[An outbreak of Q fever in The Netherlands--possible link to goats]. / Een uitbraak van Q-koorts in Nederland--mogelijk verband met geiten.

In 2007, 73 cases of Q fever were identified through reports and retrospective analyses; the affected region extended from Tilburg in the southwest to Arnhem in the northeast. The infections arose in late spring, particularly in May and June. Several spontaneous abortions due to Q fever occurred on 4 dairy goat farms in the same region. The national incidence of spontaneous abortion due to Q fever was 6 cases in 2006 and 7 in 2007. Climatically, this southern region was extraordinarily dry during April 2007. All pregnant women from a small region with the highest incidence in northeast North Brabant were called for diagnostic testing. Infected patients were followed for symptoms and ultrasound was performed as indicated. A definitive source of the infection could not yet be identified. Favourable climatic conditions were suspected as the cause for the combination of widespread dissemination among goats and transmission to humans. Q fever is a zoonosis caused by Coxiella burnetti, a microorganism dispersed in great numbers in the area in which an infected animal gives birth. C. burnetti is particularly resistant to chemical and physical factors and can disperse by air across large distances under dry climatic conditions. Q fever should be considered in patients in The Netherlands who present with lower airway infection and, in rare cases, hepatitis. Reporting atypical clusters ofpneumonia to the Municipal Health Service (GGD) is advisable. The GGD maintains close contact with Animal Health Services, which is aware of current infectious animal diseases. Targeted investigation can identify the source of infection and eliminate it. Greater awareness can prevent delays in diagnosis and treatment and help identify chronic forms at an early stage or prevent them.

Diversion of first blood volume results in a reduction of bacterial contamination for whole-blood collections.

BACKGROUND AND OBJECTIVES: In a previous study we established a reliable setpoint for the prevalence of bacteria in whole blood. In the present study we investigated the possible preventive effect, of diversion of the first 10 ml of a blood donation, on the bacterial contamination rate. MATERIALS AND METHODS: To divert the first 10 ml of a whole-blood donation, we used a special five-bag system equipped with a Composampler device. After venepuncture, the first 10 ml of a donation was sampled into a vacutainer tube. This was followed by the collection of the whole-blood unit. The extra bag allowed direct sampling of the final donation in a closed system for BacT/Alert. Whole-blood samples were taken after storage (2-14 h at 20 degrees C) and subsequent mixing. BacT/Alert culture bottles were incubated until positive, or for 7 days if negative. Confirmation and identification of positive cultures was performed according to internationally recognized standard reference methods. RESULTS: The prevalence of bacteria in whole blood, as determined by using standard collection techniques, was 0.35% (95% confidence interval 0.27-0.44%, n = 18 257). After diversion of the first 10 ml this value was significantly lower: 0.21% (P < 0.05, 95% confidence interval 0.12-0.35%, n = 7087). Most strikingly, a reduction in the frequency of staphylococcal species was observed (P < 0.02, reduction from 0.14 to 0.03%). CONCLUSIONS: Diversion of the first 10 ml of blood was shown to contribute significantly to a reduction in the prevalence of superficial skin bacteria in whole-blood units. In our opinion, blood collection systems should be adapted to use the first 10-30 ml of a whole-blood donation for testing purposes.

Determination of the degree of bacterial contamination of whole-blood collections using an automated microbe-detection system.

BACKGROUND: The prevalence of bacterial contamination in whole-blood collections, either with immediate sampling or sampling after overnight storage as whole blood at 20 degrees C, is determined. STUDY DESIGN AND METHODS: Whole blood was collected under blood bank conditions in special five-bag systems, allowing sampling in a closed system for culture bottles. Samples were taken within 2 hours after collection (Group 1) or after overnight storage of the whole blood at 20 degrees C (Group 2). Culture bottles were incubated for 7 days, and positive samples were entered on agar plates for confirmation and determination. RESULTS: In Group 1, 9219 units were tested; 27 units were positive with positive subculture, that is, 0.29 percent with a 95% CI of 0.19 to 0.42 percent. In Group 2, 9038 units were tested; 36 units were positive with positive subculture, that is, 0.39 percent with a 95% CI of 0.28 to 0.55 percent. No significant difference could be found between the two test groups. The majority of bacteria were either Staphylococcus (all coagulase-negative) or Propionibacterium species. CONCLUSION: For a total of 18,257 units, 0.34 percent (CI, 0.25-0.44) of whole-blood collections appeared to have bacterial contamination (mainly skin-derived). Overnight storage of whole blood at 20 degrees C did not have a significant effect on the prevalence of bacterial contamination.

[Rat bite fever after a bite from a tame pet rat]. / Rattenbeetziekte na een beet van een tamme huisrat.

A 43-year-old woman presented with a generalized febrile illness, an exanthema with mixed maculopapulous and pustulous eruptions on the lower halves of the extremities, elbows, knees, palms and soles. There was also severe arthralgia and asymmetric arthritis. The diagnosis was rat bite fever. The disease became manifest eight days after she was bitten by a pet rat. Rat bite fever can easily be missed, even after adequate anamnesis and physical examination, while the differential diagnostic considerations are numerous. Our patient was cured completely after intravenous administration of penicillin G. Antimicrobial therapy was completed by an oral course of doxycycline.

Sterility testing of blood products in 1994/1995 by three cooperating blood banks in The Netherlands.

BACKGROUND AND OBJECTIVES: Dutch regulations require blood banks to check the sterility of random blood components to detect contamination during preparation. MATERIALS AND METHODS: We reviewed the results of two years' testing, using standard bacteriologic methods. RESULTS: Of all tested components, 0.5% were contaminated, with Staphylococcus epidermidis being the most frequently detected microorganism. Platelet concentrates showed higher rates of contamination, especially when pooled. Leukocyte-depleted red cell concentrates showed much lower contamination than red cell concentrates that had not been leuko-depleted. CONCLUSIONS: The rate of contamination compares well with that reported by others in the literature. Since most contamination occurs from the phlebotomy site, most of the bacteria detected were derived from the skin. Leukocyte reduction lowers the rate of contamination.

[Histoplasma capsulatum infection, a manifestation of AIDS unusual for The Netherlands]. / Histoplasma capsulatum-infectie, een voor Nederland ongewone manifestatie van de ziekte AIDS.

The history of 29-year-old male from Surinam with antibodies to HIV-1 and long-lasting fever, lymphadenopathy, pain in the right upper abdomen and a granulomatous hepatitis is described. The patient suffered from disseminated histoplasmosis, a fungal disease rare in The Netherlands, which is the indicator disease for the diagnosis of AIDS (CDC-IVCI). It is stressed that in seropositive patients coming from endemic areas, including Surinam, the possibility of this disease should be considered.

Bifidobacterium, Bacteroides, and Clostridium spp. in fecal samples from breast-fed and bottle-fed infants with and without iron supplement.

Bifidobacterium, Bacteroides, and Clostridium spp. isolated from the feces of 23 neonates during the first 3 months of life were identified. Of the 23 neonates, 10 were breast fed, 6 received an infant formula with iron supplement (5 mg/liter), and 7 received the formula without iron supplement (iron concentration, less than 0.5 mg/liter). The Bifidobacterium spp. most frequently isolated from the three groups of infants were B. longum, B. breve, B. adolescentis, and B. bifidum. The bacteroides spp. most frequently isolated were B. fragilis and B. vulgatus. The most common Clostridium sp. in the three groups of infants was C. perfringens. The type of milk did not select for species of Bifidobacterium, Bacteroides, or Clostridium, except for Clostridium butyricum, which was isolated significantly more often from bottle-fed infants with iron supplement than from the other groups, and Clostridium tertium, which was more often isolated from breast-fed infants. The species of the three anaerobic genera did not persist for a long period of time in the three groups of infants.

Antibodies raised against rough mutants of Escherichia coli and Salmonella strains are opsonic only in the presence of complement.

The opsonic capacity of antisera raised in rabbits against rough (R) mutants and smooth (S) parent strains of Escherichia coli and Salmonella typhimurium were studied. All specific antibodies in the antisera belonged to the immunoglobulin G (IgG) class. Radioactively labeled bacteria were preincubated in various dilutions of antisera, in which complement was inactivated. Fresh normal rabbit serum, as a standard complement source, was used in some experiments. After preincubation, washed bacteria were added to normal human neutrophils. Opsonization of R mutants for 5 min in 5% fresh normal rabbit serum resulted in effective phagocytosis; S strains needed at least a 30-min opsonization time or 20 to 50% serum. After incubation for 5 min in diluted, homologous antisera, phagocytosis of S strains was optimal, but preincubation of R mutants in diluted, homologous antisera did not lead to amelioration of phagocytosis compared with that of bacteria preincubated in buffer only. However, when fresh normal serum was added to homologous antisera, uptake of R mutants occurred at a faster rate than that of bacteria opsonized in fresh serum alone. Using six clinical isolates of members of the family Enterobacteriaceae, we found that, with or without complement, antisera raised against E. coli J5 or S. typhimurium Re had, with the exception of one strain, no opsonic activity for these strains. Thus, the protective effect of R antisera in gram-negative bacteremia, as shown by several investigators, is unlikely to be mediated through enhanced opsonization of invading bacteria by IgG antibodies directed against these R mutants.

Complement activating and opsonic capacity of monoclonal antibodies raised against Escherichia coli O111 and its rough mutant J5.

Six monoclonal antibodies raised against Escherichia coli O111 and against its rough mutant J5 (chemotype Rc) were studied. One IgG2A, one IgM anti-J5, and one IgG2A anti-O111 monoclonal antibody did not bind to lipopolysaccharides of the homologous strain, but cross-reacted with heterologous gram-negative rods in an enzyme-linked immunosorbent assay. These three monoclonal antibodies activated complement when incubated with homologous or heterologous strains, but were opsonic neither in the presence nor in the absence of complement. The other three monoclonal antibodies were directed against lipopolysaccharide of the homologous strain, but showed no cross-reactivity. The IgG3 and one IgM anti-J5 monoclonal antibodies activated complement and were opsonic only in the presence of complement. The IgM anti-O111 monoclonal antibody activated complement and was opsonic both in the presence and absence of complement. Thus, the outcome of the interaction between bacteria, antibodies, and complement is influenced primarily by whether antibodies are directed against lipopolysaccharides or against other cell wall components.

Cross-reactivity of monoclonal antibodies against lipopolysaccharides of gram-negative bacteria.

Monoclonal antibodies were produced against Escherichia coli O111, Escherichia coli J5, and the rough (R) mutant of Salmonella typhimurium M206, and tested by enzyme-linked immunosorbent assay against lipopolysaccharides of several gram-negative strains. The monoclonal antibodies were also identified with an immunoblotting assay. Anti-Escherichia coli O111 monoclonal antibodies reacted only with homologous O antigens. Anti-J5 monoclonal antibodies cross-reacted with core lipopolysaccharide, especially with Rc lipopolysaccharide. IgM anti-J5 monoclonal antibodies showed more extensive cross-reactivity than IgG3 monoclonal antibodies. Anti-Re monoclonal antibodies cross-reacted weakly with all rough lipopolysaccharide tested. Thus, the varying specificity of these monoclonal antibodies seems to indicate that the core regions in the lipopolysaccharides of various gram-negative bacteria are not similar.

Effect of iron on serotypes and haemagglutination patterns of Escherichia coli in bottle-fed infants.

Strains of Escherichia coli isolated from faecal specimens of ten infants receiving breast milk, six receiving a cow-milk preparation with iron supplement (5 mg/l) and six the preparation without iron supplement (less than 0.5 mg/l), were serotyped and examined for their haemagglutinating activity. The Escherichia coli flora of breast-fed and bottle-fed infants consisted of one resident strain, accompanied by one or more transient strains. Changes in the serotype of the Escherichia coli flora and in the frequency of occurrence of strains associated with urinary tract infections were more often seen in bottle-fed than in breast-fed infants. In breast-fed and bottle-fed infants without iron supplement most strains of Escherichia coli were non-haemagglutinating, while most strains in infants bottle-fed with iron supplement showed mannose-resistant haemagglutination. It is concluded that human milk favours the establishment of a stable non-pathogenic Escherichia coli flora and that a low iron content in standard cow-milk preparation favours colonization with non-adherent strains of Escherichia coli.

Detection of antibodies against lipopolysaccharides of Escherichia coli and Salmonella R and S strains by immunoblotting.

Antisera raised against several smooth and rough strains of Escherichia coli and Salmonella typhimurium were tested against lipopolysaccharides (LPS) of homologous and heterologous strains. The LPS were separated by sodium dodecyl sulfate-gel electrophoresis, transferred to nitrocellulose paper, and overlaid with antisera. The results showed that antisera raised against smooth strains reacted with high- as well as low-molecular-weight bands of their corresponding LPS and showed very few cross-reactions. Anti-E. coli J5 antiserum cross-reacted with few strains in the core region. But, anti-S. typhimurium Ra antiserum cross-reacted with many more strains. When these sera were absorbed with either the homologous- or a heterologous-positive strain, reactions were abolished. It appears that reactions of anti-E. coli J5 antiserum and anti-S. typhimurium Ra antiserum with homologous and heterologous strains were not due to the same antibody. This immunoblotting technique proved to be a useful method to distinguish different antibodies in antiserum raised against LPS of gram-negative bacteria.

Effect of iron on neonatal gut flora during the first three months of life.

To study the effect of milk supplemented with iron on neonatal gut flora, faecal specimens of ten infants receiving breast milk, six receiving a cow-milk preparation supplemented with iron (5 mg/l) and seven receiving the same product without iron supplement (iron concentration less than 0.5 mg/l) were examined during the first 12 weeks of life. In breast-fed infants bifidobacteria was predominant, counts of Escherichia coli were low, and other bacteria were rarely present. Infants receiving fortified cow-milk preparation had high counts of Escherichia coli, counts and isolation frequency of bifidobacteria were low and other bacteria were frequently isolated. In those on unfortified cow-milk preparation isolation frequency of Escherichia coli, bifidobacteria and bacteroides was comparable with that in breast-fed infants; however, counts of Escherichia coli were high. It is concluded that the faecal flora of infants fed unfortified cow-milk preparation acquires characteristics of that found in breast-fed infants.

Effect of iron on neonatal gut flora during the first week of life.

Faecal specimens from 23 infants during the first week of life were compared. Ten infants received breast milk, six received cow-milk preparation supplemented with iron (+/- 5 mg/l) and seven unfortified cow-milk preparation (iron concentration less than 0.5 mg/l). Those on breast milk had low faecal pH, high counts of bifidobacteria and low counts of Enterobacteriaceae, bacteroides and clostridia. Infants receiving fortified cow-milk preparation had a high faecal pH and high counts of Enterobacteriaceae and putrefactive bacteria such as bacteroides and clostridia. Counts of bifidobacteria were also high. In those on unfortified cow-milk preparation a slow rise was observed in counts of Enterobacteriaceae followed by an increase in counts and isolation frequency of bifidobacteria: the latter was still rising on day 7. It is concluded that a low iron content in standard preparations of cow's milk enhances resistance of the neonatal gut to colonization.

Effects of lysosomotropic amines on human polymorphonuclear leucocyte function.

Lysosomotropic agents interfere with lysosome function. We studied the effects of the lysosomotropic amines: lidocaine, diphenylamine and dansylcadaverine on several functions of human polymorphonuclear leucocytes (PMN): enzyme release, phagosome-lysosome fusion, superoxide anion generation upon stimulation with opsonized bacteria, and phagocytosis and killing of opsonized Staphylococcus aureus. Lidocaine depressed all cellular functions tested. Diphenylamine reduced enzyme release and phagosome-lysosome fusion in phagocytosing PMN. This was accompanied by an increase in superoxide anion generation. Dansylcadaverine enhanced enzyme release and phagosome-lysosome fusion, and reduced superoxide anion generation. Neither of these two agents influenced bacterial uptake; bacterial killing was impaired only in dansylcadaverine treated cells. Cadaverine, an analogue that does not penetrate cells, had no effect on any of the functions tested.

Synergy between the iron chelator deferoxamine and the antimicrobial agents gentamicin, chloramphenicol, cefalothin, cefotiam and cefsulodin.

Synergy between the iron chelator deferoxamine in the presence or absence of ascorbic acid and gentamicin, chloramphenicol, cephalothin, cefotiam or cefsulodin, used against Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Klebsiella pneumoniae, proteus mirabilis and species of Salmonella, Enterobacter, Pseudomonas and Providencia, was determined by measuring the effect of the drugs and combination of drugs on growth of the bacteria in an automated turbidimeter. The combination of drugs was considered to be synergistic when the growth inhibiting effect of the combination was greater than that of the combined action of each of the drugs separately. Deferoxamine plus ascorbic acid together with either gentamicin or cefsulodin showed synergy in 10 out of 10, and 5 out of 6 cultures respectively, whereas deferoxamine plus ascorbic acid with chloramphenicol, cephalothin or cefotiam was synergistic in 6 out of 14, 5 out of 11, and 3 out of 6 cultures. This synergistic effect was much lower when microorganisms were incubated with deferoxamine combined with the various antibiotics but without ascorbic acid. Ascorbic acid alone had no synergistic effect. When deferoxamine was saturated with iron, its antibacterial effect was completely abolished.

Inhibition of bacterial multiplication by the iron chelator deferoxamine: potentiating effect of ascorbic acid.

Since iron is essential for the multiplication of microorganisms, the effect of the iron chelator deferoxamine, with or without ascorbic acid, on the growth of 43 strains of Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Alcaligenes faecalis, Neisseria meningitidis and species of Salmonella, Enterobacter, Pseudomonas and Providencia, was investigated with the use of an automated turbidimeter. Addition of deferoxamine (25-400 micrograms/ml) to the incubation medium was inhibitory in a dose-dependent fashion. At concentrations between 200-400 micrograms/ml, growth was about 25% lower than control values. However, when ascorbic acid (100 micrograms/ml) was added to the culture medium, this antimicrobial activity of deferoxamine was significantly increased to on average 75% of the control value (p less than 0.05). Ascorbic acid alone had no bacteriostatic properties. Growth in the presence of 200 micrograms/ml deferoxamine combined with 100 micrograms/ml ascorbic acid was significantly lower than that in control media without additions (p less than 0.001). Addition of ferric citrate to the culture medium at a concentration sufficient to saturate all of the deferoxamine with iron, abolished the growth inhibiting effect of deferoxamine. The results provide evidence that deferoxamine is bacteriostatic due to its capacity to deplete iron which would otherwise be used for bacterial multiplication, and that ascorbic acid enhances this antibacterial property of deferoxamine.

Semielectronic turbidimeter for automated monitoring of bacterial growth in test tubes.

An automated turbidimeter for measuring bacterial growth in ordinary test tubes is described. The device records and prints adsorbance, expressed as Klett units, of 60 cultures every 15 min. Provision is made for either aerobic or anaerobic incubation. The device is adaptable to modification, depending upon local requirements and availability of computation facilities.