RESUMEN
OBJECTIVE: To detect and characterise enterovirus RNA in skeletal muscle from patients with chronic fatigue syndrome (CFS) and to compare efficiency of muscle energy metabolism in enterovirus positive and negative CFS patients. METHODS: Quadriceps muscle biopsy samples from 48 patients with CFS were processed to detect enterovirus RNA by two stage, reverse transcription, nested polymerase chain reaction (RT-NPCR), using enterovirus group specific primer sets. Direct nucleotide sequencing of PCR products was used to characterise the enterovirus. Controls were 29 subjects with normal muscles. On the day of biopsy, each CFS patient undertook a subanaerobic threshold exercise test (SATET). Venous plasma lactate was measured immediately before and after exercise, and 30 minutes after testing. An abnormal lactate response to exercise (SATET+) was defined as an exercise test in which plasma lactate exceeded the upper 99% confidence limits for normal sedentary controls at two or more time points. RESULTS: Muscle biopsy samples from 20.8% of the CFS patients were positive for enterovirus sequences by RT-NPCR, while all the 29 control samples were negative; 58.3% of the CFS patients had a SATET+ response. Nine of the 10 enterovirus positive cases were among the 28 SATET+ patients (32.1%), compared with only one (5%) of the 20 SATET- patients. PCR products were most closely related to coxsackie B virus. CONCLUSIONS: There is an association between abnormal lactate response to exercise, reflecting impaired muscle energy metabolism, and the presence of enterovirus sequences in muscle in a proportion of CFS patients.
Asunto(s)
ADN Viral/análisis , Enterovirus/genética , Enterovirus/patogenicidad , Ejercicio Físico/fisiología , Síndrome de Fatiga Crónica/etiología , Síndrome de Fatiga Crónica/virología , Músculo Esquelético/virología , Adulto , Enterovirus Humano B/genética , Enterovirus Humano B/patogenicidad , Prueba de Esfuerzo , Femenino , Humanos , Ácido Láctico/metabolismo , Masculino , Persona de Mediana Edad , Músculo Esquelético/patología , Músculo Esquelético/fisiología , Enfermedades Musculares/metabolismo , Enfermedades Musculares/virología , Resistencia Física , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
This study was performed to detect and characterize the enterovirus present in myocardium of some patients with heart muscle disease by nucleotide sequencing of polymerase chain reaction (PCR) products after amplification with enterovirus group-specific primers. Enterovirus sequences have been detected previously in myocardium of patients with myocarditis or dilated cardiomyopathy and seem causal, although the particular virus serotypes involved have not been identified. In a prospective study of endomyocardial biopsy specimens from 35 consecutive patients with suspected heart muscle disease, enterovirus sequences from the 5' nontranslated region were amplified by reverse transcription-nested PCR using group-specific primers. This region contains both conserved and variable sequence motifs, characteristic of particular enterovirus serotypes. The nucleotide sequences of individual PCR products were determined by cycle sequencing and compared with all known sequences (GenBank/EMBOL), using the GCG software package. Endomyocardial biopsy specimens from 9 of 21 (42.9%) patients with a histologically confirmed diagnosis of dilated cardiomyopathy were positive for enterovirus by PCR, compared with only 1 of 14 (7.1%) patients with other myocardial pathological conditions (Fisher's exact probability=0.0275: odds ratio=9.75; 95% confidence interval=1.31-72.78). The nucleotide sequence of the PCR products differed, indicating no cross-contamination. However, computerized comparison showed that each had greatest homology with the 5' nontranslated region of Coxsackie B virus but contained up to 11% sequence variations compared with the prototype Coxsackie B3 strain Nancy. Parallel investigation of tissue from our mouse model of Coxsackievirus B3-induced myocarditis showed that nucleotide sequence changes are not introduced by reverse transcription or PCR. These data support the link between enteroviral infection and dilated heart muscle disease and suggest that Coxsackie B serotypes are the enteroviruses most frequently involved.
Asunto(s)
Cardiomiopatía Dilatada/virología , Infecciones por Coxsackievirus/virología , Enterovirus Humano B/genética , Corazón/virología , Miocardio/patología , ARN Viral/análisis , Adolescente , Adulto , Anciano , Cardiomiopatía Dilatada/patología , Infecciones por Coxsackievirus/patología , Cartilla de ADN/química , Enterovirus Humano B/aislamiento & purificación , Femenino , Humanos , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Estudios Prospectivos , Análisis de Secuencia de ARN , Transcripción GenéticaRESUMEN
The purpose of this study was to determine the genetic origin of a series of seven diploid hydatidiform moles with fetal red blood cells in the molar villi, normally a characteristic feature of triploid, partial hydatidiform moles. DNA was prepared from formalin-fixed, paraffin-embedded blocks of molar tissue and blood from the patient and her partner. The genetic origin of molar tissue was determined by comparing microsatellite polymorphisms in molar and parental tissue following polymerase chain reaction (PCR) amplification of DNA. In six cases, the hydatidiform mole was shown to be androgenetic in origin and therefore genetically to be a complete hydatidiform mole. In one case, the hydatidiform mole was of biparental origin, having both a maternal and a paternal contribution to the genome. We conclude that fetal red blood cells may be observed in the villi of complete hydatidiform moles. In cases where the degree of trophoblastic hyperplasia and ploidy is suggestive of a complete hydatidiform mole, the presence of fetal red blood cells alone should not be considered indicative of a diagnosis of partial hydatidiform mole.
Asunto(s)
Diploidia , Embrión de Mamíferos/patología , Eritrocitos/patología , Mola Hidatiforme/genética , Neoplasias Uterinas/genética , ADN de Neoplasias/genética , Diagnóstico Diferencial , Femenino , Humanos , Mola Hidatiforme/patología , Repeticiones de Microsatélite , Reacción en Cadena de la Polimerasa , Polimorfismo Genético , Embarazo , Neoplasias Uterinas/patologíaRESUMEN
Molluscum contagiosum virus (MCV) and vaccinia virus (VV) are serologically unrelated poxviruses with a disparate genome composition (MCV, 66% G+C; VV, 33% G+C). Molecular studies of MCV have been hindered by the inability to propagate the virus in cells cultured in vitro. We sequenced 7765 bp of MCV DNA cloned from four widely spaced regions throughout the MCV genome and identified a total of 11 potential open reading frames (ORF), designated CX1-11. These include MCV homologues of the VV genes encoding protein kinase 2, structural protein VP8, RNA polymerase 35 kDa subunit and 3beta-hydroxysteroid dehydrogenase. The position and orientation of the MCV ORFs was collinear to the VV genome, with the exception of the region around ORF CX11 which is inverted in the MCV genome.
Asunto(s)
3-Hidroxiesteroide Deshidrogenasas/genética , Cápside/genética , Proteínas de Unión al ADN/genética , ARN Polimerasas Dirigidas por ADN/genética , Genoma Viral , Molusco Contagioso/virología , Virus del Molusco Contagioso/genética , Proteínas Serina-Treonina Quinasas/genética , Virus Vaccinia/genética , Proteínas Estructurales Virales/genética , Secuencia de Aminoácidos , Secuencia de Bases , Proteínas de la Cápside , ADN Viral , Humanos , Datos de Secuencia Molecular , Molusco Contagioso/patología , Homología de Secuencia de Aminoácido , Proteínas ViralesRESUMEN
For appropriate clinical management of patients with gestational trophoblastic tumors it is important to ascertain both the nature of the causative pregnancy and the time interval between that pregnancy and the diagnosis of the tumor. It has been shown that the immediately antecedent pregnancy may not be the causative pregnancy in some cases of choriocarcinoma, particularly where there is a history of molar pregnancy. We report further studies of a case where the causative pregnancy was shown to be a hydatidiform mole, not the immediately antecedent normal term pregnancy. We describe the use of the polymerase chain reaction (PCR) to amplify short tandem repeat polymorphisms in DNA prepared from pathologic blocks of the patient's previously recognised molar pregnancy. A comparison of these polymorphisms with those in the parental and tumor DNA has enabled us to confirm that this hydatidiform mole was indeed the causative pregnancy. Molecular genetic techniques provide a rapid method of determining whether a choriocarcinoma is gestational and, if so, identifying the causative pregnancy.