[Abortions and stillbirths caused by Coxiella burnetii in goats]. / Aborte und Totgeburten bei Ziegen unter besonderer Berücksichtigung von Coxiella burnetii.

INTRODUCTION: Coxiellosis, caused by the bacterium Coxiella burnetii, is a reportable disease in animals and humans in Switzerland. The number of cases in farm animals and humans has risen continuously in recent years. The aim of this work was to investigate abortions and stillbirths in goats with a focus on C. burnetii, to identify excretory routes which pose a zoonotic risk and the excretion time after an acute infection. Besides the submitted fetuses, does were screened with a serological antibody test. In addition, excretion via milk, faeces and vaginal mucus were investigated in dams with fetuses tested positive for C. burnetii at 14-day intervals.C. burnetii were isolated in 8 cases (3× in the placenta, 2× in the abomasum, 3× in the placenta and abomasum) of 13 examined stillbirths/abortions. Ten abomasums of goat kids and 8 placentas were examined using modified Ziehl-Neelsen staining (ZN) according to Stamp simultaneously with a real-time PCR. Four of 18 samples were false negative using modified ZN staining according to Stamp in contrast to real-time PCR. Seven does had serum antibodies against Coxiella. The excretion of C. burnetii persisted for 63 days in the milk, for 96 days in the vaginal mucus and for 96 respectively 114 days in two does monitored extensively. Intermittent excretion could also be observed in the milk during these 63 days. The present study showed that confirmation of disease, respectively transmission cannot be based on a single test. Only combined serological antibody test and real-time PCR examinations of birth material, milk, feces and vaginal mucus can result in a conclusive diagnosis. In addition, the examination using modified ZN staining according to Stamp is less sensitive and specific than the real-time PCR examination.

INTRODUCTION: La coxiellose, causée par la bactérie Coxiella burnetii, est une maladie à déclaration obligatoire en Suisse qui touche les animaux et les humains. Le nombre de cas chez les animaux de rente et les humains n'a cessé d'augmenter ces dernières années. Le but de ce travail était d'étudier les avortements et la mortalité périnatale chez les chèvres avec un focus sur C. burnetii, d'en identifier les voies d'excrétion qui présentent un risque zoonotique et de déterminer le temps d'excrétion après une infection aiguë. Pour ce faire, des examens sérologiques d'anticorps ont été effectués sur les mères en parallèle des examens sur les fœtus envoyés. L'excrétion par le lait, les selles et les sécrétions vaginales ont été examinées à intervalles de 14 jours sur les mères dont les fœtus ont été testés positifs à C. burnetii. Sur les 13 mort-nés et avortements examinés, C. burnetii a été isolés dans 8 échantillons (3× dans le placenta, 2× dans la caillette, 3× dans le placenta et la caillette). Dix caillettes de chevr­eaux et 8 placentas ont été simultanément examinés en utilisant une coloration Ziehl-Neelsen (ZN), modifiée selon Stamp, et un real-time PCR. Sur les 18 échantillons examinés, 4 échantillons ont donné des faux négatifs en utilisant la coloration Ziehl-Neelsen modifiée par rapport à la real-time PCR. La sérologie a dévoilé que 7 femelles présentaient des anticorps contre Coxiella. Pour 2 femelles, suivies durant une période plus longue, l'excrétion de C. burnetii dans le lait a persisté durant 63 jours, dans les sécrétions vaginales durant 96 jours pour les 2 femelles et dans les selles durant 96 et 114 jours respectivement. Une excrétion intermittente par le lait a également pu être observée durant les 63 jours. Cette étude a démontré que la mise en évidence de la maladie respectivement de l'excrétion ne peut pas être assurée sur la base d'un seul test. Seul la combinaison de la sérologie et des examens au moyen de la real-time PCR sur les arrière-faix, le lait, les selles et les sécrétions vaginales peuvent aboutir à un diagnostic concluant. De plus, l'examen au moyen de la coloration ZN modifiée selon Stamp est moins sensible et moins spécifique que la real-time PCR.

Regulation of early peritoneal neutrophil migration by macrophage inflammatory protein-2 and mast cells in experimental peritonitis.

Neutrophil (PMN) migration into the peritoneal cavity in response to fecal peritonitis is an important mechanism of host defense against bacterial invasion. We show that the murine C-X-C (PMN-specific) chemokine, macrophage inflammatory protein-2 (MIP-2), on intraperitoneal injection in mice, causes PMN migration into the peritoneum. MIP-2 mRNA and protein were expressed by peritoneal leukocytes after cecal ligation and puncture (CLP) in mice and neutralization of MIP-2 reduced peritoneal PMN migration. A prerequisite for neutrophil-endothelial adhesion and subsequent migration from the circulation is selectin-mediated rolling. Pretreatment of mice with an anti-P-selectin antibody before intraperitoneal injection of MIP-2 significantly reduced peritoneal PMN migration. However, there are no reports that a C-X-C chemokine can up-regulate endothelial selectins. We postulated that MIP-2, when injected intraperitoneally, interacts with a cell that is known to release factors that up-regulate endothelial selectins. A likely candidate is the mast cell, which contains histamine and tumor necrosis factor alpha (TNF-alpha), and both of these factors induce selectins. Intraperitoneally injected MIP-2 caused an early significant increase in peritoneal TNF-alpha, whereas histamine levels were unaffected. In a subsequent experiment, mast cell-deficient mice and their normal controls were then injected intraperitoneally with MIP-2 or underwent CLP. Significantly fewer PMNs migrated into the peritoneal cavity in the mast cell-deficient mice after MIP-2 injection or CLP. Thus, our findings indicate that mast cells and MIP-2 are necessary for PMN migration into the peritoneum in response to intra-abdominal infection, and that MIP-2 appears to facilitate this through an increase in TNF-alpha release.

Allogeneic stem cell transplantation after conditioning with treosulfan, etoposide and cyclophosphamide for patients with ALL: a phase II-study on behalf of the German Cooperative Transplant Study Group and ALL Study Group (GMALL).

TBI-based preparative regimens are considered as standard conditioning therapy for allogeneic stem cell transplantation (AHSC) in patients with ALL. We investigated toxicity and efficacy of a non-TBI-based regimen consisting of treosulfan, etoposide and cyclophosphamide for ALL within a prospective study. Major inclusion criteria were CR and non-eligibility for TBI. Fifty patients with a median age of 46.5 years (range, 18-64) were included. Donors were HLA-identical sibling (n=8), matched (n=42) or mismatched (n=10) unrelated. The toxicity was moderate, resulting in a cumulative incidence of non-relapse mortality (NRM) at 1 year of 8% (90% confidence interval: 2-15%). Acute GvHD grade II-IV and grade III/IV was noted in 53% and 14%, respectively. Chronic GvHD at one year was seen in 41%. After a median follow-up of 24 months the cumulative incidence of relapse was 36% (90% confidence interval: 24-48) and 51% (90% confidence interval: 37-65) at 1 and 2 years, respectively. The estimated 2-year disease-free and overall survivals were 36 and 48%, respectively. Treosulfan, etoposide and cyclophosphamide followed by AHSC has a favorable toxicity profile with low NRM and therefore represents a potential alternative regimen for ALL in 1. CR (NCT00682305).

Neutrophils and renal failure.

In many diseases and acute inflammatory disorders, important components of pathological processes are linked to the neutrophils' ability to release a complex assortment of agents that can destroy normal cells and dissolve connective tissue. This review summarizes the mechanisms of tissue destruction by neutrophils and the role of kidney-specific factors that promote this effect. Nicotinamide adenine dinucleotide phosphate H (NADPH) oxidase is a membrane-associated enzyme that generates a family of reactive oxygen intermediates (ROI). There is increasing evidence that ROIs are implicated in glomerular pathophysiology: ROIs contribute to the development of proteinuria, alter glomerular filtration rate, and induce morphological changes in glomerular cells. Specific neutrophil granules contain microbicidal peptides, proteins, and proteolytic enzymes, which mediate the dissolution of extracellular matrix, harm cell structures or cell function, and induce acute and potentially irreparable damage. Although both ROI and neutrophil-derived proteases alone have the potential for tissue destruction, it is their synergism that circumvents the intrinsic barriers designed to protect the host. Even small amounts of ROI can generate hypochlorus acid (HOCl) in the presence of neutrophil-derived myeloperoxidase (MPO) and initiate the deactivation of antiproteases and activation of latent proteases, which lead to tissue damage if not properly controlled. In addition, neutrophil-derived phospholipase products such as leukotrienes and platelet-activating factor contribute to vascular changes in acute inflammation and amplify tissue damage. Increasing evidence suggests that mesangial cells and neutrophils release chemotactic substances (eg, interleukin 8), which further promote neutrophil migration to the kidney, activate neutrophils, and increase glomerular injury. Also, the expression of adhesion molecules (eg, intercellular adhesion molecule 1 on kidney-specific cells and beta-2-integrins on leukocytes) has been correlated with the degree of injury in various forms of glomerulonephritis or after ischemia and reperfusion. Together, these results suggest that neutrophils and adhesion molecules play an important role in mediating tissue injury with subsequent renal failure. Conversely, chronic renal failure reduces neutrophil function and thereby can increase susceptibility to infection and sepsis.

Sepsis and septic complications in the surgical patient: who is at risk?

The estimation of patients who are at risk for infection, sepsis, and organ dysfunction/failure is crucial not only for inclusion in treatment algorithms but also for entry into appropriate clinical trials of prophylaxis and therapy. Patients on the surgical service who have sustained major trauma or who have undergone transplantation are clearly at the greatest risk. Other immunosuppressed patients at risk for sepsis include those receiving myelosuppressive chemotherapy, those with overwhelming malignancy, and those who suffer from cirrhosis, diabetes mellitus, and severe malnutrition. We have focused on the trauma patient, in whom infection and organ failure are the leading causes of late death, major morbidity, and prolonged hospital stay. Over a 10 yr period, we have surveyed a number of host defense parameters that pertain to an adequate immune response and found a suppressed response shortly after injury in many. All were anergic to a standard skin test panel, and the duration of anergy varied with the clinical course of infection. Immunoglobulin levels were low after major injury as well as specific antibodies to both Gram-positive and Gram-negative organisms. The ability of serum from the trauma patient to opsonize heat-killed bacteria was markedly depressed 24 h after injury in those patients who subsequently died of infection. Class II major histocompatibility antigen expression on peripheral blood monocytes correlated closely with clinical outcome and led to the development of an Outcome Predictive Score. This score can identify patients within hours of hospitalization who are at risk of subsequently developing overt clinical infection and sepsis. Intervention then can be applied to such at-risk populations prior to the onset of sepsis and to evaluate the efficacy of prophylaxis. Patients in whom prophylaxis fails could be eligible for trials of therapeutic intervention as well.

Inhibition of neutrophil migration at the site of infection increases remote organ neutrophil sequestration and injury.

Up-regulation of the leukocyte beta 2 integrin, CD18, is a key event in neutrophil-endothelial adhesion and neutrophil-mediated organ injury. Inhibition of CD18 with monoclonal antibodies reduces lung and liver neutrophil sequestration in animal models of Gram-negative bacteremia or endotoxemia. However, with a persistent septic challenge, interference with host leukocyte phagocytic defense could adversely affect outcome. To assess the effects of inhibiting CD18 on organ neutrophil responses, bacteremia, and organ injury after fecal peritonitis, mice underwent cecal ligation and puncture (CLP). At the time of CLP and 12 h later, mice received intravenous anti-CD18 antibody or control IgG. At 3, 6, and 18 h after CLP, lung and liver tissue neutrophil content were measured by myeloperoxidase (MPO) assay, peritoneal cells and blood leukocytes were differentially counted, blood was cultured, and serum aspartate aminotransferase was measured. There was a significant reduction in peritoneal neutrophil migration and an increase in blood neutrophils after anti-CD18 treatment compared with results from treatment with the control antibody. In the anti-CD18-treated group, liver MPO was increased fivefold at 6 and 18 h, while lung MPO was increased two-fold at 18 h when compared with the control antibody-treated group. The anti-CD18-treated group also had an increase in bacteria cultured from the blood at 6 and 18 h and an increase in serum aminotransferase at 18 h. Our data demonstrate that peritoneal neutrophil migration in response to an endogenous fecal challenge is CD18-dependent, and that this mechanism forms a vital part of host defense. Inhibition of CD18 increased neutrophil sequestration in the liver and lung and increased liver injury. This study demonstrates a paradoxical increase in organ neutrophil sequestration using a leukocyte anti-adhesion therapy during sepsis and suggests that anti-adhesion therapies targeted towards neutrophil may worsen outcome if given during an ongoing, localized infection.

Is laparoscopic appendectomy the new 'gold standard'?

OBJECTIVES: To determine the efficacy of laparoscopic appendectomy compared with open appendectomy in patients with acute appendicitis and to compare the morbidity between the two groups. DESIGN: Prospective sampling of 102 patients who underwent diagnostic laparoscopy and laparoscopic appendectomy for acute appendicitis and retrospective hospital chart review of 204 patients who underwent open appendectomy for acute appendicitis. RESULTS: The mean +/- SD duration of surgery was 83 +/- 29 minutes in the laparoscopic group and 64 +/- 30 minutes in the open appendectomy group (P < .001). Hospital stay was shorter in the laparoscopic group (P < .04). There was no difference in the complication rate between the patients who underwent laparoscopic appendectomy (13%) and the patients who underwent open appendectomy (11%). The occurrence of postoperative ileus was correlated with the duration of operation (P < .01) but not with laparoscopic appendectomy. CONCLUSIONS: The results confirm that laparoscopic appendectomy had a longer time of surgery, a shorter hospital stay, and no difference in complications. Further investigation will likely establish that laparoscopic appendectomy can be considered the "gold standard."

Differences between bacterial species shown by simultaneous assessment of neutrophil phagocytosis and generation of reactive oxygen intermediates in trauma patients.

HYPOTHESIS: Previous studies on alterations in phagocytosis and bacterial killing after trauma have yielded conflicting results. We hypothesize that these changes are variable, depending on the species of bacteria used to assay these variables. DESIGN: Blood samples from patients were assayed by means of flow cytometry for phagocytosis and reactive oxygen intermediate generation. Several common clinical pathogens were used: Escherichia coli, Klebsiella pneumoniae, and Staphylococcus aureus. Results were compared with those from controls. SETTING: Regional level I trauma center. PATIENTS: Ten consecutive patients were studied with E. coli and K. pneumoniae. Five of these were also studied with S. aureus. Patients were 18 years of age or older, with an Injury Severity Score of 16 or more. Patients who were taking corticosteroids before hospital admission or who were administered corticosteroids before blood was drawn were not studied. Isolated head injuries or limb fractures were also excluded. Controls consisted of healthy volunteers. MAIN OUTCOME MEASURES: The ingestion of bacteria by neutrophils and the generation of reactive oxygen intermediates. RESULTS: After trauma, phagocytosis of E. coli was enhanced, whereas ingestion of K. pneumoniae was depressed. Ingestion of S aureus remained unchanged. The generation of reactive oxygen intermediates was depressed after incubation with E. coli and unchanged with K. pneumoniae, but enhanced with S. aureus. CONCLUSIONS: Neutrophil response to trauma is dependent on which bacterial species the cell is attempting to kill. This may, in part, explain why only a limited number of bacterial species cause a significant proportion of early infections after trauma.

Constituent analysis may permit improved diagnosis of intra-abdominal abscess.

BACKGROUND: Intra-abdominal abscesses (IAA) often fail to resolve with intravenous antibiotics alone and frequently require drainage. Diagnosis of IAA in postoperative patients with other likely sources of infection is very difficult. PATIENTS AND METHODS: In order to characterize IAA and identify parameters that might facilitate diagnosis, we prospectively examined peripheral blood and pus of 15 consecutive patients with IAA and compared them to samples from 34 consecutive patients with soft-tissue abscesses (STA). RESULTS: Serum interleukin (IL)-10 was elevated in IAA patients, while abnormally detectable serum IL-4 was demonstrated in the pus of both IAA and STA patients. IL-10 in IAA pus was more than 11-fold higher than in STA, whereas IL-4 in pus was similar in both types of abscesses. Both IL-4 and IL-10 were 4- to 10-fold higher in IAA and STA pus than in corresponding patient serum. Serum lysozyme was, however, significantly elevated in all abscess patients. CONCLUSIONS: The presence of IL-4 and IL-10 may indicate a T-helper 2 lymphocyte response in the etiology of abscess formation and persistence, although precise determination of T-helper 1-related cytokines is needed to verify this. Serum lysozyme and IL-10 may be reliable and relatively inexpensive diagnostic aids.

Inflammatory response after abdominal trauma, infection, or intestinal obstruction measured by oxygen radical production in peritoneal fluid.

BACKGROUND: Reactive oxygen intermediates (ROI) have been implicated in many pathophysiological processes of inflammatory tissue damage and tissue repair. In the present study we compared the production of ROI in three different types of tissue damage in surgical patients. METHODS: Peritoneal fluid specimens were harvested during the initial operation and postoperatively from 25 surgical patients with abdominal trauma, intraabdominal infection, and intestinal obstruction. The optical density at 412 nm, representing the peroxidation of hemoglobin, was measured to assess intraperitoneal ROI production. Patients were categorized into 3 groups: (A) infected patients with good outcome, (B) patients after trauma or obstruction with good outcome, and (C) patients with poor outcome due to persistent or secondary infection and multiple organ failure. Analysis of variance (ANOVA) and paired t test were used for statistical analysis. RESULTS: Overall, the ROI production decreased significantly at days 2 and 3 compared with day 0 and 1 (P = 0.0013). No initial differences of intraoperative ROI concentrations were found among the three groups; however, patients with a poor outcome showed increased ROI values after 4 to 5 days (P = 0.038) when compared with the good outcome group. CONCLUSIONS: We have demonstrated that intraperitoneal ROI production (1) can be measured in patients with intraabdominal tissue damage, (2) is not different between patients with intraabdominal infections, abdominal trauma, or intestinal obstruction, and (3) correlates with the clinical picture and the presence of an inflammatory intraabdominal focus or tissue damage.

Heparin binding protein (CAP37) is an opsonin for Staphylococcus aureus and increases phagocytosis in monocytes.

Heparin binding protein (HBP), also known as cationic antibiotic protein (CAP37) or azurocidin, is stored in azurophilic granules of neutrophils and is released to the extracellular space when granulocytes phagocytose Staphylococcus aureus. We investigated whether extracellular HBP also has the potential to increase phagocytosis of S. aureus by other phagocytes. We used flow cytometry to characterize the binding of HBP to S. aureus and to simultaneously measure phagocytosis and superoxide production of opsonized S. aureus in monocytes and granulocytes. Our results demonstrate that HBP is a strong opsonin for S. aureus, and that monocytes, but not granulocytes, increase phagocytosis of HBP-treated S. aureus. However, HBP-treated S. aureus increases the production of superoxide in both monocytes and granulocytes as compared with untreated S. aureus. These findings support the role of granulocytes in the afferent limb of inflammation and demonstrate that HBP, when released from activated granulocytes, potentiates bacterial uptake in monocytes and enhances the potential of microbial killing in monocytes and granulocytes.

The pulmonary inflammatory response to experimental fecal peritonitis: relative roles of tumor necrosis factor-alpha and endotoxin.

The roles of endotoxin (LPS) and tumor necrosis factor-alpha (TNF-alpha) in the causation of organ injury during sepsis are unclear. To study LPS and TNF-alpha in the genesis of lung inflammation after cecal ligation and puncture (CLP), we used endotoxin-resistant (C3H/HeJ) and endotoxin-sensitive mice (C3H/HeOuJ). We examined lung neutrophil sequestration, interleukin 1 (IL-1)beta mRNA expression, IL-1 beta protein expression, and injury. We also determined the expression of two C-X-C chemokine mRNAs, macrophage inflammatory protein-2 (MIP-2) and KC, in the lung to determine whether in vivo, endotoxin, or TNF-alpha are significant modulators of MIP-2 and KC mRNA expression. After CLP, increased neutrophils sequestrated in the lungs of both strains of mice and coincided with an increase in expression of IL-1 beta, MIP-2 and KC mRNAs, and IL-1 beta protein. Lung and serum TNF-alpha were significantly increased in the C3H/HeOuJ strain but not in the C3H/HeJ strain. Histologic studies of the lung revealed similar injury in both strains. Our results suggest that bacterial factors other than endotoxin cause lung neutrophil sequestration and injury after CLP and, further, that TNF-alpha production is not a prerequisite. Our findings also suggest a potential role for local pulmonary chemokine production in the control of neutrophil sequestration after CLP.

Quantitative evaluations of gap junctions in old rat brown adipose tissue after cold acclimation: a freeze-fracture and ultra-structural study.

The morphological and functional modifications of brown adipose tissue (BAT), the tissue responsible for non-shivering thermogenesis, are well established during the phases of active stimulation (i.e. neonatal period and cold acclimation) in young animals. The 'active' brown adipocytes are filled with numerous small lipid vacuoles and large mitochondria packed with cristae rich in the protonophore uncoupling protein (UCP), whereas the 'quiescent' cell shows larger, confluent vacuoles and smaller mitochondria with rarefied cristae poor of the uncoupling protein. It is well known from literature that also gap junctions (gjs), responsible for the electrical coupling among adjacent adipocytes, modify their size following the physiological stimulus in young animals. This is in agreement with the morphology of the functionally active brown adipocyte, i.e. the multilocular, UCP-positive cell. Although the presence of the BAT in old animals is well documented, less is known about its reactivity to physiological stimuli. The present work demonstrates that after cold acclimation brown adipocytes of old rats (2 years) change their ultrastructure in a similar way as in young rats. A quantitative analysis of gap junction areas on replicas obtained by the freeze fracture technique, showed that gj increase in size (mean area 53.2 vs 110.4 x 10(-3) microns2, p = 0.003). All these morphological modifications are quite similar to those observed in BAT of young and young adult rats, supporting the hypothesis of a physiological role of brown adipose tissue at every age.

[Chylothorax after blunt thoracic trauma]. / Chylothorax nach stumpfem Thoraxtrauma.

The diagnosis of chylothorax following blunt chest trauma is rare, only few cases have been reported. We describe three patients with chylothorax following blunt chest trauma. Conservative treatment consists of drainage, in severe cases mechanical ventilation with PEEP and total parenteral nutrition. In case of persisting and/or increasing chylus production, thoracotomy and ligation of the thoracic duct may be required. In all of our patients thoracostomy was the definite therapeutic modality, no thoracotomy was necessary.

[The role of transarterial embolisation in the treatment of patients with abdominal injuries]. / Transarterielle Embolisation. Stellenwert in der Behandlung von Patienten mit Abdominaltrauma.

INTRODUCTION: The role of transarterial embolisation in patients with abdominal injuries is controversial. Some trauma centres advocate routine angiography, whereas others believe in restricted indications such as increasing haematomas or persistent/recurrent haematuria. METHOD: We prospectively studied 167 patients with blunt and penetrating abdominal trauma. We used restricted indications for angiography and embolisation. RESULTS: Eleven of 167 patients with abdominal trauma (7%) were treated with angiography and embolisation., Overall, three of 11 patients (27%) with angiography and embolisation were treated emergently and eight of them (73%) at an average of 7.3 days. There were no complications due to the embolisation procedure, and all bleeding could be stopped. CONCLUSION: Transarterial angiography and embolisation is an important and safe tool in the treatment of acute abdominal injury when used for restricted indications. We believe this should not be performed as a routine procedure, especially in unstable patients.