Characterization of Escherichia fergusonii and Escherichia albertii isolated from water.

AIMS: The aim of this study was to characterize Escherichia fergusonii and Escherichia albertii isolated from water. METHODS AND RESULTS: The characterization of E. fergusonii and E. albertii isolated from water was determined using an Escherichia coli-specific uidA PCR, a tuf PCR, and with phylogenetic analysis using three housekeeping genes (adk, gyrB, and recA) from the E. coli MLST scheme, selected for their ability to discriminate among all Escherichia species. Among the 527 isolates tested, 25 (4·7%) were uidA PCR negative and tuf PCR positive. Phylogenetic analysis using adk, gyrB and recA genes showed that 6, 18 and 1 of these 25 non-E. coli Escherichia spp. isolates grouped with reference strains of E. fergusonii, E. albertii, and E. coli, respectively. Finally, the 25 non-E. coli Escherichia spp. strains isolated were investigated for the presence of pathogenic factors, comprising intimin (eae gene), cytolethal distending toxin (cdtB gene) and shiga toxin (stx gene). With the PCR primers used, the presence of eae and stx genes was not detected. However, cdtB genes types I/IV were detected for 3 (16·7%) E. albertii strains, whereas 15 of 18 (83·3%) possessed the cdtB gene types II/III/V. CONCLUSIONS: These results showed that MLST scheme allows a more accurate identification of non-E. coli species than phenotypic tests. We also showed that E. fergusonii and E. albertii represent, respectively, 0·8 and 2·5% of all Escherichia species isolated and the pathogenic cdtB genes were present in 83·3% of these strains. SIGNIFICANCE AND IMPACT OF THE STUDY: The data presented in this study provided an efficient way to correctly identify non-E. coli species contributing to our understanding of the risks associated with Escherichia species in water consumed by humans and animals. Furthermore, the results give an insight about the natural habitats of these species.

Development of a real-time PCR assay for the specific detection and identification of Streptococcus pseudopneumoniae using the recA gene.

We sequenced the evolutionarily conserved genes 16S rRNA, atpD, tuf, and recA from Streptococcus pseudopneumoniae, Streptococcus pneumoniae, Streptococcus mitis, and Streptococcus oralis. Phylogenetic analysis revealed that recA provided good resolution between these species, including discrimination of the novel species S. pseudopneumoniae. By contrast, the more conserved 16S rRNA, tuf and atpD are not sufficiently discriminatory. Therefore, recA sequences were used to develop a real-time PCR assay with a locked nucleic acid-mediated TaqMan probe for the specific detection and identification of S. pseudopneumoniae. The PCR assay showed excellent specificity and a detection limit of <10 genome copies for the detection and identification of S. pseudopneumoniae strains, which makes it a promising tool for molecular identification and epidemiological studies. In conclusion, this article describes for the first time a PCR assay for the specific identification of S. pseudopneumoniae.

Toward Automatic Label-Free Whispering Gallery Modes Biodetection with a Quantum Dot-Coated Microsphere Population.

We explore a new calibration-free approach to biodetection based on whispering gallery modes (WGMs) without a reference measure and relative shifts. Thus, the requirement to keep track of the sensor position is removed, and a freely moving population of fluorophore-doped polystyrene microspheres can now fulfill this role of sensing resonator. Breaking free from fixed surface-based biosensing promotes adhesion between the microsphere sensors and the analytes since both can now be thoroughly mixed. The 70-nm-wide spectrum of green fluorescent microbeads allows us to monitor over 20 WGMs simultaneously without needing evanescent light coupling into the microspheres, hence enabling remote sensing. Since the exact radius of each microsphere is unknown a priori, it requires algorithmic analyses to obtain a reliable result for the refractive index of a solution. We first test our approach with different solutions of alcohol in water obtaining 3 x 10(-4) precision on the refractive index at lower concentrations. Then, the solutions of bacterial spores in water yield clear evidence of biodetection in the statistical analysis of WGMs from 50 microspheres. To extend the fluorescence spectral range of our WGM sensors, we present preliminary results on coating microspheres with CdSe/ZnS quantum dots.

Diagnosing infections--current and anticipated technologies for point-of-care diagnostics and home-based testing.

In recent years, we have witnessed many transitions in healthcare systems around the globe. For example, population expansion and ageing, and the human immunodeficiency virus (HIV)-AIDS epidemics, have exerted pressure to decentralize the practice of healthcare outside of traditional settings to bring care to those in need. Upstream of patient management, diagnosis is aimed at adequately orienting medical decisions, and considerable efforts have been made to make this process faster and more efficient. However, there are several diseases and medical conditions that may/will benefit from technologies and tests that can be performed closer to the patient, at the point of care or even in the home. In this review, and in light of the paradox that technology and assay developers and healthcare officials must take into consideration for advancing human health in developed and developing countries, we present an overview of rapid diagnosis of infectious diseases at the point of care and of technologies that may contribute to enhancement of the worldwide point-of-care testing market.

Clostridium lavalense sp. nov., a glycopeptide-resistant species isolated from human faeces.

Two vancomycin-resistant, strictly anaerobic, Gram-positive, rod-shaped, spore-forming organisms (strains CCRI-9842(T) and CCRI-9929) isolated from human faecal specimens in Québec, Canada, and Australia were characterized using phenotypic, biochemical and molecular taxonomic methods. Pairwise analysis of the 16S rRNA gene sequences showed that both strains were closely related to each other genetically (displaying 99.2 % sequence similarity) and represented a previously unknown subline within the Clostridium coccoides rRNA group of organisms (rRNA cluster XIVa of the genus Clostridium). Strains CCRI-9842(T) and CCRI-9929 used carbohydrates as fermentable substrates, producing acetic acid as the major product of glucose metabolism. The novel strains were most closely related to Clostridium asparagiforme, Clostridium bolteae and Clostridium clostridioforme, but morphological, biochemical and phylogenetic studies demonstrated that they represent a previously unidentified species of the genus Clostridium. This was confirmed by the unique cellular fatty acid composition of strains CCRI-9842(T) and CCRI-9929. Therefore, on the basis of data from the polyphasic taxonomic analysis, it is proposed that strains CCRI-9842(T) and CCRI-9929 represent a novel species of the genus Clostridium, for which the name Clostridium lavalense sp. nov. is proposed. The type strain is CCRI-9842(T) (=CCUG 54291(T)=JCM 14986(T)=NML 03-A-015(T)).

Ruminococcus gauvreauii sp. nov., a glycopeptide-resistant species isolated from a human faecal specimen.

A novel strictly anaerobic, vancomycin-resistant, Gram-positive coccus (strain CCRI-16,110(T)) was isolated from a human faecal specimen. This strain was characterized using morphological, biochemical and molecular taxonomic methods. The organism was unable to hydrolyse aesculin and failed to produce acid from cellobiose, d-lactose and alpha-raffinose. Acetic acid was the sole product of glucose fermentation by the organism. On the basis of 16S rRNA and tuf gene sequence comparison, strain CCRI-16,110(T) was most closely related to species of the genus Ruminococcus and formed a hitherto unknown sublineage within the Clostridium coccoides rRNA cluster of organisms (cluster XIVa). Based on phenotypic and phylogenetic evidence, a novel species, Ruminococcus gauvreauii sp. nov., is proposed. The type strain is CCRI-16,110(T) (=NML 060141(T) =CCUG 54,292(T) =JCM 14987(T)).

Assessment of microbiological quality of drinking water from household tanks in Bermuda.

Bermuda residents collect rainwater from rooftops to fulfil their freshwater needs. The objective of this study was to assess the microbiological quality of drinking water in household tanks throughout Bermuda. The tanks surveyed were selected randomly from the electoral register. Governmental officers visited the selected household (n = 102) to collect water samples and administer a short questionnaire about the tank characteristics, the residents' habits in terms of water use, and general information on the water collecting system and its maintenance. At the same time, water samples were collected for analysis and total coliforms and Escherichia coli were determined by 2 methods (membrane filtration and culture on chromogenic media, Colilert kit). Results from the 2 methods were highly correlated and showed that approximately 90% of the samples analysed were contaminated with total coliforms in concentrations exceeding 10 CFU/100 mL, and approximately 66% of samples showed contamination with E. coli. Tank cleaning in the year prior to sampling seems to protect against water contamination. If rainwater collection from roofs is the most efficient mean for providing freshwater to Bermudians, it must not be considered a source of high quality drinking water because of the high levels of microbial contamination.

vanD and vanG-like gene clusters in a Ruminococcus species isolated from human bowel flora.

A vancomycin-resistant, anaerobic, gram-positive coccus containing the vanD and vanG-like genes (strain CCRI-16110) was isolated from a human fecal specimen during a hospital surveillance program to detect carriers of vancomycin-resistant enterococci. Comparison of the 16S rRNA gene sequence of strain CCRI-16110 with databases revealed a potentially novel Ruminococcus species that was most similar (<94% identity) to Clostridium and Ruminococcus species. Strain CCRI-16110 was highly resistant to vancomycin and teicoplanin (MICs of >256 microg/ml). The complete DNA sequence of the vanD cluster was most similar (98.2% identity) to that of Enterococcus faecium BM4339, containing the vanD1 allele. An intD gene with 99% identity with that of this E. faecium strain was found to be associated with the vanD gene cluster of this novel anaerobic bacterium. Strain CCRI-16110 also harbors genes encoding putative VanS(G), VanG, and VanT(G) proteins displaying 56, 73.6, and 55% amino acid sequence identity, respectively, compared to the corresponding proteins encoded by the vanG1 and vanG2 operons of Enterococcus faecalis BM4518 and N03-0233. This study reports for the first time an anaerobic bacterium containing the vanD gene cluster. This strain also harbors a partial vanG-like gene cluster. The presence of vanD- and vanG-containing anaerobic bacteria in the human bowel flora suggests that these bacteria may serve as a reservoir for the vanD and vanG vancomycin resistance genes.

High prevalence of glycopeptide resistance genes vanB, vanD, and vanG not associated with enterococci in human fecal flora.

The presence of Enterococcus-associated vancomycin resistance genes vanA, vanB, vanD, vanE, and vanG in rectal swabs was investigated in two hospitals using PCR. All vanA genes detected were associated with vancomycin-resistant enterococci (VRE), whereas VRE-associated vanB genes were detected in only one hospital (4.7%). However, in both hospitals, high prevalences of vanB (6.2 and 2.3%), vanD (43.8 and 26.7%), and vanG (10.5 and 6.9%) genes not associated with enterococci were found.

Characterization of a Tn5382-like transposon containing the vanB2 gene cluster in a Clostridium strain isolated from human faeces.

OBJECTIVES: During a hospital surveillance programme to detect VRE carriers, an anaerobic vancomycin-resistant bacterial strain CCRI-9842 containing a vanB gene was isolated from a human faecal specimen. In this study, we have characterized this strain and its vanB-containing element. METHODS: Strain CCRI-9842 was characterized by 16S rDNA sequencing and susceptibility testing. PCR mapping and sequencing of the vanB-containing element, as well as plasmid extraction and mating experiments, were carried out to investigate the genetic basis of vancomycin resistance in this strain. RESULTS: Strain CCRI-9842 was identified as a Clostridium species closely related to Clostridium bolteae (96.8% 16S rDNA identity). This strain was resistant to a high level of vancomycin (MIC of 256 mg/L), but was susceptible to teicoplanin and ampicillin. The complete sequence of the CCRI-9842 vanB gene exhibited 99.1% identity with that of vanB2. PCR mapping and sequencing showed that the genetic element carrying vanB2 was similar to transposon Tn5382/Tn1549. This Tn5382-like transposon forms circular intermediates and is flanked on the left and right ends by repeat sequences of at least 700 bp in the opposite direction. No plasmid was detected in this strain, suggesting that the Tn5382-like transposon was integrated into the chromosome. The vancomycin resistance was not transferable to enterococci. CONCLUSIONS: Our report shows for the first time the presence of a Tn5382-like transposon carrying vanB2 in a Clostridium species of the human intestinal flora. This suggests that the vanB2 Tn5382-like transposon is an important vector for the spread of vancomycin resistance in several bacterial species.

Laboratory detection of group B Streptococcus for prevention of perinatal disease.

Group B Streptococcus (GBS) or Streptococcus agalactiae emerged in the 1970s as the leading cause of neonatal morbidity and mortality. Today, GBS remains one of the leading causes of sepsis and meningitis in newborns despite important prevention efforts, including the issuance of recommendations for prevention of perinatal GBS disease by the American College of Obstetricians and Gynecologists, the Centers for Disease Control and Prevention, and the American Academy of Pediatrics in 1996/1997. The gastrointestinal tract is the natural human reservoir for GBS and is the likely source of vaginal colonization. GBS disease in newborns usually results from ascending spread of GBS into the amniotic fluid, which leads to neonatal colonization and to invasive disease in some infants. This review analyzes the various laboratory methods available for the detection of GBS from clinical samples collected from pregnant women and will discuss their impact in the prevention of neonatal GBS infections and in the rationalization of antibiotic use. The recent commercial availability of a rapid and highly sensitive real-time polymerase chain reaction assay suitable for the specific detection of GBS from vagino-rectal samples obtained from pregnant women during delivery, which is approved by the US Food and Drug Administration, provides improvements in the accuracy and rapidity of GBS colonization screening compared to the standard culture-based method using the recommended selective enrichment broth.

New real-time PCR assay for rapid detection of methicillin-resistant Staphylococcus aureus directly from specimens containing a mixture of staphylococci.

Molecular methods for the rapid identification of methicillin-resistant Staphylococcus aureus (MRSA) are generally based on the detection of an S. aureus-specific gene target and the mecA gene. However, such methods cannot be applied for the direct detection of MRSA from nonsterile specimens such as nasal samples without the previous isolation, capture, or enrichment of MRSA because these samples often contain both coagulase-negative staphylococci (CoNS) and S. aureus, either of which can carry mecA. In this study, we describe a real-time multiplex PCR assay which allows the detection of MRSA directly from clinical specimens containing a mixture of staphylococci in <1 h. Five primers specific to the different staphylococcal cassette chromosome mec (SCCmec) right extremity sequences, including three new sequences, were used in combination with a primer and three molecular beacon probes specific to the S. aureus chromosomal orfX gene sequences located to the right of the SCCmec integration site. Of the 1,657 MRSA isolates tested, 1,636 (98.7%) were detected with the PCR assay, whereas 26 of 569 (4.6%) methicillin-susceptible S. aureus (MSSA) strains were misidentified as MRSA. None of the 62 nonstaphylococcal bacterial species or the 212 methicillin-resistant or 74 methicillin-susceptible CoNS strains (MRCoNS and MSCoNS, respectively) were detected by the assay. The amplification of MRSA was not inhibited in the presence of high copy numbers of MSSA, MRCoNS, or MSCoNS. The analytical sensitivity of the PCR assay, as evaluated with MRSA-negative nasal specimens containing a mixture of MSSA, MRCoNS, and MSCoNS spiked with MRSA, was approximately 25 CFU per nasal sample. This real-time PCR assay represents a rapid and powerful method which can be used for the detection of MRSA directly from specimens containing a mixture of staphylococci.

Sodium lauryl sulfate, a microbicide effective against enveloped and nonenveloped viruses.

The number of individuals infected with human immunodeficiency virus (HIV) and other pathogens causing sexually transmitted diseases (STDs) is growing dramatically worldwide. Globally, heterosexual transmission may account for as much as 85-90% of new cases of HIV infection. Latex condoms represent an effective barrier against sexually transmitted pathogens, but unfortunately, their use is not generalized. Therefore, there is an urgent need to develop safe and potent topical microbicides under the control of women to efficiently reduce the spread of sexually transmitted infections. Sodium lauryl sulfate (SLS), an anionic surfactant with protein denaturing potency, is a potent inhibitor of the infectivity of several enveloped (Herpes simplex viruses, HIV-1, Semliki Forest virus) and nonenveloped (papillomaviruses, reovirus, rotavirus and poliovirus) viruses. The mechanism of action of SLS involves the solubilization of the viral envelope and/or the denaturation of envelope and/or capsid proteins. Studies have shown that SLS is not toxic for cultured cell lines of different origins at concentrations that inactivate HIV-1, herpes and human papillomavirus in vitro. In addition, intravaginal pretreatment of mice with a gel formulation containing SLS, completely protected animals against Herpes simplex virus type-2 infection. The gel formulation containing SLS was also well-tolerated following repeated intravaginal administrations to rabbits. Taken together, these data suggest that SLS represents a potential candidate for the use as a topical microbicide to prevent the sexual transmission of HIV-1, herpes, human papillomavirus and possibly other sexually transmitted pathogens. The impact of such a preventive tool on public health can be enormous.

Treatment of visceral leishmaniasis with sterically stabilized liposomes containing camptothecin.

The efficacy of 20(S)-camptothecin (CPT), free and incorporated into sterically stabilized liposomes, has been investigated in vitro against Leishmania donovani promastigotes and in vivo in a murine model of visceral leishmaniasis. Incubation of L. donovani promastigotes with free or liposomal CPT inhibited the growth of parasites in a dose-dependent manner. Tissue distribution studies revealed that the intraperitoneal administration of liposomal CPT was efficient for the delivery of high drug levels to the liver and spleen. Treatment of infected mice with intraperitoneal injections of free and liposomal CPT significantly reduced the parasite loads in the livers by 43 and 55%, respectively, compared with the loads for untreated controls. However, both treatments caused normochromic anemia and neutropenia.

Pulmonary and systemic host response to Streptococcus pneumoniae and Klebsiella pneumoniae bacteremia in normal and immunosuppressed mice.

Mortality related to bacteremic pneumonia remains high, and the role of sepsis in inflammation, pulmonary injury, and death remains unclear, mostly in leukopenic states. In the present study, the microbiology, histopathology, and host response to Streptococcus pneumoniae and Klebsiella pneumoniae infection were determined in an experimental model of bacteremia in immunocompetent and leukopenic mice. Leukocyte depletion by cyclophosphamide did not impair the early clearance of pneumococci from blood but facilitated growth in lungs. By contrast, klebsiellae rapidly grew in blood of leukopenic mice. These observations suggest that tissue-based phagocytes and circulating leukocytes, respectively, play prominent roles in S. pneumoniae and K. pneumoniae eradication. The kinetics of leukocyte recruitment in lungs during S. pneumoniae bacteremia suggested early strong inflammation in immunocompetent mice that is associated with tumor necrosis factor alpha release and histological disorders, including cell debris and surfactant in alveolar spaces. Leukocyte depletion further stimulated pulmonary capillary leakage both in S. pneumoniae and K. pneumoniae bacteremia, which seemed attributable to bacterial virulence factors. Nitric oxide production did not differ significantly among groups. Leukopenia and low platelet counts characterized the late stage of bacteremia for both strains, but only K. pneumoniae altered renal function. Understanding the pathogenesis of bacteremia will help establish beneficial therapies for both sepsis and pneumonia.

Microbiological and inflammatory factors associated with the development of pneumococcal pneumonia.

Pneumococcal pneumonia still is associated with a high mortality rate, despite appropriate antimicrobial therapy. Many gaps remain in the understanding of the pathogenesis of this deadly infection. The microbial and inflammatory events that characterize survival or death after intranasal inoculation of mice with an LD(50) inoculum of Streptococcus pneumoniae were investigated. Survival was associated with rapid bacterial clearance and low inflammation (surfactant and red blood cells in alveoli), but no neutrophil recruitment or lung tissue injury was noted. By contrast, death was preceded by strong bacterial growth that peaked 48 h after the infection and was associated with gradual increases in pulmonary levels of interleukin-6, macrophage inflammatory protein (MIP)-1alpha, MIP-2, monocyte chemoattractant protein-1, KC, and neutrophil recruitment. The injection of tumor necrosis factor-alpha or the addition of lipopolysaccharide or heat-killed S. pneumoniae to the inoculum enhanced early host response and survival. These observations may help develop appropriate markers of evolution of pneumonia, as well as new therapeutic strategies.

Sodium lauryl sulfate abrogates human immunodeficiency virus infectivity by affecting viral attachment.

The microbicidal activity of sodium lauryl sulfate (SLS) against human immunodeficiency virus type 1 (HIV-1) was studied in cultured cells. Pretreatment of HIV-1(NL4-3) with SLS decreased, in a concentration-dependent manner, its infectivity when using 1G5 as target cells. In the absence of a viral pretreatment period or when 1G5 cells were pretreated with SLS, the surfactant-induced inactivation of viral infectivity was less pronounced, especially at concentrations between 375 and 550 microM. SLS had no effect on HIV-1 when the virus was adsorbed to 1G5 cells by a 2-h incubation period. SLS almost completely inhibited the fusion process by decreasing the attachment of HIV-1 to target cells. SLS also inhibited the infectivity of HIV-1-based luciferase reporter viruses pseudotyped with the amphotropic murine leukemia virus envelope (which enters cells in a CD4-, CCR5-, and CXCR4-independent manner), indicating that SLS may inactivate other envelope viruses. In contrast, no effect was seen with vesicular stomatitis virus envelope glycoprotein G (which enters cells through receptor-mediated endocytosis) pretreated with up to 700 microM SLS. SLS also decreased, in a dose-dependent manner, the HIV-1-dependent syncytium formation between 1G5 and J1.1 cells after a 24-h incubation. The reduction of luciferase activity was more pronounced when J1.1 cells (which express HIV-1 proteins on their surface) were pretreated with SLS rather than 1G5 cells. Taken together, our results suggest that SLS could represent a candidate of choice for use in vaginal microbicides to prevent the sexual transmission of HIV and possibly other pathogens causing sexually transmitted diseases.

Development of a PCR assay for identification of staphylococci at genus and species levels.

We have developed a PCR-based assay which allows the detection of staphylococci at the genus level by targeting the tuf gene, which encodes the elongation factor Tu. Degenerate PCR primers derived from consensus regions of several tuf genes were used to amplify a target region of 884 bp from 11 representative staphylococcal species. Subsequently, the entire nucleotide sequence of these amplicons was determined. The analysis of a multiple alignment of these sequences revealed regions conserved among staphylococci but distinct from those of other gram-positive bacteria genetically related to staphylococci. PCR primers complementary to these regions could amplify specifically and efficiently a DNA fragment of 370 bp for all of 27 different staphylococcal species tested. There was no amplification with genomic DNA prepared from 53 nonstaphylococcal species tested to verify the specificity of the assay (20 gram positive and 33 gram negative). Furthermore, this assay amplified efficiently all 27 American Type Culture Collection (ATCC) staphylococcal reference strains as well as 307 clinical isolates of staphylococci from the Québec City region. Analysis of the multiple sequence alignment for the 884-bp fragment for the 11 staphylococcal species as well as comparison of the sequences for the 370-bp amplicon from five unrelated ATCC and clinical strains for each of the species S. aureus, S. epidermidis, S. haemolyticus, S. hominis, and S. saprophyticus demonstrated sufficient interspecies polymorphism to generate genus- and species-specific capture probes. This sequence information allowed the development of Staphylococcus-specific and species-specific (targeting S. aureus, S. epidermidis, S. haemolyticus, S. hominis, or S. saprophyticus) capture probes hybridizing to the 370-bp amplicon. In conclusion, this PCR assay is suitable for detection of staphylococci at both genus and species levels.

Role of chemokines and formyl peptides in pneumococcal pneumonia-induced monocyte/macrophage recruitment.

Host-derived chemoattractant factors are suggested to play crucial roles in leukocyte recruitment elicited by inflammatory stimuli in vitro and in vivo. However, in the case of acute bacterial infections, pathogen-derived chemoattractant factors are also present, and it has not yet been clarified how cross-talk between chemoattractant receptors orchestrates diapedesis of leukocytes in this context of complex chemoattractant arrays. To investigate the role of chemokine (host-derived) and formyl peptide (pathogen-derived) chemoattractants in leukocyte extravasation in life-threatening infectious diseases, we used a mouse model of pneumococcal pneumonia. We found an increase in mRNA expression of eight chemokines (RANTES, macrophage-inflammatory protein (MIP)-1alpha, MIP-1beta, MIP-2, IP-10, monocyte chemoattractant protein (MCP)-1, T cell activation 3, and KC) within the lungs during the course of infection. KC and MIP-2 protein expression closely preceded pulmonary neutrophil recruitment, whereas MCP-1 protein production coincided more closely than MIP-1alpha with the kinetics of macrophage infiltration. In situ hybridization of MCP-1 mRNA suggested that MCP-1 expression started at peribronchovascular regions and expanded to alveoli-facing epithelial cells and infiltrated macrophages. Interestingly, administration of a neutralizing Ab against MCP-1, RANTES, or MIP-1alpha alone did not prevent macrophage infiltration into infected alveoli, whereas combination of the three Abs significantly reduced macrophage infiltration without affecting neutrophil recruitment. The use of an antagonist to N-formyl peptides, N-t-Boc-Phe-D-Leu-Phe-D-Leu-Phe, reduced both macrophages and neutrophils significantly. These data demonstrate that a complex chemokine network is activated in response to pulmonary pneumococcal infection, and also suggest an important role for fMLP receptor in monocyte/macrophage recruitment in that model.

Thermoreversible gel formulations containing sodium lauryl sulfate or n-Lauroylsarcosine as potential topical microbicides against sexually transmitted diseases.

The microbicidal efficacies of two anionic surfactants, sodium lauryl sulfate (SLS) and n-lauroylsarcosine (LS), were evaluated in cultured cells and in a murine model of herpes simplex type 2 (HSV-2) intravaginal infection. In vitro studies showed that SLS and LS were potent inhibitors of the infectivity of HSV-2 strain 333. The concentrations of SLS which inhibit viral infectivity by 50% (50% inhibitory dose) and 90% (90% inhibitory dose) were 32.67 and 46.53 microM, respectively, whereas the corresponding values for LS were 141.76 and 225.30 microM. In addition, intravaginal pretreatment of mice with thermoreversible gel formulations containing 2.5% SLS or 2.5% LS prior to the inoculation of HSV-2 strain 333 completely prevented the development of genital herpetic lesions and the lethality associated with infection. Of prime interest, no infectious virus could be detected in mouse vaginal mucosa. Both formulations still provided significant protection when viral challenge was delayed until 1 h after pretreatment. Finally, intravaginal application of gel formulations containing 2.5% SLS or 2.5% LS once daily for 14 days to rabbits did not induce significant irritations to the genital mucosa, as demonstrated from macroscopic and histopathologic examinations. These results suggest that thermoreversible gel formulations containing SLS or LS could represent potent and safe topical microbicides for the prevention of HSV-2 and possibly other sexually transmitted pathogens, including human immunodeficiency virus.