Detection of three human adenovirus species in adults with acute respiratory infection in China.

Human adenoviruses (HAdVs) are recognized as causal agents in a wide range of human diseases. However, researchers lack sufficient data on the exact HAdV species and serotypes associated with adult acute respiratory tract infections (ARTIs). To detect and characterize HAdV infections in adults in China, clinical specimens were collected from 10,310 adults with ARTIs from May 2005 to July 2010. The partial HAdV hexon gene was amplified by polymerase chain reaction (PCR), sequenced, and phylogenetically analyzed. HAdVs were detected in 86 samples (0.8%), of which 67 (77.9%) were species B (HAdV-3, -7, -11, and -14), 7 (8.1%) were species C (HAdV-1, -2, and -6), and 12 (14%) were species E (HAdV-4). HAdV-3 was the most frequently detected serotype (41/86, 47.7%), followed by HAdV-7 (13/86, 15.1%), HAdV-4 (12/86, 14.0%), and HAdV-11 (11/86, 12.8%). Patients 14-25 years old (60.5%) exhibited a higher rate of adenovirus detection than older patients. Co-infections with other respiratory viruses were observed in samples positive for HAdV species B and E. Human rhinovirus was the most commonly found virus in patients with HAdV infection. These findings provide baseline data for the surveillance and control of HAdV infection in China.

Detection of human bocavirus 3 in China.

Since its first identification in 2005, four species of human bocavirus (HBoV1-4) have been documented. HBoV1 and HBoV2 have been shown to be associated with respiratory tract illnesses, as well as with acute gastroenteritis (AGE), worldwide. However, reports on the prevalence, clinical significance, and molecular characteristics of the two most newly identified HBoV species, HBoV3 and HBoV4, are very limited. To detect and characterize HBoV3 and HBoV4 infections in children with AGE in China, stool specimens were collected from 366 children with AGE. HBoVs in these samples were amplified by nested polymerase chain reaction (PCR), sequenced, and phylogenetically analyzed. HBoVs were detected in 44 samples (12%), of which nine were HBoV1, 33 were HBoV2, and two were HBoV3. HBoV4 was not detected. Most HBoV-positive samples (35/44) were co-detected with other viral pathogens. Both HBoV3 samples were co-detected with rotavirus. Analysis of the HBoV3 (46-BJ07) genome sequence indicates that HBoV3 may be a recombinant derived from HBoV1 and HBoV2 or from HBoV1 and HBoV4. To our knowledge, this is the first report of HBoV3 in China. However, it is unclear whether HBoV3 is associated with AGE because of its low detection rate in AGE patients and its co-infection with other AGE-causing viruses.

Evaluation of real-time nucleic acid sequence-based amplification for detection of Chikungunya virus in clinical samples.

The Chikungunya virus (CHIKV) is a member of the genus Alphavirus that is transmitted to humans by Aedes mosquitoes. In 2005 and 2006, the Indian Ocean island of La Réunion was hit with an unprecedented CHIKV fever outbreak that infected 300 000 people. In the present study, we describe the evaluation of real-time nucleic acid sequence-based amplification (RT-NASBA) for the detection of CHIKV in clinical samples. A co-extracted and co-amplified chimerical CHIKV RNA sequence was used as an internal control to eliminate false-negative results. The detection threshold of the assay was determined from quantified CHIKV-positive plasma, and estimated to be 200 copies per NASBA reaction. The specificity of the assay was determined using blast analyses and non-cross-reactivity using an O'nyong-nyong virus culture and 250 CHIKV RT-PCR-negative plasma samples. A 100 % specificity was found and no invalid result was obtained, showing the good quality of the nucleic acid extraction. The assay was then evaluated using 252 CHIKV-positive RT-PCR plasma samples. The samples were all tested positive, including those with low viral load. This evaluation showed that the RT-NASBA is a rapid (5 h from sample nucleic acid extraction to detection), sensitive, specific and reliable method for the routine diagnosis of CHIKV in clinical samples.

Absence of hybridization with the wild-type and mutant rpoB probes in the Genotype MTBDRplus assay detects 'disputed' rifampicin mutations.

OBJECTIVE: Mycobacterium tuberculosis isolates that fail to hybridize to at least one rpoB wild-type or any mutation probe on the Genotype MTBDRplus strip are assumed to be rifampicin-resistant. However, the precise mutation(s) are unknown. We sought to identify the mutations in isolates with such hybridization patterns and determine if the mutations are associated with resistance to rifampicin. METHODS: In this study, 275 M. tuberculosis isolates were screened with the Genotype MTBDRplus assay to identify isolates with the hybridization pattern. These isolates were sequenced and their minimum inhibitory concentrations (MIC) determined using the Bactec MGIT 960 system. RESULTS: Among the 275 isolates tested, 15 (6%) isolates with the hybridization pattern were identified. Sequencing showed that failure to hybridize to rpoB wild-type probes resulted from the presence of 'disputed' rifampicin mutations, which are mutations not always associated with a rifampicin-resistant phenotype. All, except 3/15, isolates had a rifampicin-resistant phenotype (MIC > 1 µg/mL). One of the three isolates with a rifampicin-susceptible phenotype had the same mutation at position 526 (His526Leu) as another isolate that had a rifampicin-resistant phenotype. CONCLUSION: The recommendation of the Genotype MTBDRplus assay to assume rifampicin resistance based solely on failure to hybridize to rpoB wild-type probe allows the identification of important RIF-resistant isolates. About 20% (3/15) of such isolates could be missed by relying only on the standard MGIT 960 DST assay for drug susceptibility testing.

Antioxidant response modulated by copper in healthy or parasitized carp (Cyprinus carpio L.) by Ptychobothrium sp. (Cestoda).

An increased antioxidant response (catalase, glutathione S-transferase (GST) and glutathione reductase (GRd) activities in liver and GST activity in head kidney) was observed in carp parasitized by Ptychobothrium sp. compared to healthy fish. In case of a copper contamination of these fish, the decrease in enzymatic activities observed was less pronounced in parasitized than in healthy carp.

Basic nuclear proteins of the histone-less eukaryote Crypthecodinium cohnii (Pyrrhophyta): two-dimensional electrophoresis and DNA-binding properties.

Unlike typical eukaryotes, the Dinoflagellate Crypthecodinium cohnii does not contain histones but six major basic, low molecular weight nuclear proteins which represent only 10% of the DNA mass and differ from histones in their electrophoretic and DNA-binding properties. These proteins are resolved in two-dimensional electrophoresis (AUT-PAGE x SDS-PAGE). Three proteins with an apparent molecular mass of 16, 16.5 and 17 kDa (p16, p16.5 and p17) are present in addition to the major 14 kDa basic nuclear component (HCc). HCc itself is resolved in three proteins (alpha, beta and gamma). When the proteins are not reduced with 2-mercaptoethanol before 2D-PAGE, the migration of HCc alpha, beta and gamma is modified in a way which suggests the formation of both inter- and intramolecular disulfide bridges and thus, the presence of at least two cysteines. The amino-acid analysis of HCc proteins resolved in 2D gels confirms that they are lysine-rich. HCc alpha, beta and gamma as well as p16, p16.5 and p17 are removed from isolated chromatin with 0.6 M NaCl, indicating that their affinity for DNA in vivo is lower than that of core histones. Furthermore, in vitro, they bind more tightly to single-stranded than to double-stranded DNA.

Prevalence and persistence of antibody titers to recombinant HIV-1 core and matrix proteins in HIV-1 infection.

Numerous studies have established the correlation between antibodies to the core protein p24 of HIV-1 and the progression of the acquired immunodeficiency syndrome. In this study, we analyzed the immune response to two recombinant gag proteins, p24 and p17, in order to evaluate their diagnostic or prognostic significance. Immune response to the immunodominant domain of the transmembrane glycoprotein gp41 was used as a reference. Sera collected from individuals from France and Burundi (Central Africa) at various CDC stages of HIV-1 infection were tested using three sandwich enzyme-linked immunoassays developed with a synthetic peptide corresponding to the immunodominant domain of gp41, SP gp41, or recombinant p24 and p17 cloned and expressed in Escherichia coli. These assays allowed detection of titer antibodies to the three cited antigens. Antibodies to SP gp41 were detected in every HIV-1-positive patient from France and Burundi, generally at a high and stable level. Results obtained with p24 confirmed the value of antibodies to p24 as a prognostic marker only in European and North American populations, since the African population had very high levels of these antibodies even at an advanced stage of the disease. They also confirmed that initial antibody response to p24 is more predictive of outcome than antibody titer change over time. Although antibodies to p17 decline during progression to AIDS, they are frequently absent in French patients at early, asymptomatic stages and therefore could not be used as a prognostic marker. In contrast, antibodies to p17 are significantly less common in African patients with AIDS when compared with symptomless HIV-1-infected African individuals.(ABSTRACT TRUNCATED AT 250 WORDS)

Diagnosis of zoonotic viral encephalitis.

Human infections by zoonotic encephalitis viruses are usually asymptomatic or symptoms are not specific to these viruses. Some of them have high mortality and morbidity rates and most often no specific treatment exist. This emphasizes the need for a precise identification of arboviruses in clinical specimens from humans and animals. Because these diseases are frequent in developing countries and tend to emerge or re-emerge in others, diagnostic tools must detect the broadest possible range of viruses with a high sensitivity and this is a key factor for surveillance, control of transmission and prevention through vaccination. In countries with limited diagnostic infrastructures, low-cost and easy-to-use tests are required. The diagnosis of arboviral encephalitis has been significantly improved in the recent years. Sensitive ELISA assay to detect antibodies against many arboviruses in serum or CSF are commercially available and can be used to detect early infections. Immunochromatographic rapid tests for the detection of specific IgM that could be used on fingertip blood would be valuable tools in developing countries. A limitation of these serologic assays is their lack of specificity as many arboviruses are antigenically related. Virus isolation or molecular assays from different human or animal tissues are also important diagnostic tools. Molecular assays have been extensively described in the recent years. They are very sensitive and have the advantage over cell culture that specimen transportation is less critical. Real-time detection has even improved sensitivity and reduced time-to-result. Although the utility of molecular assays for the detection of arboviruses in mosquito pools has been demonstrated, an extensive validation of their pertinence in clinical settings is still required. The use of DNA-microarrays may further extend the range of viruses that can be detected in a single test and allow isolates typing for epidemiology purposes.

Determination of anti-Toxoplasma gondii immunoglobulin G avidity: adaptation to the Vidas system (bioMérieux).

The determination of specific anti-Toxoplasma gondii IgG avidity has been proposed to improve the determination of the date of toxoplasmic seroconversion in pregnant women. In this study, we adapted this serological technique to the Vidas system (bioMérieux) using 6 M urea as the dissociating agent. We studied 356 sera, including 42 sequential sera from sero-conversions in pregnant women. Our results show that the test is easy to use, and that an avidity index higher than 0.300 allows the exclusion of a recent infection acquired less than 4 months before serum sampling.

Characterisation of the yeast Pichia membranifaciens and its possible use in the biological control of Botrytis cinerea, causing the grey mould disease of grapevine.

Pichia membranifaciens strain FY-101, isolated from grape skins, was found to be antagonistic to Botrytis cinerea, the causal organism of the grey mould disease of the grapevine. When grown together on solid as well as liquid media, the yeast brings about the inhibition of this parasitic fungus, coagulation and leakage of its cytoplasm, and suppression of its ability to produce the characteristic grey mould symptoms on the grapevine plantlets. In vitro experiments confirm that this yeast can be used as a biological control organism against B. cinerea. An account of the molecular characterisation of P. membranifaciens (complete sequence of the ITS region of its ribosomal DNA, GenBank accession No. AF 270935), as well as the interaction between B. cinerea and the yeast, are given here.

Phytotoxicity of diuron alone and in combination with copper or folpet on duckweed (Lemna minor).

The photosystem II-herbicide diuron is widely used for weed control in Champagne's vineyards. Its important use and its relative persistence make it of particular interest for ecotoxicological studies. Toxicity of diuron was assessed on Lemna minor L., a representative aquatic macrophyte regularly used for toxicological studies. Toxicity assessments were based on inhibition of growth and total chlorophyll content of L. minor cultures after 7 days. Growth was inhibited and IC(50) and IC(90) were, respectively, 25 and 60 microg l(-1), but chlorophyll content of L. minor increased in response to the herbicide. When diuron was combined with copper, growth inhibition of L. minor depended on the concentrations of both chemicals. For some concentrations, combination of these chemicals resulted in a slight (but non-significant) antagonism. Additivity was observed for all other mixtures. When diuron was combined with folpet, growth and chlorophyll content of L. minor only depended on the concentration of the herbicide. Diuron was also found to prevent the copper-induced decrease of chlorophyll content when it was combined with this metal. A multifactorial model was found more appropriate to characterize interactions between pesticides than Abott's model.

Effects of procymidone, fludioxonil and pyrimethanil on two non-target aquatic plants.

Procymidone, fludioxonil, and pyrimethanil are widely used to control the pathogenic fungus Botrytis cinerea in Champagne's vineyards. These fungicides may end up in surface waters and present potential risks for aquatic vascular plants and algae. Therefore, their toxicity was evaluated on Lemna minor and Scenedesmus acutus in six-day or 48-h tests, respectively. Based on growth and chlorophyll (Chl) content of L. minor and S. acutus cultures, the results showed that the alga was the most sensitive to the fungicides. Among the fungicides, pyrimethanil was the most toxic for L. minor, its nominal IC50 was 46.16 mg l(-1) and that of the other two was >100 mg l(-1). In contrast, pyrimethanil appeared the least toxic for S. acutus at low concentration, nominal IC50 were 22.81, 4.85, and 4.55 mg l(-1) for pyrimethanil, fludioxonil, and procymidone, respectively. Fate of the fungicides in the media was also investigated and acute toxicity of the agrochemicals is discussed in regard to concentration in the culture media. Poor solubility of procymidone and fludioxonil appeared to be partly responsible for the low toxicity of these fungicides. Based on these toxicity data and the concentrations found in ponds collecting vineyard runoff water, these pesticides should not impair the establishment of pioneer plants.

[Agranulocytosis under metapramine. A "well-known" effect without bibliographical data]. / Agranulocytose sous métapramine. Un effet "notoire" sans données bibliographiques.

Metapramine (Timaxel) is a tricyclic antidepressant drug devoid of anticholinergic properties. As other psychotropic drugs, it may induce hyperthermia and several case-reports have been published. Agranulocytosis is an exceptional side-effect of this drug. Even if several cases have been reported to Pharmacovigilance structures in France, as far as we know there is no publication of this unwanted effect. The authors reported the case of an old woman who has presented two episodes of fever and asthenia. Blood formula was both times characterized by agranulocytosis without anemia nor thrombocytopenia. The accidental rechallenge of metapramine permitted to conclude to a causal relationship between these hematological events and metapramine intake.

[Chronic cryptogenic pneumonia]. / Pneumonie chronique cryptogénique.

We report a case of a 72 year old man with the appropriate criteria for the recently identified chronic cryptogenic pneumonia: dyspnoea, cough, low general state, fever, raised sedimentation rate, localised opacities on the chest x-ray; no cause has been identified; the patient will improve on steroid therapy even though antibiotics are without effect, but relapses on stopping the steroids. A lung biopsy shows a predominant intra-alveolar fibrosis. A definitive cure can be obtained by 12 weeks of steroid therapy.

[Erythrosis interfollicularis colli de Leder]. / L'erythrosis interfollicularis colli de leder

The authors have studied clinically and histologically two cases of " erythrosis interfollicularis colli de Leder ". It's a frequent affection, principally between 40 and 50 years of age but not very known because the patients rarely consult for it. Clinically they insist on the respect of a triangulary and lozengical median zone, and on the sight in "plucked chicken skin"; histologically on the dilatation of sebaceous glands, the frequency of a folliculary corneous plug and the elastic tissue alterations. On the etiological plan, the part of the age, the sun exposure, and in their two cases on the possible photosensibilisative part of the scents.

[Familial polycythemia caused by high oxygen affinity hemoglobin. 2nd world case of hemoglobin J Providence]. / Polyglobulie familiale par hémoglobine hyperaffine pour l'oxygène. Deuxième cas mondial d'hémoglobine J Providence.

The cas observed in Auvergne and reported here raises the aetiological problem of polycythaemias. Young subjects with polycythaemia should be investigated for congenital anomaly of oxygen transport by measuring P50 and 2,3-DPG, which provides information on the oxygen-carrying capacity of haemoglobin. When confronted with familial polycythaemia due to high oxygen affinity haemoglobin, clinicians must know that a cause-effect relationship is not always demonstrable since other factors, such as tobacco-smoking in this particular case, may intervene.

Metallothionein induction in the aquatic oligochaete Tubifex tubifex exposed to the herbicide isoproturon.

Metallothioneins (MTs), are low molecular weight proteins, mainly involved in metal ion detoxification. Recently it has been demonstrated that MTs participate in several cellular functions such as regulation of growth, and anti-oxidative defenses. Moreover, pesticides can induce their synthesis. The aim of the current work was to determine the effects of isoproturon either pure or formulated as Matin (suspension containing 500 g x l(-1) isoproturon) on the metallothionein and total protein content of the aquatic worm Tubifex tubifex. MT levels in exposed worms increased significantly after 7 and 15 days of exposure to 50 mg x l(-1) of herbicide. Isoproturon reduced metal (Cu, Zn, and Cd) content of metallothioneins, and it also increased total protein content of the worms. The results suggest that MT induction may not be considered as a specific biomarker of metal exposure but it can be used as a non specific biomarker of isoproturon effect in the worm.

Stepwise implementation of a new diagnostic algorithm for multidrug-resistant tuberculosis in Haiti.

SETTING: The uptake of tests endorsed by the World Health Organization to detect and appropriately confirm multidrug-resistant tuberculosis (MDR-TB) in low-income countries remains insufficient. OBJECTIVE: To validate the implementation of line-probe assays (LPA) and liquid culture to develop an algorithm to detect MDR-TB in the challenging setting of Haiti. METHODS: Through an EXPAND-TB (Expanding Access to New Diagnostics for TB) partnership, proficiency testing and validation of 221 acid-fast bacilli positive specimens were performed. Sensitivity, cost and processing time were analysed. RESULTS: Using liquid vs. solid culture shortened the turnaround time from 54 to 19 days, with a sensitivity of 100% vs. 98.6% and a total cost reduction of 13%. LPA detected all TB and MDR-TB cases at a lower cost than culture, in a mean time of 7.5 days. CONCLUSION: The combined use of molecular and liquid culture techniques accelerates the accurate diagnosis of TB and susceptibility testing against first-line drugs in a significantly shorter time, and is less expensive. The implementation of this new algorithm could significantly and accurately improve the screening and treatment follow-up of patients affected with TB and MDR-TB.

Laboratory-based diagnosis of pneumococcal pneumonia: state of the art and unmet needs.

In view of the increasing use of pneumococcal vaccines, especially in the developing world, there is a need for appropriate diagnostics to understand the aetiology of pneumonia, to define the burden of pneumococcal disease, and to monitor vaccine efficacy and effectiveness. This article summarizes a meeting on the diagnosis, detection and serotyping of pneumococcal disease organized by PATH and Fondation Mérieux (18-20 October 2009, Fondation Mérieux Conference Centre, Les Pensières, France). Workers and experts met to discuss the gaps in the microbiology-based diagnosis of Streptococcus pneumoniae disease, with special emphasis on pneumonia. The meeting was designed to evaluate the state of the art of pneumococcal diagnostics and serotyping methodologies, identify research and development needs, and propose new guidelines to public health authorities to support the introduction of vaccines. Regarding detection, the main recommendations were to encourage chest X-rays and antigen detection in urine. Large-scale studies are needed to evaluate the diagnostic utility of test algorithms that associate chest X-rays, antigen detection in urine, S. pneumoniae quantitative PCR in nasopharyngeal aspirates and sputum, and C-reactive protein or procalcitonin measurement in blood. Efforts should be focused on proteomics to identify pneumococcus-specific antigens in urine or host markers in blood expressed during pneumonia. It was recommended to develop S. pneumoniae typing capacities, to understand the epidemiology of pneumococcal disease, and to evaluate vaccine effectiveness. Simple and effective approaches are encouraged, and new technologies based on beads, microarrays or deep sequencing should be developed to determine, in a single test capsular serotype, resistance profile and genotype.

Afriflu--international conference on influenza disease burden in Africa, 1-2 June 2010, Marrakech, Morocco.

The burden of influenza disease is to a large extent unknown for the African continent. Moreover, the interaction of influenza with common infectious diseases in Africa remains poorly described. Solid scientific evidence on the influenza disease burden in Africa is critical for the development of effective influenza vaccine policies. On 1st and 2nd June 2010 in Marrakech, Morocco, over eighty surveillance and influenza experts from 22 African countries as well as Europe and America met at the 'Afriflu' conference to discuss influenza challenges and solutions for the continent. During the meeting, participants exchanged their experiences and discussed a wide variety of topics related to influenza in Africa, including diagnosis, surveillance, epidemiology, and interventions. The meeting concluded with a pledge to improve influenza knowledge and awareness in Africa, with an emphasis on accurate determination of disease burden to help orient public health policies.