Rapid identification of Rhodococcus equi by a PCR assay targeting the choE gene

The actinomycete Rhodococcus equi is an important pathogen of horses and an emerging opportunistic pathogen of humans. Identification of R. equi by classical bacteriological techniques is sometimes difficult, and misclassification of an isolate is not uncommon. We report here on a specific PCR assay for the rapid and reliable identification of R. equi. It is based on the amplification of a fragment of the choE gene encoding cholesterol oxidase. The choE-based PCR was assessed by using a panel of strains comprising 132 isolates from different sources and of different geographical origins, all initially identified biochemically as R. equi, and 30 isolates of representative non-R. equi actinomycete species, including cholesterol oxidase producers. The expected 959-bp amplicon was observed only with R. equi isolates, as confirmed by sequencing of a variable region of the 16S RNA gene from a random sample of 20 PCR-positive isolates. All R. equi isolates gave a positive choE-based PCR result, which correlated with a high degree of conservation of the choE gene. Three of the 132 strains originally identified as R. equi were negative for the choE gene, and subsequent analysis of their 16S RNA gene sequences confirmed that they belonged to other bacterial species (Dietzia maris, Mycobacterium peregrinum, and Staphylococcus epidermidis). All non-R. equi isolates were negative by the choE-based PCR. ATCC 21387, the only known isolate of Brevibacterium sterolicum, gave a 959-bp amplicon whose DNA sequence was virtually identical to that of R. equi choE. Comparison of the 16S RNA genes indicated that ATCC 21387 should be considered an R. equi isolate.     

Connectivity patterns in tuberculosis and leprosy patients are indistinguishable from that of healthy donors

Connectivity, the self-defined interactions between antigen-recognising molecules in a network system can in part be assessed by measuring the reactivity of a given serum against an ordered set of immunoglobulin (Ig)G F(ab')2 fractions, separated by means of isoelectric focusing so that, the serum reactivity against the whole set of fractions defines a characteristic pattern of connectivity. Deviations from the normal condition (healthy donors) have so far been documented for two autoimmune diseases: systemic lupus erythematosus (SLE) and pemphigus vulgaris, as well as for human immunodeficiency virus (HIV)-1 infection. We tested here if bacterial infections lead to alterations in connectivity. In addition, we wanted to test if two antigenically related bacteria would produce similar or otherwise distinctive connectivity patterns. Connectivity analysis was applied on the sera from tuberculosis and leprosy patients and the sera from healthy donors were used as control. No statistically significant differences between the three groups studied were found. These results have implications for theories that set the origin of autoimmune diseases in microbial infections. To the best of our knowledge, this is the first attempt to analyze the connectivity status in bacterial infections.     

The C-terminal domain of the pVP2 precursor is essential for the interaction between VP2 and VP3, the capsid polypeptides of infectious bursal disease virus

The interaction between the infectious bursal disease virus (IBDV) capsid proteins VP2 and VP3 has been analyzed in vivo using baculovirus expression vectors. Data presented here demonstrate that the 71-amino acid C-terminal-specific domain of pVP2, the VP2 precursor, is essential for the establishment of the VP2-VP3 interaction. Additionally, we show that coexpression of the pVP2 and VP3 polypeptides from independent genes results in the assembly of virus-like particles (VLPs). This observation demonstrates that these two polypeptides contain the minimal information required for capsid assembly, and that this process does not require the presence of the precursor polyprotein.     

Tumor necrosis factor-alpha -308 promoter polymorphism contributes independently to HLA alleles in the severity of rheumatoid arthritis in Mexicans

The aim of this study was to determine the frequency and potential relevance of the promoter polymorphisms of the tumor necrosis factor-alpha (TNF-alpha) in the severity of rheumatoid arthritis (RA) in Mexicans. HLA-DR and polymorphisms at positions -238 and -308 of TNF-alpha gene were determined in 137 Mexican RA patients (44 with severe and 93 with non-severe RA) as well as in 169 healthy controls (99 were typed for HLA-DR). We observed an increased frequency of HLA-DR4 in severe RA compared to healthy controls (pC=0.02, OR=2.33). TNF polymorphism analysis showed a significant increased frequency of TNF -238 GG genotype in the whole group of RA patients when compared to healthy controls (pC=0.007, OR=4.71). When the analyses were carried out separately in severe and non-severe RA patients, the increased frequency of -238 GG genotype only was observed in patients with non-severe forms of the disease. Analysis of -308 polymorphism showed increased frequency of -308 T2 (A) allele in severe RA when compared to non-severe disease (pC=0.011, OR=3.29) and to healthy controls (pC=0.002, OR=3.97). The data demonstrate that -308 T2 (A) allele is associated with susceptibility to develop severe RA in Mexicans. This association could be independent from HLA-DR alleles and might be used as a prognostic marker for severe RA.     

Molecular characterization and genetic variation of the VP7 gene of human rotaviruses isolated in Paraguay

Rotavirus is the main cause of acute diarrhea in infants and young children worldwide. In Paraguay, acute diarrhea ranks fourth among the causes of mortality in children under 4 years of age. Rotavirus was detected in 93 out of 410 patients admitted to three main hospitals in Asunción, Paraguay from August 1998 to August 2000. Samples from 64 patients were analyzed by RT-PCR for G and P typing. G4P[8] (46.9%; 30/64) was the most common strain detected, followed by G9P[8] (17.2%; 11/64) and G1P[8] (10,9%; 7/64). Since G4 was predominant during the epidemiological peaks of 1998 and 1999, four samples were sequenced and all grouped into sublineage Ic. This sublineage was reported for the first time in 1998 in Argentina, southern border of Paraguay, and it was shown to be responsible for the increased prevalence of G4 during the epidemiological season of 1998 in that country. In addition, Paraguayan G1 strains grouped in different lineages (I and II). However, G9 was predominant during the rotavirus epidemiological peak of 2000, and phylogenetic analysis of five samples grouped into a common emergent/reemergent lineage that circulates worldwide. Since vaccination could reduce the severity and the number of cases of rotavirus disease, this study suggests that a vaccine containing recently isolated variants of the most prevalent types (G1-G4) together with the emerging G9 type, might be sufficient to elicit a protective immune response against the rotavirus types circulating currently in Paraguay.     

Nocardia ignorata, a new agent of human nocardiosis isolated from respiratory specimens in Europe and soil samples from Kuwait

Nocardia ignorata is a recently described species identified on the basis of a single isolate of unknown origin. Here we describe the epidemiological, phenotypic, and phylogenetic characteristics of this new species, based on five new clinical and soil isolates.     

Comparative evaluation of the effect of balloon dilatation by exercise precordial mapping and exercise 201-thallium scintigraphy

Precordial exercise electrocardiographic mapping and exercise 201-thallium scintigraphy were simultaneously used on sixteen patients before, shortly after and three months after percutaneous transluminal coronary angioplasty (PTCA) to evaluate the immediate and late results of the intervention. In spite of the limited number of observations, positive exercise precordial mapping and or a reversible 201-thallium perfusion defect after PTCA seem to be indicative of failure of balloon dilatation or of development of restenosis in a given coronary artery. "Reverse minimum", that is the minimum in the resting precordial map which disappears on exercise, may be a sign of segmental coronary artery occlusion and well functioning collateral supply.     

Potentiation of acute opioid-induced respiratory depression and reversal of tolerance by the calcium antagonist nimodipine in awake rats

Summary The interaction between sufentanil, a -opioid agonist, and the Ca²⁺ antagonist nimodipine on respiration and on the development of opioid tolerance in awake rats has been analyzed. Our previous work demonstrated that chronic treatment with nimodipine together with sufentanil increases the analgesic potency of the opioid 50 fold. Therefore, we have investigated whether the opioid-induced respiratory depression is potentiated in parallel with the analgesia. Ventilation was measured by the whole body plethysmographic method. In naive rats, sufentanil (10–80 g/kg) consistently induced a dose-dependent respiratory depression. Pretreatment with nimodipine (200 g/kg) potentiated this effect but to a lesser extent than it potentiated analgesia. After chronic administration of the opioid (2 g/h, 7 days) tolerance was manifested as a reduction in both the area under the time course curve and in the maximum effect. Nimodipine (1 g/h) administered concurrently with sufentanil for 7 days counteracted the tolerance to respiratory depression but no additional potentiation was observed. These results demonstrate that the interaction between nimodipine and sufentanil is not limited to antinociception but also exends to respiratory depression. However, compared with analgesia, the clinical relevance of a potential increase in opioid-induced respiratory depression by nimodipine may be negligible.Correspondence to: M. A. Hurlé at the above address