Hannahtoxin, the major hemorrhagin purified from king cobra (Ophiophagus hannah) venom, elicits hemorrhages in rabbits but not in mice. Two antisera against hannahtoxin were prepared: one raised against purified hannahtoxin, while the other was raised against glutaraldehyde cross-linked and detoxified hannahtoxin. The antisera were refined by pepsin digestion and ammonium sulfate precipitation. They are of approximately equal potency in their ability to neutralize the hemorrhagic activity of king cobra venom in rabbits. The antisera did not form a precipitin line with venom of snakes of the Viperidae family nor neutralize hemorrhages elicited in mice by any of these venoms. However, when the hemorrhagic activity was assayed in rabbits, both antisera were able to abolish the hemorrhages elicited by all of the venoms tested. These results suggest that hannahtoxin displays few epitopes in common with hemorrhagins of viperid venoms, except those involved in the neutralization of hemorrhagic activity in rabbits. The epitopes of viperid venom hemorrhagins involved in the neutralization reaction in rabbits are different from those in mice.
This study evaluated 2 rapid leptospirosis serological tests, Leptorapide® (Linnodee, Northern Ireland) and VISITECT®-LEPTO (Omega Diagnostics, Scotland, UK), which are commonly used in Malaysia. A total of 183 samples comprised 113 sera from leptospirosis patients, and 70 sera from other infections and healthy controls were used. The leptospirosis sera were grouped into 2 serum panels, i.e., Group I (MAT+, PCR+) and Group II (MAT+). When inconclusive results were interpreted as positives, both tests showed lower diagnostic sensitivities (≤ 34%) with Group I sera, as compared to Group II sera (Leptorapide®, 93%; VISITECT®-LEPTO, 40%). When inconclusive results were interpreted as negatives, the 2 tests showed ~20% sensitivity with both serum panels. The diagnostic specificity of VISITECT®-LEPTO (94%) was superior to Leptorapide® (69%). Since both tests had misdiagnosed a large proportion of Group I patients and showed many inconclusive results among Group II patients, they have limited diagnostic value in detecting acute leptospirosis.
1. Anaesthesia caused marked decreases in the plasma concentrations of triiodothyronine (T3) and thyroxine (T4) and in the body temperature of young fowl. 2. Exogenous T4 or a thyroid hormone secretagogue (somatostatin antiserum), increased endogenous T3 and T4 concentrations and body temperature in conscious birds and prevented the body temperature decline in anaesthetized fowl. 3. These results provide further evidence for a role of T3 and T4 in temperature regulation in birds, particularly during anaesthesia.
Snake envenomation is an important medical problem. One of the hurdles in antivenom development is the in vivo assay of antivenom potency which is expensive, gives variable results and kills many animals. We report a novel in vitro assay involving the specific binding of the postsynaptic neurotoxins (PSNTs) of elapid snakes with purified Torpedo californica nicotinic acetylcholine receptor (nAChR). The potency of an antivenom is determined by its antibody ability to bind and neutralize the PSNT, thus preventing it from binding to nAChR. The PSNT of Naja kaouthia (NK3) was immobilized on microtiter wells and nAChR was added to bind with it. The in vitro IC50 of N. kaouthia venom that inhibited 50% of nAChR binding to the immobilized NK3 was determined. Varying concentrations of antisera against N. kaouthia were separately pre-incubated with 5xIC50 of N. kaouthia venom. The remaining free NK3 were incubated with nAChR before adding to the NK3 coated plates. The in vitro and in vivo median effective ratio, ER50s of 12 batches of antisera showed correlation (R 2) of 0.9809 (p
The aim of this study was to develop an in vitro assay for use in place of in vivo assays of snake venom lethality and antivenom neutralizing potency. A novel in vitro assay has been developed based on the binding of post-synaptically acting α-neurotoxins to nicotinic acetylcholine receptor (nAChR), and the ability of antivenoms to prevent this binding. The assay gave high correlation in previous studies with the in vivo murine lethality tests (Median Lethal Dose, LD50), and the neutralization of lethality assays (Median Effective Dose, ED50) by antisera against Naja kaouthia, Naja naja and Bungarus candidus venoms. Here we show that, for the neurotoxic venoms of 20 elapid snake species from eight genera and four continents, the in vitro median inhibitory concentrations (IC50s) for α-neurotoxin binding to purified nAChR correlated well with the in vivo LD50s of the venoms (R2 = 0.8526, p < 0.001). Furthermore, using this assay, the in vitro ED50s of a horse pan-specific antiserum against these venoms correlated significantly with the corresponding in vivo murine ED50s, with R2 = 0.6896 (p < 0.01). In the case of four elapid venoms devoid or having a very low concentration of α-neurotoxins, no inhibition of nAChR binding was observed. Within the philosophy of 3Rs (Replacement, Reduction and Refinement) in animal testing, the in vitro α-neurotoxin-nAChR binding assay can effectively substitute the mouse lethality test for toxicity and antivenom potency evaluation for neurotoxic venoms in which α-neurotoxins predominate. This will greatly reduce the number of mice used in toxicological research and antivenom production laboratories. The simpler, faster, cheaper and less variable in vitro assay should also expedite the development of pan-specific antivenoms against various medically important snakes in many parts of the world.
Burkholderia pseudomallei is the etiological agent of melioidosis, a severe infectious disease of humans and animals. The role of the bacterium's proteins expressed in vivo during human melioidosis continues to remain an enigma. This study's aim was to identify B. pseudomallei target proteins that elicit the humoral immune response in infected humans. A small insert genomic expression library was constructed and immunoscreened to identify peptides that reacted exclusively with melioidosis patients' sera. Sero-positive clones expressing immunogenic peptides were sequenced and annotated, and shown to represent 109 proteins involved in bacterial cell envelope biogenesis, cell motility and secretion, transcription, amino acid, ion and protein metabolism, energy production, DNA repair and unknown hypothetical proteins. Western blot analysis of three randomly selected full-length immunogenic polypeptides with patients' sera verified the findings of the immunome screening. The patients' humoral immune response to the 109 proteins suggests the induction or significant upregulation of these proteins in vivo during human infection and thus may play a role in the pathogenesis of B. pseudomallei. Identification of B. pseudomallei immunogens has shed new light on the elucidation of the bacterium's pathogenesis mechanism and disease severity. These immunogens can be further evaluated as prophylactic and serodiagnostic candidates as well as drug targets.
A seroepidemiologic survey of Plasmodium vivax and Plasmodium falciparum transmission was conducted in 94 Orang Asli children and adults. The prevalence of malaria was 46% in this population, and infections due to P. vivax and P. falciparum occurred with equal frequency. Multi-species infection was common, particularly in children less than 10 years of age. Circumsporozoite (CS) antibodies to P. vivax were detected by ELISA, using the recombinant protein NS181V20, in sera from 53-95% of all subjects in this study. The specificity of reactivity to NS181V20 was confirmed by immunofluorescence using air-dried sporozoites. CS antibodies to P. falciparum were present in less than 50% of the population less than 30 years of age. These data support further testing of this protein as a candidate vivax vaccine.