Detectable Bacteremia Research Articles

A Rare Case of Bivalvular Infective Endocarditis and Mural Vegetation Caused by Bartonella quintana

Bartonella quintana is a rare but increasingly recognized cause of culture-negative infective endocarditis. We report a patient who presented with features of congestive heart failure without associated fever or detectable bacteremia. Diagnostic work-up revealed infective endocarditis due to B. quintana causing a large right atrial mural vegetation along with severe mitral regurgitation and severe aortic regurgitation, for which the patient had surgical valve replacements. The case highlights the indolent nature of the disease, leading to late presentation and challenging management, especially in a vulnerable patient population.

Duration of tick attachment necessary for transmission of Anaplasma phagocytophilum by Ixodes scapularis (Acari: Ixodidae) nymphs

This study assessed the duration of tick attachment necessary for a successful transmission of Anaplasma phagocytophilum by an infected I. scapularis nymph. Individual nymphs were placed upon BALB/c mice and allowed to feed for predetermined time intervals of 4 to 72 h. Ticks removed from mice at predetermined intervals were tested by PCR for verification of infection and evaluation of the bacterial load. The success of pathogen transmission to mice was assessed by blood-PCR at 7, 14 and 21 days postinfestation, and IFA at 21 days postinfestation. Anaplasma phagocytophilum infection was documented in 10-30 % of mice, from which ticks were removed within the first 20 h of feeding. However, transmission success was ≥70% if ticks remained attached for 36 h or longer. Notably, none of the PCR-positive mice that were exposed to infected ticks for 4 to 8 h and only half of PCR-positive mice exposed for 24 h developed antibodies within 3 weeks postinfestation. On the other hand, all mice with detectable bacteremia after being infested for 36 h seroconverted. This suggests that although some of the ticks removed prior to 24 h of attachment succeed in injecting a small amount of A. phagocytophilum, this amount is insufficient for stimulating humoral immunity and perhaps for establishing disseminated infection in BALB/c mice. Although A. phagocytophilum may be present in salivary glands of unfed I. scapularis nymphs, the amount of A. phagocytophilum initially contained in saliva appears insufficient to cause sustainable infection in a host. Replication and, maybe, reactivation of the agent for 12-24 h in a feeding tick is required before a mouse can be consistently infected.

Bacteremia after Endodontic Procedures in Patients with Heart Disease: Culture and Molecular Analyses

IntroductionInfective endocarditis (IE) is still associated with high mortality, and antibiotic prophylaxis strategies are under intense debate. We evaluated the incidence of bacteremia after root canal preparation in teeth with necrotic pulps and apical periodontitis. MethodsBlood samples were taken before and 5 and 30 minutes after endodontic treatment in teeth with apical periodontitis from individuals at high (n = 21) or no risk (n = 11) for IE. The former received prophylactic antibiotic therapy. Bacteriologic samples were taken from root canals before chemomechanical preparation to confirm pulp infection. Samples were subjected to aerobic and anaerobic culture and quantitative real-time polymerase chain reaction (qPCR), the latter to determine the total bacterial and streptococcal levels. ResultsCulture revealed no bacteremia in all individuals. Analysis by qPCR showed that bacterial DNA occurred in all root canal samples. qPCR showed a similar incidence of bacteremia between patients who received or did not receive prophylactic antibiotic therapy (P > .05). In blood samples taken 5 minutes after endodontic procedures, bacteria were detected in 2 of 11 (18%) individuals not taking antibiotics and in 4 of 21 (19%) patients under prophylaxis. After 30 minutes, the incidence of bacteremia decreased to 2 of 21 (10%) in patients taking antibiotics and was undetectable in patients at no risk of IE. The incidence of bacteremia by streptococci was identical as that for total bacteria. ConclusionsNo detectable bacteremia was evident by culture after treatment of infected root canals. Molecular analysis revealed bacterial DNA and streptococci in blood from some patients without a significant difference between individuals receiving or not receiving antibiotic prophylaxis.

Revisiting the Concept of Targeting Only Bacillus anthracis Toxins as a Treatment for Anthrax.

Protective antigen (PA)-based vaccines are effective in preventing the development of fatal anthrax disease both in humans and in relevant animal models. The Bacillus anthracis toxins lethal toxin (lethal factor [LF] plus PA) and edema toxin (edema factor [EF] plus PA) are essential for the establishment of the infection, as inactivation of these toxins results in attenuation of the pathogen. Since the toxins reach high toxemia levels at the bacteremic stages of the disease, the CDC's recommendations include combining antibiotic treatment with antitoxin (anti-PA) immunotherapy. We demonstrate here that while treatment with a highly potent neutralizing monoclonal antibody was highly efficient as postexposure prophylaxis treatment, it failed to protect rabbits with any detectable bacteremia (≥10 CFU/ml). In addition, we show that while PA vaccination was effective against a subcutaneous spore challenge, it failed to protect rabbits against systemic challenges (intravenous injection of vegetative bacteria) with the wild-type Vollum strain or a toxin-deficient mutant. To test the possibility that additional proteins, which are secreted by the bacteria under pathogenicity-stimulating conditions in vitro, may contribute to the vaccine's potency, we immunized rabbits with a secreted protein fraction from a toxin-null mutant. The antiserum raised against the secreted fraction reacts with the bacteria in an immunofluorescence assay. Immunization with the secreted protein fraction did not protect the rabbits against a systemic challenge with the fully pathogenic bacteria. Full protection was obtained only by a combined vaccination with PA and the secreted protein fraction. Therefore, these results indicate that an effective antiserum treatment in advanced stages of anthrax must include toxin-neutralizing antibodies in combination with antibodies against bacterial cell targets.

Vitamin A deficient mice are highly susceptible to Streptococcus pneumoniae and exhibit a poor response to vaccination with Prevnar-13

Abstract Streptococcus pneumoniae (pneumococcus) is frequently found in the human respiratory tract and can cause a wide range of diseases from mild otitis to fatal pneumonia or meningitis. It is estimated that pneumococcus kills 1.6 million people each year, approximately half of whom are children in developing nations. Given that vitamin A deficiency (VAD) occurs in both developed and developing countries, we sought to explore the relationship between VAD and pneumococcal disease. Previously, we have shown that mice with VAD fail to clear respiratory virus infections efficiently, resulting in a prolonged state of inflammation. Here we show that mice with VAD are more likely to develop a severe lung infection following intranasal infection with pneumococcus compared to vitamin A sufficient controls. We also found that VAD increases morbidity and mortality from secondary pneumococcal infections in influenza-infected mice. This increased sensitivity to pneumococcal infections led us to investigate the effectiveness of the human pneumococcal vaccine Prevnar-13 (PCV) in the context of VAD. Vaccinated mice with VAD mounted only minimal IgM and IgG antibody responses toward PCV compared to controls. Upon challenge with an invasive strain (TIGR4) of pneumococcus, vaccinated control animals were nearly 100% protected from disease with no detectable bacteremia at 24 hours post-infection, whereas nearly 1/3 of vaccinated VAD animals developed bacteremia within 24 hours and ultimately succumbed to the infection. Our results suggest that clinical analyses of children in both developed and developing countries should be conducted to ensure that PCV successfully protects against pneumococcal disease in individuals with VAD.

Incidence of bacteremia following upper gastrointestinal endoscopy and biopsy in healthy dogs before, during, and after treatment with omeprazole

To determine the incidence of bacteremia, as detected by routine methods for bacterial culture of blood samples, following routine endoscopic biopsy of the stomach and duodenum in healthy research dogs and to determine whether treatment with omeprazole administration affected the incidence of bacteremia. 8 healthy purpose-bred research dogs. All dogs underwent gastroduodenoscopy with biopsy at 4 points: twice prior to treatment with omeprazole, once following 15 days of omeprazole treatment (20 mg, PO, q 12 h), and once 14 days after treatment ceased. Dogs had a mean ± SD body weight of 18.6 ± 2.0 kg. Blood samples were aseptically obtained at 3 points during each procedure (before, immediately following, and 24 hours after endoscopy), and routine aerobic and anaerobic bacterial culture of blood was performed. 96 cultures were attempted for each culture method, yielding positive results of aerobic culture for 2 dogs at separate time points and no positive results of anaerobic culture. Routine gastrointestinal endoscopy with biopsy in healthy dogs did not result in a detectable bacteremia in most dogs. Treatment with the gastric acid-suppressing medication omeprazole did not affect the incidence of bacteremia as detected via standard techniques.

Investigation of bacteremia induced by removal of orthodontic mini-implants

The aim of this study was to investigate potential occurrence of bacteremia in orthodontic patients after removal of miniscrews.The study group comprised 30 healthy subjects (17 males, 13 females) with a mean age of 24.1 years treated with self-ligating fixed appliances and mini-implant anchorage. Two 20 ml venous blood samples were obtained prior to and 30-60 seconds after miniscrew explantation following an aseptic technique. Blood culturing in aerobic and anaerobic conditions was carried out by means of the BACTEC blood culture analyzer. Microbiological analysis showed that none of the pre- and post-operative samples exhibited detectable bacteremia. Future research should be focused on determining the collective bacteremic effect of a sequence of orthodontic procedures including miniscrew placement or removal, typically performed during a single treatment session.

Protection from reinfection in “Candidatus Mycoplasma turicensis”-infected cats and characterization of the immune response

“Candidatus Mycoplasma turicensis” (CMt) is a hemoplasma species of felids. Recent evidence has shown that cats that overcome bacteremia may be protected from reinfection. The purposes of this study were to (1) re-inoculate ostensibly recovered cats, (2) evaluate the immune response and (3) assess CMt tissue loads. Fifteen specified pathogen-free cats were subcutaneously inoculated with CMt: 10 cats (group A) had previously undergone bacteremia and recovered, and 5 naïve cats (group B) served as controls. CMt infections were monitored by real-time PCR using blood and tissue, and the humoral immune response was assessed using DnaK ELISA. Cytokine mRNA expression levels were measured by real-time PCR, and lymphocyte subsets were detected by flow cytometry. The cats in group A were protected from reinfection (no detectable bacteremia) and showed a transient decrease in antibodies. Eosinophilia was noted in cats from group A. The cats from group B became PCR-positive and seroconverted. All of the tissues analyzed from the cats in group B but none of the tissues analyzed from the cats in group A were CMt PCR-positive. Significant changes were observed in the expression of tumor necrosis factor-α, interferon-γ, interleukin-4 and the Th2/Th1 ratio in both groups. The cats from group A occasionally showed higher numbers of CD4+, CD8+, CD4+CD25+ and CD5+MHCII+ T lymphocytes than the control cats. In conclusion, this study describes, for the first time, the occurrence of immunological protection within the same hemoplasma species. Furthermore, the immune response during CMt infections appeared to be skewed toward the Th2 type.

Chronic "Candidatus Mycoplasma turicensis" infection

"Candidatus Mycoplasma turicensis" infects felids. The pathogenesis of "Candidatus M. turicensis" chronic infection is poorly understood. The goals of the present study were to (1) induce reactivation of the infection in chronic carrier cats by attempted immunosuppression, (2) identify potential tissue sequestration using real-time TaqMan® PCR and (3) monitor the humoral immune response by DnaK enzyme-linked immunosorbent assay (ELISA). Ten specified pathogen-free cats that had ostensibly recovered from experimental "Candidatus M. turicensis" infection were used: five cats (group 1) received high dose methylprednisolone (attempted immunosuppression), while five cats served as untreated controls (group 2). Besides weekly blood samples, tissue samples were collected from bone marrow, kidney, liver and salivary glands at selected time points. The cats in group 1 had significantly lower lymphocyte counts and higher blood glucose levels after methylprednisolone administration than the controls. After methylprednisolone administration one blood and three tissue samples from cats in group 1 tested PCR-positive; before the administration, only one sample was positive. All other samples tested PCR-negative. All cats stayed seropositive; the antibody levels of the cats in group 1 showed a significant transient decrease after methylprednisolone administration. This is the first study to report the presence of "Candidatus M. turicensis" in tissues of chronically infected cats and the persistence of anti-feline hemoplasma antibodies in the absence of detectable bacteremia. Methylprednisolone administration did not lead to a significant reactivation of the infection. Our results enhance the knowledge of "Candidatus M. turicensis" infection pathogenesis and are clinically relevant to the prognosis of hemoplasma-infected cats.

Bacteraemia due to dental flossing

The aims of this study were to (1) investigate the incidence of bacteraemia following flossing in subjects with chronic periodontitis or periodontal health; (2) identify the micro-organisms in detected bacteraemias; and (3) identify any patient or clinical factors associated with such bacteraemia. Baseline blood samples were obtained from 30 individuals with chronic periodontitis (17 M:13 F, 29-75 years) and 30 with periodontal health (17 M:13 F, 28-71 years) following a non-invasive examination. Each subject's teeth were then flossed in a standardized manner and blood samples obtained 30 s and 10 min. after flossing cessation. Blood samples were cultured in a BACTEC system and positive samples subcultured for identification. Forty per cent of periodontitis subjects and 41% of periodontally healthy subjects tested positive for bacteraemia following flossing. Viridans streptococci, which are commonly implicated in infective endocarditis (IE), were isolated from 19% of positive subjects and accounted for 35% of microbial isolates. Twenty per cent of subjects had a detectable bacteraemia at 10 min. post-flossing. No patient or clinical factors were significantly associated with post-flossing bacteraemia. Dental flossing can produce bacteraemia in periodontally healthy and periodontally diseased individuals at a rate comparable with that caused by some dental treatments for which antibiotic prophylaxis is given to prevent IE.

Primary Gluteal Abscess Due to Salmonella typhimurium: A Case Report and Review of the Literature

Infections due to nontyphoidal salmonellae are common, and their incidence has been increasing in the last few years. In this case report, we document the unusual finding of a gluteal abscess caused by Salmonella typhimurium in the absence of detectable bacteremia. Fluoroquinolones remain an important treatment option for salmonellosis, and to avoid potential treatment failures, we underscore the importance of routine screening for nalidixic acid susceptibility with all extraintestinal Salmonella isolates. To our knowledge, this is the first published case of S typhimurium gluteal abscess in the absence of bacteremia and risk factors for salmonellosis.

Detection of bacteraemias during non-surgicalroot canal treatment

Some dental procedures initiate a bacteraemia. In certain compromised patients, this bacteraemia may lead to distant site infections, most notably infective endocarditis. To investigate whether a detectable bacteraemia was produced during non-surgical root canal therapy. Thirty patients receiving non-surgical root canal therapy were studied. Three blood samples were taken per patient: pre-operatively, peri-operatively and post-operatively. In addition, a paper point sample was collected from the root canal. The blood samples were cultured by pour plate and blood bottle methods. The isolated organisms were identified by standard techniques. Blood samples were analysed for the presence of bacterial DNA by the polymerase chain reaction (PCR). In two cases where the same species of organism was identified in the root canal and the bloodstream, the isolates were typed by pulsed field gel electrophoresis (PFGE). By conventional culturing, a detectable bacteraemia was present in 9 (30%) of the 30 patients who had no positive pre-operative control blood sample. In 7 (23.3%) patients, the same species of organism was identified in both the bloodstream and in the paper point sample from the root canal system. Overall, PCR gave lower detection rates compared with conventional culture, with 10 of 90 (11%) of the blood samples displaying bacterial DNA. PFGE typing was undertaken for two pairs of culture isolates from blood and paper points; these were found to be genetically identical. Non-surgical root canal treatment may invoke a detectable bacteraemia.

Prospective Studies ofBartonellaof Rodents. Part II. Diverse Infections in a Single Rodent Community

The genetic diversity of Bartonella species within a small mammal community and in individual cotton rats (Sigmodon hispidus) was examined by trapping, capturing, sampling, and releasing of marked animals over a 17-month interval. Based on sequence analyses of the Bartonella gltA gene, amplicons separated into four genogroups (A, B, C, and Pin) containing 11 variants. Although the prevalence of bacteremia due to different genogroups/variants of Bartonella was temporally variable, variants of genogroup A predominated during each sampling period. Multiple gltA variants were often (20.5% of individuals) isolated from a single cotton rat blood sample; a maximum of five variants was recovered from an individual during its sampling history. Among 92 cotton rats bacteremic at two or more sampling dates, 34 rats retained a single genetic variant, alone or in mixed infection, throughout their sampling history. The temporal course of individual infections was complex as the succession of gltA variants was variable and detectable bacteremia was often intermittent. No antibodies (titer of >1:8) were detected to homologous strains of Bartonella recovered from individual cotton rats during their sampling history. The temporal course of Bartonella infections could result from a single, persistent, and potentially multi-genogroup/variant infection, during which variants differentially dominate the detectable bacteremia.

Bacteremia following surgical dental extraction with an emphasis on anaerobic strains.

Our aim was to investigate bacteremia caused by surgical extraction of partly erupted mandibular third molars. From 16 young adults, bacterial samples were taken from the third-molar pericoronal pocket and post-operatively from the extraction socket, and blood samples were drawn from the ante-cubital vein up to 30 min after surgery. Of the subjects, 88% had detectable bacteremia-50% 1 min after the incision, 44% immediately after extraction. The respective percentages at 10, 15, and 30 min were 44%, 25%, and 13%. Blood cultures contained 31 species (74% anaerobes), with 3.9 +/- 2.6 species isolated per subject. Most prevalent were the anaerobes Prevotella, Eubacterium, and Peptostreptococcus sp. and the aerobes viridans-group streptococci and Streptococcus milleri group. Any species found in the blood was also isolated from the mouth, from 93% of the pericoronal pockets and from 43% of the extraction sockets. Surgical dental extraction clearly causes bacteremia of a high frequency and lasting longer than thus far assumed.

Inflammatory consequences of the translocation of bacteria and endotoxin to mesenteric lymph nodes

Background: Translocation of intestinal bacteria to mesenteric lymph nodes (MLNs) has been documented in humans under a variety of circumstances, yet its clinical significance remains to be established. The aim of this study was to correlate detectable translocation to MLNs of bacteria and endotoxin with local and systemic signs of inflammation. Methods: From each of 10 patients with carcinoma of the cecal region two MLNs were harvested prior to resection. The presence of bacteria and endotoxin in the lymphatic tissue and blood was determined by culture methods and DNA preparation (PCR) and by a Limulus assay, respectively. Inflammatory mediators were determined in plasma and in MLN homogenates. Results: Viable bacteria were detected in MLNs of 7 patients and in 9 of 20 lymph nodes. PCR revealed traces of bacteria in 4 patients and in 6 of their MLNs. Combining both modalities, the translocation rate was 80% and 55% for patients and MLNs, respectively. There was no detectable bacteremia. Endotoxin was found in the plasma of 7 patients and in 9 MLNs from 5 patients. There was no correlation between culture findings and endotoxin concentrations. Moreover, bacteriological data did not correspond to local or systemic inflammation. The group of MLN with detectable endotoxin differed significantly from LPS-negative nodes with respect to interleukin-6, interleukin-10, and sCD14. Systemic concentrations of endotoxin and inflammatory parameters did not correspond to levels within MLNs. Conclusion: Translocation to MLNs occurs in patients with cecal carcinoma. This, however, seems not to be of major clinical significance if no additional physiologic insults are encountered. Irrespective of the presence of bacteria, there are variations in inflammatory reactions between lymph nodes from one and the same patient, probably reflecting fluctuating response mechanisms to low-grade translocation.

Experimental infection of cotton rats with three naturally occurring Bartonella species.

The kinetics of infection and humoral immune response of laboratory-bred cotton rats (Sigmodon hispidus) challenged with three Bartonella spp. recovered from the blood of naturally infected cotton rats captured in Georgia (USA) are described. Bartonella spp. infection, as determined by bacteremia, occurred in all 18 cotton rats inoculated with live Bartonella of each species at either a low dose, 10(3) colony-forming units (CFU's), or high dose, 10(7) CFU. Cotton rats inoculated with lower doses of Bartonella spp. developed higher bacteremia that persisted for longer periods than in those inoculated with high doses. Peak bacteremia varied among Bartonella spp, ranging from 10(4) to 10(6) CFUs per 1.0 ml of blood. Antibody measured by immunofluorescence assays using species-specific antigens indicated more rapidly rising and higher antibody titers in cotton rats challenged with high doses vs. low doses and with inactivated bacteria vs. live bacteria. Each group of rats produced high IgG titers to the homologous challenge antigen; low or unmeasurable cross-reactivity was detected to heterologous Bartonella antigens. Exposure of cotton rats to a specific Bartonella sp. resulted in protection, as measured by detectable bacteremia, in eight of nine animals challenged with the same Bartonella sp. used initially; no evidence of resistance to secondary challenge with different Bartonella spp. was obtained. Cross-protection between Bartonella spp., isolated from the same rodent species, may not occur.

Gut-derived mesenteric lymph but not portal blood increases endothelial cell permeability and promotes lung injury after hemorrhagic shock.

To determine whether gut-derived factors leading to organ injury and increased endothelial cell permeability would be present in the mesenteric lymph at higher levels than in the portal blood of rats subjected to hemorrhagic shock. This hypothesis was tested by examining the effect of portal blood plasma and mesenteric lymph on endothelial cell monolayers and the interruption of mesenteric lymph flow on shock-induced lung injury. The absence of detectable bacteremia or endotoxemia in the portal blood of trauma victims casts doubt on the role of the gut in the generation of multiple organ failure. Nevertheless, previous experimental work has clearly documented the connection between shock and gut injury as well as the concept of gut-induced sepsis and distant organ failure. One explanation for this apparent paradox would be that gut-derived inflammatory factors are reaching the lung and systemic circulation via the gut lymphatics rather than the portal circulation. Human umbilical vein endothelial cell monolayers, grown in two-compartment systems, were exposed to media, sham-shock, or postshock portal blood plasma or lymph, and permeability to rhodamine (10K) was measured. Sprague-Dawley rats were subjected to 90 minutes of sham or actual shock and shock plus lymphatic division (before and after shock). Lung permeability, pulmonary myeloperoxidase levels, alveolar apoptosis, and bronchoalveolar fluid protein content were used to quantitate lung injury. Postshock lymph increased endothelial cell monolayer permeability but not postshock plasma, sham-shock lymph/plasma, or medium. Lymphatic division before hemorrhagic shock prevented shock-induced increases in lung permeability to Evans blue dye and alveolar apoptosis and reduced pulmonary MPO levels. In contrast, division of the mesenteric lymphatics at the end of the shock period but before reperfusion ameliorated but failed to prevent increased lung permeability, alveolar apoptosis, and MPO accumulation. Gut barrier failure after hemorrhagic shock may be involved in the pathogenesis of shock-induced distant organ injury via gut-derived factors carried in the mesenteric lymph rather than the portal circulation.

Does Bacteremia Occur During High Pressure Lavage of Contaminated Wounds?

The risk of bacteremia secondary to high pressure lavage of contaminated wounds was assessed. Twenty canines were divided randomly into four treatment groups. A 10-cm incision was made over the left shoulder of each dog. The deltoideus muscle was disrupted and traumatized. Groups A and B (n = 8) had wound contamination with 1.4 x 10(9) Staphylococcus aureus followed 75 minutes later by high pressure lavage or bulb syringe irrigation, respectively. Groups C and D (n = 2) had no contamination, followed by the same treatment. Bacterial counts were obtained before and after wound irrigation. Blood cultures were obtained before, during, and 15 minutes after irrigation. Positive control cultures were obtained during injection of bacteria into the antecubital vein. A detectable bacteremia did not occur during or after high pressure lavage or bulb syringe irrigation of acute contaminated wounds but did occur in 18 of 20 positive controls. Bacterial levels were reduced by an average of 70% +/- 10% by high pressure lavage and 44% +/- 50% by bulb irrigation. Reduction of wound bacteria was achieved more consistently with high pressure lavage than with bulb syringe irrigation.

Clinical definition for invasive Salmonella infection in African children.

Invasive salmonellosis is common among children in tropical Africa, typically presenting as a nonspecific febrile illness that is difficult to distinguish clinically from malaria. This study examines the performance of a clinical definition devised to aid its recognition among children ages 1 to 15 years presenting to a mission hospital in rural Zaire. Invasive salmonellosis was defined by: (1) illness requiring admission to hospital in the opinion of an experience pediatrician; (2) history of fever for 5 or more days; (3) no focus of infection on clinical examination; and (4) negative or only scanty positive thick film for malarial parasites. Children fulfilling all these criteria were treated with ciprofloxacin after culture of blood and feces. The primary outcome measure was blood culture-confirmed salmonellosis. Secondary measures were final clinical diagnosis and serologic evidence of recent salmonellosis. Of 120 children fulfilling the definition, 55 (46%) were bacteremia; in 46 (38%) Salmonella species were isolated. In the majority of the nonbacteremic children no definite cause for the fever could be found. Salmonella serology supported invasive salmonellosis as the diagnosis in 62% of the nonbacteremic children. Salmonella serology suggested that invasive salmonellosis without detectable bacteremia was common. The addition of blood culture-proved and serologically diagnosed cases indicates that the definition has a specificity of at least 60%.

Characterization of a mutant of Haemophilus influenzae type b lacking the P2 major outer membrane protein

An isogenic mutant of Haemophilus influenzae type b (Hib) lacking the ability to express the P2 major outer membrane (porin) protein was constructed and characterized in various model systems. Linker insertion mutagenesis of a cloned Hib DNA insert containing the P2 structural gene was used in conjunction with a genetic transformation system to obtain a transformant unreactive with a P2-specific monoclonal antibody. This transformant was shown to lack detectable P2 protein by both protein staining and immunoblot methods. The P2 mutant exhibited a generation time in complex broth medium that was significantly longer than that of the wild-type parent strain. The P2 mutant was also unable to produce detectable bacteremia in infant rats after intraperitoneal challenge, while the wild-type parent strain produced bacteremia in all animals challenged with this strain. Reintroduction of a wild-type copy of the P2 gene into this mutant yielded a transformant strain that had a generation time in vitro identical to that of the wild-type parent strain and that was also fully virulent in the infant rat model. These findings suggest that the ability to synthesize the P2 protein may be necessary but not sufficient for full expression of virulence by this pathogen.