Adaptation and validation of antibody-ELISA using dried blood spots on filter paper for epidemiological surveys of tsetse-transmitted trypanosomosis in cattle

The indirect enzyme-linked immunosorbent assay (ELISA) for the detection of anti-trypanosomal antibodies in bovine serum was adapted for use with dried blood spots on filter paper. Absorbance (450 nm) results for samples were expressed as percent positivity, i.e. percentage of the median absorbance result of four replicates of the strong positive control serum. The antibody-ELISA was evaluated in Zambia for use in epidemiological surveys of the prevalence of tsetse-transmitted bovine trypanosomosis. Known negative samples (sera, n = 209; blood spots, n = 466) were obtained from cattle from closed herds in tsetse-free areas close to Lusaka. Known positive samples (sera, n = 367; blood spots, n = 278) were obtained from cattle in Zambia's Central, Lusaka and Eastern Provinces, diagnosed as being infected with Trypanosoma brucei, T. congolense, or T. vivax using the phase-contrast buffy-coat technique or Giemsa-stained thick and thin blood smears. For sera (at a cut-off value of 23.0% positivity) sensitivity and specificity were 86.1 and 95.2%, respectively. For bloodspots (at a cut-off value of 18.8% positivity) sensitivity and specificity were 96.8 and 95.7%, respectively. The implications of persistence of antibodies following treatment or self-cure are discussed.     

Differential acetylcholine and GABA release from cultured chick retina cells

In the present work we investigated the mechanisms controlling the release of acetylcholine (ACh) and of gamma-aminobutyric acid (GABA) from cultures of amacrine-like neurons, containing a subpopulation of cells which are simultaneously GABAergic and cholinergic. We found that 81.2 +/- 2.8% of the cells present in the culture were stained immunocytochemically with an antibody against choline acetyltransferase, and 38.5 +/- 4.8% of the cells were stained with an antibody against GABA. Most of the cells containing GABA (87.0 +/- 2.9%) were cholinergic. The release of acetylcholine and GABA was mostly Ca2+-dependent, although a significant release of [3H]GABA occurred by reversal of its transporter. Potassium evoked the Ca2+-dependent release of [3H]GABA and [3H]acetylcholine, with EC50 of 31.0 +/- 1.0 mm and 21.6 +/- 1.1 mm, respectively. The Ca2+-dependent release of [3H]acetylcholine was significantly inhibited by 1 micrometer tetrodotoxin and by low (30 nm) omega-conotoxin GVIA (omega-CgTx GVIA) concentrations, or by high (300 nm) nitrendipine (Nit) concentrations. On the contrary, the release of [14C]GABA was reduced by 30 nm nitrendipine, or by 500 nm omega-CgTx GVIA, but not by this toxin at 30 nm. The release of either transmitters was unaffected by 200 nm omega-Agatoxin IVA (omega-Aga IVA), a toxin that blocks P/Q-type voltage-sensitive Ca2+ channels (VSCC). The results show that Ca2+-influx through omega-CgTx GVIA-sensitive N-type VSCC and through Nit-sensitive L-type VSCC induce the release of ACh and GABA. However, the significant differences observed regarding the Ca2+ channels involved in the release of each neurotransmitter suggest that in amacrine-like neurons containing simultaneously GABA and acetylcholine the two neurotransmitters may be released in distinct regions of the cells, endowed with different populations of VSCC.     

Changes in bacterial species composition in enrichment cultures with various dilutions of inoculum as monitored by denaturing gradient gel electrophoresis

Denaturing gradient gel electrophoresis revealed changes in the bacterial species obtained from enrichment cultures with different inoculum dilutions. This inoculum dilution enrichment approach may facilitate the detection and isolation of a greater number of bacterial species than traditional enrichment techniques.     

Mesenteric venous thrombosis after appendicectomy in a child: clinical case and review of the literature

Mesenteric venous thrombosis (MVT) after appendicectomy has not been reported in the pediatric literature. This complication is associated with significant morbidity and mortality rates in adults. The report of this case attempts to analyze the features found in an appendicectomized child documented by radiological examination and surgery before and after starting heparin therapy. Anticoagulation treatment should be started as soon as possible and continued for a long time to decrease the risk of relapse.     

Human DDX3X Unwinds Japanese Encephalitis and Zika Viral 5′ Terminal Regions

Flavivirus genus includes many deadly viruses such as the Japanese encephalitis virus (JEV) and Zika virus (ZIKV). The 5′ terminal regions (TR) of flaviviruses interact with human proteins and such interactions are critical for viral replication. One of the human proteins identified to interact with the 5′ TR of JEV is the DEAD-box helicase, DDX3X. In this study, we in vitro transcribed the 5′ TR of JEV and demonstrated its direct interaction with recombinant DDX3X (K_d of 1.66 ± 0.21 µM) using microscale thermophoresis (MST). Due to the proposed structural similarities of 5′ and 3′ TRs of flaviviruses, we investigated if the ZIKV 5′ TR could also interact with human DDX3X. Our MST studies suggested that DDX3X recognizes ZIKV 5′ TR with a K_d of 7.05 ± 0.75 µM. Next, we performed helicase assays that suggested that the binding of DDX3X leads to the unwinding of JEV and ZIKV 5′ TRs. Overall, our data indicate, for the first time, that DDX3X can directly bind and unwind in vitro transcribed flaviviral TRs. In summary, our work indicates that DDX3X could be further explored as a therapeutic target to inhibit Flaviviral replication     

Fatty acid composition of oils from 21 species of marine fish,freshwater fish and shellfish

The fatty acid composition of body lipids was determined by GLC for 14 species of saltwater fish, three species of freshwater fish and four species of shellfish. In addition, liver lipids of two species and egg lipids of one species were analyzed for comparison with the fish body lipids. The various species ranged from lean to fatty and contained from 0.7~15.5% oil in the tissues. Certain major fatty acids were found to vary widely among the species, as follows: 1.6~8.0% myristic, 9.5~33.4% palmitic, 2.0~11.2% palmitoleic, 5.2~29.1% oleic, 0.7~10.5% eicosenoic, 5.0~21.5% eicosapentaenoic, 0.2~11.6% docosenoic and 5.9~26.2% docosahexaenoic acids. Analyses of two separate mullet-oil samples illustrated the wide differences that are possible for a single species caught during different seasons. Significant differences in the amt of particular fatty acids were found in comparing freshwater-fish analyses with analyses for marine fish. Oysters and scallops showed large amt of pentaenoic and hexaenoic acids in their oils. Presented in part at the AOCS Meeting, New York, 1960.