Rapid determination of macrolide and lincosamide resistance in group B streptococcus isolated from vaginal-rectal swabs.

OBJECTIVE: Our objective was to assess the ability of real-time PCR to predict in vitro resistance in isolates of group B streptococcus (GBS). METHODS: The first real-time PCR assays for the genes known to confer resistance to erythromycin and clindamycin in GBS were developed. Three hundred and forty clinical GBS isolates were assessed with these assays and compared with conventional disk diffusion. RESULTS: The presence of an erythromycin ribosome methylation gene (ermB or ermTR variant A) predicted in vitro constitutive or inducible resistance to clindamycin with a sensitivity of 93% (95% CI 86%-97%), specificity of 90% (95% CI 85%-93%), positive predictive value of 76% (95% CI 67%-84%), and negative predictive value of 97% (95% CI 94%-99%). CONCLUSION: This rapid and simple assay can predict in vitro susceptibility to clindamycin within two hours of isolation as opposed to 18-24 hours via disk diffusion. The assay might also be used to screen large numbers of batched isolates to establish the prevalence of resistance in a given area.

The cotton rat: an underutilized animal model for human infectious diseases can now be exploited using specific reagents to cytokines, chemokines, and interferons.

The cotton rat represents the best or only animal model for a large number of human infectious diseases, and it may be unique among small laboratory animals in its susceptibility to several potential agents of bioterrorism. Although the cotton rat is a reliable model to define pathologic changes produced during infection with human pathogens, the lack of specific reagents has precluded a more extensive analysis of the molecular basis of pathogenesis. Here, we report the cloning of 24 cotton rat genes encoding various cytokines, chemokines, and interferons (IFNs). Analysis of the expression of most of these genes was performed by RT-PCR in cotton rat macrophages during treatment with lipopolysaccharide (LPS) and in cotton rat lungs after infection with influenza virus. The availability of these reagents will provide the tools for molecular analysis of pathogenesis and immune responses to a wide variety of pathogens and set the basis for the development of new prophylactic and therapeutic strategies against human infectious diseases.

Respiratory syncytial virus (RSV) infection induces cyclooxygenase 2: a potential target for RSV therapy.

Cyclooxygenases (COXs) are rate-limiting enzymes that initiate the conversion of arachidonic acid to prostanoids. COX-2 is the inducible isoform that is up-regulated by proinflammatory agents, initiating many prostanoid-mediated pathological aspects of inflammation. The roles of cyclooxygenases and their products, PGs, have not been evaluated during respiratory syncytial virus (RSV) infection. In this study we demonstrate that COX-2 is induced by RSV infection of human lung alveolar epithelial cells with the concomitant production of PGs. COX-2 induction was dependent on the dose of virus and the time postinfection. PG production was inhibited preferentially by NS-398, a COX-2-specific inhibitor, and indomethacin, a pan-COX inhibitor, but not by SC-560, a COX-1-specific inhibitor. In vivo, COX-2 mRNA expression and protein production were strongly induced in the lungs and cells derived from bronchioalveolar lavage of cotton rats infected with RSV. The pattern of COX-2 expression in vivo in lungs is cyclical, with a final peak on day 5 that correlates with maximal histopathology. Treatment of cotton rats with indomethacin significantly mitigated lung histopathology produced by RSV. The studies described in this study provide the first evidence that COX-2 is a potential therapeutic target in RSV-induced disease.

The cotton rat provides a useful small-animal model for the study of influenza virus pathogenesis.

Influenza A virus continues to cause annual epidemics. The emergence of avian viruses in the human population poses a pandemic threat, and has highlighted the need for more effective influenza vaccines and antivirals. Development of such therapeutics would be enhanced by the use of a small-animal model that is permissive for replication of human influenza virus, and for which reagents are available to dissect the host response. A model is presented of nasal and pulmonary infection in adult inbred cotton rats (Sigmodon hispidus) that does not require viral 'adaptation'. It was previously demonstrated that animals infected intranasally with 10(7) TCID50 of a recent H3N2 influenza, A/Wuhan/359/95, have increased breathing rates. In this report it is shown that this is accompanied by weight loss and decreased temperature. Virus replication peaked within 24 h in the lung, with peak titres proportional to the infecting dose, clearing by day 3. Replication was more permissive in nasal tissues, and persisted for 6 days. Pulmonary pathology included early bronchiolar epithelial cell damage, followed by extensive alveolar and interstitial pneumonia, which persisted for nearly 3 weeks. Interleukin 1 alpha (IL1alpha), alpha interferon (IFN-alpha), IL6, tumour necrosis factor alpha (TNF-alpha), GROalpha and MIP-1beta mRNA were elevated soon after infection, and expression coincided with virus replication. A biphasic response was observed for RANTES, IFN-gamma, IL4, IL10 and IL12-p40, with increased mRNA levels early during virus replication followed by a later increase that coincided with pulmonary inflammation. These results indicate that cotton rats will be useful for further studies of influenza pathogenesis and immunity.

Comparison of corticosteroids for treatment of respiratory syncytial virus bronchiolitis and pneumonia in cotton rats.

Triamcinolone acetonide, methylprednisolone, and dexamethasone were each evaluated in combination with palivizumab (Synagis) for the therapy of established respiratory syncytial virus infection in the cotton rat. Triamcinolone and methylprednisolone proved to be more effective than dexamethasone in reducing lung pathology. No recurrence of viral replication or pulmonary pathology followed the cessation of therapy.

Cytokine and chemokine gene expression after primary and secondary respiratory syncytial virus infection in cotton rats.

The induction of pro- and anti-inflammatory cytokines and chemokines was studied in the lungs of cotton rats after primary or secondary infection with respiratory syncytial virus (RSV). Increases in messenger RNA (mRNA) levels of all genes analyzed were observed during the course of primary infection. In general, mRNA expression peaked between postinfection days 1 and 4 and returned to near-normal levels by day 10. During secondary infection, the expression of some genes (i.e., interferon [IFN]-gamma and interleukin [IL]-10) began earlier, some (i.e., IL-1beta and macrophage inflammatory protein-1beta) began later, and some (i.e., IL-1beta, IL-10, growth-regulated protein, and tumor necrosis factor-alpha) showed prolonged expression, whereas 2 genes (i.e., IFN-alpha and IL-6) were not expressed. This study presents evidence of different kinetics of expression of inflammatory mediators during primary and secondary infection that likely coincide with innate and adaptive immune response and complement previous observations that emphasize the role of inflammation in the pathogenesis of RSV disease.