valve stenosis were present in 78.4% and 43.8% and 25.5% and 18.8% of adults and children, respectively. At most recent follow-up, despite multiple lipid-lowering treatment, low-density lipoprotein cholesterol goals were achieved in only a minority of adults and children. Query of the Family Heart Database identified 277 individuals with profiles similar to patients with genetically confirmed HoFH. Advanced lipid-lowering treatments were prescribed for 18%; 40% were on no lipid-lowering treatment; atherosclerotic cardiovascular disease was reported in 20%; familial hypercholesterolemia diagnosis was uncommon. Conclusions Only patients with the most severe HoFH phenotypes are diagnosed early. HoFH remains challenging to treat. Results from the Family Heart Database indicate HoFH is systemically underdiagnosed and undertreated. Earlier screening, aggressive lipid-lowering treatments, and guideline implementation are required to reduce disease burden in HoFH.
Assuntos
Anticolesterolemiantes , Aterosclerose , Doenças Cardiovasculares , Hipercolesterolemia Familiar Homozigota , Hiperlipoproteinemia Tipo II , Estados Unidos/epidemiologia , Humanos , Doenças Cardiovasculares/tratamento farmacológico , Hiperlipoproteinemia Tipo II/diagnóstico , Hiperlipoproteinemia Tipo II/epidemiologia , Hiperlipoproteinemia Tipo II/genética , LDL-Colesterol , Aterosclerose/diagnóstico , Aterosclerose/epidemiologia , Aterosclerose/genética , Sistema de Registros , Anticolesterolemiantes/uso terapêutico , HomozigotoRESUMO
PURPOSE: Polymerase chain reaction (PCR) is a molecular technique for the diagnosis of ocular infectious disease. In this large patient sample and multiyear study, the impact of PCR for detecting infectious agents from ocular samples was reviewed in comparison with nonmolecular diagnostic techniques. DESIGN: A retrospective laboratory review of PCR testing. PARTICIPANTS: Three thousand fifty-six patient samples with a differential of ocular infection. METHODS: The daily laboratory logs for diagnostic testing were reviewed for PCR, cell culture isolation, shell vial isolation, and Acanthamoeba isolation from January 1997 through May 2010 for herpes simplex virus (HSV), adenovirus, varicella zoster virus (VZV), Chlamydia trachomatis, Acanthamoeba, and infrequent pathogens of intraocular inflammation. MAIN OUTCOME MEASURES: Incidence of the positive presence of ocular infectious agents. RESULTS: Polymerase chain reaction results were positive more often than culture results for HSV (P = 0.0001), VZV (P = 0.00001), C. trachomatis (P = 0.00005), and Acanthamoeba (P = 0.04). For adenovirus, cell culture isolation results were positive more often than PCR results (P = 0.001). Polymerase chain reaction was the primary diagnostic test for detecting cytomegalovirus and Toxoplasma. CONCLUSIONS: The current study demonstrated the importance of PCR as a routine diagnostic test for detecting both common and infrequent ocular pathogens. Cell culture isolation is still a definitive test for adenovirus and a confirmatory test for HSV and Acanthamoeba.
Assuntos
Infecções Oculares/microbiologia , Infecções Oculares/parasitologia , Reação em Cadeia da Polimerase , Acanthamoeba/isolamento & purificação , Adenoviridae/isolamento & purificação , Linhagem Celular Tumoral , Chlamydia trachomatis/isolamento & purificação , Citomegalovirus/isolamento & purificação , Herpesvirus Humano 3/isolamento & purificação , Humanos , Reação em Cadeia da Polimerase/normas , Estudos Retrospectivos , Simplexvirus/isolamento & purificação , Toxoplasma/isolamento & purificaçãoRESUMO
Many Serratia marcescens strains produce the red pigment prodigiosin, which has antimicrobial and anti-tumor properties. Previous reports suggest that cyclic AMP (cAMP) is a positive regulator of prodigiosin production. Supporting this model, the addition of glucose to growth medium inhibited pigment production in rich and minimal media. Unexpectedly, we observed highly elevated levels of prodigiosin production in isogenic strains with mutations in genes involved in cAMP production (cyaA and crr) and in cAMP-dependent transcriptional signaling (crp). Multicopy expression of the Escherichia coli cAMP-phosphodiesterase gene, cpdA, also conferred a striking increase in prodigiosin production. Exogenous cAMP decreased both pigment production and pigA-lacZ transcription in the wild-type (WT) strain, and pigA-lacZ transcription was significantly increased in a crp mutant relative to WT. Suppressor and epistasis analysis indicate that the hyperpigment phenotype was dependent upon pigment biosynthetic genes (pigA, pigB, pigC, pigD and pigM). These experiments establish cAMP as a negative regulator of prodigiosin production in S. marcescens.
Assuntos
AMP Cíclico/metabolismo , Regulação da Expressão Gênica , Prodigiosina/biossíntese , Serratia marcescens/fisiologia , Proteínas de Bactérias/genética , Meios de Cultura/química , Deleção de Genes , Glucose/metabolismo , Serratia marcescens/metabolismoRESUMO
Confirmation of Acanthamoeba keratitis by laboratory diagnosis is the first step in the treatment of this vision-threatening disease. Two real-time PCR TaqMan protocols (the Rivière and Qvarnstrom assays) were developed for the detection of genus-specific Acanthamoeba DNA but lacked clinical validation. We have adapted these assays for the Cepheid SmartCycler II system (i) by determining their real-time PCR limits of detection and amplification efficiencies, (ii) by determining their ability to detect trophozoites and cysts, and (iii) by testing a battery of positive and negative samples. We also examined the inhibitory effects of a number of commonly used topical ophthalmic drugs on real-time PCR. The results of the real-time PCR limit of detection and amplification efficiency of the Rivière and Qvarnstrom assays were 11.3 DNA copies/10 microl and 94% and 43.8 DNA copies/10 microl and 92%, respectively. Our extraction protocol enabled us to detect 0.7 Acanthamoeba cysts/10 microl and 2.3 Acanthamoeba trophozoites/10 microl by both real-time PCR assays. The overall agreement between the assays was 97.0%. The clinical sensitivity and specificity of both real-time PCR assays based on culture were 100% (7 of 7) and 100% (37 of 37), respectively. Polyhexamethylene biguanide was the only topical drug that demonstrated PCR inhibition, with a minimal inhibitory dilution of 1/640 and an amplification efficiency of 72.7%. Four clinical samples were Acanthamoeba culture negative and real-time PCR positive. Our results indicate that both real-time PCR assays could be used to diagnose Acanthamoeba keratitis. Polyhexamethylene biguanide can inhibit PCR, and we suggest that specimen collection occur prior to topical treatment to avoid possible false-negative results.
Assuntos
Ceratite por Acanthamoeba/diagnóstico , Acanthamoeba/genética , Parasitologia/métodos , Reação em Cadeia da Polimerase/métodos , Acanthamoeba/isolamento & purificação , Animais , Biguanidas/farmacologia , Primers do DNA/genética , Inibidores Enzimáticos/farmacologia , Humanos , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
OBJECTIVE: To compare the SmartCycler II system (Cepheid, Sunnyvale, Calif) results with those of standard cell culture, to compare the SmartCycler II system results with those of a dedicated polymerase chain reaction facility, and to establish the SmartCycler II system as a polymerase chain reaction method for detecting viral and chlamydial DNA from ocular specimens. METHODS: True-positive samples (test-positive specimens based on standard testing) and true-negative samples (test-negative specimens based on standard testing) were processed for polymerase chain reaction using the SmartCycler II system for adenovirus, herpes simplex virus type 1, varicella-zoster virus, and Chlamydia trachomatis. Sensitivity, specificity, positive predictive value, negative predictive value, and efficiency were based on the testing of true-positive and true-negative specimens. RESULTS: The descriptive statistics for adenovirus, herpes simplex virus type 1, varicella-zoster virus, and C trachomatis were, respectively, as follows: sensitivity, 85%, 98%, 100%, and 94%; specificity, 98%, 100%, 100%, and 100%; positive predictive value, 98%, 100%, 100%, and 100%; negative predictive value, 85%, 91%, 100%, and 98%; and efficiency, 92%, 95%, 100%, and 99%. Test sensitivity for the SmartCycler II system was equivalent to that from a central molecular laboratory. CONCLUSION: The descriptive statistics of the SmartCycler II system obtained in a small laboratory were comparable to those of a central molecular laboratory for detecting viruses and Chlamydia species. Clinical Relevance Polymerase chain reaction has great potential in the routine diagnosis of ocular infections in any conventional laboratory.