Visual detection of filaria-specific IgG4 in urine using red-colored high density latex beads

The use of urine for the immunodiagnosis of lymphatic filariasis has a definite advantage: the sample collection is not invasive and thus well accepted by people. Urine-based ELISA to detect filaria-specific IgG4 has been used successfully. However, ELISA requires equipment such as a microplate reader, which is often not available in most endemic areas. We have developed a new visual immunodiagnosis that detects urinary IgG4 using red-colored latex beads (bead test). The sensitivity was 87.2% when ICT antigen test positive people were regarded as the standard (136/156), and the specificity was 97.2% with the non-endemic people in Japan and Bangladesh, and the urine ELISA negatives in Sri Lanka (1264/1300). In a prevalence study, the bead test could detect filarial infection more effectively than ICT test among young children in Sri Lanka, indicating the usefulness of the visual test in epidemiological studies.     

Development of a highly sensitive ELISA for aldosterone in mouse urine: Validation in physiological and pathophysiological states of aldosterone excess and depletion

BackgroundClinical studies have established aldosterone as a critical physiological and pathophysiological factor in salt and water homeostasis, blood pressure control and in heart failure. Genetic and physiological studies of mice are used to model these processes. A sensitive and specific assay for aldosterone is therefore needed to monitor adrenocortical activity in murine studies of renal function and cardiovascular diseases.MethodsAntibodies against aldosterone were raised in sheep as previously described. HRP-Donkey-anti-sheep IgG enzyme tracer was produced in our laboratory using the Lightning-Link HRP technique. Aldosterone ELISA protocol was validated and optimised to achieve the best sensitivity. The assay was validated by analysing the urine of mice collected under various experimental conditions designed to stimulate or suppress aldosterone in the presence of other potentially interfering steroid hormones.ResultsCross-reactivity with the steroids most likely to interfere was minimal: corticosterone = 0.0028%, cortisol = 0.0006%, DOC = 0.0048% except for 5α-dihydro-aldosterone = 1.65%. Minimum detection limit of this ELISA was 5.2 pmole/L (1.5 pg/mL). The validity of urinary aldosterone ELISA was confirmed by the excellent correlation between results obtained before and after solvent extraction and HPLC separation step (Y = 1.092X + 0.03, R² = 0.995, n = 54). Accuracy studies, parallelism and imprecision data were determined and all found to be satisfactory. Using this assay, mean urinary aldosterone levels were (i) approximately 60-fold higher in females than males mice; (ii) increased 6-fold by dietary sodium restriction; (iii) increased 10-fold by ACTH infusion and (iv) reduced by >60% in Cyp11b1 null mice.ConclusionWe describe an ELISA for urinary aldosterone that is suitable for repeated non-invasive measurements in mice. Female aldosterone levels are higher than males. Unlike humans, most aldosterone in mouse urine is not conjugated. Increased levels were noted in response to dietary sodium restriction and ACTH treatment. The sensitivity of the assay is sufficient to detect suppressed levels in mouse models of congenital adrenal hyperplasia.     

Diagnosis of caseous lymphadenitis by ELISA in naturally infected goats from Venezuela

Due to the absence of previous reports, the goal of this work was to detect caseous lymphadenitis (CLA) in goat flocks from Venezuela using an indirect immunoenzymatic assay (ELISA). Eighteen farms were randomly selected in Falcon State, North-Western Venezuela. Blood samples were taken from 259 goats, 65 of them with abscesses. Experimental inoculations were made to healthy kids with 0.5 mL inocula containing 4.7 × 10⁵ of Corynebacterium pseudotuberculosis to observe the kinetics of antibody response. Immunoenzymatic assays were carried out using exotoxin of C. pseudotuberculosis as antigen. Antibody response in experimentally inoculated animals was detected 2 weeks after infection. Of 259 field goat sera, 55.98% were positive by ELISA. Of 65 goats with abscesses, 67.69% had CLA demonstrated by bacteriological methods; from these, 72.73% showed antibodies by ELISA. Of the remaining goats negative to CLA, 47.62% had antibodies by ELISA. Sensitivity was calculated in 72.73% and specificity in 67.74%. The immunoenzymatic assay applied in this research could be useful to detect CLA in naturally infected goat flocks from Venezuela.     

Hyperuricemia and metabolic syndrome in children with overweight and obesity

ObjectiveTo study the prevalence of hyperuricemia in children with overweight or obesity and analyze the relation with metabolic syndrome and the diseases that define it.Materials and methodsThis is a cross-sectional prevalence study in 148 children recruited from pediatric endocrinology consultation, with overweight or obesity (12 ± 3 years, 48% boys, BMI 31.8 ± 6.1). We measured BMI, waist-height, waist circumference, blood pressure with standard instrumentation and glucose (fasting and after overload with 75 g), insulin resistance, cholesterol HDL, triglycerides and uric acid.ResultsThe prevalence of hyperuricemia was 53%. Patients with hyperuricemia had greater BMI (33.9 vs 30.6, p = 0.003), plus waist circumference (101.4 vs 91.1 cm, p < 0.001), higher blood pressure: systolic (123.4 vs 111.9 mm Hg, p < 0.001), diastolic (78.2 vs 68.7 mm Hg, p < 0.001). They presented greater blood glucose after overload oral glucose (107.5 vs 100.7 mg/dl, p = 0.03), insulin was higher (29.2 vs 20.7 mg/dl, p = 0.001) as well as HOMA IR (6.5 vs 4.4, p < 0.001) and HDL levels were lower (49.5 vs 54.4 mg/dl, p = 0.02).Uric acid's level which most is the likely diagnosis of metabolic syndrome corresponds to 5.4 mg/dl in our sample (sensitivity: 64% and specificity 62%).ConclusionThe prevalence of hyperuricemia in children with overweight and obesity is high. In the group of patients with obesity and hyperuricemia, we found out that the parameters measured to diagnose with metabolic syndrome were less favorable. Uric acid's level from where there is a higher possibility to see metabolic syndrome is 5.4 mg/dl.     

Intérêt de la scintigraphie rénale au DMSA dans les pyélonéphrites aiguës de l’enfant

IntroductionAcute pyelonephritis is responsible for significant morbidity during the acute phase and distant irreversible kidney damage. However, diagnosis is often difficult in children. The aim of our study was to evaluate in children hospitalized for suspected acute pyelonephritis prevalence of acute lesions revealed by scintigraphy with ^99mTc labeled dimercaptosuccinic acid (DMSA) and the relationship between renal parenchymal lesions and different clinical factors and biological risks.MethodsThis is a prospective study collecting 40 children admitted with suspected infection of the high urinary tract. All have benefited from DMSA renal scintigraphy.ResultsThe average age of our patients was 3.25 years. The DMSA scan showed signs of acute pyelonephritis in 32.5% of cases. Taking this examination as a reference, fever was the most sensitive clinical signs (76.9%), but the specificity was only 48.1%. CRP (≥ 20 mg/mL) was the most sensitive (92.3%) but with a specificity of 40.7% organic signs. A high CRP was associated with acute pyelonephritis significantly (P = 0.0033). Sixty-three percent of refluxing kidneys were associated with acute pyelonephritis versus 20.8% of non-refluxing kidneys, with a significant difference (P = 0.007).ConclusionIn case of urinary tract infection in children, a complete clinical and laboratory table may be sufficient to retain the diagnosis of acute pyelonephritis. However, this possibility is rare. In the opposite case, a DMSA scan should be performed to confirm the diagnosis.     

Selection of antimalarial drug resistance after intermittent preventive treatment of infants and children (IPTi/c) in Senegal

Senegal has since 2003 used sulphadoxine-pyrimethamine (SP) for Intermittent Preventive Treatment (IPT) of malaria in risk groups. However, the large-scale IPT strategy may result in increasing drug resistance. Our study investigated the possible impact of SP-IPT given to infants and children on the prevalence of SP-resistant haplotypes in the Plasmodium falciparum genes Pfdhfr and Pfdhps, comparing sites with and without IPTi/c. P. falciparum positives samples (n = 352) were collected from children under 5 years of age during two cross-sectional surveys in 2010 and 2011 in three health districts (two on IPTi/c and one without IPTi/c intervention) located in the southern part of Senegal. The prevalence of SP-resistance-related haplotypes in Pfdhfr and Pfdhps was determined by nested PCR followed by sequence-specific oligonucleotide probe (SSOP)–ELISA. The prevalence of the Pfdhfr double mutant haplotypes (CNRN and CICN) was stable between years at < 10% in the control group (P = 0.69), while it rose significantly in the IPTi/c group from 2% in 2010 to 20% in 2011 (P = 0.008). The prevalence of the Pfdhfr triple mutant haplotype (CIRN) increased in both groups, but only significantly in the IPTi/c group from 41% to 65% in 2011 (P = 0.005). Conversely, the Pfdhps 437G mutation decreased in both groups from 44.6% to 28.6% (P = 0.07) and from 66.7% to 47.5% (P = 0.02) between 2010 and 2011 in the control and the IPTi/c groups, respectively. Combined with Pfdhfr, there was a weak trend for decreasing prevalence of quadruple mutants (triple Pfdhfr + Pfdhps 437G) in both groups (P = 0.15 and P = 0.34). During the two cross-sectional surveys, some significant changes were observed in the SP-resistance-related genes. However, since these changes were observed in the two groups, the IPTi/c strategy does only seem to have limited impact on resistance development and other factors as well. However, continuous monitoring will be needed, due to the up-scaling of the IPTi/c strategy in Senegal according to WHO recommendations.     

Evaluación de la utilidad diagnóstica de la biopsia ungueal en las onicomicosis

BackgroundOnychomycosis affects between 2% to 30% of the world population. Nail biopsy may help in making a diagnosis and can distinguish between invasion and colonisation.AimsTo evaluate the diagnostic usefulness of nail biopsy with Periodic acid–Schiff (PAS) staining for onychomycosis, compared to direct KOH examination, culture and its combination in a reference laboratory in Colombia.MethodsThe study included 66 patients in whom a blind and independent reading of the three tests was performed. The usefulness was defined based on the validity (sensitivity, specificity, Youden's index, likelihood ratios), performance (predictive values) efficiency (proportion of correctly diagnosed patients), and reproducibility (kappa coefficient).ResultsThe mean age of the patients was 55 ± 16 years, and included 76% women. The direct tests with KOH were positive in 66.7% (n = 44), 62.1% (n = 41) were positive with culture, and 56.1% (n = 37) with the biopsy. The main causal agents were non-dermatophytes moulds in 36.4% (n = 24). The most frequent species were Neoscytalidium dimidiatum (n = 11), Trichophyton rubrum (n = 11), and Candida parapsilosis (n = 13). The sensitivity of nail biopsy, when compared to the standard (KOH and/or culture), was 71%, specificity 83%, Youden's index 0.54, positive likelihood ratio 4.25, negative likelihood ratio 0.35, positive predictive value 92%, negative predictive value 52%, efficiency 74% and kappa coefficient 0.45. When biopsy was evaluated only in patients with onycholysis of the nail plate greater than 50%, all the parameters of diagnostic usefulness increased.ConclusionsThe overall usefulness of the biopsy was moderate for patients with more severe symptomatology, which makes its use advisable in cases of extensive onycholysis, and when discriminating colonisation from invasion is required.     

Effects of 5 rounds of mass drug administration with diethylcarbamazine and albendazole on filaria-specific IgG4 titers in urine: 6-year follow-up study in Sri Lanka

ELISA for filaria-specific IgG4 in urine (urine ELISA) was applied to children in 7 schools in Sri Lanka, before and after 5 rounds of annual mass drug administration (MDA). The pre-treatment IgG4 prevalence in 2002 was 3.20%, which decreased to 0.91% in 2003 after the first MDA (P<0.001), and finally to 0.36% in 2007 after the 5th MDA. Among 5-10 year-old children, the prevalence decreased from 3.37% in 2002 to 0.51% in 2003 (P=0.009). A pattern of IgG4 titer distribution according to age and its yearly change could also provide useful information in drug efficacy analysis. In 2008, new samples from eleven 2006/07 urine ELISA-positive students and their family members (total n=56) were examined by ICT antigen test, microfilaria test, and urine ELISA. No infection was confirmed among them. Urine ELISA will be useful in monitoring elimination/resurgence in a post-MDA low endemic situation.     

TEP au 18-FDG et carcinose péritonéale d’origine ovarienne : valeurs diagnostique et pronostique avant chimio-hyperthermie intra-péritonéale (CHIP)

AimEvaluate 18-FDG PET/CT diagnostic and prognostic value before HIPEC.Materials and methodsThis retrospective monocentric study included 38 patients with recurrent (n = 27) or primary (n = 11) ovarian cancer who were blinded reviewed for the presence of peritoneal lesions on PET/CT before HIPEC treatment. The results were compared to surgical and histopathological findings, and to survival curves.ResultsThirty-two patients had peritoneal carcinomatosis based on surgical and histopathological findings, and 19 based on PET. There was no suspicious site of abnormal FGD uptake in the supradiaphragmatic regions. The sensitivity and specificity were respectively 56.2 and 100%. Among the 14 false negative patients, 11 had infracentimetric peritoneal implants, and most of them (n = 13) had chemotherapy before HIPEC. There was significantly more patients treated by chemotherapy in the negative PET group (P = 0.02). Even if the event rate observed was higher in the positive PET group for both event free and overall survival (respectively 68% vs. 64% and 26.3% vs. 17.6%), no significant difference was observed using the survival curves (respectively P = 0.62 and 0.59).ConclusionFDG PET/CT before HIPEC showed excellent specificity and lower sensitivity, due to small peritoneal implants and probably to chemotherapy before HIPEC. No significant prognostic value of FDG PET was observed in our study. FDG PET could be considered as a useful tool for detecting distant metastasis with an impact on therapeutic management.     

Intérêt de la tomographie par émission de positons (TEP) au 18FDG dans le suivi des patients traités pour carcinome épidermoïde des voies aérodigestives supérieures (VADS) en rémission clinique

IntroductionPosttreatment follow-up of head and neck squamous cell carcinoma (HNSCC) recurrence is a diagnostic challenge. Tissue distortions from radiation and surgery can obscure early detection of recurrence by conventional follow-up approaches such as physical examination (PE), computed tomography, and magnetic resonance imaging. A number of studies have shown that ¹⁸Fluoro-fluorodeoxyglucose (¹⁸FDG) Positron emission tomography (PET) may be an effective technique for the detection of persistent, recurrent, and distant metastatic HNSCC after treatment. The aim of this prospective study was to determine the benefits (sensitivity, specificity, predictive values, and accuracy) of ¹⁸FDG PET using hybrid PET–Computed tomography system (PET/CT) in the detection of HNSCC subclinical locoregional recurrence and distant metastases, in patients 12 months after curative treatment with a negative conventional follow up.Materials and MethodsNinety-one patients cured from head and neck squamous cell carcinoma (HNSCC) without any clinical element for recurrence were included. Whole-body ¹⁸FDG PET/CT examination was performed 11.6 ± 4.4 months after the end of the treatment. The gold standard was histopathology or 6 months imaging follow-up.ResultsThe whole-body ¹⁸FDG PET/CT of the 91 patients in this study consisted of 52 negative and 39 positive results. Nine of these patients who exhibited abnormal ¹⁸FDG uptake in head and neck area did not have subsequently proven recurrent HNSCC (false positive). Thirty had proven recurrence (true positive). All 52 patients with negative readings of ¹⁸FDG PET/CT remained free of disease at 6 months (true negative). The sensitivity and specificity of ¹⁸FDG PET/CT in this study for the diagnosis of HNSCC recurrence were 100% (30/30) and 85% (52/61) respectively. The positive predictive value was 77% (30/39). The negative predictive value was 100% (52/52). The overall accuracy was 90% (82/91).ConclusionThe results of our study confirm the high effectiveness of ¹⁸FDG PET/CT in assessment of HNSCC recurrence. It suggests that this modality is more accurate than conventional follow-up PE alone in the assessment of patient recurrence after previous curative treatment for HNSCC. Therefore, a PET study could be systematically proposed at 12 months after the end of the treatment.     

Blood lead level and its relationship to essential elements in preschool children from Nanning,China

ObjectiveOur study aimed to assess the distribution of blood lead level and its relationship to essential elements in preschool children in an urban area of China.Design and methodsA total of 6741 children aged 0- to 6-year-old were recruited. Levels of lead, zinc, copper, iron, calcium, and magnesium in whole blood samples were determined using atomic absorption spectrometry.ResultsThe mean blood lead level (BLL) and the prevalence of BLL ≥ 10 μg/dl (5.26 ± 4.08 μg/dl and 6.84%, respectively) increased with age gradually, and there was a gender-difference for blood lead, copper, zinc and iron levels. Compared with the group of children who had BLLs < 5 μg/dl, the groups of 5 ≤ BLLs < 10 μg/dl and 10 ≤ BLLs < 15 μg/dl showed higher blood zinc, iron and magnesium levels, and a lower blood calcium level. A positive correlation of lead with zinc, iron and magnesium, and a negative correlation of lead with calcium were found in the group of children with BLL < 5 μg/dl.ConclusionAge- and gender-differences were found when assessing the BLL and intoxication prevalence in preschool children. Metabolic disorder of essential elements was found even with a low level of lead exposure.     

Circulating soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) as diagnostic and prognostic marker in neonatal sepsis

ObjectiveTriggering receptor expressed on myeloid cells-1 (TREM-1) is an important receptor involved in the innate inflammatory response and sepsis. We assessed soluble TREM-1 (sTREM-1) in 112 septic neonates (63 culture-positive and 49 culture-negative) and 40 healthy controls as a potential early diagnostic and prognostic marker for neonatal sepsis (NS).MethodsStudied neonates were evaluated for early- or late-onset sepsis using clinical and laboratory indicators upon admission. sTREM-1 was measured on initial sepsis evaluation and at 48 h after antibiotic therapy. For ethical reasons, cord blood samples were collected from control neonates and only samples from neonates that proved to be healthy by clinical examination and laboratory analysis were further analyzed for sTREM-1.ResultsBaseline sTREM-1 levels were significantly elevated in culture-proven (1461.1 ± 523 pg/mL) and culture-negative sepsis (1194 ± 485 pg/mL) compared to controls (162.2 ± 61 pg/mL) with no significant difference between both septic groups. Culture-positive or negative septic preterm neonates had significantly higher sTREM-1 compared to full term neonates. sTREM-1 was significantly higher in neonates with early sepsis than late sepsis and was associated with high mortality. sTREM-1 was significantly decreased 48 h after antibiotic therapy compared to baseline or levels in neonates with persistently positive cultures. sTREM-1 was positively correlated to white blood cells (WBCs), absolute neutrophil count, immature/total neutrophil (I/T) ratio, C-reactive protein (hs-CRP) and sepsis score while negatively correlated to gestational age and weight. hs-CRP and sepsis score were independently related to sTREM-1 in multiregression analysis. sTREM-1 cutoff value of 310 pg/mL could be diagnostic for NS with 100% sensitivity and specificity (AUC, 1.0 and 95% confidence interval [CI], 0.696–1.015) while the cutoff value 1100 pg/mL was predictive of survival with 100% sensitivity and 97% specificity (AUC, 0.978 and 95% CI, 0.853–1.13). However, hs-CRP cutoff 13.5 mg/L could be diagnostic for NS with a sensitivity of 76% and specificity of 72% (AUC, 0.762 and 95% CI, 0.612–0.925) and levels were not related to survival as no significant difference was found between dead and alive septic neonates.ConclusionsElevated sTREM-1 could be considered an early marker for NS that reflects sepsis severity and poor prognosis.     

Comparison of accuracy of ultrasonography,progesterone, and pregnancy-associated glycoprotein tests for pregnancy diagnosis in semidomesticated reindeer

The aim of the study was to compare transrectal ultrasound with progesterone (P4) and pregnancy-associated glycoproteins (PAGs) as pregnancy detection methods for semidomesticated reindeer (Rangifer tarandus tarandus) in field conditions. Female reindeer (n = 195) were scanned transrectally by a 7.5-MHz linear array transducer, and blood was sampled either in December 2005 (n = 33), December 2006 (n = 92), or January 2007 (n = 70) during early or mid gestation. Plasma levels of P4 and PAGs were assessed by radioimmunoassay (RIA). Based on calving records, the sensitivity, specificity, predictive values, and the overall accuracy of the three tests were calculated. The overall calving rate calculated from the calving records was 86.2%. The overall accuracy of transrectal ultrasound was 99.5%. The sensitivity and specificity of transrectal ultrasound were 99.4% and 100%, respectively. In the plasma P4 test, the threshold level of 5.0 nmol/L gave the highest overall accuracy (94.9%). The sensitivity of the P4 test decreased from 96.4% to 81.5%, when the threshold level increased from 5.0 nmol/L to 8.0 nmol/L, while the specificity remained at 85.2% over the range of these cutoff values. The overall accuracy of the plasma PAG test decreased from 96.4% to 64.1% when the plasma PAG threshold level increased from 0.5 ng/mL to 3.5 ng/mL, whereas sensitivity decreased from 99.4% to 58.3%. Specificity increased from 77.8% to 100% when the plasma PAG threshold level reached 3.0 ng/mL. Transrectal ultrasound showed higher diagnostic values than those of plasma P4-RIA and PAG-RIA in diagnosing pregnancy of reindeer, with the advantage that diagnoses can be made in real time in field conditions.     

The blood lead levels of outpatients aged 1–18 years from Lu’an,China, 2012–2014

To evaluate blood lead levels (BLLs) and possible influencing factors among children and adolescents in Lu’an, we collected blood samples of 3266 outpatients aged 1–18 years from January 2012 to December 2014 and BLLs were determined by atomizer absorption spectrophotometer. The results showed that the geometric mean (GM) of BLLs was 29.53 μg/L (95% CI: 29.00–30.06 μg/L) and the prevalence of BLLs ≥50 and 100 μg/L were 17.7% and 0.2%, respectively. The GM BLLs and prevalence of BLLs ≥50 μg/L were increased with age before 7 years old and then slightly decreased, and, contrary to previous studies, they were the highest at Jan–Feb and the lowest at Sep–Oct. From 2012 to 2014, the GM BLLs were significantly increased (P < 0.001) while the prevalence of BLLs ≥50 μg/L was decreased but with no significant difference (P > 0.05). These results suggested that although the BLLs of Lu’an children were lower than most areas of China and several neighboring cities as previously reported, they were still higher than that of developed countries. Meanwhile, the GM BLLs of children and adolescents from Lu’an have a trend of increase in recent years. A lot of future works need to be done to identify the risk factors for lead exposure.     

Simultaneous quantitative determination of olmesartan and hydrochlorothiazide in human plasma and urine by liquid chromatography coupled to tandem mass spectrometry

A specific, sensitive and rapid method based on high performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) was developed for the simultaneous determination of olmesartan (OLM) and hydrochlorothiazide (HCTZ) in human plasma and urine. Solid-phase extraction (SPE) was used to isolate the analytes from biological matrices followed by injection of the extracts onto a C₁₈ column with isocratic elution. Detection was carried out on a triple quadrupole tandem mass spectrometer in multiple reaction monitoring (MRM) mode using negative electrospray ionization (ESI). The method was validated over the concentration range of 1.00–1000 ng/mL and 5.00–5000 ng/mL for OLM in human plasma and urine as well as 0.500–200 ng/mL and 25.0–25,000 ng/mL for HCTZ in human plasma and urine, respectively. Inter- and intra-run precision of OLM and HCTZ were less than 15% and the accuracy was within 85–115% for both plasma and urine. The average extraction recoveries were 96.6% and 92.7% for OLM, and 87.2% and 72.1% for HCTZ in human plasma and urine, respectively. The linearity, recovery, matrix effect and stability were validated for OLM/HCTZ in human plasma and urine.     

Determination of melamine and cyanuric acid in human urine by a liquid chromatography tandem mass spectrometry

Melamine was found to be the etiological factor for the urinary stones epidemic in infants and young children in China in 2008. Urine level of melamine and its analog cyanuric acid may be useful markers for the evaluation of toxic effects. Liquid chromatography tandem mass spectrometry methods for the individual determination of melamine and cyanuric acid in human urine are described. Using isotope labeled internal standards during liquid–liquid extraction, the method was fully validated by verifying specificity, linearity, LLOQ, intra- and inter-assay precision and accuracy, matrix effect, recovery and stability. Calibration curves with good linearity (r = 0.9999) over the concentration range from 10 to 5000 ng/ml, intra-assay precision <10% and inter-assay precision <15%, accuracy between 93.0 and 111.6% were obtained with multiple reaction monitoring mode for melamine and cyanuric acid in human urine. The methods were successfully applied to the analysis of urine samples collected from 86 infants and 110 adults.     

Immunocapture of circulating Schistosoma mansoni cathepsin B antigen (Sm31) by anti-Sm31 polyclonal antibodies

Adult Schistosoma mansoni parasites have the capacity to degrade ingested host hemoglobin and other host plasma proteins by using a series of gut proteolytic enzymes, including cathepsin B; this enzyme is released to the host intravascular environment during regurgitations of adult worms. Cathepsin B becomes thus a circulating parasite component that has been shown to be specifically recognized as the Sm31 antigen by antibodies present in most S. mansoni infected patients. Taking advantage of this immunological property, we attempted here to immunocapture Sm31 from sera of infected patients using specific polyclonal rabbit antibodies raised against a highly enriched preparation of Sm31 and detect its intrinsic proteolytic activity using a previously described solid-phase procedure called Cysteine Protease Immuno Assay (CPIA). To produce highly specific anti-Sm31/cathepsin B antibodies, cathepsin B (Sm31 or SmCB) was enriched more than 3000-folds from an adult worm preparation using a series of conventional biochemical steps including ion exchange and affinity chromatography. Anti-cathepsin B antibodies were generated by immunizing rabbits with the enriched cathepsin B fraction; these antibodies recognized a band of Mr. ~ 31 kDa in Western-blot (WB) analysis of this fraction and were able to capture, in a modified CPIA procedure, Sm31/SmCB present in sera from infected Venezuelan patients living in low endemic areas for schistosomiasis. CPIA showed 100% sensitivity and 100% specificity; representing a new diagnostic tool to detect circulating Sm31 antigen in actual infections.     

Serum evaluation of soluble interferon-α/β receptor and high-sensitivity C-reactive protein for diagnosis of the patients with gastrointestinal and hepatobiliary-pancreatic cancer

Serum soluble interferon-α/β receptor (sIFN-α/βR) and high-sensitivity C-reactive protein (hs-CRP) levels were evaluated in the patients with gastrointestinal and hepatobiliary-pancreatic cancer. We compared the sensitivity and specificity of serum sIFN-α/βR with that of serum hs-CRP and evaluated the two diagnostic parameters in combination. Serum sIFN-α/βR levels were measured in 92 patients and 25 healthy individuals by enzyme-linked immunosorbent assay. The diagnoses were 37 cases of hepatocellular carcinoma, 17 cases of pancreatic cancer, 15 cases of colon cancer, 13 cases of biliary tract cancer, and 10 cases of gastric cancer. Serum levels of sIFN-α/βR and hs-CRP were significantly higher in the patients than in healthy individuals (p < 0.05). The optimal cut-off values of sIFN-α/βR and hs-CRP were 3600 pg/ml and 0.5 μg/ml, respectively. The sensitivity and specificity for these thresholds were 94.6% and 88.0%, whereas positive predictive and negative predictive values were 96.7% and 81.5%. These results suggest that a combination of serum sIFN-α/βR and hs-CRP thresholds may be more reliable diagnostic parameter for gastrointestinal and hepatobiliary-pancreatic cancer.     

A comparative evaluation of efficacy of chemotherapy,immunotherapy and immunochemotherapy in visceral leishmaniasis-an experimental study

Visceral leishmaniasis (VL) represents the second most challenging infectious disease worldwide, leading to nearly 500,000 new cases and 60,000 deaths annually. Ninety per cent of VL cases occur in five countries namely Bangladesh, India, Nepal, Sudan and Brazil. No licensed vaccine is available till date against any form of leishmaniasis. High toxicity and increasing resistance to the current chemotherapeutic regimens have further complicated the situation in VL endemic regions of the world. To combat this situation, immunochemotherapy can provide a solution. In the present study, an attempt has been made to assess the in vivo antileishmanial efficacy of chemotherapy, immunotherapy and immunochemotherapy with the use of a first generation antigen Killed Leishmania donovani (KLD) along with a standard drug sodium stibogluconate (SSG) and a newly tested antileishmanial cisplatin. Inbred BALB/c mice were infected with 10⁷ promastigotes/0.1 ml of Leishmania donovani. A month after infection, these animals were given specific immunotherapy (KLD/KLD + MPL-A) or chemotherapy (SSG/cisplatin) or immunochemotherapy (SSG + KLD/SSG + KLD + MPL-A/cisplatin + KLD/cisplatin + KLD + MPL-A). Animals were sacrificed on 1, 15 and 30^th day post treatment. The efficacy of these combinations was assessed in terms of parasite load and by immunological investigations. Infected mice and normal mice served as controls. Results showed that combination of drug and KLD significantly reduced the parasite burden, enhanced the DTH (Delayed Type Hypersensitivity) responses, showed increased levels of IgG2a and decreased levels of IgG1 as compared to mice given chemotherapy or immunotherapy alone. Further maximum protection was provided by SSG + KLD + MPL-A and it was most effective as depicted by 98.5% reduction in parasite load, a potent increase in IFN-γ levels and a significant decrease in IL-10 and IL-4 levels thus skewing the immune response towards Th1 type. Hence, immunochemotherapy is more effective in control of VL in comparison to chemotherapy or immunotherapy.