Abnormal scattergrams and cell population data generated by fully automated hematological analyzers: New tools for screening malaria infection?

INTRODUCTION: Malaria is a life-threatening infectious disease, which has been for long confined to specific endemic areas. Nevertheless, the recent increase in immigration flows from endemic regions and imported cases has reemphasized many diagnostic challenges in Western countries, thus paving the way to introduce rapid and accurate strategies for screening subjects with suspected Malaria infection. Therefore, the aim of this article was to describe our recent experience with Sysmex XN-module for rapid screening of subjects with suspected Malaria. METHODS: Fourteen patients admitted to the Emergency Department (Papa Giovanni XXIII Hospital Bergamo, Italy) with a clinical suspicion of Malaria infection were evaluated, along with 1047 control samples. The analysis of peripheral blood was performed with XN-module, and results were then compared to optical microscopy. RESULTS: Nine patients were positive to Plasmodim falciparum, 3 to Plasmodim vivax, one to Plasmodim ovale, and one to Plasmodim malarie. Characteristic abnormalities could be observed in both white blood cell differential (WDF) and white cell nucleated (WNR) scattergrams (sensitivity 0.64 and specificity 1.0) in 9 samples with parasites at gametocyte or schizos stage irrespective of Plasmodium species and parasitic index, while characteristic scattergram abnormalities could not be seen in the 5 samples containing only parasites at the trophozoites stage. In these cases, specific variations of some cell population data (CPD) could be recorded (sensitivity 1.00 and specificity 0.91). CONCLUSION: The peculiar abnormalities observed in CPDs, WDF, and WNR-scattergrams may raise a definite suspicion of Malaria infection. Further studies should then be planned for validating these preliminary findings and assessing whether these specific abnormalities may be incorporated in rapid and inexpensive Malaria diagnostic algorithms.

Two-site evaluation of the diagnostic performance of the Sysmex XN Body Fluid (BF) module for cell count and differential in Cerebrospinal Fluid.

INTRODUCTION: Cellular analysis in cerebrospinal fluid (CSF) provides important diagnostic information in many pathological settings. The aim of this two-site study was to evaluate the Sysmex XN Body Fluid mode (XN-BF) for cell analysis of CSF compared to light microscopy (LM). METHODS: Two hundred and seven consecutive CSF samples were analyzed in parallel with XN-BF and LM. The study also included the estimation of the limit of blank (LoB), limit of detection (LoD), limit of quantitation (LoQ), carry-over and linearity of XN-BF module. RESULTS: LoQ of white blood cells (WBC) was 3×106  cells/L; linearity was good and carry-over negligible. XN-BF parameters were compared to LM for the following cell classes: total cells, WBC, polymorphonuclear (PMN), and mononuclear (MN) cells. The bias ranged from 1.3 to 15.2×106  cells/L. The receiver operating characteristics curve analysis for WBC showed an area under the curve of 0.98, and the global diagnostic agreement was 95% at a cutoff of 5×106  cells/L. CONCLUSIONS: XN-BF provides rapid and accurate counts in clinically relevant ranges of CSF values, thus providing a valuable alternative to conventional LM analysis. However, microscopic review remains advisable in samples with abnormal cell counts or high fluorescent (HF-BF) cell parameter exceeding 5×106  cells/L.

Evaluation and comparison of automated hematology analyzer, flow cytometry, and digital morphology analyzer for monocyte counting.

INTRODUCTION: This study was aimed to evaluate monocyte counts on Sysmex XN-9000, Sysmex CyFlow Space System, and Sysmex DI60 and compare the performance of these systems with the reference optical microscopy (OM) assessment. METHODS: In all, 55 peripheral blood samples, collected in K3 EDTA tubes, were analyzed with XN-9000, CyFlow System (FlowDiff1 and 2), DI60, and OM. Within-run imprecision was carried out using normal samples. Data comparison was performed with Passing-Bablok regression and Bland-Altman plots. RESULTS: The within-run imprecision of monocyte count on XN, FlowDiff, OM, and DI60 ranged between 1.9% for FlowDiff 2 and 22.1% for DI60. The Passing-Bablok regression analysis of absolute count yielded slopes comprised between 0.93 (FlowDiff2 vs DI60) and 1.21 (DI60 vs OM), whereas the intercepts ranged between -0.002 (FlowDiff 1 vs FlowDiff 2) and 0.13 (FlowDiff1 and 2 vs DI60). Bland-Altman plots in absolute values yielded absolute bias comprised between -0.01 × 109 /L (FlowDiff 1 vs FlowDiff 2; DI60 vs OM) and 0.15 × 109 (XN-module vs DI60). CONCLUSION: The results of this analytical evaluation suggest that flow cytometry generates monocyte counts suitable for routine clinical use. OM or DI60 analysis may be useful for identifying morphologic abnormalities, but does not achieve a satisfactory level of accuracy for enumerating blood cells types such as monocytes, which are usually very low in peripheral blood.

IgA anti-actin antibodies ELISA in coeliac disease: a multicentre study.

BACKGROUND: Previous studies have demonstrated that serum anti-actin antibodies are a reliable marker of intestinal damage severity in coeliac disease. AIMS: To validate in a multicentre study the clinical usefulness of serum IgA anti-actin antibody ELISA and its possible use in monitoring intestinal mucosa lesions during gluten-free diet. PATIENTS AND METHODS: Four centres recruited 205 newly diagnosed coeliac disease patients with villous atrophy, 80 healthy controls and 81 "disease" controls. Twelve coeliac disease patients on gluten-free diet but with persistent symptoms underwent serum IgA anti-actin antibody assay and intestinal histology evaluation. IgA anti-actin antibody ELISA was performed with a commercial kit. All coeliac disease patients underwent intestinal histology study. RESULTS: IgA anti-actin antibodies showed a sensitivity of 80% and a specificity of 85% in the diagnosis of coeliac disease patients with villous atrophy. The area under the receiving operator curve for anti-actin antibodies was 0.873 [95% C.I. 0.805-0.899]. Serum anti-actin antibodies values were significantly higher in coeliac disease patients than in healthy or "disease" controls (P<0.0001). Serum anti-actin antibodies were positive in 41 of the 60 coeliac disease patients with mild intestinal histology lesions (69%) and in 123 of the 145 with severe lesions (85.3%) (P<0.05). There was a significant inverse correlation between anti-actin antibody values and the villi/crypts ratio (r=-0.423; P<0.0001). In the 12 coeliac disease patients on gluten-free diet who underwent re-evaluation as they were persistently symptomatic, intestinal histology showed three cases with persistent villous atrophy: all of these were positive for serum anti-actin antibodies ELISA, whereas both serum anti-tTG and EmAs were negative. The other nine patients showed normal intestinal villi and were negative for serum anti-actin antibodies. CONCLUSIONS: Anti-actin antibodies are a reliable marker of severe intestinal mucosa damage in coeliac disease patients and a simple ELISA technique offers an accurate method for their determination. These antibodies seem to be a very reliable marker of persistent intestinal damage in coeliac disease patients.

Analytical comparison between two hematological analyzer systems: CAL-8000 vs. XN-9000.

INTRODUCTION: This study was aimed to compare the analytical performance of traditional and new parameters and morphological flags of CAL-8000 and XN-9000. The automated differential leukocyte count (DIFF profile) and morphological flags were compared with optical microscopy (OM). METHODS: A total of 1025 peripheral blood samples, collected in K3 EDTA tubes, were analyzed by CAL-8000, by XN-9000, and by OM. Within-run imprecision was performed in low cellularity samples. The comparison was made using Spearman's correlation, Passing-Bablok regression, Bland-Altman bias, and Cohen's K test. RESULTS: Within-run imprecision in low cellularity samples yielded reproducible data between the instruments (imprecision was higher than 10% on samples with platelet count <21 × 109 /L using impedance technology). Passing-Bablok regression (CAL-8000 vs. XN-9000) yielded slopes ranging between 0.2 to 1.16 and intercepts from -6.54 to 21.63. The bias for leukocytes parameters ranged from -1.8% to -82.2%, the red blood cell parameters from -2.9% to 3.1%, platelets parameters from -27.8% to 26%, and reticulocyte parameters from -115.3% to 4.5%. The comparison of morphological flags yielded a K value always <0.55. The DIFF profile vs. OM had a Passing-Bablok regression with slopes ranging between 0.34 to 1.00 and intercepts from -0.01% to 0.11 and bias ranging from -42.9% to 2.6% for XN-9000 parameters and from -2.7% to 35.0% for CAL-8000 parameters. The comparison of morphological flags showed a K value ranging from 0.35 to 0.77 for XN-9000 and from 0.17 to 0.54 for CAL-8000. CONCLUSION: Differences exist between the two analyzers, especially in the generation of morphology flags, thus emphasizing the need of pursuing a major degree of harmonization and/or adopting instrument-specific reference ranges.

Reliability of automated synovial fluid cell counting with Mindray BC-6800 body fluid mode.

INTRODUCTION: The enumeration and differentiation of nuclear elements in synovial fluid is a cornerstone for diagnosis and follow-up of many orthopedic and rheumatologic diseases. In this study, we evaluated the analytical performance of Mindray BC-6800 BF mode (BC-6800-BF) for synovial fluid analysis. METHODS: Overall, 78 synovial fluids were collected and analyzed with both BC-6800-BF and light microscopy. The study also entailed the assessment of limit of blank (LoB), limit of detection (LoD), limit of quantification (LoQ), carryover and linearity. RESULTS: The LoB for the parameters total cells and white blood cells was 6 × 106 cells/L, and the LoD and LoQ were instead 15 and 16 × 106 cells/L, respectively. Linearity was excellent and carryover was negligible. The agreement between BC-6800-BF and light microscopy was satisfactory for all samples pretreated with hyaluronidase, displaying a bias between -5.9% and 8.2%. CONCLUSIONS: The use of BC-6800-BF for synovial fluid analysis enables rapid and accurate assessment, especially for total cell and polymorphonuclear counts. The use of BC-6800-BF may therefore allow the replacement of optical analysis, especially in samples pretreated with hyaluronidase, thus allowing its routine use for the screening of synovial specimens.

In-vitro activity of cefpodoxime proxetil (RU 51807): a comparative disk diffusion study on isolates from geriatric patients.

997 strains isolated from clinical specimens arrived at the "Pio Albergo Trivulzio" microbiology laboratory were tested using disks of cefpodoxime, amoxicillin + clavulanic acid, cefaclor, cefuroxime, ofloxacin, cotrimoxazole, ceftriaxone and cefalexin. Gram-positive strains were tested also with erythromycin, while gram-negative bacteria were tested against aztreonam. Cefpodoxime overall activity was well above the effectiveness of the other oral cephalosporins and on the same order as ceftriaxone and ofloxacin. Cefpodoxime proved to be also more active than the combination amoxicillin-clavulanic acid.

Monoclonal antibody-defined T lymphocyte subpopulations in monoclonal gammapathy of undetermined significance.

We used monoclonal antibodies of the OK series to study T lymphocyte subpopulations in 55 patients with monoclonal gammapathy of undetermined significance (MGUS) and in 40 healthy control subjects, with the aim to investigate if alterations in T lymphocyte subpopulations occur also in MGUS. Mean OKT3+ and OKT8+ cell counts were higher (p less than 0.01 and p less than 0.001, respectively) and the mean OKT4/OKT8 ratio was lower (p less than 0.02) in MGUS patients than in the control subjects. MGUS with the IgM-type monoclonal immunoglobulin (IgM-MGUS) showed the most evident derangement of T lymphocyte subpopulations, i.e., a significant increase of OKT8+ cells and a significant reduction of the OKT4/OKT8 ratio. OKT4+ and OKT8+ cells were significantly increased in patients with high paraprotein concentration (above 16 g/l). Our data suggest that alterations of T lymphocytes are present in MGUS, and that they are similar to those observed in malignant lymphoproliferative disorders.

Interrelationships between body mass and lipid and lipoprotein triglycerides and cholesterol in obese women.

In 72 obese women body mass index positively correlated with age and both showed multiple correlations with serum lipids and lipoprotein lipids. After adjustment for age (partial correlation procedure), body mass index resulted to be positively correlated with serum triglycerides, VLDL lipids, HDL-triglycerides and negatively correlated with HDL-cholesterol. The decrease in HDL-cholesterol concentration along with the increase in body weight was due to the reduction of cholesterol in HDL2 subfraction, while HDL3-cholesterol did not show any significant correlation with body mass index. The negative correlation between HDL2-cholesterol and body mass index was independent of other lipoprotein variables and in particular of VLDL lipid levels that were, as expected, inversely related to HDL2-cholesterol. HDL2-cholesterol is believed to be a strong protective factor against atherosclerosis, while doubts exist on the meaning of VLDL lipids as risk factors. Thus, the decrease in HDL2-cholesterol concentration, rather than the increase in VLDL lipids, might give a rational basis to the high incidence rate of vascular disease in obese subjects.

Fenofibrate therapy of hypertriglyceridaemia. Differential effects on LDL cholesterol level in type IV and in type IIb primary hyperlipoproteinaemia.

Twenty five hypertriglyceridaemic patients (16 Type IV and 9 Type IIb) were treated with fenofibrate 300 mg/d. In Type IV patients serum triglycerides and VLDL cholesterol decreased, while LDL and HDL cholesterol rose significantly. In Type IIb patients, triglycerides and total, VLDL, IDL and LDL cholesterol were significantly reduced by the treatment. The correction of hypertriglyceridaemia by fenofibrate seems, therefore, to induce different changes in lipoproteins in Type IIb and in Type IV hyperlipoproteinaemic patients. The practical implications of these results are discussed.

Changes in serum lipoprotein pattern following bezafibrate. Differential effects in type IIa and in type IIb hyperlipoproteinemic patients.

Sixteen type IIb and 26 type IIa hyperlipoproteinemic patients were treated with bezafibrate 200 mg t.i.d. After 1 month VLDL-TG decreased by 54% in type IIb and by 43% in type IIa; LDL-C decreased by 13% in type IIb and by 23% in type IIa patients. HDL3-C rose in both group of patients, while HDL2-C significantly increased (52%) only in patients with phenotype IIb. The change in LDL-C and in HDL2-C concentrations resulted to be related to the pretreatment VLDL lipid concentration and to its change during the treatment. Twenty patients were treated with bezafibrate for 4 months. On the average, the changes in lipoprotein lipid concentration attained at the end of the first month of therapy remained substantially constant in the following 3 months.

Serum lipoprotein subfractions before and during low-calorie diet in obese women.

Twenty-five obese women were put on low-calorie diet for one month. The mean weight loss was 2.7 kg and was accompanied by a 9% decrease of LDL-cholesterol. LDL-apolipoprotein B (in 13 subjects) showed a 16% fall and LDL-triglyceride levels did not change. The results suggest both a decrease of LDL concentration and a relative increase of triglyceride-rich particles in the density range of 1.006-1.063. HDL2-cholesterol showed a 17% decrease and HDL3-cholesterol a 16% increase. Total and lipoprotein triglycerides were not modified. The changes of lipoprotein pattern suggest a sluggish VLDL catabolism by lipoprotein lipase. In 10 women followed for 6 months the decrease of LDL-cholesterol seen after one month of diet was no more present at the 6th month, while the decrease of HDL2-cholesterol and the increase of HDL3-cholesterol were still evident. At the end of the 6th month a slight but significant decrease of total and VLDL-triglycerides also occurred.

Relationship between metabolic control and HDL2-cholesterol in type I diabetic patients.

The relationship of the degree of metabolic control to the serum lipoprotein pattern has been studied in 62 young insulin treated diabetic patients. Stable HbA1 and 24-h mean blood glucose, but not daily insulin dosage, were positively correlated to VLDL-TG (p less than 0.001) and negatively correlated to HDL2-C (p less than 0.05). HDL2-C was higher in patients with HbA1 less than 10% than in patients with HbA1 greater than 10%. In 13 patients, in whom an improvement of metabolic control was achieved, a significant increase of HDL2-C was recorded after three months. Our data are consistent with the view that in insulin-treated patients, good metabolic control is associated with low VLDL-TG and high HDL2-C levels.

Relationship of serum triglyceride concentration to lipoprotein composition and concentration in normolipidemic and hyperlipidemic subjects.

In a series of 438 subjects (184 normolipidemics and 254 hyperlipidemics) the relationship among serum concentration of triglycerides, lipoprotein lipids and apoproteins A-I and B has been evaluated. The results show that as serum triglyceride level increases, VLDL rise and become enriched in triglycerides. The increase of VLDL is associated with a reduction of serum levels of LDL and HDL which appear to be rich in triglycerides and poor in cholesterol. The decrease in serum HDL level is mainly due to a reduction in serum concentration of the HDL2 subfraction. The triglyceride content of HDL2 and HDL3 rises with increasing serum triglycerides. The increase in serum triglyceride concentration seems then to be associated with a complex metabolic derangement which involves all the lipoprotein fractions.

[Myocardial protection in valvular surgery by retroperfusion of the coronary sinus with cardioplegic solution in valve surgery. Immediate results and technical considerations]. / Protezione miocardica nella chirurgia valvolare mediante retroperfusione del seno coronarico con soluzione cardioplegica nella chirurgia valvolare. Risultati immediati e considerazioni tecniche.

Retrograde coronary sinus perfusion as a means of delivering cardioplegia was evaluated in 20 patients undergoing cardiac valve surgery. Other 10 patients, undergoing similar operation with antegrade coronary cardioplegic perfusion served as a control. Results showed no significant differences between the two groups of patients in terms of: myocardial cooling; pre-ischemic and post-perfusion myocardial lactate extraction; post-operative clinical findings (myocardial infarction occurrence, need for inotropic support, mortality). We conclude that retroperfusion through the coronary sinus is a safe and effective alternative of cardioplegic delivery. Since it simplifies the operative procedure (it allows the cardioplegia delivery at any desired time of the operation, without discontinuing the procedure), we recommended its use during valvular cardiac operations, especially on aortic valve replacement.