Diagnosis of Apis dorsata venom allergy: use of recombinant allergens of Apis mellifera and a passive basophil activation test.

BACKGROUND: Allergy to Apis dorsata (Giant Asian Honeybee) venom is the commonest insect allergy in Sri Lanka and South East Asia. However, laboratory diagnosis is difficult as the pure venom and diagnostic reagents are not commercially available. OBJECTIVE: This study assessed the use of four recombinant allergens of A. mellifera venom and the passive basophil activation test in the diagnosis of A. dorsata venom anaphylaxis. METHODS: Serum IgE levels to four recombinant allergens of A. mellifera, rApi m 1, 2, 5 and 10 were assessed and compared with serum IgE to the crude venom of A. mellifera or V. vulgaris by Phadia ImmunoCAP, in patients who developed anaphylaxis to A. dorsata stings. Basophil activation in response to venom of A. dorsata or V. affinis was assessed using a passive basophil activation test. Association of the severity of the reaction with basophil activation was compared. RESULTS: rApi m 1 and 10 combinedly had significant correlation (r = 0.722; p < 0.001) with the crude venom of A. mellifera (Western honeybee) and a higher positivity rate of 90% (27/30). Whereas, IgE reactivity to rApi m 2 or 5 had significant correlation (p = 0.02 and p = 0.005 respectively) with V. vulgaris crude venom. All 30 (100%) were positive to A. dorsata venom in passive BAT; 70% (21/30) had over 80% activation, 96.7% (29/30) had over 60% activation and 100% had over 50% activation. Percentage activation of basophils in patients who had mild or moderate reactions (n = 20) was significantly low (p = 0.02) from that of patients who had severe reactions (n = 10). CONCLUSIONS: rApi m 1 and 10 when combined was sensitive for the diagnosis of A. dorsata allergy. This combination had the lowest cross-reactivity rate with Vespula vulgaris. The passive BAT is highly sensitive in A. dorsata allergy. The basophil reactivity was significantly higher in severe anaphylaxis compared to mild/moderate anaphylaxis. This finding should be further explored in further studies.

Bee and wasp stings in Deniyaya; a series of 322 cases.

OBJECTIVES: To describe wasp and bee species that sting humans, analyse risk factors and clinical features. METHODS: A prospective observational study was conducted on patients presenting to Base Hospital Deniyaya with suspected bee and wasp stings from 2011 to 2013. Data were gathered using a questionnaire and specimens of offending insects collected for identification. When the insect specimen was unavailable, identification was made by the victim selecting (without prompting) from several dead specimens presented by the first author. RESULTS: There were 322 patients (mean age: 42.5 years, SD: 15.1, 173 [53.7%] males). Insects were brought by 55 (17%) and 267 (83%) were identified using specimens. All occurred during day-time, 142 (44.1%) during August and September, and 200 victims (62%) were tea plantation workers. Majority (78.9%) reported a localized painful self-limiting swelling without systemic features and 15 (4.6%) developed anaphylactic shock. None died. Five specimens were available from those in anaphylactic shock (four Apis dorsata, one Ropalidia marginata). Vespa tropica stinging caused a characteristic skin lesion. Of the 55 specimens, 46 (83.6%) were Apis dorsata (Giant honey-bee, ''Bambara''), 8 (14.5%) Vespa tropica (Greater banded hornet, ''Debara'') and one Ropalidia marginata (Paper wasp, ''Kaladuruwa''). CONCLUSIONS: Only three hymenoptera species stings were reported. Risk factors included day-time outdoor activities, occupation (tea plantation workers) and period of year. The latter may be due to pollen season when the insects are found in abundance. Only 4.6% of the patients developed anaphylactic shock. Vespa tropica stings led to a unique skin lesion.

Hump-nosed pit viper (Hypnale hypnale and Hypnale zara) envenoming in Deniyaya, Sri Lanka: Epidemiological and clinical features from a prospective observational study.

Hypnale hypnale, Hypnale zara and Hypnale nepa are the three species of Hump nosed pit vipers that are implicated in human bites in Sri Lanka. H. zara and H. nepa are two endemic species to the country. The objective of the study was to characterize epidemiology and clinical features after different Hump nosed pit viper species bites. A prospective observational study was conducted in Base Hospital Deniyaya from 2013 to 2015. Hump nosed pit viper as the offending snake was identified when the victims brought live or dead specimens. Species identification of dead specimen was done by a herpetologist. Clinical details were recorded during the hospital stay. 83 Hump nosed pit viper bite patients (19-81 years) were studied. Fifty two dead specimens were identified as 39 of H. zara and 13 of H. hypnale by the herpetologist. No H. nepa was identified. Thirty one live snakes were identified as Hump nosed pit vipers and released in to the wild. Envenoming was reported throughout the year. Eighty (96.4%) were daytime bites. Hands (43.4%), feet (55.4%) and arms (1.2%) were affected sites. Sixty two (74.7%) were bitten in tea plantations. All had pain over bite site. Overall, sixty nine (83.1%) had local swelling, seven (8.4%) extensive limb swelling, nineteen (22.9%) haemorrhagic blisters, and seven (8.4%) regional lymphadenopathy. Four (4.8%) patients had incoagulable blood by 20WBCT. H. zara bites had 28.2% haemorrhagic blisters, 7.7% extensive limb swelling and 2.6% coagulopathy. H. hypnale bites had 7.7% each extensive limb swelling and coagulopathy and 23.1% haemorrhagic blisters. None developed elevated serum creatinine. Neurotoxicity was not noted. Hump nosed pit viper bites caused substantial morbidity. Risk factors included day time outdoor activities and occupation. Pain, extensive limb swelling, bite site swelling, haemorrhagic blisters, regional lymphadenopathy and coagulopathy were prominent clinical features. Nephrotoxicity and neurotoxicity were notably absent. Clinical features varied depending on the species implicated.

Sri Lankan green pit viper (Trimeresurus trigonocephalus) bites in Deniyaya: A clinico-epidemiological study.

Green pit viper (Trimeresurus trigonocephalus) is endemic to Sri Lanka and bites are reported from tropical rain forest and plantation areas. As scant data exists on clinico-epidemiological features of Sri Lankan green pit viper bite, objective of this study was to address this knowledge gap. A prospective observational study was conducted in Base Hospital Deniyaya from October 2013 to September 2015. Green pit viper as the offending snake was identified when the victims brought live or dead specimens. When the specimen was not available, green pit viper was identified by the victim pointing to preserved specimens and photographs of green pit viper and different similar appearing snakes. Clinical details were recorded during the hospital stay. Twenty four green pit viper bite patients (17-68 years) were studied. All cases were daytime bites despite green pit viper being a nocturnally active snake. Sixteen patients (67%) were bitten while they were working in tea plantation. Hands (42%), feet, (33%), forearms (8%), legs (8%), thighs (4%) and shoulders (4%) were the sites of bites. Thirteen (54%) had extensive limb swelling. Nine (38%) had local swelling and two had no swelling. Three (12.5%) developed haemorrhagic blisters. Four (17%) had regional lymphadenopathy. Two (8%) developed non clotting blood by the TWBCT. Green pit viper bites caused substantial morbidity. Risk factors included day time outdoor activities in bordering villages of tropical rain forests and occupation. Pain, extensive limb swelling and bite site swelling were prominent clinical features. However haemorrhagic blisters, regional lymphadenopathy and coagulopathy were reported. Nephrotoxicity and neurotoxicity were notably absent.

IgE cross-reactivity of phospholipase A2 and hyaluronidase of Apis dorsata (Giant Asian Honeybee) and Apis mellifera (Western Honeybee) venom: Possible use of A. mellifera venom for diagnosis of patients allergic to A. dorsata venom.

Diagnostic and therapeutic reagents are unavailable for anaphylaxis arising from stings by Apis dorsata. Venom profiles and cross-reactivity of A. dorsata and Apis mellifera were compared, to ascertain whether venom of A. mellifera can be used for diagnosis in A. dorsata allergy. Both venom profiles were similar by High Performance Liquid Chromatography and SDS-PAGE. Sera of 29 of 30 (96.7%) patients with anaphylaxis to A. dorsata stings had IgE to the phospholipase-2 (PLA2) doublet (15 and 16 kDa) of A. dorsata venom by immunoblot, compared to 26 of 30 (86.7%) with the PLA2 of A. mellifera and a purified preparation of PLA2. Twelve patients (40%) with severe anaphylaxis had IgE reactivity to a 39 kDa protein band of venom of both species, a third band, identified in immunoblot as hyaluronidase. The cross-reactivity of PLA2 and hyaluronidase of A. dorsata and A. mellifera were further confirmed by immunoblot inhibition results. Twenty five of 30 (83.3%) of our patients had positive venom specific IgE (>0.35 KUA/L) reactivity to Phadia ImmunoCAPs of A. mellifera venom. The observed IgE cross reactivity suggests the possibility of using A. mellifera venom as a diagnostic test for A. dorsata venom allergy.