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Titolo:
IN-SITU DETECTION OF HUMAN CYTOMEGALOVIRUS DNA IN GASTROINTESTINAL BIOPSIES FROM AIDS PATIENTS BY MEANS OF VARIOUS PCR-DERIVED METHODS
Autore:
MUSSO O; SOMMER P; DROUET E; COTTE L; NEYRA M; GRIMAUD JA; CHEVALLIER M;
Indirizzi:
INST PASTEUR LYON,CNRS URA 1459,UNITE PATHOL CELLULAIRE,AVE TONY GARNIER F-69365 LYON 07 FRANCE INST PASTEUR LYON,CNRS URA 1459,UNITE PATHOL CELLULAIRE F-69365 LYON 07 FRANCE INST PASTEUR,UNIT BACTERIOL VIROL F-69365 LYON 07 FRANCE INSERM U271,UNITE RECH CLIN,SIDA F-69003 LYON FRANCE HOP HOTEL DIEU,UNITE HEPATOGASTROENTEROL F-69003 LYON FRANCE
Titolo Testata:
Journal of virological methods
fascicolo: 2, volume: 56, anno: 1996,
pagine: 125 - 137
SICI:
0166-0934(1996)56:2<125:IDOHCD>2.0.ZU;2-2
Fonte:
ISI
Lingua:
ENG
Soggetto:
POLYMERASE CHAIN-REACTION; ACQUIRED-IMMUNODEFICIENCY-SYNDROME; NUCLEIC-ACID HYBRIDIZATION; VIRUS INFECTION; INSITU HYBRIDIZATION; AMPLIFICATION; SEQUENCES; DIGOXIGENIN; CULTURE; COLITIS;
Keywords:
PCR; NONRADIOACTIVE ISH; CYTOMEGALOVIRUS; PCR IN SITU; AIDS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Science Citation Index Expanded
Science Citation Index Expanded
Citazioni:
41
Recensione:
Indirizzi per estratti:
Citazione:
O. Musso et al., "IN-SITU DETECTION OF HUMAN CYTOMEGALOVIRUS DNA IN GASTROINTESTINAL BIOPSIES FROM AIDS PATIENTS BY MEANS OF VARIOUS PCR-DERIVED METHODS", Journal of virological methods, 56(2), 1996, pp. 125-137

Abstract

The development of new in situ assessment of HCMV disease on endoscopical gastrointestinal biopsies from AIDS patients is described and compared with the viral load measured by semiquantitative solution-phase PCR (SQ-PCR). Ten biopsies were examined by viral isolation, standard histology, in situ hybridization (ISH), in situ PCR-hybridization (PCR-ISH) and SQ-PCR, using the same target sequence. The methods developed for in situ HCMV detection were HCMV primers, the plasmid pCMV 406-S, a vector-free-digoxigenin-labelled HCMV-362 probe and the pSK + MCS nonsense probe. Paraffin-embedded MRCS cells, either HCMV-infected or uninfected served as controls of specificity for ISH. beta-Actin primers were designed as markers of DNA integrity. Computerized models of the PCR, solution-phase and in situ PCR on formalin-fixed DNA indicatedthat HCMV and beta-actin primers were efficient and specific. Nine biopsies were negative for HCMV by histology and virus isolation. SQ-PCRrevealed 80 000; 80 and <80 HCMV genomic equivalents in 6, 2 and 2 biopsies, respectively. In 8 biopsies, both ISH and PCR-ISH identified positive nuclei in the intestinal epithelium, with sparing of the lamina propria. This indicates that an improvement in in situ methods can help the timely diagnosis of HCMV infection. Direct in situ PCR with beta-actin primers showed a positive signal in all the nuclei in the tissue sections, whereas omission of Tag polymerase resulted in an absence of signal, implying optimal in situ PCR. The data suggest an early-stage reactivation of HCMV, possibly harboured in the intestinal epithelium.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 20/09/20 alle ore 19:53:20