Occupational allergy to flowers: immunoblot analysis of allergens in freesia, gerbera and chrysanthemum pollen.

High exposure to pollen from ornamental flowers can induce an IgE-mediated occupational allergy in florists and horticulture workers. We investigated IgE-binding antigens in chrysanthemum, freesia and gerbera pollen by immunoblot analysis and analysed the cross-reactivity of these pollen with birch, grass and mugwort pollen. In immunoblots with chrysanthemum pollen, major IgE-binding structures were seen with a molecular weight (MW) of approximately 25, 45 and 65 kD. In the immunoblots with freesia pollen, IgE from freesia pollen was directed against two proteins with an MW of approximately 15 kD. Most sera showed IgE binding to an approximately 15 kD band in gerbera pollen; with some sera additional bands were seen in the range of 30-50 kD. IgE binding to chrysanthemum pollen was inhibited by mugwort pollen only, whereas IgE binding to freesia pollen was suppressed by birch, grass and mugwort pollen. The inhibitory activity of birch and grass pollen extract on IgE binding to gerbera pollen extract was serum dependent and ranged from no inhibition to complete inhibition. Occupational exposure to many different flowers induced IgE against all three types of pollen. Exposure in greenhouses to gerbera flowers elicited mainly IgE against gerbera pollen. Mugwort pollen extract inhibited IgE binding to pollen from all three flowers.

Potential value of serum total IgE for differentiation between autoimmune pancreatitis and pancreatic cancer.

Autoimmune pancreatitis (AIP) is associated with a marked elevation of serum total IgG4 . Although there is evidence of autoimmunity in AIP, there are also signs of an allergic nature of its pathogenesis. Therefore, we determined both IgE and IgG4 in 13 patients with AIP, in 12 patients with pancreatic carcinoma and in 14 patients with atopic allergy and investigated the relationship between IgE and IgG4 . Total IgG4 was determined by automated nephelometry and total IgE by automated enzyme fluoroimmunoassay. Both total IgE and total IgG4 levels in patients with AIP were significantly higher than those in patients with pancreatic carcinoma (P = 0.0004 and P = 0.015, respectively). There was a significant correlation between the total IgE and total IgG4 levels in patients with AIP and patients with atopic allergy (r(s) =0.82, P=0.0006 and r(s) =0.88, P < 0.0001, respectively). The IgE/ IgG4 ratio in sera from patients with atopic allergy was significantly different (P = 0.0012) from this ratio in sera from patients with AIP. These results suggest that analysis of total IgE in serum might be useful in the differentiation between autoimmune pancreatitis and pancreatic carcinoma.

Occupational allergy in strawberry greenhouse workers.

BACKGROUND: Employees in strawberry greenhouses are highly exposed to several (potential) allergenic agents. However, no occupational allergy in this branch has been described before. First, the presence of work-related allergic symptoms in strawberry workers was explored. Second, we aimed to prove the concept that an IgE-mediated allergy could be responsible for work-related symptoms. To test the possibility of an IgE response secondary to cross-reactivity to birch or grass pollen, inhibition experiments were performed. METHODS: First, a questionnaire survey concerning work-related allergic symptoms among strawberry workers in the Netherlands was carried out. Second, 3 workers with work-related symptoms were investigated in detail. Skin tests, serum-specific IgE tests with home-made extracts of strawberry pollen and other possible allergenic agents of the strawberry greenhouse environment were executed. Furthermore, immunoblots and nasal provocations with strawberry pollen extract were performed. In addition, inhibition experiments were performed. RESULTS: 29 of 75 questionnaire respondents (38.7%) reported work-related symptoms. Sensitization to strawberry pollen was found in skin tests in all 3 employees with work-related symptoms. ELISA and immunoblotting with strawberry pollen showed positive results in 2 employees. Birch and grass pollen failed to inhibit IgE binding to strawberry pollen in 1 of 2 employees. Partial inhibition was seen in the second employee. Nasal provocation validated clinically relevant allergy to these pollens in 2 of 3 subjects. CONCLUSIONS: Allergic symptoms attributable to the workplace are present among a proportion of strawberry greenhouse employees. An IgE-mediated occupational allergy to strawberry pollen may contribute to these symptoms.

Serum tryptase and SCORMA (SCORing MAstocytosis) Index as disease severity parameters in childhood and adult cutaneous mastocytosis.

BACKGROUND: Skin lesions are the predominant clinical feature of the commonest form of mastocytosis. Mastocytosis is classified according to World Health Organization criteria. Determination of the levels of mast-cell mediators or their metabolites reflects the mast-cell burden. The extent of cutaneous mastocytosis can be assessed clinically using a scoring system (SCORing MAstocytosis; SCORMA Index) that we have developed. OBJECTIVE: Serum tryptase levels were compared with the SCORMA Index in a large group of paediatric and adult patients to investigate whether there was any correlation between the two. METHODS: The SCORMA Index in 64 patients (31 children and 33 adults) was compared with serum tryptase levels. The results of the first visit at which SCORMA and tryptase were evaluated were analysed. RESULTS: There was a positive correlation between the SCORMA Index and serum tryptase levels, indicating the value of the SCORMA Index in the assessment of mastocytosis with skin involvement. CONCLUSION: The results of this study showed that the SCORMA Index is a useful tool for evaluating the severity of cutaneous mastocytosis. The correlation between the SCORMA Index and serum tryptase levels underlines the benefit of the SCORMA Index as a clinical tool. Repeated SCORMA Index measurements can provide a rapid impression of changes in the clinical state of mastocytosis. This is particularly relevant in children, because taking blood samples from this group is much more difficult. The well-established methods for evaluation of disease severity may be expanded by the rapid SCORMA Index method.

Pancreatic fibrosis correlates with exocrine pancreatic insufficiency after pancreatoduodenectomy.

BACKGROUND: Obstruction of the pancreatic duct can lead to pancreatic fibrosis. We investigated the correlation between the extent of pancreatic fibrosis and the postoperative exocrine and endocrine pancreatic function. METHODS: Fifty-five patients who were treated for pancreatic and periampullary carcinoma and 19 patients with chronic pancreatitis were evaluated. Exocrine pancreatic function was evaluated by fecal elastase-1 test, while endocrine pancreatic function was assessed by plasma glucose level. The extent of fibrosis, duct dilation and endocrine tissue loss was examined histopathologically. RESULTS: A strong correlation was found between pancreatic fibrosis and elastase-1 level less than 100 microg/g (p < 0.0001), reflecting severe exocrine pancreatic insufficiency. A strong correlation was found between pancreatic fibrosis and endocrine tissue loss (p < 0.0001). Neither pancreatic fibrosis nor endocrine tissue loss were correlated with the development of postoperative diabetes mellitus. Duct dilation alone was neither correlated with exocrine nor with endocrine function loss. CONCLUSION: The majority of patients develop severe exocrine pancreatic insufficiency after pancreatoduodenectomy. The extent of exocrine pancreatic insufficiency is strongly correlated with preoperative fibrosis. The loss of endocrine tissue does not correlate with postoperative diabetes mellitus. Preoperative dilation of the pancreatic duct per se does not predict exocrine or endocrine pancreatic insufficiency postoperatively.

Elevated levels of plasma matrix metalloproteinase-9 in patients with atopic dermatitis: a pilot study.

Matrix metalloproteinase (MMP)-9 has been shown to play a role in the infiltration of inflammatory cells in various tissues. It is thus part of the pathogenesis of many inflammatory diseases, including asthma and allergic rhinitis/conjunctivitis. We compared plasma MMP-9 levels of 20 patients with atopic dermatitis (AD) with that of 17 control subjects. Additional outcome parameters consisted of the modified objective SCORing Atopic Dermatitis and the Three Item Severity score in patients, and peripheral blood leucocytes and eosinophils in both groups. Plasma MMP-9 levels were found to be significantly higher in patients compared with controls, supporting a role for MMP-9 in the pathogenesis of AD.

Comparison of serum tryptase and urine N-methylhistamine in patients with suspected mastocytosis.

BACKGROUND: The disease extent of mastocytosis can be assessed by measurement of mediators or their metabolites, secreted from mast cells. In the present study, we compared results of urinary N-methylhistamine measurements with analysis of total tryptase in serum from patients with suspected mastocytosis. METHODS: Tryptase in serum was determined with the UniCAP tryptase fluor-enzyme-immunoassay, according to the manufacturers' instructions (Pharmacia, Woerden, Netherlands). N-methylhistamine in urine was determined by competitive radioimmunoassay, according to the manufacturers' instructions (Pharmacia). RESULTS: A significant correlation between serum tryptase and urine N-methylhistamine was found both for 138 patients aged 14 or older (Spearman Rank r(s)=0.43, p<0.0001) and for 23 younger patients (Spearman Rank r(s)=0.46, p=0.0267). The between-run coefficient of variation of the tryptase assay was half (6.7%) of the one (13%) found with the urinary N-methylhistamine assay. Both for urine N-methylhistamine and serum tryptase, a significant difference was found between corresponding biopsies with an increased number of mast cell aggregates and biopsies without such an increase. The difference between tryptase levels however was stronger (Mann-Whitney: p=0.0012) than the difference between N-methylhistamine levels (Mann-Whitney: p=0.0140). CONCLUSION: Serum tryptase discriminates better than urinary N-methylhistamine between patients with an increased number of mast cell aggregates and persons without such an increase.

Automated counting of white blood cells in synovial fluid.

OBJECTIVES: To evaluate the performance of automated leucocyte (white blood cell; WBC) counting by comparison with manual counting. METHODS: The number of WBC was determined in heparinized synovial fluid samples by the use of (i) a standard urine cytometer (Kova) and a microscope (reference method) and (ii) a haematology analyser (Sysmex XE-2100; WBC/BASO and DIFF channels). Imprecision within and between days was determined by replicate analysis of a low (level A; WBC approximately 0.560 x 10(9)/l) and a high (level B; WBC approximately 1.081 x 10(9)/l) dedicated synovial fluid control (Quantimetrix). RESULTS: The WBC count of the DIFF channel was highly correlated with the WBC count of the microscopic reference method (r = 0.99; WBC analyser = 0.870 x WBC reference method + 0.413). In contrast, no agreement existed between WBC counts generated by the WBC/BASO channel of the analyser and the reference method (r = 0.52; WBC analyser = 0.008 x WBC reference method + 0.079). Within-day imprecision (4-7%) and between-day imprecision (10%) of the haematology analyser were smaller than the within-day imprecision (12%) and the between-day imprecision (20-22%) of the manual reference method. For manual counting, inter-observer coefficients of variation were 35.9% (control level A) and 21.0% (control level B). CONCLUSIONS: The WBC count in synovial fluid can be reliably determined using the DIFF channel of the Sysmex XE-2100. Automated counting of WBC in synovial fluid offers more precise and faster results than manual counting.

Primary sensitization to sweet bell pepper pollen in greenhouse workers with occupational allergy.

BACKGROUND: In a previous investigation, a high prevalence of allergy to sweet bell pepper pollen was found among exposed horticulture workers. Allergy to plant-derived food is often the consequence of primary sensitization to common pollen allergens. OBJECTIVE: We therefore investigated the cross-reactivity between sweet bell pepper pollen and pollen from grass, birch or mugwort. METHOD: We selected 10 sera from greenhouse workers who had, besides specific IgE against sweet bell pepper pollen, also IgE to grass, birch or mugwort pollen. Cross-reactivity was tested by the inhibition of IgE binding to solid-phase coupled sweet bell pepper pollen extract. The 10 sera were also analysed for IgE binding to sweet bell pepper pollen by immunoblotting. RESULTS: With these sera, no or small inhibition of IgE binding to sweet bell pepper pollen extract was observed with grass, birch and mugwort pollen. With immunoblotting, major IgE-binding structures were seen at 14, 29 and 69 kDa in sweet bell pepper pollen extract. CONCLUSION: The results of our study demonstrate that sweet bell pepper pollen contains allergens that have no or limited cross-reactivity with common pollen allergens. With sera from the 10 patients tested, sensitization to sweet bell pepper pollen was not the consequence of primary sensitization to common pollen allergens.

Urinary N-methylhistamine as an indicator of bone marrow involvement in mastocytosis.

Thirty-seven patients with mastocytosis and unexplained elevated levels of urinary N-methylhistamine who were undergoing bone marrow biopsy were studied with respect to the diagnosis of mastocytosis and the manifestations of the disease. These patients were from a group of 66 patients from whom a bone marrow biopsy was obtained and urinary N-methylhistamine levels were measured in the period 1990-1998. In seven (19%) of the 37 patients, mastocytosis was limited to the skin. Five (14%) of the 37 patients showed accumulation of mast cells in the bone marrow without characteristic skin lesions, whereas seven (19%) of the 37 patients showed increased numbers of mast cells both in the skin and the bone marrow. Eighteen (49%) of the 37 patients with elevated N-methylhistamine did not have mast cell accumulation in either the skin or the bone marrow biopsy. The median level of N-methylhistamine in the urine of patients with mastocytosis limited to the skin was 245 micro mol/mol creatinine. The average level of N-methylhistamine was 509 micro mol/mol creatinine in patients with mast cell accumulation in the bone marrow and cutaneous mastocytosis. There was a significant difference in the levels of N-methylhistamine in patients with mast cell accumulation in the bone marrow biopsy compared with those without. The likelihood of mastocytosis with mast cell accumulation in the bone marrow biopsy at a given level of N-methylhistamine was calculated. It was established that an N-methylhistamine level of 297 micro mol/mol creatinine or higher may be considered as a threshold indicator for obtaining a bone marrow biopsy in patients suspected of mastocytosis with mast cell accumulation in the bone marrow. For practical purposes, we propose to consider the cut-off level of approximately 300 micro mol/mol N-methylhistamine creatinine for this assay.

Prevalence of sensitization to the predatory mite Amblyseius cucumeris as a new occupational allergen in horticulture.

BACKGROUND: Protection against thrips, a common pest in bell pepper horticulture is effectively possible without pesticides by using the commercially available predatory mite Amblyzeius cucumeris (Ac). The prevalence of sensitization to Ac among exposed greenhouse employees and its clinical relevance was studied. METHODS: Four hundred and seventytwo employees were asked to fill in a questionnaire and were tested on location. Next to RAST, skin prick tests (SPTs) were performed with common inhalant allergens, the storage mite Tyrophagus putrescentiae (Tp) which serves as a temporary food source during the cultivation process and Ac. Furthermore, nasal challenge tests with Ac were carried out in 23 sensitized employees. RESULTS: SPTs positive to Ac were found in 109 employees (23%). Work-related symptoms were reported by 76.1%. Sensitization to Tp was found in 62 employees of whom 48 were also sensitized to Ac. Immunoglobulin (Ig)E-mediated allergy to inhalant allergens appeared to be an important risk factor for sensitization to Ac. Employees with rhinitis symptoms showed a significantly higher response to all Ac doses during the nasal challenge test compared with employees without rhinitis symptoms. CONCLUSIONS: The predatory mite Ac is a new occupational allergen in horticulture which can cause an IgE-mediated allergy in exposed employees. It is biologically active on the mucous membranes of the nose and therefore clinically relevant for the development of work-related symptoms.

Prevalence of occupational allergy to bell pepper pollen in greenhouses in the Netherlands.

BACKGROUND: An increasing number of allergic complaints appear to have occurred among bell pepper greenhouse employees. OBJECTIVE: The aim of this study was to estimate the prevalence of work-related allergic symptoms and the prevalence of sensitization to specific occupational allergens and its determinants. METHODS: We studied 472 employees who were invited to answer an extensive questionnaire and to be tested on location with inhalant allergens and home-made extracts of the bell pepper plant. In addition, peak expiratory flow monitoring and RASTs were performed. RESULTS: Work-related symptoms were reported in 53.8% of all cases. Sensitization to the bell pepper plant was found in 35.4%. Positive reactions to leaf, stem and/or juice, however, were associated in nearly 90% with sensitization to pollen, which appeared to be most important allergen of the plant. Sensitization to the bell pepper plant and inhalant atopy were considered the most important risk factors for the occurrence of work-related symptoms of the upper airways (PRR 2.63, CI 2.11-3.25 and PRR 2.25, CI 1.82-2.79) as well as of the lower airways (PRR 4.08, CI 2.38-7.00 and PRR 3.16, CI 1.87-5.33). CONCLUSION: There is a surprisingly high prevalence of work-related respiratory symptoms (53.8%) in bell pepper horticulture. In two-thirds of the employees, symptoms at work were associated with an IgE-mediated allergy due to the high and chronic exposure to bell pepper pollen. Complaints at work without specific sensitization to bell pepper pollen can be caused by non-specific hyper-reactivity or atopy to other occupational allergens. The extent of this occupational allergy has important consequences for the health care of this large, still growing occupational group.

Immunoblot analysis of IgE-binding antigens in paprika and tomato pollen.

BACKGROUND: The high incidence of occupational allergy in horticulture has only recently been recognized. We determined IgE against pollen and fruit from paprika and tomato plants in sera from 3 greenhouse workers and in 3 sera from food-allergic patients. METHODS: Proteins in extracts of paprika and tomato pollen were incubated with patients' sera after covalent coupling of these proteins to agarose beads, or in immunoblots. RESULTS: IgE against paprika pollen, but no IgE against tomato pollen, was found in serum from 2 greenhouse workers who worked with paprika plants only. IgE binding of these 2 sera to agarose-bound paprika pollen extract could be inhibited by paprika pollen but not by tomato pollen extract. A greenhouse worker, who cultivated tomato plants, had IgE against both tomato and paprika pollen. IgE binding of this serum to agarose-bound paprika pollen extract could be inhibited by both paprika pollen and tomato pollen extract. Three food-allergic patients also had IgE against tomato and paprika pollen. IgE from 2 food-allergic patients recognized IgE-binding structures in paprika or tomato pollen that were also present in fruit from the corresponding plant. In contrast, no substantial cross-reactivity was observed between paprika pollen and fruit towards IgE from 3 greenhouse workers. In 4 of 5 sera that were positive in the paprika pollen immunoblot major IgE binding to allergens of about 30 and 64 kD occurred. CONCLUSION: The presence of IgE against paprika or tomato pollen is not restricted to workers in horticulture; IgE against these pollen can also be present in food-allergic patients who have serum IgE against paprika and/or tomato fruit.

Allergy to bumblebee venom. II. IgE cross-reactivity between bumblebee and honeybee venom.

To obtain more information on IgE cross-reactivity between bumblebee venom and honeybee venom, we tested sera from venom-sensitized patients for specific IgE against venoms from the European bumblebee (Bombus terrestris), the North American bumblebee (Megabombus pennsylvanicus), and the honeybee (Apis mellifera). RAST, RAST-inhibition, and immunoblotting experiments indicate that bumblebee venom and honeybee venom contain venom-specific IgE-binding epitopes. These results suggest that immunotherapy using honeybee venom may not be effective in all bumblebee venom-allergic patients. Our experiments also revealed differences in IgE binding for venom from European and American bumblebees.

[Efficiency of mixed RASTs in serological screening of possibly allergic patients]. / Doelmatigheid van mengsel-RASTs bij oriënterend serologisch onderzoek van patiënten met een mogelijke allergie.

OBJECTIVE: To determine the age distribution of results of serological allergy screening using mixed-allergen radioallergoabsorbent tests (RASTs), and to determine the cost-effectiveness of mix RASTs. DESIGN: Retrospective. SETTING: University Hospital of Rotterdam; Dijkzigt Hospital and Sophia Children's Hospital, the Netherlands. METHODS: An analysis was made of the results of all determinations requested in a period of 3 years with use of the CAP Phadiatop test (aimed at IgE against a mixture of inhalant allergens) and the CAP f x 5 food mix test (aimed at IgE against a mixture of food allergens). RESULTS: The CAP Phadiatop test was positive most strongly and most frequently in patients aged 7 to 30 years. From the age of 2 years, there was a sharp rise of the number of positive CAP Phadiatop tests, but IgE against inhalant allergens was also found in over 10% of the children aged 0 and 1 year. Up to and including the age category of 6 years, over one-third of the sera submitted had a positive CAP f x 5 food mix test. From a budget point of view, preliminary screening with a mix RAST is the more advantageous the lower the allergy prevalence and the higher the mean number of CAP-RASTs to be requested per serum. CONCLUSION: A substantial saving of laboratory costs can be achieved by having mix RASTs such as CAP Phadiatop and CAP f x 5 food mix tests precede determinations of specific IgE against separate allergens; these savings will be the higher the smaller the proportion of positive results and the higher the mean number of separate RASTs to be requested per serum.

Prevalence of natural rubber latex allergy (type I and type IV) in laboratory workers in The Netherlands.

The objective of the study was to study the prevalence of Type IV and Type I allergy to natural rubber latex (NRL) in a population at risk in the Netherlands. Laboratory workers regularly using gloves were invited to complete a questionnaire and to be tested. We performed patch tests with standard contact allergens, rubber additives, glove powder and pieces of 4 gloves; prick tests with inhalant allergens, glove extracts, glove powder and fruit extracts; and RASTs. Glove-related hand dermatitis was reported in 36.9% of the individuals interviewed. A positive patch test result for rubber additives was seen in only 6.6%. Glove-related urticaria, rhinoconjunctivitis and/or asthma were reported in 24.6% of all cases. Confirmation of an IgE-mediated reaction was achieved in 8.3% by prick test with glove extracts and 5.0% by RAST. No reaction to glove powder was noticed in patch testing or in prick testing. A high prevalence rate of glove-related symptoms and NRL Type I allergy was found in laboratory workers exposed to rubber gloves. Surprisingly, there was no co-existence of Type I and Type IV allergy in this population.

Nasal responsiveness to allergen and histamine in patients with perennial rhinitis with and without a late phase response.

BACKGROUND: In the lower airways an association has been found between early phase reaction (EPR), late phase reaction (LPR), and bronchial hyperreactivity. However, this association has not been shown for the upper airways in nasal pollen challenge studies. A study was undertaken to determine whether the EPR, LPR, and nasal hyperreactivity are related in perennial allergic rhinitis. METHODS: Twenty four patients with rhinitis who were allergic to house dust mite (HDM) were challenged with HDM extract. The nasal response was monitored by symptom scores and nasal lavages for up to 9.5 hours after challenge and concentrations of albumin, tryptase, and eosinophil cationic protein (ECP) in the lavage fluid were measured. Thirteen patients (defined as dual responders) had increased symptom scores between 3.5 and 9.5 hours compared with the baseline score. The other 11 patients (defined as early responders) showed an isolated EPR only. Nasal hyperreactivity was determined by nasal histamine challenge 24 hours later. RESULTS: Dual responders showed a significantly higher symptom score, albumin influx, and tryptase release during the EPR. During the late phase (3.5-9.5 hours) albumin influx was significantly increased at most time points and ECP release was significantly higher at 9.5 hours in the dual responder group. Dual responders showed a significantly stronger response to all doses of histamine. The area under the curve (AUC) of symptom scores during EPR and LPR and the AUC of the histamine dose response were significantly correlated (EPR-LPR: r = 0.49, p < 0.01; EPR-histamine: r = 0.75, p < 0.001; LPR-histamine: r = 0.66, p < 0.001). CONCLUSIONS: In patients with perennial allergic rhinitis the nasal responses to allergen and histamine are associated. Dual responders have an increased EPR, increased levels of mediators, and increased allergen-induced hyperreactivity.