Boettcherisca peregrina (Diptera: Sarcophagidae): A flesh fly species of medical and forensic importance

@#Boettcherisca peregrine, as a fly with the necrophagous habits found on human corpses and a vector of disease or parasitic, myiasis-producing agent, is a significant flesh fly species in forensic entomology and medical context. This study reviewed the various aspects of this fly species, including morphology, bionomics, molecular analysis, medical and forensic entomology involvement, such as morphological characteristics of larva, puparia and adult, developmental rate of larvae, the effects of heavy metal (such as Cd and Cu) on the growth and developmentin of larvae, and the impact of some specific stimulis on the labellar chemosensory hair of B. peregrina. Species identification, gene and functions, myiasis and forensic case of this species were also outlined. Therefore, the paper has an important implication for improving the role of B. peregrina in medicine and forensic science.

A quadriplex PCR assay for rapid detection of diarrhoeacausing parasitic protozoa from spiked stool samples

@#Diarrhoea is a leading killer of children, accounting for 9% of all deaths among children under age 5 worldwide and 3% in Malaysia in 2015. A large proportion of diarrhoea illnesses among children in developing countries are ascribed to an unknown etiology because microscopic examination was the only available technique which has low detection limits. The proposed study aimed to evaluate a new quadriplex PCR assay to detect parasitic pathogens namely E. histolytica, G. lamblia and C. parvum which considered responsible for the majority of human infections. Three set of specific primer pairs were designed for detection of parasitic pathogens. Quadriplex PCR assay was optimized and an internal amplification control was incorporated to check for PCR inhibitors in samples. The PCR assay was evaluated using spiked stool samples. Specific primer pairs were successfully designed and simultaneously amplified the targeted genes. The analytical sensitivity of the quadriplex PCR at the DNA level was found to be 50 ng DNA. The analytical specificity was evaluated with 11 reference protozoal and bacterial strains and was found to be 100%. We concluded that the developed quadriplex PCR assay was rapid and gave results within 5 hours which is essential for the identification of parasitic pathogen and might be useful as an additional diagnostic tool whenever time is important in the diagnosis of parasite that cause diarrhoea.

Identification of IgE and IgG1 specific antigens in Echinococcus granulosus cyst fluid

Cystic echinococcosis (CE) is an anthropozoonotic disease with worldwide distribution and is caused by the cestode Echinococcus granulosus. Anaphylactic shock induced by CE rupture is a serious complication especially in patients with hydatid infections, as the resulting leakage of fluid contains highly toxic endogenous antigen. We aimed to isolate and identify the antigens of specific IgE and IgG1 (sIgE and sIgG1) in E. granulosus cyst fluid (EgCF). Crude antigen for EgCF was prepared from E. granulosus-infected sheep liver. Antigens were separated and identified by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1D SDS-PAGE), two-dimensional gel electrophoresis (2-DE), and immunoblotting. Results of 1D SDS-PAGE and immunoblotting showed that 40.5 kDa protein was the major antigen of sIgE, and 35.5 kDa protein was the major antigen of sIgG1 in EgCF. Results of 2-DE and immunoblotting showed that main antigens of sIgE in EgCF were four proteins with pI values ranging from 6.5 to 9.0 and a molecular weight of 40.5 kDa. Main antigens of sIgG1 in EgCF were five proteins with pI values ranging from 6.5 to 9.0 and a molecular weight of 35.5 kDa. The antigens identified for sIgE and sIgG1 can provide critical insights into cellular and molecular mechanisms underlying anaphylactic shock induced by CE.

The consensus sequence of FAMLF alternative splice variants is overexpressed in undifferentiated hematopoietic cells

The familial acute myeloid leukemia related factor gene (FAMLF) was previously identified from a familial AML subtractive cDNA library and shown to undergo alternative splicing. This study used real-time quantitative PCR to investigate the expression of the FAMLF alternative-splicing transcript consensus sequence (FAMLF-CS) in peripheral blood mononuclear cells (PBMCs) from 119 patients with de novo acute leukemia (AL) and 104 healthy controls, as well as in CD34+ cells from 12 AL patients and 10 healthy donors. A 429-bp fragment from a novel splicing variant of FAMLF was obtained, and a 363-bp consensus sequence was targeted to quantify total FAMLF expression. Kruskal-Wallis, Nemenyi, Spearman's correlation, and Mann-Whitney U-tests were used to analyze the data. FAMLF-CS expression in PBMCs from AL patients and CD34+ cells from AL patients and controls was significantly higher than in control PBMCs (P<0.0001). Moreover, FAMLF-CS expression in PBMCs from the AML group was positively correlated with red blood cell count (rs =0.317, P=0.006), hemoglobin levels (rs =0.210, P=0.049), and percentage of peripheral blood blasts (rs =0.256, P=0.027), but inversely correlated with hemoglobin levels in the control group (rs =–0.391, P<0.0001). AML patients with high CD34+ expression showed significantly higher FAMLF-CS expression than those with low CD34+ expression (P=0.041). Our results showed that FAMLF is highly expressed in both normal and malignant immature hematopoietic cells, but that expression is lower in normal mature PBMCs.

Sequence variation and bioinformatics analysis of Toxoplasma gondii GRA16 Gene

Toxoplasmosis is caused by the intracellular protozoan Toxoplasma gondii. It is anopportunistic zoonosis in warm-blooded animals and humans, with a worldwide distribution. Toxoplasma gondii dense granule protein 16 (TgGRA16) can modulate some functions in host cells and is considered a significant virulent factor of the parasite. The present study reports sequence variation in TgGRA16 gene among T. gondii strains from different hosts and geographical locations, and the construction of phylogenetic relationships of these T. gondii strains based on sequences of TgGRA16, and analysis of B cell epitopes in TgGRA16. Our results showed that all TgGRA16 gene sequences were 1518 bp and the C+G contents ranged from 52.17% to 52.59%. Sequence variation in the TgGRA16 gene was 0-1.51%. Phylogenetic analysis revealed that TgGRA16 gene sequence could not be used to differentiate the different T. gondii genotypes. Six B cell epitopes were predicted in TgGRA16. These results indicated that TgGRA16 gene is not an ideal marker for studying genetic relationships of T. gondii isolates, but may represent a good vaccine candidate against toxoplasmosis.

A retrospective clinical study of bevacizumab combined with gemcibabine or paclitaxel in the treatment of recurrent ovarian cancer.

BACKGROUND: Bevacizumab, a recombinant humanized monoclonal antibody that blocks angiogenesis by inhibiting vascular endothelial growth factor A, was described to be effective in the treatment of recurrent or platinum‑resistance ovarian cancer. The present retrospective study was performed to further evaluate the clinical efficacy and toxicity of bevacizumab in the treatment of Chinese recurrent ovarian cancer patients who had been previously treated by platinum‑based chemotherapy. MATERIALS AND METHODS: We reviewed the hospital database and finally included 26 recurrent ovarian cancer patients who were treated with bevacizumab combined with gemcibabine or paclitaxel or single agent. All included patients received >3 cycle of bevacizumab treatment. The tumor response, overall survival, and toxicities were documented. RESULTS: Under the treatment of bevacizumab combined with gemcibabine or paclitaxel, 2 complete response (7.7%), 8 partial response (30.8%), 7 stable disease (26.9%) and 9 progression disease (34.6%) was documented with the objective response rate of 38.5% and disease control rate of 65.4%. The median overall survival from the first application of bevacizumab was 15.3 months [Figure 1] for all of the 26 patients. The median overall survival time was 16.2 and 14.0 months for bevacizumab + gemcitabine and bevacizumab + paclitaxel treatment schedule respectively. The overall survival was not different between bevacizumab + gemcitabine and bevacizumab + paclitaxel treatment regimen hazard ratio = 0.80 (95% confidence interval: 0.32–2, P = 0.64). The hypertension and proteinuria were the major bevacizumab related toxicities. CONCLUSIONS: Bevacizumab combined with gemcibabine or paclitaxel was a promising treatment schedule for platinum‑resistance recurrent ovarian cancer.

Yields from blood cultures of patients with suspected paratyphoid fever A

The yield and speed of detection of Salmonella enterica serotype Paratyphi A from the blood of patients with suspected paratyphoid fever A in 13 500 paired aerobic and anaerobic bottles (AEB, ANB) that were each filled with 5 ml of blood by the BacT/ALERT 3D system were compared, and the blood bacterial counts of 1 000 probable patients were estimated by pour plate method. A total of 4 060 isolates were recovered, of these, 3 149 were recovered from both AEB and ANB, 461 from the AEB only, and 450 from the ANB only. The estimating median bacterial count in blood from 400 patients was 0.5 CFU/ml. The research findings demonstrate that the blood volume drawn is an important factor determining the yields from blood cultures. Growth of significantly more isolates was detected earlier in AEB.

Multiple factor analysis of metachronous upper urinary tract transitional cell carcinoma after radical cystectomy

Transitional cell carcinoma (TCC) of the urothelium is often multifocal and subsequent tumors may occur anywhere in the urinary tract after the treatment of a primary carcinoma. Patients initially presenting a bladder cancer are at significant risk of developing metachronous tumors in the upper urinary tract (UUT). We evaluated the prognostic factors of primary invasive bladder cancer that may predict a metachronous UUT TCC after radical cystectomy. The records of 476 patients who underwent radical cystectomy for primary invasive bladder TCC from 1989 to 2001 were reviewed retrospectively. The prognostic factors of UUT TCC were determined by multivariate analysis using the COX proportional hazards regression model. Kaplan-Meier analysis was also used to assess the variable incidence of UUT TCC according to different risk factors. Twenty-two patients (4.6 percent). developed metachronous UUT TCC. Multiplicity, prostatic urethral involvement by the bladder cancer and the associated carcinoma in situ (CIS) were significant and independent factors affecting the occurrence of metachronous UUT TCC (P = 0.0425, 0.0082, and 0.0006, respectively). These results were supported, to some extent, by analysis of the UUT TCC disease-free rate by the Kaplan-Meier method, whereby patients with prostatic urethral involvement or with associated CIS demonstrated a significantly lower metachronous UUT TCC disease-free rate than patients without prostatic urethral involvement or without associated CIS (log-rank test, P = 0.0116 and 0.0075, respectively). Multiple tumors, prostatic urethral involvement and associated CIS were risk factors for metachronous UUT TCC, a conclusion that may be useful for designing follow-up strategies for primary invasive bladder cancer after radical cystectomy.

Beneficial effect of recombinant human growth hormone on the intestinal mucosa barrier of septic rats

The objective of the present study was to investigate the effects of recombinant human growth hormone (rhGH) on the intestinal mucosa barrier of septic rats and explore its possible mechanism. Female Sprague-Dawley rats were randomized into three groups: control, Escherichia coli-induced sepsis (S) and treatment (T) groups. Groups S and T were subdivided into subgroups 1d and 3d, respectively. Expression of liver insulin-like growth factor-1 (IGF-1) mRNA, Bcl-2 and Bax protein levels and the intestinal Bax/Bcl-2 ratio, and plasma GH and IGF-1 levels were determined. Histological examination of the intestine was performed and bacterial translocation was determined. rhGH significantly attenuated intestinal mucosal injuries and bacterial translocation in septic rats, markedly decreased Bax protein levels, inhibited the decrease of Bcl-2 protein expression and maintained the Bax/Bcl-2 ratio in the intestine. rhGH given after sepsis significantly improved levels of plasma GH (T1d: 1.28 ± 0.24; T3d: 2.14 ± 0.48 æg/L vs S1d: 0.74 ± 0.12; S3d: 0.60 ± 0.18 æg/L; P < 0.05) and IGF-1 (T1d: 168.94 ± 65.67; T3d: 201.56 ± 64.98 æg/L vs S1d: 116.72 ± 13.96; S3d: 107.50 ± 23.53 æg/L; P < 0.05) and expression of liver IGF-1 mRNA (T1d: 0.98 ± 0.20; T3d: 1.76 ± 0.17 vs S1d: 0.38 ± 0.09; S3d: 0.46 ± 0.10; P < 0.05). These findings indicate that treatment with rhGH had beneficial effects on the maintenance of the integrity of the intestinal mucosa barrier in septic rats.

Evaluation of recombinant chitinase antigen in serological diagnosis and surveillance of lymphatic filariasis.

Apply recombinant chitinase fusion protein antigen, enzyme-linked immunosorbent assays examined anti-filarial antibodies and evaluated of useful value in serological diagnosis and surveillance of lymphatic filariasis. The test jirds were immunized and infected by chitinase and B. malayi third stage larvae respectively. Functional protein molecular of chitinase was analyzed by SDS-PAGE, Western blot. The result shown that jirds from microfilaremia (mf) and donors with Mf were directly to react with chitinase antigen that positive rate was 100%, but Mf-xt antigen was only 80%. Normal jirds and persons sera from unepidemic control donors all were negative. False positives of 5% and 20% reacted with chitinase and Mf-xt antigens respectively. The results indicate that recombinant chitinase antigen is suitable for detection of active occult or patent lymphatic filariasis with daytime blood samples in residents of endemic areas, is easy to be performed and inexpensive.

Bronchial responsiveness of aged asthmatic patients to bronchodilator and methacholine.

Geriatric asthma is characterized by prolonged illness, lower remission rate, poor response to therapy and higher mortality rate. We studied bronchodilator response and methacholine challenge in 25 aged non-smoking asthmatic patients; thirty-two young asthmatic patients were included as control. The elderly patients had poorer baseline pulmonary function and were more responsive to a bronchodilator than the younger patients. The response to bronchoprovocation did not show any difference between the two groups. Our findings suggested that the airways of elderly asthmatics are as sensitive as those of younger patients and should not be under-treated.

The modulatory effect of antigen- and PAF-induced asthmatic reaction by aerosol administration of OKY-046 in guinea pigs.

The therapeutic effect of a thromboxane A2 (TXA2) synthetase inhibitor on asthma is still controversial. This study was aimed at clarifying its effect on asthmatic reactions in guinea pigs. Both ovalbumin (OVA)- and platelet activating factor (PAF)-induced dual phase airway spasm and hyperreactivity in guinea pigs were used as the asthma model. Our results demonstrated that aerosol administration of OKY-046 could inhibit both OVA- and PAF-induced late phase bronchoconstriction and airway hyperreactivity to methacholine in OVA sensitized guinea pigs. PAF administration could also induced dual phase bronchoconstriction in normal guinea pigs. Similarly, late phase airway spasm and airway hyperreactivity after PAF exposure was also blocked by OKY-046. In conclusion, aerosol administration of OKY-046 is a safe and effective way to modulate OVA- and PAF-induced asthmatic reactions. The protective effect of OKY-046 on OVA- and PAF-induced late phase bronchoconstriction and airway hyperreactivity indicates that TXA2 might play an important role in the late phase asthmatic reaction and airway hyperreactivity. The normalization of PAF-induced airway hyperreactivity by OKY-046 also indicates that PAF induced airway inflammation might be through the generation of TXA2.

The inhibitory effect of methotrexate on PAF-induced neutrophil and eosinophil locomotion in asthmatic patients.

We have tested the effect of methotrexate (MTX) on platelet activating factor (PAF)-induced neutrophil and eosinophil locomotion, neutrophil leukotriene B4 (LTB4) generation and mononuclear cell DNA synthesis. Neutrophils from patients treated with low dose methotrexate showed reduced PAF-induced chemotactic responses (727.8 +/- 72.2/10 HPF vs 481.9 +/- 87.3/10 HPF, p < 0.05). Both MTX and the specific PAF antagonist BN-52021 significantly inhibited PAF-induced eosinophil and neutrophil locomotion in a dose-dependent manner. MTX also reduced calcium ionophore-driven LTB4 generation from the neutrophils of asthmatics (358.9 +/- 39.5 pg/10(6) cells vs 240.1 +/- 29.1 pg/10(6) cells, p < 0.05) and attenuated PHA-induced mononuclear DNA synthesis as shown by a reduction in 3H-thymidine uptake and propidium iodide staining. These findings support the view that the beneficial effects of MTX in asthma may be due not only to its anti-mitotic effects on the proliferation of mononuclear cells but also to direct effects on granulocyte locomotion and production of LTB4.

The modulatory effect of immunoglobulin G on the CD23 and HLA-DR expression and cytokine production in different groups of asthmatic patients.

The therapeutic effect and mechanism of action of immunoglobulin G (IgG) on bronchial asthma are not defined. Recently, it has been proposed that mononuclear cell (MNC) infiltration in the airway plays a role in the pathogenesis of asthma. In this study, we evaluated the effect of IgG on the cell receptor expression and cytokine production of MNC from two groups (young atopic and old non-atopic) of stable asthmatic patients. MNCs from both asthmatic patients and normal healthy individuals were obtained after Ficoll-Hypaque separation. Cells were cultured in serum free AIM-V medium, with or without phytohemagglutinin (PHA, 5 micrograms/ml) and IgG (100 micrograms/ml). After culture, MNCs were harvested and stained with monoclonal antibodies for HLA-DR (Ia), CD23 and CD3. MNC supernatants were collected for IL-2 and IL-4 measurement. The results showed an enhancing effect of IgG on young atopic MNC proliferation when stimulated with PHA. The production of IL-2 and IL-4 from MNCs were significantly higher in old non-atopic asthmatics after PHA stimulation. The CD23, but not HLA-DR, expression on CD3 positive T cells and cytokines (IL-2 and IL-4) production were increased by IgG when stimulated with PHA in young atopic asthmatics. To the contrary, the effect of IgG on PHA stimulated MNC proliferation, CD23 and HLA-DR expression on CD23 positive T cells in old non-atopic asthmatics were trivial. Only IL-4 production can be significantly inhibited by IgG. These results suggested that the therapeutic effect of IgG on asthmatics might be variable in different groups of asthmatics.(ABSTRACT TRUNCATED AT 250 WORDS)

Enhanced eosinophil luminol-dependent chemiluminescence and complement receptor expression by platelet-activating factor and interleukin-5.

The cytokine interleukin-5 (IL-5) and the lipid mediator platelet-activating factor (PAF) have both been shown to be involved in eosinophil differentiation and activation. We have measured and compared the effect of PAF and IL-5 on human eosinophils in terms of their luminol-dependent chemiluminescence (CL) response and their expression of complement receptors, CR1 and CR3. Both IL-5 and PAF enhanced the eosinophil CL response. The optimal concentrations were 40 U/ml for IL-5, and 10(-6) M for PAF. The priming effect of IL-5 was slow and reached a maximal response after 90 minutes incubation. In contrast, the effect of PAF peaked early and declined during incubation. In the complement receptor study, only PAF was able to enhance CR3 expression (p < 0.05) while the effect of IL-5 on eosinophil complement receptor expression was negligible. These results provide evidence that both inflammatory mediator (PAF) and cytokine (IL-5) can activate eosinophils but the effects of IL-5 and PAF on eosinophil CL response appear to be distinct. The activation of eosinophils by PAF and IL-5 may occur through different mechanisms.

Survival and infectivity of Brugia malayi microfilariae after cryopreservation.

Methods were studied for the cryopreservation of microfilariae of periodic Brugia malayi. RPMI-1640 tissue culture medium containing 6% dimethyl sulfoxide (DMSO) and 15% newborn calf serum was used as cryoprotectant. Samples were frozen slowly in the vapor phase of liquid nitrogen prior to emersion in liquid nitrogen (-196 degrees C). The freezing rate was -0.5 to -1.0 degrees C per minute, microfilariae remained viable for as long as, 212 and 375 days, survival rates were 94 to 98% and they were infective to Aedes togoi mosquitos. The infective larvae (L3) were obtained for 10-11 days after feeding at 28 degrees C room-temperature and the infection rate of L3 in test mosquitos was 22.4-30.6%. All DMSO should be removed from the freezing medium to restore microfilariae activity after freezing.

Evaluación del valor nutricional de leches de soya preparadas a partir de granos de soya tratados con microondas / Evaluation of the nutritive value of soybean milk prepared with microwave-treated soybeans

Se prepararon extractos acuosos de soya a partir de granos previamente sometidos a microondas, para la inactivación casi completa de la lipoxigenasa. Las leches de soya así obtenidas se evaluaron nutricionalmente. Todas las leches sometidas a estudio presentaron menor contenido de proteína, lípidos, ceniza y sólidos totales, en comparación con el alimento control, preparado a partir de granos no tratados por microondas. La leche obtenida de granos de soya con 8.7% de humedad inicial, irradiados con microondas durante 240 segundos, acusó el mejor puntaje químico de aminoácidos esenciales y la mayor disponibilidad aparente de metionina y PER. La inactivación completa de la actividad inhibidora de la tripsina se obtuvo en la leche preparada a partir de granos de soya con 56.8% de humedad, tratados por microondas durante 180 segundos. Por otro lado, la leche obtenida de granos de soya con 38.8% de humedad, irradiados con microondas durante 180 segundos, resultó ser la de mayor digestibilidad de la proteína in vitro

Blood meals of mosquito vectors of filariasis in Guizhou Province, China.

The blood meals of mosquito vectors of W. bancrofti and B. malayi were determined by performing CIEP of the eluant of crushed mosquitoes in filter paper against rabbit anti-human, cow and pig sera. The mosquitoes were collected from houses, cowsheds and pigpens in two counties in Guizhou Province. It was shown that all three species fed on blood from humans, cows and pigs with different preference. While An sinensis fed more on cows, Cx. fatigans and An. lesteri fed on the hosts that were nearby, i.e., Cx. fatigans caught from households fed more on humans, and those collected from cowsheds or pigpens fed more on cows and pigs, respectively.

The influence of microtiter plates from different suppliers on lymphocyte proliferation.

Microtiter plates have been popularly used for lymphocyte culture, but the influence of culture plates from different sources has not been investigated. In this study, the degree of mitogen-induced cell proliferation was investigated using six different brands of flat-bottomed plates. Lymphocytes from twelve normal donors were cultured for 96 hours with several mitogens including PHA, Con A and PWM. Spontaneous cell proliferation was slow and it did not differ significantly among the different plates. However, mitogen-induced cell proliferation showed a wide variation among the six types of plates used. The importance of selecting certain kinds of plates for specific purposes is emphasized.

Effects of gastric cancer cells on lymphocyte proliferation.

Lack of lymphocyte infiltration into gastric cancer tissue appears to be an ominous prognostic indicator. The effects of gastric cancer cells on PHA-induced lymphocyte proliferation were studied. Peripheral lymphocytes were co-cultured for 72 hours with either gastric cancer cells or normal mucosal cells. Pairs of cancerous and normal mucosal cells from stomachs of eight patients, were separately co-cultured with peripheral lymphocytes either from patients or from normal volunteers. The degree of PHA-induced lymphocyte proliferation was measured by 3H-thymidine incorporation. The lymphocyte proliferation was inhibited by the presence of either gastric cancerous or normal mucosal cells in a dose-related manner. The lymphocytes from the normals proliferated twice as much as did the lymphocytes from the patients. The isotope incorporation occurred in lymphocytes rather than in gastric cells since the later incorporated insignificant amounts of isotope. There was no difference between gastric cancerous or normal mucosal cells inhibiting the proliferation of either normal or patients' lymphocytes (p greater than 0.05). In conclusion, gastric cancerous cells (up to 10(6)/ml) have no enhanced inhibition on lymphocyte proliferation when compared with normal gastric mucosal cells.