Interest of human papillomavirus DNA quantification and genotyping in paired cervical and urine samples to detect cervical lesions.

BACKGROUND: Cervical cancer is caused by persistent infection with high-risk human papillomavirus (HR-HPV). Conventional human papillomavirus (HPV) testing requires cervical sampling. However, vaginal and urine self-sampling methods are more acceptable for patients and result in increased participation when they are available in screening programs. In this context, we have developed a non-invasive screening method via the detection of HPV DNA in urine samples. PURPOSE: To compare HPV viral loads and genotypes in paired cervical and urine samples, and to assess correlation between virological and cytological results in women seeking gynecological consultation. METHODS: Paired urine and cervical specimens were collected and analyzed from 230 of 245 women participating in the previously described prospective PapU study. HPV DNA detection and quantification were performed using a real-time PCR method with short fragment PCR primers. Genotyping was carried out using the INNO-LiPA HPV genotyping assay. RESULTS: The prevalence of HPV in the 230 paired urine and cervical smear samples was 42 and 49 %, respectively. Overall agreement for HPV positivity and negativity between the paired samples was 90 % (κ = 0.80). High HPV viral load in both cervical and urine samples was associated with cytological abnormalities. HPV-positive women were mostly infected with HR-HPV types. The agreement between high- and low-risk HPV (LR-HPV) detection in both samples was 97 % (κ = 0.95 for HR-HPV and κ = 0.97 for LR-HPV). CONCLUSIONS: High concordance rates for HPV-DNA quantification and high/low-risk HPV genotyping in paired urine/cervical samples suggest that urinary HPV DNA testing could be useful for cervical lesion screening.

HPV and head and neck cancer.

Head and neck cancer is frequent worldwide and oropharyngeal locations are presently sharply on the increase, in relation with an increasing incidence of oropharyngeal infection by oncogenic type-16 human papillomavirus (HPV). The clinical and biologic profile of these patients is distinct from that of other oropharyngeal carcinoma patients, with earlier onset, cystic cervical nodes and basaloid carcinoma histopathology. Detection of intratumoral viral DNA is essential to confirm the role of HPV, and E6/E7 mRNA expression is the most relevant indicator for stratification. Several methods can reveal intratumoral oncogenic HPV DNA, but PCR with hybridization is the most sensitive and most widely used. According to several reports, prognosis in terms of survival and locoregional control is better in HPV-positive oropharyngeal carcinoma than in oropharyngeal carcinoma associated with smoking and alcohol consumption. The future lies in vaccination, but further studies will determine whether the rate of oropharyngeal carcinoma falls in women vaccinated against cervical cancer.

Human papillomavirus infection of the cervix uteri in women attending a Health Examination Center of the French social security.

Since human papillomavirus (HPV) is the central causal factor in cervical cancer, understanding the epidemiology of this infection constitutes an important step towards development of strategies for prevention. Six hundred and fifty seven cervical samples were tested for HPV using PCR with consensus primers (MY09/MY11), by genotyping (restriction and sequencing analyses) and by cervical cytology, from women who attended a Health Examination Center of the French social security. Women with no cervical smear as well as women with cytological abnormalities within the last 3 years were recruited. HPV DNA was detected in 7.3% of the women (5.3% for high-risk, 2.4% for low-risk, and 0.5% for unknown risk types) including 6 (0.9%) mixed infections. Fifteen different genotypes were detected, of which genotypes 16 (22.2%), 58 (13.0%), 18 (11.1%), 30 (9.2%), and 33 (9.2%) were the most prevalent. In age group 17-25 years, we found the highest frequencies for both any (22.1%) and high-risk (14.7%) HPV, and prevalences gradually decreased with age. 5.2% of low-grade squamous intraepithelial lesion, 0.3% of high-grade squamous intraepithelial lesion, and 1.2% of atypical squamous cells of undetermined significance were found. The frequencies of high risk and all HPV types were significantly higher in squamous intraepithelial lesions than in those with normal and reactive/reparative changes (P < 0.0001). The prevalence of high-risk HPV in the atypical squamous cells of undetermined significance/low-grade squamous intraepithelial lesion group (28.6%) was significantly higher than in the normal and reactive/reparative changes groups (3.4%) (P < 0.0001). HPV detection was associated with younger age, single marital and non-pregnant status (P < 0.0001), premenopausal status (P = 0.0004), and contraception (P = 0.0008). Marital status (OR 4.5; 95% CI = 2.3-9.0) and tobacco consumption (OR 3.0; 95% CI = 1.6-5.7) were predictive independent factors of HPV infection. The French system of Health Examination Centers might be of interest for following women regularly, especially those with a low socioeconomic status.

Surveillance network for herpes simplex virus resistance to antiviral drugs: 3-year follow-up.

Herpes simplex virus (HSV) infections are very common in the general population and among immunocompromised patients. Acyclovir (ACV) is an effective treatment which is widely used. We deemed it essential to conduct a wide and coordinated survey of the emergence of ACV-resistant HSV strains. We have formed a network of 15 virology laboratories which have isolated and identified, between May 1999 and April 2002, HSV type 1 (HSV-1) and HSV-2 strains among hospitalized subjects. The sensitivity of each isolate to ACV was evaluated by a colorimetric test (C. Danve, F. Morfin, D. Thouvenot, and M. Aymard, J. Virol. Methods 105:207-217, 2002). During this study, 3900 isolated strains among 3357 patients were collected; 55% of the patients were immunocompetent. Only six immunocompetent patients excreted ACV-resistant HSV strains (0.32%), including one female patient not treated with ACV who was infected primary by an ACV-resistant strain. Among the 54 immunocompromised patients from whom ACV-resistant HSV strains were isolated (3.5%), the bone marrow transplantation patients showed the highest prevalence of resistance (10.9%), whereas among patients infected by human immunodeficiency virus, the prevalence was 4.2%. In 38% of the cases, the patients who excreted the ACV-resistant strains were treated with foscarnet (PFA), and 61% of them developed resistance to PFA. The collection of a large number of isolates enabled an evaluation of the prevalence of resistance of HSV strains to antiviral drugs to be made. This prevalence has remained stable over the last 10 years, as much among immunocompetent patients as among immunocompromised patients.

Fatal adenovirus type 7b infection in a child with Smith-Lemli-Opitz syndrome.

Adenovirus type 7 causes worldwide respiratory tract infections, mainly in children. Severe systemic infections can occur, especially in immunocompromised patients and in patients with underlying chronic diseases. This report describes the first case of a fatal disseminated adenovirus type 7 infection in a child with Smith-Lemli-Opitz syndrome, a rare autosomal recessive disorder due to a primary enzymatic defect in cholesterol metabolism. Nasopharyngeal secretions and autopsy specimens including liver, lung, pleural fluid, and rectum were collected for viral culture. Adenovirus serotype 7 strains were obtained from all anatomic sites, except the liver. All these clinical isolates were analyzed using restriction endonuclease digestion of the genome, identifying them as genome type 7b, a virulent type. In this case, the fatal evolution could have been accelerated by the presence of an immunodeficiency although immunodeficiency is not included in the definition of Smith-Lemli-Opitz syndrome. The frequent recurrent banal infections in Smith-Lemli-Opitz syndrome could be prevented by a cholesterol supplementation regimen. Finally, this report emphasizes the need for efficient therapy for disseminated adenovirus infections, especially for virulent genome types.

Nucleotide sequence analysis of human T-cell lymphotropic virus type I pX and LTR regions from patients with sicca syndrome.

Human T-cell lymphotropic virus type I (HTLV-I) is associated with adult T-cell leukemia (ATL) and tropical spastic paraparesis/HTLV-I-associated myelopathy (TSP/HAM). Other inflammatory disorders may occur in HTLV-I-infected patients, such as sicca syndrome resembling Sjögren's syndrome. The sicca syndrome may be the unique clinical manifestation of HTLV-I infection, but is associated frequently with TSP/HAM, which could suggest that sicca syndrome might be an early event in disease progression to TSP/HAM in some cases. We investigated whether peculiar pX and LTR mutations could be related to sicca syndrome, or might argue the existence of clinical progression to TSP/HAM. pX, especially pX(I), pX(II), and pX(IV) ORFs corresponding to Tax cytotoxic T-lymphocyte epitopes, and LTR regions from Caribbean patients who have sicca syndrome with or without TSP/HAM, ATL patients, and healthy carriers were sequenced. The sequences were aligned and compared with ATK-1 prototype and published sequences. LTR sequences exhibited 1.5-2.4% of divergence with ATK-1. pX-sequenced regions showed a lower homology within p12(I) encoding sequences. Only few mutations were found within functionally important regions, but were not associated specifically with the clinical status. Finally, no mutations that could be related to sicca syndrome or argue the existence of clinical progression to TSP/HAM were found. It would be of interest to study the clinical evolution of HTLV-I-sicca syndrome in patients and to determine HTLV-I sequences from peripheral blood and salivary glands at different stages.

Genital recurrent infection occurring 6 months after meningitis due to the same herpes simplex virus type 2 (HSV-2) strain evidence by restriction endonuclease analysis.

Herpes simplex virus type 2 (HSV-2) is more often sexually transmitted and associated with genital recurrent infection. However, HSV-2 neurological manifestations such as meningitis were already reported. We describe a case of meningitis due to HSV-2, preceded by signs suggesting a common cystitis, in a woman with no history of primary or recurrent genital infection. Six months later genital herpetic lesions occurred. One HSV-2 strain was obtained from cerebrospinal fluid (CSF) and another from genital lesions. The molecular comparative analysis using restriction endonuclease digestion patterns showed the similarity of the two strains. Our report illustrates that HSV-2 infections are underdiagnosed and that molecular techniques can be of value in clarifying the physiopathology of HSV diseases.

ORL-1 and mu opioid receptor antisera label different fibers in areas involved in pain processing.

Mu opioid receptors (MOR) mediate the analgesic effects of opioid drugs such as morphine. The opioid receptor-like (ORL-1) receptor is structurally related to opioid receptors and the ORL-1 receptor agonist, orphanin FQ/nociceptin, induces analgesia at the spinal level, but appears to recruit different circuitry than that used by mu opioids. When administered intracerebroventricularly, orphanin FQ/nociceptin produces hyperalgesia and/or reverses opioid analgesia. The functionally distinct actions elicited by MOR and ORL-1 receptors, which activate similar intracellular signaling systems and show similar regional distributions, could be explained by their differential cellular localization. By using double label immunohistochemistry and confocal microscopy, the present study investigates the distribution of MOR and ORL-1 receptors in regions of the rat nervous system that are involved with nociceptive processing. In general co-localization of MOR and ORL-1 receptor immunoreactivity was not observed in either perikarya or neuropil in the dorsal root ganglia, nor in the Lissauer's tract and superficial laminae of the spinal cord. Likewise, there was no evidence for co-localization of these receptors within the periaqueductal gray, the nucleus raphe magnus, the gigantocellular reticular nucleus, and the nucleus of the solitary tract. These observations indicate that MOR and ORL-1 receptors are expressed predominantly on different fiber systems in these regions. This differential distribution is consistent with the distinct pharmacology of ORL-1 and MOR receptor agonists and suggests that the antisera to MOR and ORL-1 receptors may provide useful markers for further investigations of analgesic and counteranalgesic pathways modulating pain perception.

mu-Opioid receptor internalization: opiate drugs have differential effects on a conserved endocytic mechanism in vitro and in the mammalian brain.

mu-Opioid receptors are the pharmacological targets of endogenous opioid peptides and morphine-like alkaloid drugs. Previous studies of transfected cells and peripheral neurons indicate that opioid receptors are rapidly internalized after activation by the alkaloid agonist etorphine but not after activation by morphine. To determine whether opioid receptors in the central nervous system are regulated by a similar process of agonist-selective internalization, mu-opioid receptors were examined in rat brain neurons after treatment of animals with opioid drugs. Internalized mu receptors were observed within 30 min after intraperitoneal injection of the alkaloid agonist etorphine, and this process was blocked by the antagonist naloxone. Colocalization of internalized opioid receptors with transferrin receptors in confocal optical sections indicated that receptor internalization observed in vivo is mediated by a membrane trafficking pathway similar to that observed previously in vitro using transfected human embryonic kidney 293 cells. Morphine failed to induce detectable rapid internalization of receptors, even when administered to animals at doses far in excess of those required to induce analgesia. To quantify these agonist-selective differences and to analyze an array of opioid ligands for their ability to trigger internalization, we used flow cytometry on stably transfected 293 cells. These studies indicated that the different effects of individual agonists are not correlated with their potencies for receptor activation and that a variety of clinically important agonists differ significantly in their relative abilities to stimulate the rapid internalization of opioid receptors.

[Rhinovirus infections in hospitalized children: a 3-year study]. / Infections à rhinovirus chez des enfants hospitalisés: étude sur une période de 3 ans.

BACKGROUND: Rhinoviruses (RH) are responsible for acute respiratory illnesses, mainly in the upper respiratory tract. POPULATION AND METHODS: 3,152 children aged under 16 years, admitted to the Paediatrics department of the University Hospital Centre of Poitiers from January 1, 1993 to December 31, 1995 with ear, nose and throat (ENT) and/or respiratory symptoms were systematically investigated. One hundred and forty-five RH strains were isolated from nasopharyngeal secretions of 87 boys and 58 girls (mean age: 20.3 months). Among these, 92 (63.4%) were less than 1 year of age. Bacteriological investigations were done for 29 patients when a concomitant bacterial infection was suspected. RESULTS: RH infection rate was maximum before 1 year of age (median age: 6.5 months) and decreased with age. RH were isolated throughout the 3 years, with a first peak from February to April, and a second one in autumn. The main symptoms were sibilants (27.6%) and cough (24.1%). Sibilants were more frequently associated in children under 12 months of age (P = 0.01). Sometimes, ophthalmologic or digestive symptoms were present. Three children with respiratory distress were transferred to the reanimation ward. In addition, a RH strain was isolated from a child who died of sudden infant death. Thirty-four children (23.4%) were co-infected by one or several viruses; the most frequently detected were the respiratory syncytial virus (41.2%) and the adenoviruses (35.3%). Twenty-nine children were infected by two viruses and five by three. Associated bacterial infections were diagnosed in 23 children, especially conjunctivitis due to Haemophilus influenzae (21.7%). Among these children, eight had a multiple viral infection. CONCLUSION: RH have a limited pathogenicity but can be associated with serious illnesses among infants and children.

[Epidemic of viral gastroenteritis in an elderly community]. / Epidémie de gastro-entérite virale dans une collectivité de sujets âgés.

Rotaviruses and adenoviruses are the main aetiologic pathogens of gastroenteritis in infants. Adults may also be concerned with usually mild clinical symptoms. An epidemic of viral gastroenteritis occurred in a long-term hospitalization ward from July 25 to August 21, 1991: 101 patients. This study involved 26 females and 6 males with a mean age of 86 years (range 70-101 years), presenting clinical symptoms of gastroenteritis. The clinical attack rate was 27.7%. Outcome was favourable for the majority of patients who recovered in 4-5 days. The main clinical signs were diarrhoea (90.6%), hyperthermia (18.8%) and vomiting (18.8%). Virology investigations gave the diagnosis of 8 rotavirus infections, 6 adenovirus infections and 2 rotavirus and adenovirus mixed infections. Among staff members, 3 rotavirus and one adenovirus infections were diagnosed. The precise origin of the epidemic could not be determined from rotavirus electropherotypes obtained from stools of elderly and paediatric patients hospitalized during the same period. This outbreak recalls the viral involvement in diarrhoeal episodes of elderly people. These episodes of viral gastroenteritis are responsible for high morbidity in the elderly and may upset a precarious physiological state.

Unclassifiable high grade malignant T-cell lymphoma with clonal evolution.

We report a case of leukemic malignant T-cell lymphoma with mixed small and large cells. The small cells displayed a mature CD8-positive phenotype, a diploid DNA distribution by cell cycle analysis, and structural karyotypic abnormalities. Large cells were near triploid, showed additional structural cytogenetic abnormalities and a more immature membrane phenotype without CD8 expression. Altogether, these data provide suggestive evidence for a clonal evolution from a mature small cell T-cell lymphoma to a more immature large cell proliferation.

Polymerase chain reaction amplified HTLV-I, HIV-1 and HIV-2 DNA fragments in subjects with mixed retroviral infections.

Serum and peripheral blood mononuclear cells from eight patients from the Ivory Coast with positive screening test results for retroviral infections were studied by serology (ELISA, Western blot (WB), synthetic peptide test), cell co-culture, and polymerase chain reaction (PCR). Two HIV-2 infections with indeterminate interpretation on HIV-1 WB were detected, two were clear dual HIV-1/HIV-2 infections, three were ambiguous mixed HIV-1/HIV-2 infections, and one was a triple retroviral infection by HTLV-I, HIV-1 and HIV-2. Four slow/low HIV-1 strains were isolated at the expense of HTLV-I and HIV-2 strains. The ELISA tests were found to be very sensitive. Indeterminate WB interpretations were frequent (HTLV-I, four; HIV-1, three; HIV-2, two). PCR provided clear evidence of multiple retroviral infections in three cases and enabled interpretation of indeterminate WB samples in three cases. One sample presented a puzzling pattern with positive PCR results for HIV-1 and HIV-2 associated with negative or indeterminate serological results. Thus, our data emphasise the need to analyse serological as well as virological markers to gain better insight on mixed retroviral infections, especially in endemic areas such as West Africa.

[Necrotizing enterocolitis in a premature infant]. / Entérite aiguë nécrosante du prématuré: rôle possible d'un adénovirus.

We report a case of necrotizing enterocolitis in a 30 weeks old premature infant fed with breast milk. No ischemic etiology was present. Bacteriological and parasitological investigations were found to be negative. Only, a positive latex agglutination test for Adenoviruses was detected in stools collected the day of the bloody diarrhea. Detailed microbiological examinations are emphasized by this unusual finding because the role of pathogens in this disease deserves further understanding.