Incubation of whole blood at 39°C augments gamma interferon (IFN-γ)-induced protein 10 and IFN-γ responses to Mycobacterium tuberculosis antigens

A rarely challenged dogma in cell-mediated immune (CMI) assays is the incubation temperature, 37°C. Fever augments proinflammatory immune responses in vivo, and the aim of this study was to explore whether incubation at fever-range temperature could increase antigen-specific biomarker responses. We compared CMI responses following incubation of whole blood at 37°C and 39°C. Whole blood was obtained from (i) 34 healthy subjects whose blood was incubated with TB10.4 antigen, present in the Mycobacterium bovis bacillus Calmette-Guérin vaccine and many environmental mycobacteria; (ii) 8 TB patients and 8 controls incubated with Mycobacterium tuberculosis-specific antigens in the QuantiFERON-TB Gold test (QFT-IT); and (iii) from both groups incubated with a T cell mitogen. T cell responses (gamma interferon [IFN-γ]) and responses from antigen-presenting cells (IFN-γ-induced protein 10 [IP-10]) were determined. We further evaluated the effect of adding interleukin-7 (IL-7) and blocking IL-10 during incubation. In TB patients, IFN-γ and IP-10 levels were increased 4.1- and 3.4-fold, respectively, at 39°C incubation (P < 0.001). Similar results were seen after mitogen stimulation. In subjects responding to TB10.4, the effects were less pronounced and significant only for IP-10. Incubation at 39°C increased IP-10 and IFN-γ responsiveness to both antigens and mitogen in persons with baseline or initial low responses. Adding IL-7 and blocking IL-10 augmented the effects in synergy with fever-range temperature. Incubation at fever-range temperature vividly increases CMI responsiveness to antigen stimulation in vitro in tuberculosis patients and may increase the sensitivity of CMI assays.     

Serum YKL‐40 and gestational diabetes – an observational cohort study

To examine serum YKL‐40 in women developing gestational diabetes mellitus (GDM). In the present large observational cohort study of 1179 pregnant women, we determined serum YKL‐40 four times during pregnancy (at gestational age 12, 20, 25, and 32 weeks). Pregnancy outcome was obtained from medical records. Sixty‐eight women (5.8%) developed GDM. Serum YKL‐40 increased from gestational age (GA) 12 weeks and the following weeks in the women who developed GDM and was independent of BMI, parity, and maternal age (OR = 2.69, 95% CI: 1.45–5.00, p = 0.002). No association was found between serum YKL‐40 and the oral glucose tolerance test results. In conclusion, YKL‐40 significantly increased in pregnant women with GDM compared with women without GDM, probably reflecting the low‐grade inflammation of GDM. However, we did not find an association between serum concentrations of YKL‐40 in early pregnancy and the development of GDM and thus we conclude that YKL‐40 alone is not usable as a biomarker for early prediction of GDM.     

Unusual Distribution of Burkholderia cepacia Complex Species in Danish Cystic Fibrosis Clinics May Stem from Restricted Transmission between Patients

Forty-four of 48 Burkholderia cepacia complex strains cultured from Danish cystic fibrosis patients were Burkholderia multivorans, a distribution of species that has not been reported before. Although cases of cross infections were demonstrated, no major epidemic clone was found. The species distribution may represent the sporadic acquisition of bacteria from the environment.Burkholderia cepacia and related bacteria have emerged as significant pathogens in cystic fibrosis (CF) patients due to the risk of cepacia syndrome (a fatal necrotizing pneumonia with bacteremia), the organism''s innate multiresistance to antibiotics, and the transmissibility of bacterial strains between patients by social contact (10, 15). The genus Burkholderia encompasses more than 50 validly published species that can be divided into four groups (21). Strains colonizing the respiratory tract of CF patients are predominantly members of the B. cepacia complex (BCC), with 17 formally named species (23). Chronic infections typically involve a single strain, although strain displacements have been demonstrated (24).Most or all species of the BCC can colonize the lower airways of CF patients, although some of them are infrequently demonstrated. Studies from North America, Europe, and Australasia have shown that Burkholderia cenocepacia is the dominant species being recovered from 46 to 90% of colonized patients (4, 6, 13, 17, 19, 20). Different situations have been described in Lisbon, where contamination of saline solutions used in inhalant therapy by CF patients has resulted in a predominance of B. cepacia (5), and in France, where a small excess of B. multivorans (52%) over B. cenocepacia (45%) has been reported (3).Danish CF patients are treated in two centers, and respiratory cultures are routinely performed at the monthly visit to the outpatient clinic. Four hundred thirty-one patients were alive by 1 January 2007, and 24 (5.6%) were chronically infected with BCC species. A chronic infection was defined as the isolation of BCC bacteria from more than half of sputum cultures for more than 6 months (modified “Leeds criteria” for chronic Pseudomonas aeruginosa infection [14]), and/or the development of ≥2 precipitins measured by crossed immunoelectrophoresis (18). A total of 52 Danish CF patients are known to have been intermittently (n = 11) or chronically (n = 41) infected with BCC bacteria (Fig. ​(Fig.1).1). Intermittently colonized patients may be underrepresented in data from before the routine use of colistin-containing selective agar plates (9), and some of the recent BCC acquisitions may be reclassified as chronic infections with time. In retrospect, the first Danish patient was chronically infected with BCC in the late 1970s (18), but few cases were identified until 1990. The increased rate of BCC colonization after 1990 may be secondary to the widespread use of inhaled colistin for P. aeruginosa infection, which was introduced in the 1980s (11). Since 1993, the rate has stabilized at around three new cases per year (43 BCC acquisitions during 14 years) (Fig. ​(Fig.1).1). In the same time period, 174 Danish patients have been diagnosed with CF (on average, 12.4 ± 4.4 [mean ± standard deviation] new patients per year; range, 6 to 22).Open in a separate windowFIG. 1.Cumulative numbers of Danish CF patients experiencing a first-time isolation of BCC bacteria, separated by status (open bars, chronic infections; gray bars, intermittent colonizations).BCC isolates from 9 intermittently colonized and 39 chronically infected patients were available for characterization. One isolate per patient, cultured between 1994 and 2006, was included in the study. Allocation to species within the BCC was performed by partial atpD and recA sequencing (1); occasional isolates with no PCR product from either amplification were subjected to partial sequencing of fur (16). Two independent sequence-based identifications were thus obtained for all BCC isolates. Only three species were identified in Danish patients, and B. multivorans accounted for more than 90% of the isolates (Table ​(Table1).1). Pulsed-field gel electrophoresis (PFGE) genotypes were assessed after digestion with Xba and SpeI and interpreted as described previously (22). Thirty-eight BCC genotypes were disclosed by both enzymes, and five of the genotypes were identified in more than one patient (two to four patients). Some of these small clusters were epidemiologically related and probably reflect cases of cross infections. Two pairs of siblings each carried the same strain, and one additional patient harbored the same genotype as the two siblings treated in that CF center. Between 1994 and 2003, chronic infections with BCC of a single genotype were established in 4 patients treated in one center. A fourth cluster was composed of patients treated at both of the two Danish CF centers; a possible epidemiological relationship between these three patients was unknown. No patient-to-patient transmission could have occurred in the fifth cluster, where the same genotype was intermittently detected in two patients in 1994 and 1999, respectively, and established a chronic infection in a third patient in 2005. All BCC genotypes identified in more than one patient were B. multivorans.

TABLE 1.

Specific identification of 48 BCC strains isolated from Danish CF patients

Characterization of copulatory behavior in female mice: evidence for paced mating

In this study we characterized female mouse sexual behavior using a pacing paradigm similar to that used to evaluate sexual behavior in female rats. A pacing chamber was designed for use with mice and we compared the sexual behavior of female mice that were tested in both pacing and nonpacing paradigms and under different hormone conditions. We found that, like rats, female mice do pace their copulatory behavior by altering the temporal sequence of copulatory events. Female mice take longer to return to the male after an ejaculation, compared to either a mount or intromission. However, it is still unclear if female-paced mating serves the same functions as it does in female rats. More work is needed to confirm that paced mating induces hormonal changes needed for pregnancy as is the case in rats.     

Identification of viral agents associated with diarrhea in young children during a winter season in Beijing, China.

BACKGROUND: Viral diarrhea remains a major cause of childhood morbidity and mortality worldwide. Although rotavirus was extensively studied in China, few comprehensive studies of all viral agents related to diarrhea in children have been conducted. OBJECTIVES: Our study was performed to investigate the role of enteric viruses in acute diarrhea in our country and to evaluate methods that could be used in routine diagnostics. STUDY DESIGN: One hundred stool samples were collected from children under 5 years of age seeking medical care for acute diarrhea during the winter season 2000/2001 in Beijing Children's Hospital. All specimens were initially screened microscopically for leucocytes/red blood cells. Samples with negative results were analyzed for virus presence using commercial EIAs and/or in-house RT-PCRs. RESULTS: At least one viral agent was found in 67% of the specimens. The frequency of rotavirus, astrovirus, norovirus and enteric adenovirus was 59%, 8%, 6% and 2%, respectively. Dual infections were found in 9.0% (6/67) of the positive samples. The results from rotavirus and astrovirus EIAs were concordant with those of rotavirus and astrovirus RT-PCRs. CONCLUSIONS: Enteric viruses play an important role in pediatric diarrhea during the winter season in China. A combination of microscopic examination of stool samples with specific EIA assays to detect virus antigen in stool specimens may be suitable for routine diagnostics.     

European seroepidemiology network 2: Standardisation of assays for seroepidemiology of varicella zoster virus.

BACKGROUND: The aim of the European Sero-Epidemiology Network (ESEN2) is to harmonise the serological surveillance of vaccine-preventable diseases in Europe. OBJECTIVE: To allow comparison of antibody prevalence in different countries by standardising results into common units. STUDY DESIGN: For varicella zoster virus (VZV), a reference laboratory established a panel of 148 samples, characterised by indirect enzyme-immunoassay (ELISA), indirect immunofluorescence, and complement fixation test. Fifty-seven samples were also studied by the fluorescence antibody to membrane antigen test. The geometric mean of the antibody activity (GMAA) obtained from four ELISA determinations was used to characterise each sample of the panel as positive (GMAA: >100 mIU/ml), equivocal (GMAA: 50-100 mIU/ml) or negative (GMAA: <50 mIU/ml) for antibody to VZV (anti-VZV). Thirteen laboratories, using five different ELISA tests, tested the panel. RESULTS: Agreement with the reference laboratory was above 85% in all cases, and the R(2) values obtained from regression analysis of the quantitative results were always higher than 0.87. Finally, the regression equations could be used to convert national values into a common unitage. CONCLUSION: This study confirmed that results for anti-VZV obtained by different ELISA methods can be converted into common units, enabling the comparison of the seroprevalence profiles obtained in the participant countries.