Elsebeth Tvenstrup Jensen

Some bacterial parameters influencing the neutrophil oxidative burst response to Pseudomonas aeruginosa biofilms.

Persistence of bacteria in spite of a normal host immune system and relevant antibiotic treatment is a key problem in many chronic infections, such as the bronchopulmonary P. aeruginosa infection in cystic fibrosis patients. The capability of bacteria to establish themselves in microcolonies or biofilms is an important protective mechanism of the microorganisms. We examined the human PMN oxidative burst response to P. aeruginosa in biofilm and in planktonic form. The PMN chemiluminescence response to P. aeruginosa in biofilms was reduced to 30.5–47.5% (p<0.04) and the superoxide response to 85.9% (p<0.02) of the response to equivalent numbers of planktonic bacteria. Mechanical disruption of the biofilms before the assays elicited a significantly increased response in the chemiluminescence experiments and to nonopsonized biofilms in the superoxide anion experiments. We conclude that biofilm bacteria, although able to stimulate the PMN, result in a reduced, suboptimal response leading to lack of efficient eradication of the bacteria in the chronic infection.

The relationship between prior cervical conization, cervical microbial colonization and preterm premature rupture of the membranes.

The occurrence of prior cervical conization and the cervical microbial colonization was investigated in 38 women with idiopathic preterm labor, 35 women with preterm premature rupture of the membranes (PPROM) and 75 normal pregnant women at 26-34 weeks of gestation. Data were analyzed by Fishers exact test (two-tailed). The frequency of prior cervical conization was significantly higher in PPROM patients compared to normal pregnant women (P < 0.001) and to patients in preterm labor (P < 0.01). Lactobacilli occurred with a lower frequency in patients with PPROM compared to patients in preterm labor (P < 0.05) and control patients (P = 0.0543)-and with a lower frequency in patients with prior cervical conization (P < 0.05). All other microorganisms occurred with the same frequencies in all groups. The absence of lactobacilli may indicate changes in the cervical flora, which could increase the risk of PPROM. Prior cervical conization may impair the antimicrobial defense-mechanisms in the cervix, which could facilitate ascending microbial colonization. This may lead to a release of prostaglandins and proteolytic enzymes and subsequently preterm labor and rupture of the membranes.

Experimental Salmonella typhimurium infections in rats

The course of experimentally induced Salmonella typhimurium infection was studied in three groups of inbred LEW rats: homozygous +/+, athymic rnu/rnu and isogeneic thymus‐grafted rnu/rnu rats. In the first experiment the animals were inoculated intraperitoneally with 108 bacteria and all animals became severely septicemic and died within a week of inoculation, irrespective of presence or absence of thymus. In the second experiment the animals were inoculated with 106 bacteria, and both euthymic and thymus‐grafted animals responded with high titres of anti bacterial antibodies while these were very low in the athymic nude animals. Polyclonal antibody production was only observed in the euthymic animals and only regarding IgG. Athymic rats were not able to clear the infection, while the thymus‐grafted animals reacted like euthymic rats: Very few animals housed the bacteria four weeks after inoculation, and no bacteria could be detected after four months. Immunohistochemical studies of lymphoid organs revealed that the infection caused a drop in the percentages of T non‐helper cells, indicating low suppressor activity. The study shows that athymic nude rats are well suited for studies of infectious diseases in immune deficient hosts, and that whole neonatally isogeneic thymus grafting is a good way of reconstituting these animals immunologically.

Induction of oxidative burst response in human neutrophils by immune complexes made in vitro of lipopolysaccharide and hyperimmune serum from chronically infected patients

Purified lipopolysaccharide (LPS) from Pseudomonas aeruginosa was used as an antigen for immune complex (IC) formation in vitro together with hyperimmune sera from chronically P. aeruginosa‐infected patients with cystic fibrosis (CF). P. aeruginosa LPS by itself did not induce an oxidative burst in human neutrophil granulocytes (PMN)s measured by chemiluminescence (CL). This was also the case using hyperimmune CF serum alone. In contrast, P. aeruginosa LPS together with CF serum did induce a CL response. The CL responses varied depending on the sera used for IC formation, and were reduced when protein A preabsorbed sera were used. PEG precipitation of the ICs from the mixture increased the CL response. These findings indicate that the CL responses induced by the mixture of P. aeruginosa LPS and CF serum were due to IC formation and an Fc‐mediated stimulation of the PMNs. It is concluded that ICs made from sera of chronically infected CF patients and purified P. aeruginosa LPS are biologically active in terms of activating PMNs, and may contribute to the lung tissue damage seen in this group of patients.

Experimental Salmonella typhimurium infections in rats. III. Transfer of immunity with primed lymphocyte subpopulations.

Immunization against a lethal dose of Salmonella typhimurium was studied in athymic and thymus‐bearing LEW rats. Active immunization was performed with formalin‐killed whole cell vaccine or sublethal infection prior to the lethal infection. After vaccination with killed bacteria the euthymic animals produced antibodies against S. typhimurium, but neither the euthymic nor the athymic animals survived the infection. After non‐lethal infection euthymic and thymus‐grafted nude rats were not affected by the second and otherwise lethal bacterial dose, and had high antibody titres. All the athymic nude rats died after the second and lethal bacterial challenge. Passive immunization with plasma from immunized euthymic animals did not protect any of the animals against the lethal bacterial dose. However, all animals survived when treated with large doses of spleen cells from immunized euthymic rats. Both athymic and thymus‐bearing animals treated with primed spleen cells had high antibody titres. The percentages of splenic and lymph node T lymphocytes in primed spleen cell‐treated athymic rats were comparable to those found in euthymic and thymus‐grafted animals. Treatment with primed spleen cells from immunized thymus grafted animals provided only limited protective effect, and treatment with cells from athymic animals had no effect. The study shows that although isogeneic thymus‐grafted nude rats become resistent to reinfection with S. typhimurium, only large doses of spleen cells from immunized euthymic animals can be used for passive transfer of immunity.

Immunity to experimental Salmonella typhimurium infections in rats

The protective effect of primed CD4+ T lymphocytes against a lethal dose of 108 viable Salmonella typhimurium was studied in Lewis rats. Primed CD4+ T lymphocytes were obtained by inoculating Lewis rats with a non‐lethal dose of 106 viable S. typhimurium. Four weeks after the infection, spleen CD4+ T lymphocytes were separated using magnetic microspheres coated with an antibody against the CD4 molecule (W3/25). Subsequent sorting into activated and non‐activated subpopulations using the p55 α‐chain of the interleukin‐2 receptor (CD25) as an activation marker was performed by a fluorescence‐activated cell sorter. Untreated Lewis rats were injected with 104 different primed CD4+ T‐cell populations 24 h prior to the lethal dose of 108 viable S. typhimurium. Blood samples were drawn from the orbital plexus 1, 2, 3, and 4 weeks after the infection, and analysed for specific IgM and IgG antibodies. Cell sorting revealed that 2/3 of the primed CD4+ T lymphocytes expressed high levels of CD25. Cell transfer revealed that both CD25high and CD25low expression populations could induce immunity against a lethal dose of S. typhimurium, whilst antibody analysis revealed that antibody levels were not correlated with protection against S. typhimurium infections, although it showed that a higher and more persistent level of specific IgG antibodies was produced in animals receiving the CD4+CD25high fraction. It is concluded that 104 primed CD4+ T lymphocytes can induce immunity in animals challenged with a lethal dose of S. typhimurium and that antibodies do not seem to be correlated with the immunity induced. The CD4+CD25high fraction was, however, associated with a higher and more persistent level of specific IgG antibodies.

Group B streptococcal chorioamnionitis and neonatal septicemia following 8 days pivampicillin and metronidazol prophylaxis after premature rupture of membranes; a case report

A case of preterm premature rupture of the membranes (PPROM) in the 31st week of gestation is reported. Initial cultures from the cervix and urine were without pathogenic microorganisms. After 8 days of prophylactic pivampicillin and metronidazol, culture from the cervix showed profuse growth of Group B Streptococci (GBS) and the patient developed symptoms of chorioamnionitis. Cesarean section was performed and the infant presented GBS-septicemia. In spite of continued treatment with pivampicillin, culture from the cervix on day 6 post partum still showed profuse growth of GBS. Prolonged prophylactic per oral administration of broad-spectrum antibiotics after PPROM may not always protect against infectious complications. Literature is reviewed, and it is discussed whether the applied regimen in some cases even may favour the occurrence of serious infections.

Mechanism of Action of Antibiotics in Chronic Pulmonary Pseudomonas Infection

Publisher Summary This chapter describes the rationale based on pathogenesis, the principles of treatment, and the mechanism of action of antibiotics in chronic pulmonary P. aeruginosa infection, based on the experience obtained in cystic fibrosis (CF) patients. These principles are also used to treat other chronic obstructive pulmonary disease (COPD) patients suffering from such infections in the Danish CF Centre. Chronic P. aeruginosa infection in CF patients is an endobronchiolitis caused by bacteria producing a biofilm that makes them resistant to antibiotics, and to the defense mechanisms of the body. The characteristics of bacterial biofilms are briefly described. Improved knowledge about the pathology of recurrent and chronic infections and about the intensive treatment has continued to improve the survival and the quality of life of CF patients. The described methods can be adapted to other COPD patients with chronic P. aeruginosa colonization and infection, and hopefully will improve and normalize the life of these patients as well.

Uro‐genital microbial colonization and threatening preterm delivery

Objective. To examine whether there is a relationship between the urogenital microbial colonization and threatening preterm delivery.